Alcohols enhance caerulein-induced zymogen activation in pancreatic acinar cells

PubMed Central

LU, ZHAO; KARNE, SURESH; KOLODECIK, THOMAS; GORELICK, FRED S.

2010-01-01

Activation of zymogens within the pancreatic acinar cell is an early feature of acute pancreatitis. Supraphysiological concentrations of cholecystokinin (CCK) cause zymogen activation and pancreatitis. The effects of the CCK analog, caerulein, and alcohol on trypsin and chymotrypsin activation in isolated pancreatic acini were examined. Caerulein increased markers of zymogen activation in a time- and concentration-dependent manner. Notably, trypsin activity reached a peak value within 30 min, then diminished with time, whereas chymotrypsin activity increased with time. Ethanol (35 mM) sensitized the acinar cells to the effects of caerulein (10−10 to 10−7 M) on zymogen activation but had no effect alone. The effects of ethanol were concentration dependent. Alcohols with a chain length of ≥2 also sensitized the acinar cell to caerulein; the most potent was butanol. Branched alcohols (2-propanol and 2-butanol) were less potent than aliphatic alcohols (1-propanol and 1-butanol). The structure of an alcohol is related to its ability to sensitize acinar cells to the effects of caerulein on zymogen activation. PMID:11842000

An update on pancreatic pathophysiology (do we have to rewrite pancreatic pathophysiology?).

PubMed

Hammer, Heinz F

2014-02-01

This review focuses on seven aspects of physiology and pathophysiology of the exocrine pancreas that have been intensively discussed and studied within the past few years: (1) the role of neurohormonal mechanisms like melatonin, leptin, or ghrelin in the stimulation of pancreatic enzyme secretion; (2) the initiation processes of acute pancreatitis, like fusion of zymogen granules with lysosomes leading to intracellular activation of trypsinogen by the lysosomal enzyme cathepsin B, or autoactivation of trypsinogen; (3) the role of genes in the pathogenesis of acute pancreatitis; (4) the role of alcohol and constituents of alcoholic beverages in the pathogenesis of acute pancreatitis; (5) the role of pancreatic hypertension, neuropathy, and central mechanisms for the pathogenesis of pain in chronic pancreatitis; (6) the relation between exocrine pancreatic function and diabetes mellitus; and (7) pathophysiology, diagnosis and treatment of pancreatic steatorrhea.

Zymogen proteolysis within the pancreatic acinar cell is associated with cellular injury.

PubMed

Grady, T; Mah'Moud, M; Otani, T; Rhee, S; Lerch, M M; Gorelick, F S

1998-11-01

The pathological activation of digestive zymogens within the pancreatic acinar cell probably plays a central role in initiating many forms of pancreatitis. To examine the relationship between zymogen activation and acinar cell injury, we investigated the effects of secretagogue treatment on isolated pancreatic acini. Immunofluorescence studies using antibodies to the trypsinogen-activation peptide demonstrated that both CCK (10(-7) M) hyperstimulation and bombesin (10(-5) M) stimulation of isolated acini resulted in trypsinogen processing to trypsin. These treatments also induced the proteolytic processing of procarboxypeptidase A1 to carboxypeptidase A1 (CA1). After CCK hyperstimulation, most CA1 remained in the acinar cell. In contrast, the CA1 generated by bombesin was released from the acinar cell. CCK hyperstimulation of acini was associated with cellular injury, whereas bombesin treatment did not induce injury. These studies suggest that 1) proteolytic zymogen processing occurs within the pancreatic acinar cell and 2) both zymogen activation and the retention of enzymes within the acinar cell may be required to induce injury.

X-ray microanalysis of exocrine glands in animal models for cystic fibrosis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mueller, R.M.R.; Roomans, G.M.

1985-01-01

Elemental distribution and ultrastructure of the submandibular gland, the parotid gland and the pancreas were investigated in three suggested animal models of the disease cystic fibrosis: the chronically reserpinized rat, the chronically isoproterenol-treated rat, and the chronically pilocarpine-treated rat. To elucidate the cellular mechanism underlying the effects of these treatments, chronic effects of specific alpha - and beta -adrenergic agonists, as well as acute effects of reserpine and various agonists were also investigated. Reserpine, isoproterenol, and pilocarpine cause an increase in the calcium concentration in submandibular gland acinar cells, due to an increased calcium content of the intracellular mucus. Inmore » the parotid gland, reserpine and isoproterenol cause a decrease of the calcium concentration in acinar cells, due to a lower calcium content of the zymogen granules. In the submandibular gland, a decreased cellular Na concentration was noted after chronic treatment with isoproterenol or pilocarpine, and after a single dose of reserpine or isoproterenol. Ultrastructural changes in the exocrine glands investigated included excessive accumulation of intracellular secretory material and formation of abnormal uncondensed secretion granules. A common pattern in the animal models appears to be (1) inhibition of secretion resulting in intracellular accumulation of secretory material, (2) synthesis of secretory macromolecules with altered cation-binding properties.« less

Roles of CUB and LDL receptor class A domain repeats of a transmembrane serine protease matriptase in its zymogen activation

PubMed Central

Inouye, Kuniyo; Tomoishi, Marie; Yasumoto, Makoto; Miyake, Yuka; Kojima, Kenji; Tsuzuki, Satoshi; Fushiki, Tohru

2013-01-01

Matriptase is a type II transmembrane serine protease containing two complement proteases C1r/C1s–urchin embryonic growth factor–bone morphogenetic protein domains (CUB repeat) and four low-density lipoprotein receptor class A domains (LDLRA repeat). The single-chain zymogen of matriptase has been found to exhibit substantial protease activity, possibly causing its own activation (i.e. conversion to a disulfide-linked two-chain fully active form), although the activation seems to be mediated predominantly by two-chain molecules. Our aim was to assess the roles of CUB and LDLRA repeats in zymogen activation. Transient expression studies of soluble truncated constructs of recombinant matriptase in COS-1 cells showed that the CUB repeat had an inhibitory effect on zymogen activation, possibly because it facilitated the interaction of two-chain molecules with a matriptase inhibitor, hepatocyte growth factor activator inhibitor type-1. By contrast, the LDLRA repeat had a promoting effect on zymogen activation. The effect of the LDLRA repeat seems to reflect its ability to increase zymogen activity. The proteolytic activities were higher in pseudozymogen forms of recombinant matriptase containing the LDLRA repeat than in a pseudozymogen without the repeat. Our findings provide new insights into the roles of these non-catalytic domains in the generation of active matriptase. PMID:23038671

Pharmacological and genetic inhibition of calcineurin protects against carbachol-induced pathological zymogen activation and acinar cell injury.

PubMed

Muili, Kamaldeen A; Ahmad, Mahwish; Orabi, Abrahim I; Mahmood, Syeda M; Shah, Ahsan U; Molkentin, Jeffery D; Husain, Sohail Z

2012-04-15

Acute pancreatitis is a major health burden for which there are currently no targeted therapies. Premature activation of digestive proenzymes, or zymogens, within the pancreatic acinar cell is an early and critical event in this disease. A high-amplitude, sustained rise in acinar cell Ca(2+) is required for zymogen activation. We previously showed in a cholecystokinin-induced pancreatitis model that a potential target of this aberrant Ca(2+) signaling is the Ca(2+)-activated phosphatase calcineurin (Cn). However, in this study, we examined the role of Cn on both zymogen activation and injury, in the clinically relevant condition of neurogenic stimulation (by giving the acetylcholine analog carbachol) using three different Cn inhibitors or Cn-deficient acinar cells. In freshly isolated mouse acinar cells, pretreatment with FK506, calcineurin inhibitory peptide (CiP), or cyclosporine (CsA) blocked intra-acinar zymogen activation (n = 3; P < 0.05). The Cn inhibitors also reduced leakage of lactate dehydrogenase (LDH) by 79%, 62%, and 63%, respectively (n = 3; P < 0.05). Of the various Cn isoforms, the β-isoform of the catalytic A subunit (CnAβ) was strongly expressed in mouse acinar cells. For this reason, we obtained acinar cells from CnAβ-deficient mice (CnAβ-/-) and observed an 84% and 50% reduction in trypsin and chymotrypsin activation, respectively, compared with wild-type controls (n = 3; P < 0.05). LDH release in the CnAβ-deficient cells was reduced by 50% (n = 2; P < 0.05). The CnAβ-deficient cells were also protected against zymogen activation and cell injury induced by the cholecystokinin analog caerulein. Importantly, amylase secretion was generally not affected by either the Cn inhibitors or Cn deficiency. These data provide both pharmacological and genetic evidence that implicates Cn in intra-acinar zymogen activation and cell injury during pancreatitis.

Pharmacological and genetic inhibition of calcineurin protects against carbachol-induced pathological zymogen activation and acinar cell injury

PubMed Central

Muili, Kamaldeen A.; Ahmad, Mahwish; Orabi, Abrahim I.; Mahmood, Syeda M.; Shah, Ahsan U.; Molkentin, Jeffery D.

2012-01-01

Acute pancreatitis is a major health burden for which there are currently no targeted therapies. Premature activation of digestive proenzymes, or zymogens, within the pancreatic acinar cell is an early and critical event in this disease. A high-amplitude, sustained rise in acinar cell Ca2+ is required for zymogen activation. We previously showed in a cholecystokinin-induced pancreatitis model that a potential target of this aberrant Ca2+ signaling is the Ca2+-activated phosphatase calcineurin (Cn). However, in this study, we examined the role of Cn on both zymogen activation and injury, in the clinically relevant condition of neurogenic stimulation (by giving the acetylcholine analog carbachol) using three different Cn inhibitors or Cn-deficient acinar cells. In freshly isolated mouse acinar cells, pretreatment with FK506, calcineurin inhibitory peptide (CiP), or cyclosporine (CsA) blocked intra-acinar zymogen activation (n = 3; P < 0.05). The Cn inhibitors also reduced leakage of lactate dehydrogenase (LDH) by 79%, 62%, and 63%, respectively (n = 3; P < 0.05). Of the various Cn isoforms, the β-isoform of the catalytic A subunit (CnAβ) was strongly expressed in mouse acinar cells. For this reason, we obtained acinar cells from CnAβ-deficient mice (CnAβ−/−) and observed an 84% and 50% reduction in trypsin and chymotrypsin activation, respectively, compared with wild-type controls (n = 3; P < 0.05). LDH release in the CnAβ-deficient cells was reduced by 50% (n = 2; P < 0.05). The CnAβ-deficient cells were also protected against zymogen activation and cell injury induced by the cholecystokinin analog caerulein. Importantly, amylase secretion was generally not affected by either the Cn inhibitors or Cn deficiency. These data provide both pharmacological and genetic evidence that implicates Cn in intra-acinar zymogen activation and cell injury during pancreatitis. PMID:22323127

Intracellular proteolysis of pancreatic zymogens.

PubMed Central

Gorelick, F. S.; Modlin, I. M.; Leach, S. D.; Carangelo, R.; Katz, M.

1992-01-01

Activation of pancreatic digestive zymogens within the pancreatic acinar cell may be an early event in the development of pancreatitis. To detect such activation, an immunoblot assay has been developed that measures the relative amounts of inactive zymogens and their respective active enzyme forms. Using this assay, high doses of cholecystokinin or carbachol were found to stimulate the intracellular conversion of at least three zymogens (procarboxypeptidase A1, procarboxypeptidase B, and chymotrypsinogen 2) to their active forms. Thus, this conversion may be a generalized phenomenon of pancreatic zymogens. The conversion is detected within ten minutes of treatment and is not associated with changes in acinar cell morphology; it has been predicted that the lysosomal thiol protease, cathepsin B, may initiate this conversion. Small amounts of cathepsin B are found in the secretory pathway, and cathepsin B can activate trypsinogen in vitro; however, exposure of acini to a thiol protease inhibitor (E64) did not block this conversion. Conversion was inhibited by the serine protease inhibitor, benzamidine, and by raising the intracellular pH, using chloroquine or monensin. This limited proteolytic conversion appears to require a low pH compartment and a serine protease activity. After long periods of treatment (60 minutes), the amounts of the active enzyme forms began to decrease; this observation suggested that the active enzyme forms were being degraded. Treatment of acini with E64 reduced this late decrease in active enzyme forms, suggesting that thiol proteases, including lysosomal hydrolases, may be involved in the degradation of the active enzyme forms. These findings indicate that pathways for zymogen activation as well as degradation of active enzyme forms are present within the pancreatic acinar cell. Images FIG. 1 FIG. 6 PMID:1340058

Proliferative capability of parietal and zymogen cells.

PubMed Central

Chen, K Y; Withers, H R

1975-01-01

Six to eight week old male mice of C3Hf/Bu strain were killed and studied at various times from 30 minutes to 30 days after tritiated thymidine (3H-TdR) injection. Labelling of parietal and zymogen cells was observed in autoradiographic histological specimens. There were 590 +/- 22 gastric glands per circumference at the body of the stomach. Parietal cells were counted as 16-2 (14-0-18-4) per gland or 9600 per circumference. There were 11-4 (9-4-13-7) zymogen cells per gland or 6700 per circumference. Two labelled immature parietal cells per circumference were seen 30 minutes after 3H-TdR injection, equivalent to a ratio of 1:5000; more mature labelled parietal cells were seen at later times up to 30 days. There was evidence for proliferation during maturation and downward migration of cells toward the body and the lower part of the gastric gland. Our data support earlier evidence that immature parietal cells constantly supply mature parietal cells through migration from a proliferative zone. Whether the parietal cells are derived from the same stem cell compartment as surface epithelium cells is unclear at the present time. An average of 2-6 zymogen cells per circumference, or 1:2500, was found to be labelled. While most zymogen cells were not proliferating, cells entered the proliferating cycle at random. Based on the findings reported in this paper and the radiation responses of both parietal and zymogen cells, it is postulated that they form part of a slow renewal system. Images Fig. 1 (cont.) Fig. 1 (cont.) Fig. 1 (cont.) Fig. 1 Fig. 2 (cont.) Fig. 2 (cont.) Fig. 2 PMID:1213947

Sericin ameliorated dysmorphic mitochondria in high-cholesterol diet/streptozotocin rat by antioxidative property.

PubMed

Ampawong, Sumate; Isarangkul, Duangnate; Aramwit, Pornanong

2017-02-01

Sericin has been implicated in lower cholesterolemic effect due to its properties with several mechanisms. Mitochondria are one of the most important targets to be affected in high blood cholesterol and glucose conditions. The protective role of sericin on mitochondria remains doubtful. To examine this role, electron microscopic, histopathologic, immunohistochemical, and biochemical studies were performed in a high-cholesterol diet/streptozotocin rat model. The results demonstrated that sericin reduced blood cholesterol without hypoglycemic effect. Sericin alleviated dysmorphic mitochondria in heart and liver but not in kidney and also decreased peculiar endoplasmic reticulum in the exocrine pancreas. In addition, sericin decreased hepatic steatosis and preserved zymogen granule referable to the decline of reactive oxygen species production in hepatic mitochondrial extraction and down-regulation of malondialdehyde expression in the liver and exocrine pancreas however irrelevant to lipase activity. This study suggests that sericin has antioxidative property to reduce blood cholesterol by means of diminishing fat deposit in hepatocyte and improves mitochondria and endoplasmic reticulum integrities. [Box: see text].

Stress-activated protein kinase activation is the earliest direct correlate to the induction of secretagogue-induced pancreatitis in rats.

PubMed

Grady, T; Dabrowski, A; Williams, J A; Logsdon, C D

1996-10-03

We compared the cellular events induced by hyperstimulation of rats with caerulein which induces acute pancreatitis, to bombesin, which does not induce pancreatitis. Both secretogogues induced the intracellular activation of trypsinogen and the colocalization of lysosomal hydrolases and zymogen granules within 10-15 minutes. These data indicate that these parameters, previously thought to be crucial initiating events of pancreatitis, are not definitive cellular markers of the disease. We then compared the abilities of the two secretagogues to activate stress-activated protein kinase (SAPK). Significant effects of caerulein hyperstimulation on SAPK activity were observed within 5 minutes, the maximum (57-fold) activation was evident after 15 minutes, and levels remained above control for at least 3 hours. In comparison, hyperstimulation with bombesin induced a maximal 5-fold increase of SAPK activity which returned to basal within one hour. These data indicate that SAPK activity is the earliest and best correlated cellular marker associated with secretagogue-induced pancreatitis.

Intracellular activation of digestive zymogens in rat pancreatic acini. Stimulation by high doses of cholecystokinin.

PubMed Central

Leach, S D; Modlin, I M; Scheele, G A; Gorelick, F S

1991-01-01

The mechanism by which digestive zymogens become activated during acute pancreatitis remains poorly understood. Given the ability for cholecystokinin (CCK) to induce pancreatitis in vivo, the effects of high dose CCK on preparations of isolated pancreatic acini were examined. Using an immunologic technique for the detection of zymogen activation, CCK was found to stimulate the conversion of procarboxypeptidase A1 to a 35-kD form having the same net charge and electrophoretic mobility as purified recombinant carboxypeptidase A1. This enhanced conversion was proportional to the dose of CCK (maximal at 100 nM), and time dependent. CCK also produced changes in the electrophoretic mobility of procarboxypeptidase B and chymotrypsinogen 2 immunoreactivity, consistent with activation of these zymogens. These events were detectable only within acinar cell pellets and not in the incubation medium, suggesting an intracellular site of conversion. The conversion of procarboxypeptidase A1 to its active form was inhibited by pretreatment with the weak base chloroquine (40 microM) and the protonophore monensin (10 microM). This conversion was also inhibited by pretreatment with the serine protease inhibitor benzamidine (10 mM) but not the cysteine protease inhibitor E64 (100 microM). The results suggest that high dose CCK stimulates the intracellular activation of digestive zymogens within isolated pancreatic acini. This event appears to require an acidic subcellular compartment and serine protease activity. Images PMID:1985109

Pepsinogen D

PubMed Central

Lee, D.; Ryle, A. P.

1967-01-01

Methods are described for the isolation and purification of pepsinogen D, a minor zymogen occurring to the extent of about 5% of the potential proteolytic activity in neutral extracts of the pig gastric mucosa. The physical and chemical properties of this zymogen indicate that it is very similar to, if not identical with, dephosphopepsinogen. ImagesFig. 3. PMID:4167464

Kinetic Dissection of the Pre-existing Conformational Equilibrium in the Trypsin Fold*

PubMed Central

Vogt, Austin D.; Chakraborty, Pradipta; Di Cera, Enrico

2015-01-01

Structural biology has recently documented the conformational plasticity of the trypsin fold for both the protease and zymogen in terms of a pre-existing equilibrium between closed (E*) and open (E) forms of the active site region. How such plasticity is manifested in solution and affects ligand recognition by the protease and zymogen is poorly understood in quantitative terms. Here we dissect the E*-E equilibrium with stopped-flow kinetics in the presence of excess ligand or macromolecule. Using the clotting protease thrombin and its zymogen precursor prethrombin-2 as relevant models we resolve the relative distribution of the E* and E forms and the underlying kinetic rates for their interconversion. In the case of thrombin, the E* and E forms are distributed in a 1:4 ratio and interconvert on a time scale of 45 ms. In the case of prethrombin-2, the equilibrium is shifted strongly (10:1 ratio) in favor of the closed E* form and unfolds over a faster time scale of 4.5 ms. The distribution of E* and E forms observed for thrombin and prethrombin-2 indicates that zymogen activation is linked to a significant shift in the pre-existing equilibrium between closed and open conformations that facilitates ligand binding to the active site. These findings broaden our mechanistic understanding of how conformational transitions control ligand recognition by thrombin and its zymogen precursor prethrombin-2 and have direct relevance to other members of the trypsin fold. PMID:26216877

[Molecular authentication of Jinyinhua formula granule by using allele-specific PCR].

PubMed

Jiang, Chao; Tu, Li-Chan; Yuan, Yuan; Huang, Lu-Qi; Gao, Wei; Jin, Yan

2017-07-01

Traditional authentication method is hard to identify herb's authenticity of traditional Chinese medicine(TCM) formula granules because they have lost all their morphological characteristics. In this study, a new allele-specific PCR method was established for identifying the authentication of Jinyinhua formula granule (made from Lonicerae Japonicae Flos) based on an SNP site in trnL-trnF fragment. Genomic DNA was successfully extracted from Lonicerae Japonicae Flos and its formula granules by using an improved spin column method and then PCR was performed with the designed primer. Approximately 110 bp specific bands was obtained only in the authentic Lonicerae Japonicae Flos and its formula granules, while no bands were found in fake mixed products. In addition, the PCR product sequence was proved from Lonicerae Japonicae Flos trnL-trnF sequence by using BLAST method. Therefore, DNA molecular authentication method could make up the limitations of character identification method and microscopic identification, and quickly identify herb's authenticity of TCM formula granules, with enormous potential for market supervision and quality control. Copyright© by the Chinese Pharmaceutical Association.

The Zymogen-Enteropeptidase System: A Practical Approach to Study the Regulation of Enzyme Activity by Proteolytic Cleavage

ERIC Educational Resources Information Center

Pizauro, Joao M., Jr.; Ferro, Jesus A.; de Lima, Andrea C. F.; Routman, Karina S.; Portella, Maria Celia

2004-01-01

The present research describes an efficient procedure to obtain high levels of trypsinogen and chymotrypsinogen by using a simple, rapid, and easily reproducible method. The extraction process and the time-course of activation of zymogens can be carried out in a single laboratory period, without sophisticated equipment. The main objective was to…

Immunohistochemical localization of hepatopancreatic phospholipase in gastropods mollusc, Littorina littorea and Buccinum undatum digestive cells

PubMed Central

2011-01-01

Background Among the digestive enzymes, phospholipase A2 (PLA2) hydrolyzes the essential dietary phospholipids in marine fish and shellfish. However, we know little about the organs that produce PLA2, and the ontogeny of the PLA2-cells. Accordingly, accurate localization of PLA2 in marine snails might afford a better understanding permitting the control of the quality and composition of diets and the mode of digestion of lipid food. Results We have previously producted an antiserum reacting specifically with mSDPLA2. It labeled zymogen granules of the hepatopancreatic acinar cells and the secretory materials of certain epithelial cells in the depths of epithelial crypts in the hepatopancreas of snail. To confirm this localization a laser capture microdissection was performed targeting stained cells of hepatopancreas tissue sections. A Western blot analysis revealed a strong signal at the expected size (30 kDa), probably corresponding to the PLA2. Conclusions The present results support the presence of two hepatopancreatic intracellular and extracellular PLA2 in the prosobranchs gastropods molluscs, Littorina littorea and Buccinum undatum and bring insights on their localizations. PMID:22114916

Proteolysis in isolated autophagic vacuoles from the rat pancreas. Effects of chloroquine administration.

PubMed

Yucel-Lindberg, T; Jansson, H; Glaumann, H

1991-01-01

Administration of the antimalaria drug chloroquine increased the number of autophagic vacuoles (AVs) in the rat pancreas. Ultrastructural analysis showed that AVs contained segregated organelles such as mitochondria, zymogen granules, peroxisomes and small portions of cytoplasm. The maximum number of AVs was observed after 3 h of chloroquine treatment. The effect lasted for 12 h and almost disappeared after 16 h. The increase in AVs caused by chloroquine made it possible to isolate them in a discontinuous Metrizamide gradient with high purity. The proteolytic capacity of the AVs isolated after different chloroquine exposure times was measured after prelabeling pancreatic proteins with an injection of L-(1-14C)leucine 16 h before sacrifice. Protein degradation in isolated AVs increased during the first 6 h of chloroquine exposure and then returned to control values 16 h after the administration. In addition, the activities of two lysosomal enzymes, acid phosphatase and cathepsin B, increased in the AV-fractions following chloroquine treatment. It is concluded that the augmented proteolysis in the isolated AVs is due to a combination of increased substrate content and increased proteolytic lysosomal enzyme activities.

Chronic ethanol feeding modulates the synthesis of digestive enzymes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ponnappa, B.C.; Hoek, J.B.; Rubin, E.

1987-05-01

The effects of chronic ethanol feeding on pancreatic protein synthesis were investigated. Protein synthesis was assessed by studying the rate of incorporation of /sup 3/H-leucine into TCA-precipitable proteins in isolated pancreatic acini from rats. Chronic ethanol ingestion increased the rate of pancreatic protein synthesis by 2-4 fold. The onset of the increase in protein synthesis was detectable two days after ethanol feeding, reached a maximum after 7 days and remained unchanged after 4 months on the ethanol-containing diet. The rate of synthesis of individual digestive enzymes was studied by SDS-PAGE on extracts obtained from purified zymogen granules. Ethanol feeding inducedmore » an increase in the rate of synthesis of most of the digestive enzymes; chymotrypsinogen, trypsinogen and an unidentified protein were increased to a greater extent than other digestive enzymes. By contrast, the synthesis of amylase was selectively decreased after ethanol feeding. These results suggest that chronic ethanol ingestion has specific effects on the rate of synthesis of individual digestive enzymes in the exocrine pancreas.« less

Discovery of a highly selective chemical inhibitor of matrix metalloproteinase-9 (MMP-9) that allosterically inhibits zymogen activation.

PubMed

Scannevin, Robert H; Alexander, Richard; Haarlander, Tara Mezzasalma; Burke, Sharon L; Singer, Monica; Huo, Cuifen; Zhang, Yue-Mei; Maguire, Diane; Spurlino, John; Deckman, Ingrid; Carroll, Karen I; Lewandowski, Frank; Devine, Eric; Dzordzorme, Keli; Tounge, Brett; Milligan, Cindy; Bayoumy, Shariff; Williams, Robyn; Schalk-Hihi, Celine; Leonard, Kristi; Jackson, Paul; Todd, Matthew; Kuo, Lawrence C; Rhodes, Kenneth J

2017-10-27

Aberrant activation of matrix metalloproteinases (MMPs) is a common feature of pathological cascades observed in diverse disorders, such as cancer, fibrosis, immune dysregulation, and neurodegenerative diseases. MMP-9, in particular, is highly dynamically regulated in several pathological processes. Development of MMP inhibitors has therefore been an attractive strategy for therapeutic intervention. However, a long history of failed clinical trials has demonstrated that broad-spectrum MMP inhibitors have limited clinical utility, which has spurred the development of inhibitors selective for individual MMPs. Attaining selectivity has been technically challenging because of sequence and structural conservation across the various MMPs. Here, through a biochemical and structural screening paradigm, we have identified JNJ0966, a highly selective compound that inhibited activation of MMP-9 zymogen and subsequent generation of catalytically active enzyme. JNJ0966 had no effect on MMP-1, MMP-2, MMP-3, MMP-9, or MMP-14 catalytic activity and did not inhibit activation of the highly related MMP-2 zymogen. The molecular basis for this activity was characterized as an interaction of JNJ0966 with a structural pocket in proximity to the MMP-9 zymogen cleavage site near Arg-106, which is distinct from the catalytic domain. JNJ0966 was efficacious in reducing disease severity in a mouse experimental autoimmune encephalomyelitis model, demonstrating the viability of this therapeutic approach. This discovery reveals an unprecedented pharmacological approach to MMP inhibition, providing an opportunity to improve selectivity of future clinical drug candidates. Targeting zymogen activation in this manner may also allow for pharmaceutical exploration of other enzymes previously viewed as intractable drug targets. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Identification of the Haloarchaeal Phasin (PhaP) That Functions in Polyhydroxyalkanoate Accumulation and Granule Formation in Haloferax mediterranei

PubMed Central

Cai, Shuangfeng; Cai, Lei; Liu, Hailong; Liu, Xiaoqing; Han, Jing; Zhou, Jian

2012-01-01

The polyhydroxyalkanoate (PHA) granule-associated proteins (PGAPs) are important for PHA synthesis and granule formation, but currently little is known about the haloarchaeal PGAPs. This study focused on the identification and functional analysis of the PGAPs in the haloarchaeon Haloferax mediterranei. These PGAPs were visualized with two-dimensional gel electrophoresis (2-DE) and identified by matrix-assisted laser desorption ionization–tandem time of flight mass spectrometry (MALDI-TOF/TOF MS). The most abundant protein on the granules was identified as a hypothetical protein, designated PhaP. A genome-wide analysis revealed that the phaP gene is located upstream of the previously identified phaEC genes. Through an integrative approach of gene knockout/complementation and fermentation analyses, we demonstrated that this PhaP is involved in PHA accumulation. The ΔphaP mutant was defective in both PHA biosynthesis and cell growth compared to the wild-type strain. Additionally, transmission electron microscopy results indicated that the number of PHA granules in the ΔphaP mutant cells was significantly lower, and in most of the ΔphaP cells only a single large granule was observed. These results demonstrated that the H. mediterranei PhaP was the predominant structure protein (phasin) on the PHA granules involved in PHA accumulation and granule formation. In addition, BLASTp and phylogenetic results indicate that this type of PhaP is exclusively conserved in haloarchaea, implying that it is a representative of the haloarchaeal type PHA phasin. PMID:22247127

Intracisternal granules in the adipokinetic cells of locusts are not degraded and apparently function as supplementary stores of secretory material.

PubMed

Harthoorn, L F; Diederen, J H; Oudejans, R C; Verstegen, M M; Vullings, H G; Van der Horst, D J

2000-01-01

The intracisternal granules in locust adipokinetic cells appear to represent accumulations of secretory material within cisternae of the rough endoplasmic reticulum. An important question is whether these granules are destined for degradation or represent stores of (pro)hormones. Two strategies were used to answer this question. First, cytochemistry was applied to elucidate the properties of intracisternal granules. The endocytic tracers horseradish peroxidase and wheat-germ agglutinin-conjugated horseradish peroxidase were used to facilitate the identification of endocytic, autophagic, and lysosomal organelles, which may be involved in the degradation of intracisternal granules. No intracisternal granules could be found within autophagosomes, and granules fused with endocytic and lysosomal organelles were not observed, nor could tracer be found within the granules. The lysosomal enzyme acid phosphatase was absent from the granules. Second, biochemical analysis of the content of intracisternal granules revealed that these granules contain prohormones as well as hormones. Prohormones were present in relatively higher amounts compared with ordinary secretory granules. Since the intracisternal granules in locust adipokinetic cells are not degraded and contain intact (pro)hormones it is concluded that they function as supplementary stores of secretory material.

[Ultrastructure of granulocytes of bony fishes (orders Salmoniformes, Cypriniformes, Perciformes)].

PubMed

Flerova, E A; Balabanova, L V

2013-01-01

Analysis of data on utrastructure of granulocytes of freshwater and marine bony fish of orders Salmoniformes, Cypriniformes, and Perciformes showed that in all studied species there were revealed two types of granulocytes - neutrophils and eosinophils. The exception was the bluefish Pomatomus saltatrix L. whose pronephros hemopoietic tissue was found to contain one type of the granulocytic line - neutrophils. The identification parameters of granular leukocytes are specific granules filling the cytoplasm. The main form of specific granules in neutrophils of bony fish of various phylogenetic groups is an elongated granule with different distribution of fibrils or a granule that has crystalloid formed from fibrils. The main form of eosinophil granules - large, electron-dense, homogenous.

Enzymatic aspects in ENT cancer-Matrix metalloproteinases

PubMed Central

Zamfir Chiru, AA; Popescu, CR; Gheorghe, DC

2014-01-01

Abstract The study of ENT cancer allows the implementation of molecular biology methods in diagnosis, predicting the evolution of the disease and suggesting a certain treatment. MMPs are proteolytic enzymes, zinc dependent endopeptidases, secreted by tissues and proinflammatory cells that play a role in the clearance of cell surface receptors. They are expressed as zymogens (inactive forms). Proteolytic enzymes cleave zymogens generating active forms. They are involved in cell proliferation, adhesion, differentiation, migration, angiogenesis, apoptosis and host defense. PMID:25408759

Structure and dynamics of zymogen human blood coagulation factor X.

PubMed

Venkateswarlu, Divi; Perera, Lalith; Darden, Tom; Pedersen, Lee G

2002-03-01

The solution structure and dynamics of the human coagulation factor X (FX) have been investigated to understand the key structural elements in the zymogenic form that participates in the activation process. The model was constructed based on the 2.3-A-resolution x-ray crystallographic structure of active-site inhibited human FXa (PDB:1XKA). The missing gamma-carboxyglutamic acid (GLA) and part of epidermal growth factor 1 (EGF1) domains of the light chain were modeled based on the template of GLA-EGF1 domains of the tissue factor (TF)-bound FVIIa structure (PDB:1DAN). The activation peptide and other missing segments of FX were introduced using homology modeling. The full calcium-bound model of FX was subjected to 6.2 ns of molecular dynamics simulation in aqueous medium using the AMBER6.0 package. We observed significant reorientation of the serine-protease (SP) domain upon activation leading to a compact multi-domain structure. The solution structure of zymogen appears to be in a well-extended conformation with the distance between the calcium ions in the GLA domain and the catalytic residues estimated to be approximately 95 A in contrast to approximately 83 A in the activated form. The latter is in close agreement with fluorescence studies on FXa. The S1-specificity residues near the catalytic triad show significant differences between the zymogen and activated structures.

Novel insights into RNP granules by employing the trypanosome's microtubule skeleton as a molecular sieve.

PubMed

Fritz, Melanie; Vanselow, Jens; Sauer, Nadja; Lamer, Stephanie; Goos, Carina; Siegel, T Nicolai; Subota, Ines; Schlosser, Andreas; Carrington, Mark; Kramer, Susanne

2015-09-18

RNP granules are ribonucleoprotein assemblies that regulate the post-transcriptional fate of mRNAs in all eukaryotes. Their exact function remains poorly understood, one reason for this is that RNP granule purification has not yet been achieved. We have exploited a unique feature of trypanosomes to prepare a cellular fraction highly enriched in starvation stress granules. First, granules remain trapped within the cage-like, subpellicular microtubule array of the trypanosome cytoskeleton while soluble proteins are washed away. Second, the microtubules are depolymerized and the granules are released. RNA sequencing combined with single molecule mRNA FISH identified the short and highly abundant mRNAs encoding ribosomal mRNAs as being excluded from granules. By mass spectrometry we have identified 463 stress granule candidate proteins. For 17/49 proteins tested by eYFP tagging we have confirmed the localization to granules, including one phosphatase, one methyltransferase and two proteins with a function in trypanosome life-cycle regulation. The novel method presented here enables the unbiased identification of novel RNP granule components, paving the way towards an understanding of RNP granule function. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

A novel protein kinase D inhibitor attenuates early events of experimental pancreatitis in isolated rat acini.

PubMed

Thrower, Edwin C; Yuan, Jingzhen; Usmani, Ashar; Liu, Yannan; Jones, Courtney; Minervini, Samantha N; Alexandre, Martine; Pandol, Stephen J; Guha, Sushovan

2011-01-01

Novel protein kinase C isoforms (PKC δ and ε) mediate early events in acute pancreatitis. Protein kinase D (PKD/PKD1) is a convergent point of PKC δ and ε in the signaling pathways triggered through CCK or cholinergic receptors and has been shown to activate the transcription factor NF-κB in acute pancreatitis. For the present study we hypothesized that a newly developed PKD/PKD1 inhibitor, CRT0066101, would prevent the initial events leading to pancreatitis. We pretreated isolated rat pancreatic acinar cells with CRT0066101 and a commercially available inhibitor Gö6976 (10 μM). This was followed by stimulation for 60 min with high concentrations of cholecystokinin (CCK, 0.1 μM), carbachol (CCh, 1 mM), or bombesin (10 μM) to induce initial events of pancreatitis. PKD/PKD1 phosphorylation and activity were measured as well as zymogen activation, amylase secretion, cell injury and NF-κB activation. CRT0066101 dose dependently inhibited secretagogue-induced PKD/PKD1 activation and autophosphorylation at Ser-916 with an IC(50) ∼3.75-5 μM but had no effect on PKC-dependent phosphorylation of the PKD/PKD1 activation loop (Ser-744/748). Furthermore, CRT0066101 reduced secretagogue-induced zymogen activation and amylase secretion. Gö6976 reduced zymogen activation but not amylase secretion. Neither inhibitor affected basal zymogen activation or secretion. CRT0066101 did not affect secretagogue-induced cell injury or changes in cell morphology, but it reduced NF-κB activation by 75% of maximal for CCK- and CCh-stimulated acinar cells. In conclusion, CRT0066101 is a potent and specific PKD family inhibitor. Furthermore, PKD/PKD1 is a potential mediator of zymogen activation, amylase secretion, and NF-κB activation induced by a range of secretagogues in pancreatic acinar cells.

Requirements for the identification of dense-core granules.

PubMed

Meldolesi, Jacopo; Chieregatti, Evelina; Luisa Malosio, Maria

2004-01-01

Dense-core granules (DCGs), cytoplasmic organelles competent for regulated exocytosis, show considerable heterogeneity depending upon the specificity of their expressing cells--primarily neurons and neurosecretory cells. DCGs have been mainly identified by detecting their cargo molecules, often members of the granin family, and using conventional electron microscopy and immunocytochemistry. However, by a critical analysis of the various stages of DCG "life" within neurosecretory cells, we have highlighted several specific molecular and functional properties that are common to all these organelles. We propose that these properties be considered as strict requirements for the identification of DCGs.

Zinc toxicosis in a free-flying trumpeter swan (Cygnus buccinator)

USGS Publications Warehouse

Carpenter, J.W.; Andrews, G.A.; Beyer, W.N.

2004-01-01

A trumpeter swan (Cygnus buccinator) was observed near it mill pond in Picher, Oklahoma. USA. It became weakened and emaciated after about 1 mo, was captured with little resistance, and taken into captivity for medical care. Serum chemistry results were consistent with hepatic, renal, and muscular damage. Serum zinc concentration was elevated at 11.2 parts per million (ppm). The swan was treated for suspected heavy-metal poisoning, but died overnight. Gross postmortem findings were emaciation and pectoral muscle atrophy. Histopathologic lesions in the pancreas included mild diffuse disruption of acinar architecture, severe diffuse depletion or absence of zymogen granules, occasional apoptotic bodies ics in acinar epithelial cells, and mild interstitial and capsular fibrosis. Zinc concentration in pancreas was 3,200 ppm wet weight, and was similar to that reported in the pancreases of waterfowl known to be killed by zinc toxicity. Zinc concentrations in liver (154 ppm) and kidneys (145 ppm) also were elevated. Acute tubular necrosis of the collecting tubules of the kidneys was also possibly due to zinc toxicity. To the authors' knowledge, this is the first confirmed case of zinc poisoning in a trumpeter swan associated with mining wastes..

Biochemical, transcriptomic and proteomic analyses of digestion in the scorpion Tityus serrulatus: insights into function and evolution of digestion in an ancient arthropod.

PubMed

Fuzita, Felipe J; Pinkse, Martijn W H; Patane, José S L; Juliano, Maria A; Verhaert, Peter D E M; Lopes, Adriana R

2015-01-01

Scorpions are among the oldest terrestrial arthropods and they have passed through small morphological changes during their evolutionary history on land. They are efficient predators capable of capturing and consuming large preys and due to envenomation these animals can become a human health challenge. Understanding the physiology of scorpions can not only lead to evolutionary insights but also is a crucial step in the development of control strategies. However, the digestive process in scorpions has been scarcely studied. In this work, we describe the combinatory use of next generation sequencing, proteomic analysis and biochemical assays in order to investigate the digestive process in the yellow scorpion Tityus serrulatus, mainly focusing in the initial protein digestion. The transcriptome generated database allowed the quantitative identification by mass spectrometry of different enzymes and proteins involved in digestion. All the results suggested that cysteine cathepsins play an important role in protein digestion. Two digestive cysteine cathepsins were isolated and characterized presenting acidic characteristics (pH optima and stability), zymogen conversion to the mature form after acidic activation and a cross-class inhibition by pepstatin. A more elucidative picture of the molecular mechanism of digestion in a scorpion was proposed based on our results from Tityus serrulatus. The midgut and midgut glands (MMG) are composed by secretory and digestive cells. In fasting animals, the secretory granules are ready for the next predation event, containing enzymes needed for alkaline extra-oral digestion which will compose the digestive fluid, such as trypsins, astacins and chitinase. The digestive vacuoles are filled with an acidic proteolytic cocktail to the intracellular digestion composed by cathepsins L, B, F, D and legumain. Other proteins as lipases, carbohydrases, ctenitoxins and a chitolectin with a perithrophin domain were also detected. Evolutionarily, a large gene duplication of cathepsin L occurred in Arachnida with the sequences from ticks being completely divergent from other arachnids probably due to the particular selective pressures over this group.

Identification of human cysteine-rich secretory protein 3 (CRISP-3) as a matrix protein in a subset of peroxidase-negative granules of neutrophils and in the granules of eosinophils.

PubMed

Udby, Lene; Calafat, Jero; Sørensen, Ole E; Borregaard, Niels; Kjeldsen, Lars

2002-09-01

Cysteine-rich secretory protein 3 (CRISP-3; also known as SGP28) was originally discovered in human neutrophilic granulocytes. We have recently developed a sensitive sandwich enzyme-linked immunosorbent assay for CRISP-3 and demonstrated the presence of CRISP-3 in exocrine secretions. To investigate the subcellular localization and mobilization of CRISP-3 in human neutrophils, we performed subcellular fractionation of resting and activated neutrophils on three-layer Percoll density gradients, release-studies of granule proteins in response to different secretagogues, and double-labeling immunogold electron microscopy. CRISP-3 was found to be localized in a subset of granules with overlapping characteristics of specific and gelatinase granules and mobilized accordingly, thus confirming the hypothesis that peroxidase-negative granules exist as a continuum from specific to gelatinase granules regarding protein content and mobilization. CRISP-3 was found to be a matrix protein, which is stored in granules as glycosylated and as unglycosylated protein. The subcellular distribution of the two forms of CRISP-3 was identical. In addition, CRISP-3 was found as a granule protein in eosinophilic granulocytes. The presence of CRISP-3 in peroxidase-negative granules of neutrophils, in granules of eosinophils, and in exocrine secretions indicates a role in the innate host defense.

Proteome Profile of Starch Granules Purified from Rice (Oryza sativa) Endosperm.

PubMed

Xing, Shihai; Meng, Xiaoxi; Zhou, Lihui; Mujahid, Hana; Zhao, Chunfang; Zhang, Yadong; Wang, Cailin; Peng, Zhaohua

2016-01-01

Starch is the most important food energy source in cereals. Many of the known enzymes involved in starch biosynthesis are partially or entirely granule-associated in the endosperm. Studying the proteome of rice starch granules is critical for us to further understand the mechanisms underlying starch biosynthesis and packaging of starch granules in rice amyloplasts, consequently for the improvement of rice grain quality. In this article, we developed a protocol to purify starch granules from mature rice endosperm and verified the quality of purified starch granules by microscopy observations, I2 staining, and Western blot analyses. In addition, we found the phenol extraction method was superior to Tris-HCl buffer extraction method with respect to the efficiency in recovery of starch granule associated proteins. LC-MS/MS analysis showed identification of already known starch granule associated proteins with high confidence. Several proteins reported to be involved in starch synthesis in prior genetic studies in plants were also shown to be enriched with starch granules, either directly or indirectly, in our studies. In addition, our results suggested that a few additional candidate proteins may also be involved in starch synthesis. Furthermore, our results indicated that some starch synthesis pathway proteins are subject to protein acetylation modification. GO analysis and KEGG pathway enrichment analysis showed that the identified proteins were mainly located in plastids and involved in carbohydrate metabolism. This study substantially advances the understanding of the starch granule associated proteome in rice and post translational regulation of some starch granule associated proteins.

Freshwater catfish jundiá (Rhamdia quelen) larvae are prepared to digest inert feed at the exogenous feeding onset: physiological and histological assessments.

PubMed

Silveira, Jenniffer; Silva, Carlos Peres; Cargnin-Ferreira, Eduardo; Alexandre, Daniel; Elias, Mariele Abádia; Fracalossi, Débora Machado

2013-12-01

This study assessed the morphological development of jundiá larvae's digestive system and digestive proteolytic activity. Specific serine proteinases activities varied over time, with the highest peak at 12 h after hatching (AH), which corresponded to 296.38 ± 84.20 mU mg⁻¹ for trypsin and 315.45 ± 42.16 mU mg⁻¹ for chymotrypsin. Specific aspartic proteinases activities increased up to the start of weaning, oscillated during that phase, but showed a consistent increase after that, resulting in the highest specific activity at 252 h AH (7.88 ± 0.68 mU mg⁻¹). Gel assays showed different molecular forms, especially of serine proteinases. Histology showed the gastrointestinal tract development onset at 0 h AH and open mouth at 4 h AH. At 16 h AH, the following differentiation of the digestive tract was evident: oropharyngeal cavity, esophagus, liver, pancreas, stomach, and intestine. At 40 h AH, zymogen granules in the pancreas were observed, and at 48 h AH, mucus in the digestive tract and gastric glands in the stomach. Findings indicate that jundiá has a functional stomach before the end of vitelline reserves. Therefore, jundiá larvae are probably capable to digest inert feed at the exogenous feeding onset.

Outbreak of infectious pancreatic necrosis virus (IPNV) in farmed rainbow trout in China.

PubMed

Zhu, Ling; Wang, Xingli; Wang, Kaiyu; Yang, Qian; He, Jie; Qin, Zhenyang; Geng, Yi; Ouyang, Ping; Huang, Xiaoli

2017-06-01

Infectious pancreatic necrosis virus (IPNV) is a member of the Aquabirnavirus genus, which caused mass mortality (nearly 100%) in farmed rainbow trout (Oncorhynchus mykiss) in aquaculture farms in 2016, China. Major clinical signs included decreased appetite, mucous-like stools, and darkened pigmentation. Pathological changes in moribund fish were observed, such as marked vacuolar degeneration of the pancreatic cells with pyknotic nucleus and decreased zymogen granules, severe hemorrhage in the liver, and tumidness of respiratory epithelium in gills. In addition, the tissue fluid of diseased fish could produce a cytopathic effect (CPE) in RTG-2 cells. The presence of specific 206bp fragments by the reverse transcription-polymerase chain reaction (RT-PCR) using tissue homogenate of diseased fish and supernatant of infected cells revealed that IPNV could be confirmed. The pathogenicity test of cell culture supernatant detected cumulative mortality of 80%, and the clinical symptoms observed in the moribund and dead fish were similar to the naturally infected fish. Furthermore, the sequence analysis of VP2 gene showed that the isolated virus strain belonged to genogroup 1, and 97% homology with the Mexican IPNV isolate was found. To our knowledge, this is the first report on IPNV natural infection in the southwest of China. Copyright © 2017 Elsevier B.V. All rights reserved.

Extensive Basal Level Activation of Complement Mannose-Binding Lectin-Associated Serine Protease-3: Kinetic Modeling of Lectin Pathway Activation Provides Possible Mechanism.

PubMed

Oroszlán, Gábor; Dani, Ráhel; Szilágyi, András; Závodszky, Péter; Thiel, Steffen; Gál, Péter; Dobó, József

2017-01-01

Serine proteases (SPs) are typically synthesized as precursors, termed proenzymes or zymogens, and the fully active form is produced via limited proteolysis by another protease or by autoactivation. The lectin pathway of the complement system is initiated by mannose-binding lectin (MBL)-associated SPs (MASP)-1, and MASP-2, which are known to be present as proenzymes in blood. The third SP of the lectin pathway, MASP-3, was recently shown to be the major activator, and the exclusive "resting blood" activator of profactor D, producing factor D, the initiator protease of the alternative pathway. Because only activated MASP-3 is capable of carrying out this cleavage, it was presumed that a significant fraction of MASP-3 must be present in the active form in resting blood. Here, we aimed to detect active MASP-3 in the blood by a more direct technique and to quantitate the active to zymogen ratio. First, MASPs were partially purified (enriched) from human plasma samples by affinity chromatography using immobilized MBL in the presence of inhibitors. Using this MASP pool, only the zymogen form of MASP-1 was detected by Western blot, whereas over 70% MASP-3 was in an activated form in the same samples. Furthermore, the active to zymogen ratio of MASP-3 showed little individual variation. It is enigmatic how MASP-3, which is not able to autoactivate, is present mostly as an active enzyme, whereas MASP-1, which has a potent autoactivation capability, is predominantly proenzymic in resting blood. In an attempt to explain this phenomenon, we modeled the basal level fluid-phase activation of lectin pathway proteases and their subsequent inactivation by C1 inhibitor and antithrombin using available and newly determined kinetic constants. The model can explain extensive MASP-3 activation only if we assume efficient intracomplex activation of MASP-3 by zymogen MASP-1. On the other hand, the model is in good agreement with the fact that MASP-1 and -2 are predominantly proenzymic and some of them is present in the form of inactive serpin-protease complexes. As an alternative hypothesis, MASP-3 activation by proprotein convertases is also discussed.

Proteome Profile of Starch Granules Purified from Rice (Oryza sativa) Endosperm

PubMed Central

Xing, Shihai; Meng, Xiaoxi; Zhou, Lihui; Mujahid, Hana; Zhao, Chunfang; Zhang, Yadong; Wang, Cailin; Peng, Zhaohua

2016-01-01

Starch is the most important food energy source in cereals. Many of the known enzymes involved in starch biosynthesis are partially or entirely granule-associated in the endosperm. Studying the proteome of rice starch granules is critical for us to further understand the mechanisms underlying starch biosynthesis and packaging of starch granules in rice amyloplasts, consequently for the improvement of rice grain quality. In this article, we developed a protocol to purify starch granules from mature rice endosperm and verified the quality of purified starch granules by microscopy observations, I2 staining, and Western blot analyses. In addition, we found the phenol extraction method was superior to Tris-HCl buffer extraction method with respect to the efficiency in recovery of starch granule associated proteins. LC-MS/MS analysis showed identification of already known starch granule associated proteins with high confidence. Several proteins reported to be involved in starch synthesis in prior genetic studies in plants were also shown to be enriched with starch granules, either directly or indirectly, in our studies. In addition, our results suggested that a few additional candidate proteins may also be involved in starch synthesis. Furthermore, our results indicated that some starch synthesis pathway proteins are subject to protein acetylation modification. GO analysis and KEGG pathway enrichment analysis showed that the identified proteins were mainly located in plastids and involved in carbohydrate metabolism. This study substantially advances the understanding of the starch granule associated proteome in rice and post translational regulation of some starch granule associated proteins. PMID:27992503

Identification of proteins associated with polyhydroxybutyrate granules from Herbaspirillum seropedicae SmR1--old partners, new players.

PubMed

Tirapelle, Evandro F; Müller-Santos, Marcelo; Tadra-Sfeir, Michelle Z; Kadowaki, Marco A S; Steffens, Maria B R; Monteiro, Rose A; Souza, Emanuel M; Pedrosa, Fabio O; Chubatsu, Leda S

2013-01-01

Herbaspirillum seropedicae is a diazotrophic ß-Proteobacterium found associated with important agricultural crops. This bacterium produces polyhydroxybutyrate (PHB), an aliphatic polyester, as a carbon storage and/or source of reducing equivalents. The PHB polymer is stored as intracellular insoluble granules coated mainly with proteins, some of which are directly involved in PHB synthesis, degradation and granule biogenesis. In this work, we have extracted the PHB granules from H. seropedicae and identified their associated-proteins by mass spectrometry. This analysis allowed us to identify the main phasin (PhaP1) coating the PHB granule as well as the PHB synthase (PhbC1) responsible for its synthesis. A phbC1 mutant is impaired in PHB synthesis, confirming its role in H. seropedicae. On the other hand, a phaP1 mutant produces PHB granules but coated mainly with the secondary phasin (PhaP2). Furthermore, some novel proteins not previously described to be involved with PHB metabolism were also identified, bringing new possibilities to PHB function in H. seropedicae.

Identification of Proteins Associated with Polyhydroxybutyrate Granules from Herbaspirillum seropedicae SmR1 - Old Partners, New Players

PubMed Central

Tirapelle, Evandro F.; Müller-Santos, Marcelo; Tadra-Sfeir, Michelle Z.; Kadowaki, Marco A. S.; Steffens, Maria B. R.; Monteiro, Rose A.; Souza, Emanuel M.; Pedrosa, Fabio O.; Chubatsu, Leda S.

2013-01-01

Herbaspirillum seropedicae is a diazotrophic ß-Proteobacterium found associated with important agricultural crops. This bacterium produces polyhydroxybutyrate (PHB), an aliphatic polyester, as a carbon storage and/or source of reducing equivalents. The PHB polymer is stored as intracellular insoluble granules coated mainly with proteins, some of which are directly involved in PHB synthesis, degradation and granule biogenesis. In this work, we have extracted the PHB granules from H. seropedicae and identified their associated-proteins by mass spectrometry. This analysis allowed us to identify the main phasin (PhaP1) coating the PHB granule as well as the PHB synthase (PhbC1) responsible for its synthesis. A phbC1 mutant is impaired in PHB synthesis, confirming its role in H. seropedicae. On the other hand, a phaP1 mutant produces PHB granules but coated mainly with the secondary phasin (PhaP2). Furthermore, some novel proteins not previously described to be involved with PHB metabolism were also identified, bringing new possibilities to PHB function in H. seropedicae. PMID:24086439

Identification of a multifunctional protein, PhaM, that determines number, surface to volume ratio, subcellular localization and distribution to daughter cells of poly(3-hydroxybutyrate), PHB, granules in Ralstonia eutropha H16.

PubMed

Pfeiffer, Daniel; Wahl, Andreas; Jendrossek, Dieter

2011-11-01

A two-hybrid approach was applied to screen for proteins with the ability to interact with PHB synthase (PhaC1) of Ralstonia eutropha. The H16_A0141 gene (phaM) was identified in the majority of positive clones. PhaM (26.6 kDa) strongly interacted with PhaC1 and with phasin PhaP5 but not with PhaP1 or other PHB granule-associated proteins. A ΔphaM mutant accumulated only one or two large PHB granules instead of three to six medium-sized PHB granules of the wild type, and distribution of granules to daughter cells was disordered. All three phenotypes (number, size and distribution of PHB granules) were reversed by reintroduction of phaM. Purified PhaM revealed DNA-binding properties in gel mobility shift experiments. Expression of a fusion of the yellow fluorescent protein (eYfp) with PhaM resulted in formation of many small fluorescent granules that were bound to the nucleoid region. Remarkably, an eYfp-PhaP5 fusion localized at the cell poles in a PHB-negative background and overexpression of eYfp-PhaP5 in the wild type conferred binding of PHB granules to the cell poles. In conclusion, subcellular localization of PHB granules in R. eutropha depends on a concerted expression of at least three PHB granule-associated proteins, namely PhaM, PhaP5 and PHB synthase PhaC1. © 2011 Blackwell Publishing Ltd.

Ultrastructural identification of Langerhans cells in normal swine epidermis.

PubMed Central

Romano, J; Balaguer, L

1991-01-01

Langerhans cells of the epidermis of 6-month-old white crossbred farm pigs were identified by electron microscopy. Ultrastructurally they were similar to those described in other mammals. They were present in basal and suprabasal layers and were characterised by a lobulated nucleus and an electrolucent cytoplasm with occasional dendritic processes, and the absence of tonofilaments and specialised unions with surrounding keratinocytes. They were specifically identified by the presence of characteristic rod or racquet-shaped intracytoplasmic granules. Intraepidermal clear cells without specific granules were present, although no melanocytes were observed. This is the first report of the presence of Birbeck granules in porcine Langerhans cells. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:1817140

The three-dimensional structure of the native ternary complex of bovine pancreatic procarboxypeptidase A with proproteinase E and chymotrypsinogen C.

PubMed Central

Gomis-Rüth, F X; Gómez, M; Bode, W; Huber, R; Avilés, F X

1995-01-01

The metalloexozymogen procarboxypeptidase A is mainly secreted in ruminants as a ternary complex with zymogens of two serine endoproteinases, chymotrypsinogen C and proproteinase E. The bovine complex has been crystallized, and its molecular structure analysed and refined at 2.6 A resolution to an R factor of 0.198. In this heterotrimer, the activation segment of procarboxypeptidase A essentially clamps the other two subunits, which shield the activation sites of the former from tryptic attack. In contrast, the propeptides of both serine proproteinases are freely accessible to trypsin. This arrangement explains the sequential and delayed activation of the constituent zymogens. Procarboxypeptidase A is virtually identical to the homologous monomeric porcine form. Chymotrypsinogen C displays structural features characteristic for chymotrypsins as well as elastases, except for its activation domain; similar to bovine chymotrypsinogen A, its binding site is not properly formed, while its surface located activation segment is disordered. The proproteinase E structure is fully ordered and strikingly similar to active porcine elastase; its specificity pocket is occluded, while the activation segment is fixed to the molecular surface. This first structure of a native zymogen from the proteinase E/elastase family does not fundamentally differ from the serine proproteinases known so far. Images PMID:7556081

Highly Conserved Arg Residue of ERFNIN Motif of Pro-Domain is Important for pH-Induced Zymogen Activation Process in Cysteine Cathepsins K and L.

PubMed

Aich, Pulakesh; Biswas, Sampa

2018-06-01

Pro-domain of a cysteine cathepsin contains a highly conserved Ex 2 Rx 2 Fx 2 Nx 3 Ix 3 N (ERFNIN) motif. The zymogen structure of cathepsins revealed that the Arg(R) residue of the motif is a central residue of a salt-bridge/H-bond network, stabilizing the scaffold of the pro-domain. Importance of the arginine is also demonstrated in studies where a single mutation (Arg → Trp) in human lysosomal cathepsin K (hCTSK) is linked to a bone-related genetic disorder "Pycnodysostosis". In the present study, we have characterized in vitro Arg → Trp mutant of hCTSK and the same mutant of hCTSL. The R → W mutant of hCTSK revealed that this mutation leads to an unstable zymogen that is spontaneously activated and auto-proteolytically degraded rapidly. In contrast, the same mutant of hCTSL is sufficiently stable and has proteolytic activity almost like its wild-type counterpart; however it shows an altered zymogen activation condition in terms of pH, temperature and time. Far and near UV circular dichroism and intrinsic tryptophan fluorescence experiments have revealed that the mutation has minimal effect on structure of the protease hCTSL. Molecular modeling studies shows that the mutated Trp31 in hCTSL forms an aromatic cluster with Tyr23 and Trp30 leading to a local stabilization of pro-domain and supplements the loss of salt-bridge interaction mediated by Arg31 in wild-type. In hCTSK-R31W mutant, due to presence of a non-aromatic Ser30 residue such interaction is not possible and may be responsible for local instability. These differences may cause detrimental effects of R31W mutation on the regulation of hCTSK auto-activation process compared to altered activation process in hCTSL.

Screening and identification of α-glucosidase inhibitors from Shenqi Jiangtang Granule by ultrafiltration liquid chromatography and mass spectrometry.

PubMed

Zhang, Hui; Zhang, Xiaojing; Jiang, Huijie; Xu, Cong; Tong, Shengqiang; Yan, Jizhong

2018-02-01

Shenqi Jiangtang Granule, a well-known traditional Chinese herbal preparation, has been widely used for the treatment of type II diabetes mellitus. In this work, an ultrafiltration liquid chromatography with quadrupole time-of-flight mass spectrometry method was proposed for the rapid identification of bioactive ingredients from Shenqi Jiangtang Granule using α-glucosidase as an example. First, the chemical profile of this preparation was clarified, including 37 saponins, 17 flavonoids, 37 lignans, and seven other compounds. After incubation with α-glucosidase in vitro, the methanol extract with an IC 50 value of 0.19 mg/mL exhibited significant inhibitory activity. Then, 18 specific binding peaks were screened, and 15 peaks were identified. Among these, ten compounds were reported to have potential α-glucosidase inhibitory activity for the first time. Subsequently, the inhibitory activities of these active compounds were evaluated by ultraviolet spectrophotometry with p-nitrophenyl α-d-glucopyranoside as a substrate. As a result, gomisin J and gomisin D exhibited stronger α-glucosidase inhibitory activities than other active compounds with IC 50 values of 77.69 and 133.85 μM, respectively. The results demonstrated that the integrated ultrafiltration liquid chromatography with mass spectrometry method was an effective and powerful tool for the discovery of active ingredients in Shenqi Jiangtang Granule. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Identification of Genes Encoding Granule-Bound Starch Synthase Involved in Amylose Metabolism in Banana Fruit

PubMed Central

Liu, Weixin; Xu, Biyu; Jin, Zhiqiang

2014-01-01

Granule-bound starch synthase (GBSS) is responsible for amylose synthesis, but the role of GBSS genes and their encoded proteins remains poorly understood in banana. In this study, amylose content and GBSS activity gradually increased during development of the banana fruit, and decreased during storage of the mature fruit. GBSS protein in banana starch granules was approximately 55.0 kDa. The protein was up-regulated expression during development while it was down-regulated expression during storage. Six genes, designated as MaGBSSI-1, MaGBSSI-2, MaGBSSI-3, MaGBSSI-4, MaGBSSII-1, and MaGBSSII-2, were cloned and characterized from banana fruit. Among the six genes, the expression pattern of MaGBSSI-3 was the most consistent with the changes in amylose content, GBSS enzyme activity, GBSS protein levels, and the quantity or size of starch granules in banana fruit. These results suggest that MaGBSSI-3 might regulate amylose metabolism by affecting the variation of GBSS levels and the quantity or size of starch granules in banana fruit during development or storage. PMID:24505384

Biochemical, Transcriptomic and Proteomic Analyses of Digestion in the Scorpion Tityus serrulatus: Insights into Function and Evolution of Digestion in an Ancient Arthropod

PubMed Central

Fuzita, Felipe J.; Pinkse, Martijn W. H.; Patane, José S. L.; Juliano, Maria A.; Verhaert, Peter D. E. M.; Lopes, Adriana R.

2015-01-01

Scorpions are among the oldest terrestrial arthropods and they have passed through small morphological changes during their evolutionary history on land. They are efficient predators capable of capturing and consuming large preys and due to envenomation these animals can become a human health challenge. Understanding the physiology of scorpions can not only lead to evolutionary insights but also is a crucial step in the development of control strategies. However, the digestive process in scorpions has been scarcely studied. In this work, we describe the combinatory use of next generation sequencing, proteomic analysis and biochemical assays in order to investigate the digestive process in the yellow scorpion Tityus serrulatus, mainly focusing in the initial protein digestion. The transcriptome generated database allowed the quantitative identification by mass spectrometry of different enzymes and proteins involved in digestion. All the results suggested that cysteine cathepsins play an important role in protein digestion. Two digestive cysteine cathepsins were isolated and characterized presenting acidic characteristics (pH optima and stability), zymogen conversion to the mature form after acidic activation and a cross-class inhibition by pepstatin. A more elucidative picture of the molecular mechanism of digestion in a scorpion was proposed based on our results from Tityus serrulatus. The midgut and midgut glands (MMG) are composed by secretory and digestive cells. In fasting animals, the secretory granules are ready for the next predation event, containing enzymes needed for alkaline extra-oral digestion which will compose the digestive fluid, such as trypsins, astacins and chitinase. The digestive vacuoles are filled with an acidic proteolytic cocktail to the intracellular digestion composed by cathepsins L, B, F, D and legumain. Other proteins as lipases, carbohydrases, ctenitoxins and a chitolectin with a perithrophin domain were also detected. Evolutionarily, a large gene duplication of cathepsin L occurred in Arachnida with the sequences from ticks being completely divergent from other arachnids probably due to the particular selective pressures over this group. PMID:25875018

First record of the Granulate Ambrosia Beetle, Xylosandrus crassiusculus (Coleoptera: Curculionidae, Scolytinae), in the Iberian Peninsula.

PubMed

Gallego, Diego; Lencina, José Luis; Mas, Hugo; Ceveró, Julia; Faccoli, Massimo

2017-06-06

The Granulate Ambrosia Beetle Xylosandrus crassiusculus, an alien species of Asian origin, was recorded for first time in the Iberian Peninsula. Many specimens were collected in October 2016 in the Valencia region (Spain) from infested carob trees. The species is included in the EPPO Alert List as causing serious damage in many Mediterranean regions. A key for the morphological identification of the Xylosandrus species occurring in Europe is also reported.

Caerulin-induced pancreatitis in rats: Histological and genetic expression changes from acute phase to recuperation

PubMed Central

Magaña-Gómez, Javier; López-Cervantes, Guillermo; de la Barca, Ana María Calderón

2006-01-01

AIM: To study the histological and pancreatitis-associated protein mRNA accumulation changes of pancreas from acute phase of caerulin-induced pancreatitis to recuperation in rats. METHODS: Acute pancreatitis was induced by caerulein in male Wistar rats and followed up for 90 d by histological and mRNA analyses of pancreas. Pancreases were dissected at 0, 9, 24 h and 3, 5, 15, 30, 60, 90 d post-induction. Edema (E), polymorphonuclear neutrophil (PMN) infiltration, cytoplasmic vacuolization (V), zymogen granule depletion (ZD) and acinar disorganization (AD) were microscopically evaluated. Accumulation of pancreatitis-associated protein (PAP) and L13A mRNAs were quantified by real-time PCR. RESULTS: The main histological changes appeared at 9 h post-induction for PMN infiltration and cytoplasmic V, while at 24 h and 3 d for E and ZD, respectively. All the parameters were recovered after 5 d, except for ZD which delayed more than 30 d. The main AD was observed after 15 d and values returned to normal after 30 d. Similarly to histological changes, accumulation of the PAP mRNA was increased at 9 h with the highest accumulation at 24 h and differences disappeared after 5 d. CONCLUSION: From the acute phase to recuperation of pancreatitis, regeneration and re-differentiation of pancreas occur and PAP expression is exclusively an acute response of pancreatitis. PMID:16810747

Extensive molecular analysis suggested the strong genetic heterogeneity of idiopathic chronic pancreatitis.

PubMed

Sofia, Valentina Maria; Da Sacco, Letizia; Surace, Cecilia; Tomaiuolo, Anna Cristina; Genovese, Silvia; Grotta, Simona; Gnazzo, Maria; Petrocchi, Stefano; Ciocca, Laura; Alghisi, Federico; Montemitro, Enza; Martemucci, Luigi; Elce, Ausilia; Lucidi, Vincenzina; Castaldo, Giuseppe; Angioni, Adriano

2016-05-26

Genetic features of Chronic Pancreatitis (CP) have been extensively investigated mainly testing genes associated to the trypsinogen activation pathway. However, different molecular pathways involving other genes may be implicated in CP pathogenesis. 80 patients with Idiopathic CP were investigated using Next Generation Sequencing approach with a panel of 70 genes related to six different pancreatic pathways: premature activation of trypsinogen; modifier genes of Cystic Fibrosis phenotype; pancreatic secretion and ion homeostasis; Calcium signalling and zymogen granules exocytosis; autophagy; autoimmune pancreatitis related genes. We detected mutations in 34 out of 70 genes examined; 64/80 patients (80.0%) were positive for mutations in one or more genes, 16/80 patients (20.0%) had no mutations. Mutations in CFTR were detected in 32/80 patients (40.0%) and 22 of them exhibited at least one mutation in genes of other pancreatic pathways. Of the remaining 48 patients, 13/80 (16.3%) had mutations in genes involved in premature activation of trypsinogen and 19/80 (23.8%) had mutations only in genes of the other pathways: 38/64 patients positive for mutations showed variants in two or more genes (59.3%). Our data, although to be extended with functional analysis of novel mutations, suggest a high rate of genetic heterogeneity in chronic pancreatitis and that trans-heterozygosity may predispose to the idiopathic CP phenotype.

Adenomatous ductal proliferation/hyperplasia in the parotid gland associated without any other pathological lesions; a report and survey of the literatures.

PubMed

Adhikari, Bhoj Raj; Nishimura, Michiko; Takimoto, Kosuke; Harada, Fumiya; Onishi, Aya; Hiraki, Daichi; Paudel, Durga; Neopane, Puja; Utsunomiya, Masafumi; Morikawa, Tetsuro; Yoshida, Koki; Sato, Jun; Shakya, Mamata; Nakayama, Eiji; Nagayasu, Hiroki; Abiko, Yoshihiro

2018-05-15

Adenomatous ductal proliferation/hyperplasia (ADP/H) is a rare hyperplastic condition of the salivary gland. It is mostly associated with other salivary gland pathologies such as tumors and inflammations, and is incidentally found in tissue sections during histopathological examinations of those diseases. Herein, we report a case of ADP/H in the parotid gland not associated with any other pathological lesions, and present a review of the literature on this condition. A 60-year-old Japanese female complained of swelling on the left side of parotid region. Clinical examination revealed a swelling on the lower lobe of the left parotid gland. The lesion was firm but non-tender and was not attached to adjacent structures. A clinical diagnosis of benign salivary gland tumor was reached, and surgical excision was performed under general anesthesia. Histopathological examination revealed an intact parotid gland capsule with isomorphic and basaloid cells within scanty cytoplasm. In addition, an admixture of hyperplasia and proliferation of the intercalated ducts, the presence of zymogen granules, the absence of solid nests, and a peripheral palisaded arrangement of the cells were observed. Based on these findings, a diagnosis of ADP/H was confirmed. ADP/H is a non-tumorous lesion; therefore, tumor involvement should be ruled out before the diagnosis is reached.

Characterization and identification of the proteins bound to two types of polyhydroxyalkanoate granules in Pseudomonas sp. 61-3.

PubMed

Hokamura, Ayaka; Fujino, Kanako; Isoda, Yoshiko; Arizono, Koji; Shiratsuchi, Hideki; Matsusaki, Hiromi

2015-01-01

Pseudomonas sp. 61-3 accumulates two types of polyhydroxyalkanoates (PHAs), poly(3-hydroxybutyrate) [P(3HB)], and poly(3HB-co-3-hydroxyalkanoates) [P(3HB-co-3HA)], and some proteins associated with their PHA granules have been identified. To date, PhaFPs (GA36) and PhaIPs (GA18) were identified from P(3HB-co-3HA) granules. In this study, the gene encoding GA24 associated with P(3HB) granule was identified as phbPPs. PhbPPs was composed of 192 amino acids with a calculated molecular mass of 20.4 kDa and was assumed to be a phasin. phbFPs gene and unknown ORF were also found on phb locus. PhbFPs was anticipated to be the transcriptional repressor of phbPPs gene. PhbPPs was bound to the P(3HB-co-3HA) granules with 3HB composition of more than 87 mol%, and PhaIPs and PhaFPs were bound to the P(3HB-co-3HA) granules with 3HA (C6-C12) composition of more than 13 mol% in the producing cells, suggesting that localization of these proteins is attributed to the monomer compositions of the copolymers.

Venom Protein C activators as diagnostic agents for defects of protein C System.

PubMed

Ramzan, Faiqah; Asmat, Andleeb

2018-06-18

Background Protein C is a vitamin K dependent plasma zymogen. It prevents clotting by inhibiting clotting by inactivating factor V and factor VIII. Protein C activation pathway involves three steps: (i) Activation of protein C; (ii) Inhibition of coagulation through inactivating factor V and VIII by activated protein C and (iii) Inhibition of activated protein C by plasma protease inhibitors specific for this enzyme. Proteinases converts the zymogen Protein C (PC) of vertebrates into activated PC, which has been detected in several snake venoms. Most PC activators have been purified from venom of snake species belonging to the genera of the Agkistrodon complex. Unlike the physiological thrombin-catalyzed PC activation reaction which requires thrombomodulin as a cofactor, most snake venom activators directly convert the zymogen PC into the catalytically active form which can easily be determined by means of coagulation or chromogenic substrate techniques. Conclusion The fast-acting PC activator Protac® from Agkistrodon contortrix (southern copperhead snake) venom has been found to have broad application in diagnostic practice for the determination of disorders in the PC pathway. Recently, screening assays for the PC pathway have been introduced, based on the observation that the PC pathway is probably the most important physiological barrier against thrombosis. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Identification of key nitrous oxide production pathways in aerobic partial nitrifying granules.

PubMed

Ishii, Satoshi; Song, Yanjun; Rathnayake, Lashitha; Tumendelger, Azzaya; Satoh, Hisashi; Toyoda, Sakae; Yoshida, Naohiro; Okabe, Satoshi

2014-10-01

The identification of the key nitrous oxide (N2O) production pathways is important to establish a strategy to mitigate N2O emission. In this study, we combined real-time gas-monitoring analysis, (15)N stable isotope analysis, denitrification functional gene transcriptome analysis and microscale N2O concentration measurements to identify the main N2O producers in a partial nitrification (PN) aerobic granule reactor, which was fed with ammonium and acetate. Our results suggest that heterotrophic denitrification was the main contributor to N2O production in our PN aerobic granule reactor. The heterotrophic denitrifiers were probably related to Rhodocyclales bacteria, although different types of bacteria were active in the initial and latter stages of the PN reaction cycles, most likely in response to the presence of acetate. Hydroxylamine oxidation and nitrifier denitrification occurred, but their contribution to N2O emission was relatively small (20-30%) compared with heterotrophic denitrification. Our approach can be useful to quantitatively examine the relative contributions of the three pathways (hydroxylamine oxidation, nitrifier denitrification and heterotrophic denitrification) to N2O emission in mixed microbial populations. © 2014 Society for Applied Microbiology and John Wiley & Sons Ltd.

Identification of molting fluid carboxypeptidase A (MF-CPA) in Bombyx mori.

PubMed

Ote, Manabu; Mita, Kazuei; Kawasaki, Hideki; Daimon, Takaaki; Kobayashi, Masahiko; Shimada, Toru

2005-07-01

Using microarray analyses, we identified carboxypeptidase A (MF-CPA), which was induced during pupal ecdysis in the wing discs of Bombyx mori. Here, we report the functional characterization of MF-CPA. MF-CPA has amino acid sequence similarities with the proteins in the carboxypeptidase A/B subfamily, from human to nematode. The MF-CPA gene is expressed during the molting periods in the epithelial tissues. MF-CPA is detected in the molting fluid, which fills the space between the old and new cuticle during molting. By Western blot analysis, we show that MF-CPA is secreted as a zymogen and processed in the molting fluid. Recombinant MF-CPA expressed in the insect cells has carboxypeptidase A activity. We propose that MF-CPA degrades the proteins from the old cuticle during the molting periods and contributes to recycling of the amino acids.

Comparison of DNA extraction methods for meat analysis.

PubMed

Yalçınkaya, Burhanettin; Yumbul, Eylem; Mozioğlu, Erkan; Akgoz, Muslum

2017-04-15

Preventing adulteration of meat and meat products with less desirable or objectionable meat species is important not only for economical, religious and health reasons, but also, it is important for fair trade practices, therefore, several methods for identification of meat and meat products have been developed. In the present study, ten different DNA extraction methods, including Tris-EDTA Method, a modified Cetyltrimethylammonium Bromide (CTAB) Method, Alkaline Method, Urea Method, Salt Method, Guanidinium Isothiocyanate (GuSCN) Method, Wizard Method, Qiagen Method, Zymogen Method and Genespin Method were examined to determine their relative effectiveness for extracting DNA from meat samples. The results show that the salt method is easy to perform, inexpensive and environmentally friendly. Additionally, it has the highest yield among all the isolation methods tested. We suggest this method as an alternative method for DNA isolation from meat and meat products. Copyright © 2016 Elsevier Ltd. All rights reserved.

Ultrastructural Evidence of Serous Gland Polymorphism in the Skin of the Tungara Frog Engystomops pustulosus (Anura Leptodactylidae).

PubMed

Delfino, Giovanni; Giachi, Filippo; Malentacchi, Cecilia; Nosi, Daniele

2015-09-01

Three types of serous products were detected in the syncytial cutaneous glands of the leptodactylid tungara frog, Engystomops pustulosus: type Ia, granules with wide halos and variable density cores; type Ib, high density granules without halos; and type II, vesicles containing a finely dispersed product. Ultrastructural evidence revealed that these products were manufactured by different serous gland types and excluded that they represented different steps in the secretory cycle of a single gland type. Indeed, secretory maturation affecting the products released by the Golgi apparatus proceeded through different mechanisms: confluence (vesicles), interactions between syncytium and secretory product (type Ib granules), and a combination of both processes (type Ia granules). In conclusion, this investigation of secretory maturation was shown to be a suitable approach for the identification of serous gland polymorphism and demonstrated that the tungara frog belongs to the minority of anuran species characterized by this peculiar morpho-functional trait. © 2015 Wiley Periodicals, Inc.

Identification of inorganic and organic species of phosphorus and its bio-availability in nitrifying aerobic granular sludge.

PubMed

Huang, Wenli; Cai, Wei; Huang, He; Lei, Zhongfang; Zhang, Zhenya; Tay, Joo Hwa; Lee, Duu-Jong

2015-01-01

Phosphorus (P) recovery from sewage sludge is necessary for a sustainable development of the environment and thus the society due to gradual depletion of non-renewable P resources. Aerobic granular sludge is a promising biotechnology for wastewater treatment, which could achieve P-rich granules during simultaneous nitrification and denitrification processes. This study aimed to disclose the changes in inorganic and organic P species and their correlation with P mobility and bio-availability in aerobic granules. Two identical square reactors were used to cultivate aerobic granules, which were operated for 120 days with influent ammonia nitrogen (NH₄-N) of 100 mg/L before day 60 and then increased to 200 mg/L during the subsequent 60 days (chemical oxygen demand (COD) was kept constant at 600 mg/L). The aerobic granules exhibited excellent COD removal and nitrification efficiency. Results showed that inorganic P (IP) was about 61.4-67.7% of total P (TP) and non-apatite inorganic P (NAIP) occupied 61.9-70.2% of IP in the granules. The enrichment amount of NAIP and apatite P (AP) in the granules had strongly positive relationship with the contents of metal ions, i.e. Fe and Ca, respectively accumulated in the granules. X-ray diffraction (XRD) analysis and solution index calculation demonstrated that hydroxyapatite (Ca₅(PO₄)₃(OH)) and iron phosphate (Fe₇(PO₄)₆) were the major P minerals in the granules. Organic P (OP) content maintained around 7.5 mg per gram of biomass in the aerobic granules during the 120 days' operation. Monoester phosphate (21.8% of TP in extract), diester phosphate (1.8%) and phosphonate (0.1%) were identified as OP species by Phosphorus-31 nuclear magnetic resonance (³¹P NMR). The proportion of NAIP + OP to TP was about 80% in the granules, implying high potentially mobile and bio-available P was stored in the nitrifying aerobic granules. The present results provide a new insight into the characteristics of P species in aerobic granules, which could be helpful for developing P removal and recovery techniques through biological wastewater treatment.

Chemical fingerprint of Ganmaoling granule by double-wavelength ultra high performance liquid chromatography and ultra high performance liquid chromatography with quadrupole time-of-flight mass spectrometry.

PubMed

Lou, Qiong; Ye, Xiaolan; Zhou, Yingyi; Li, Hua; Song, Fenyun

2015-06-01

A method incorporating double-wavelength ultra high performance liquid chromatography with quadrupole time-of-flight mass spectrometry was developed for the investigation of the chemical fingerprint of Ganmaoling granule. The chromatographic separations were performed on an ACQUITY UPLC HSS C18 column (2.1 × 50 mm, 1.8 μm) at 30°C using gradient elution with water/formic acid (1%) and acetonitrile at a flow rate of 0.4 mL/min. A total of 11 chemical constituents of Ganmaoling granule were identified from their molecular weight, UV spectra, tandem mass spectrometry data, and retention behavior by comparing the results with those of the reference standards or literature. And 25 peaks were selected as the common peaks for fingerprint analysis to evaluate the similarities among 25 batches of Ganmaoling granule. The results of principal component analysis and orthogonal projection to latent structures discriminant analysis showed that the important chemical markers that could distinguish the different batches were revealed as 4,5-di-O-caffeoylquinic acid, 3,5-di-O-caffeoylquinic acid, and 4-O-caffeoylquinic acid. This is the first report of the ultra high performance liquid chromatography chemical fingerprint and component identification of Ganmaoling granule, which could lay a foundation for further studies of Ganmaoling granule. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Preparation and evaluation of highly drug-loaded fine globular granules using a multi-functional rotor processor.

PubMed

Iwao, Yasunori; Kimura, Shin-Ichiro; Ishida, Masayuki; Mise, Ryohei; Yamada, Masaki; Namiki, Noriyuki; Noguchi, Shuji; Itai, Shigeru

2015-01-01

The manufacture of highly drug-loaded fine globular granules eventually applied for orally disintegrating tablets has been investigated using a unique multi-functional rotor processor with acetaminophen, which was used as a model drug substance. Experimental design and statistical analysis were used to evaluate potential relationships between three key operating parameters (i.e., the binder flow rate, atomization pressure and rotating speed) and a series of associated micromeritics (i.e., granule mean size, proportion of fine particles (106-212 µm), flowability, roundness and water content). The results of multiple linear regression analysis revealed several trends, including (1) the binder flow rate and atomization pressure had significant positive and negative effects on the granule mean size value, Carr's flowability index, granular roundness and water content, respectively; (2) the proportion of fine particles was positively affected by the product of interaction between the binder flow rate and atomization pressure; and (3) the granular roundness was negatively and positively affected by the product of interactions between the binder flow rate and the atomization pressure, and the binder flow rate and rotating speed, respectively. The results of this study led to the identification of optimal operating conditions for the preparation of granules, and could therefore be used to provide important information for the development of processes for the manufacture of highly drug-loaded fine globular granules.

Starch granule evidence for the earliest potato use in North America

PubMed Central

Louderback, Lisbeth A.; Pavlik, Bruce M.

2017-01-01

The prehistory of wild potato use, leading to its domestication and diversification, has been well-documented in, and confined to, South America. At least 20 tuber-bearing, wild species of Solanum are known from North and Central America, yet their importance in ancient diets has never been assessed from the archaeological record. Here, we report the earliest evidence of wild potato use in North America at 10,900–10,100 calendar years (cal) B.P. in the form of well-preserved starch granules extracted from ground stone tools at North Creek Shelter, southern Utah. These granules have been identified as those of Solanum jamesii Torr. (Four Corners potato), a tuber-bearing species native to the American Southwest. Identification was based on applying five strictly defined diagnostic characteristics (eccentric hilum, longitudinal fissure, lack of fissure branching, fissure ratio, and maximum granule size) to each of 323 archaeological granules. Of those, nine were definitively assigned to S. jamesii based on possession of all characteristics, and another 61 were either likely or possibly S. jamesii depending on the number of characteristics they possessed. The oldest granules were found in substratum 4k (10,900–10,100 cal B.P.). Younger deposits, dating to ∼6,900 cal B.P., also contained tools with S. jamesii granules, indicating at least 4,000 y of intermittent use. Ethnographic and historical accounts extend the period of use to more than 10,000 y. The question then arises as to whether some S. jamesii populations could have undergone transport, cultivation, and eventual domestication over such a long period of time. PMID:28673982

Identification of the region of a 14-kilodalton protein of Rhodococcus ruber that is responsible for the binding of this phasin to polyhydroxyalkanoic acid granules.

PubMed Central

Pieper-Fürst, U; Madkour, M H; Mayer, F; Steinbüchel, A

1995-01-01

The function of the polyhydroxyalkanoic acid (PHA) granule-associated GA14 protein of Rhodococcus ruber was investigated in Escherichia coli XL1-Blue, which coexpressed this protein with the polyhydroxybutyric acid (PHB) biosynthesis operon of Alcaligenes eutrophus. The GA14 protein had no influence on the biosynthesis rate of PHB in E. coli XL1-Blue(pSKCO7), but this recombinant E. coli strain formed smaller PHB granules than were formed by an E. coli strain that expressed only the PHB operon. Immunoelectron microscopy with GA14-specific antibodies demonstrated the binding of GA14 protein to these mini granules. In a previous study, two hydrophobic domains close to the C terminus of the GA14 protein were analyzed, and a working hypothesis that suggested an anchoring of the GA14 protein in the phospholipid monolayer surrounding the PHA granule core by these hydrophobic domains was developed (U. Pieper-Fürst, M. H. Madkour, F. Mayer, and A. Steinbüchel, J. Bacteriol. 176:4328-4337, 1994). This hypothesis was confirmed by the construction of C-terminally truncated variants of the GA14 protein lacking the second or both hydrophobic domains and by the demonstration of their inability to bind to PHB granules. Further confirmation of the hypothesis was obtained by the construction of a fusion protein composed of the acetaldehyde dehydrogenase II of A. eutrophus and the C terminus of the GA14 protein containing both hydrophobic domains and by its affinity to native and artificial PHB granules. PMID:7730285

[Application of quality by design in granulation process for ginkgo leaf tablet (Ⅱ)： identification of critical quality attributes].

PubMed

Xu, Bing; Cui, Xiang-Long; Yang, Chan; Wang, Xin; Shi, Xin-Yuan; Qiao, Yan-Jiang

2017-03-01

Quality by design (QbD) highlights the concept of "begin with the end", which means to thoroughly understand the target product quality first, and then guide pharmaceutical process development and quality control throughout the whole manufacturing process. In this paper, the Ginkgo biloba granules intermediates were taken as the research object, and the requirements of the tensile strength of tablets were treated as the goals to establish the methods for identification of granules' critical quality attributes (CQAs) and establishment of CQAs' limits. Firstly, the orthogonal partial least square (OPLS) model was adopted to build the relationship between the micromeritic properties of 29 batches of granules and the tensile strength of ginkgo leaf tablets, and thereby the potential critical quality attributes (pCQAs) were screened by variable importance in the projection (VIP) indexes. Then, a series of OPLS models were rebuilt by reducing pCQAs variables one by one in view of the rule of VIP values from low to high in sequence. The model performance results demonstrated that calibration and predictive performance of the model had no decreasing trend after variables reduction. In consideration of the results from variables selection as well as the collinearity test and testability of the pCQAs, the median particle size (D₅₀) and the bulk density (Da) were identified as critical quality attributes (CQAs). The design space of CQAs was developed based on a multiple linear regression model established between the CQAs (D₅₀ and Da) and the tensile strength. The control constraints of the CQAs were determined as 170 μm< D₅₀<500 μm and 0.30 g•cm⁻³

Study on the Multi-marker Components Quantitative HPLC Fingerprint of the Compound Chinese Medicine Wuwei Changyanning Granule

PubMed Central

Yang, Xian; Yang, Shui-Ping; Zhang, Xue; Yu, Xiao-Dong; He, Qi-Yi; Wang, Bo-Chu

2014-01-01

The aim of this paper is to develop a rapid and highly sensitive quantitative HPLC fingerprint method with multiple indicators by using the Compound Chinese Medicine Wuwei Changyanning granule and 5 herbs in the prescription. The quantitative fingerprint chromatogram with multiple indicators was investigated. і)6 compositions included rutin, gallic acid, chlorogenic acid, atractylenolide Ⅰ, pachymic acid and apigenin, which originated from 5 herbs respectively, were selected as quantitative compositions, and their contents were determined using HPLC from 11 batches granules and the corresponding 5 medicinal materials. ⅱ) The precision, stability and repeatability of fingerprinting were investigated. In addition, common peaks number, the percentage of non-common peaks and similarity were also studied. Among them, 21 common peaks in the granule could find the source of peaks from the 5 herbs, among of 10 peaks from Niuerfeng, 9 peaks from Laliao, 3 peaks from Baishu, 3 peaks from Fuling and 5 peaks from Guanghuoxiang. The results showed that the identification method of fingerprinting was reliable. PMID:25587307

Sericin ameliorated dysmorphic mitochondria in high-cholesterol diet/streptozotocin rat by antioxidative property

PubMed Central

Ampawong, Sumate; Isarangkul, Duangnate

2016-01-01

Sericin has been implicated in lower cholesterolemic effect due to its properties with several mechanisms. Mitochondria are one of the most important targets to be affected in high blood cholesterol and glucose conditions. The protective role of sericin on mitochondria remains doubtful. To examine this role, electron microscopic, histopathologic, immunohistochemical, and biochemical studies were performed in a high-cholesterol diet/streptozotocin rat model. The results demonstrated that sericin reduced blood cholesterol without hypoglycemic effect. Sericin alleviated dysmorphic mitochondria in heart and liver but not in kidney and also decreased peculiar endoplasmic reticulum in the exocrine pancreas. In addition, sericin decreased hepatic steatosis and preserved zymogen granule referable to the decline of reactive oxygen species production in hepatic mitochondrial extraction and down-regulation of malondialdehyde expression in the liver and exocrine pancreas however irrelevant to lipase activity. This study suggests that sericin has antioxidative property to reduce blood cholesterol by means of diminishing fat deposit in hepatocyte and improves mitochondria and endoplasmic reticulum integrities. Impact statement The present work provides new insights regarding the antioxidative effect of sericin in (i) reducing blood cholesterol, (ii) improving liver and heart mitochondrial structures, (iii) maintaining endoplasmic reticulum integrity in exocrine pancreatic glands, and (iv) inhibiting fat deposition in the liver. Electron microscopic, histopathologic, immunohistochemical, and biochemical studies were performed. All of the results demonstrate the efficacy of sericin as a candidate for development of a functional food or adjunctive therapeutic agent against non-communicable diseases such as hypercholesterolemia. PMID:27903836

Genetic and functional characterization of PCSK1.

PubMed

Choquet, Hélène; Stijnen, Pieter; Creemers, John W M

2011-01-01

PC1/3 is a neuroendocrine-specific member of the mammalian subtilisin-like proprotein convertase family. This seven-member family is involved in the endoproteolytic cleavage of a large number of precursor proteins including prohormones, proneuropeptides, zymogens, and proreceptors. PC1/3 is synthesized as a zymogen, proPC1/3, and its propeptide is rapidly and autocatalytically cleaved in the endoplasmic reticulum. The mature protein is sorted and stored in dense-core secretory vesicles, together with its substrates. Compound-inactivating mutations in the PCSK1 gene, which encodes PC1/3, cause monogenic obesity. Furthermore, the contribution of two common nonsynonymous variants in PCSK1 to polygenic obesity risk has recently been established. Additional rare variants have been identified in non-consanguineous extremely obese Europeans but functional characterization has not yet been described. Sequencing efforts of larger cohorts of obese patients might reveal more variants conferring risk of obesity.

Tissue distribution and subcellular localizations determine in vivo functional relationship among prostasin, matriptase, HAI-1, and HAI-2 in human skin.

PubMed

Lee, Shiao-Pieng; Kao, Chen-Yu; Chang, Shun-Cheng; Chiu, Yi-Lin; Chen, Yen-Ju; Chen, Ming-Hsing G; Chang, Chun-Chia; Lin, Yu-Wen; Chiang, Chien-Ping; Wang, Jehng-Kang; Lin, Chen-Yong; Johnson, Michael D

2018-01-01

The membrane-bound serine proteases prostasin and matriptase and the Kunitz-type protease inhibitors HAI-1 and HAI-2 are all expressed in human skin and may form a tightly regulated proteolysis network, contributing to skin pathophysiology. Evidence from other systems, however, suggests that the relationship between matriptase and prostasin and between the proteases and the inhibitors can be context-dependent. In this study the in vivo zymogen activation and protease inhibition status of matriptase and prostasin were investigated in the human skin. Immunohistochemistry detected high levels of activated prostasin in the granular layer, but only low levels of activated matriptase restricted to the basal layer. Immunoblot analysis of foreskin lysates confirmed this in vivo zymogen activation status and further revealed that HAI-1 but not HAI-2 is the prominent inhibitor for prostasin and matriptase in skin. The zymogen activation status and location of the proteases does not support a close functional relation between matriptase and prostasin in the human skin. The limited role for HAI-2 in the inhibition of matriptase and prostasin is the result of its primarily intracellular localization in basal and spinous layer keratinocytes, which probably prevents the Kunitz inhibitor from interacting with active prostasin or matriptase. In contrast, the cell surface expression of HAI-1 in all viable epidermal layers renders it an effective regulator for matriptase and prostasin. Collectively, our study suggests the importance of tissue distribution and subcellular localization in the functional relationship between proteases and protease inhibitors.

Crystal structure of prethrombin-1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Zhiwei; Pelc, Leslie A.; Di Cera, Enrico

2010-11-15

Prothrombin is the zymogen precursor of the clotting enzyme thrombin, which is generated by two sequential cleavages at R271 and R320 by the prothrombinase complex. The structure of prothrombin is currently unknown. Prethrombin-1 differs from prothrombin for the absence of 155 residues in the N-terminal domain and is composed of a single polypeptide chain containing fragment 2 (residues 156-271), A chain (residues 272-320), and B chain (residues 321-579). The X-ray crystal structure of prethrombin-1 solved at 2.2-{angstrom} resolution shows an overall conformation significantly different (rmsd = 3.6 {angstrom}) from that of its active form meizothrombin desF1 carrying a cleavage atmore » R320. Fragment 2 is rotated around the y axis by 29{sup o} and makes only few contacts with the B chain. In the B chain, the oxyanion hole is disrupted due to absence of the I16-D194 ion pair and the Na{sup +} binding site and adjacent primary specificity pocket are highly perturbed. A remarkable feature of the structure is that the autolysis loop assumes a helical conformation enabling W148 and W215, located 17 {angstrom} apart in meizothrombin desF1, to come within 3.3 {angstrom} of each other and completely occlude access to the active site. These findings suggest that the zymogen form of thrombin possesses conformational plasticity comparable to that of the mature enzyme and have significant implications for the mechanism of prothrombin activation and the zymogen {yields} protease conversion in trypsin-like proteases.« less

Failure to Deliver and Translate-New Insights into RNA Dysregulation in ALS.

PubMed

Coyne, Alyssa N; Zaepfel, Benjamin L; Zarnescu, Daniela C

2017-01-01

Amyotrophic Lateral Sclerosis (ALS) is a progressive and fatal neurodegenerative disease affecting both upper and lower motor neurons. The molecular mechanisms underlying disease pathogenesis remain largely unknown. Multiple genetic loci including genes involved in proteostasis and ribostasis have been linked to ALS providing key insights into the molecular mechanisms underlying disease. In particular, the identification of the RNA binding proteins TDP-43 and fused in sarcoma (FUS) as causative factors of ALS resulted in a paradigm shift centered on the study of RNA dysregulation as a major mechanism of disease. With wild-type TDP-43 pathology being found in ~97% of ALS cases and the identification of disease causing mutations within its sequence, TDP-43 has emerged as a prominent player in ALS. More recently, studies of the newly discovered C9orf72 repeat expansion are lending further support to the notion of defects in RNA metabolism as a key factor underlying ALS. RNA binding proteins are involved in all aspects of RNA metabolism ranging from splicing, transcription, transport, storage into RNA/protein granules, and translation. How these processes are affected by disease-associated mutations is just beginning to be understood. Considerable work has gone into the identification of splicing and transcription defects resulting from mutations in RNA binding proteins associated with disease. More recently, defects in RNA transport and translation have been shown to be involved in the pathomechanism of ALS. A central hypothesis in the field is that disease causing mutations lead to the persistence of RNA/protein complexes known as stress granules. Under times of prolonged cellular stress these granules sequester specific mRNAs preventing them from translation, and are thought to evolve into pathological aggregates. Here we will review recent efforts directed at understanding how altered RNA metabolism contributes to ALS pathogenesis.

Characterization and identification of multiple constituents in Yinhuang granules by high-performance liquid chromatography with diode-array and time-of-flight mass spectrometry detection.

PubMed

Liu, E-Hu; Liu, Qun; Chu, Chu; Li, Ping

2011-10-01

A fast high-performance liquid chromatography (HPLC) method with diode-array detection (DAD) and time-of-flight mass spectrometry (TOF/MS) has been developed for the analysis of multi-constituent in Yinhuang granules, a well-known combined herbal remedy prepared from the extract mixtures of Flos Lonicerae and Radix Scutellariae. The fast HPLC analysis was performed on an Agilent ZorBax SB-C(18) column (4.6×50 mm, 1.8 μm) and 0.2% aqueous formic acid and acetonitrile was the optimum mobile phase for gradient elution in 17 min, which is five times faster than the performance of conventional columns packed with 5.0 μm particles. With various fragmentor voltages in TOF/MS, accurate mass measurements (<5 ppm error) for molecular ions and characteristic fragment ions represented reliable identification criteria for different constituents. A total of 28 compounds, including nine phenolic acids, three iridoid glycosides and nine saponins from Flos Lonicerae and seven flavonoids from Radix Scutellariae, were identified or tentatively characterized in the extract of Yinhuang granules. The established fast HPLC-DAD-TOF/MS method turns out to be useful and efficient for quality control of this commonly used Chinese herbal preparation. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Two new Haemoproteus species (Haemosporida: Haemoproteidae) from columbiform birds.

PubMed

Valkiūnas, Gediminas; Iezhova, Tatjana A; Evans, Edward; Carlson, Jenny S; Martínez-Gómez, Juan E; Sehgal, Ravinder N M

2013-06-01

Here we describe Haemoproteus (Haemoproteus) multivolutinus n. sp. from a tambourine dove (Turtur timpanistria) of Uganda and Haemoproteus (Haemoproteus) paramultipigmentatus n. sp. (Haemosporida, Haemoproteidae) from the Socorro common ground dove (Columbina passerina socorroensis) of Socorro Island, Mexico. These parasites are described based on the morphology of their blood stages and segments of the mitochondrial cytochrome b gene that can be used for molecular identification and diagnosis of these species. Gametocytes of H. multivolutinus possess rod-like pigment granules and are evenly packed with volutin, which masks pigment granules and darkly stains both macro- and microgametocytes in the early stages of their development. Based on these 2 characters, H. multivolutinus can be readily distinguished from other species of hemoproteids parasitizing columbiform (Columbiformes) birds. Haemoproteus paramultipigmentatus resembles Haemoproteus multipigmentatus; it can be distinguished from the latter parasite primarily due to the broadly ovoid shape of its young gametocytes and significantly fewer pigment granules in its fully developed gametocytes. We provide illustrations of blood stages of the new species, and phylogenetic analyses identify DNA lineages closely related to these parasites. Cytochrome b lineages of Haemoproteus multivolutinus and H. paramultipigmentatus cluster with hippoboscid-transmitted lineages of hemoproteids; thus these parasites likely belong to the subgenus Haemoproteus. We emphasize the importance of using cytochrome b sequences in conjunction with thorough microscopic descriptions to facilitate future identification of these and other avian hemosporidian species.

[Investigation on production process quality control of traditional Chinese medicine--Banlangen granule as an example].

PubMed

Tan, Manrong; Yan, Dan; Qiu, Lingling; Chen, Longhu; Yan, Yan; Jin, Cheng; Li, Hanbing; Xiao, Xiaohe

2012-04-01

For the quality management system of herbal medicines, intermediate and finished products it exists the " short board" effect of methodologies. Based on the concept of process control, new strategies and new methods of the production process quality control had been established with the consideration of the actual production of traditional Chinese medicine an the characteristics of Chinese medicine. Taking Banlangen granule as a practice example, which was effective and widespread application, character identification, determination of index components, chemical fingerprint and biometrics technology were sequentially used respectively to assess the quality of Banlangen herbal medicines, intermediate (water extraction and alcohol precipitation) and finished product. With the transfer rate of chemical information and biological potency as indicators, the effectiveness and transmission of the above different assessments and control methods had been researched. And ultimately, the process quality control methods of Banlangen granule, which were based on chemical composition analysis-biometric analysis, had been set up. It can not only validly solute the current status that there were many manufacturers varying quality of Banlangen granule, but also ensure and enhance its clinical efficacy. Furthermore it provided a foundation for the construction of the quality control of traditional Chinese medicine production process.

Structural insights into activation and inhibition of histo-aspartic protease (HAP) from Plasmodium falciparum

PubMed Central

Bhaumik, Prasenjit; Xiao, Huogen; Hidaka, Koushi; Gustchina, Alla; Kiso, Yoshiaki; Yada, Rickey Y.; Wlodawer, Alexander

2012-01-01

Histo-aspartic protease (HAP) from Plasmodium falciparum offers a promising target for the development of novel antimalarial drugs. HAP exhibits high sequence similarity to pepsin-like aspartic proteases, but one of the two catalytic aspartates, Asp32, is replaced by histidine. Crystal structures of the truncated zymogen of HAP and of the complex of the mature enzyme with inhibitor KNI-10395 have been determined at 2.1 and 2.5 Å resolution, respectively. As in other proplasmepsins, the propeptide of the zymogen interacts with the C-terminal domain of the enzyme, forcing the N- and C- terminal domains apart, thereby separating His32 and Asp215 and preventing formation of the mature active site. In the inhibitor complex the enzyme forms a tight domain-swapped dimer, not previously seen in any aspartic proteases. The inhibitor is found in an unprecedented conformation resembling the letter “U”, stabilized by two intramolecular hydrogen bonds. Surprisingly, the location and conformation of the inhibitor are similar to the fragment of helix 2 comprising residues 34p–38p in the prosegments of the zymogens of gastric aspartic proteases; a corresponding helix assumes a vastly different orientation in proplasmepsins. Each inhibitor molecule is in contact with two molecules of HAP, interacting with the carboxylate group of the catalytic Asp215 of one HAP protomer through a water molecule, while also making a direct hydrogen bond to Glu278A′ of the other protomer. A comparison of the shifts in the positions of the catalytic residues in the inhibitor complex presented here with those published previously gives further hints regarding the enzymatic mechanism of HAP. PMID:21928835

Structural Insights into the Activation and Inhibition of Histo-Aspartic Protease from Plasmodium falciparum

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bhaumik, Prasenjit; Xiao, Huogen; Hidaka, Koushi

2012-09-17

Histo-aspartic protease (HAP) from Plasmodium falciparum is a promising target for the development of novel antimalarial drugs. The sequence of HAP is highly similar to those of pepsin-like aspartic proteases, but one of the two catalytic aspartates, Asp32, is replaced with histidine. Crystal structures of the truncated zymogen of HAP and of the complex of the mature enzyme with inhibitor KNI-10395 have been determined at 2.1 and 2.5 {angstrom} resolution, respectively. As in other proplasmepsins, the propeptide of the zymogen interacts with the C-terminal domain of the enzyme, forcing the N- and C-terminal domains apart, thereby separating His32 and Asp215more » and preventing formation of the mature active site. In the inhibitor complex, the enzyme forms a tight domain-swapped dimer, not previously seen in any aspartic proteases. The inhibitor is found in an unprecedented conformation resembling the letter U, stabilized by two intramolecular hydrogen bonds. Surprisingly, the location and conformation of the inhibitor are similar to those of the fragment of helix 2 comprising residues 34p-38p in the prosegments of the zymogens of gastric aspartic proteases; a corresponding helix assumes a vastly different orientation in proplasmepsins. Each inhibitor molecule is in contact with two molecules of HAP, interacting with the carboxylate group of the catalytic Asp215 of one HAP protomer through a water molecule, while also making a direct hydrogen bond to Glu278A' of the other protomer. A comparison of the shifts in the positions of the catalytic residues in the inhibitor complex presented here with those published previously gives further hints regarding the enzymatic mechanism of HAP.« less

Charge-Triggered Membrane Insertion of Matrix Metalloproteinase-7, Supporter of Innate Immunity and Tumors.

PubMed

Prior, Stephen H; Fulcher, Yan G; Koppisetti, Rama K; Jurkevich, Alexander; Van Doren, Steven R

2015-11-03

Matrix metalloproteinase-7 (MMP-7) sheds signaling proteins from cell surfaces to activate bacterial killing, wound healing, and tumorigenesis. The mechanism targeting soluble MMP-7 to membranes has been investigated. Nuclear magnetic resonance structures of the zymogen, free and bound to membrane mimics without and with anionic lipid, reveal peripheral binding to bilayers through paramagnetic relaxation enhancements. Addition of cholesterol sulfate partially embeds the protease in the bilayer, restricts its diffusion, and tips the active site away from the bilayer. Its insertion of hydrophobic residues organizes the lipids, pushing the head groups and sterol sulfate outward toward the enzyme's positive charge on the periphery of the enlarged interface. Fluorescence probing demonstrates a similar mode of binding to plasma membranes and internalized vesicles of colon cancer cells. Binding of bilayered micelles induces allosteric activation and conformational change in the auto-inhibitory peptide and the adjacent scissile site, illustrating a potential intermediate in the activation of the zymogen. Copyright © 2015 Elsevier Ltd. All rights reserved.

Monitoring fluidized bed drying of pharmaceutical granules.

PubMed

Briens, Lauren; Bojarra, Megan

2010-12-01

Placebo granules consisting of lactose monohydrate, corn starch, and polyvinylpyrrolidone were prepared using de-ionized water in a high-shear mixer and dried in a conical fluidized bed dryer at various superficial gas velocities. Acoustic, vibration, and pressure data obtained over the course of drying was analyzed using various statistical, frequency, fractal, and chaos techniques. Traditional monitoring methods were also used for reference. Analysis of the vibration data showed that the acceleration levels decreased during drying and reached a plateau once the granules had reached a final moisture content of 1–2 wt.%; this plateau did not differ significantly between superficial gas velocities, indicating a potential criterion to support drying endpoint identification. Acoustic emissions could not reliably identify the drying endpoint. However, high kurtosis values of acoustic emissions measured in the filtered air exhaust corresponded to high entrainment rates. This could be used for process control to adjust the fluidization gas velocity to allow drying to continue rapidly while minimizing entrainment and possible product losses.

Assessing the Potential for Bioremediation through Formation and Fate of Metal Rich Granules in the Terrestrial Environment

DTIC Science & Technology

2011-05-01

laboratory ► biochemistry and analytical laboratories BUILDING STRONG® Impacts of Lead (Pb) • Most common metal contaminant on US Army small arms...Diffraction (XRD): Crystalline phase identification through fingerprinting Tungsten and Calcium in Snail Shell W Identificati on of Selenium and

Comparison of PC12 and Cerebellar Granule Cell Cultures for Evaluating Neurite Outgrowth Using High Content Screening

EPA Science Inventory

Development of high-throughput assays for chemical screening and hazard identification is a pressing priority worldwide. One approach uses in vitro, cell-based assays which recapitulate biological events observed in vivo. Neurite outgrowth is one such critical cellular process un...

Comparative proteome analysis reveals four novel polyhydroxybutyrate (PHB) granule-associated proteins in Ralstonia eutropha H16.

PubMed

Sznajder, Anna; Pfeiffer, Daniel; Jendrossek, Dieter

2015-03-01

Identification of proteins that were present in a polyhydroxybutyrate (PHB) granule fraction isolated from Ralstonia eutropha but absent in the soluble, membrane, and membrane-associated fractions revealed the presence of only 12 polypeptides with PHB-specific locations plus 4 previously known PHB-associated proteins with multiple locations. None of the previously postulated PHB depolymerase isoenzymes (PhaZa2 to PhaZa5, PhaZd1, and PhaZd2) and none of the two known 3-hydroxybutyrate oligomer hydrolases (PhaZb and PhaZc) were significantly present in isolated PHB granules. Four polypeptides were found that had not yet been identified in PHB granules. Three of the novel proteins are putative α/β-hydrolases, and two of those (A0671 and B1632) have a PHB synthase/depolymerase signature. The third novel protein (A0225) is a patatin-like phospholipase, a type of enzyme that has not been described for PHB granules of any PHB-accumulating species. No function has been ascribed to the fourth protein (A2001), but its encoding gene forms an operon with phaB2 (acetoacetyl-coenzyme A [CoA] reductase) and phaC2 (PHB synthase), and this is in line with a putative function in PHB metabolism. The localization of the four new proteins at the PHB granule surface was confirmed in vivo by fluorescence microscopy of constructed fusion proteins with enhanced yellow fluorescent protein (eYFP). Deletion of A0671 and B1632 had a minor but detectable effect on the PHB mobilization ability in the stationary growth phase of nutrient broth (NB)-gluconate cells, confirming the functional involvement of both proteins in PHB metabolism. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

Comparative Proteome Analysis Reveals Four Novel Polyhydroxybutyrate (PHB) Granule-Associated Proteins in Ralstonia eutropha H16

PubMed Central

Sznajder, Anna; Pfeiffer, Daniel

2014-01-01

Identification of proteins that were present in a polyhydroxybutyrate (PHB) granule fraction isolated from Ralstonia eutropha but absent in the soluble, membrane, and membrane-associated fractions revealed the presence of only 12 polypeptides with PHB-specific locations plus 4 previously known PHB-associated proteins with multiple locations. None of the previously postulated PHB depolymerase isoenzymes (PhaZa2 to PhaZa5, PhaZd1, and PhaZd2) and none of the two known 3-hydroxybutyrate oligomer hydrolases (PhaZb and PhaZc) were significantly present in isolated PHB granules. Four polypeptides were found that had not yet been identified in PHB granules. Three of the novel proteins are putative α/β-hydrolases, and two of those (A0671 and B1632) have a PHB synthase/depolymerase signature. The third novel protein (A0225) is a patatin-like phospholipase, a type of enzyme that has not been described for PHB granules of any PHB-accumulating species. No function has been ascribed to the fourth protein (A2001), but its encoding gene forms an operon with phaB2 (acetoacetyl-coenzyme A [CoA] reductase) and phaC2 (PHB synthase), and this is in line with a putative function in PHB metabolism. The localization of the four new proteins at the PHB granule surface was confirmed in vivo by fluorescence microscopy of constructed fusion proteins with enhanced yellow fluorescent protein (eYFP). Deletion of A0671 and B1632 had a minor but detectable effect on the PHB mobilization ability in the stationary growth phase of nutrient broth (NB)-gluconate cells, confirming the functional involvement of both proteins in PHB metabolism. PMID:25548058

LYSOSOMAL FRACTIONS FROM TRANSITIONAL EPITHELIUM

PubMed Central

Kanczak, Norbert M.; Krall, Joseph I.; Hayes, E. Russell; Elliott, Willard B.

1965-01-01

Histochemical data suggested that the so called lipoid granules of transitional epithelium in some species are equivalent to lysosomes. Scrapings of bovine and canine transitional epithelium were subjected to differential centrifugation to confirm this identification biochemically. Fractions of rat liver, the classic source of lysosomes, were also prepared by the same methods to compare with the fractions obtained from urinary epithelium. In contrast to rat liver, uroepithelial fractions with a high relative specific activity for hydrolases were sedimented before the heavy mitochondria. Microscopically, these fractions contained the highest proportion of lipoid granules. The size and sedimentation characteristics of lysosomes from transitional epithelium more closely resembled those of lysosomes derived from rat kidney than those isolated from liver. PMID:14326111

Novel mutant alleles of the starch synthesis gene TaSSIVb-D result in the reduction of starch granule number per chloroplast in wheat.

PubMed

Guo, Huijun; Liu, Yunchuan; Li, Xiao; Yan, Zhihui; Xie, Yongdun; Xiong, Hongchun; Zhao, Linshu; Gu, Jiayu; Zhao, Shirong; Liu, Luxiang

2017-05-08

Transient starch provides carbon and energy for plant growth, and its synthesis is regulated by the joint action of a series of enzymes. Starch synthesis IV (SSIV) is one of the important starch synthase isoforms, but its impact on wheat starch synthesis has not yet been reported due to the lack of mutant lines. Using the TILLING approach, we identified 54 mutations in the wheat gene TaSSIVb-D, with a mutation density of 1/165 Kb. Among these, three missense mutations and one nonsense mutation were predicted to have severe impacts on protein function. In the mutants, TaSSIVb-D was significantly down-regulated without compensatory increases in the homoeologous genes TaSSIVb-A and TaSSIVb-B. Altered expression of TaSSIVb-D affected granule number per chloroplast; compared with wild type, the number of chloroplasts containing 0-2 granules was significantly increased, while the number containing 3-4 granules was decreased. Photosynthesis was affected accordingly; the maximum quantum yield and yield of PSII were significantly reduced in the nonsense mutant at the heading stage. These results indicate that TaSSIVb-D plays an important role in the formation of transient starch granules in wheat, which in turn impact the efficiency of photosynthesis. The mutagenized population created in this study allows the efficient identification of novel alleles of target genes and could be used as a resource for wheat functional genomics.

Vesicular trafficking of immune mediators in human eosinophils revealed by immunoelectron microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Melo, Rossana C.N., E-mail: rossana.melo@ufjf.edu.br; Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, 330 Brookline Avenue, CLS 943, Boston, MA 02215; Weller, Peter F.

Electron microscopy (EM)-based techniques are mostly responsible for our current view of cell morphology at the subcellular level and continue to play an essential role in biological research. In cells from the immune system, such as eosinophils, EM has helped to understand how cells package and release mediators involved in immune responses. Ultrastructural investigations of human eosinophils enabled visualization of secretory processes in detail and identification of a robust, vesicular trafficking essential for the secretion of immune mediators via a non-classical secretory pathway associated with secretory (specific) granules. This vesicular system is mainly organized as large tubular-vesicular carriers (Eosinophil Sombreromore » Vesicles – EoSVs) actively formed in response to cell activation and provides a sophisticated structural mechanism for delivery of granule-stored mediators. In this review, we highlight the application of EM techniques to recognize pools of immune mediators at vesicular compartments and to understand the complex secretory pathway within human eosinophils involved in inflammatory and allergic responses. - Highlights: • Application of EM to understand the complex secretory pathway in human eosinophils. • EM techniques reveal an active vesicular system associated with secretory granules. • Tubular vesicles are involved in the transport of granule-derived immune mediators.« less

PGE2 induces oenocytoid cell lysis via a G protein-coupled receptor in the beet armyworm, Spodoptera exigua

USDA-ARS?s Scientific Manuscript database

Eicosanoids mediate cellular and humoral immune responses in the beet armyworm, Spodoptera exigua, including activation of prophenoloxidase (PPO). PPO activation begins with release of its inactive zymogen, PPO, from oenocytoids in response to prostaglandins (PGs). Based on the biomedical literatur...

Phosphorus detection in vitrified bacteria by cryo-STEM annular dark-field analysis.

PubMed

Wolf, Sharon Grayer; Rez, Peter; Elbaum, Michael

2015-11-01

Bacterial cells often contain dense granules. Among these, polyphosphate bodies (PPBs) store inorganic phosphate for a variety of essential functions. Identification of PPBs has until now been accomplished by analytical methods that required drying or chemically fixing the cells. These methods entail large electron doses that are incompatible with low-dose imaging of cryogenic specimens. We show here that Scanning Transmission Electron Microscopy (STEM) of fully hydrated, intact, vitrified bacteria provides a simple means for mapping of phosphorus-containing dense granules based on quantitative sensitivity of the electron scattering to atomic number. A coarse resolution of the scattering angles distinguishes phosphorus from the abundant lighter atoms: carbon, nitrogen and oxygen. The theoretical basis is similar to Z contrast of materials science. EDX provides a positive identification of phosphorus, but importantly, the method need not involve a more severe electron dose than that required for imaging. The approach should prove useful in general for mapping of heavy elements in cryopreserved specimens when the element identity is known from the biological context. © 2015 The Authors Journal of Microscopy © 2015 Royal Microscopical Society.

Induction and identification of a small-granule, high-amylose mutant in cassava (Manihot esculenta Crantz).

PubMed

Ceballos, Hernán; Sánchez, Teresa; Denyer, Kay; Tofiño, Adriana P; Rosero, Elvia A; Dufour, Dominique; Smith, Alison; Morante, Nelson; Pérez, Juan C; Fahy, Brendan

2008-08-27

Only two mutations have been described in the literature, so far, regarding starch and root quality traits in cassava. This article reports on an induced mutation in this crop, first identified in 2006. Botanical seed from five different cassava families were irradiated with gamma rays. Seed was germinated, transplanted to the field (M1 plants), and self-pollinated to produce the M2 generation. Abnormal types regarding starch granule morphology were identified during the single plant evaluation of M2 genotypes. To confirm these characteristics, selected genotypes were cloned and a second evaluation, based on cloned plants obtained from vegetative multiplication, was completed in September 2007. Two M2 genotypes presented small starch granules, but only one could be fully characterized, presenting a granule size of 5.80 +/- 0.33 microm compared with three commercial clones with granule sizes ranging from 13.97 +/- 0.12 to 18.73 +/- 0.10 microm and higher-than-normal amylose content (up to 30.1% in cloned plants harvested in 2007, as compared with the typical values for "normal" cassava starch of around 19.8%). The gels produced by the starch of these plants did not show any viscosity when analyzed with the rapid viscoanalyzers (5% suspension), and the gels had low clarity. Low viscosity could be observed at higher concentrations (8 or 10% suspensions). Preliminary results suggest that the mutation may be due to a lesion in a gene encoding one of the isoforms of isoamylase (probably isa1 or isa2).

Characteristics of a Bacteriocin Derived from Streptococcus faecalis var. zymogenes Antagonistic to Diplococcus peumoniae

PubMed Central

Bottone, Edward; Allerhand, Jona; Pisano, Michael A.

1971-01-01

A bacteriocin-producing strain of Streptococcus faecalis var. zymogenes (E-1) was isolated from clinical material (conjunctiva). The active substance differed from bacteriocins described by other investigators primarily in its spectrum of antibacterial activity, especially by its marked inhibition of Diplococcus pneumoniae. The E-1 bacteriocin also inhibited nonhemolytic strains of enterococci as well as one-third of the Viridans group of streptococcal strains investigated. The degree of inhibition, however, as indicated by the size of the zones against the latter organisms, was significantly reduced. No activity was detected against any of the strains belonging to the following groups of bacteria: hemolytic enterococci, beta-hemolytic streptococci, nonhemolytic streptococci, staphylococci, and various gram-negative species. Similarly, three strains each of Bacillus cereus and Listeria monocytogenes and one strain of Erysipelothrix insidiosa were not inhibited. The bacteriocin was able to diffuse through bacterial membranes as well as cellulose dialyzer tubing. It was inactivated by heating to 80 C for 20 min but resisted inactivation by either trypsin or chloroform. Images PMID:4398532

In vitro evidence of a tissue factor-independent mode of action of recombinant factor VIIa in hemophilia.

PubMed

Augustsson, Cecilia; Persson, Egon

2014-11-13

Successful competition of activated factor VII (FVIIa) with zymogen factor VII (FVII) for tissue factor (TF) and loading of the platelet surface with FVIIa are plausible driving forces behind the pharmacological effect of recombinant FVIIa (rFVIIa) in hemophilia patients. Thrombin generation measurements in platelet-rich hemophilia A plasma revealed competition for TF, which potentially could reduce the effective (r)FVIIa:TF complex concentration and thereby attenuate factor Xa production. However, (auto)activation of FVII apparently counteracted the negative effect of zymogen binding; a small impact was observed at endogenous concentrations of FVII and FVIIa but was virtually absent at pharmacological amounts of rFVIIa. Moreover, corrections of the propagation phase in hemophilia A required rFVIIa concentrations above the range where a physiological level of FVII was capable to downregulate thrombin generation. These data strongly suggest that rFVIIa acts independently of TF in hemophilia therapy and that FVII displacement by rFVIIa is a negligible mechanistic component. © 2014 by The American Society of Hematology.

Protein Biomarkers Associated With Growth And Synaptogenesis In a cell culture model of neuronal development

EPA Science Inventory

Cerebellar granule cells (CGC) provide a homogenous population of cells which can be used as an in vitro model for studying the cellular processes involved in the normal development of the CNS. They may also be useful for hazard identification as in vitro screens fo...

OsBT1 encodes an ADP-glucose transporter involved in starch synthesis and compound granule formation in rice endosperm

PubMed Central

Li, Sanfeng; Wei, Xiangjin; Ren, Yulong; Qiu, Jiehua; Jiao, Guiai; Guo, Xiuping; Tang, Shaoqing; Wan, Jianmin; Hu, Peisong

2017-01-01

Starch is the main storage carbohydrate in higher plants. Although several enzymes and regulators for starch biosynthesis have been characterized, a complete regulatory network for starch synthesis in cereal seeds remains elusive. Here, we report the identification and characterization of the rice Brittle1 (OsBT1) gene, which is expressed specifically in the developing endosperm. The osbt1 mutant showed a white-core endosperm and a significantly lower grain weight than the wild-type. The formation and development of compound starch granules in osbt1 was obviously defective: the amyloplast was disintegrated at early developmental stages and the starch granules were disperse and not compound in the endosperm cells in the centre region of osbt1 seeds. The total starch content and amylose content was decreased and the physicochemical properties of starch were altered. Moreover, the degree of polymerization (DP) of amylopectin in osbt1 was remarkably different from that of wild-type. Map-based cloning of OsBT1 indicated that it encodes a putatively ADP-glucose transporter. OsBT1 coded protein localizes in the amyloplast envelope membrane. Furthermore, the expression of starch synthesis related genes was also altered in the osbt1 mutant. These findings indicate that OsBT1 plays an important role in starch synthesis and the formation of compound starch granules. PMID:28054650

Questions on unusual Mimivirus-like structures observed in human cells.

PubMed

Lusi, Elena Angela; Maloney, Dan; Caicci, Federico; Guarascio, Paolo

2017-01-01

Background: Mimiviruses or giant viruses that infect amoebas have the ability to retain the Gram stain, which is usually used to colour bacteria. There is some evidence suggesting that Mimiviruses can also infect human cells. Guided by these premises, we performed a routine Gram stain on a variety of human specimens to see if we could detect the same Gram positive blue granules that identify Mimiviruses in the amoebas.  Methods: We analysed 24 different human specimens (liver, brain, kidney, lymph node and ovary) using Gram stain histochemistry, electron microscopy immunogold, high resolution mass spectrometry and protein identification.  Results: We detected in the human cells Gram positive granules that were distinct from bacteria. The fine blue granules displayed the same pattern of the Gram positive granules that diagnose Mimiviruses in the cytoplasm of the amoebas. Electron microscopy confirmed the presence of human Mimiviruses-like structures and mass spectrometry identified histone H4 peptides, which had the same footprints as giant viruses. However, some differences were noted: the Mimivirus-like structures identified in the human cells were ubiquitous and manifested a distinct mammalian retroviral antigenicity.  Conclusions: Our main hypotheses are that the structures could be either giant viruses having a retroviral antigenicity or ancestral cellular components having a viral origin. However, other possible alternatives have been proposed to explain the nature and function of the newly identified structures.

Clostridium perfringens epsilon toxin targets granule cells in the mouse cerebellum and stimulates glutamate release.

PubMed

Lonchamp, Etienne; Dupont, Jean-Luc; Wioland, Laetitia; Courjaret, Raphaël; Mbebi-Liegeois, Corinne; Jover, Emmanuel; Doussau, Frédéric; Popoff, Michel R; Bossu, Jean-Louis; de Barry, Jean; Poulain, Bernard

2010-09-30

Epsilon toxin (ET) produced by C. perfringens types B and D is a highly potent pore-forming toxin. ET-intoxicated animals express severe neurological disorders that are thought to result from the formation of vasogenic brain edemas and indirect neuronal excitotoxicity. The cerebellum is a predilection site for ET damage. ET has been proposed to bind to glial cells such as astrocytes and oligodendrocytes. However, the possibility that ET binds and attacks the neurons remains an open question. Using specific anti-ET mouse polyclonal antibodies and mouse brain slices preincubated with ET, we found that several brain structures were labeled, the cerebellum being a prominent one. In cerebellar slices, we analyzed the co-staining of ET with specific cell markers, and found that ET binds to the cell body of granule cells, oligodendrocytes, but not astrocytes or nerve endings. Identification of granule cells as neuronal ET targets was confirmed by the observation that ET induced intracellular Ca(2+) rises and glutamate release in primary cultures of granule cells. In cultured cerebellar slices, whole cell patch-clamp recordings of synaptic currents in Purkinje cells revealed that ET greatly stimulates both spontaneous excitatory and inhibitory activities. However, pharmacological dissection of these effects indicated that they were only a result of an increased granule cell firing activity and did not involve a direct action of the toxin on glutamatergic nerve terminals or inhibitory interneurons. Patch-clamp recordings of granule cell somata showed that ET causes a decrease in neuronal membrane resistance associated with pore-opening and depolarization of the neuronal membrane, which subsequently lead to the firing of the neuronal network and stimulation of glutamate release. This work demonstrates that a subset of neurons can be directly targeted by ET, suggesting that part of ET-induced neuronal damage observed in neuronal tissue is due to a direct effect of ET on neurons.

Clostridium perfringens Epsilon Toxin Targets Granule Cells in the Mouse Cerebellum and Stimulates Glutamate Release

PubMed Central

Lonchamp, Etienne; Dupont, Jean-Luc; Wioland, Laetitia; Courjaret, Raphaël; Mbebi-Liegeois, Corinne; Jover, Emmanuel; Doussau, Frédéric; Popoff, Michel R.; Bossu, Jean-Louis; de Barry, Jean; Poulain, Bernard

2010-01-01

Epsilon toxin (ET) produced by C. perfringens types B and D is a highly potent pore-forming toxin. ET-intoxicated animals express severe neurological disorders that are thought to result from the formation of vasogenic brain edemas and indirect neuronal excitotoxicity. The cerebellum is a predilection site for ET damage. ET has been proposed to bind to glial cells such as astrocytes and oligodendrocytes. However, the possibility that ET binds and attacks the neurons remains an open question. Using specific anti-ET mouse polyclonal antibodies and mouse brain slices preincubated with ET, we found that several brain structures were labeled, the cerebellum being a prominent one. In cerebellar slices, we analyzed the co-staining of ET with specific cell markers, and found that ET binds to the cell body of granule cells, oligodendrocytes, but not astrocytes or nerve endings. Identification of granule cells as neuronal ET targets was confirmed by the observation that ET induced intracellular Ca2+ rises and glutamate release in primary cultures of granule cells. In cultured cerebellar slices, whole cell patch-clamp recordings of synaptic currents in Purkinje cells revealed that ET greatly stimulates both spontaneous excitatory and inhibitory activities. However, pharmacological dissection of these effects indicated that they were only a result of an increased granule cell firing activity and did not involve a direct action of the toxin on glutamatergic nerve terminals or inhibitory interneurons. Patch-clamp recordings of granule cell somata showed that ET causes a decrease in neuronal membrane resistance associated with pore-opening and depolarization of the neuronal membrane, which subsequently lead to the firing of the neuronal network and stimulation of glutamate release. This work demonstrates that a subset of neurons can be directly targeted by ET, suggesting that part of ET-induced neuronal damage observed in neuronal tissue is due to a direct effect of ET on neurons. PMID:20941361

Localization of Poly(3-Hydroxybutyrate) (PHB) Granule-Associated Proteins during PHB Granule Formation and Identification of Two New Phasins, PhaP6 and PhaP7, in Ralstonia eutropha H16

PubMed Central

Pfeiffer, Daniel

2012-01-01

Poly(3-hydroxybutyrate) (PHB) granules are covered by a surface layer consisting of mainly phasins and other PHB granule-associated proteins (PGAPs). Phasins are small amphiphilic proteins that determine the number and size of accumulated PHB granules. Five phasin proteins (PhaP1 to PhaP5) are known for Ralstonia eutropha. In this study, we identified three additional potential phasin genes (H16_B1988, H16_B2296, and H16_B2326) by inspection of the R. eutropha genome for sequences with “phasin 2 motifs.” To determine whether the corresponding proteins represent true PGAPs, fusions with eYFP (enhanced yellow fluorescent protein) were constructed. Similar fusions of eYFP with PhaP1 to PhaP5 as well as fusions with PHB synthase (PhaC1), an inactive PhaC1 variant (PhaC1-C319A), and PhaC2 were also made. All fusions were investigated in wild-type and PHB-negative backgrounds. Colocalization with PHB granules was found for all PhaC variants and for PhaP1 to PhaP5. Additionally, eYFP fusions with H16_B1988 and H16_B2326 colocalized with PHB. Fusions of H16_B2296 with eYFP, however, did not colocalize with PHB granules but did colocalize with the nucleoid region. Notably, all fusions (except H16_B2296) were soluble in a ΔphaC1 strain. These data confirm that H16_B1988 and H16_B2326 but not H16_B2296 encode true PGAPs, for which we propose the designation PhaP6 (H16_B1988) and PhaP7 (H16_B2326). When localization of phasins was investigated at different stages of PHB accumulation, fusions of PhaP6 and PhaP7 were soluble in the first 3 h under PHB-permissive conditions, although PHB granules appeared after 10 min. At later time points, the fusions colocalized with PHB. Remarkably, PHB granules of strains expressing eYFP fusions with PhaP5, PhaP6, or PhaP7 localized predominantly near the cell poles or in the area of future septum formation. This phenomenon was not observed for the other PGAPs (PhaP1 to PhaP4, PhaC1, PhaC1-C319A, and PhaC2) and indicated that some phasins can have additional functions. A chromosomal deletion of phaP6 or phaP7 had no visible effect on formation of PHB granules. PMID:22923598

Localization of poly(3-hydroxybutyrate) (PHB) granule-associated proteins during PHB granule formation and identification of two new phasins, PhaP6 and PhaP7, in Ralstonia eutropha H16.

PubMed

Pfeiffer, Daniel; Jendrossek, Dieter

2012-11-01

Poly(3-hydroxybutyrate) (PHB) granules are covered by a surface layer consisting of mainly phasins and other PHB granule-associated proteins (PGAPs). Phasins are small amphiphilic proteins that determine the number and size of accumulated PHB granules. Five phasin proteins (PhaP1 to PhaP5) are known for Ralstonia eutropha. In this study, we identified three additional potential phasin genes (H16_B1988, H16_B2296, and H16_B2326) by inspection of the R. eutropha genome for sequences with "phasin 2 motifs." To determine whether the corresponding proteins represent true PGAPs, fusions with eYFP (enhanced yellow fluorescent protein) were constructed. Similar fusions of eYFP with PhaP1 to PhaP5 as well as fusions with PHB synthase (PhaC1), an inactive PhaC1 variant (PhaC1-C319A), and PhaC2 were also made. All fusions were investigated in wild-type and PHB-negative backgrounds. Colocalization with PHB granules was found for all PhaC variants and for PhaP1 to PhaP5. Additionally, eYFP fusions with H16_B1988 and H16_B2326 colocalized with PHB. Fusions of H16_B2296 with eYFP, however, did not colocalize with PHB granules but did colocalize with the nucleoid region. Notably, all fusions (except H16_B2296) were soluble in a ΔphaC1 strain. These data confirm that H16_B1988 and H16_B2326 but not H16_B2296 encode true PGAPs, for which we propose the designation PhaP6 (H16_B1988) and PhaP7 (H16_B2326). When localization of phasins was investigated at different stages of PHB accumulation, fusions of PhaP6 and PhaP7 were soluble in the first 3 h under PHB-permissive conditions, although PHB granules appeared after 10 min. At later time points, the fusions colocalized with PHB. Remarkably, PHB granules of strains expressing eYFP fusions with PhaP5, PhaP6, or PhaP7 localized predominantly near the cell poles or in the area of future septum formation. This phenomenon was not observed for the other PGAPs (PhaP1 to PhaP4, PhaC1, PhaC1-C319A, and PhaC2) and indicated that some phasins can have additional functions. A chromosomal deletion of phaP6 or phaP7 had no visible effect on formation of PHB granules.

New advances in cell physiology and pathophysiology of the exocrine pancreas.

PubMed

Mössner, Joachim

2010-01-01

This review provides some aspects on the physiology of stimulation and inhibition of pancreatic digestive enzyme secretion and the pathophysiology of pancreatic acinar cell function leading to pancreatitis. Cholecystokinin (CCK) stimulates both directly via CCK-A receptors on acinar cells and indirectly via CCK-B receptors on nerves, followed by acetylcholine release, pancreatic enzyme secretion. It is still not known whether CCK-A receptors exist in human acinar cells, in contrast to acinar cells of rodents where CCK-A receptors have been well described. CCK has numerous actions both in the periphery and in the central nervous systems. CCK inhibits gastric motility and regulates satiety. Another major function of CCK is stimulation of gallbladder contraction. This function enables that bile acids act simultaneously with pancreatic lipolytic enzymes. Secretin is a major stimulator of bicarbonate secretion. Trypsinogen is activated by the gut mucosal enzyme enterokinase. The other pancreatic proenzymes are activated by trypsin. Termination of enzyme secretion may be regulated by negative feedback mechanisms via destruction of CCK-releasing peptides by trypsin. Furthermore, the ileum may act as a brake by release of inhibitory hormones such as PYY and somatostatin. In the pathophysiology of acute pancreatitis, fusion of zymogen granules with lysosomes leading to intracellular activation of trypsinogen is regarded as an initiation step. This activation of trypsinogen may be caused by the lysosomal enzyme cathepsin B. However, autoactivation of trypsinogen itself may be a possibility in pathogenesis. Autoactivation is enhanced in certain mutations of trypsinogen. Furthermore, an imbalance of protease inhibitors and active proteases may be involved. The role of pancreatic lipolytic enzymes, the role of bicarbonate secretion, and toxic Ca(2+) signals by excessive liberation from the endoplasmic reticulum have to be discussed in the pathogenesis of acute pancreatitis. Copyright © 2011 S. Karger AG, Basel.

Histological study of the gastrointestinal tract in longfin yellowtail (Seriola rivoliana) larvae.

PubMed

Teles, Andressa; Salas-Leiva, Joan; Alvarez-González, Carlos Alfonso; Gisbert, Enric; Ibarra-Castro, Leonardo; Urbiola, Juan Carlos Pérez; Tovar-Ramírez, Dariel

2017-12-01

This work contributes basic knowledge on larval development of Seriola rivoliana. A histological study describes the development of the digestive tract and accessory glands in S. rivoliana larvae reared under laboratory conditions at 24 °C from hatching to 30 days post-hatching (DPH). At hatching (2.6 ± 0.12 mm), larvae had an undifferentiated digestive tract with a closed straight tube and a large yolk sac with an oil globule. The liver and pancreas were observed at 1 and 2 days, and the mouth and anus opened at day 2. Enriched rotifers were visible in their digestive tract. At the beginning of the pre-flexion stage, a mixed nutritional period was observed. At day 3, exogenous feeding began; the digestive tract became differentiated into the buccopharynx, esophagus, an undifferentiated stomach, and the intestines. Zymogen granules were visible in the exocrine pancreas. At day 4, supranuclear vacuoles were present in the posterior intestine, indicating the beginning of intracellular digestion. At day 5, goblet cells were present in the esophagus and became functional at day 7 in the esophagus and intestine. The buccopharynx goblet cells developed at day 15. The presence of gastric glands and differentiation of the stomach in the fundic, cardiac, and pyloric regions during the post-flexion stage occurred at day 20. This was the onset of the juvenile period and the beginning of weaning; however, a long co-feeding phase is recommended. Pyloric caeca were observed at day 30 (13.6 ± 1.6 mm). These results provide valuable information on S. rivoliana larvae biology and digestive physiology, which should be useful to improve cultivation techniques and identify ecological features involved in ontogeny.

EPI64B Acts as a GTPase-activating Protein for Rab27B in Pancreatic Acinar Cells*

PubMed Central

Hou, Yanan; Chen, Xuequn; Tolmachova, Tatyana; Ernst, Stephen A.; Williams, John A.

2013-01-01

The small GTPase Rab27B localizes to the zymogen granule membranes and plays an important role in regulating protein secretion by pancreatic acinar cells, as does Rab3D. A common guanine nucleotide exchange factor (GEF) for Rab3 and Rab27 has been reported; however, the GTPase-activating protein (GAP) specific for Rab27B has not been identified. In this study, the expression in mouse pancreatic acini of two candidate Tre-2/Bub2/Cdc16 (TBC) domain-containing proteins, EPI64 (TBC1D10A) and EPI64B (TBC1D10B), was first demonstrated. Their GAP activity on digestive enzyme secretion was examined by adenovirus-mediated overexpression of EPI64 and EPI64B in isolated pancreatic acini. EPI64B almost completely abolished the GTP-bound form of Rab27B, without affecting GTP-Rab3D. Overexpression of EPI64B also enhanced amylase release. This enhanced release was independent of Rab27A, but dependent on Rab27B, as shown using acini from genetically modified mice. EPI64 had a mild effect on both GTP-Rab27B and amylase release. Co-overexpression of EPI64B with Rab27B can reverse the inhibitory effect of Rab27B on amylase release. Mutations that block the GAP activity decreased the inhibitory effect of EPI64B on the GTP-bound state of Rab27B and abolished the enhancing effect of EPI64B on the amylase release. These data suggest that EPI64B can serve as a potential physiological GAP for Rab27B and thereby participate in the regulation of exocytosis in pancreatic acinar cells. PMID:23671284

Leucine Affects α-Amylase Synthesis through PI3K/Akt-mTOR Signaling Pathways in Pancreatic Acinar Cells of Dairy Calves.

PubMed

Guo, Long; Liang, Ziqi; Zheng, Chen; Liu, Baolong; Yin, Qingyan; Cao, Yangchun; Yao, Junhu

2018-05-23

Dietary nutrient utilization, particularly starch, is potentially limited by digestion in dairy cow small intestine because of shortage of α-amylase. Leucine acts as an effective signal molecular in the mTOR signaling pathway, which regulates a series of biological processes, especially protein synthesis. It has been reported that leucine could affect α-amylase synthesis and secretion in ruminant pancreas, but mechanisms have not been elaborated. In this study, pancreatic acinar (PA) cells were used as a model to determine the cellular signal of leucine influence on α-amylase synthesis. PA cells were isolated from newborn Holstein dairy bull calves and cultured in Dulbecco's modifed Eagle's medium/nutrient mixture F12 liquid media containing four leucine treatments (0, 0.23, 0.45, and 0.90 mM, respectively), following α-amylase activity, zymogen granule, and signal pathway factor expression detection. Rapamycin, a specific inhibitor of mTOR, was also applied to PA cells. Results showed that leucine increased ( p < 0.05) synthesis of α-amylase as well as phosphorylation of PI3K, Akt, mTOR, and S6K1 while reduced ( p < 0.05) GCN2 expression. Inhibition of mTOR signaling downregulated the α-amylase synthesis. In addition, the extracellular leucine dosage significantly influenced intracellular metabolism of isoleucine ( p < 0.05). Overall, leucine regulates α-amylase synthesis through promoting the PI3K/Akt-mTOR pathway and reducing the GCN2 pathway in PA cells of dairy calves. These pathways form the signaling network that controls the protein synthesis and metabolism. It would be of great interest in future studies to explore the function of leucine in ruminant nutrition.

Ultrastructural evaluation of adenocarcinomas derived from apocrine glands of the anal sac associated with hypercalcemia in dogs.

PubMed Central

Meuten, D. J.; Capen, C. C.; Kociba, G. J.; Chew, D. J.; Cooper, B. J.

1982-01-01

Adenocarcinomas derived from apocrine glands of the anal sac and associated with persistent hypercalcemia in dogs were composed of tumor cells with numerous profiles of rough endoplasmic reticulum, clusters of free ribosomes, and a prominent Golgi apparatus. Neoplastic cells contained microtubules, microfilaments, tonofibrils, and had two types of electron-dense granules. Large lysosomelike dense bodies ranged from 0.6 to 2.2 microns in diameter and had a poorly delineated limiting membrane. Small granules (150-400 nm in diameter) had a sharply delineated limiting membrane with a narrow submembranous space and a homogeneous dense core. These smaller granules usually were located near the apexes of neoplastic cells, whereas the larger granules were situated near the base of cells. Apocrine cells in glands of the anal sac from control dogs that were in the secretory phase were columnar and had large dilated profiles of rough endoplasmic reticulum. Membranes of the endoplasmic reticulum fused with the plasmalemma and appeared to secrete their product directly into the lumens of acini, characteristic of merocrine secretion. Apical blebs of electron-lucent cytoplasm pinched off from nonneoplastic aprocine cells and were released into glandular lumens. Similar electron-lucent cytoplasmic blebs were present at the apexes of tumor cells. Myoepithelial cells were present between the epithelial cells and basement membrane in normal apocrine glands and were absent in neoplasms derived from these glands. Identification of the contents of the secretory-like granules in tumor cells and characterization of the hypercalcemic factor in the plasma or tumor tissue from dogs with this syndrome will help explain the pathogenesis of hypercalcemia associated with malignancy in animals and man. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Figure 8 Figure 9 Figure 10 Figure 11 PMID:7200729

Ultrastructural evaluation of adenocarcinomas derived from apocrine glands of the anal sac associated with hypercalcemia in dogs.

PubMed

Meuten, D J; Capen, C C; Kociba, G J; Chew, D J; Cooper, B J

1982-05-01

Adenocarcinomas derived from apocrine glands of the anal sac and associated with persistent hypercalcemia in dogs were composed of tumor cells with numerous profiles of rough endoplasmic reticulum, clusters of free ribosomes, and a prominent Golgi apparatus. Neoplastic cells contained microtubules, microfilaments, tonofibrils, and had two types of electron-dense granules. Large lysosomelike dense bodies ranged from 0.6 to 2.2 microns in diameter and had a poorly delineated limiting membrane. Small granules (150-400 nm in diameter) had a sharply delineated limiting membrane with a narrow submembranous space and a homogeneous dense core. These smaller granules usually were located near the apexes of neoplastic cells, whereas the larger granules were situated near the base of cells. Apocrine cells in glands of the anal sac from control dogs that were in the secretory phase were columnar and had large dilated profiles of rough endoplasmic reticulum. Membranes of the endoplasmic reticulum fused with the plasmalemma and appeared to secrete their product directly into the lumens of acini, characteristic of merocrine secretion. Apical blebs of electron-lucent cytoplasm pinched off from nonneoplastic aprocine cells and were released into glandular lumens. Similar electron-lucent cytoplasmic blebs were present at the apexes of tumor cells. Myoepithelial cells were present between the epithelial cells and basement membrane in normal apocrine glands and were absent in neoplasms derived from these glands. Identification of the contents of the secretory-like granules in tumor cells and characterization of the hypercalcemic factor in the plasma or tumor tissue from dogs with this syndrome will help explain the pathogenesis of hypercalcemia associated with malignancy in animals and man.

Development of immunoassays for human urokinase

NASA Technical Reports Server (NTRS)

Atassi, M. Zouhair

1988-01-01

Radioimmune assays (RIA) and enzyme linked immune assays for measurement of pro-urokinase and the two active forms of the enzyme were developed. Polyclonal and monoclonal antibodies, with desired specificities against preselected synthetic regions of urokinase (UK), were obtained by immunization with the respective synthetic peptides and used to develop RIA for zymogen and the two activated forms of UK.

Identification of an abundant 56 kDa protein implicated in food allergy as granule-bound starch synthase

USDA-ARS?s Scientific Manuscript database

Rice, the staple food of South and East Asian counties, is considered to be hypoallergenic. However, several clinical studies have documented rice-induced allergy in sensitive patients. Rice proteins with molecular weights of 14-16 kDa, 26 kDa, 33 kDa and 56 kDa have been identified as allergens. Re...

Distribution Profile of Inositol 1,4,5-Trisphosphate Receptor/Ca2+ Channels in α and β Cells of Pancreas: Dominant Localization in Secretory Granules and Common Error in Identification of Secretory Granule Membranes.

PubMed

Hur, Yong Suk; Yoo, Seung Hyun

2015-01-01

The α and β cells of pancreatic islet release important hormones in response to intracellular Ca increases that result from Ca releases through the inositol 1,4,5-trisphoshate receptor (IP3R)/Ca channels. Yet no systematic studies on distribution of IP3R/Ca channels have been done, prompting us to investigate the distribution of all 3 IP3R isoforms. Immunogold electron microscopy was performed to determine the presence and the relative concentrations of all 3 IP3R isoforms in 2 major organelles secretory granules (SGs) and the endoplasmic reticulum of α and β cells of rat pancreas. All 3 IP3R isoforms were present in SG membranes of both cells, and the IP3R concentrations in SGs were ∼2-fold higher than those in the endoplasmic reticulum. Moreover, large halos shown in the electron microscope images of insulin-containing SGs of β cells were gap spaces that resulted from separation of granule membranes from the surrounding cytoplasm. These results strongly suggest the important roles of SGs in IP3-induced, Ca-dependent regulatory secretory pathway in pancreas. Moreover, the accurate location of SG membranes of β cells was further confirmed by the location of another integral membrane protein synaptotagmin V and of membrane phospholipid PI(4,5)P2.

Identification and analysis of chemical constituents and rat serum metabolites in Suan-Zao-Ren granule using ultra high performance liquid chromatography quadrupole time-of-flight mass spectrometry combined with multiple data processing approaches.

PubMed

Du, Yiyang; He, Bosai; Li, Qing; He, Jiao; Wang, Di; Bi, Kaishun

2017-07-01

Suan-Zao-Ren granule is widely used to treat insomnia in China. However, because of the complexity and diversity of the chemical compositions in traditional Chinese medicine formula, the comprehensive analysis of constituents in vitro and in vivo is rather difficult. In our study, an ultra high performance liquid chromatography with quadrupole time-of-flight mass spectrometry and the PeakView® software, which uses multiple data processing approaches including product ion filter, neutral loss filter, and mass defect filter, method was developed to characterize the ingredients and rat serum metabolites in Suan-Zao-Ren granule. A total of 101 constituents were detected in vitro. Under the same analysis conditions, 68 constituents were characterized in rat serum, including 35 prototype components and 33 metabolites. The metabolic pathways of main components were also illustrated. Among them, the metabolic pathways of timosaponin AI were firstly revealed. The bioactive compounds mainly underwent the phase I metabolic pathways including hydroxylation, oxidation, hydrolysis, and phase II metabolic pathways including sulfate conjugation, glucuronide conjugation, cysteine conjugation, acetycysteine conjugation, and glutathione conjugation. In conclusion, our results showed that this analysis approach was extremely useful for the in-depth pharmacological research of Suan-Zao-Ren granule and provided a chemical basis for its rational. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The defensive secretion of Carabus lefebvrei Dejean 1826 pupa (Coleoptera, Carabidae): gland ultrastructure and chemical identification.

PubMed

Giglio, Anita; Brandmayr, Pietro; Dalpozzo, Renato; Sindona, Giovanni; Tagarelli, Antonio; Talarico, Federica; Brandmayr, Tullia Zetto; Ferrero, Enrico A

2009-05-01

This study documents the defensive function of flavored humor secreted by the abdominal glands of Carabus lefebvrei pupae. The morphology and the ultrastructure of these glands were described and the volatile compounds of glands secretion were identified by gas chromatography/mass spectrometry. The ultrastructure analysis shows an acinose complex formed by about 50 clusters. Each cluster has 20 glandular units and the unit-composed of one secretory and one canal cell lying along a duct-belongs to the class 3 cell type of Quennedey (1998). In the cytoplasm, the secretory cell contains abundant rough endoplasmatic reticula, glycogen granules, numerous mitochondria, and many well-developed Golgi complexes producing electron-dense secretory granules. Mitochondria are large, elongated, and often adjoining electronlucent vesicles. The kind and the origin of secretory granules varying in size and density were discussed. The chemical analysis of the gland secretion revealed the presence of a mixture of low molecular weight terpenes, ketones, aldehydes, alcohols, esters, and carboxylic acids. Monoterpenes, especially linalool, were the major products. We supposed that ketones, aldehydes, alcohols, esters, and carboxylic acids have a deterrent function against the predators and monoterpenes provide a prophylaxis function against pathogens. (c) 2008 Wiley-Liss, Inc.

Guidelines on common cold for traditional Chinese medicine based on pattern differentiation.

PubMed

Jiao, Yang; Liu, Jianping; Jiang, Liangduo; Liu, Qingquan; Li, Xiaoli; Zhang, Shunan; Zhao, Baixiao; Wang, Tianfang

2013-08-01

To establish the guidelines on common cold treated with Traditional Chinese Medicine (TCM) in terms of pattern identification. The guidelines were formulated by using the basic patterns common cold in China Pharmacopeia integrated with findings from systematic literature review and the experts' consensus on the issue in question. Common cold was divided into four patterns in the guidelines. The medications were recommended respectively: Ganmaoqingre granule for wind-cold exterior syndrome, Yinqiaojiedu granule for wind-heat exterior syndrome, Huoxiangzhengqi Wan for summer-heat dampness exterior syndrome and Shensu Wan for wind-cold exterior syndrome accompanied with Qi deficiency. The guidelines were primarily derived from the practice experience of TCM and the experts' consensus. The process was not strictly evidence-based because of lacking enough clinical studies. Further refinement of the guidelines should be needed as more studies are available.

Pancreatitis in wild zinc-poisoned waterfowl

USGS Publications Warehouse

Sileo, Louis; Beyer, W. Nelson; Mateo, Rafael

2003-01-01

Four waterfowl were collected in the TriState Mining District (Oklahoma, Kansas and Missouri, USA), an area known to be contaminated with lead, cadmium and zinc (Zn). They were part of a larger group of 20 waterfowl collected to determine the exposure of birds to metal contamination at the site. The four waterfowl (three Branta canadensis, one Anas platyrhynchos) had mild to severe degenerative abnormalities of the exocrine pancreas, as well as tissue (pancreas, liver) concentrations of Zn that were considered toxic. The mildest condition was characterized by generalized atrophy of exocrine cells that exhibited cytoplasmic vacuoles and a relative lack of zymogen. The most severe condition was characterized by acini with distended lumens and hyperplastic exocrine tissue that completely lacked zymogen; these acini were widely separated by immature fibrous tissue. Because the lesions were nearly identical to the lesions reported in chickens and captive waterfowl that had been poisoned with ingested Zn, and because the concentrations of Zn in the pancreas and liver of the four birds were consistent with the concentrations measured in Zn-poisoned birds, we concluded that these waterfowl were poisoned by Zn. This may be the first reported case of zinc poisoning in free-ranging wild birds poisoned by environmental Zn.

Metabolic efficiency and turnover of soil microbial communities in biodegradation tests.

PubMed Central

Shen, J; Bartha, R

1996-01-01

Biodegradability screening tests of soil commonly measure 14CO2 evolution from radiolabeled test compounds, and glucose has often served as a positive control. When constant amounts of radiolabel were added to soil in combination with increasing amounts of unlabeled substrates, glucose and some related hexoses behaved in an anomalous manner. In contrast to that of formate, benzoate, n-hexadecane, or bis(2-ethylhexyl) phthalate, dilution of glucose radiocarbon with unlabeled glucose increased rather than decreased the rate and extent of 14CO2 evolution. [14C]glucose incorporation into biomass and Vmax values were consistent with the interpretation that application of relatively high concentrations of glucose to soil shifts the balance of the soil microbial community from the autochthonous (humus-degrading) to the zymogeneous (opportunistic) segment. The higher growth and turnover rates that define zymogeneous microorganisms, combined with a lower level of carbon incorporation into their biomass, result in the evolution of disproportionate percentages of 14CO2. When used as positive controls, glucose and related hexoses may raise the expectations for percent 14CO2 evolution to levels that are not realistic for other biodegradable compounds. PMID:8779580

Metaplastic Cells in the Stomach Arise, Independently of Stem Cells, via Dedifferentiation or Transdifferentiation of Chief Cells.

PubMed

Radyk, Megan D; Burclaff, Joseph; Willet, Spencer G; Mills, Jason C

2018-03-01

Spasmolytic polypeptide-expressing metaplasia (SPEM) develops in patients with chronic atrophic gastritis due to infection with Helicobacter pylori; it might be a precursor to intestinal metaplasia and gastric adenocarcinoma. Lineage tracing experiments of the gastric corpus in mice have not established whether SPEM derives from proliferating stem cells or differentiated, post-mitotic zymogenic chief cells in the gland base. We investigated whether differentiated cells can give rise to SPEM using a nongenetic approach in mice. Mice were given intraperitoneal injections of 5-fluorouracil, which blocked gastric cell proliferation, plus tamoxifen to induce SPEM. Based on analyses of molecular and histologic markers, we found SPEM developed even in the absence of cell proliferation. SPEM therefore did not arise from stem cells. In histologic analyses of gastric resection specimens from 10 patients with adenocarcinoma, we found normal zymogenic chief cells that were transitioning into SPEM cells only in gland bases, rather than the proliferative stem cell zone. Our findings indicate that SPEM can arise by direct reprogramming of existing cells-mainly of chief cells. Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.

Correlation between loose density and compactibility of granules prepared by various granulation methods.

PubMed

Murakami, H; Yoneyama, T; Nakajima, K; Kobayashi, M

2001-03-23

The objectives of this study were to prepare the lactose granules by various granulation methods using polyethylene glycol 6000 (PEG 6000) as a binder and to evaluate the effects of granulation methods on the compressibility and compactibility of granules in tabletting. Lactose was granulated by seven granulation methods -- four wet granulations including wet massing granulation, wet high-speed mixer granulation, wet fluidized bed granulation and wet tumbling fluidized bed granulation; and three melt granulations including melt high-speed mixer granulation, melt fluidized bed granulation and melt tumbling fluidized bed granulation. The loose density, angle of repose, granule size distribution, mean diameter of granules, and the tensile strength and porosity of tablets were evaluated. The compactibilities of granules were varied by the granulation methods. However, the difference in compactibility of granules could not be explained due to the difference in compressibility, since there was no difference in Heckel plots due to granulation methods. Among their granule properties, the loose density of granules seemed to have a correlation with the tablet strength regardless of the granulation methods.

Ultrahigh and High Resolution Structures and Mutational Analysis of Monomeric Streptococcus pyogenes SpeB Reveal a Functional Role for the Glycine-rich C-terminal Loop

DOE Office of Scientific and Technical Information (OSTI.GOV)

González-Páez, Gonzalo E.; Wolan, Dennis W.

2012-09-05

Cysteine protease SpeB is secreted from Streptococcus pyogenes and has been studied as a potential virulence factor since its identification almost 70 years ago. Here, we report the crystal structures of apo mature SpeB to 1.06 {angstrom} resolution as well as complexes with the general cysteine protease inhibitor trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane and a novel substrate mimetic peptide inhibitor. These structures uncover conformational changes associated with maturation of SpeB from the inactive zymogen to its active form and identify the residues required for substrate binding. With the use of a newly developed fluorogenic tripeptide substrate to measure SpeB activity, we determined IC{sub 50}more » values for trans-epoxysuccinyl-L-leucylamido(4-guanidino)butane and our new peptide inhibitor and the effects of mutations within the C-terminal active site loop. The structures and mutational analysis suggest that the conformational movements of the glycine-rich C-terminal loop are important for the recognition and recruitment of biological substrates and release of hydrolyzed products.« less

The Lys-Asp-Tyr Triad within the Mite Allergen Der p 1 Propeptide Is a Critical Structural Element for the pH-Dependent Initiation of the Protease Maturation.

PubMed

Chevigné, Andy; Campizi, Vincenzo; Szpakowska, Martyna; Bourry, David; Dumez, Marie-Eve; Martins, José C; Matagne, André; Galleni, Moreno; Jacquet, Alain

2017-05-20

The major house dust mite allergen, Der p 1, is a papain-like cysteine protease expressed as an inactive precursor, proDer p 1, carrying an N-terminal propeptide with a unique structure. The maturation of the zymogen into an enzymatically-active form of Der p 1 is a multistep autocatalytic process initiated under acidic conditions through conformational changes of the propeptide, leading to the loss of its inhibitory ability and its subsequent gradual cleavage. The aims of this study were to characterize the residues present in the Der p 1 propeptide involved in the initiation of the zymogen maturation process, but also to assess the impact of acidic pH on the propeptide structure, the activity of Der p 1 and the fate of the propeptide. Using various complementary enzymatic and structural approaches, we demonstrated that a structural triad K17p-D51p-Y19p within the N-terminal domain of the propeptide is essential for its stabilization and the sensing of pH changes. Particularly, the protonation of D51p under acidic conditions unfolds the propeptide through disruption of the K17p-D51p salt bridge, reduces its inhibition capacity and unmasks the buried residues K17p and Y19p constituting the first maturation cleavage site of the zymogen. Our results also evidenced that this triad acts in a cooperative manner with other propeptide pH-responsive elements, including residues E56p and E80p, to promote the propeptide unfolding and/or to facilitate its proteolysis. Furthermore, we showed that acidic conditions modify Der p 1 proteolytic specificity and confirmed that the formation of the first intermediate represents the limiting step of the in vitro Der p 1 maturation process. Altogether, our results provide new insights into the early events of the mechanism of proDer p 1 maturation and identify a unique structural triad acting as a stabilizing and a pH-sensing regulatory element.

Label-free distinguishing between neurons and glial cells based on two-photon excited fluorescence signal of neuron perinuclear granules

NASA Astrophysics Data System (ADS)

Du, Huiping; Jiang, Liwei; Wang, Xingfu; Liu, Gaoqiang; Wang, Shu; Zheng, Liqin; Li, Lianhuang; Zhuo, Shuangmu; Zhu, Xiaoqin; Chen, Jianxin

2016-08-01

Neurons and glial cells are two critical cell types of brain tissue. Their accurate identification is important for the diagnosis of psychiatric disorders such as depression and schizophrenia. In this paper, distinguishing between neurons and glial cells by using the two-photon excited fluorescence (TPEF) signals of intracellular intrinsic sources was performed. TPEF microscopy combined with TUJ-1 and GFAP immunostaining and quantitative image analysis demonstrated that the perinuclear granules of neurons in the TPEF images of brain tissue and the primary cultured cortical cells were a unique characteristic of neurons compared to glial cells which can become a quantitative feature to distinguish neurons from glial cells. With the development of miniaturized TPEF microscope (‘two-photon fiberscopes’) imaging devices, TPEF microscopy can be developed into an effective diagnostic and monitoring tool for psychiatric disorders such as depression and schizophrenia.

The Mealiness and Quality of Herbal Medicine: Licorice for Example.

PubMed

Liu, Xueying; Hou, Weilong; Dou, Deqiang

2017-01-01

The morphological identification is an effective and simple quality evaluation method in Chinese drugs, and the traits of mealiness and color were widely used in the commercial market of Chinese drugs. The objective of this study was to explore the correlation between mealiness of herbal drugs and its quality; licorice was selected as an example. The mealiness of licorice was graded by its weight; meanwhile, the content of glycyrrhizic acid and liquiritin was determined by high-performance liquid chromatography-diode-array detection method; the content of polysaccharides, soluble sugars, pectin, total starch, amylose, and amylopectin was measured by colorimetric method; and the number and diameter of starch granule were observed by microscope. The results showed that the mealiness of licorice which collected from wild and cultivated plants is positively correlated with the content of glycyrrhizic acid, liquiritin, the ratio of amylose to total starch, and the number of starch granules whose diameter was over 5 μm. However, the mealiness is negatively correlated with the total starch. Further, the formation mechanism of starch granule was discussed. It is for the first time to report the positive correlation between the mealiness and the starch granule size, the ratio of amylose to total starch, which can provide rationality for the quality evaluation using the character of mealiness in herbal medicine. It is a convenient method to justify the quality of herbal medicine. To explore the correlation between mealiness of herbal drugs and its quality, licorice was selected as an example. The result indicated that the effective constituent is correlated with mealiness of licorice. Abbreviations Used: TCM: Traditional Chinese Medicine.

A Patatin-Like Protein Associated with the Polyhydroxyalkanoate (PHA) Granules of Haloferax mediterranei Acts as an Efficient Depolymerase in the Degradation of Native PHA

PubMed Central

Liu, Guiming; Hou, Jing; Cai, Shuangfeng; Zhao, Dahe; Cai, Lei; Han, Jing; Zhou, Jian

2015-01-01

The key enzymes and pathways involved in polyhydroxyalkanoate (PHA) biosynthesis in haloarchaea have been identified in recent years, but the haloarchaeal enzymes for PHA degradation remain unknown. In this study, a patatin-like PHA depolymerase, PhaZh1, was determined to be located on the PHA granules in the haloarchaeon Haloferax mediterranei. PhaZh1 hydrolyzed the native PHA (nPHA) [including native polyhydroxybutyrate (nPHB) and native poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (nPHBV) in this study] granules in vitro with 3-hydroxybutyrate (3HB) monomer as the primary product. The site-directed mutagenesis of PhaZh1 indicated that Gly16, Ser47 (in a classical lipase box, G-X-S47-X-G), and Asp195 of this depolymerase were essential for its activity in nPHA granule hydrolysis. Notably, phaZh1 and bdhA (encoding putative 3HB dehydrogenase) form a gene cluster (HFX_6463 to _6464) in H. mediterranei. The 3HB monomer generated from nPHA degradation by PhaZh1 could be further converted into acetoacetate by BdhA, indicating that PhaZh1-BdhA may constitute the first part of a PHA degradation pathway in vivo. Interestingly, although PhaZh1 showed efficient activity and was most likely the key enzyme in nPHA granule hydrolysis in vitro, the knockout of phaZh1 had no significant effect on the intracellular PHA mobilization, implying the existence of an alternative PHA mobilization pathway(s) that functions effectively within the cells of H. mediterranei. Therefore, identification of this patatin-like depolymerase of haloarchaea may provide a new strategy for producing the high-value-added chiral compound (R)-3HB and may also shed light on the PHA mobilization in haloarchaea. PMID:25710370

Volutin granules of Eimeria parasites are acidic compartments and have physiological and structural characteristics similar to acidocalcisomes

PubMed Central

Medeiros, Lia Carolina Soares; Gomes, Fabio; Maciel, Luis Renato Maia; Seabra, Sergio Henrique; Docampo, Roberto; Moreno, Silvia; Plattner, Helmut; Hentschel, Joachim; Kawazoe, Urara; Barrabin, Hector; de Souza, Wanderley; DaMatta, Renato Augusto; Miranda, Kildare

2012-01-01

The structural organization of parasites has been the subject of investigation by many groups and has lead to the identification of structures and metabolic pathways that may represent targets for anti-parasitic drugs. A specific group of organelles named acidocalcisomes has been identified in a number of organisms, including the apicomplexan parasites such as Toxoplasma and Plasmodium, where they have been shown to be involved in cation homeostasis, polyphosphate metabolism, and osmoregulation. Their structural counterparts in the apicomplexan parasite Eimeria have not been fully characterized. In this work, the ultrastructural and chemical properties of acidocalcisomes in Eimeria were characterized. Electron microscopy analysis of Eimeria parasites showed the dense organelles called volutin granules similar to acidocalcisomes. Immunolocalization of the vacuolar proton pyrophosphatase, considered as a marker for acidocalcisomes, showed labeling in vesicles of size and distribution similar to the dense organelles seen by electron microscopy. Spectrophotometric measurements of the kinetics of proton uptake showed a vacuolar proton pyrophosphatase activity. X-ray mapping revealed significant amounts of Na, Mg, P, K, Ca, and Zn in their matrix. The results suggest that volutin granules of Eimeria parasites are acidic, dense organelles and possess structural and chemical properties analogous to those of other acidocalcisomes, suggesting a similar functional role in these parasites. PMID:21699625

Revisiting the mechanism of coagulation factor XIII activation and regulation from a structure/functional perspective

PubMed Central

Gupta, Sneha; Biswas, Arijit; Akhter, Mohammad Suhail; Krettler, Christoph; Reinhart, Christoph; Dodt, Johannes; Reuter, Andreas; Philippou, Helen; Ivaskevicius, Vytautas; Oldenburg, Johannes

2016-01-01

The activation and regulation of coagulation Factor XIII (FXIII) protein has been the subject of active research for the past three decades. Although discrete evidence exists on various aspects of FXIII activation and regulation a combinatorial structure/functional view in this regard is lacking. In this study, we present results of a structure/function study of the functional chain of events for FXIII. Our study shows how subtle chronological submolecular changes within calcium binding sites can bring about the detailed transformation of the zymogenic FXIII to its activated form especially in the context of FXIIIA and FXIIIB subunit interactions. We demonstrate what aspects of FXIII are important for the stabilization (first calcium binding site) of its zymogenic form and the possible modes of deactivation (thrombin mediated secondary cleavage) of the activated form. Our study for the first time provides a structural outlook of the FXIIIA2B2 heterotetramer assembly, its association and dissociation. The FXIIIB subunits regulatory role in the overall process has also been elaborated upon. In summary, this study provides detailed structural insight into the mechanisms of FXIII activation and regulation that can be used as a template for the development of future highly specific therapeutic inhibitors targeting FXIII in pathological conditions like thrombosis. PMID:27453290

Mechanism of mitochondrial permeability transition pore induction and damage in the pancreas: inhibition prevents acute pancreatitis by protecting production of ATP

PubMed Central

Mukherjee, Rajarshi; Mareninova, Olga A; Odinokova, Irina V; Huang, Wei; Murphy, John; Chvanov, Michael; Javed, Muhammad A; Wen, Li; Booth, David M; Cane, Matthew C; Awais, Muhammad; Gavillet, Bruno; Pruss, Rebecca M; Schaller, Sophie; Molkentin, Jeffery D; Tepikin, Alexei V; Petersen, Ole H; Pandol, Stephen J; Gukovsky, Ilya; Criddle, David N; Gukovskaya, Anna S

2016-01-01

Objective Acute pancreatitis is caused by toxins that induce acinar cell calcium overload, zymogen activation, cytokine release and cell death, yet is without specific drug therapy. Mitochondrial dysfunction has been implicated but the mechanism not established. Design We investigated the mechanism of induction and consequences of the mitochondrial permeability transition pore (MPTP) in the pancreas using cell biological methods including confocal microscopy, patch clamp technology and multiple clinically representative disease models. Effects of genetic and pharmacological inhibition of the MPTP were examined in isolated murine and human pancreatic acinar cells, and in hyperstimulation, bile acid, alcoholic and choline-deficient, ethionine-supplemented acute pancreatitis. Results MPTP opening was mediated by toxin-induced inositol trisphosphate and ryanodine receptor calcium channel release, and resulted in diminished ATP production, leading to impaired calcium clearance, defective autophagy, zymogen activation, cytokine production, phosphoglycerate mutase 5 activation and necrosis, which was prevented by intracellular ATP supplementation. When MPTP opening was inhibited genetically or pharmacologically, all biochemical, immunological and histopathological responses of acute pancreatitis in all four models were reduced or abolished. Conclusions This work demonstrates the mechanism and consequences of MPTP opening to be fundamental to multiple forms of acute pancreatitis and validates the MPTP as a drug target for this disease. PMID:26071131

Circular trimers of gelatinase B/matrix metalloproteinase-9 constitute a distinct population of functional enzyme molecules differentially regulated by tissue inhibitor of metalloproteinases-1

PubMed Central

Vandooren, Jennifer; Born, Benjamin; Solomonov, Inna; Zajac, Ewa; Saldova, Radka; Senske, Michael; Ugarte-Berzal, Estefanía; Martens, Erik; Van den Steen, Philippe E.; Van Damme, Jo; Garcia-Pardo, Angeles; Froeyen, Matheus; Deryugina, Elena I.; Quigley, James P.; Moestrup, Søren K.; Rudd, Pauline M.; Sagi, Irit; Opdenakker, Ghislain

2015-01-01

Gelatinase B/matrix metalloproteinase-9 (MMP-9) (EC 3.4.24.35) cleaves many substrates and is produced by most cell types as a zymogen, proMMP-9, in complex with the tissue inhibitor of metalloproteinases-1 (TIMP-1). Natural proMMP-9 occurs as monomers, homomultimers, and heterocomplexes, but our knowledge about the overall structure of proMMP-9 monomers and multimers is limited. We investigated biochemical, biophysical, and functional characteristics of zymogen and activated forms of MMP-9 monomers and multimers. In contrast to a conventional notion of a dimeric nature of MMP-9 homomultimers, we demonstrate that these are reduction-sensitive trimers. Based on the information from electrophoresis, atomic force microscopy (AFM) and transmission electron microscopy (TEM), we generated a 3Dstructure model of the proMMP-9 trimer. Remarkably, the proMMP-9 trimers possessed a 50-fold higher affinity for TIMP-1 than the monomers. In vivo, this finding was reflected in a higher extent of TIMP-1 inhibition of angiogenesis induced by trimers versus monomers. Our results show that proMMP-9 trimers constitute a novel structural and functional entity that is differentially regulated by TIMP-1. PMID:25360794

Influence of granulating method on physical and mechanical properties, compression behavior, and compactibility of lactose and microcrystalline cellulose granules.

PubMed

Horisawa, E; Danjo, K; Sunada, H

2000-06-01

The physical and mechanical properties of lactose (LC) and microcrystalline cellulose (MCC) granules prepared by various granulating methods were determined, and their effects on the compression and strength of the tablets were examined. From the force-displacement curve obtained in a crushing test on a single granule, all LC granules appeared brittle, and MCC granules were somewhat plastically deformable. Inter-granular porosity epsilon inter clearly decreased with greater spherical granule shape for both materials. Decrease in intragranular porosity epsilon intra enhanced the crushing force of a single granule Fg. Agitating granulation brought about the most compactness and hardness of granules. In granule compression tests, the initial slope of Heckel plots K1 appeared closely related to ease of filling voids in a granule bed by the slippage or rolling of granules. The reciprocal of the slope in the succeeding step 1/K2 in compression of MCC granules indicated positive correlation to Fg, while in LC granules, no such obvious relation was evident. 1/K2 differed only slightly among granulating methods. Tensile strength of tablets Tt obtained by compression of various LC granules was low as a whole and was little influenced by granulating method. For MCC granules, which are plastically deformable, tablet strength greatly depended on granulation. Granules prepared by extruding or dry granulation gave strong tablets. Tablets prepared from granules made by the agitating method showed particularly low Tt. From stereomicroscopic observation, the contact area between granule particles in a tablet appeared smaller; this would explain the decrease in inter-granular bond formation.

Analysis of the release process of phenylpropanolamine hydrochloride from ethylcellulose matrix granules V. Release properties of ethylcellulose layered matrix granules.

PubMed

Fukui, Atsuko; Fujii, Ryuta; Yonezawa, Yorinobu; Sunada, Hisakazu

2008-04-01

In the pharmaceutical preparation of a controlled release drug, it is very important and necessary to understand the release properties. In previous papers, a combination of the square-root time law and cube-root law equations was confirmed to be a useful equation for qualitative treatment. It was also confirmed that the combination equation could analyze the release properties of layered granules as well as matrix granules. The drug release property from layered granules is different from that of matrix granules. A time lag occurs before release, and the entire release property of layered granules was analyzed using the combination of the square-root time law and cube-root law equations. It is considered that the analysis method is very useful and efficient for both matrix and layered granules. Comparing the granulation methods, it is easier to control the manufacturing process by tumbling granulation (method B) than by tumbling-fluidized bed granulation (method C). Ethylcellulose (EC) layered granulation by a fluidized bed granulator might be convenient for the preparation of controlled release dosage forms as compared with a tumbling granulator, because the layered granules prepared by the fluidized bed granulator can granulate and dry at the same time. The time required for drying by the fluidized bed granulator is shorter than that by the tumbling granulator, so the fluidized bed granulator is convenient for preparation of granules in handling and shorter processing time than the tumbling granulator. It was also suggested that the EC layered granules prepared by the fluidized bed granulator were suitable for a controlled release system as well as the EC matrix granules.

Detection of component segregation in granules manufactured by high shear granulation with over-granulation conditions using near-infrared chemical imaging.

PubMed

Koide, Tatsuo; Nagato, Takuya; Kanou, Yoshiyuki; Matsui, Kou; Natsuyama, Susumu; Kawanishi, Toru; Hiyama, Yukio

2013-01-30

The objective of this study was to evaluate the high shear granulation process using near-infrared (NIR) chemical imaging technique and to make the findings available for pharmaceutical development. We prepared granules and tablets made under appropriate- and over-granulation conditions with high shear granulation and observed these granules and tablets using NIR chemical imaging system. We found an interesting phenomenon: lactose agglomeration and segregation of ingredients occurred in experimental tablets when over-granulation conditions, including greater impeller rotation speeds and longer granulation times, were employed. Granules prepared using over-granulation conditions were larger and had progressed to the consolidation stage; segregation between ethenzamide and lactose occurred within larger granules. The segregation observed here is not detectable using conventional analytical technologies such as high pressure liquid chromatography (HPLC) because the content of the granules remained uniform despite the segregation. Therefore, granule visualization using NIR chemical imaging is an effective method for investigating and evaluating the granulation process. Copyright © 2012 Elsevier B.V. All rights reserved.

MODster: Namespaces and Redirection for Earth Science Data

NASA Astrophysics Data System (ADS)

Frew, J.; Metzger, D.; Slaughter, P.

2005-12-01

MODster is a distributed, decentralized inventory server for Earth science data granules (standard units of data content and structure.) MODster connects data granule users (people who know which specific granule they want, but who don't know who has it or how to get it) with data granule providers (people or institutions that keep granules accessible online.) * If you're a provider, you can tell MODster which granules you have and where they live (i.e., their URLs.) * If you're a user, you can ask MODster for a granule, and it will transparently redirect your request to whomever has it. The key to making this work is a standard granule namespace. A granule namespace is a naming convention that associates particular names with particular granules, regardless of where those granules live. Different Earth science data products have their own granule namespaces. For example, in the MODIS granule namespace, the granule name "MOD43A2.A1998365.h5.v8.001.1999001090020.hdf" always refers to version 1 of the 5th horizontal and 8th vertical tile of the Level 3 16-day Bi-directional Reflectance Distribution Function product, acquired by the MODIS Terra sensor on 31 December 1998 and generated on 01 January 1999 at 9:00:20 AM. A MODster URL is simply a standard way of referring to a data product namespace and one of its granules. MODster URLs have the general form "http://server/namespace/granule" where "granule" is a granule name that conforms to a granule namespace, "namespace" is a MODster namespace, which is the name of a granule namespace whose conventions are known to MODster, and "server" is a MODster server, which is an HTTP server that can redirect namespace/granule requests to granule providers. A MODster URL with no granule component gets a description of the MODster namespace, its authority (the persons or institutions responsible for documenting and maintaining the naming convention), and also any services for that MODster namespace that the MODster server supports. Our current MODster implementation allows granule providers to explicitly register their granules, and can also crawl provider sites looking for granules whose names match specific rules or regular expressions.

PHYSICAL PROPERTIES OF LARGE AND SMALL GRANULES IN SOLAR QUIET REGIONS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu Daren; Xie Zongxia; Hu Qinghua

The normal mode observations of seven quiet regions obtained by the Hinode spacecraft are analyzed to study the physical properties of granules. An artificial intelligence technique is introduced to automatically find the spatial distribution of granules in feature spaces. In this work, we investigate the dependence of granular continuum intensity, mean Doppler velocity, and magnetic fields on granular diameter. We recognized 71,538 granules by an automatic segmentation technique and then extracted five properties: diameter, continuum intensity, Doppler velocity, and longitudinal and transverse magnetic flux density to describe the granules. To automatically explore the intrinsic structures of the granules in themore » five-dimensional parameter space, the X-means clustering algorithm and one-rule classifier are introduced to define the rules for classifying the granules. It is found that diameter is a dominating parameter in classifying the granules and two families of granules are derived: small granules with diameters smaller than 1.''44, and large granules with diameters larger than 1.''44. Based on statistical analysis of the detected granules, the following results are derived: (1) the averages of diameter, continuum intensity, and Doppler velocity in the upward direction of large granules are larger than those of small granules; (2) the averages of absolute longitudinal, transverse, and unsigned flux density of large granules are smaller than those of small granules; (3) for small granules, the average of continuum intensity increases with their diameters, while the averages of Doppler velocity, transverse, absolute longitudinal, and unsigned magnetic flux density decrease with their diameters. However, the mean properties of large granules are stable; (4) the intensity distributions of all granules and small granules do not satisfy Gaussian distribution, while that of large granules almost agrees with normal distribution with a peak at 1.04 I{sub 0}.« less

Propeptides are sufficient to regulate organelle-specific pH-dependent activation of furin and proprotein convertase 1/3.

PubMed

Dillon, Stephanie L; Williamson, Danielle M; Elferich, Johannes; Radler, David; Joshi, Rajendra; Thomas, Gary; Shinde, Ujwal

2012-10-12

The proprotein convertases (PCs) furin and proprotein convertase 1/3 (PC1) cleave substrates at dibasic residues along the eukaryotic secretory/endocytic pathway. PCs are evolutionarily related to bacterial subtilisin and are synthesized as zymogens. They contain N-terminal propeptides (PRO) that function as dedicated catalysts that facilitate folding and regulate activation of cognate proteases through multiple-ordered cleavages. Previous studies identified a histidine residue (His69) that functions as a pH sensor in the propeptide of furin (PRO(FUR)), which regulates furin activation at pH~6.5 within the trans-Golgi network. Although this residue is conserved in the PC1 propeptide (PRO(PC1)), PC1 nonetheless activates at pH~5.5 within the dense core secretory granules. Here, we analyze the mechanism by which PRO(FUR) regulates furin activation and examine why PRO(FUR) and PRO(PC1) differ in their pH-dependent activation. Sequence analyses establish that while both PRO(FUR) and PRO(PC1) are enriched in histidines when compared with cognate catalytic domains and prokaryotic orthologs, histidine content in PRO(FUR) is ~2-fold greater than that in PRO(PC1), which may augment its pH sensitivity. Spectroscopy and molecular dynamics establish that histidine protonation significantly unfolds PRO(FUR) when compared to PRO(PC1) to enhance autoproteolysis. We further demonstrate that PRO(FUR) and PRO(PC1) are sufficient to confer organelle sensing on folding and activation of their cognate proteases. Swapping propeptides between furin and PC1 transfers pH-dependent protease activation in a propeptide-dictated manner in vitro and in cells. Since prokaryotes lack organelles and eukaryotic PCs evolved from propeptide-dependent, not propeptide-independent prokaryotic subtilases, our results suggest that histidine enrichment may have enabled propeptides to evolve to exploit pH gradients to activate within specific organelles. Copyright © 2012 Elsevier Ltd. All rights reserved.

Serological diagnosis and prognosis of severe acute pancreatitis by analysis of serum glycoprotein 2.

PubMed

Roggenbuck, Dirk; Goihl, Alexander; Hanack, Katja; Holzlöhner, Pamela; Hentschel, Christian; Veiczi, Miklos; Schierack, Peter; Reinhold, Dirk; Schulz, Hans-Ulrich

2017-05-01

Glycoprotein 2 (GP2), the pancreatic major zymogen granule membrane glycoprotein, was reported to be elevated in acute pancreatitis in animal models. Enzyme-linked immunosorbent assays (ELISAs) were developed to evaluate human glycoprotein 2 isoform alpha (GP2a) and total GP2 (GP2t) as specific markers for acute pancreatitis in sera of 153 patients with acute pancreatitis, 26 with chronic pancreatitis, 125 with pancreatic neoplasms, 324 with non-pancreatic neoplasms, 109 patients with liver/biliary disease, 67 with gastrointestinal disease, and 101 healthy subjects. GP2a and GP2t levels were correlated with procalcitonin and C-reactive protein in 152 and 146 follow-up samples of acute pancreatitis patients, respectively. The GP2a ELISA revealed a significantly higher assay accuracy in contrast to the GP2t assay (sensitivity ≤3 disease days: 91.7%, specificity: 96.7%, positive likelihood ratio [LR+]: 24.6, LR-: 0.09). GP2a and GP2t levels as well as prevalences were significantly elevated in early acute pancreatitis (≤3 disease days) compared to all control cohorts (p<0.05, respectively). GP2a and GP2t levels were significantly higher in patients with severe acute pancreatitis at admission compared with mild cases (p<0.05, respectively). Odds ratio for GP2a regarding mild vs. severe acute pancreatitis with lethal outcome was 7.8 on admission (p=0.0222). GP2a and GP2t levels were significantly correlated with procalcitonin [Spearman's rank coefficient of correlation (ρ)=0.21, 0.26; p=0.0110, 0.0012; respectively] and C-reactive protein (ρ=0.37, 0.40; p<0.0001; respectively). Serum GP2a is a specific marker of acute pancreatitis and analysis of GP2a can aid in the differential diagnosis of acute upper abdominal pain and prognosis of severe acute pancreatitis.

Vesicular trafficking of immune mediators in human eosinophils revealed by immunoelectron microscopy.

PubMed

Melo, Rossana C N; Weller, Peter F

2016-10-01

Electron microscopy (EM)-based techniques are mostly responsible for our current view of cell morphology at the subcellular level and continue to play an essential role in biological research. In cells from the immune system, such as eosinophils, EM has helped to understand how cells package and release mediators involved in immune responses. Ultrastructural investigations of human eosinophils enabled visualization of secretory processes in detail and identification of a robust, vesicular trafficking essential for the secretion of immune mediators via a non-classical secretory pathway associated with secretory (specific) granules. This vesicular system is mainly organized as large tubular-vesicular carriers (Eosinophil Sombrero Vesicles - EoSVs) actively formed in response to cell activation and provides a sophisticated structural mechanism for delivery of granule-stored mediators. In this review, we highlight the application of EM techniques to recognize pools of immune mediators at vesicular compartments and to understand the complex secretory pathway within human eosinophils involved in inflammatory and allergic responses. Copyright © 2016 Elsevier Inc. All rights reserved.

Isolation and identification of poly beta hydroxybutyric acid accumulating bacteria of Staphylococcal sp. and characterization of biodegradable polyester.

PubMed

Roy, Bappaditya; Banerjee, Rajat; Chatterjee, Sumana

2009-04-01

Staphylococcus sp. strain BP/SU1, capable of degrading the biopolymer and utilize it as a source of carbon and energy, was isolated from activated sludge using METABOLIX (MBX D411G). It was found that this strain was capable of accumulating poly(3-hydroxybutyric acid) P(3-HB), as granule poly (3-hydroxybutyric acid), p(3-HB), inclusion bodies when grown under suitable nutrient conditions. These strains could sustain cell growth up to a dry mass of 9.24 g/l with a doubling time of 8 to 10 hr and could accumulate P(3-HB) as granular inclusion bodies to a cell dry weight of more than 12%. P(3-HB) accumulated by this organism was isolated and characterized through NMR, FT-IR spectroscopy, UV Spectroscopy, Mass spectroscopy and Differential Scanning Calorimetry. P(3-HB) granules so isolated showed physical and chemical properties that should be possessed by a superior quality thermoplastic biopolymer.

Identification of Neuregulin-2 as a novel stress granule component.

PubMed

Kim, Jin Ah; Jayabalan, Aravinth Kumar; Kothandan, Vinoth Kumar; Mariappan, Ramesh; Kee, Younghoon; Ohn, Takbum

2016-08-01

Stress Granules (SGs) are microscopically visible, phase dense aggregates of translationally stalled messenger ribonucleoprotein (mRNP) complexes formed in response to distinct stress conditions. It is generally considered that SG formation is induced to protect cells from conditions of stress. The precise constituents of SGs and the mechanism through which SGs are dynamically regulated in response to stress are not completely understood. Hence, it is important to identify proteins which regulate SG assembly and disassembly. In the present study, we report Neuregulin-2 (NRG2) as a novel component of SGs; furthermore, depletion of NRG2 potently inhibits SG formation. We also demonstrate that NRG2 specifically localizes to SGs under various stress conditions. Knockdown of NRG2 has no effect on stress-induced polysome disassembly, suggesting that the component does not influence early step of SG formation. It was also observed that reduced expression of NRG2 led to marginal increase in cell survival under arsenite-induced stress. [BMB Reports 2016; 49(8): 449-454].

Identification of Neuregulin-2 as a novel stress granule component

PubMed Central

Kim, Jin Ah; Jayabalan, Aravinth Kumar; Kothandan, Vinoth Kumar; Mariappan, Ramesh; Kee, Younghoon; Ohn, Takbum

2016-01-01

Stress Granules (SGs) are microscopically visible, phase dense aggregates of translationally stalled messenger ribonucleoprotein (mRNP) complexes formed in response to distinct stress conditions. It is generally considered that SG formation is induced to protect cells from conditions of stress. The precise constituents of SGs and the mechanism through which SGs are dynamically regulated in response to stress are not completely understood. Hence, it is important to identify proteins which regulate SG assembly and disassembly. In the present study, we report Neuregulin-2 (NRG2) as a novel component of SGs; furthermore, depletion of NRG2 potently inhibits SG formation. We also demonstrate that NRG2 specifically localizes to SGs under various stress conditions. Knockdown of NRG2 has no effect on stress-induced polysome disassembly, suggesting that the component does not influence early step of SG formation. It was also observed that reduced expression of NRG2 led to marginal increase in cell survival under arsenite-induced stress. [BMB Reports 2016; 49(8): 449-454] PMID:27345716

The life cycle of platelet granules.

PubMed

Sharda, Anish; Flaumenhaft, Robert

2018-01-01

Platelet granules are unique among secretory vesicles in both their content and their life cycle. Platelets contain three major granule types-dense granules, α-granules, and lysosomes-although other granule types have been reported. Dense granules and α-granules are the most well-studied and the most physiologically important. Platelet granules are formed in large, multilobulated cells, termed megakaryocytes, prior to transport into platelets. The biogenesis of dense granules and α-granules involves common but also distinct pathways. Both are formed from the trans -Golgi network and early endosomes and mature in multivesicular bodies, but the formation of dense granules requires trafficking machinery different from that of α-granules. Following formation in the megakaryocyte body, both granule types are transported through and mature in long proplatelet extensions prior to the release of nascent platelets into the bloodstream. Granules remain stored in circulating platelets until platelet activation triggers the exocytosis of their contents. Soluble N -ethylmaleimide-sensitive factor attachment protein receptor (SNARE) proteins, located on both the granules and target membranes, provide the mechanical energy that enables membrane fusion during both granulogenesis and exocytosis. The function of these core fusion engines is controlled by SNARE regulators, which direct the site, timing, and extent to which these SNAREs interact and consequently the resulting membrane fusion. In this review, we assess new developments in the study of platelet granules, from their generation to their exocytosis.

Mechanism of the formation of hollow spherical granules using a high shear granulator.

PubMed

Asada, Takumi; Nishikawa, Mitsunori; Ochiai, Yasushi; Noguchi, Shuji; Kimura, Shin-Ichiro; Iwao, Yasunori; Itai, Shigeru

2018-05-30

Recently, we have developed a novel granulation technology to manufacture hollow spherical granules (HSGs) for controlled-release formulations; however, the mechanism of the granulation is still unclear. The aim of this study is to determine the mechanism of the formation of the HSGs using a high shear granulator. Samples of granulated material were collected at various times during granulation and were investigated using scanning electron microscope and X-ray computed tomography. It was observed that the granulation proceeded by drug layering to the polymer, followed by formation of a hollow in the granule. In addition, it was also found that generation of a crack in the adhered drug layer and air flow into the granules might be involved in forming the hollow in the structure. Observation of the granulation of formulations with different types of drugs and polymers indicated that negative pressure in the granules occurred and the granules caved in when the hollow was formed. The hollow-forming speed and the shell density of the hollow granules depended on the particular drug and polymer. Taken together, the granulation mechanism of HSGs was determined and this information will be valuable for HSGs technology development. Copyright © 2018 Elsevier B.V. All rights reserved.

Porosome: The Universal Secretory Portal in Cells

NASA Astrophysics Data System (ADS)

Jena, Bhanu

2012-10-01

In the past 50 years it was believed that during cell secretion, membrane-bound secretory vesicles completely merge at the cell plasma membrane resulting in the diffusion of intra-vesicular contents to the cell exterior and the compensatory retrieval of the excess membrane by endocytosis. This explanation made no sense or logic, since following cell secretion partially empty vesicles accumulate as demonstrated in electron micrographs. Furthermore, with the ``all or none'' mechanism of cell secretion by complete merger of secretory vesicle membrane at the cell plasma membrane, the cell is left with little regulation and control of the amount of content release. Moreover, it makes no sense for mammalian cells to possess such `all or none' mechanism of cell secretion, when even single-cell organisms have developed specialized and sophisticated secretory machinery, such as the secretion apparatus of Toxoplasma gondii, the contractile vacuoles in paramecium, or the various types of secretory structures in bacteria. Therefore, in 1993 in a News and Views article in Nature, E. Neher wrote ``It seems terribly wasteful that, during the release of hormones and neurotransmitters from a cell, the membrane of a vesicle should merge with the plasma membrane to be retrieved for recycling only seconds or minutes later.'' This conundrum in the molecular mechanism of cell secretion was finally resolved in 1997 following discovery of the ``Porosome,'' the universal secretory machinery in cells. Porosomes are supramolecular lipoprotein structures at the cell plasma membrane, where membrane-bound secretory vesicles transiently dock and fuse to release inravesicular contents to the outside during cell secretion. In the past decade, the composition of the porosome, its structure and dynamics at nm resolution and in real time, and its functional reconstitution into artificial lipid membrane, have all been elucidated. Since porosomes in exocrine and neuroendocrine cells measure 100-180 nm, and only 20-45% increase in porosome diameter is demonstrated following the docking and fusion of 0.2-1.2 μm in diameter secretory vesicles, it is concluded that secretory vesicles ``transiently'' dock and fuse, rather than completely merge at the base of the porosome complex to release their contents to the outside. In agreement, it has been demonstrated that ``secretory granules are recaptured largely intact after stimulated exocytosis in cultured endocrine cells''; that ``single synaptic vesicles fuse transiently and successively without loss of identity''; and that``zymogen granule (the secretory vesicle in exocrine pancreas) exocytosis is characterized by long fusion pore openings and preservation of vesicle lipid identity.'' In this presentation, the discovery of the porosome, resulting in a paradigm shift in our understanding of cell secretion will be briefly discussed.

Dimerization of Matrix Metalloproteinase-2 (MMP-2)

PubMed Central

Koo, Bon-Hun; Kim, Yeon Hyang; Han, Jung Ho; Kim, Doo-Sik

2012-01-01

Matrix metalloproteinase-2 (MMP-2) functions in diverse biological processes through the degradation of extracellular and non-extracellular matrix molecules. Because of its potential for tissue damage, there are several ways to regulate MMP-2 activity, including gene expression, compartmentalization, zymogen activation, and enzyme inactivation by extracellular inhibitors. Enzyme regulation through zymogen activation is important for the regulation of MMP-2 activity. In our previous studies, we showed that thrombin directly cleaved the propeptide of MMP-2 at specific sites for enzyme activation. We also demonstrated that heparan sulfate was required for thrombin-mediated activation of pro-MMP-2 by binding to thrombin, presumably through conformational changes at the active site of the enzyme. This suggests a regulatory mechanism for thrombin-mediated activation of pro-MMP-2. In this study, we found that MMP-2 formed a reduction-sensitive homodimer in a controlled manner and that Ca2+ ion was essential for homodimerization of MMP-2. Homodimerization was not associated with protein kinase C-mediated phosphorylation of MMP-2. MMP-2 formed a homodimer through an intermolecular disulfide bond between Cys102 and the neighboring Cys102. Homodimerization of MMP-2 enhanced thrombin-mediated activation of pro-MMP-2. Moreover, the MMP-2 homodimer could cleave a small peptide substrate without removal of the propeptide. Taken together, our experimental data suggest a novel regulatory mechanism for pro-MMP-2 activation that is modulated through homodimerization of MMP-2. PMID:22577146

Digestive peptidase evolution in holometabolous insects led to a divergent group of enzymes in Lepidoptera.

PubMed

Dias, Renata O; Via, Allegra; Brandão, Marcelo M; Tramontano, Anna; Silva-Filho, Marcio C

2015-03-01

Trypsins and chymotrypsins are well-studied serine peptidases that cleave peptide bonds at the carboxyl side of basic and hydrophobic L-amino acids, respectively. These enzymes are largely responsible for the digestion of proteins. Three primary processes regulate the activity of these peptidases: secretion, precursor (zymogen) activation and substrate-binding site recognition. Here, we present a detailed phylogenetic analysis of trypsins and chymotrypsins in three orders of holometabolous insects and reveal divergent characteristics of Lepidoptera enzymes in comparison with those of Coleoptera and Diptera. In particular, trypsin subsite S1 was more hydrophilic in Lepidoptera than in Coleoptera and Diptera, whereas subsites S2-S4 were more hydrophobic, suggesting different substrate preferences. Furthermore, Lepidoptera displayed a lineage-specific trypsin group belonging only to the Noctuidae family. Evidence for facilitated trypsin auto-activation events were also observed in all the insect orders studied, with the characteristic zymogen activation motif complementary to the trypsin active site. In contrast, insect chymotrypsins did not seem to have a peculiar evolutionary history with respect to their mammal counterparts. Overall, our findings suggest that the need for fast digestion allowed holometabolous insects to evolve divergent groups of peptidases with high auto-activation rates, and highlight that the evolution of trypsins led to a most diverse group of enzymes in Lepidoptera. Copyright © 2015 Elsevier Ltd. All rights reserved.

Exosites in the substrate specificity of blood coagulation reactions.

PubMed

Bock, P E; Panizzi, P; Verhamme, I M A

2007-07-01

The specificity of blood coagulation proteinases for substrate, inhibitor, and effector recognition is mediated by exosites on the surfaces of the catalytic domains, physically separated from the catalytic site. Some thrombin ligands bind specifically to either exosite I or II, while others engage both exosites. The involvement of different, overlapping constellations of exosite residues enables binding of structurally diverse ligands. The flexibility of the thrombin structure is central to the mechanism of complex formation and the specificity of exosite interactions. Encounter complex formation is driven by electrostatic ligand-exosite interactions, followed by conformational rearrangement to a stable complex. Exosites on some zymogens are in low affinity proexosite states and are expressed concomitant with catalytic site activation. The requirement for exosite expression controls the specificity of assembly of catalytic complexes on the coagulation pathway, such as the membrane-bound factor Xa*factor Va (prothrombinase) complex, and prevents premature assembly. Substrate recognition by prothrombinase involves a two-step mechanism with initial docking of prothrombin to exosites, followed by a conformational change to engage the FXa catalytic site. Prothrombin and its activation intermediates bind prothrombinase in two alternative conformations determined by the zymogen to proteinase transition that are hypothesized to involve prothrombin (pro)exosite I interactions with FVa, which underpin the sequential activation pathway. The role of exosites as the major source of substrate specificity has stimulated development of exosite-targeted anticoagulants for treatment of thrombosis.

A novel two-chain IGF-II-derived peptide from purified β-cell granules.

PubMed

Buchanan, Christina M; Phillips, Anthony R J; Cooper, Garth J S

2010-10-01

Insulin-like growth factor II (IGF-II) is a potent mitogen that regulates prenatal growth and development in both humans and rodents. Its role in post-natal life is less clear although immunohistochemical studies have observed IGF-II-like immunoreactivity (IGF-II-LI) associated with insulin-producing pancreatic β-cells. Here we isolated secretory granules from a β-cell line, βTC6-F7, and characterized the nature of the IGF-II-LI located therein. Secretory granules were isolated from cultured mouse βTC6-F7 cells by ultracentrifugation. Granule protein content was separated by reversed-phase HPLC, and assayed for IGF-II (radioimmunoassay) prior to identification by gas-phase NH(2)-terminal sequencing and MALDI-TOF MS. Effects of glucose incorporation into muscle glycogen were determined by incubating with isolated rat soleus muscle strips. βTC6-F7 cells contained 60 ± 8 pmol of IGF-II-LI per 10⁶ cells compared to 340 ± 44 pmol insulin-LI per 10⁶ cells. IGF-II immunoreactive fractions were found to contain an IGF-II-like molecule with a molecular mass of 6847.6 Da. The protein was found to be a two-chain insulin-like product of Igf2 that corresponds to mouse des(37-40)IGF-II, which we termed 'vesiculin'. This molecule was also detectable in βTC6-F7 cells by intact-cell mass spectrometry. Mouse vesiculin evoked concentration-dependent stimulation of muscle glycogen synthesis ex vivo with an EC(50) value of 131 nM ± 1.35. Vesiculin, des(37-40)IGF-II, is a novel two-chain insulin-like hormone and the major "IGF-II-like" peptide found in purified mouse βTC6-F7 secretory granules. It stimulated ex vivo muscle glycogen synthesis with an efficacy greater than or equal to the intrinsic potency of IGF-II when compared to insulin derived from the same species. Copyright © 2010 Growth Hormone Research Society. Published by Elsevier Ltd. All rights reserved.

Investigation of Physicochemical Drug Properties to Prepare Fine Globular Granules Composed of Only Drug Substance in Fluidized Bed Rotor Granulation.

PubMed

Mise, Ryohei; Iwao, Yasunori; Kimura, Shin-Ichiro; Osugi, Yukiko; Noguchi, Shuji; Itai, Shigeru

2015-01-01

The effect of some drug properties (wettability and particle size distribution) on granule properties (mean particle size, particle size distribution, sphericity, and granule strength) were investigated in a high (>97%) drug-loading formulation using fluidized bed rotor granulation. Three drugs: acetaminophen (APAP); ibuprofen (IBU); and ethenzamide (ETZ) were used as model drugs based on their differences in wettability and particle size distribution. Granules with mean particle sizes of 100-200 µm and a narrow particle size distribution (PSD) could be prepared regardless of the drug used. IBU and ETZ granules showed a higher sphericity than APAP granules, while APAP and ETZ granules exhibited higher granule strength than IBU. The relationship between drug and granule properties suggested that the wettability and the PSD of the drugs were critical parameters affecting sphericity and granule strength, respectively. Furthermore, the dissolution profiles of granules prepared with poorly water-soluble drugs (IBU and ETZ) showed a rapid release (80% release in 20 min) because of the improved wettability with granulation. The present study demonstrated for the first time that fluidized bed rotor granulation can prepare high drug-loaded (>97%) globular granules with a mean particle size of less than 200 µm and the relationship between physicochemical drug properties and the properties of the granules obtained could be readily determined, indicating the potential for further application of this methodology to various drugs.

Phenotypic properties and microbial diversity of methanogenic granules from a full-scale upflow anaerobic sludge bed reactor treating brewery wastewater.

PubMed

Díaz, Emiliano E; Stams, Alfons J M; Amils, Ricardo; Sanz, José L

2006-07-01

Methanogenic granules from an anaerobic bioreactor that treated wastewater of a beer brewery consisted of different morphological types of granules. In this study, the microbial compositions of the different granules were analyzed by molecular microbiological techniques: cloning, denaturing gradient gel electrophoresis and fluorescent in situ hybridization (FISH), and scanning and transmission electron microscopy. We propose here that the different types of granules reflect the different stages in the life cycle of granules. Young granules were small, black, and compact and harbored active cells. Gray granules were the most abundant granules. These granules have a multilayer structure with channels and void areas. The core was composed of dead or starving cells with low activity. The brown granules, which were the largest granules, showed a loose and amorphous structure with big channels that resulted in fractured zones and corresponded to the older granules. Firmicutes (as determined by FISH) and Nitrospira and Deferribacteres (as determined by cloning and sequencing) were the predominant Bacteria. Remarkably, Firmicutes could not be detected in the brown granules. The methanogenic Archaea identified were Methanosaeta concilii (70 to 90% by FISH and cloning), Methanosarcina mazei, and Methanospirillum spp. The phenotypic appearance of the granules reflected the physiological condition of the granules. This may be valuable to easily select appropriate seed sludges to start up other reactors.

The combined effect of wet granulation process parameters and dried granule moisture content on tablet quality attributes.

PubMed

Gabbott, Ian P; Al Husban, Farhan; Reynolds, Gavin K

2016-09-01

A pharmaceutical compound was used to study the effect of batch wet granulation process parameters in combination with the residual moisture content remaining after drying on granule and tablet quality attributes. The effect of three batch wet granulation process parameters was evaluated using a multivariate experimental design, with a novel constrained design space. Batches were characterised for moisture content, granule density, crushing strength, porosity, disintegration time and dissolution. Mechanisms of the effect of the process parameters on the granule and tablet quality attributes are proposed. Water quantity added during granulation showed a significant effect on granule density and tablet dissolution rate. Mixing time showed a significant effect on tablet crushing strength, and mixing speed showed a significant effect on the distribution of tablet crushing strengths obtained. The residual moisture content remaining after granule drying showed a significant effect on tablet crushing strength. The effect of moisture on tablet tensile strength has been reported before, but not in combination with granulation parameters and granule properties, and the impact on tablet dissolution was not assessed. Correlations between the energy input during granulation, the density of granules produced, and the quality attributes of the final tablets were also identified. Understanding the impact of the granulation and drying process parameters on granule and tablet properties provides a basis for process optimisation and scaling. Copyright © 2016 Elsevier B.V. All rights reserved.

Microbial community variation in cryoconite granules on Qaanaaq Glacier, NW Greenland.

PubMed

Uetake, Jun; Tanaka, Sota; Segawa, Takahiro; Takeuchi, Nozomu; Nagatsuka, Naoko; Motoyama, Hideaki; Aoki, Teruo

2016-09-01

Cryoconite granules are aggregations of microorganisms with mineral particles that form on glacier surfaces. To understand the processes by which the granules develop, this study focused on the altitudinal distribution of the granules and photosynthetic microorganisms on the glacier, bacterial community variation with granules size and environmental factors affecting the growth of the granules. Size-sorted cryoconite granules collected from five different sites on Qaanaaq Glacier were analyzed. C and N contents were significantly higher in large (diameter greater than 250 μm) granules than in smaller (diameter 30-249 μm) granules. Bacterial community structures, based on 16S rRNA gene amplicon sequencing, were different between the smaller and larger granules. The filamentous cyanobacterium Phormidesmis priestleyi was the dominant bacterial species in larger granules. Multivariate analysis suggests that the abundance of mineral particles on the glacier surface is the main factor controlling growth of these cyanobacteria. These results show that the supply of mineral particles on the glacier enhances granule development, that P. priestleyi is likely the key species for primary production and the formation of the granules and that the bacterial community in the granules changes over the course of the granule development. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Cytoplasmic RNA Granules in Somatic Maintenance.

PubMed

Moujaber, Ossama; Stochaj, Ursula

2018-05-30

Cytoplasmic RNA granules represent subcellular compartments that are enriched in protein-bound RNA species. RNA granules are produced by evolutionary divergent eukaryotes, including yeast, mammals, and plants. The functions of cytoplasmic RNA granules differ widely. They are dictated by the cell type and physiological state, which in turn is determined by intrinsic cell properties and environmental factors. RNA granules provide diverse cellular functions. However, all of the granules contribute to aspects of RNA metabolism. This is exemplified by transcription, RNA storage, silencing, and degradation, as well as mRNP remodeling and regulated translation. Several forms of cytoplasmic mRNA granules are linked to normal physiological processes. For instance, they may coordinate protein synthesis and thereby serve as posttranscriptional "operons". RNA granules also participate in cytoplasmic mRNA trafficking, a process particularly well understood for neurons. Many forms of RNA granules support the preservation of somatic cell performance under normal and stress conditions. On the other hand, severe insults or disease can cause the formation and persistence of RNA granules that contribute to cellular dysfunction, especially in the nervous system. Neurodegeneration and many other diseases linked to RNA granules are associated with aging. Nevertheless, information related to the impact of aging on the various types of RNA granules is presently very limited. This review concentrates on cytoplasmic RNA granules and their role in somatic cell maintenance. We summarize the current knowledge on different types of RNA granules in the cytoplasm, their assembly and function under normal, stress, or disease conditions. Specifically, we discuss processing bodies, neuronal granules, stress granules, and other less characterized cytoplasmic RNA granules. Our focus is primarily on mammalian and yeast models, because they have been critical to unravel the physiological role of various RNA granules. RNA granules in plants and pathogens are briefly described. We conclude our viewpoint by summarizing the emerging concepts for RNA granule biology and the open questions that need to be addressed in future studies. © 2018 S. Karger AG, Basel.

The herpes simplex virus 2 virion-associated ribonuclease vhs interferes with stress granule formation.

PubMed

Finnen, Renée L; Hay, Thomas J M; Dauber, Bianca; Smiley, James R; Banfield, Bruce W

2014-11-01

In a previous study, it was observed that cells infected with herpes simplex virus 2 (HSV-2) failed to accumulate stress granules (SGs) in response to oxidative stress induced by arsenite treatment. As a follow-up to this observation, we demonstrate here that disruption of arsenite-induced SG formation by HSV-2 is mediated by a virion component. Through studies on SG formation in cells infected with HSV-2 strains carrying defective forms of UL41, the gene that encodes vhs, we identify vhs as a virion component required for this disruption. Cells infected with HSV-2 strains producing defective forms of vhs form SGs spontaneously late in infection. In addition to core SG components, these spontaneous SGs contain the viral immediate early protein ICP27 as well as the viral serine/threonine kinase Us3. As part of these studies, we reexamined the frameshift mutation known to reside within the UL41 gene of HSV-2 strain HG52. We demonstrate that this mutation is unstable and can rapidly revert to restore wild-type UL41 following low-multiplicity passaging. Identification of the involvement of virion-associated vhs in the disruption of SG formation will enable mechanistic studies on how HSV-2 is able to counteract antiviral stress responses early in infection. In addition, the ability of Us3 to localize to stress granules may indicate novel roles for this viral kinase in the regulation of translation. Eukaryotic cells respond to stress by rapidly shutting down protein synthesis and storing mRNAs in cytoplasmic stress granules (SGs). Stoppages in protein synthesis are problematic for all viruses as they rely on host cell machinery to synthesize viral proteins. Thus, many viruses target SGs for disruption or modification. Infection by herpes simplex virus 2 (HSV-2) was previously observed to disrupt SG formation induced by oxidative stress. In this follow-up study, we identify virion host shutoff protein (vhs) as a viral protein involved in this disruption. The identification of a specific viral protein involved in disrupting SG formation is a key step toward understanding how HSV-2 interacts with these antiviral structures. Additionally, this understanding may provide insights into the biology of SGs that may find application in studies on human motor neuron degenerative diseases, like amyotrophic lateral sclerosis (ALS), which may arise as a result of dysregulation of SG formation. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Numerical analysis of temperature field in the high speed rotary dry-milling process

NASA Astrophysics Data System (ADS)

Wu, N. X.; Deng, L. J.; Liao, D. H.

2018-01-01

For the effect of the temperature field in the ceramic dry granulation. Based on the Euler-Euler mathematical model, at the same time, made ceramic dry granulation experiment equipment more simplify and established physical model, the temperature of the dry granulation process was simulated with the granulation time. The relationship between the granulation temperature and granulation effect in dry granulation process was analyzed, at the same time, the correctness of numerical simulation was verified by measuring the fluidity index of ceramic bodies. Numerical simulation and experimental results showed that when granulation time was 4min, 5min, 6min, maximum temperature inside the granulation chamber was: 70°C, 85°C, 95°C. And the equilibrium of the temperature in the granulation chamber was weakened, the fluidity index of the billet particles was: 56.4. 89.7. 81.6. Results of the research showed that when granulation time was 5min, the granulation effect was best. When the granulation chamber temperature was more than 85°C, the fluidity index and the effective particles quantity of the billet particles were reduced.

CaMKIIalpha 3' untranslated region-directed mRNA translocation in living neurons: visualization by GFP linkage

NASA Technical Reports Server (NTRS)

Rook, M. S.; Lu, M.; Kosik, K. S.

2000-01-01

The CaMKIIalpha mRNA extends into distal hippocampal dendrites, and the 3' untranslated region (3'UTR) is sufficient to mediate this localization. We labeled the 3'UTR of the CaMKIIalpha mRNA in hippocampal cultures by using a green fluorescent protein (GFP)/MS2 bacteriophage tagging system. The CaMKIIalpha 3'UTR formed discrete granules throughout the dendrites of transfected cells. The identity of the fluorescent granules was verified by in situ hybridization. Over 30 min time periods these granules redistributed without a net increase in granule number; with depolarization there is a tendency toward increased numbers of granules in the dendrites. These observations suggest that finer time resolution of granule motility might reveal changes in the motility characteristics of granules after depolarization. So that motile granules could be tracked, shorter periods of observation were required. The movements of motile granules can be categorized as oscillatory, unidirectional anterograde, or unidirectional retrograde. Colocalization of CaMKIIalpha 3'UTR granules and synapses suggested that oscillatory movements allowed the granules to sample several local synapses. Neuronal depolarization increased the number of granules in the anterograde motile pool. Based on the time frame over which the granule number increased, the translocation of granules may serve to prepare the dendrite for mounting an adequate local translation response to future stimuli. Although the resident pool of granules can respond to signals that induce local translation, the number of granules in a dendrite might reflect its activation history.

Effect of granular activated carbon on the aerobic granulation of sludge and its mechanism.

PubMed

Tao, Jia; Qin, Lian; Liu, Xiaoying; Li, Bolin; Chen, Junnan; You, Juan; Shen, Yitian; Chen, Xiaoguo

2017-07-01

The granulation of activated sludge and effect of granular activated carbon (GAC) was investigated under the alternative anaerobic and aerobic conditions. The results showed that GAC accelerated the granulation, but had no obvious effect on the bacterial community structure of granules. The whole granulation process could be categorized into three phases, i.e. lag, granulation and granule maturation phase. During lag period GAC provided nuclei for sludge to attach, and thus enhanced the morphological regularization of sludge. During granulation period the granule size increased significantly due to the growth of bacteria in granules. GAC reduced the compression caused by the inter-particle collisions and thus accelerate the granulation. GAC has no negative effect on the performance of SBR, and thus efficient simultaneous removal of COD, nitrogen and phosphorus were obtained during most of the operating time. Copyright © 2017. Published by Elsevier Ltd.

Breakage and drying behaviour of granules in a continuous fluid bed dryer: Influence of process parameters and wet granule transfer.

PubMed

De Leersnyder, F; Vanhoorne, V; Bekaert, H; Vercruysse, J; Ghijs, M; Bostijn, N; Verstraeten, M; Cappuyns, P; Van Assche, I; Vander Heyden, Y; Ziemons, E; Remon, J P; Nopens, I; Vervaet, C; De Beer, T

2018-03-30

Although twin screw granulation has already been widely studied in recent years, only few studies addressed the subsequent continuous drying which is required after wet granulation and still suffers from a lack of detailed understanding. The latter is important for optimisation and control and, hence, a cost-effective practical implementation. Therefore, the aim of the current study is to increase understanding of the drying kinetics and the breakage and attrition phenomena during fluid bed drying after continuous twin screw granulation. Experiments were performed on a continuous manufacturing line consisting of a twin-screw granulator, a six-segmented fluid bed dryer, a mill, a lubricant blender and a tablet press. Granulation parameters were fixed in order to only examine the effect of drying parameters (filling time, drying time, air flow, drying air temperature) on the size distribution and moisture content of granules (both of the entire granulate and of size fractions). The wet granules were transferred either gravimetrically or pneumatically from the granulator exit to the fluid bed dryer. After a certain drying time, the moisture content reached an equilibrium. This drying time was found to depend on the applied airflow, drying air temperature and filling time. The moisture content of the granules decreased with an increasing drying time, airflow and drying temperature. Although smaller granules dried faster, the multimodal particle size distribution of the granules did not compromise uniform drying of the granules when the target moisture content was achieved. Extensive breakage of granules was observed during drying. Especially wet granules were prone to breakage and attrition during pneumatic transport, either in the wet transfer line or in the dry transfer line. Breakage and attrition of granules during transport and drying should be anticipated early on during process and formulation development by performing integrated experiments on the granulator, dryer and mill. Copyright © 2018 Elsevier B.V. All rights reserved.

Cellular stress induces cytoplasmic RNA granules in fission yeast.

PubMed

Nilsson, Daniel; Sunnerhagen, Per

2011-01-01

Severe stress causes plant and animal cells to form large cytoplasmic granules containing RNA and proteins. Here, we demonstrate the existence of stress-induced cytoplasmic RNA granules in Schizosaccharomyces pombe. Homologs to several known protein components of mammalian processing bodies and stress granules are found in fission yeast RNA granules. In contrast to mammalian cells, poly(A)-binding protein (Pabp) colocalizes in stress-induced granules with decapping protein. After glucose deprivation, protein kinase A (PKA) is required for accumulation of Pabp-positive granules and translational down-regulation. This is the first demonstration of a role for PKA in RNA granule formation. In mammals, the translation initiation protein eIF2α is a key regulator of formation of granules containing poly(A)-binding protein. In S. pombe, nonphosphorylatable eIF2α does not block but delays granule formation and subsequent clearance after exposure to hyperosmosis. At least two separate pathways in S. pombe appear to regulate stress-induced granules: pka1 mutants are fully proficient to form granules after hyperosmotic shock; conversely, eIF2α does not affect granule formation in glucose starvation. Further, we demonstrate a Pka1-dependent link between calcium perturbation and RNA granules, which has not been described earlier in any organism.

Deficiency of Starch Synthase IIIa and IVb Alters Starch Granule Morphology from Polyhedral to Spherical in Rice Endosperm1

PubMed Central

Toyosawa, Yoshiko; Kawagoe, Yasushi; Matsushima, Ryo; Ogawa, Masahiro; Fukuda, Masako; Kumamaru, Toshihiro; Okazaki, Yozo; Kusano, Miyako; Saito, Kazuki; Toyooka, Kiminori; Sato, Mayuko; Ai, Yongfeng; Fujita, Naoko

2016-01-01

Starch granule morphology differs markedly among plant species. However, the mechanisms controlling starch granule morphology have not been elucidated. Rice (Oryza sativa) endosperm produces characteristic compound-type granules containing dozens of polyhedral starch granules within an amyloplast. Some other cereal species produce simple-type granules, in which only one starch granule is present per amyloplast. A double mutant rice deficient in the starch synthase (SS) genes SSIIIa and SSIVb (ss3a ss4b) produced spherical starch granules, whereas the parental single mutants produced polyhedral starch granules similar to the wild type. The ss3a ss4b amyloplasts contained compound-type starch granules during early developmental stages, and spherical granules were separated from each other during subsequent amyloplast development and seed dehydration. Analysis of glucan chain length distribution identified overlapping roles for SSIIIa and SSIVb in amylopectin chain synthesis, with a degree of polymerization of 42 or greater. Confocal fluorescence microscopy and immunoelectron microscopy of wild-type developing rice seeds revealed that the majority of SSIVb was localized between starch granules. Therefore, we propose that SSIIIa and SSIVb have crucial roles in determining starch granule morphology and in maintaining the amyloplast envelope structure. We present a model of spherical starch granule production. PMID:26747287

High resolution of heterogeneity among human neutrophil granules: physical, biochemical, and ultrastructural properties of isolated fractions.

PubMed

Rice, W G; Kinkade, J M; Parmley, R T

1986-08-01

Previous studies on the fractionation of human neutrophil granules have identified two major populations: myeloperoxidase (MPO)-containing azurophil, or primary, granules and MPO-deficient specific, or secondary, granules. Peripheral blood neutrophils from individual donors were lysed in sucrose-free media by either hypotonic shock or nitrogen cavitation. Using a novel two-gradient Percoll density centrifugation system, the granule-rich postnuclear supernatant was rapidly (ten minutes) and reproducibly resolved into 13 granule fractions (L1 through L8 and H1 through H5). Granule flotation and recentrifugation experiments on both continuous, self-generated and multiple-step gradients using individual and mixed isolated fractions demonstrated that the banding patterns were isopycnic and nonartifactual. Isolated granules were intact based on the findings that biochemical latency of several granule enzymes was greater than 95%, and thin-sectioned electron micrographs demonstrated intact granule profiles. Biochemical analyses of the granule marker proteins MPO, beta-glucuronidase, lysozyme, and lactoferrin indicated that a number of the fractions were related to the major azurophil and specific granule populations. Lactoferrin was found in ten of 13 fractions (L1 through L8, H1 to H2), whereas MPO was found in every fraction. Consistent with these biochemical data, all fractions exhibited varying degrees of heterogeneity based on ultrastructural morphology and cytochemistry, including diaminobenzidine (DAB) reactivity for peroxidase and periodate-thiocarbohydrazide-silver proteinate (PA-TCH-SP) staining for complex glycoconjugates. A variable but significant percentage (23% to 70%) of the granules in fractions L1 through L8 and H1 and H2 showed DAB reactivity, while about 90% of the granules in fractions H3 through H5 were peroxidase positive. These results demonstrated that DAB-reactive granules spanned the entire range of granule size and density. Ultrastructural PA-TCH-SP staining of isolated granule fractions revealed patterns similar to those of granules in intact neutrophils at different stages of development. Granules from human acute promyelocytic leukemia cells (HL-60) exhibited a surprisingly low density compared with typical azurophil granules from normal, mature neutrophils. The data suggest that both functional and maturational differences contribute to granule heterogeneity, and provide a new practical and conceptual framework for further defining the phenomenon of neutrophil granule heterogeneity.

Evaluation of beta-lactose, PVP K12 and PVP K90 as excipients to prepare piroxicam granules using two wet granulation techniques.

PubMed

Albertini, Beatrice; Cavallari, Cristina; Passerini, Nadia; González-Rodríguez, M L; Rodriguez, Lorenzo

2003-11-01

The present investigation aimed at evaluating the use of different excipients, beta-lactose and polyvinylpyrrolidone of two molecular weights (PVP K12 and PVP K90), in the production of improved release piroxicam granules, by wet granulation using both water and steam as granulation liquid. The formulations examined were: piroxicam (Px)/beta-lactose; Px/PVP K12 and Px/PVP K90, each one at a 1:9 weight ratio. The most significant difference between beta-lactose and PVP is that, using the first excipient, both steam and water granules were produced while, when PVP were employed, only steam granules were obtained. Image analysis revealed that beta-lactose steam granules had a larger surface area with respect to water granules, whereas lower values of this parameter were observed in PVP-s granules, confirming the Scanning Electron Microscopy micrographs and the fractal analysis results. As regards the enhancement of the dissolution profiles, the best result was obtained using beta-lactose steam granules followed by PVP K12 ones, even if the reactive dimension values indicated that during the dissolution process PVP K12 granules modified the surface more than beta-lactose granules. As regards PVP K90, this excipient was the one less influencing the granule morphology and the dissolution behaviour. Differential Scanning Calorimetry analysis suggested the partial amorphisation of the drug in the granules containing the three excipients. This result was then confirmed by X-ray powder diffraction analysis. Therefore, beta-lactose and PVP K12 could be proposed as useful excipients to enhance the dissolution rate of Px from granules prepared using the steam granulation technique.

Influence of metronidazole particle properties on granules prepared in a high-shear mixer-granulator.

PubMed

Di Martino, Piera; Censi, Roberta; Malaj, Ledjan; Martelli, Sante; Joiris, Etienne; Barthélémy, Christine

2007-02-01

Metronidazole is a good example of high-dose drug substance with poor granulating and tableting properties. Tablets are generally produced by liquid granulation; however, the technological process failure is quite frequent. In order to verify how the metronidazole particle characteristics can influence granule properties, three metronidazole batches differing for crystal habit, mean particle size, BET surface area and wettability were selected, primarily designed according to their different elongation ratio: needle-shaped, stick-shaped, and isodimensional. In the presence of lactose monohydrate and pregelatinized maize starch, respectively as diluent and binder, they were included in a formula for wet granulation in a high-shear mixer-granulator. In order to render the process comparable as far as possible, all parameters and experimental conditions were maintained constant. Four granule batches were obtained: granules from placebo (G-placebo), granules from needle-shaped crystals (G-needle-shaped), granules from stick-shaped crystals (G-stick-shaped), and granules from isodimensional crystals (G-isodimensional). Different granule properties were considered, in particular concerning porosity, friability, loss on drying (LOD), and flowability. In order to study their tabletability and compressibility, the different granules obtained were then compressed in a rotary press. The best tabletability was obtained with the isodimensional batch, while the poorest was exhibited by the stick-shaped one. Differences in tabletability are in good accordance with compressibility results: to a better tabletability corresponds an important granule ability to undergo a volume reduction as a result of an applied pressure. In particular, it was proposed that the greatest compressibility of the G-isodimensional must be related to the greatest granule porosity percentage.

Evaluation of the physicochemical properties and compaction behavior of melt granules produced in microwave-induced and conventional melt granulation in a single pot high shear processor.

PubMed

Loh, Z H; Sia, B Y; Heng, Paul W S; Lee, C C; Liew, Celine V

2011-12-01

Recently, microwave-induced melt granulation was shown to be a promising alternative to conventional melt granulation with improved process monitoring capabilities. This study aimed to compare the physicochemical and compaction properties of granules produced from microwave-induced and conventional melt granulation. Powder admixtures comprising equivalent proportions by weight of lactose 200 M and anhydrous dicalcium phosphate were granulated with polyethylene glycol 3350 under the influence of microwave-induced and conventional heating in a 10-L single pot high shear processor. The properties of the granules and compacts produced from the two processes were compared. Relative to conventional melt granulation, the rates at which the irradiated powders heated up in microwave-induced melt granulation were lower. Agglomerate growth proceeded at a slower rate, and this necessitated longer massing durations for growth induction. These factors prompted greater evaporative moisture losses from the melt granules. Additionally, nonuniform heating of the powders under the influence of microwaves led to increased inter-batch variations in the binder contents of resultant melt granules and a reliance of content homogeneity on massing duration. Agglomerate growth proceeded more rapidly under the influence of conventional heating due to the enhanced heating capabilities of the powders. Melt granules produced using the conventional method possessed higher moisture contents and improved content homogeneity. The compaction behavior of melt granules were affected by their mean sizes, porosities, flow properties, binder, and moisture contents. The last two factors were responsible for the disparities in compaction behavior of melt granules produced from microwave-induced and conventional melt granulation.

Effect of different excipients on the physical characteristics of granules and tablets with carbamazepine prepared with polyethylene glycol 6000 by fluidized hot-melt granulation (FHMG).

PubMed

Kraciuk, Radosław; Sznitowska, Malgorzata

2011-12-01

The objective of this study was to investigate the properties of granules and tablets with carbamazepine which were prepared employing a fluidized hot-melt granulation (FHMG) technique. The FHMG process was carried out at 65°C. Macrogol 6000 (PEG 6000) was used as a binder at the content 10% (w/w) of the granulated mass. Granules containing up to 70% (w/w) of the drug and 20-90% (w/w) of a filler (lactose, mannitol, calcium hydrogen phosphate (Di-Cafos), pregelatinized starch, and microcrystalline cellulose (MCC)) were produced. When the drug content was 30% (w/w), the yield of the process was satisfying (>95%) and flowability of the granules was better than placebo granules or drug-loaded granules prepared by wet granulation. Type of a filler had strong impact on physical properties of granules, and size distribution of the particles was the most homogenous when lactose or Di-Cafos were used. The FHMG technique enabled preparation of granules with better compressability compared with the wet-granulated product or with non-granulated powders. Tablets with shorter disintegration time than 10 min were obtained with 2.0% crospovidone added as a disintegrant. In comparison to tablets prepared from the wet-granulated mass, employment of the FHMG method resulted in tablets with faster dissolution of carbamazepine (more than 80% of the drug released within 15 min). This was achieved with mannitol or lactose/MCC, as fillers.

[Preparation and evaluation of taste masked orally disintegrating tablets with granules made by the wet granulation method].

PubMed

Kawano, Yayoi; Ito, Akihiko; Sasatsu, Masanaho; Machida, Yoshiharu; Onishi, Hiraku

2010-12-01

Using furosemide (FU) as a model drug, we examined the wet granulation method as a way to improve the taste masking and physical characteristics of orally disintegrating tablets (ODTs). In the wet granulation method, yogurt powder (YO) was used as a corrective and maltitol (MA) was used as a binding agent. The taste masked FU tablets were prepared using the direct compression method. Microcrystalline cellulose (Avicel® PH-302) and mannitol were added as excipients at a mixing ratio of 1/1 by weight. Based on the results of sensory test on taste, the prepared granules markedly improved the taste of FU, and a sufficient masking effect was obtained at the YO/FU ratio of 1 or more. Furthermore, it was found that the masking effect achieved by YO granules made with the wet granulation method was similar to or better than that produced by the granules made with dry granulation method. All types of tablets displayed sufficient hardness (over 3.5×10(-2) kN), and rapidly disintegrating tablets were obtained with YO granules produced at a mixing ratio of FU/YO=1/1, which disintegrated within 20 s. Disintegration time lengthened as the mixing ratio of YO to FU increased. In the mixing ratio of FU/YO=1/1, the hardness of tablets with granules made by the wet granulation method exceeded that of tablets with granules made by the dry granulation method, with minimal differences in disintegration time. The hardness and disintegration time of the tablets with granules made by the wet granulation method could be controlled by varying the compression force. In conclusion, YO was found to be a useful additive for masking unpleasant tastes. FU ODTs with improved taste, rapid disintegration and greater hardness could be prepared with YO-containing granules made by the wet granulation method using MA as a binding agent.

Rapid Feedforward Inhibition and Asynchronous Excitation Regulate Granule Cell Activity in the Mammalian Main Olfactory Bulb

PubMed Central

Burton, Shawn D.

2015-01-01

Granule cell-mediated inhibition is critical to patterning principal neuron activity in the olfactory bulb, and perturbation of synaptic input to granule cells significantly alters olfactory-guided behavior. Despite the critical role of granule cells in olfaction, little is known about how sensory input recruits granule cells. Here, we combined whole-cell patch-clamp electrophysiology in acute mouse olfactory bulb slices with biophysical multicompartmental modeling to investigate the synaptic basis of granule cell recruitment. Physiological activation of sensory afferents within single glomeruli evoked diverse modes of granule cell activity, including subthreshold depolarization, spikelets, and suprathreshold responses with widely distributed spike latencies. The generation of these diverse activity modes depended, in part, on the asynchronous time course of synaptic excitation onto granule cells, which lasted several hundred milliseconds. In addition to asynchronous excitation, each granule cell also received synchronous feedforward inhibition. This inhibition targeted both proximal somatodendritic and distal apical dendritic domains of granule cells, was reliably recruited across sniff rhythms, and scaled in strength with excitation as more glomeruli were activated. Feedforward inhibition onto granule cells originated from deep short-axon cells, which responded to glomerular activation with highly reliable, short-latency firing consistent with tufted cell-mediated excitation. Simulations showed that feedforward inhibition interacts with asynchronous excitation to broaden granule cell spike latency distributions and significantly attenuates granule cell depolarization within local subcellular compartments. Collectively, our results thus identify feedforward inhibition onto granule cells as a core feature of olfactory bulb circuitry and establish asynchronous excitation and feedforward inhibition as critical regulators of granule cell activity. SIGNIFICANCE STATEMENT Inhibitory granule cells are involved critically in shaping odor-evoked principal neuron activity in the mammalian olfactory bulb, yet little is known about how sensory input activates granule cells. Here, we show that sensory input to the olfactory bulb evokes a barrage of asynchronous synaptic excitation and highly reliable, short-latency synaptic inhibition onto granule cells via a disynaptic feedforward inhibitory circuit involving deep short-axon cells. Feedforward inhibition attenuates local depolarization within granule cell dendritic branches, interacts with asynchronous excitation to suppress granule cell spike-timing precision, and scales in strength with excitation across different levels of sensory input to normalize granule cell firing rates. PMID:26490853

Difference in distribution of membrane proteins between low- and high-density secretory granules in parotid acinar cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fujita-Yoshigaki, Junko; Katsumata, Osamu; Matsuki, Miwako

Secretory granules (SGs) are considered to be generated as immature granules and to mature by condensation of their contents. In this study, SGs of parotid gland were separated into low-, medium-, and high-density granule fractions by Percoll-density gradient centrifugation, since it was proposed that the density corresponds to the degree of maturation. The observation with electron microscopy showed that granules in the three fractions were very similar. The average diameter of high-density granules was a little but significantly larger than that of low-density granules. Although the three fractions contained amylase, suggesting that they are all SGs, distribution of membrane proteinsmore » was markedly different. Syntaxin6 and VAMP4 were localized in the low-density granule fraction, while VAMP2 was concentrated in the high-density granule fraction. Immunoprecipitation with anti-syntaxin6 antibody caused coprecipitation of VAMP2 from the medium-density granule fraction without solubilization, but not from Triton X-100-solubilized fraction, while VAMP4 was coprecipitated from both fractions. Therefore, VAMP2 is present on the same granules, but is separated from syntaxin6 and VAMP4, which are expected to be removed from immature granules. These results suggest that the medium-density granules are intermediates from low- to high-density granules, and that the membrane components of SGs dynamically change by budding and fusion during maturation.« less

PHB granules are attached to the nucleoid via PhaM in Ralstonia eutropha.

PubMed

Wahl, Andreas; Schuth, Nora; Pfeiffer, Daniel; Nussberger, Stephan; Jendrossek, Dieter

2012-11-16

Poly(3-hydroxybutyrate) (PHB) granules are important storage compounds of carbon and energy in many prokaryotes which allow survival of the cells in the absence of suitable carbon sources. Formation and subcellular localization of PHB granules was previously assumed to occur randomly in the cytoplasm of PHB accumulating bacteria. However, contradictionary results on subcellular localization of PHB granules in Ralstonia eutropha were published, recently. Here, we provide evidence by transmission electron microscopy that PHB granules are localized in close contact to the nucleoid region in R. eutropha during growth on nutrient broth. Binding of PHB granules to the nucleoid is mediated by PhaM, a PHB granule associated protein with phasin-like properties that is also able to bind to DNA and to phasin PhaP5. Over-expression of PhaM resulted in formation of many small PHB granules that were always attached to the nucleoid region. In contrast, PHB granules of ∆phaM strains became very large and distribution of granules to daughter cells was impaired. Association of PHB granules to the nucleoid region was prevented by over-expression of PhaP5 and clusters of several PHB granules were mainly localized near the cell poles. Subcellular localization of PHB granules is controlled in R. eutropha and depends on the presence and concentrations of at least two PHB granule associated proteins, PhaM and PhaP5.

Sodium phenylbutyrate coated granules (Pheburane). Defective urea synthesis: a welcome formulation.

PubMed

2015-02-01

Compared with Ammonaps granules, Pheburane coated granules mask the unpleasant taste of sodium phenylbutyrate. A more precise dosing device is provided with the coated granules than with the uncoated granules (Ammonaps).

Rab3A, a possible marker of cortical granules, participates in cortical granule exocytosis in mouse eggs

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bello, Oscar Daniel; Cappa, Andrea Isabel; Paola, Matilde de

Fusion of cortical granules with the oocyte plasma membrane is the most significant event to prevent polyspermy. This particular exocytosis, also known as cortical reaction, is regulated by calcium and its molecular mechanism is still not known. Rab3A, a member of the small GTP-binding protein superfamily, has been implicated in calcium-dependent exocytosis and is not yet clear whether Rab3A participates in cortical granules exocytosis. Here, we examine the involvement of Rab3A in the physiology of cortical granules, particularly, in their distribution during oocyte maturation and activation, and their participation in membrane fusion during cortical granule exocytosis. Immunofluorescence and Western blotmore » analysis showed that Rab3A and cortical granules have a similar migration pattern during oocyte maturation, and that Rab3A is no longer detected after cortical granule exocytosis. These results suggested that Rab3A might be a marker of cortical granules. Overexpression of EGFP-Rab3A colocalized with cortical granules with a Pearson correlation coefficient of +0.967, indicating that Rab3A and cortical granules have almost a perfect colocalization in the egg cortical region. Using a functional assay, we demonstrated that microinjection of recombinant, prenylated and active GST-Rab3A triggered cortical granule exocytosis, indicating that Rab3A has an active role in this secretory pathway. To confirm this active role, we inhibited the function of endogenous Rab3A by microinjecting a polyclonal antibody raised against Rab3A prior to parthenogenetic activation. Our results showed that Rab3A antibody microinjection abolished cortical granule exocytosis in parthenogenetically activated oocytes. Altogether, our findings confirm that Rab3A might function as a marker of cortical granules and participates in cortical granule exocytosis in mouse eggs. - Highlights: • Rab3A has a similar migration pattern to cortical granules in mouse oocytes. • Rab3A can be a marker of cortical granules. • Active Rab3A triggered cortical granule exocytosis. • Blocking endogenous Rab3A inhibits cortical granule exocytosis. • Rab3A participates in cortical reaction in mouse oocytes.« less

Surface Localization of Zein Storage Proteins in Starch Granules from Maize Endosperm1

PubMed Central

Mu-Forster, Chen; Wasserman, Bruce P.

1998-01-01

Starch granules from maize (Zea mays) contain a characteristic group of polypeptides that are tightly associated with the starch matrix (C. Mu-Forster, R. Huang, J.R. Powers, R.W. Harriman, M. Knight, G.W. Singletary, P.L. Keeling, B.P. Wasserman [1996] Plant Physiol 111: 821–829). Zeins comprise about 50% of the granule-associated proteins, and in this study their spatial distribution within the starch granule was determined. Proteolysis of starch granules at subgelatinization temperatures using the thermophilic protease thermolysin led to selective removal of the zeins, whereas granule-associated proteins of 32 kD or above, including the waxy protein, starch synthase I, and starch-branching enzyme IIb, remained refractory to proteolysis. Granule-associated proteins from maize are therefore composed of two distinct classes, the surface-localized zeins of 10 to 27 kD and the granule-intrinsic proteins of 32 kD or higher. The origin of surface-localized δ-zein was probed by comparing δ-zein levels of starch granules obtained from homogenized whole endosperm with granules isolated from amyloplasts. Starch granules from amyloplasts contained markedly lower levels of δ-zein relative to granules prepared from whole endosperm, thus indicating that δ-zein adheres to granule surfaces after disruption of the amyloplast envelope. Cross-linking experiments show that the zeins are deposited on the granule surface as aggregates. In contrast, the granule-intrinsic proteins are prone to covalent modification, but do not form intermolecular cross-links. We conclude that individual granule intrinsic proteins exist as monomers and are not deposited in the form of multimeric clusters within the starch matrix. PMID:9536075

Insensitivity of compaction properties of brittle granules to size enlargement by roller compaction.

PubMed

Wu, Sy-Juen; Sun, Changquan 'Calvin'

2007-05-01

Pharmaceutical granules prepared by roller compaction often exhibit significant loss of tabletability, that is, reduction in tensile strength, when compared to virgin powder. This may be attributed to granule size enlargement for highly plastic materials, for example, microcrystalline cellulose. The sensitivity of powder compaction properties on granule size variations impacts the robustness of the dry granulation process. We hypothesize that such sensitivity of compaction properties on granule size is minimum for brittle materials because extensive fracture of brittle granules during compaction minimizes differences in initial granule size. We tested the hypothesis using three common brittle excipients. Results show that the fine (44-106 microm), medium (106-250 microm), and coarse (250-500 microm) granules exhibit essentially identical tabletability below a certain critical compaction pressure, 100, 140, and 100 MPa for spray-dried lactose monohydrate, anhydrous dibasic calcium phosphate, and mannitol, respectively. Above respective critical pressure, tabletability lines diverge with smaller granules exhibiting slightly higher tablet tensile strength at identical compaction conditions. Overall, tabletability of brittle granules is insensitive to granule size enlargement. The results provide a scientific basis to the common practice of incorporating brittle filler to a typical tablet formulation processed by roller compaction granulation. (c) 2007 Wiley-Liss, Inc. and the American Pharmacists Association.

Roll compaction/dry granulation: comparison between roll mill and oscillating granulator in dry granulation.

PubMed

Sakwanichol, Jarunee; Puttipipatkhachorn, Satit; Ingenerf, Gernot; Kleinebudde, Peter

2012-01-01

Different experimental factorial designs were employed to evaluate granule properties obtained from oscillating granulator and roll mill. Four oscillating-granulator parameters were varied, i.e. rotor speed, oscillating angle, aperture of mesh screen and rotor type. Six roll-mill parameters that were throughput, speed ratio in both first and second stages, gap between roll pair in both stages and roll-surface texture were also investigated. Afterwards, the granule properties obtained from two milling types with similar median particle size were compared. All milling parameters in both milling types affected significantly the median particle size, size distribution and amount of fine particles (P < 0.05), except the rotor types of oscillating granulator on fines. Only three milling parameters influenced significantly the flowability (P < 0.05). These were the throughput and the gap size in the first stage of roll mill and the sieve size of oscillating granulator. In comparison between milling types, the differences of granule properties were not practically relevant. However, the roll mill had much higher capacity than the oscillating granulator about seven times, resulting in improving energy savings per unit of product. Consequently, the roll mill can be applied instead of oscillating granulator for roll compaction/dry granulation technique.

An innovative method for the preparation of high API-loaded hollow spherical granules for use in controlled-release formulation.

PubMed

Asada, Takumi; Kobiki, Mitsuaki; Ochiai, Yasushi; Iwao, Yasunori; Itai, Shigeru

2017-05-15

The aim of this study was to prepare controlled-release (CR) granules with suitable particle strength, flowability, particle size distribution (PSD) and density characteristics for blending with other excipients. We also wanted these CR granules to contain large quantities of active pharmaceutical ingredient (API). A high shear mixer was used to mix an API with various polymers at various feed ratios, and the resulting granulated materials were sprayed with solvent. The wet granules were dried using a fluidized bed dryer to give CR granules. The API content of the granules was determined to be 95wt%. The granules were found to be spherical in shape with smooth surfaces by scanning electron microscopy. The inner structure of each granule was determined to be hollow by X-ray computed tomography, highlighting the unusual mechanism of this granulation process. The PSD of the granules was found to be dependent on that of the constituent polymer, and a narrow PSD was obtained by adjusting the PSD of the polymer. The dissolution profile of the granules was also dependent on the constituent polymer. Taken together, these results show that we have successfully developed a new manufacturing technology for the simple and low-cost preparation of ideal CR granules. Copyright © 2017 Elsevier B.V. All rights reserved.

Enhancement of ibuprofen dissolution via wet granulation with beta-cyclodextrin.

PubMed

Ghorab, M K; Adeyeye, M C

2001-08-01

The purpose was to investigate the effect of wet granulation with beta-cyclodextrin (betaCD) on the enhancement of ibuprofen (IBU) dissolution. The effect of the granulation variables on the physical properties as well as the dissolution of tablets prepared from these granules was also examined. Granulation was performed using three granulating solvents: water, ethanol (95 vol%), and isopropanol. Granules were either oven-dried for 2 h or air-dried for 3 days. The granules or respective physical mixtures were compressed into tablets. Powder X-ray diffraction showed that oven-dried granulation resulted in less amorphous entities thatfacilitated IBU-betaCD complexation in solution and enhanced the dissolution of the corresponding tablets compared to the physical mixture with or without oven drying. In contrast, air-dried granulation did not cause any differences in the X-ray diffraction pattern (crystallinity) or the dissolution compared to the physical mixture without drying. Isopropanol and water, as granulating solvents, enhanced the dissolution of the oven-dried batches more than ethanol. The Differential scanning calorimetry (DSC) and Thermogravimetric analysis (TGA) data showed that tablets prepared from oven-dried granules, but not air-dried granules, had lower AH values and percent loss in weight, respectively, than those prepared from the physical mixture as a result of the expulsion of the water molecules from the betaCD cavity and enhancement of the complexation in solution. These results showed that oven-dried granulation of IBU and betaCD provided faster IBU dissolution than the physical mixture; air-dried granulation did not substantially affect the dissolution of IBU.

The biological significance of storage granules in rat parathyroid cells.

PubMed

Setoguti, T; Inoue, Y; Wild, P

1995-10-01

Both prosecretory and storage granules are concomitantly formed at the trans Golgi network including the innermost Golgi cisterna. Prosecretory granules develop into small secretory granules that release their contents by exocytosis finely regulated by a complex mechanism for maintaining calcium homeostasis. In the rat parathyroid cells, storage granules are large secretory granules storing parathyroid hormone for an emergency supply. The hormone is rapidly discharged by exocytosis when serum calcium concentration is decreased. The granules are constantly produced even under conditions of low serum calcium concentration in the regions of 8 mg/dl. The granule content is constantly hydrolyzed when not discharged, leading to a decreased core and finally to the formation of vacuolar bodies. The fate of the vacuolar bodies is unknown. Hypercalcemic conditions accelerate hydrolysis. The threshold value of calcium concentration required for the release of storage granule contents is between 8.0 and 7.5 mg/dl and that of calcium concentration for accelerating degradation of storage granules is about 11.5 mg/dl. Sympathetic stimulation causes storage granules to be discharged regardless of hypercalcemia or hypocalcemia. Parasympathetic stimulation accelerates hydrolysis. The degradation of storage granules seems to be closely associated with an intracellular regulatory mechanism for parathyroid hormone secretion.

Specific aerobic granules can be developed in a completely mixed tank reactor by bioaugmentation using micro-mycelial pellets of Phanerochaete chrysosporium.

PubMed

Hailei, Wang; Ping, Li; Qianlong, Jin; Ge, Qin

2014-03-01

Aerobic granules were firstly developed in a completely mixed tank reactor (CMTR) by seeding micro-mycelial pellets (MMPs) of Phanerochaete chrysosporium. During phenol wastewater treatment, sludge granulation rate reached 67 % after 15-day operation. The granules in CMTR are different from aerobic granules described in literature in morphology, and a majority of them are rod-shaped or rodlike sludge besides spherical granules. The polymorphic granules, having no essential difference with aerobic granules previously reported, achieve advantages over conventional activated sludge in settling ability, biomass concentration, density, integrity coefficient and removal ability to phenol wastewater. The optimized parameters for sludge granulation in CMTR including temperature, inoculum quantity, rotary speed and superficial air upflow velocity are 30 °C, 5–7 g/l, 150 rpm, and 0.5 cm/s, respectively. Analysis on sludge granulation mechanism indicates that MMPs not only result in the formation of aerobic granules containing MMPs as nuclei, but also induce the formation of biogranules which do not have MMP at their cores. The work challenges the general belief that the homogenous circular flow pattern of microbial aggregates is necessary for aerobic sludge granulation.

PHB granules are attached to the nucleoid via PhaM in Ralstonia eutropha

PubMed Central

2012-01-01

Background Poly(3-hydroxybutyrate) (PHB) granules are important storage compounds of carbon and energy in many prokaryotes which allow survival of the cells in the absence of suitable carbon sources. Formation and subcellular localization of PHB granules was previously assumed to occur randomly in the cytoplasm of PHB accumulating bacteria. However, contradictionary results on subcellular localization of PHB granules in Ralstonia eutropha were published, recently. Results Here, we provide evidence by transmission electron microscopy that PHB granules are localized in close contact to the nucleoid region in R. eutropha during growth on nutrient broth. Binding of PHB granules to the nucleoid is mediated by PhaM, a PHB granule associated protein with phasin-like properties that is also able to bind to DNA and to phasin PhaP5. Over-expression of PhaM resulted in formation of many small PHB granules that were always attached to the nucleoid region. In contrast, PHB granules of ∆phaM strains became very large and distribution of granules to daughter cells was impaired. Association of PHB granules to the nucleoid region was prevented by over-expression of PhaP5 and clusters of several PHB granules were mainly localized near the cell poles. Conclusion Subcellular localization of PHB granules is controlled in R. eutropha and depends on the presence and concentrations of at least two PHB granule associated proteins, PhaM and PhaP5. PMID:23157596

Lithium granule ablation and penetration during ELM pacing experiments at DIII-D

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lunsford, R.; Bortolon, A.; Roquemore, A. L.

At DIII-D, lithium granules were radially injected into the plasma at the outer midplane to trigger and pace edge localized modes (ELMs). Granules ranging in size from 300 to 1000 microns were horizontally launched into H-mode discharges with velocities near 100 m/s, and granule to granule injection frequencies less than 500 Hz. While the smaller granules were only successful in triggering ELMs approximately 20% of the time, the larger granules regularly demonstrated ELM triggering efficiencies of greater than 80%. A fast visible camera looking along the axis of injection observed the ablation of the lithium granules. We used the durationmore » of ablation as a benchmark for a neutral gas shielding calculation, and approximated the ablation rate and mass deposition location for the various size granules, using measured edge plasma profiles as inputs. In conclusion, this calculation suggests that the low triggering efficiency of the smaller granules is due to the inability of these granules to traverse the steep edge pressure gradient region and reach the top of the pedestal prior to full ablation.« less

Lithium granule ablation and penetration during ELM pacing experiments at DIII-D

DOE PAGES

Lunsford, R.; Bortolon, A.; Roquemore, A. L.; ...

2016-05-25

At DIII-D, lithium granules were radially injected into the plasma at the outer midplane to trigger and pace edge localized modes (ELMs). Granules ranging in size from 300 to 1000 microns were horizontally launched into H-mode discharges with velocities near 100 m/s, and granule to granule injection frequencies less than 500 Hz. While the smaller granules were only successful in triggering ELMs approximately 20% of the time, the larger granules regularly demonstrated ELM triggering efficiencies of greater than 80%. A fast visible camera looking along the axis of injection observed the ablation of the lithium granules. We used the durationmore » of ablation as a benchmark for a neutral gas shielding calculation, and approximated the ablation rate and mass deposition location for the various size granules, using measured edge plasma profiles as inputs. In conclusion, this calculation suggests that the low triggering efficiency of the smaller granules is due to the inability of these granules to traverse the steep edge pressure gradient region and reach the top of the pedestal prior to full ablation.« less

Visualization and understanding of the granulation liquid mixing and distribution during continuous twin screw granulation using NIR chemical imaging.

PubMed

Vercruysse, Jurgen; Toiviainen, Maunu; Fonteyne, Margot; Helkimo, Niko; Ketolainen, Jarkko; Juuti, Mikko; Delaet, Urbain; Van Assche, Ivo; Remon, Jean Paul; Vervaet, Chris; De Beer, Thomas

2014-04-01

Over the last decade, there has been increased interest in the application of twin screw granulation as a continuous wet granulation technique for pharmaceutical drug formulations. However, the mixing of granulation liquid and powder material during the short residence time inside the screw chamber and the atypical particle size distribution (PSD) of granules produced by twin screw granulation is not yet fully understood. Therefore, this study aims at visualizing the granulation liquid mixing and distribution during continuous twin screw granulation using NIR chemical imaging. In first instance, the residence time of material inside the barrel was investigated as function of screw speed and moisture content followed by the visualization of the granulation liquid distribution as function of different formulation and process parameters (liquid feed rate, liquid addition method, screw configuration, moisture content and barrel filling degree). The link between moisture uniformity and granule size distributions was also studied. For residence time analysis, increased screw speed and lower moisture content resulted to a shorter mean residence time and narrower residence time distribution. Besides, the distribution of granulation liquid was more homogenous at higher moisture content and with more kneading zones on the granulator screws. After optimization of the screw configuration, a two-level full factorial experimental design was performed to evaluate the influence of moisture content, screw speed and powder feed rate on the mixing efficiency of the powder and liquid phase. From these results, it was concluded that only increasing the moisture content significantly improved the granulation liquid distribution. This study demonstrates that NIR chemical imaging is a fast and adequate measurement tool for allowing process visualization and hence for providing better process understanding of a continuous twin screw granulation system. Copyright © 2013 Elsevier B.V. All rights reserved.

Formulation of a poorly water-soluble drug in sustained-release hollow granules with a high viscosity water-soluble polymer using a fluidized bed rotor granulator.

PubMed

Asada, Takumi; Yoshihara, Naoki; Ochiai, Yasushi; Kimura, Shin-Ichiro; Iwao, Yasunori; Itai, Shigeru

2018-04-25

Water-soluble polymers with high viscosity are frequently used in the design of sustained-release formulations of poorly water-soluble drugs to enable complete release of the drug in the gastrointestinal tract. Tablets containing matrix granules with a water-soluble polymer are preferred because tablets are easier to handle and the multiple drug-release units of the matrix granules decreases the influences of the physiological environment on the drug. However, matrix granules with a particle size of over 800 μm sometimes cause a content uniformity problem in the tableting process because of the large particle size. An effective method of manufacturing controlled-release matrix granules with a smaller particle size is desired. The aim of this study was to develop tablets containing matrix granules with a smaller size and good controlled-release properties, using phenytoin as a model poorly water-soluble drug. We adapted the recently developed hollow spherical granule granulation technology, using water-soluble polymers with different viscosities. The prepared granules had an average particle size of 300 μm and sharp particle size distribution (relative width: 0.52-0.64). The values for the particle strength of the granules were 1.86-1.97 N/mm 2 , and the dissolution profiles of the granules were not affected by the tableting process. The dissolution profiles and the blood concentration levels of drug released from the granules depended on the viscosity of the polymer contained in the granules. We succeeded in developing the desired controlled-release granules, and this study should be valuable in the development of sustained-release formulations of poorly water-soluble drugs. Copyright © 2018 Elsevier B.V. All rights reserved.

Factor X/Xa elicits protective signaling responses in endothelial cells directly via PAR-2 and indirectly via endothelial protein C receptor-dependent recruitment of PAR-1.

PubMed

Bae, Jong-Sup; Yang, Likui; Rezaie, Alireza R

2010-11-05

We recently demonstrated that the Gla domain-dependent interaction of protein C with endothelial protein C receptor (EPCR) leads to dissociation of the receptor from caveolin-1 and recruitment of PAR-1 to a protective signaling pathway. Thus, the activation of PAR-1 by either thrombin or PAR-1 agonist peptide elicited a barrier-protective response if endothelial cells were preincubated with protein C. In this study, we examined whether other vitamin K-dependent coagulation protease zymogens can modulate PAR-dependent signaling responses in endothelial cells. We discovered that the activation of both PAR-1 and PAR-2 in endothelial cells pretreated with factor FX (FX)-S195A, but not other procoagulant protease zymogens, also results in initiation of protective intracellular responses. Interestingly, similar to protein C, FX interaction with endothelial cells leads to dissociation of EPCR from caveolin-1 and recruitment of PAR-1 to a protective pathway. Further studies revealed that, FX activated by factor VIIa on tissue factor bearing endothelial cells also initiates protective signaling responses through the activation of PAR-2 independent of EPCR mobilization. All results could be recapitulated by the receptor agonist peptides to both PAR-1 and PAR-2. These results suggest that a cross-talk between EPCR and an unknown FX/FXa receptor, which does not require interaction with the Gla domain of FX, recruits PAR-1 to protective signaling pathways in endothelial cells.

Factor XI and Contact Activation as Targets for Antithrombotic Therapy

PubMed Central

Gailani, David; Bane, Charles E.; Gruber, Andras

2015-01-01

Summary The most commonly used anticoagulants produce therapeutic antithrombotic effects either by inhibiting thrombin or factor Xa, or by lowering the plasma levels of the precursors of these key enzymes, prothrombin and factor X. These drugs do not distinguish between thrombin generation contributing to thrombosis from thrombin generation required for hemostasis. Thus, anticoagulants increase bleeding risk, and many patients who would benefit from therapy go untreated because of comorbidities that place them at unacceptable risk for hemorrhage. Studies in animals demonstrate that components of the plasma contact activation system contribute to experimentally-induced thrombosis, despite playing little or no role in hemostasis. Attention has focused on factor XII, the zymogen of a protease (factor XIIa) that initiates contact activation when blood is exposed to foreign surfaces; and factor XI, the zymogen of the protease factor XIa, which links contact activation to the thrombin generation mechanism. In the case of factor XI, epidemiologic data indicate this protein contributes to stroke and venous thromboembolism, and perhaps myocardial infarction, in humans. A phase 2 trial showing that reduction of factor XI may be more effective than low-molecular-weight heparin at preventing venous thrombosis during knee replacement surgery provides proof of concept for the premise that an antithrombotic effect can be uncoupled from an anticoagulant effect in humans by targeting components of contact activation. Here we review data on the role of factor XI and factor XII in thrombosis, and results of pre-clinical and human trials for therapies targeting these proteins. PMID:25976012

Physical, electrochemical, and thermal properties of granulated natural graphite as anodes for Li-ion batteries.

PubMed

Jo, Yong Nam; Park, Min-Sik; Kim, Jae-Hun; Kim, Young-Jun

2013-05-01

Two different types of granulated graphites were synthesized by blending and kneading of natural graphite with pitch followed by sintering methods. The electrochemical performances of granulated graphites were investigated as anode materials for use in Li-ion batteries. The blending type granulated graphite possesses a large amount of cavities and voids, while the kneading type granulated graphite has a relatively compact microstructure, which is responsible for a high tap density. Both granulated graphites show improved the initial coulombic efficiencies as a result of decrease of surface area by the granulations. In particular, the kneading type granulated graphite exhibits an excellent rate-capability without significant capacity loss. In addition, the thermal stabilities of both granulated graphites were also improved, which could be attributed to the decrease of active surface area due to pitch coating.

New Insights into PhaM-PhaC-Mediated Localization of Polyhydroxybutyrate Granules in Ralstonia eutropha H16

PubMed Central

Bresan, Stephanie

2017-01-01

ABSTRACT The formation and localization of polyhydroxybutyrate (PHB) granules in Ralstonia eutropha are controlled by PhaM, which interacts both with the PHB synthase (PhaC) and with the bacterial nucleoid. Here, we studied the importance of proline and lysine residues of two C-terminal PAKKA motifs in PhaM for their importance in attaching PHB granules to DNA by in vitro and in vivo methods. Substitution of the lysine residues but not of the proline residues resulted in detachment of formed PHB granules from the nucleoid. Instead, formation of PHB granule clusters at polar regions of the rod-shaped cells and an unequal distribution of PHB granules to daughter cells were observed. The formation of PHB granules was studied by the expression of chromosomally anchored gene fusions of fluorescent proteins with PhaM and PhaC in different backgrounds. PhaM and PhaC fusions showed a distinct colocalization at formed PHB granules in the nucleoid region of the wild type. In a ΔphaC background, PhaM and the catalytically inactive PhaCC319A protein were not able to form fluorescent foci, indicating that correct positioning requires the formation of PHB. Furthermore, time-lapse experiments revealed that PhaC and PhaM proteins detach from formed PHB granules at later stages, resulting in a nonhomogeneous population of PHB granules. This could explain why growth of individual PHB granules stops under PHB-permissive conditions at a certain size. IMPORTANCE PHB granules are storage compounds for carbon and energy in many prokaryotes. Equal distribution of accumulated PHB granules during cell division is therefore important for optimal fitness of the daughter cells. In R. eutropha, PhaM is responsible for maximal activity of PHB synthase, for initiation of PHB granule formation at discrete regions in the cells, and for association of formed PHB granules with the nucleoid. Here we found that four lysine residues of C-terminal PhaM sequence motifs are essential for association of PHB granules with the nucleoid. Furthermore, we followed PHB granule formation by time-lapse microscopy and provide evidence for aging of PHB granules that is manifested by detachment of previously PHB granule-associated PhaM and PHB synthase. PMID:28389545

Osseointegration of alumina bioceramic granules: A comparative experimental study

NASA Astrophysics Data System (ADS)

Rerikh, V. V.; Avetisyan, A. R.; Zaydman, A. M.; Anikin, K. A.; Bataev, V. A.; Nikulina, A. A.; Sadovoy, M. A.; Aronov, A. M.; Semantsova, E. S.

2016-08-01

To perform a comparative analysis of osseointegration of bioceramic alumina-based granules, hydroxyapatite-based granules, and deproteinized bone granules. The experiment was conducted on 52 adult male Kyoto-Wistar rats weighing 350 to 520 g. The animals were divided into five matched groups that differed only in the type of an implanted material. The granules were implanted in the lumbar vertebral bodies and in the distal right femur of each laboratory animal. Two months after surgery, the animals were euthanized, followed by tissue sampling for morphological studies. An examination of specimens from the groups with implanted alumina granules revealed the newly formed trabecular bone with remodeling signs. The bone tissue filled the intragranular space, tightly adhering to the granule surface. There was no connective tissue capsule on the border between bone tissue and alumina granules. Cylindrical bioceramic alumina-based granules with an open internal channel have a higher strength surpassing than that of analogs and the osseointegration ability close to that of hydroxyapatite and deproteinized bone granules.

Formation of artificial granules for proving gelation as the main mechanism of aerobic granulation in biological wastewater treatment.

PubMed

Li, Yun; Yang, Shu-Fang; Zhang, Jian-Jun; Li, Xiao-Yan

2014-01-01

In this study, gelation-facilitated biofilm formation as a new mechanism is proposed for the phenomenon of aerobic granulation in biological wastewater treatment. To obtain an experimental proof for the gelation-based theory, the granulation process was simulated in a chemical system using latex particles for bacterial cells and organic polymers (alginate and peptone) for extracellular polymeric substances (EPS) in a solution with the addition of cations (Ca²⁺, Mg²⁺ and Fe³⁺). The results showed that at a low alginate content (70 mg g⁻¹ mixed liquid suspended solids (MLSS)) flocculation was observed in the suspension with loose flocs. At a higher alginate content (180 mg g⁻¹ MLSS), together with discharge of small flocs, formation of artificial gel granules was successfully achieved leading to granulation. The artificial granules show a morphological property similar to that of actual microbial granules. However, if the protein content increased, granulation became difficult with little gel formation. The experimental work demonstrates the importance of the bonding interactions between EPS functional groups and cations in gel formation and granulation. The laboratory results on the formation of artificial granules provide a sound proof for the theory of gelation-facilitated biofilm formation as the main mechanism for aerobic granulation in sludge suspensions.

Granule Formation Mechanisms within an Aerobic Wastewater System for Phosphorus Removal▿ †

PubMed Central

Barr, Jeremy J.; Cook, Andrew E.; Bond, Phillip L.

2010-01-01

Granular sludge is a novel alternative for the treatment of wastewater and offers numerous operational and economic advantages over conventional floccular-sludge systems. The majority of research on granular sludge has focused on optimization of engineering aspects relating to reactor operation with little emphasis on the fundamental microbiology. In this study, we hypothesize two novel mechanisms for granule formation as observed in three laboratory scale sequencing batch reactors operating for biological phosphorus removal and treating two different types of wastewater. During the initial stages of granulation, two distinct granule types (white and yellow) were distinguished within the mixed microbial population. White granules appeared as compact, smooth, dense aggregates dominated by 97.5% “Candidatus Accumulibacter phosphatis,” and yellow granules appeared as loose, rough, irregular aggregates with a mixed microbial population of 12.3% “Candidatus Accumulibacter phosphatis” and 57.9% “Candidatus Competibacter phosphatis,” among other bacteria. Microscopy showed white granules as homogeneous microbial aggregates and yellow granules as segregated, microcolony-like aggregates, with phylogenetic analysis suggesting that the granule types are likely not a result of strain-associated differences. The microbial community composition and arrangement suggest different formation mechanisms occur for each granule type. White granules are hypothesized to form by outgrowth from a single microcolony into a granule dominated by one bacterial type, while yellow granules are hypothesized to form via multiple microcolony aggregation into a microcolony-segregated granule with a mixed microbial population. Further understanding and application of these mechanisms and the associated microbial ecology may provide conceptual information benefiting start-up procedures for full-scale granular-sludge reactors. PMID:20851963

Development of a discriminative biphasic in vitro dissolution test and correlation with in vivo pharmacokinetic studies for differently formulated racecadotril granules.

PubMed

Deng, Jia; Staufenbiel, Sven; Hao, Shilei; Wang, Bochu; Dashevskiy, Andriy; Bodmeier, Roland

2017-06-10

The purpose of this study was to discriminate the release behavior from three differently formulated racecadotril (BCS II) granules and to establish an in vitro-in vivo correlation. Three granule formulations of the lipophilic drug were prepared with equivalent composition but prepared with different manufacturing processes (dry granulation, wet granulation with or without binder). In vitro release of the three granules was investigated using a biphasic dissolution system (phosphate buffer pH6.8 and octanol) and compared to the conventional single phase USP II dissolution test performed under sink and non-sink conditions. In vivo studies with each granule formulation were performed in rats. Interestingly, the granule formulations exhibited pronouncedly different behavior in the different dissolution systems depending on different wetting and dissolution conditions. Single phase USP II dissolution tests lacked discrimination. In contrast, remarkable discrimination between the granule formulations was observed in the octanol phase of biphasic dissolution system with a rank order of release from granules prepared by wet granulation with binder>wet granulation without binder>dry granulation. This release order correlated well with the wettability of these granules. An excellent correlation was also established between in vitro release in the octanol phase of the biphasic test and in vivo data (R 2 =0.999). Compared to conventional dissolution methods, the biphasic method provides great potential to discriminate between only minor formulation and process changes within the same dosage form for poorly soluble drugs. Copyright © 2017 Elsevier B.V. All rights reserved.

Modification of drug release from acetaminophen granules by melt granulation technique - consideration of release kinetics.

PubMed

Uhumwangho, M U; Okor, R S

2006-01-01

Acetaminophen granules have been formed by a melt granulation process with the objective of retarding drug release for prolonged action formulations. The waxes used were goat wax, carnuba wax and glyceryl monostearate. In the melt granulation procedure, acetaminophen powder was triturated with the melted waxes and passed through a sieve of mesh 10 (aperture size 710 microm). The content of wax in resulting granules ranged from 10 to 40%w/w. Acetaminophen granules were also formed by the convectional method of wet granulation with starch mucilage (20%w/w). The granules were subjected to in-vitro drug release tests. The release data were subjected to analysis by three different well-established mathematical models (release kinetics) namely, - zero order flux, first order, and the Higuchi square root of time relationship. The convectional granules exhibited an initial zero order flux (first 55%) followed by a first order release profile (the remaining 45%). The pattern of drug release from the melt granulations was consistent with the first order kinetic and the Higuchi square root of time relationship, indicating a diffusion-controlled release mechanism. The first order release rate constant of the convectional granules was 1.95 +/- 0.02 h(-1). After melt granulation (wax content, 20%w/w) the rate constants dropped drastically to 0.130+/-0.001 h(-1) (goat wax), 0.120+/-0.003 h(-1) (carnuba wax), and 0.130+/-0.002 h(-1) (glyceryl monosterate) indicating that all three waxes were equivalent in retarding drug release from the melt granulations.

State of the art of aerobic granulation in continuous flow bioreactors.

PubMed

Kent, Timothy R; Bott, Charles B; Wang, Zhi-Wu

In the wake of the success of aerobic granulation in sequential batch reactors (SBRs) for treating wastewater, attention is beginning to turn to continuous flow applications. This is a necessary step given the advantages of continuous flow treatment processes and the fact that the majority of full-scale wastewater treatment plants across the world are operated with aeration tanks and clarifiers in a continuous flow mode. As in SBRs, applying a selection pressure, based on differences in either settling velocity or the size of the biomass, is essential for successful granulation in continuous flow reactors (CFRs). CFRs employed for aerobic granulation come in multiple configurations, each with their own means of achieving such a selection pressure. Other factors, such as bioaugmentation and hydraulic shear force, also contribute to aerobic granulation to some extent. Besides the formation of aerobic granules, long-term stability of aerobic granules is also a critical issue to be addressed. Inorganic precipitation, special inocula, and various operational optimization strategies have been used to improve granule long-term structural integrity. Accumulated studies reviewed in this work demonstrate that aerobic granulation in CFRs is capable of removing a wide spectrum of contaminants and achieving properties generally comparable to those in SBRs. Despite the notable research progress made toward successful aerobic granulation in lab-scale CFRs, to the best of our knowledge, there are only three full-scale tests of the technique, two being seeded with anammox-supported aerobic granules and the other with conventional aerobic granules; two other process alternatives are currently in development. Application of settling- or size-based selection pressures and feast/famine conditions are especially difficult to implement to these and similar mainstream systems. Future research efforts needs to be focused on the optimization of the granule-to-floc ratio, enhancement of granule activity, improvement of long-term granule stability, and a better understanding of aerobic granulation mechanisms in CFRs, especially in full-scale applications. Copyright © 2018 Elsevier Inc. All rights reserved.

Neutrophil-derived hyperresistinemia in severe acute streptococcal infections.

PubMed

Johansson, Linda; Linnér, Anna; Sundén-Cullberg, Jonas; Haggar, Axana; Herwald, Heiko; Loré, Karin; Treutiger, Carl-Johan; Norrby-Teglund, Anna

2009-09-15

The concept of neutrophil activation and degranulation as important contributors to disease pathology in invasive group A streptococcal infections has recently been emphasized. This study focuses on two of the most severe streptococcal manifestations, toxic shock syndrome and necrotizing fasciitis, and the newly described proinflammatory molecule resistin, known to derive from adipocytes and monocytes. We demonstrate for the first time that these conditions are characterized by hyperresistinemia in circulation as well as at the local site of infection. Importantly, analyses of patient tissue biopsies and whole blood revealed that neutrophils represent a novel and dominant source of resistin in bacterial septic shock. This was confirmed by the identification of resistin within neutrophil azurophilic granules. In vitro assays using primary neutrophils showed that resistin release was readily triggered by streptococcal cell wall components and by the streptococcal M1 protein, but not by the potent streptococcal superantigens. This is the first report demonstrating that resistin is released from neutrophils in response to microbial stimuli, which adds resistin to the neutrophil granule proteins that are likely to contribute to the pathologic inflammatory responses associated with severe streptococcal infections.

Identification of Grandchildless Loci Whose Products Are Required for Normal Germ-Line Development in the Nematode Caenorhabditis Elegans

PubMed Central

Capowski, E. E.; Martin, P.; Garvin, C.; Strome, S.

1991-01-01

To identify genes that encode maternal components required for development of the germ line in the nematode Caenorhabditis elegans, we have screened for mutations that confer a maternal-effect sterile or ``grandchildless'' phenotype: homozygous mutant hermaphrodites produced by heterozygous mothers are themselves fertile, but produce sterile progeny. Our screens have identified six loci, defined by 21 mutations. This paper presents genetic and phenotypic characterization of four of the loci. The majority of mutations, those in mes-2, mes-3 and mes-4, affect postembryonic germ-line development; the progeny of mutant mothers undergo apparently normal embryogenesis but develop into agametic adults with 10-1000-fold reductions in number of germ cells. In contrast, mutations in mes-1 cause defects in cytoplasmic partitioning during embryogenesis, and the resulting larvae lack germ-line progenitor cells. Mutations in all of the mes loci primarily affect the germ line, and none disrupt the structural integrity of germ granules. This is in contrast to grandchildless mutations in Drosophila melanogaster, all of which disrupt germ granules and affect abdominal as well as germ-line development. PMID:1783292

Granulation of fine powder

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Ching-Fong

A mixture of fine powder including thorium oxide was converted to granulated powder by forming a first-green-body and heat treating the first-green-body at a high temperature to strengthen the first-green-body followed by granulation by crushing or milling the heat-treated first-green-body. The granulated powder was achieved by screening through a combination of sieves to achieve the desired granule size distribution. The granulated powder relies on the thermal bonding to maintain its shape and structure. The granulated powder contains no organic binder and can be stored in a radioactive or other extreme environment. The granulated powder was pressed and sintered to formmore » a dense compact with a higher density and more uniform pore size distribution.« less

The effect of the chopper on granules from wet high-shear granulation using a PMA-1 granulator.

PubMed

Briens, Lauren; Logan, Ryan

2011-12-01

Chopper presence and then chopper speed was varied during wet high shear granulation of a placebo formulation using a PMA-1 granulator while also varying the impeller speed. The granules were extensively analyzed for differences due to the chopper. The effect of the chopper on the granules varied with impeller speed from no effect at a low impeller speed of 300 rpm to flow interruptions at an impeller speed of 700 rpm to minimal impact at very high impeller speeds as caking at the bowl perimeter obscured the effect of the chopper on the flow pattern. Differences in the granule flowability were minimal. However, it was concluded that the largest fraction of optimal granules would be obtained at an impeller speed of 700 rpm with the chopper at 1,000 rpm allowing balances between flow establishment, segregation, and centrifugal forces.

Characteristics of aerobic granules grown on glucose and acetate in sequential aerobic sludge blanket reactors.

PubMed

Tay, J H; Liu, Q S; Liu, Y

2002-08-01

Aerobic granules were cultivated in two column-type sequential aerobic sludge blanket reactors fed with glucose and acetate, respectively. The characteristics of aerobic granules were investigated. Results indicated that the glucose- and acetate-fed granules have comparable characteristics in terms of settling velocity, size, shape, biomass density, hydrophobicity, physical strength, microbial activity and storage stability. Substrate component does not seem to be a key factor on the formation of aerobic granules. However, microbial diversity of the granules is closely associated with the carbon sources supplied to the reactors. Compared with the conventional activated sludge flocs, aerobic granules exhibit excellent physical characteristics that would be essential for industrial application. This research provides a complete set of characteristics data of aerobic granules grown on glucose and acetate, which would be useful for further development of aerobic granules-based compact bioreactor for handling high strength organic wastewater.

Starch-Branching Enzymes Preferentially Associated with A-Type Starch Granules in Wheat Endosperm1

PubMed Central

Peng, Mingsheng; Gao, Ming; Båga, Monica; Hucl, Pierre; Chibbar, Ravindra N.

2000-01-01

Two starch granule-bound proteins (SGP), SGP-140 and SGP-145, were preferentially associated with A-type starch granules (>10 μm) in developing and mature wheat (Triticum aestivum) kernels. Immunoblotting and N-terminal sequencing suggested that the two proteins were different variants of SBEIc, a 152-kD isoform of wheat starch-branching enzyme. Both SGP-140 and SGP-145 were localized to the endosperm starch granules but were not found in the endosperm soluble fraction or pericarp starch granules younger than 15 d post anthesis (DPA). Small-size starch granules (<10 μm) initiated before 15 DPA incorporated SGP-140 and SGP-145 throughout endosperm development and grew into full-size A-type starch granules (>10 μm). In contrast, small-size starch granules harvested after 15 DPA contained only low amounts of SGP-140 and SGP-145 and developed mainly into B-type starch granules (<10 μm). Polypeptides of similar mass and immunologically related to SGP-140 and/or SGP-145 were also preferentially incorporated into A-type starch granules of barley (Hordeum vulgare), rye (Secale cereale), and triticale (× Triticosecale Wittmack) endosperm, which like wheat endosperm have a bimodal starch granule size distribution. PMID:10982441

Comparison of waxy and normal potato starch remaining granules after chemical surface gelatinization: pasting behavior and surface morphology.

PubMed

Huang, Junrong; Chen, Zhenghong; Xu, Yalun; Li, Hongliang; Liu, Shuxing; Yang, Daqing; Schols, Henk A

2014-02-15

To understand the contribution of granule inner portion to the pasting property of starch, waxy potato starch and two normal potato starches and their acetylated starch samples were subjected to chemical surface gelatinization by 3.8 mol/L CaCl2 to obtain remaining granules. Native and acetylated, original and remaining granules of waxy potato starch had similar rapid visco analyzer (RVA) pasting profiles, while those of two normal potato starches behaved obviously different from each other. All remaining granules had lower peak viscosity than the corresponding original granules. Contribution of waxy potato starch granule's inner portion to the peak viscosity was significant more than those of normal potato starches. The shell structure appearing on the remaining granule surface for waxy potato starch was smoother and thinner than that for normal potato starches as observed by scanning electron microscopy, indicating a more regular structure of shell and a more ordered packing of shell for waxy potato starch granules. The blocklet size of waxy potato starch was smaller and more uniform than those of normal potato starches as shown by atomic force microscopy images of original and remaining granules. In general, our results provided the evidence for the spatial structure diversity between waxy and normal potato starch granules: outer layer and inner portion of waxy potato starch granule had similar structure, while outer layer had notably different structure from inner portion for normal potato starch granule. Copyright © 2013 Elsevier Ltd. All rights reserved.

The role of two isoenzymes of alpha-amylase of Araucaria araucana (Araucariaceae) on the digestion of starch granules during germination.

PubMed

Waghorn, Juana J; del Pozo, Talía; Acevedo, Elba A; Cardemil, Liliana A

2003-03-01

Starch is the principal reserve of Araucaria araucana seeds, and it is hydrolysed during germination mainly by alpha-amylase. There are several alpha-amylase isoenzymes whose patterns change in the embryo and in the megagametophyte from the one observed in quiescent seeds (T(0)) to a different one observed 90 h after imbibition (T(90)). The objective of this research was to study the roles of two purified alpha-amylase isoenzymes by in vitro digestion of starch granules extracted from the tissues at two times of imbibition: one is abundant in quiescent seeds and the other is abundant after 90 h of imbibition. The isoenzymes digested the starch granules of their own stage of germination better, since the isoenzyme T(0) digested starch granules mainly from quiescent seeds, while the isoenzyme T(90) digested starch mainly at 90 h of imbibition. The sizes of the starch granule and the tissue from which these granules originated make a difference to digestion by the isoenzymes. Embryonic isoenzyme T(0) digested large embryonic starch granules better than small and medium-sized granules, and better than those isolated from megagametophytes. Similarly isoenzyme T(90) digested small embryonic starch granules better than medium-sized and large granules, and better than those isolated from megagametophytes. However, a mixture of partially purified megagametophytic isoenzymes T(0) and T(90) digested the megagametophytic granules better than those isolated from embryos. Studies of in vitro sequential digestion of starch granules with these isoenzymes corroborated their specificity. The isoenzyme T(90) digested starch granules previously digested by the isoenzyme T(0). This suggests that in vivo these two isoenzymes may act sequentially in starch granule digestion.

Reducing the startup time of aerobic granular sludge reactors through seeding floccular sludge with crushed aerobic granules.

PubMed

Pijuan, Maite; Werner, Ursula; Yuan, Zhiguo

2011-10-15

One of the main challenging issues for the aerobic granular sludge technology is the long startup time when dealing with real wastewaters. This study presents a novel strategy to reduce the time required for granulation while ensuring a high level of nutrient removal. This new approach consists of seeding the reactor with a mixture of crushed aerobic granules and floccular sludge. The effectiveness of the strategy was demonstrated using abattoir wastewater, containing nitrogen and phosphorus at approximately 250 mgN/L and 30 mgP/L, respectively. Seven different mixtures of crushed granules and floccular sludge at granular sludge fractions (w/w in dry mass) of 0%, 5%, 10%, 15%, 25%, 30% and 50% were used to start eight granulation processes. The granulation time (defined as the time when the 10th percentile bacterial aggregate size is larger than 200 μm) displayed a strong dependency on the fraction of granular sludge. The shortest granulation time of 18 days was obtained with 50% crushed granules, in comparison with 133 days with 5% crushed granules. Full granulation was not achieved in the two trials without seeding with crushed granules. In contrast to the 100% floccular sludge cases, where a substantial loss of biomass occurred during granulation, the biomass concentration in all other trails did not decrease during granulation. This allowed that good nitrogen removal was maintained in all the reactors during the granulation process. However, enhanced biological phosphorus removal was achieved in only one of the eight trials. This was likely due to the temporary accumulation of nitrite, a strong inhibitor of polyphosphate accumulating organisms. Copyright © 2011 Elsevier Ltd. All rights reserved.

High-rate hydrogenotrophic methanogenesis for biogas upgrading: the role of anaerobic granules.

PubMed

Xu, Heng; Gong, Shufen; Sun, Yuanzi; Ma, Hailing; Zheng, Mingyue; Wang, Kaijun

2015-01-01

Hydrogenotrophic methanogenesis has been proved to be a feasible biological method for biogas upgrading. To improve its performance, the feasibility of typical anaerobic granules as the inoculum was investigated in both batch and continuous experiments. The results from batch experiments showed that glucose-acclimated granules seemed to perform better than granules acclimated to acidified products (AP, i.e. acetate, propionate and ethanol) in in situ biogas upgrading systems and a slightly higher H2 consumption rate (1.5 mmol H2 g VSS(-1) h(-1)) was obtained for glucose-acclimated granules. For AP-acclimated granules, the inhibition on anaerobic digestion and pH increase (up to 9.55±0.16) took place, and the upgrading performance was adversely affected. In contrast, better performance for AP-acclimated granules was observed in ex situ systems, possibly due to their higher hydrogenotrophic methanogenic activities (HMA). Moreover, when gas-liquid mass transfer limitations were alleviated, the upgrading performance was significantly improved (three-fold) for both glucose-acclimated and AP-acclimated granules. The HMA of anaerobic granules could be further enhanced to improve biogas upgrading performance via continuous cultivation with H2/CO2 as the sole substrate. During the three months' cultivation, secondary granulation and microbial population shift were observed, but anaerobic granules still remained intact and their HMA increased from 0.2 to 0.6 g COD g VSS(-1) d(-1). It indicated that the formation of hydrogenotrophic methanogenic granules, a new type of anaerobic granules specialized for high-rate hydrogenotrophic methanogenesis and biogas upgrading, might be possible. Conclusively, anaerobic granules showed great potential for biogas upgrading.

Investigation of internal structure of fine granules by microtomography using synchrotron X-ray radiation.

PubMed

Noguchi, Shuji; Kajihara, Ryusuke; Iwao, Yasunori; Fujinami, Yukari; Suzuki, Yoshio; Terada, Yasuko; Uesugi, Kentaro; Miura, Keiko; Itai, Shigeru

2013-03-10

Computed tomography (CT) using synchrotron X-ray radiation was evaluated as a non-destructive structural analysis method for fine granules. Two kinds of granules have been investigated: a bromhexine hydrochloride (BHX)-layered Celphere CP-102 granule coated with pH-sensitive polymer Kollicoat Smartseal 30-D, and a wax-matrix granule constructed from acetaminophen (APAP), dibasic calcium phosphate dehydrate, and aminoalkyl methacrylate copolymer E (AMCE) manufactured by melt granulation. The diameters of both granules were 200-300 μm. CT analysis of CP-102 granule could visualize the laminar structures of BHX and Kollicoat layers, and also visualize the high talc-content regions in the Kollicoat layer that could not be detected by scanning electron microscopy. Moreover, CT analysis using X-ray energies above the absorption edge of Br specifically enhanced the contrast in the BHX layer. As for granules manufactured by melt granulation, CT analysis revealed that they had a small inner void space due to a uniform distribution of APAP and other excipients. The distribution of AMCE revealed by CT analysis was also found to involve in the differences of drug dissolution from the granules as described previously. These observations demonstrate that CT analysis using synchrotron X-ray radiation is a powerful method for the detailed internal structure analysis of fine granules. Copyright © 2013 Elsevier B.V. All rights reserved.

Homologs of PROTEIN TARGETING TO STARCH Control Starch Granule Initiation in Arabidopsis Leaves[OPEN

PubMed Central

David, Laure C.; Abt, Melanie; Lu, Kuan-Jen

2017-01-01

The molecular mechanism that initiates the synthesis of starch granules is poorly understood. Here, we discovered two plastidial proteins involved in granule initiation in Arabidopsis thaliana leaves. Both contain coiled coils and a family-48 carbohydrate binding module (CBM48) and are homologs of the PROTEIN TARGETING TO STARCH (PTST) protein; thus, we named them PTST2 and PTST3. Chloroplasts in mesophyll cells typically contain five to seven granules, but remarkably, most chloroplasts in ptst2 mutants contained zero or one large granule. Chloroplasts in ptst3 had a slight reduction in granule number compared with the wild type, while those of the ptst2 ptst3 double mutant contained even fewer granules than ptst2. The ptst2 granules were larger but similar in morphology to wild-type granules, but those of the double mutant had an aberrant morphology. Immunoprecipitation showed that PTST2 interacts with STARCH SYNTHASE4 (SS4), which influences granule initiation and morphology. Overexpression of PTST2 resulted in chloroplasts containing many small granules, an effect that was dependent on the presence of SS4. Furthermore, isothermal titration calorimetry revealed that the CBM48 domain of PTST2, which is essential for its function, interacts with long maltooligosaccharides. We propose that PTST2 and PTST3 are critical during granule initiation, as they bind and deliver suitable maltooligosaccharide primers to SS4. PMID:28684429

Chromaffin granules in the rat adrenal medulla release their secretory content in a particulate fashion.

PubMed

Crivellato, Enrico; Belloni, Anna; Nico, Beatrice; Nussdorfer, Gastone G; Ribatti, Domenico

2004-03-01

Exocytosis is considered the main route of granule discharge in chromaffin cells. We recently provided ultrastructural evidence suggesting that piecemeal degranulation (PMD) occurs in mouse adrenal chromaffin cells. In the present study, we processed rat adrenal glands for transmission electron microscopy (TEM), and examined chromaffin cells for changes characteristic of PMD. Both adrenaline (A)- and noradrenaline (NA)-storing cells express ultrastructural features suggestive of a slow and particulate mode of granule discharge. In adrenaline-containing cells, some granules present enlarged dimensions accompanied by eroded or dissolved matrices. Likewise, a number of granules in NA-releasing cells show content reduction with variably expanded granule chambers. Dilated, empty granule containers are recognizable in the cytoplasm of both cell types. Characteristically, altered granules and empty containers are seen intermingled with normal, resting granules. In addition, chromaffin granules often show irregular profiles, with budding or tail-like projections of their limiting membranes. Thirty 150-nm-diameter membrane-bound vesicles with a moderately electron-dense or -lucent internal structure are observable in the cytoplasm of both cell types. These vesicles are seen among the granules and some of them are fused with the perigranule membranes in the process of attachment to or budding from the granules. These data add further support to the concept that PMD may be an alternative secretory pathway in adrenal chromaffin cells. Copyright 2004 Wiley-Liss, Inc.

Layered growth with bottom-spray granulation for spray deposition of drug.

PubMed

Er, Dawn Z L; Liew, Celine V; Heng, Paul W S

2009-07-30

The gap in scientific knowledge on bottom-spray fluidized bed granulation has emphasized the need for more studies in this area. This paper comparatively studied the applicability of a modified bottom-spray process and the conventional top-spray process for the spray deposition of a micronized drug during granulation. The differences in circulation pattern, mode of growth and resultant granule properties between the two processes were highlighted. The more ordered and consistent circulation pattern of particles in a bottom-spray fluidized bed was observed to give rise to layered granule growth. This resulted in better drug content uniformity among the granule batches and within a granule batch. The processes' sensitivities to wetting and feed material characteristics were also compared and found to differ markedly. Less robustness to differing process conditions was observed for the top-spray process. The resultant bottom-spray granules formed were observed to be less porous, more spherical and had good flow properties. The bottom-spray technique can thus be potentially applied for the spray deposition of drug during granulation and was observed to be a good alternative to the conventional technique for preparing granules.

Injected mass deposition thresholds for lithium granule instigated triggering of edge localized modes on EAST

NASA Astrophysics Data System (ADS)

Lunsford, R.; Sun, Z.; Maingi, R.; Hu, J. S.; Mansfield, D.; Xu, W.; Zuo, G. Z.; Diallo, A.; Osborne, T.; Tritz, K.; Canik, J.; Huang, M.; Meng, X. C.; Gong, X. Z.; Wan, B. N.; Li, J. G.; the EAST Team

2018-03-01

The ability of an injected lithium granule to promptly trigger an edge localized mode (ELM) has been established in multiple experiments. By horizontally injecting granules ranging in diameter from 200 microns to 1 mm in diameter into the low field side of EAST H-mode discharges we have determined that granules with diameter  >600 microns are successful in triggering ELMs more than 95% of the time. It was also demonstrated that below 600 microns the triggering efficiency decreased roughly with granule size. Granules were radially injected from the outer midplane with velocities ~80 m s-1 into EAST upper single null discharges with an ITER like tungsten monoblock divertor. These granules were individually tracked throughout their injection cycle in order to determine their efficacy at triggering an ELM. For those granules of sufficient size, ELM triggering was a prompt response to granule injection. By simulating the granule injection with an experimentally benchmarked neutral gas shielding (NGS) model, the ablatant mass deposition required to promptly trigger an ELM is calculated and the fractional mass deposition is determined.

Compressibility and compactibility of granules produced by wet and dry granulation.

PubMed

Bacher, C; Olsen, P M; Bertelsen, P; Sonnergaard, J M

2008-06-24

The bulk properties, compactibility and compressibility of granules produced by wet and dry granulation were compared applying a rotary tablet press, three different morphological forms of calcium carbonate and two particle sizes of sorbitol. Granules from both granulation methods possessed acceptable flow properties; however, the ground (Mikhart) and cubic (Scoralite) calcium carbonate demonstrated better die-filling abilities in the tablet press than the scalenhedral calcium carbonate (Sturcal). The wet processed granules showed in general larger compression properties. This was explained as these granules were mechanical stronger and had a higher initial porosity. In some cases, a large particle surface area of calcium carbonate and sorbitol resulted in a small, insignificant improvement of the consolidation characteristics. A correlation between the compression and compaction characteristics was demonstrated.

Soft and Hard Textured Wheat Differ in Starch Properties as Indicated by Trimodal Distribution, Morphology, Thermal and Crystalline Properties

PubMed Central

Kumar, Rohit; Kumar, Aman; Sharma, Nand Kishor; Kaur, Navneet; Chunduri, Venkatesh; Chawla, Meenakshi; Sharma, Saloni; Singh, Kashmir; Garg, Monika

2016-01-01

Starch and proteins are major components in the wheat endosperm that affect its end product quality. Between the two textural classes of wheat i.e. hard and soft, starch granules are loosely bound with the lipids and proteins in soft wheat due to higher expression of interfering grain softness proteins. It might have impact on starch granules properties. In this work for the first time the physiochemical and structural properties of different sized starch granules (A-, B- and C-granules) were studied to understand the differences in starches with respect to soft and hard wheat. A-, B- and C-type granules were separated with >95% purity. Average number and proportion of A-, B-, and C-type granules was 18%, 56%, 26% and 76%, 19%, 5% respectively. All had symmetrical birefringence pattern with varied intensity. All displayed typical A-type crystallites. A-type granules also showed V-type crystallinity that is indicative of starch complexes with lipids and proteins. Granules differing in gelatinization temperature (ΔH) and transition temperature (ΔT), showed different enthalpy changes during heating. Substitution analysis indicated differences in relative substitution pattern of different starch granules. Birefringence, percentage crystallinity, transmittance, gelatinization enthalpy and substitution decreased in order of A>B>C being higher in hard wheat than soft wheat. Amylose content decreased in order of A>B>C being higher in soft wheat than hard wheat. Reconstitution experiment showed that starch properties could be manipulated by changing the composition of starch granules. Addition of A-granules to total starch significantly affected its thermal properties. Effect of A-granule addition was higher than B- and C-granules. Transmittance of the starch granules paste showed that starch granules of hard wheat formed clear paste. These results suggested that in addition to differences in protein concentration, hard and soft wheat lines have differences in starch composition also. PMID:26824830

Soft and Hard Textured Wheat Differ in Starch Properties as Indicated by Trimodal Distribution, Morphology, Thermal and Crystalline Properties.

PubMed

Kumar, Rohit; Kumar, Aman; Sharma, Nand Kishor; Kaur, Navneet; Chunduri, Venkatesh; Chawla, Meenakshi; Sharma, Saloni; Singh, Kashmir; Garg, Monika

2016-01-01

Starch and proteins are major components in the wheat endosperm that affect its end product quality. Between the two textural classes of wheat i.e. hard and soft, starch granules are loosely bound with the lipids and proteins in soft wheat due to higher expression of interfering grain softness proteins. It might have impact on starch granules properties. In this work for the first time the physiochemical and structural properties of different sized starch granules (A-, B- and C-granules) were studied to understand the differences in starches with respect to soft and hard wheat. A-, B- and C-type granules were separated with >95% purity. Average number and proportion of A-, B-, and C-type granules was 18%, 56%, 26% and 76%, 19%, 5% respectively. All had symmetrical birefringence pattern with varied intensity. All displayed typical A-type crystallites. A-type granules also showed V-type crystallinity that is indicative of starch complexes with lipids and proteins. Granules differing in gelatinization temperature (ΔH) and transition temperature (ΔT), showed different enthalpy changes during heating. Substitution analysis indicated differences in relative substitution pattern of different starch granules. Birefringence, percentage crystallinity, transmittance, gelatinization enthalpy and substitution decreased in order of A>B>C being higher in hard wheat than soft wheat. Amylose content decreased in order of A>B>C being higher in soft wheat than hard wheat. Reconstitution experiment showed that starch properties could be manipulated by changing the composition of starch granules. Addition of A-granules to total starch significantly affected its thermal properties. Effect of A-granule addition was higher than B- and C-granules. Transmittance of the starch granules paste showed that starch granules of hard wheat formed clear paste. These results suggested that in addition to differences in protein concentration, hard and soft wheat lines have differences in starch composition also.

Characterization and taste-masking evaluation of acetaminophen granules: comparison between different preparation methods in a high-shear mixer.

PubMed

Albertini, Beatrice; Cavallari, Cristina; Passerini, Nadia; Voinovich, Dario; González-Rodríguez, Marisa L; Magarotto, Lorenzo; Rodriguez, Lorenzo

2004-02-01

The aim of this study was to prepare and to investigate acetaminophen taste-masked granules obtained in a high-shear mixer using three different wet granulation methods (method A: water granulation, method B: granulation with a polyvinylpyrrolidone (PVP) binding solution and method C: steam granulation). The studied formulation was: acetaminophen 15%, alpha-lactose monohydrate 30%, cornstarch 45%, polyvinylpyrrolidone K30 5% and orange flavour 5% (w/w). In vitro dissolution studies, performed at pH 6.8, showed that steam granules enabled the lower dissolution rate in comparison to the water and binding solution granules; these results were then confirmed by their lower surface reactivity (D(R)) during the dissolution process. Moreover, the results of the gustatory sensation test performed by six volunteers confirmed the taste-masking effects of the granules, especially steam granules (P<0.001). Morphological, fractal and porosity analysis were then performed to explain the dissolution profiles and the results of the gustatory sensation test. Scanning electron microscopy (SEM) analysis revealed the smoother and the more regular surface of steam granules with respect to the samples obtained using methods A and B; these results were also confirmed by their lower fractal dimension (D(s)) and porosity values. Finally, differential scanning calorimetry (DSC) results showed a shift of the melting point of the drug, which was due to the simple mixing of the components and not to the granulation processes. In conclusion, the steam granulation technique resulted a suitable method to comply the purpose of this work, without modifying the availability of the drug.

Continuous twin screw granulation of controlled release formulations with various HPMC grades.

PubMed

Vanhoorne, V; Janssens, L; Vercruysse, J; De Beer, T; Remon, J P; Vervaet, C

2016-09-25

HPMC is a popular matrix former to formulate tablets with extended drug release. Tablets with HPMC are preferentially produced by direct compression. However, granulation is often required prior to tableting to overcome poor flowability of the formulation. While continuous twin screw granulation has been extensively evaluated for granulation of immediate release formulations, twin screw granulation of controlled release formulations including the dissolution behavior of the formulations received little attention. Therefore, the influence of the HPMC grade (viscosity and substitution degree) and the particle size of theophylline on critical quality attributes of granules (continuously produced via twin screw granulation) and tablets was investigated in the current study. Formulations with 20 or 40% HPMC, 20% theophylline and lactose were granulated with water at fixed process parameters via twin screw granulation. The torque was influenced by the viscosity and substitution degree of HPMC, but was not a limiting factor for the granulation process. An optimal L/S ratio was selected for each formulation based on the granule size distribution. The granule size distributions were influenced by the substitution degree and concentration of HPMC and the particle size of theophylline. Raman and UV spectroscopic analysis on 8 sieve fractions of granules indicated an inhomogeneous distribution of theophylline over the size fractions. However, this phenomenon was not correlated with the hydration rate or viscosity of HPMC. Controlled release of theophylline could be obtained over 24h with release profiles close to zero-order. The release of theophylline could be tailored via selection of the substitution degree and viscosity of HPMC. Copyright © 2016 Elsevier B.V. All rights reserved.

Stochastic Seeding Coupled with mRNA Self-Recruitment Generates Heterogeneous Drosophila Germ Granules.

PubMed

Niepielko, Matthew G; Eagle, Whitby V I; Gavis, Elizabeth R

2018-06-18

The formation of ribonucleoprotein assemblies called germ granules is a conserved feature of germline development. In Drosophila, germ granules form at the posterior of the oocyte in a specialized cytoplasm called the germ plasm, which specifies germline fate during embryogenesis. mRNAs, including nanos (nos) and polar granule component (pgc), that function in germline development are localized to the germ plasm through their incorporation into germ granules, which deliver them to the primordial germ cells. Germ granules are nucleated by Oskar (Osk) protein and contain varying combinations and quantities of their constituent mRNAs, which are organized as spatially distinct, multi-copy homotypic clusters. The process that gives rise to such heterogeneous yet organized granules remains unknown. Here, we show that individual nos and pgc transcripts can populate the same nascent granule, and these first transcripts then act as seeds, recruiting additional like transcripts to form homotypic clusters. Within a granule, homotypic clusters grow independently of each other but depend on the simultaneous acquisition of additional Osk. Although granules can contain multiple clusters of a particular mRNA, granule mRNA content is dominated by cluster size. These results suggest that the accumulation of mRNAs in the germ plasm is controlled by the mRNAs themselves through their ability to form homotypic clusters; thus, RNA self-association drives germ granule mRNA localization. We propose that a stochastic seeding and self-recruitment mechanism enables granules to simultaneously incorporate many different mRNAs while ensuring that each becomes enriched to a functional threshold. Copyright © 2018 Elsevier Ltd. All rights reserved.

Extended-release mesalamine granules for ulcerative colitis.

PubMed

Love, Bryan L; Miller, April D

2012-11-01

To evaluate the efficacy and safety of extended-release mesalamine granules in the maintenance of remission in ulcerative colitis (UC). Literature was obtained through searches of MEDLINE (1990-June 2012) using the terms mesalamine granules, ulcerative colitis, Apriso, and Salofalk. Bibliographies from retrieved articles were searched for additional citations. All English-language articles reporting on use of extended-release mesalamine granules in humans identified through the search were evaluated and included. The preferred initial treatment for induction and maintenance of remission in mild to moderate UC is agents from the 5-aminosalicylate class (balsalazide, mesalamine, olsalazine, sulfasalazine). Mesalamine granules are available as an encapsulated product in the US and as a nonencapsulated formulation in Europe. Data evaluating encapsulated mesalamine granules for induction of remission are lacking; however, the European mesalamine granule formulation has been evaluated for induction of remission. Patients receiving mesalamine granules for induction achieved clinical and endoscopic remission more frequently than those receiving placebo. Two pivotal, randomized, double-blind, placebo-controlled, multicenter studies have evaluated encapsulated mesalamine granules for maintenance in 562 adults in remission from UC. In both studies, the proportion of patients who remained relapse-free at 6 months was higher for those receiving encapsulated mesalamine granules than placebo. Mesalamine granules are well tolerated, with headache, nausea, and upper respiratory infections being the most frequently reported adverse effects. Current evidence supports the use of extended-release mesalamine granules for maintenance of remission in mild to moderate UC. Further studies are necessary to examine the ideal dose and regimen of encapsulated mesalamine granules for induction of remission in UC.

The morphology of solar granulations and dark networks

NASA Astrophysics Data System (ADS)

Graves, J. Elon; Pierce, A. Keith

1986-08-01

Solar granules are classified into four groups based on shape and splitting by sharp rifts crossing them. Grains are classified as: single granules varying in size from 1/8 to 3 in., single granules embayed by a broad dark area or possessing a central darkening, single granules split by very narrow rifts which are significantly narrower than the intergranular lanes, and complexes of granules displaying a daisy pattern. The formation and growth of 'white-light dark networks' are also discussed

Statistical properties of solar granulation derived from the SOUP instrument on Spacelab 2

NASA Technical Reports Server (NTRS)

Title, A. M.; Tarbell, T. D.; Topka, K. P.; Ferguson, S. H.; Shine, R. A.

1989-01-01

Computer algorithms and statistical techniques were used to identify, measure, and quantify the properties of solar granulation derived from movies collected by the Solar Optical Universal Polarimeter on Spacelab 2. The results show that there is neither a typical solar granule nor a typical granule evolution. A granule's evolution is dependent on local magnetic flux density, its position with respect to the active region plage, its position in the mesogranulation pattern, and the evolution of granules in its immediate neighborhood.

Effect of suspension property on granule morphology and compaction behavior

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hae-Weon Lee, Guesup Song, In-Sik Suk

1995-12-31

Granule morphology is an important factor during dry pressing, since it has great influences on die flowability, compaction ratio, and resulting green microstructure. Granule morphology and packing structure of ultrafine Si{sub 3}N{sub 4} particles in the granule were optimized during spray drying by adjusting the suspension structure. The particle packing structure of spray-dried granule was investigated with suspension structure. The effects of granule morphology and its particle packing structure on compaction and resultant sintering behavior were evaluated.

Simultaneous Comparison of Two Roller Compaction Techniques and Two Particle Size Analysis Methods.

PubMed

Saarinen, Tuomas; Antikainen, Osmo; Yliruusi, Jouko

2017-11-01

A new dry granulation technique, gas-assisted roller compaction (GARC), was compared with conventional roller compaction (CRC) by manufacturing 34 granulation batches. The process variables studied were roll pressure, roll speed, and sieve size of the conical mill. The main quality attributes measured were granule size and flow characteristics. Within granulations also the real applicability of two particle size analysis techniques, sieve analysis (SA) and fast imaging technique (Flashsizer, FS), was tested. All granules obtained were acceptable. In general, the particle size of GARC granules was slightly larger than that of CRC granules. In addition, the GARC granules had better flowability. For example, the tablet weight variation of GARC granules was close to 2%, indicating good flowing and packing characteristics. The comparison of the two particle size analysis techniques showed that SA was more accurate in determining wide and bimodal size distributions while FS showed narrower and mono-modal distributions. However, both techniques gave good estimates for mean granule sizes. Overall, SA was a time-consuming but accurate technique that provided reliable information for the entire granule size distribution. By contrast, FS oversimplified the shape of the size distribution, but nevertheless yielded acceptable estimates for mean particle size. In general, FS was two to three orders of magnitude faster than SA.

The use of Rheology Combined with Differential Scanning Calorimetry to Elucidate the Granulation Mechanism of an Immiscible Formulation During Continuous Twin-Screw Melt Granulation.

PubMed

Monteyne, Tinne; Heeze, Liza; Mortier, Severine Therese F C; Oldörp, Klaus; Cardinaels, Ruth; Nopens, Ingmar; Vervaet, Chris; Remon, Jean-Paul; De Beer, Thomas

2016-10-01

Twin screw hot melt granulation (TS HMG) is a valuable, but still unexplored alternative to continuous granulation of moisture sensitive drugs. However, knowledge of the material behavior during TS HMG is crucial to optimize the formulation, process and resulting granule properties. The aim of this study was to evaluate the agglomeration mechanism during TS HMG using a rheometer in combination with differential scanning calorimetry (DSC). An immiscible drug-binder formulation (caffeine-Soluplus(®)) was granulated via TS HMG in combination with thermal and rheological analysis (conventional and Rheoscope), granule characterization and Near Infrared chemical imaging (NIR-CI). A thin binder layer with restricted mobility was formed on the surface of the drug particles during granulation and is covered by a second layer with improved mobility when the Soluplus(®) concentration exceeded 15% (w/w). The formation of this second layer was facilitated at elevated granulation temperatures and resulted in smaller and more spherical granules. The combination of thermal and rheological analysis and NIR-CI images was advantageous to develop in-depth understanding of the agglomeration mechanism during continuous TS HMG and provided insight in the granule properties as function of process temperature and binder concentration.

Studies on the compressibility of wax matrix granules of acetaminophen and their admixtures with various tableting bases.

PubMed

Uhumwangho, M U; Okor, R S

2006-04-01

Matrix granules of acetaminophen have been formed by a melt granulation process whereby the acetaminophen powder was triturated with the melted wax--goat wax, glyceryl monostearate or carnuba wax. The compressibility of the matrix granules and their admixture, with diluent granules (lactose, alpha-cellulose or microcrystalline cellulose) was investigated. The granules were compressed to tablets at a constant load (30 arbitrary units on the load scale) of a manesty single punch machine. Resulting tablets were evaluated for tensile strength (T) and disintegration times (DT). Granule flow was determined by measuring their angle of repose when allowed to fall freely on a level surface. Matrix granules prepared by melt granulation with goat wax or glyceryl monostearate were too sticky and therefore did not flow at all. They were also poorly compressible (T values = 0.20MN/m2). Inclusion of the diluent remarkably improved granule flow property and compressibility. The T values of the tablets (measure of compressibility) increased from about 0.24 to 0.65 MN/m2 during increase in diluent (lactose) content from 20 to 80 %w/w. Microcrystalline cellulose and alpha-cellulose were more effective than lactose in promoting compressibility of the granules. By contrast the matrix granules formed with carnuba wax were free flowing (angle of repose, 18.60). Addition of the diluent further improved flowability slightly. The matrix granules (without a diluent) were readily compressible (T value, 1.79MN/m2). Addition of the diluent (80%w/w) reduced T values (MN/m2) slightly to 1.32 (lactose), 1.48 (alpha-cellulose) and 1.74 (microcrystalline cellulose). Tablets of the matrix granules only, disintegrated rapidly within 3 minutes. DT was further reduced to <30 s by addition of any of the diluents. The indication is that the inclusion of the diluents studied can be used to improve the compressibility of the otherwise poorly compressible matrix granules. Based on the flowability, compressibility, and disintegration data, carnuba wax proved most promising in the melt granulation of the test drug for sustained release applications.

Improved tabletability after a polymorphic transition of delta-mannitol during twin screw granulation.

PubMed

Vanhoorne, V; Bekaert, B; Peeters, E; De Beer, T; Remon, J-P; Vervaet, C

2016-06-15

In most formulations processed via continuous twin screw granulation microcrystalline cellulose (MCC) and/or lactose are used as excipients, but mannitol is also a preferred excipient for wet granulation and tableting due to its non-hygroscopicity and inertness. Therefore, the aim of the current study was to investigate the influence of process parameters on critical quality attributes of granules (moisture content, solid state, morphology, size distribution, specific surface area, friability, flowability and hygroscopicity) and tablets (tensile strength and friability) after twin screw granulation of δ-mannitol. The δ-polymorph was selected since a moisture-induced transformation to β-mannitol was observed during batch wet granulation, which exhibited a unique morphology with a large surface area and improved tabletability. A full factorial experimental design was performed, varying screw speed (400-900rpm), granulation temperature (25-40°C), number of kneading elements (6 or 12) and liquid-to-solid (L/S) ratio, on the granulation unit of a ConsiGma™-25 line (a continuous powder-to-tablet manufacturing system). After tray drying the granules were milled and tableted. The results showed that the polymorphic transition from δ- to β-mannitol also occurred during twin screw granulation, although the residence time and L/S ratios were much lower in continuous twin screw granulation compared to batch processing. However, the polymorphic transition was not complete in all experiments and depended on the L/S ratio, screw speed and number of kneading elements. Nevertheless all granules exhibited the unique morphology linked to the polymorphic transition and had a superior tabletability compared to granules produced with β-mannitol as starting material. This was attributed to enhanced plastic deformation of the granules manufactured using δ-mannitol as starting material. In addition, it was concluded that mannitol was granulated via a different mechanism than other, less-soluble, excipients (e.g. lactose, microcrystalline cellulose) due to its high solubility and dissolution rate as the influence of process parameters on the mannitol granule characteristics was different. Copyright © 2016 Elsevier B.V. All rights reserved.

Elucidation of solution state complexation in wet-granulated oven-dried ibuprofen and beta-cyclodextrin: FT-IR and 1H-NMR studies.

PubMed

Ghorab, M K; Adeyeye, M C

2001-08-01

The effect of oven-dried wet granulation on the complexation of beta-cyclodextrin with ibuprofen (IBU) in solution was investigated using Fourier transform infrared spectroscopy (FT-IR), proton nuclear magnetic resonance (1H NMR), and molecular modeling. Granulation was carried out using 5 mL of three different granulating solvents; water, ethanol (95% v/v), and isopropanol and the granules were oven-dried at 60 degrees C for 2 h. The granules were compared to oven-dried physical mixture and conventionally prepared complex. Phase solubility study was performed to investigate the stability of the granulation-formed complexes in solution. FT-IR was used to examine the complexation in the granules while 1H NMR, and molecular modeling studies were carried out to determine the mechanism of complexation in the water-prepared granules. The solubility studies suggested a 1:1 complex between IBU and betaCD. It also showed that the stability of the complex in solution was in the following order with respect to the granulating solvents: ethanol > water > isopropanol. The FT-IR study revealed a shift in the carboxylic acid stretching band and decrease in the intensities of the C-H bending bands of the isopropyl group and the out-of-plane aromatic ring, of IBU, in granules compared to the oven-dried physical mixture. This indicated that granules might have some extent of solid state complexation that could further enhance dissolution and the IBU-betaCD solution state complexation. 1H NMR showed that water prepared oven-dried granules had a different 1H NMR spectrum compared to similarly made oven-dried physical mixture, indicative of complexation in the former. The 1H NMR and the molecular modeling studies together revealed that solution state complexation from the granules occurred by inclusion of the isopropyl group together with part of the aromatic ring of IBU into the betaCD cavity probably through its wider side. These results indicate that granulation process induced faster complexation in solution which enhances the solubility and the dissolution rate of poorly soluble drugs. The extent of complexation in the granules was dependent on the type of solvent used.

Fast formation of aerobic granules by combining strong hydraulic selection pressure with overstressed organic loading rate.

PubMed

Liu, Yong-Qiang; Tay, Joo-Hwa

2015-09-01

The combined strong hydraulic selection pressure (HSP) with overstressed organic loading rate (OLR) as a fast granulation strategy was used to enhance aerobic granulation. To investigate the wide applicability of this strategy to different scenarios and its relevant mechanism, different settling times, different inoculums, different exchange ratios, different reactor configurations, and different shear force were used in this study. It was found that clear granules were formed within 24 h and steady state reached within three days when the fast granulation strategy was used in a lab-scale reactor seeded with well settled activated sludge (Reactor 2). However, granules appeared after 2-week operation and reached steady state after one month at the traditional step-wise decreased settling time from 20 to 2 min with OLR of 6 g COD/L·d (Reactor 1). With the fast granulation strategy, granules appeared within 24 h even with bulking sludge as seed to start up Reactor 3, but 6-day lag phase was observed compared with Reactor 2. Both Reactor 2 and Reactor 3 experienced sigmoidal growth curve in terms of biomass accumulation and granule size increase after granulation. In addition, the reproducible results in pilot-scale reactors (Reactor 5 and Reactor 6) with diameter of 20 cm and height/diameter ratio (H/D) of 4 further proved that reactor configuration and fluid flow pattern had no effect on the aerobic granulation when the fast granulation strategy was employed, but biomass accumulation experienced a short lag phase too in Reactor 5 and Reactor 6. Although overstressed OLR was favorable for fast granulation, it also led to the fluffy granules after around two-week operation. However, the stable 6-month operation of Reactor 3 demonstrated that the rapidly formed granules were able to maintain long-term stability by reducing OLR from 12 g COD/L·d to 6 g COD/L·d. A mechanism of fast granulation with the strategy of combined strong HSP and OLR was proposed to explain results and guide the operation with this fast strategy. Copyright © 2015 Elsevier Ltd. All rights reserved.

Distribution of pancreatic elastase and metalloproteinase in vertebrates.

PubMed

Yoshinaka, R; Sato, M; Tsuchiya, N; Ikeda, S

1986-01-01

Elastase-like enzymes were detected as zymogens in all of the pancreatic extracts from the gummy shark, bullhead shark, angel shark, smooth hammerhead, bestel, rainbow trout, carp, eel, Japanese mackerel, yellowtail, sea bass, parrotfish, bullfrog, chicken, bluewhite dolphin, hog, rat, cat, and dog. The distribution of pancreatic elastase and metalloproteinase was examined on the basis of the effect of specific inhibitors on elastase like-activity in each extract. The results indicate that pancreatic elastases are present in all the species examined and pancreatic metalloproteinases are present only in the teleost fishes.

Fish-borne Zoonotic Trematode Metacercariae in the Republic of Korea

PubMed Central

2009-01-01

The prevalence of fish-borne trematodes (FBT), including Clonorchis sinensis, is still high in riverside areas of the Republic of Korea. The author reviewed the detection and identification methods, differential keys, fish intermediate hosts, and morphological characteristics of FBT metacercariae. FBT metacercariae found in freshwater fish are classified mainly into 4 families, i.e., Opisthorchiidae, Heterophyidae, Echinostomatidae, and Clinostomidae. The metacercariae of C. sinensis, found in 40 species of freshwater fish, are elliptical and 0.15-0.17 × 0.13-0.15 mm in size, have nearly equal sized oral and ventral suckers, brownish pigment granules, and an O-shaped excretory bladder. Their general morphologies are similar to those of Metorchis orientalis (except in the thickness of the cyst wall). Metagonimus spp. (M. yokogawai, M. takahashii, and M. miyatai) metacercariae are subglobular or disc-shaped, and 0.14-0.16 mm in diameter. They have yellow-brownish pigment granules, a ventral sucker deflectively located from median, and a V-shaped excretory bladder. The metacercariae and fish intermediate hosts of Centrocestus armatus, Clinostomum complanatum, and 3 echinostomatid flukes (Echinostoma hortense, E. cinetorchis, and Echinochasmus japonicus) were summarized. FBT metacercariae detected in brackish water fish are mainly members of the Heterophyidae. The morphological characters, identification keys, and fish intermediate hosts of 7 species (Heterophyes nocens, Heterophyopsis continua, Pygidiopsis summa, Stellantchasmus falcatus, Stictodora fuscata, Stictodora lari, and Acanthotrema felis) were also reviewed. The contents treated in this study will provide assistance at the laboratory bench level to those working on recovery of metacercariae from fish hosts and identifying them. PMID:19885326

Modeling of lithium granule injection in NSTX using M3D-C1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fil, A.; Kolemen, E.; Ferraro, N.

In this paper, we present simulations of pedestal control by lithium granule injection (LGI) in NSTX. A model for small granule ablation has been implemented in the M3D-C1 code (Jardin et al 2012 Comput. Sci. Discovery 5 014002), allowing the simulation of realistic lithium granule injections. 2D and 3D simulations of Li injections in NSTX H-mode plasmas are performed and the effect of granule size, injection angle and velocity on the pedestal gradient increase is studied. The amplitude of the local pressure perturbation caused by the granules is found to be highly dependent on the solid granule size. Adjusting themore » granule injection velocity allows one to inject more particles at the pedestal top. 3D simulations show the destabilization of high order MHD modes whose amplitude is directly linked to the localized pressure perturbation, which is found to depend on the toroidal localization of the granule density source.« less

Autophagy meets fused in sarcoma-positive stress granules.

PubMed

Matus, Soledad; Bosco, Daryl A; Hetz, Claudio

2014-12-01

Mutations in fused in sarcoma and/or translocated in liposarcoma (FUS, TLS or FUS) are linked to familial cases of amyotrophic lateral sclerosis (ALS). Mutant FUS selectively accumulates into discrete cytosolic structures known as stress granules under various stress conditions. In addition, mutant FUS expression can alter the dynamics and morphology of stress granules. Although the link between mutant FUS and stress granules is well established, the mechanisms modulating stress granule formation and disassembly in the context of ALS are poorly understood. In this issue of Neurobiology of Aging, Ryu et al. uncover the impact of autophagy on the potential toxicity of mutant FUS-positive stress granules. The authors provide evidence indicating that enhanced autophagy activity reduces the number of stress granules, which in the case of cells containing mutant FUS-positive stress granules, is neuroprotective. Overall, this study identifies an intersection between the proteostasis network and alterations in RNA metabolism in ALS through the dynamic assembly and disassembly of stress granules. Copyright © 2014 Elsevier Inc. All rights reserved.

Internal structure of normal maize starch granules revealed by chemical surface gelatinization.

PubMed

Pan, D D; Jane, J I

2000-01-01

Normal maize starch was fractionated into two sizes: large granules with diameters more than 5 microns and small granules with diameters less than 5 microns. The large granules were surface gelatinized by treating them with an aqueous LiCl solution (13 M) at 22-23 degrees C. Surface-gelatinized remaining granules were obtained by mechanical blending, and gelatinized surface starch was obtained by grinding with a mortar and a pestle. Starches of different granular sizes and radial locations, obtained after different degrees of surface gelatinization, were subjected to scanning electron microscopy, iodine potentiometric titration, gel-permeation chromatography, and amylopectin branch chain length analysis. Results showed that the remaining granules had a rough surface with a lamella structure. Amylose was more concentrated at the periphery than at the core of the granule. Amylopectin had longer long B-chains at the core than at the periphery of the granule. Greater proportions of the long B-chains were present at the core than at the periphery of the granule.

New Class of Cargo Protein in Tetrahymena thermophila Dense Core Secretory Granules

PubMed Central

Haddad, Alex; Bowman, Grant R.; Turkewitz, Aaron P.

2002-01-01

Regulated exocytosis of dense core secretory granules releases biologically active proteins in a stimulus-dependent fashion. The packaging of the cargo within newly forming granules involves a transition: soluble polypeptides condense to form water-insoluble aggregates that constitute the granule cores. Following exocytosis, the cores generally disassemble to diffuse in the cell environment. The ciliates Tetrahymena thermophila and Paramecium tetraurelia have been advanced as genetically manipulatable systems for studying exocytosis via dense core granules. However, all of the known granule proteins in these organisms condense to form the architectural units of lattices that are insoluble both before and after exocytosis. Using an approach designed to detect new granule proteins, we have now identified Igr1p (induced during granule regeneration). By structural criteria, it is unrelated to the previously characterized lattice-forming proteins. It is distinct in that it is capable of dissociating from the insoluble lattice following secretion and therefore represents the first diffusible protein identified in ciliate granules. PMID:12456006

Modeling of lithium granule injection in NSTX using M3D-C1

DOE PAGES

Fil, A.; Kolemen, E.; Ferraro, N.; ...

2017-04-06

In this paper, we present simulations of pedestal control by lithium granule injection (LGI) in NSTX. A model for small granule ablation has been implemented in the M3D-C1 code (Jardin et al 2012 Comput. Sci. Discovery 5 014002), allowing the simulation of realistic lithium granule injections. 2D and 3D simulations of Li injections in NSTX H-mode plasmas are performed and the effect of granule size, injection angle and velocity on the pedestal gradient increase is studied. The amplitude of the local pressure perturbation caused by the granules is found to be highly dependent on the solid granule size. Adjusting themore » granule injection velocity allows one to inject more particles at the pedestal top. 3D simulations show the destabilization of high order MHD modes whose amplitude is directly linked to the localized pressure perturbation, which is found to depend on the toroidal localization of the granule density source.« less

Biomineralisation by earthworms - an investigation into the stability and distribution of amorphous calcium carbonate.

PubMed

Hodson, Mark E; Benning, Liane G; Demarchi, Bea; Penkman, Kirsty E H; Rodriguez-Blanco, Juan D; Schofield, Paul F; Versteegh, Emma A A

Many biominerals form from amorphous calcium carbonate (ACC), but this phase is highly unstable when synthesised in its pure form inorganically. Several species of earthworm secrete calcium carbonate granules which contain highly stable ACC. We analysed the milky fluid from which granules form and solid granules for amino acid (by liquid chromatography) and functional group (by Fourier transform infrared (FTIR) spectroscopy) compositions. Granule elemental composition was determined using inductively coupled plasma-optical emission spectroscopy (ICP-OES) and electron microprobe analysis (EMPA). Mass of ACC present in solid granules was quantified using FTIR and compared to granule elemental and amino acid compositions. Bulk analysis of granules was of powdered bulk material. Spatially resolved analysis was of thin sections of granules using synchrotron-based μ-FTIR and EMPA electron microprobe analysis. The milky fluid from which granules form is amino acid-rich (≤ 136 ± 3 nmol mg -1 (n = 3; ± std dev) per individual amino acid); the CaCO 3 phase present is ACC. Even four years after production, granules contain ACC. No correlation exists between mass of ACC present and granule elemental composition. Granule amino acid concentrations correlate well with ACC content (r ≥ 0.7, p ≤ 0.05) consistent with a role for amino acids (or the proteins they make up) in ACC stabilisation. Intra-granule variation in ACC (RSD = 16%) and amino acid concentration (RSD = 22-35%) was high for granules produced by the same earthworm. Maps of ACC distribution produced using synchrotron-based μ-FTIR mapping of granule thin sections and the relative intensity of the ν 2 : ν 4 peak ratio, cluster analysis and component regression using ACC and calcite standards showed similar spatial distributions of likely ACC-rich and calcite-rich areas. We could not identify organic peaks in the μ-FTIR spectra and thus could not determine whether ACC-rich domains also had relatively high amino acid concentrations. No correlation exists between ACC distribution and elemental concentrations determined by EMPA. ACC present in earthworm CaCO 3 granules is highly stable. Our results suggest a role for amino acids (or proteins) in this stability. We see no evidence for stabilisation of ACC by incorporation of inorganic components. Graphical abstractSynchrotron-based μ-FTIR mapping was used to determine the spatial distribution of amorphous calcium carbonate in earthworm-produced CaCO 3 granules.

Imaging exocytosis of single glucagon-like peptide-1 containing granules in a murine enteroendocrine cell line with total internal reflection fluorescent microscopy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ohara-Imaizumi, Mica; Aoyagi, Kyota; Akimoto, Yoshihiro

To analyze the exocytosis of glucagon-like peptide-1 (GLP-1) granules, we imaged the motion of GLP-1 granules labeled with enhanced yellow fluorescent protein (Venus) fused to human growth hormone (hGH-Venus) in an enteroendocrine cell line, STC-1 cells, by total internal reflection fluorescent (TIRF) microscopy. We found glucose stimulation caused biphasic GLP-1 granule exocytosis: during the first phase, fusion events occurred from two types of granules (previously docked granules and newcomers), and thereafter continuous fusion was observed mostly from newcomers during the second phase. Closely similar to the insulin granule fusion from pancreatic {beta} cells, the regulated biphasic exocytosis from two typesmore » of granules may be a common mechanism in glucose-evoked hormone release from endocrine cells.« less

High organic loading influences the physical characteristics of aerobic sludge granules.

PubMed

Moy, B Y-P; Tay, J-H; Toh, S-K; Liu, Y; Tay, S T-L

2002-01-01

The effect of high organic loading rate (OLR) on the physical characteristics of aerobic granules was studied. Two column-type sequential aerobic sludge blanket reactors were fed with either glucose or acetate as the main carbon source, and the OLR was gradually raised from 6 to 9, 12 and 15 kg chemical oxygen demand (COD) m(-3) d(-1). Glucose-fed granules could sustain the maximum OLR tested. At a low OLR, these granules exhibited a loose fluffy morphology dominated by filamentous bacteria. At higher OLRs, these granules became irregularly shaped, with folds, crevices and depressions. In contrast, acetate-fed granules had a compact spherical morphology at OLRs of 6 and 9 kg COD m(-3) d(-1), with better settling and strength characteristics than glucose-fed granules at similar OLRs. However, acetate-fed granules could not sustain high OLRs and disintegrated when the OLR reached 9 kg COD m(-3) d(-1). The compact regular microstructure of the acetate-fed granules appeared to limit mass transfer of nutrients at an OLR of 9 kg COD m(-3) d(-1). The looser filamentous microstructure of the glucose-fed granules and the subsequent irregular morphology delayed the onset of diffusion limitation and allowed significantly higher OLRs to be attained. SIGNIFICNACE AND IMPACT OF THE STUDY: High organic loading rates are possible with aerobic granules. This research would be helpful in the development of aerobic granule-based systems for high-strength wastewaters.

Granulation of snow: From tumbler experiments to discrete element simulations

NASA Astrophysics Data System (ADS)

Steinkogler, Walter; Gaume, Johan; Löwe, Henning; Sovilla, Betty; Lehning, Michael

2015-06-01

It is well known that snow avalanches exhibit granulation phenomena, i.e., the formation of large and apparently stable snow granules during the flow. The size distribution of the granules has an influence on flow behavior which, in turn, affects runout distances and avalanche velocities. The underlying mechanisms of granule formation are notoriously difficult to investigate within large-scale field experiments, due to limitations in the scope for measuring temperatures, velocities, and size distributions. To address this issue we present experiments with a concrete tumbler, which provide an appropriate means to investigate granule formation of snow. In a set of experiments at constant rotation velocity with varying temperatures and water content, we demonstrate that temperature has a major impact on the formation of granules. The experiments showed that granules only formed when the snow temperature exceeded -1∘C. No evolution in the granule size was observed at colder temperatures. Depending on the conditions, different granulation regimes are obtained, which are qualitatively classified according to their persistence and size distribution. The potential of granulation of snow in a tumbler is further demonstrated by showing that generic features of the experiments can be reproduced by cohesive discrete element simulations. The proposed discrete element model mimics the competition between cohesive forces, which promote aggregation, and impact forces, which induce fragmentation, and supports the interpretation of the granule regime classification obtained from the tumbler experiments. Generalizations, implications for flow dynamics, and experimental and model limitations as well as suggestions for future work are discussed.

Statistical properties of solar granulation from the SOUP instrument on Spacelab 2

NASA Astrophysics Data System (ADS)

Topka, K.; Title, A.; Tarbell, T.; Ferguson, S.; Shine, R.

1988-11-01

The Solar Optical Universal Polarimeter (SOUP) on Spacelab 2 collected movies of solar granulation completely free from atmospheric blurring, and are not degraded by pointint jitter (the pointing stability was 0.003 sec root mean square). The movies illustrate that the solar five minute oscillation has a major role in the appearance of solar granulation and that exploding granules are a common feature of the granule evolution. Using 3-D Fourier filtering techniques the oscillations were removed and it was demonstrated that the autocorrelation lifetime of granulation is a factor of two greater in magnetic field regions than in field-free quiet sun. Horizontal velocities were measured and flow patterns were observed on the scale of meso- and super granulation. In quiet regions the mean flow velocity is 370 m/s while in the magnetic regions it is about 125 m/s. It was also found that the root mean square (RMS) fluctuating horizonal velocity field is substantially greater in quiet sun than in strong magnetic field regions. By superimposing the location of exploding granules on the average flow maps it was found that they appear almost exclusively in the center of mesogranulation size flow cells. Because of the nonuniformity of the distribution of exploding granules, the evolution of the granulation pattern in mesogranule cell centers and boundaries differs fundamentally. It is clear from this study there is neither a typical granule nor a typical granule evolution.

Statistical properties of solar granulation from the SOUP instrument on Spacelab 2

NASA Technical Reports Server (NTRS)

Topka, K.; Title, A.; Tarbell, T.; Ferguson, S.; Shine, R.

1988-01-01

The Solar Optical Universal Polarimeter (SOUP) on Spacelab 2 collected movies of solar granulation completely free from atmospheric blurring, and are not degraded by pointint jitter (the pointing stability was 0.003 sec root mean square). The movies illustrate that the solar five minute oscillation has a major role in the appearance of solar granulation and that exploding granules are a common feature of the granule evolution. Using 3-D Fourier filtering techniques the oscillations were removed and it was demonstrated that the autocorrelation lifetime of granulation is a factor of two greater in magnetic field regions than in field-free quiet sun. Horizontal velocities were measured and flow patterns were observed on the scale of meso- and super granulation. In quiet regions the mean flow velocity is 370 m/s while in the magnetic regions it is about 125 m/s. It was also found that the root mean square (RMS) fluctuating horizonal velocity field is substantially greater in quiet sun than in strong magnetic field regions. By superimposing the location of exploding granules on the average flow maps it was found that they appear almost exclusively in the center of mesogranulation size flow cells. Because of the nonuniformity of the distribution of exploding granules, the evolution of the granulation pattern in mesogranule cell centers and boundaries differs fundamentally. It is clear from this study there is neither a typical granule nor a typical granule evolution.

Investigation on the pitting of potato starch granules during high frequency ultrasound treatment.

PubMed

Bai, Wenzhe; Hébraud, Pascal; Ashokkumar, Muthupandian; Hemar, Yacine

2017-03-01

In this paper, the pitting of potato starch granules in aqueous suspensions (1%) by high-frequency high-power ultrasound (850kHz at a power of 0.2W, 2W or 3.7W; and also 500kHz and 1MHz at a power of 2W) is reported. The number of pits per starch granules was found to be independent of the amylose content of starches, and the surface properties of starch granules as modified through SDS and ethanol washing. At 850kHz, the maximum number of pits per starch granule, for both normal and waxy starches, did not exceed 11. However, a close inspection of fractionated starch granules based on their sizes showed that there is an optimum granule size for which a maximum pit number is obtained. For example, starch granules with diameter size range of ∼15 to ∼30μm had a maximum pit number (between 10 and 20 pits per granule) when sonicated (2W, 850kHz and 30min); while sonication of small (<10μm) and very large (>45μm) granules resulted in a smaller number of pits per granule (∼5). Further, the maximum number of pits per granules is also found to be proportional to the ultrasound frequency, with values of approximately 7, 10 and 11 at 0.50, 0.85, and 1MHz, respectively. FTIR measurements did not show any breakup of starch molecules. Copyright © 2016 Elsevier B.V. All rights reserved.

In vitro and in vivo evaluation of medicinal carbon granules and tablet on the adsorption of acetaminophen.

PubMed

Yamamoto, Kenta; Onishi, Hiraku; Ito, Akihiko; Machida, Yoshiharu

2007-01-10

Medicinal carbon (MC) granules were prepared by wet granulation using maltitol (MT), and the MC tablet was produced by compression of the granules. The physical properties and the in vitro adsorption capacity for AA of the formulations were examined. Further, the effects of MC alone and the granules on gastrointestinal absorption of AA were examined in rats when they were administered intragastrically at 15 or 45 min after the intragastrical administration of AA. AA was rapidly adsorbed by MC, and the maximum adsorption capacity of MC was 0.329g AA per gram MC. The granules and tablet exhibited adequate strength, and the tablet disintegrated rapidly. The granules and tablet showed similar adsorption profiles, but somewhat lower adsorption capacity than MC alone. MC alone and granules administered at 15 min reduced the AUC(0-infinity) significantly against the control (no treatment); however, the suppression effect on the plasma concentration was lower with the granules than with MC alone. Thus, granules and tablet are useful as a compact dosage form of MC; though the reduced adsorption capacity must be taken into account in order to expect efficacy equivalent to that of MC alone.

Investigation of the effect of impeller speed on granules formed using a PMA-1 high shear granulator.

PubMed

Logan, R; Briens, L

2012-11-01

Impeller speed was varied from 300 to 1500 rpm during the wet high shear granulation of a placebo formulation using a new vertical shaft PharmaMATRIX-1 granulator. The resulting granules were extensively analysed for differences caused by the varying impeller speed with emphasis on flowability. Microscopy showed that initial granules were formed primarily from microcrystalline cellulose at all tested impeller speeds. At low impeller speed of 300 rpm in the "bumpy" flow regime, forces from the impeller were insufficient to incorporate all the components of the formulation into the granules and to promote granule growth to a size that significantly improved flowability. The "roping" flow regime at higher impeller speeds promoted granule growth to a median particle size of at least 100 µm that improved the flowability of the mixture. Particle size distribution measurements and advanced indicators based on avalanching behavior, however, showed that an impeller speed of 700 rpm produced the largest fraction of optimal granules with the best flowability potential. This impeller speed allowed good development of "roping" flow for sufficient mixing, collision rates and kinetic energy for collisions while minimizing excessive centrifugal forces that promote buildup around the bowl perimeter.

Starch synthase 4 is essential for coordination of starch granule formation with chloroplast division during Arabidopsis leaf expansion

PubMed Central

Crumpton-Taylor, Matilda; Pike, Marilyn; Lu, Kuan-Jen; Hylton, Christopher M; Feil, Regina; Eicke, Simona; Lunn, John E; Zeeman, Samuel C; Smith, Alison M

2013-01-01

Arabidopsis thaliana mutants lacking the SS4 isoform of starch synthase have strongly reduced numbers of starch granules per chloroplast, suggesting that SS4 is necessary for the normal generation of starch granules. To establish whether it plays a direct role in this process, we investigated the circumstances in which granules are formed in ss4 mutants. Starch granule numbers and distribution and the accumulation of starch synthase substrates and products were investigated during ss4 leaf development, and in ss4 mutants carrying mutations or transgenes that affect starch turnover or chloroplast volume. We found that immature ss4 leaves have no starch granules, but accumulate high concentrations of the starch synthase substrate ADPglucose. Granule numbers are partially restored by elevating the capacity for glucan synthesis (via expression of bacterial glycogen synthase) or by increasing the volumes of individual chloroplasts (via introduction of arc mutations). However, these granules are abnormal in distribution, size and shape. SS4 is an essential component of a mechanism that coordinates granule formation with chloroplast division during leaf expansion and determines the abundance and the flattened, discoid shape of leaf starch granules. PMID:23952675

Comparison of low-shear and high-shear granulation processes: effect on implantable calcium phosphate granule properties.

PubMed

Chevalier, E; Viana, M; Cazalbou, S; Chulia, D

2009-10-01

Calcium phosphate porous ceramics present a great interest not only as complex bone defect fillers but also as drug delivery systems. Most of the methods described in the literature to fabricate pellets are based on compaction, casting into spherical molds, or on processes such as liquid immiscibility or foaming. Despite wet granulation is used in a wide range of applications in pharmaceuticals, food, detergents, fertilizers, and minerals, it is not applied in the biomaterial field to produce granules. In this study physicochemical and in vitro drug delivery properties of implantable calcium phosphate granules, produced by two wet agglomeration processes, were compared. Pellets obtained by high shear granulation (granulation in a Mi-Pro apparatus) were shown to be more spherical and less friable than granules elaborated by low shear process (granulation in a Kenwood apparatus). Although Mi-Pro pellets had a slightly lower porosity compared to Kenwood granules, ibuprofen loading efficiency and dissolution profiles were not statistically different and the release mechanism was mainly controlled by diffusion, in both cases. Mi-Pro pellets appeared to be better candidates as bone defect fillers and local drug delivery systems as far as they were more spherical and less friable than Kenwood agglomerates.

Granule size control and targeting in pulsed spray fluid bed granulation.

PubMed

Ehlers, Henrik; Liu, Anchang; Räikkönen, Heikki; Hatara, Juha; Antikainen, Osmo; Airaksinen, Sari; Heinämäki, Jyrki; Lou, Honxiang; Yliruusi, Jouko

2009-07-30

The primary aim of the study was to investigate the effects of pulsed liquid feed on granule size. The secondary aim was to increase knowledge of this technique in granule size targeting. Pulsed liquid feed refers to the pump changing between on- and off-positions in sequences, called duty cycles. One duty cycle consists of one on- and off-period. The study was performed with a laboratory-scale top-spray fluid bed granulator with duty cycle length and atomization pressure as studied variables. The liquid feed rate, amount and inlet air temperature were constant. The granules were small, indicating that the powder has only undergone ordered mixing, nucleation and early growth. The effect of atomizing pressure on granule size depends on inlet air relative humidity, with premature binder evaporation as a reason. The duty cycle length was of critical importance to the end product attributes, by defining the extent of intermittent drying and rewetting. By varying only the duty cycle length, it was possible to control granule nucleation and growth, with a wider granule size target range in increased relative humidity. The present study confirms that pulsed liquid feed in fluid bed granulation is a useful tool in end product particle size targeting.

Stimulation of microtubule-based transport by nucleation of microtubules on pigment granules

PubMed Central

Semenova, Irina; Gupta, Dipika; Usui, Takeo; Hayakawa, Ichiro; Cowan, Ann; Rodionov, Vladimir

2017-01-01

Microtubule (MT)-based transport can be regulated through changes in organization of MT transport tracks, but the mechanisms that regulate these changes are poorly understood. In Xenopus melanophores, aggregation of pigment granules in the cell center involves their capture by the tips of MTs growing toward the cell periphery, and granule aggregation signals facilitate capture by increasing the number of growing MT tips. This increase could be explained by stimulation of MT nucleation either on the centrosome or on the aggregate of pigment granules that gradually forms in the cell center. We blocked movement of pigment granules to the cell center and compared the MT-nucleation activity of the centrosome in the same cells in two signaling states. We found that granule aggregation signals did not stimulate MT nucleation on the centrosome but did increase MT nucleation activity of pigment granules. Elevation of MT-nucleation activity correlated with the recruitment to pigment granules of a major component of MT-nucleation templates, γ-tubulin, and was suppressed by γ-tubulin inhibitors. We conclude that generation of new MT transport tracks by concentration of the leading pigment granules provides a positive feedback loop that enhances delivery of trailing granules to the cell center. PMID:28381426

To Evaluate the Effect of Solvents and Different Relative Humidity Conditions on Thermal and Rheological Properties of Microcrystalline Cellulose 101 Using METHOCEL™ E15LV as a Binder.

PubMed

Jagia, Moksh; Trivedi, Maitri; Dave, Rutesh H

2016-08-01

The solvent used for preparing the binder solution in wet granulation can affect the granulation end point and also impact the thermal, rheological, and flow properties of the granules. The present study investigates the effect of solvents and percentage relative humidity (RH) on the granules of microcrystalline cellulose (MCC) with hydroxypropyl methyl cellulose (HPMC) as the binder. MCC was granulated using 2.5% w/w binder solution in water and ethanol/water mixture (80:20 v/v). Prepared granules were dried until constant percentage loss on drying, sieved, and further analyzed. Dried granules were exposed to different percentage RH for 48 h at room temperature. Powder rheometer was used for the rheological and flow characterization, while thermal effusivity and differential scanning calorimeter were used for thermal analysis. The thermal effusivity values for the wet granules showed a sharp increase beginning 50% w/w binder solution in both cases, which reflected the over-wetting of granules. Ethanol/water solvent batches showed greater resistance to flow as compared to the water solvent batches in the wet granule stage, while the reverse was true for the dried granule stage, as evident from the basic flowability energy values. Although the solvents used affected the equilibration kinetics of moisture content, the RH-exposed granules remained unaffected in their flow properties in both cases. This study indicates that the solvents play a vital role on the rheology and flow properties of MCC granules, while the different RH conditions have little or no effect on them for the above combination of solvent and binder.

CERES Search and Subset Tool

Atmospheric Science Data Center

2016-06-24

... data granules using a high resolution spatial metadata database and directly accessing the archived data granules. Subset results are ... data granules using a high resolution spatial metadata database and directly accessing the archived data granules. Subset results are ...

Whi3, an S. cerevisiae RNA-binding protein, is a component of stress granules that regulates levels of its target mRNAs.

PubMed

Holmes, Kristen J; Klass, Daniel M; Guiney, Evan L; Cyert, Martha S

2013-01-01

RNA binding proteins (RBPs) are vital to the regulation of mRNA transcripts, and can alter mRNA localization, degradation, translation, and storage. Whi3 was originally identified in a screen for small cell size mutants, and has since been characterized as an RBP. The identification of Whi3-interacting mRNAs involved in mediating cellular responses to stress suggested that Whi3 might be involved in stress-responsive RNA processing. We show that Whi3 localizes to stress granules in response to glucose deprivation or heat shock. The kinetics and pattern of Whi3 localization in response to a range of temperatures were subtly but distinctly different from those of known components of RNA processing granules. Deletion of Whi3 resulted in an increase in the relative abundance of Whi3 target RNAs, either in the presence or absence of heat shock. Increased levels of the CLN3 mRNA in whi3Δ cells may explain their decreased cell size. Another mRNA target of Whi3 encodes the zinc-responsive transcription factor Zap1, suggesting a role for Whi3 in response to zinc stress. Indeed, we found that whi3Δ cells have enhanced sensitivity to zinc toxicity. Together our results suggest an expanded model for Whi3 function: in addition to its role as a regulator of the cell cycle, Whi3 may have a role in stress-dependent RNA processing and responses to a variety of stress conditions.

Injected mass deposition thresholds for lithium granule instigated triggering of edge localized modes on EAST

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lunsford, R.; Sun, Zhen; Maingi, Rajesh

The ability of an injected lithium granule to promptly trigger an edge localized mode (ELM) has been established in multiple experiments. By horizontally injecting granules ranging in diameter from 200 microns to 1mm in diameter into the low field side of EAST H-mode discharges we have determined that granules with diameter > 600 microns are successful in triggering ELMs more than 95% of the time. Granules were radially injected from the outer midplane with velocities ~ 80 m/s into EAST upper-single null discharges with an ITER like tungsten monoblock divertor. ELM triggering was a prompt response to granule injection, andmore » for granules of a sufficient size there was no evidence of a "trigger lag" phenomenon as observed in full metal machines. We also demonstrated that the triggering efficiency decreased with granule size during dynamic size scans. These granules were individually tracked throughout their injection cycle in order to determine their efficacy at triggering an ELM. Furthermore, by simulating the granule injection with an experimentally benchmarked neutral gas shielding (NGS) model, the ablatant mass deposition required to promptly trigger an ELM is calculated and the fractional mass deposition is determined. Simulated 900 micron granules capable of triggering an ELM show a peaked mass deposition of 3.9 x 10 17 atoms per mm of penetration at a depth of approximately 5 cm past the separatrix.« less

The importance of binder moisture content in Metformin HCL high-dose formulations prepared by moist aqueous granulation (MAG).

PubMed

Takasaki, Hiroshi; Yonemochi, Etsuo; Ito, Masanori; Wada, Koichi; Terada, Katsuhide

2015-01-01

The aim of this study was to evaluate binders to improve the flowability of granulates and compactibility of Metformin HCL (Met) using the moist aqueous granulation (MAG) process. The effect of the binder moisture content on granulate and tablet quality was also evaluated. Vinylpyrrolidone-vinyl acetate copolymer (Kollidon VA64 fine: VA64), polyvidone (Povidone K12: PVP), hydroxypropyl cellulose (HPC SSL SF: HPC) and hydroxypropyl methylcellulose (Methocel E5 LV: HPMC) were evaluated as binders. These granulates, except for HPMC, had a lower yield pressure than Met active pharmaceutical ingredient (API). HPMC Met was not sufficiently granulated with low water volume. No problems were observed with the VA64 Met granulates during the tableting process. However, HPC Met granulates had a bowl-forming tendency, and PVP Met granulates had the tendency to stick during the tableting process. These bowl-forming and sticking tendencies may have been due to the low moisture absorbency of HPC and the high volume of bound water of PVP, respectively. VA64 Met granulates had the highest ambient moisture content (bulk water, bound water) and moisture absorbency. It was concluded that the type of binder used for the Met MAG process has an impact on granulate flow and compactibility, as well as moisture absorbency and maintenance of moisture balance.

The importance of binder moisture content in Metformin HCL high-dose formulations prepared by moist aqueous granulation (MAG)

PubMed Central

Takasaki, Hiroshi; Yonemochi, Etsuo; Ito, Masanori; Wada, Koichi; Terada, Katsuhide

2015-01-01

The aim of this study was to evaluate binders to improve the flowability of granulates and compactibility of Metformin HCL (Met) using the moist aqueous granulation (MAG) process. The effect of the binder moisture content on granulate and tablet quality was also evaluated. Vinylpyrrolidone–vinyl acetate copolymer (Kollidon VA64 fine: VA64), polyvidone (Povidone K12: PVP), hydroxypropyl cellulose (HPC SSL SF: HPC) and hydroxypropyl methylcellulose (Methocel E5 LV: HPMC) were evaluated as binders. These granulates, except for HPMC, had a lower yield pressure than Met active pharmaceutical ingredient (API). HPMC Met was not sufficiently granulated with low water volume. No problems were observed with the VA64 Met granulates during the tableting process. However, HPC Met granulates had a bowl-forming tendency, and PVP Met granulates had the tendency to stick during the tableting process. These bowl-forming and sticking tendencies may have been due to the low moisture absorbency of HPC and the high volume of bound water of PVP, respectively. VA64 Met granulates had the highest ambient moisture content (bulk water, bound water) and moisture absorbency. It was concluded that the type of binder used for the Met MAG process has an impact on granulate flow and compactibility, as well as moisture absorbency and maintenance of moisture balance. PMID:26779418

Impact of functionalized particle structure on roll compaction/dry granulation and tableting of calcium carbonate.

PubMed

Grote, Simon; Kleinebudde, Peter

2018-06-10

The influence of a functionalized raw material particle structure on the granulation behavior and tabletabilty of calcium carbonate (CaCO 3 ) was investigated. Therefore, a milled grade of CaCO 3 was compared to different binary mixtures of milled and functionalized CaCO 3 . Relevant properties of raw materials, ribbons and granules were measured. The starting materials and two fractions of dry granules were compressed to tablets. The tabletability of granules was compared to that of the powders and the influence of specific compaction force and granule size on tablet tensile strength was evaluated. Adding functionalized particles drastically influenced the granulation and tableting behavior of CaCO 3 . Increasing proportions increased the ribbon porosity and granule size. Tensile strength of tablets from powder mixtures and granules was increased as well. Nevertheless, adding functionalized CaCO 3 led to a loss in tabletability induced by a previous compaction step to an extent depending on its proportion in the formulation. A clear influence of the particle morphology on granulation and tableting behavior was demonstrated by the study. The functionalized structure showed aspects of a more plastic deformation behavior. Adding functionalized CaCO 3 to a mixture, even in small amounts, seemed to be beneficial to increase granule size and tablet strength. Copyright © 2018 Elsevier B.V. All rights reserved.

Injected mass deposition thresholds for lithium granule instigated triggering of edge localized modes on EAST

DOE PAGES

Lunsford, R.; Sun, Zhen; Maingi, Rajesh; ...

2017-12-19

The ability of an injected lithium granule to promptly trigger an edge localized mode (ELM) has been established in multiple experiments. By horizontally injecting granules ranging in diameter from 200 microns to 1mm in diameter into the low field side of EAST H-mode discharges we have determined that granules with diameter > 600 microns are successful in triggering ELMs more than 95% of the time. Granules were radially injected from the outer midplane with velocities ~ 80 m/s into EAST upper-single null discharges with an ITER like tungsten monoblock divertor. ELM triggering was a prompt response to granule injection, andmore » for granules of a sufficient size there was no evidence of a "trigger lag" phenomenon as observed in full metal machines. We also demonstrated that the triggering efficiency decreased with granule size during dynamic size scans. These granules were individually tracked throughout their injection cycle in order to determine their efficacy at triggering an ELM. Furthermore, by simulating the granule injection with an experimentally benchmarked neutral gas shielding (NGS) model, the ablatant mass deposition required to promptly trigger an ELM is calculated and the fractional mass deposition is determined. Simulated 900 micron granules capable of triggering an ELM show a peaked mass deposition of 3.9 x 10 17 atoms per mm of penetration at a depth of approximately 5 cm past the separatrix.« less

The influence of granulation on super disintegrant performance.

PubMed

Zhao, Na; Augsburger, Larry L

2006-02-01

The purpose of this study is to identify the causes of efficiency loss of super disintegrants following granulation or reworking. Two processes, precompression and prewetting, were proposed to simulate the processes during dry and wet granulation, respectively. The disintegration efficiency of the resulting disintegrant granules was tested in model formulations composed of dicalcium phosphate and lactose with the unprocessed disintegrants as controls. No significant difference was shown in the intrinsic swelling and the water uptake abilities of all super disintegrants following dry granulation. However, a significant decrease was observed for both Primojel and Polyplasdone XL10 in the rate of water being absorbed into the tablet matrix following wet granulation, but not for Ac-Di-Sol. United States Pharmacopeia (USP) disintegration testing without disc revealed a significant increase in disintegration time for tablets formulated with dry granulated Primojel and Polyplasdone XL10 and all wet granulated disintegrants. The increase in particle size following granulation appears to be the cause of the loss in disintegration efficiency. In conclusion, Ac-Di-Sol is less affected by both precompression and prewetting. The efficiency of Primojel and Polyplasdone XL10 is highly dependent on their particle size. Descreasing the particle size tends to increase their efficiency. Due to the size increase following granulation, a higher addition level of super disintegrant is required to ensure fast and uniform disintegration of tablets prepared by granulation.

Aspergillus oryzae AoSO is a novel component of stress granules upon heat stress in filamentous fungi.

PubMed

Huang, Hsiang-Ting; Maruyama, Jun-ichi; Kitamoto, Katsuhiko

2013-01-01

Stress granules are a type of cytoplasmic messenger ribonucleoprotein (mRNP) granule formed in response to the inhibition of translation initiation, which typically occurs when cells are exposed to stress. Stress granules are conserved in eukaryotes; however, in filamentous fungi, including Aspergillus oryzae, stress granules have not yet been defined. For this reason, here we investigated the formation and localization of stress granules in A. oryzae cells exposed to various stresses using an EGFP fusion protein of AoPab1, a homolog of Saccharomyces cerevisiae Pab1p, as a stress granule marker. Localization analysis showed that AoPab1 was evenly distributed throughout the cytoplasm under normal growth conditions, and accumulated as cytoplasmic foci mainly at the hyphal tip in response to stress. AoSO, a homolog of Neurospora crassa SO, which is necessary for hyphal fusion, colocalized with stress granules in cells exposed to heat stress. The formation of cytoplasmic foci of AoSO was blocked by treatment with cycloheximide, a known inhibitor of stress granule formation. Deletion of the Aoso gene had effects on the formation and localization of stress granules in response to heat stress. Our results suggest that AoSO is a novel component of stress granules specific to filamentous fungi. The authors would specially like to thank Hiroyuki Nakano and Kei Saeki for generously providing experimental and insightful opinions.

Behavior of hollow balls containing granules bouncing repeatedly off the ground

NASA Astrophysics Data System (ADS)

Hu, Min; Mu, Qing-song; Luo, Ning; Li, Gang; Peng, Ning-bo

2013-07-01

An experimental study of the behavior of hollow balls filled with some granules (mung beans or millets) bouncing repeatedly off a static flat horizontal surface is presented. We observed that the bounce number of the ball is limited and decreases regularly with an increasing number of granules. Moreover, for two balls containing a different kind of granules, their bounce numbers are basically equal when the two balls have the same mass of granules. While there is no clear relationship between the first rebound height of one ball and the number of granules, there appears an exponential decay of the second rebound height with an increase of the granule number. Furthermore, a two-dimensional numerical model has been created to find out the law of the ball's rebound height and the dissipation law of the granule nested system. A generalized prediction equation to reasonably explain the law of the bounce number has also been proposed.

An investigation of the influence of process and formulation variables on mechanical properties of high shear granules using design of experiment.

PubMed

Mangwandi, Chirangano; Adams, Michael J; Hounslow, Michael J; Salman, Agba D

2012-05-10

Being able to predict the properties of granules from the knowledge of the process and formulation variables is what most industries are striving for. This research uses experimental design to investigate the effect of process variables and formulation variables on mechanical properties of pharmaceutical granules manufactured from a classical blend of lactose and starch using hydroxypropyl cellulose (HPC) as the binder. The process parameters investigated were granulation time and impeller speed whilst the formulation variables were starch-to-lactose ratio and HPC concentration. The granule properties investigated include granule packing coefficient and granule strength. The effect of some components of the formulation on mechanical properties would also depend on the process variables used in granulation process. This implies that by subjecting the same formulation to different process conditions results in products with different properties. Copyright © 2012 Elsevier B.V. All rights reserved.

Distribution of binder in granules produced by means of twin screw granulation.

PubMed

Fonteyne, Margot; Fussell, Andrew Luke; Vercruysse, Jurgen; Vervaet, Chris; Remon, Jean Paul; Strachan, Clare; Rades, Thomas; De Beer, Thomas

2014-02-28

According to the quality by design principle processes may not remain black-boxes and full process understanding is required. The granule size distribution of granules produced via twin screw granulation is often found to be bimodal. The aim of this study was to gain a better understanding of binder distribution within granules produced via twin screw granulation in order to investigate if an inhomogeneous spread of binder is causing this bimodal size distribution. Theophylline-lactose-polyvinylpyrrolidone K30 (PVP) (30-67.5-2.5%, w/w) was used as a model formulation. The intra-granular distribution of PVP was evaluated by means of hyperspectral coherent anti-Stokes Raman scattering (CARS) microscopy. For the evaluated formulation, no PVP rich zones were detected when applying a lateral spatial resolution of 0.5 μm, indicating that PVP is homogenously distributed within the granules. Copyright © 2013 Elsevier B.V. All rights reserved.

Neuronal RNA granules: a link between RNA localization and stimulation-dependent translation

NASA Technical Reports Server (NTRS)

Krichevsky, A. M.; Kosik, K. S.

2001-01-01

RNA granules are a macromolecular structure observed in neurons, where they serve as motile units that translocate mRNAs. Isolated RNA granules are highly enriched in Staufen protein and ultrastructurally contain densely packed clusters of ribosomes. With depolarization, many mRNAs, including those involved in plasticity, rapidly shift from the RNA granule fraction to polysomes. Depolarization reorganizes granules and induces a less compact organization of their ribosomes. RNA granules are not translationally competent, as indicated by the failure to incorporate radioactive amino acids and the absence of eIF4E, 4G, and tRNAs. We concluded that RNA granules are a local storage compartment for mRNAs under translational arrest but are poised for release to actively translated pools. Local release of mRNAs and ribosomes from granules may serve as a macromolecular mechanism linking RNA localization to translation and synaptic plasticity.

The converse magnetoelectric coupling in asymmetric granule/matrix composite film with Ni/PZT component

NASA Astrophysics Data System (ADS)

Chen, Bo; Su, Ning-Ning; Cui, Wen-Li; Yan, Shi-Nong

2018-04-01

In this work, a type of asymmetric granule/matrix composite film is designed, where the Ni granule is dispersed in PZT matrix, meanwhile the top and bottom electrode is constituted by Au and SRO respectively. Predicted through the electrostatic screening model and mean field approximation, considerable electrostatic charge is induced on Ni granule surface by ferroelectric PZT polarization. Predicted through the spin splitting model and spherical shell approximation, both the magnetization and magnetic anisotropy of Ni granule are modulated by ferroelectric PZT polarization. As the volume fraction of Ni granule is increased, the electric modulation of magnetization and magnetic anisotropy is reduced and enhanced respectively. As the dimension of granule/matrix composite is varied, such modulation is retained. Due to the large area-volume ratio of nano-granule, this work benefits to realize the converse magnetoelectric coupling in nanoscale.

Application of tumbling melt granulation (TMG) method to prepare controlled-release fine granules.

PubMed

Maejima, T; Kubo, M; Osawa, T; Nakajima, K; Kobayashi, M

1998-03-01

The tumbling melt granulation (TMG) method was applied to prepare controlled-release fine granules of diltiazem hydrochloride (DH). The entire process, from the preparation of the cores by the adherence of DH to the sucrose crystal to the subsequent coating of the controlled-release layer, was performed without using any solvent. A mixture of meltable material, talc, and ethylcellulose was used for the controlled-release layer and controlled-release fine granules approximately 400 microns in diameter were obtained with excellent producibility. The dissolution rate of DH from these fine granules was similar to that of a once-a-day dosage form obtained in the market; further, the dependency of the dissolution profile on pH of the media was less. Thus, it was concluded that this TMG method was very useful for preparing not only controlled-release beads of granule size (usually 500 to 1400 microns) but also fine granules.

Clarithromycin highly-loaded gastro-floating fine granules prepared by high-shear melt granulation can enhance the efficacy of Helicobacter pylori eradication.

PubMed

Aoki, Hajime; Iwao, Yasunori; Mizoguchi, Midori; Noguchi, Shuji; Itai, Shigeru

2015-05-01

In an effort to develop a new gastro-retentive drug delivery system (GRDDS) without a large amount of additives, 75% clarithromycin (CAM) loaded fine granules were prepared with three different hydrophobic binders by high-shear melt granulation and their properties were evaluated. Granules containing the higher hydrophobic binder showed sustained drug release and were able to float over 24h. The synchrotron X-ray CT measurement indicated that both the high hydrophobicity of the binder and the void space inside the granules might be involved in their buoyancy. In an in vivo experiment, the floating granules more effectively eradicated Helicobacter pylori than a CAM suspension by remaining in the stomach for a longer period. In short, CAM highly-loaded gastro-floating fine granules can enhance the eradication efficiency of H. pylori compared with CAM alone. Copyright © 2015 Elsevier B.V. All rights reserved.

Determining the Influence of Granule Size on Simulation Parameters and Residual Shear Stress Distribution in Tablets by Combining the Finite Element Method into the Design of Experiments.

PubMed

Hayashi, Yoshihiro; Kosugi, Atsushi; Miura, Takahiro; Takayama, Kozo; Onuki, Yoshinori

2018-01-01

The influence of granule size on simulation parameters and residual shear stress in tablets was determined by combining the finite element method (FEM) into the design of experiments (DoE). Lactose granules were prepared using a wet granulation method with a high-shear mixer and sorted into small and large granules using sieves. To simulate the tableting process using the FEM, parameters simulating each granule were optimized using a DoE and a response surface method (RSM). The compaction behavior of each granule simulated by FEM was in reasonable agreement with the experimental findings. Higher coefficients of friction between powder and die/punch (μ) and lower by internal friction angle (α y ) were generated in the case of small granules, respectively. RSM revealed that die wall force was affected by α y . On the other hand, the pressure transmissibility rate of punches value was affected not only by the α y value, but also by μ. The FEM revealed that the residual shear stress was greater for small granules than for large granules. These results suggest that the inner structure of a tablet comprising small granules was less homogeneous than that comprising large granules. To evaluate the contribution of the simulation parameters to residual stress, these parameters were assigned to the fractional factorial design and an ANOVA was applied. The result indicated that μ was the critical factor influencing residual shear stress. This study demonstrates the importance of combining simulation and statistical analysis to gain a deeper understanding of the tableting process.

Impact of screw configuration on the particle size distribution of granules produced by twin screw granulation.

PubMed

Vercruysse, J; Burggraeve, A; Fonteyne, M; Cappuyns, P; Delaet, U; Van Assche, I; De Beer, T; Remon, J P; Vervaet, C

2015-02-01

Twin screw granulation (TSG) has been reported by different research groups as an attractive technology for continuous wet granulation. However, in contrast to fluidized bed granulation, granules produced via this technique typically have a wide and multimodal particle size distribution (PSD), resulting in suboptimal flow properties. The aim of the current study was to evaluate the impact of granulator screw configuration on the PSD of granules produced by TSG. Experiments were performed using a 25 mm co-rotating twin screw granulator, being part of the ConsiGma™-25 system (a fully continuous from-powder-to-tablet manufacturing line from GEA Pharma Systems). Besides the screw elements conventionally used for TSG (conveying and kneading elements), alternative designs of screw elements (tooth-mixing-elements (TME), screw mixing elements (SME) and cutters) were investigated using an α-lactose monohydrate formulation granulated with distilled water. Granulation with only conveying elements resulted in wide and multimodal PSD. Using kneading elements, the width of the PSD could be partially narrowed and the liquid distribution was more homogeneous. However, still a significant fraction of oversized agglomerates was obtained. Implementing additional kneading elements or cutters in the final section of the screw configuration was not beneficial. Furthermore, granulation with only TME or SME had limited impact on the width of the PSD. Promising results were obtained by combining kneading elements with SME, as for these configurations the PSD was narrower and shifted to the size fractions suitable for tableting. Copyright © 2015 Elsevier B.V. All rights reserved.

Differential compaction behaviour of roller compacted granules of clopidogrel bisulphate polymorphs.

PubMed

Khomane, Kailas S; Bansal, Arvind K

2014-09-10

In the present work, in-die and out-of-die compaction behaviour of dry-granulated powders of clopidogrel bisulphate (CLP) polymorphs, form I and form II, was investigated using a fully instrumented rotary tablet press. Each polymorph was compacted at three different roller pressures [70.3 (S1), 105.5 (S2) and 140.6 (S3)kgf/cm(2)], and obtained granules were characterized for their physico-mechanical properties. Compaction data were analyzed for out-of-die compressibility, tabletability and compactibility profiles, and in-die Heckel, Kawakita and Walker analysis. The roller compacted granules of both forms showed markedly different tabletting behaviour. Roller pressure exhibited a trend on compaction behaviour of form I granules, whereas, in case of form II, the effect was insignificant. Tabletability of the six granule batches follows the order; I_S1>I_S2>I_S3>II_S1≈II_S2≈II_S3. In case of form I, the reduced tabletability of the granules compacted at higher roller pressure was attributed to the decreased compressibility and plastic deformation. This was confirmed by compressibility plot and various mathematical parameters derived from Heckel (Py), Kawakita (1/b) and Walker (W) equations. The reduced tabletability of form I granules was due to 'granule hardening' during roller compaction. On the other hand, insignificant effect of roller compaction on tabletting behaviour of form II granules was attributed to brittle fragmentation. The extensive fragmentation of granules offered new 'clean' surfaces and higher contact points that negated the effect of granule hardening. Copyright © 2014 Elsevier B.V. All rights reserved.

A Quasi-Periodic Linear Feeder for the Impurity Granular Injection on DIII-D

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nagy, Alex; Bortolon, A.; Brown, W.

Injection of solid nonfuel pellets has been actively used as a tool for pacing and mitigation of edge localized modes (ELMs). In DIII-D, effective ELM pacing has been demonstrated by the high-frequency injection of lithium and carbon submillimeter spheres, using the impurity granule injector (IGI). This device injects granules into the plasma at speeds up to 150 m/s, through impact with a rotating impeller. In the IGI, high-frequency granule delivery was accomplished through a vibrational granule dropper, in which high time-average rates are obtained at the cost of lack of period control. Here, we present a new in-line granule feeder,more » capable of delivering granules of size 0.2-2 mm with no restriction of material properties, at quasi-periodic rates of up to 150 Hz, for 0.7-mm diameter lithium granules (600 Hz using 0.3-mm granules). The new dropper mechanism combines two piezo-in-line units; one which feeds the impeller and one which circulates granules that are filtered out of the feeder path. A remotely adjustable filter eliminates granules that are stacked, oversized, or side by side allowing the formation of a single moving granule injection line. The granules fall off the in-line feeder exit one at a time, achieving a quasi-periodic delivery rate proportional to the exit speed. At higher rates, the periodicity deteriorates. This behavior was studied using high-speed cameras and electrostatic measurements, and it was found that at drop rates <60 Hz, the granule delivery period has a variation of ±25% which appears to be caused by gaps which develop in the last centimeter of the injection line, as granules exit off the moving track. The linear feeder concept is robust against bridge instabilities and clogging issues, thanks to the simple diverter filter and constant recirculation of granules. Furthermore, the open-top design of the device facilitates refilling the device from separate reservoirs and has easy access for directly monitoring operation and adjustment. This paper describes the details of the in-line feeder design, along with several design iterations. The goal is a robust in-vacuum mechanism that can deliver granule flow ranging from a single particle to a line of particles at 150 per second, using different sizes and materials from the same apparatus.« less

A Quasi-Periodic Linear Feeder for the Impurity Granular Injection on DIII-D

DOE PAGES

Nagy, Alex; Bortolon, A.; Brown, W.; ...

2018-04-10

Injection of solid nonfuel pellets has been actively used as a tool for pacing and mitigation of edge localized modes (ELMs). In DIII-D, effective ELM pacing has been demonstrated by the high-frequency injection of lithium and carbon submillimeter spheres, using the impurity granule injector (IGI). This device injects granules into the plasma at speeds up to 150 m/s, through impact with a rotating impeller. In the IGI, high-frequency granule delivery was accomplished through a vibrational granule dropper, in which high time-average rates are obtained at the cost of lack of period control. Here, we present a new in-line granule feeder,more » capable of delivering granules of size 0.2-2 mm with no restriction of material properties, at quasi-periodic rates of up to 150 Hz, for 0.7-mm diameter lithium granules (600 Hz using 0.3-mm granules). The new dropper mechanism combines two piezo-in-line units; one which feeds the impeller and one which circulates granules that are filtered out of the feeder path. A remotely adjustable filter eliminates granules that are stacked, oversized, or side by side allowing the formation of a single moving granule injection line. The granules fall off the in-line feeder exit one at a time, achieving a quasi-periodic delivery rate proportional to the exit speed. At higher rates, the periodicity deteriorates. This behavior was studied using high-speed cameras and electrostatic measurements, and it was found that at drop rates <60 Hz, the granule delivery period has a variation of ±25% which appears to be caused by gaps which develop in the last centimeter of the injection line, as granules exit off the moving track. The linear feeder concept is robust against bridge instabilities and clogging issues, thanks to the simple diverter filter and constant recirculation of granules. Furthermore, the open-top design of the device facilitates refilling the device from separate reservoirs and has easy access for directly monitoring operation and adjustment. This paper describes the details of the in-line feeder design, along with several design iterations. The goal is a robust in-vacuum mechanism that can deliver granule flow ranging from a single particle to a line of particles at 150 per second, using different sizes and materials from the same apparatus.« less

Promiscuous interactions and protein disaggregases determine the material state of stress-inducible RNP granules

PubMed Central

Kroschwald, Sonja; Maharana, Shovamayee; Mateju, Daniel; Malinovska, Liliana; Nüske, Elisabeth; Poser, Ina; Richter, Doris; Alberti, Simon

2015-01-01

RNA-protein (RNP) granules have been proposed to assemble by forming solid RNA/protein aggregates or through phase separation into a liquid RNA/protein phase. Which model describes RNP granules in living cells is still unclear. In this study, we analyze P bodies in budding yeast and find that they have liquid-like properties. Surprisingly, yeast stress granules adopt a different material state, which is reminiscent of solid protein aggregates and controlled by protein disaggregases. By using an assay to ectopically nucleate RNP granules, we further establish that RNP granule formation does not depend on amyloid-like aggregation but rather involves many promiscuous interactions. Finally, we show that stress granules have different properties in mammalian cells, where they show liquid-like behavior. Thus, we propose that the material state of RNP granules is flexible and that the solid state of yeast stress granules is an adaptation to extreme environments, made possible by the presence of a powerful disaggregation machine. DOI: http://dx.doi.org/10.7554/eLife.06807.001 PMID:26238190

The design of free structure granular mappings: the use of the principle of justifiable granularity.

PubMed

Pedrycz, Witold; Al-Hmouz, Rami; Morfeq, Ali; Balamash, Abdullah

2013-12-01

The study introduces a concept of mappings realized in presence of information granules and offers a design framework supporting the formation of such mappings. Information granules are conceptually meaningful entities formed on a basis of a large number of experimental input–output numeric data available for the construction of the model. We develop a conceptually and algorithmically sound way of forming information granules. Considering the directional nature of the mapping to be formed, this directionality aspect needs to be taken into account when developing information granules. The property of directionality implies that while the information granules in the input space could be constructed with a great deal of flexibility, the information granules formed in the output space have to inherently relate to those built in the input space. The input space is granulated by running a clustering algorithm; for illustrative purposes, the focus here is on fuzzy clustering realized with the aid of the fuzzy C-means algorithm. The information granules in the output space are constructed with the aid of the principle of justifiable granularity (being one of the underlying fundamental conceptual pursuits of Granular Computing). The construct exhibits two important features. First, the constructed information granules are formed in the presence of information granules already constructed in the input space (and this realization is reflective of the direction of the mapping from the input to the output space). Second, the principle of justifiable granularity does not confine the realization of information granules to a single formalism such as fuzzy sets but helps form the granules expressed any required formalism of information granulation. The quality of the granular mapping (viz. the mapping realized for the information granules formed in the input and output spaces) is expressed in terms of the coverage criterion (articulating how well the experimental data are “covered” by information granules produced by the granular mapping for any input experimental data). Some parametric studies are reported by quantifying the performance of the granular mapping (expressed in terms of the coverage and specificity criteria) versus the values of a certain parameters utilized in the construction of output information granules through the principle of justifiable granularity. The plots of coverage–specificity dependency help determine a knee point and reach a sound compromise between these two conflicting requirements imposed on the quality of the granular mapping. Furthermore, quantified is the quality of the mapping with regard to the number of information granules (implying a certain granularity of the mapping). A series of experiments is reported as well.

Docking is not a prerequisite but a temporal constraint for fusion of secretory granules.

PubMed

Kasai, Kazuo; Fujita, Takuji; Gomi, Hiroshi; Izumi, Tetsuro

2008-07-01

We examined secretory granule dynamics using total internal reflection fluorescence microscopy in normal pancreatic beta cells and their mutants devoid of Rab27a and/or its effector, granuphilin, which play critical roles in the docking and recruitment of insulin granules to the plasma membrane. In the early phase of glucose stimulation in wild-type cells, we observed marked fusion of granules recruited from a relatively distant area, in parallel with that from granules located underneath the plasma membrane. Furthermore, despite a lack of granules directly attached to the plasma membrane, both spontaneous and evoked fusion was increased in granuphilin-null cells. In addition to these granuphilin-null phenotypes, Rab27a/granuphilin doubly deficient cells showed the decreases in granules located next to the docked area and in fusion from granules near the plasma membrane in the early phase of glucose-stimulated secretion, similar to Rab27a-mutated cells. Thus, the two proteins play nonoverlapping roles in insulin exocytosis: granuphilin acts on the granules underneath the plasma membrane, whereas Rab27a acts on those in a more distal area. These findings demonstrate that, in contrast to our conventional understanding, stable attachment of secretory granules to the plasma membrane is not prerequisite but temporally inhibitory for both spontaneous and evoked fusion.

TDP-43 is directed to stress granules by sorbitol, a novel physiological osmotic and oxidative stressor.

PubMed

Dewey, Colleen M; Cenik, Basar; Sephton, Chantelle F; Dries, Daniel R; Mayer, Paul; Good, Shannon K; Johnson, Brett A; Herz, Joachim; Yu, Gang

2011-03-01

TDP-43, or TAR DNA-binding protein 43, is a pathological marker of a spectrum of neurodegenerative disorders, including amyotrophic lateral sclerosis and frontotemporal lobar degeneration with ubiquitin-positive inclusions. TDP-43 is an RNA/DNA-binding protein implicated in transcriptional and posttranscriptional regulation. Recent work also suggests that TDP-43 associates with cytoplasmic stress granules, which are transient structures that form in response to stress. In this study, we establish sorbitol as a novel physiological stressor that directs TDP-43 to stress granules in Hek293T cells and primary cultured glia. We quantify the association of TDP-43 with stress granules over time and show that stress granule association and size are dependent on the glycine-rich region of TDP-43, which harbors the majority of pathogenic mutations. Moreover, we establish that cells harboring wild-type and mutant TDP-43 have distinct stress responses: mutant TDP-43 forms significantly larger stress granules, and is incorporated into stress granules earlier, than wild-type TDP-43; in striking contrast, wild-type TDP-43 forms more stress granules over time, but the granule size remains relatively unchanged. We propose that mutant TDP-43 alters stress granule dynamics, which may contribute to the progression of TDP-43 proteinopathies.

Distribution of a viscous binder during high shear granulation--sensitivity to the method of delivery and its impact on product properties.

PubMed

Tan, Bernice Mei Jin; Loh, Zhi Hui; Soh, Josephine Lay Peng; Liew, Celine Valeria; Heng, Paul Wan Sia

2014-01-02

Binder distribution in the powder mass during high shear granulation is especially critical with the use of viscous liquid binders and with short processing times. A viscous liquid binder was delivered into the powder mass at two flow rates using three methods: pouring, pumping and spraying from a pressure pot. Binder content analyses at the scale of individual granules were conducted to investigate the impact of different delivery conditions on the homogeneity of binder distribution. There was clear evidence of non-uniformity of binder content among individual granules across all delivery conditions, particularly for the fast rates of delivery. Poorer reproducibility values of tablet thickness and disintegration time were observed when binder was poured but this may be overcome by pumping or spraying from the pressure pot. Greater homogeneity of binder distribution occurred with the slow rates of delivery and led to the earlier onset of granule growth and a consequent increase in granule size. Larger granule size and lower proportion of fines were in turn associated with increased granule bulk density and improvement of granule flow. In conclusion, delivery of a viscous binder at a slow rate either by pumping or via a pressure pot was most desirable during granulation. Copyright © 2013 Elsevier B.V. All rights reserved.

Common spectrum of polypeptides occurs in secretion granule membranes of different exocrine glands

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cameron, R.S.; Cameron, P.L.; Castle, J.D.

1986-10-01

A highly purified membrane preparation from rat parotid secretion granules has been used as a comparative probe to examine the extent of compositional overlap in granule membranes of three other exocrine secretory tissues - pancreatic, lacrimal, and submandibular - from several standpoints. First, indirect immunofluorescent studies using a polyclonal polyspecific anti-parotid granule membrane antiserum has indicated a selective staining of granule membrane profiles in all acinar cells of all tissues. Second, highly purified granule membrane subfractions have been isolated from each exocrine tissue; comparative two-dimensional (isoelectric focusing; SDS) PAGE of radioiodinated granule membranes has identified 10-15 polypeptides of identical pImore » and apparent molecular mass. These species are likely to be integral membrane components since they are not extracted by either saponin-sodium sulfate or sodium carbonate (pH 11.5) treatments, and they do not have counterparts in the granule content. Finally, the identity among selected parotid and pancreatic radioiodinated granule membrane polypeptides has been documented using two-dimensional peptide mapping of chymotryptic and tryptic digests. These findings clearly indicate that exocrine secretory granules, irrespective of the nature of stored secretion, comprise a type of vesicular carrier with a common (and probably refined) membrane composition. Conceivably, the polypeptides identified carry out general functions related to exocrine secretion.« less

Critical evaluation of root causes of the reduced compactability after roll compaction/dry granulation.

PubMed

Mosig, Johanna; Kleinebudde, Peter

2015-03-01

The influence of lubrication and particle size on the reduced compactability after dry granulation was investigated. Powder cellulose, lactose, magnesium carbonate, and two types of microcrystalline cellulose were roll compacted, granulated, and sieved into particle fractions. Particle fractions were compressed into tablets using internal and external lubrication. Internal lubrication resulted in an overlubrication of the granule material compared with the powder material. This resulted in extraordinary high reduction of compactability after dry granulation for lubricant-sensitive materials. The granule size can cause differences in strength, whereby the degree of this effect was material dependent. The loss in strength with increasing compaction force was comparable for different particles sizes of one material, suggesting a change in material properties independently of the size. Granule hardening could be one reason as for higher compaction forces the integrity of the granule structure survived the compression step. The results demonstrated that granule lubrication mainly influence the degree of the reduced compactability after dry granulation and must be considered for the evaluation of mechanism for this phenomenon. Hardening of the material as well as size enlargement will cause the loss in strength after recompression, but the influence of both depends strongly on the material. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

A comparative study of the influence of alpha-lactose monohydrate particle morphology on granule and tablet properties after roll compaction/dry granulation.

PubMed

Grote, Simon; Kleinebudde, Peter

2018-05-29

The influence of particle morphology and size of alpha-lactose monohydrate on dry granules and tablets was studied. Four different morphologies were investigated: Two grades of primary crystals, which differed in their particle size and structure (compact crystals vs. agglomerates). The materials were roll compacted at different specific compaction forces and changes in the particle size distribution and the specific surface area were measured. Afterwards, two fractions of granules were pressed to tablets and the tensile strength was compared to that from tablets compressed from the raw materials. The specific surface area was increased induced by roll compaction/dry granulation for all materials. At increased specific compaction forces, the materials showed sufficient size enlargement. The morphology of lactose determined the strength of direct compressed tablets. In contrast, the strength of granule tablets was leveled by the previous compression step during roll compaction/dry granulation. Thus, the tensile strength of tablets compressed directly from the powder mixtures determined whether materials exhibited a loss in tabletability after roll compaction/dry granulation or not. The granule size had only a slight influence on the strength of produced tablets. In some cases, the fraction of smaller granules showed a higher tensile strength compared to the larger fraction.

Joint Effects of Granule Size and Degree of Substitution on Octenylsuccinated Sweet Potato Starch Granules As Pickering Emulsion Stabilizers.

PubMed

Li, Jinfeng; Ye, Fayin; Lei, Lin; Zhou, Yun; Zhao, Guohua

2018-05-02

The granules of sweet potato starch were size fractionated into three portions with significantly different median diameters ( D 50 ) of 6.67 (small-sized), 11.54 (medium-sized), and 16.96 μm (large-sized), respectively. Each portion was hydrophobized at the mass-based degrees of substitution (DS m ) of approximately 0.0095 (low), 0.0160 (medium), and 0.0230 (high). The Pickering emulsion-stabilizing capacities of modified granules were tested, and the resultant emulsions were characterized. The joint effects of granule size and DS m on emulsifying capacity (EC) were investigated by response surface methodology. For small-, medium-, and large-sized fractions, their highest emulsifying capacities are comparable but, respectively, encountered at high (0.0225), medium (0.0158), and low (0.0095) DS m levels. The emulsion droplet size increased with granule size, and the number of freely scattered granules in emulsions decreased with DS m . In addition, the term of surface density of the octenyl succinic group (SD -OSG ) was first proposed for modified starch granules, and it was proved better than DS m in interpreting the emulsifying capacities of starch granules with varying sizes. The present results implied that, as the particulate stabilizers, the optimal DS m of modified starch granules is size specific.

Denitrifying capability and community dynamics of glycogen accumulating organisms during sludge granulation in an anaerobic-aerobic sequencing batch reactor

PubMed Central

Bin, Zhang; Bin, Xue; Zhigang, Qiu; Zhiqiang, Chen; Junwen, Li; Taishi, Gong; Wenci, Zou; Jingfeng, Wang

2015-01-01

Denitrifying capability of glycogen accumulating organisms (GAOs) has received great attention in environmental science and microbial ecology. Combining this ability with granule processes would be an interesting attempt. Here, a laboratory-scale sequencing batch reactor (SBR) was operated to enrich GAOs and enable sludge granulation. The results showed that the GAO granules were cultivated successfully and the granules had denitrifying capability. The batch experiments demonstrated that all NO3−-N could be removed or reduced, some amount of NO2−-N were accumulated in the reactor, and N2 was the main gaseous product. SEM analysis suggested that the granules were tightly packed with a large amount of tetrad-forming organisms (TFOs); filamentous bacteria served as the supporting structures for the granules. The microbial community structure of GAO granules was differed substantially from the inoculant conventional activated sludge. Most of the bacteria in the seed sludge grouped with members of Proteobacterium. FISH analysis confirmed that GAOs were the predominant members in the granules and were distributed evenly throughout the granular space. In contrast, PAOs were severely inhibited. Overall, cultivation of the GAO granules and utilizing their denitrifying capability can provide us with a new approach of nitrogen removal and saving more energy. PMID:26257096

Denitrifying capability and community dynamics of glycogen accumulating organisms during sludge granulation in an anaerobic-aerobic sequencing batch reactor.

PubMed

Bin, Zhang; Bin, Xue; Zhigang, Qiu; Zhiqiang, Chen; Junwen, Li; Taishi, Gong; Wenci, Zou; Jingfeng, Wang

2015-08-10

Denitrifying capability of glycogen accumulating organisms (GAOs) has received great attention in environmental science and microbial ecology. Combining this ability with granule processes would be an interesting attempt. Here, a laboratory-scale sequencing batch reactor (SBR) was operated to enrich GAOs and enable sludge granulation. The results showed that the GAO granules were cultivated successfully and the granules had denitrifying capability. The batch experiments demonstrated that all NO3(-)-N could be removed or reduced, some amount of NO2(-)-N were accumulated in the reactor, and N2 was the main gaseous product. SEM analysis suggested that the granules were tightly packed with a large amount of tetrad-forming organisms (TFOs); filamentous bacteria served as the supporting structures for the granules. The microbial community structure of GAO granules was differed substantially from the inoculant conventional activated sludge. Most of the bacteria in the seed sludge grouped with members of Proteobacterium. FISH analysis confirmed that GAOs were the predominant members in the granules and were distributed evenly throughout the granular space. In contrast, PAOs were severely inhibited. Overall, cultivation of the GAO granules and utilizing their denitrifying capability can provide us with a new approach of nitrogen removal and saving more energy.

Texture Analysis of Poly-Adenylated mRNA Staining Following Global Brain Ischemia and Reperfusion

PubMed Central

Szymanski, Jeffrey J.; Jamison, Jill T.; DeGracia, Donald J.

2011-01-01

Texture analysis provides a means to quantify complex changes in microscope images. We previously showed that cytoplasmic poly-adenylated mRNAs form mRNA granules in post-ischemic neurons and that these granules correlated with protein synthesis inhibition and hence cell death. Here we utilized the texture analysis software MaZda to quantify mRNA granules in photomicrographs of the pyramidal cell layer of rat hippocampal region CA3 around 1 hour of reperfusion after 10 min of normothermic global cerebral ischemia. At 1 hour reperfusion, we observed variations in the texture of mRNA granules amongst samples that were readily quantified by texture analysis. Individual sample variation was consistent with the interpretation that animal-to-animal variations in mRNA granules reflected the time-course of mRNA granule formation. We also used texture analysis to quantify the effect of cycloheximide, given either before or after brain ischemia, on mRNA granules. If administered before ischemia, cycloheximide inhibited mRNA granule formation, but if administered after ischemia did not prevent mRNA granulation, indicating mRNA granule formation is dependent on dissociation of polysomes. We conclude that texture analysis is an effective means for quantifying the complex morphological changes induced in neurons by brain ischemia and reperfusion. PMID:21477879

Spatial patterning of P granules by RNA-induced phase separation of the intrinsically-disordered protein MEG-3

PubMed Central

Smith, Jarrett; Calidas, Deepika; Schmidt, Helen; Lu, Tu; Rasoloson, Dominique; Seydoux, Geraldine

2016-01-01

RNA granules are non-membrane bound cellular compartments that contain RNA and RNA binding proteins. The molecular mechanisms that regulate the spatial distribution of RNA granules in cells are poorly understood. During polarization of the C. elegans zygote, germline RNA granules, called P granules, assemble preferentially in the posterior cytoplasm. We present evidence that P granule asymmetry depends on RNA-induced phase separation of the granule scaffold MEG-3. MEG-3 is an intrinsically disordered protein that binds and phase separates with RNA in vitro. In vivo, MEG-3 forms a posterior-rich concentration gradient that is anti-correlated with a gradient in the RNA-binding protein MEX-5. MEX-5 is necessary and sufficient to suppress MEG-3 granule formation in vivo, and suppresses RNA-induced MEG-3 phase separation in vitro. Our findings suggest that MEX-5 interferes with MEG-3’s access to RNA, thus locally suppressing MEG-3 phase separation to drive P granule asymmetry. Regulated access to RNA, combined with RNA-induced phase separation of key scaffolding proteins, may be a general mechanism for controlling the formation of RNA granules in space and time. DOI: http://dx.doi.org/10.7554/eLife.21337.001 PMID:27914198

Denitrifying capability and community dynamics of glycogen accumulating organisms during sludge granulation in an anaerobic-aerobic sequencing batch reactor

NASA Astrophysics Data System (ADS)

Bin, Zhang; Bin, Xue; Zhigang, Qiu; Zhiqiang, Chen; Junwen, Li; Taishi, Gong; Wenci, Zou; Jingfeng, Wang

2015-08-01

Denitrifying capability of glycogen accumulating organisms (GAOs) has received great attention in environmental science and microbial ecology. Combining this ability with granule processes would be an interesting attempt. Here, a laboratory-scale sequencing batch reactor (SBR) was operated to enrich GAOs and enable sludge granulation. The results showed that the GAO granules were cultivated successfully and the granules had denitrifying capability. The batch experiments demonstrated that all NO3--N could be removed or reduced, some amount of NO2--N were accumulated in the reactor, and N2 was the main gaseous product. SEM analysis suggested that the granules were tightly packed with a large amount of tetrad-forming organisms (TFOs); filamentous bacteria served as the supporting structures for the granules. The microbial community structure of GAO granules was differed substantially from the inoculant conventional activated sludge. Most of the bacteria in the seed sludge grouped with members of Proteobacterium. FISH analysis confirmed that GAOs were the predominant members in the granules and were distributed evenly throughout the granular space. In contrast, PAOs were severely inhibited. Overall, cultivation of the GAO granules and utilizing their denitrifying capability can provide us with a new approach of nitrogen removal and saving more energy.

Tryptophanol-derived oxazolopiperidone lactams: identification of a hit compound as NMDA receptor antagonist.

PubMed

Pereira, Nuno A L; Sureda, Francesc X; Esplugas, Roser; Pérez, Maria; Amat, Mercedes; Santos, Maria M M

2014-08-01

N-Methyl-D-aspartate receptors (NMDAR) exacerbated activation leads to neuron death through a phenomenon called excitotoxicity. These receptors are implicated in several neurological diseases (e.g., Alzheimer and Parkinson) and thus represent an important therapeutic target. We herein describe the study of enantiopure tryptophanol-derived oxazolopiperidone lactams as NMDA receptor antagonists. The most active hit exhibited an IC50 of 63.4 μM in cultured rat cerebellar granule neurons thus being 1.5 fold more active than clinically approved NMDA antagonist amantadine (IC50=92 μM). Copyright © 2014 Elsevier Ltd. All rights reserved.

Corin mutations K317E and S472G from preeclamptic patients alter zymogen activation and cell surface targeting. [Corrected].

PubMed

Dong, Ningzheng; Zhou, Tiantian; Zhang, Yue; Liu, Meng; Li, Hui; Huang, Xiaoyi; Liu, Zhenzhen; Wu, Yi; Fukuda, Koichi; Qin, Jun; Wu, Qingyu

2014-06-20

Corin is a membrane-bound serine protease that acts as the atrial natriuretic peptide (ANP) convertase in the heart. Recent studies show that corin also activates ANP in the pregnant uterus to promote spiral artery remodeling and prevent pregnancy-induced hypertension. Two CORIN gene mutations, K317E and S472G, were identified in preeclamptic patients and shown to have reduced activity in vitro. In this study, we carried out molecular modeling and biochemical experiments to understand how these mutations impair corin function. By molecular modeling, the mutation K317E was predicted to alter corin LDL receptor-2 module conformation. Western blot analysis of K317E mutant in HEK293 cells showed that the mutation did not block corin expression on the cell surface but inhibited corin zymogen activation. In contrast, the mutation S472G was predicted to abolish a β-sheet critical for corin frizzled-2 module structure. In Western blot analysis and flow cytometry, S472G mutant was not detected on the cell surface in transfected HEK293 cells. By immunostaining, the S472G mutant was found in the ER, indicating that the mutation S472G disrupted the β-sheet, causing corin misfolding and ER retention. Thus, these results show that mutations in the CORIN gene may impair corin function by entirely different mechanisms. Together, our data provide important insights into the molecular basis underlying corin mutations that may contribute to preeclampsia in patients. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

RNA in development: how ribonucleoprotein granules regulate the life cycles of pathogenic protozoa.

PubMed

Kramer, Susanne

2014-01-01

Ribonucleoprotein (RNP) granules are important posttranscriptional regulators of messenger RNA (mRNA) fate. Several types of RNP granules specifically regulate gene expression during development of multicellular organisms and are commonly referred to as germ granules. The function of germ granules is not entirely understood and probably diverse, but it is generally agreed that one main function is posttranscriptional regulation of gene expression during early development, when transcription is silent. One example is the translational repression of maternally derived mRNAs in oocytes. Here, I hope to show that the need for regulation of gene expression by RNP granules is not restricted to animal development, but plays an equally important role during the development of pathogenic protozoa. Apicomplexa and Trypanosomatidae have complex life cycles with frequent host changes. The need to quickly adapt gene expression to a new environment as well as the ability to suppress translation to survive latencies is critical for successful completion of life cycles. Posttranscriptional gene regulation is not necessarily simpler in protozoa. Apicomplexa surprise with the presence of micro RNA (miRNAs) and upstream open reading frames (µORFs). Trypanosomes have an unusually large repertoire of different RNP granule types. A better understanding of RNP granules in protozoa may help to gain insight into the evolutionary origin of RNP granules: Trypanosomes for example have two types of granules with interesting similarities to animal germ granules. © 2013 John Wiley & Sons, Ltd.

Formation of tRNA granules in the nucleus of heat-induced human cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miyagawa, Ryu; Department of Biological Science, Graduate School of Science, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8654; Mizuno, Rie

Highlights: Black-Right-Pointing-Pointer tRNAs are tranlocated into the nucleus in heat-induced HeLa cells. Black-Right-Pointing-Pointer tRNAs form the unique granules in the nucleus. Black-Right-Pointing-Pointer tRNA ganules overlap with nuclear stress granules. -- Abstract: The stress response, which can trigger various physiological phenomena, is important for living organisms. For instance, a number of stress-induced granules such as P-body and stress granule have been identified. These granules are formed in the cytoplasm under stress conditions and are associated with translational inhibition and mRNA decay. In the nucleus, there is a focus named nuclear stress body (nSB) that distinguishes these structures from cytoplasmic stress granules.more » Many splicing factors and long non-coding RNA species localize in nSBs as a result of stress. Indeed, tRNAs respond to several kinds of stress such as heat, oxidation or starvation. Although nuclear accumulation of tRNAs occurs in starved Saccharomyces cerevisiae, this phenomenon is not found in mammalian cells. We observed that initiator tRNA{sup Met} (Meti) is actively translocated into the nucleus of human cells under heat stress. During this study, we identified unique granules of Meti that overlapped with nSBs. Similarly, elongator tRNA{sup Met} was translocated into the nucleus and formed granules during heat stress. Formation of tRNA granules is closely related to the translocation ratio. Then, all tRNAs may form the specific granules.« less

Gastroretentive extended-release floating granules prepared using a novel fluidized hot melt granulation (FHMG) technique.

PubMed

Zhai, H; Jones, D S; McCoy, C P; Madi, A M; Tian, Y; Andrews, G P

2014-10-06

The objective of this work was to investigate the feasibility of using a novel granulation technique, namely, fluidized hot melt granulation (FHMG), to prepare gastroretentive extended-release floating granules. In this study we have utilized FHMG, a solvent free process in which granulation is achieved with the aid of low melting point materials, using Compritol 888 ATO and Gelucire 50/13 as meltable binders, in place of conventional liquid binders. The physicochemical properties, morphology, floating properties, and drug release of the manufactured granules were investigated. Granules prepared by this method were spherical in shape and showed good flowability. The floating granules exhibited sustained release exceeding 10 h. Granule buoyancy (floating time and strength) and drug release properties were significantly influenced by formulation variables such as excipient type and concentration, and the physical characteristics (particle size, hydrophilicity) of the excipients. Drug release rate was increased by increasing the concentration of hydroxypropyl cellulose (HPC) and Gelucire 50/13, or by decreasing the particle size of HPC. Floating strength was improved through the incorporation of sodium bicarbonate and citric acid. Furthermore, floating strength was influenced by the concentration of HPC within the formulation. Granules prepared in this way show good physical characteristics, floating ability, and drug release properties when placed in simulated gastric fluid. Moreover, the drug release and floating properties can be controlled by modification of the ratio or physical characteristics of the excipients used in the formulation.

High shear mixing granulation of ibuprofen and beta-cyclodextrin: effects of process variables on ibuprofen dissolution.

PubMed

Ghorab, Mohamed K; Adeyeye, Moji Christianah

2007-10-19

The aims of the study were to evaluate the effect of high shear mixer (HSM) granulation process parameters and scale-up on wet mass consistency and granulation characteristics. A mixer torque rheometer (MTR) was employed to evaluate the granulating solvents used (water, isopropanol, and 1:1 vol/vol mixture of both) based on the wet mass consistency. Gral 25 and mini-HSM were used for the granulation. The MTR study showed that the water significantly enhanced the beta-cyclodextrin (beta CD) binding tendency and the strength of liquid bridges formed between the particles, whereas the isopropanol/water mixture yielded more suitable agglomerates. Mini-HSM granulation with the isopropanol/water mixture (1:1 vol/vol) showed a reduction in the extent of torque value rise by increasing the impeller speed as a result of more breakdown of agglomerates than coalescence. In contrast, increasing the impeller speed of the Gral 25 resulted in higher torque readings, larger granule size, and consequently, slower dissolution. This was due to a remarkable rise in temperature during Gral granulation that reduced the isopropanol/water ratio in the granulating solvent as a result of evaporation and consequently increased the beta CD binding strength. In general, the HSM granulation retarded ibuprofen dissolution compared with the physical mixture because of densification and agglomeration. However, a successful HSM granulation scale-up was not achieved due to the difference in the solvent mixture's effect from 1 scale to the other.

Second Harmonic Generation Mediated by Aligned Water in Starch Granules.

PubMed

Cisek, Richard; Tokarz, Danielle; Krouglov, Serguei; Steup, Martin; Emes, Michael J; Tetlow, Ian J; Barzda, Virginijus

2014-12-26

The origin of second harmonic generation (SHG) in starch granules was investigated using ab initio quantum mechanical modeling and experimentally examined using polarization-in, polarization-out (PIPO) second harmonic generation microscopy. Ab initio calculations revealed that the largest contribution to the SHG signal from A- and B-type allomorphs of starch originates from the anisotropic organization of hydroxide and hydrogen bonds mediated by aligned water found in the polymers. The hypothesis was experimentally tested by imaging maize starch granules under various hydration and heat treatment conditions that alter the hydrogen bond network. The highest SHG intensity was found in fully hydrated starch granules, and heat treatment diminished the SHG intensity. The PIPO SHG imaging showed that dried starch granules have a much higher nonlinear optical susceptibility component ratio than fully hydrated granules. In contrast, deuterated starch granules showed a smaller susceptibility component ratio demonstrating that SHG is highly sensitive to the organization of the hydroxyl and hydrogen bond network. The polarization SHG imaging results of potato starch granules, representing starch allomorph B, were compared to those of maize starch granules representing allomorph A. The results showed that the amount of aligned water was higher in the maize granules. Nonlinear microscopy of starch granules provides evidence that varying hydration conditions leads to significant changes in the nonlinear susceptibility ratio as well as the SHG intensity, supporting the hypothesis from ab initio calculations that the dominant contribution to SHG is due to the ordered hydroxide and hydrogen bond network.

Digestion of Starch Granules from Maize, Potato and Wheat by Larvae of the the Yellow Mealworm, Tenebrio molitor and the Mexican Bean Weevil, Zabrotes subfasciatus

PubMed Central

Meireles, Elaine A.; Carneiro, Cíntia N. B.; DaMatta, Renato A.; Samuels, Richard I.; Silva, Carlos P.

2009-01-01

Scanning electron microscopy images were taken of starch granules from different sources following exposure in vivo and in vitro to gut α-amylases isolated from Tenebrio molitor L. (Coleoptera: Tenebrionidae) and Zabrotes subfasciatus Boheman (Coleoptera: Bruchidae). One α-amylase was isolated from whole larval midguts of T. molitor using non-denaturing SDS-PAGE, while two other α-amylase fractions were isolated from whole larval midguts of Z. subfasciatus using hydrophobic interaction chromatography., Digested starch granules from larvae fed on maize, potato or wheat were isolated from midgut contents. Combinations of starch granules with isolated α-amylases from both species showed similar patterns of granule degradation. In vitro enzymatic degradation of maize starch granules by the three different α-amylase fractions began by creating small holes and crater-like areas on the surface of the granules. Over time, these holes increased in number and area resulting in extensive degradation of the granule structure. Granules from potato did not show formation of pits and craters on their surface, but presented extensive erosion in their interior. For all types of starch, as soon as the interior of the starch granule was reached, the inner layers of amylose and amylopectin were differentially hydrolyzed, resulting in a striated pattern. These data support the hypothesis that the pattern of starch degradation depends more on the granule type than on the α-amylase involved. PMID:19619014

[Study on spray-drier preparation technology of weitai granules using orthogonal experiments].

PubMed

Qu, Cai-Hong; Yang, Li; Chen, Zhi-Liang

2006-04-01

To screen preparation technique in order to raise the end-product and economical efficiency of spray-drier preparation technology of weitai granules. Newly Fluid-bed-spray-drier-granulation technique was adoped and taken extracting technique, temperature of exit and entry and the matching of accessories as inspecting factors, two levels of each factors, end-product and the water content of semi-finished weitai granules as inspecting marker, the best preparation technique of weitai granules was screened by orthogonal desing. Among the 3 factors, the matching of accessories was most notalbe (P < 0.01), next was the temperature of exit and entry (P < 0.05). However, the extracting technique was of little importance (P > 0.05). The optimum spray-drier granulation technique of weitai granules is A3B1C3.

New gentle-wing high-shear granulator: impact of processing variables on granules and tablets characteristics of high-drug loading formulation using design of experiment approach.

PubMed

Fayed, Mohamed H; Abdel-Rahman, Sayed I; Alanazi, Fars K; Ahmed, Mahrous O; Tawfeek, Hesham M; Al-Shdefat, Ramadan I

2017-10-01

The aim of this work was to study the application of design of experiment (DoE) approach in defining design space for granulation and tableting processes using a novel gentle-wing high-shear granulator. According to quality-by-design (QbD) prospective, critical attributes of granules, and tablets should be ensured by manufacturing process design. A face-centered central composite design has been employed in order to investigate the effect of water amount (X 1 ), impeller speed (X 2 ), wet massing time (X 3 ), and water addition rate (X 4 ) as independent process variables on granules and tablets characteristics. Acetaminophen was used as a model drug and granulation experiments were carried out using dry addition of povidone k30. The dried granules have been analyzed for their size distribution, density, and flow pattern. Additionally, the produced tablets have been investigated for; weight uniformity, breaking force, friability and percent capping, disintegration time, and drug dissolution. Results of regression analysis showed that water amount, impeller speed and wet massing time have significant (p < .05) effect on granules and tablets characteristics. However, the water amount had the most pronounced effect as indicated by its higher parameter estimate. On the other hand, water addition rate showed a minimal impact on granules and tablets properties. In conclusion, water amount, impeller speed, and wet massing time could be considered as critical process variables. Thus, understanding the relationship between these variables and quality attributes of granules and corresponding tablets provides the basis for adjusting granulation variables in order to optimize product performance.

Assessment of bacterial and structural dynamics in aerobic granular biofilms

PubMed Central

Weissbrodt, David G.; Neu, Thomas R.; Kuhlicke, Ute; Rappaz, Yoan; Holliger, Christof

2013-01-01

Aerobic granular sludge (AGS) is based on self-granulated flocs forming mobile biofilms with a gel-like consistence. Bacterial and structural dynamics from flocs to granules were followed in anaerobic-aerobic sequencing batch reactors (SBR) fed with synthetic wastewater, namely a bubble column (BC-SBR) operated under wash-out conditions for fast granulation, and two stirred-tank enrichments of Accumulibacter (PAO-SBR) and Competibacter (GAO-SBR) operated at steady-state. In the BC-SBR, granules formed within 2 weeks by swelling of Zoogloea colonies around flocs, developing subsequently smooth zoogloeal biofilms. However, Zoogloea predominance (37–79%) led to deteriorated nutrient removal during the first months of reactor operation. Upon maturation, improved nitrification (80–100%), nitrogen removal (43–83%), and high but unstable dephosphatation (75–100%) were obtained. Proliferation of dense clusters of nitrifiers, Accumulibacter, and Competibacter from granule cores outwards resulted in heterogeneous bioaggregates, inside which only low abundance Zoogloea (<5%) were detected in biofilm interstices. The presence of different extracellular glycoconjugates detected by fluorescence lectin-binding analysis showed the complex nature of the intracellular matrix of these granules. In the PAO-SBR, granulation occurred within two months with abundant and active Accumulibacter populations (56 ± 10%) that were selected under full anaerobic uptake of volatile fatty acids and that aggregated as dense clusters within heterogeneous granules. Flocs self-granulated in the GAO-SBR after 480 days during a period of over-aeration caused by biofilm growth on the oxygen sensor. Granules were dominated by heterogeneous clusters of Competibacter (37 ± 11%). Zoogloea were never abundant in biomass of both PAO- and GAO-SBRs. This study showed that Zoogloea, Accumulibacter, and Competibacter affiliates can form granules, and that the granulation mechanisms rely on the dominant population involved. PMID:23847600

Detection and Analysis of the Quality of Ibuprofen Granules

NASA Astrophysics Data System (ADS)

Yu-bin, Ji; Xin, LI; Guo-song, Xin; Qin-bing, Xue

2017-12-01

The Ibuprofen Granules comprehensive quality testing to ensure that it is in accordance with the provisions of Chinese pharmacopoeia. With reference of Chinese pharmacopoeia, the Ibuprofen Granules is tested by UV, HPLC, in terms of grain size checking, volume deviation, weight loss on drying detection, dissolution rate detection, and quality evaluation. Results indicated that Ibuprofen Granules conform to the standards. The Ibuprofen Granules are qualified and should be permitted to be marketed.

Carbon, nitrogen and phosphorus removal mechanisms of aerobic granules.

PubMed

Sarma, Saurabh Jyoti; Tay, Joo-Hwa

2018-04-10

Aerobic granules are the potential tools to develop modern wastewater treatment technologies with improved nutrient removal efficiency. These granules have several promising advantages over conventional activated sludge-based wastewater treatment processes. This technology has the potential of reducing the infrastructure and operation costs of wastewater treatment by 25%, energy requirement by 30%, and space requirement by 75%. The nutrient removal mechanisms of aerobic granules are slightly different from that of the activated sludge. For instance, unlike activated sludge process, according to some reports, as high as 70% of the total phosphorus removed by aerobic granules were attributed to precipitation within the granules. Similarly, aerobic granule-based technology reduces the total amount of sludge produced during wastewater treatment. However, the reason behind this observation is unknown and it needs further explanations based on carbon and nitrogen removal mechanisms. Thus, as a part of the present review, a set of new hypotheses have been proposed to explain the peculiar nutrient removal mechanisms of the aerobic granules.

Native granule associated short chain length polyhydroxyalkanoate synthase from a marine derived Bacillus sp. NQ-11/A2.

PubMed

Prabhu, Nimali N; Santimano, Maria Celisa; Mavinkurve, Suneela; Bhosle, Saroj N; Garg, Sandeep

2010-01-01

A rapidly growing marine derived Bacillus sp. strain NQ-11/A2, identified as Bacillus megaterium, accumulated 61% polyhydroxyalkanoate by weight. Diverse carbon sources served as substrates for the accumulation of short chain length polyhydroxyalkanoate. Three to nine granules either single or attached as buds could be isolated intact from each cell. Maximum activity of polyhydroxyalkanoate synthase was associated with the granules. Granule-bound polyhydroxyalkanoate synthase had a K(m) of 7.1 x 10(-5) M for DL-beta-hydroxybutyryl-CoA. Temperature and pH optima for maximum activity were 30 degrees C and 7.0, respectively. Sodium ions were required for granule-bound polyhydroxyalkanoate synthase activity and inhibited by potassium. Granule-bound polyhydroxyalkanoate synthase was apparently covalently bound to the polyhydroxyalkanoate-core of the granules and affected by the chaotropic reagent urea. Detergents inhibited the granule-bound polyhydroxyalkanoate synthase drastically whilst glycerol and bovine serum albumin stabilized the synthase.

Artificial Intelligence Tools for Scaling Up of High Shear Wet Granulation Process.

PubMed

Landin, Mariana

2017-01-01

The results presented in this article demonstrate the potential of artificial intelligence tools for predicting the endpoint of the granulation process in high-speed mixer granulators of different scales from 25L to 600L. The combination of neurofuzzy logic and gene expression programing technologies allowed the modeling of the impeller power as a function of operation conditions and wet granule properties, establishing the critical variables that affect the response and obtaining a unique experimental polynomial equation (transparent model) of high predictability (R 2 > 86.78%) for all size equipment. Gene expression programing allowed the modeling of the granulation process for granulators of similar and dissimilar geometries and can be improved by implementing additional characteristics of the process, as composition variables or operation parameters (e.g., batch size, chopper speed). The principles and the methodology proposed here can be applied to understand and control manufacturing process, using any other granulation equipment, including continuous granulation processes. Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Sustained-release progesterone vaginal suppositories 1--development of sustained-release granule--.

PubMed

Nakayama, Ayako; Sunada, Hisakazu; Okamoto, Hirokazu; Furuhashi, Kaoru; Ohno, Yukiko; Ito, Mikio

2009-02-01

Progesterone (P) is an important hormone for the establishment of pregnancy, and its administration is useful for luteal insufficiency. Considering the problems of commercially available oral and injection drugs, hospital-formulated vaginal suppositories are clinically used. However, since the half-life of P suppositories is short, it is difficult to maintain its constant blood concentration. To sustain drug efficacy and prevent side-effects, we are attempting to develop sustained-release suppositories by examining the degree of sustained-release of active ingredients. In this study, we examined the combinations of granulation methods and release systems for the preparation of sustained-release granules of P, and produced 13 types of sustained-release granules. We also examined the diameter, content, and dissolution of each type of granules, and confirmed that the sustained-release of all types of granules was satisfactory. Among the sustained-release granules, we selected granules with a content and a degree of sustained-release suitable for sustained-release suppositories.

Preparation of orally disintegrating tablets with taste-masking function: masking effect in granules prepared with correctives using the dry granulation method and evaluation of tablets prepared using the taste-masked granules.

PubMed

Kawano, Yayoi; Ito, Akihiko; Sasatsu, Masanaho; Machida, Yoshiharu

2010-01-01

We investigated several methods of taste masking in the preparation of orally disintegrating tablets (ODTs), using furosemide (FU) as a model drug. Four types of FU preparations were prepared: granules with maltitol (MA), granules with yogurt powder (YO), a physical mixture of FU and MA, and a physical mixture of FU and YO. All taste-masking granules were prepared using the dry granulation method. The taste of each type of preparation was evaluated. All four preparations markedly improved the taste of the FU tablets, but the mixing ratios of the correctives did not affect the masking effect. No difference in masking effect was found between MA and YO in the physical mixtures, but the masking effect in the granules with YO was superior to that of the granules with MA. Taste-masked FU tablets were prepared using the direct compression method; crystalline cellulose (Avicel PH-302) and mannitol were added as excipients at the mixing ratio of 1/1. All four types of tablets displayed sufficient hardness, but MA-containing tablets were harder than YO-containing tablets. The hardness of the tablets prepared from YO granules increased as the YO content increased. The most rapidly disintegrating tablets were those of YO granules prepared at a mixing ratio of FU/YO=1/1, which disintegrated within 20 s, followed by the tablets of MA granules prepared at a mixing ratio of FU/MA=1/1. The disintegration times of the tablets made from physical mixtures, in contrast, were longer than 200 s. Disintegration time lengthened as the mixing ratio of YO or MA increased. The hardness and disintegration time of these tablets could be controlled by varying the compression pressure. We found that YO is more useful than MA in masking unpleasant tastes and confirmed that orally disintegrating tablets with taste-masking function can be prepared using granules of YO prepared using the dry granulation method as a new corrective.

Multi-species impurity granule injection and mass deposition projections in NSTX-U discharges

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lunsford, R.; Bortolon, A.; Roquemore, A. L.

Here, by employing a neutral gas shielding (NGS) model to characterize impurity granule injection, the ablation rates for three different species of granule: lithium, boron, and carbon, are determined. Utilizing the duration of ablation events recorded on experiments performed at DIII-D to calibrate the NGS model, we quantify the ablation rate with respect to the plasma density profile. The species-specific granule shielding constant is then used to model granule ablation within NSTX-U discharges. Simulations of 300, 500 and 700 micron diameter granules injected at 50 m s –1 are presented for NSTX-U L-mode type plasmas, as well as H-mode dischargesmore » with low natural ELM frequency. Additionally, ablation calculations of 500 micron granules of each species are presented at velocities ranging from 50–150 m s –1. In H-mode discharges these simulations show that the majority of the injected granule is ablated within or just past the edge steep gradient region. At this radial position, the perturbation to the background plasma generated by the ablating granule can lead to conditions advantageous for the rapid triggering of ELM crashes.« less

Multi-species impurity granule injection and mass deposition projections in NSTX-U discharges

DOE PAGES

Lunsford, R.; Bortolon, A.; Roquemore, A. L.; ...

2017-05-16

Here, by employing a neutral gas shielding (NGS) model to characterize impurity granule injection, the ablation rates for three different species of granule: lithium, boron, and carbon, are determined. Utilizing the duration of ablation events recorded on experiments performed at DIII-D to calibrate the NGS model, we quantify the ablation rate with respect to the plasma density profile. The species-specific granule shielding constant is then used to model granule ablation within NSTX-U discharges. Simulations of 300, 500 and 700 micron diameter granules injected at 50 m s –1 are presented for NSTX-U L-mode type plasmas, as well as H-mode dischargesmore » with low natural ELM frequency. Additionally, ablation calculations of 500 micron granules of each species are presented at velocities ranging from 50–150 m s –1. In H-mode discharges these simulations show that the majority of the injected granule is ablated within or just past the edge steep gradient region. At this radial position, the perturbation to the background plasma generated by the ablating granule can lead to conditions advantageous for the rapid triggering of ELM crashes.« less

Evaluating scale-up rules of a high-shear wet granulation process.

PubMed

Tao, Jing; Pandey, Preetanshu; Bindra, Dilbir S; Gao, Julia Z; Narang, Ajit S

2015-07-01

This work aimed to evaluate the commonly used scale-up rules for high-shear wet granulation process using a microcrystalline cellulose-lactose-based low drug loading formulation. Granule properties such as particle size, porosity, flow, and tabletability, and tablet dissolution were compared across scales using scale-up rules based on different impeller speed calculations or extended wet massing time. Constant tip speed rule was observed to produce slightly less granulated material at the larger scales. Longer wet massing time can be used to compensate for the lower shear experienced by the granules at the larger scales. Constant Froude number and constant empirical stress rules yielded granules that were more comparable across different scales in terms of compaction performance and tablet dissolution. Granule porosity was shown to correlate well with blend tabletability and tablet dissolution, indicating the importance of monitoring granule densification (porosity) during scale-up. It was shown that different routes can be chosen during scale-up to achieve comparable granule growth and densification by altering one of the three parameters: water amount, impeller speed, and wet massing time. © 2015 Wiley Periodicals, Inc. and the American Pharmacists Association.

Pharmaceutical production of tableting granules in an ultra-small-scale high-shear granulator as a pre-formulation study.

PubMed

Ogawa, Tatsuya; Uchino, Tomohiro; Takahashi, Daisuke; Izumi, Tsuyoshi; Otsuka, Makoto

2012-11-01

In some of drug developments, the amount of bulk drug powder to use in early stages is limited and it is not easy to supply a sufficient drug amount for conventional preparation methods. Therefore, an ultra-small-scale high-shear granulator (less than 5 g) (USG) was developed and applied to small-scale granulation as a pre-formulation. The sample powder consisted of 66.5% lactose, 28.5% microcrystalline cellulose and 5.0% hydroxypropylcellulose. The granules were obtained to agitate 5 g of the sample powder with 1.0 mL of water at 300 rpm for 5 min after pre-powder mixing for 3 min by the USG and the manual hand (HM) methods. The granules were evaluated by the 10% and 90% accumulated particle size and the recoveries of the granules and the powder solid. Median particle size for the USG and the HM methods was 159.2 ± 2.3 and 270.9 ± 14.9 µm, respectively. The USG method had a narrower particle size distribution than those by the HM method. The recovery of the granules by USG was significantly larger than that by the HM method. Characteristics of all of the granules indicated that the USG method could produce higher quality granules within a shorter time than the HM methods.

PtdIns(4,5)P2 is not required for secretory granule docking.

PubMed

Omar-Hmeadi, Muhmmad; Gandasi, Nikhil R; Barg, Sebastian

2018-06-01

Phosphoinositides (PtdIns) play important roles in exocytosis and are thought to regulate secretory granule docking by co-clustering with the SNARE protein syntaxin to form a docking receptor in the plasma membrane. Here we tested this idea by high-resolution total internal reflection imaging of EGFP-labeled PtdIns markers or syntaxin-1 at secretory granule release sites in live insulin-secreting cells. In intact cells, PtdIns markers distributed evenly across the plasma membrane with no preference for granule docking sites. In contrast, syntaxin-1 was found clustered in the plasma membrane, mostly beneath docked granules. We also observed rapid accumulation of syntaxin-1 at sites where granules arrived to dock. Acute depletion of plasma membrane phosphatidylinositol (4,5) bisphosphate (PtdIns(4,5)P 2 ) by recruitment of a 5'-phosphatase strongly inhibited Ca 2+ -dependent exocytosis, but had no effect on docked granules or the distribution and clustering of syntaxin-1. Cell permeabilization by α-toxin or formaldehyde-fixation caused PtdIns marker to slowly cluster, in part near docked granules. In summary, our data indicate that PtdIns(4,5)P 2 accelerates granule priming, but challenge a role of PtdIns in secretory granule docking or clustering of syntaxin-1 at the release site. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Three novel proteins co-localise with polyhydroxybutyrate (PHB) granules in Rhodospirillum rubrum S1.

PubMed

Narancic, Tanja; Scollica, Elisa; Cagney, Gerard; O'Connor, Kevin E

2018-04-01

Polyhydroxybutyrate (PHB), a biodegradable polymer accumulated by bacteria is deposited intracellularly in the form of inclusion bodies often called granules. The granules are supramolecular complexes harbouring a varied number of proteins on their surface, which have specific but incompletely characterised functions. By comparison with other organisms that produce biodegradable polymers, only two phasins have been described to date for Rhodosprillum rubrum, raising the possibility that more await discovery. Using a comparative proteomics strategy to compare the granules of wild-type R. rubrum with a PHB-negative mutant housing artificial PHB granules, we identified four potential PHB granules' associated proteins. These were: Q2RSI4, an uncharacterised protein; Q2RWU9, annotated as an extracellular solute-binding protein; Q2RQL4, annotated as basic membrane lipoprotein; and Q2RQ51, annotated as glucose-6-phosphate isomerase. In silico analysis revealed that Q2RSI4 harbours a Phasin_2 family domain and shares low identity with a single-strand DNA-binding protein from Sphaerochaeta coccoides. Fluorescence microscopy found that three proteins Q2RSI4, Q2EWU9 and Q2RQL4 co-localised with PHB granules. This work adds three potential new granule associated proteins to the repertoire of factors involved in bacterial storage granule formation, and confirms that proteomics screens are an effective strategy for discovery of novel granule associated proteins.

Deubiquitylases USP5 and USP13 are recruited to and regulate heat-induced stress granules through their deubiquitylating activities.

PubMed

Xie, Xuan; Matsumoto, Shunsuke; Endo, Akinori; Fukushima, Toshiaki; Kawahara, Hiroyuki; Saeki, Yasushi; Komada, Masayuki

2018-04-12

Stress granules are transient cytoplasmic foci induced by various stresses that contain translation-stalled mRNAs and RNA-binding proteins. They are proposed to modulate mRNA translation and stress responses. Here, we show that the deubiquitylases USP5 and USP13 are recruited to heat-induced stress granules. Heat-induced stress granules also contained K48- and K63-linked ubiquitin chains. Depletion of USP5 or USP13 resulted in elevated ubiquitin chain levels and accelerated assembly of heat-induced stress granules, suggesting that these enzymes regulate the stability of the stress granules through their ubiquitin isopeptidase activity. Moreover, disassembly of heat-induced stress granules after returning the cells to normal temperatures was markedly repressed by individual depletion of USP5 or USP13. Finally, overexpression of a ubiquitin mutant lacking the C-terminal diglycine motif caused the accumulation of unanchored ubiquitin chains and the repression of the disassembly of heat-induced stress granules. As unanchored ubiquitin chains are preferred substrates for USP5, we suggest that USP5 regulates the assembly and disassembly of heat-induced stress granules by mediating the hydrolysis of unanchored ubiquitin chains while USP13 regulates stress granules through deubiquitylating protein-conjugated ubiquitin chains.This article has an associated First Person interview with the first author of the paper. © 2018. Published by The Company of Biologists Ltd.

Role of the Caenorhabditis elegans multidrug resistance gene, mrp-4, in gut granule differentiation.

PubMed

Currie, Erin; King, Brian; Lawrenson, Andrea L; Schroeder, Lena K; Kershner, Aaron M; Hermann, Greg J

2007-11-01

Caenorhabditis elegans gut granules are lysosome-related organelles with birefringent contents. mrp-4, which encodes an ATP-binding cassette (ABC) transporter homologous to mammalian multidrug resistance proteins, functions in the formation of gut granule birefringence. mrp-4(-) embryos show a delayed appearance of birefringent material in the gut granule but otherwise appear to form gut granules properly. mrp-4(+) activity is required for the extracellular mislocalization of birefringent material, body-length retraction, and NaCl sensitivity, phenotypes associated with defective gut granule biogenesis exhibited by embryos lacking the activity of GLO-1/Rab38, a putative GLO-1 guanine nucleotide exchange factor GLO-4, and the AP-3 complex. Multidrug resistance protein (MRP)-4 localizes to the gut granule membrane, consistent with it playing a direct role in the transport of molecules that compose and/or facilitate the formation of birefringent crystals within the gut granule. However, MRP-4 is also present in oocytes and early embryos, and our genetic analyses indicate that its site of action in the formation of birefringent material may not be limited to just the gut granule in embryos. In a search for genes that function similarly to mrp-4(+), we identified WHT-2, another ABC transporter that acts in parallel to MRP-4 for the formation of birefringent material in the gut granule.

HIGH-SHEAR GRANULATION PROCESS: INFLUENCE OF PROCESSING PARAMETERS ON CRITICAL QUALITY ATTRIBUTES OF ACETAMINOPHEN GRANULES AND TABLETS USING DESIGN OF EXPERIMENT APPROACH.

PubMed

Fayed, Mohamed H; Abdel-Rahman, Sayed I; Alanazi, Fars K; Ahmed, Mahrous O; Tawfeek, Hesham M; Al-Shedfat, Ramadan I

2017-01-01

Application of quality by design (QbD) in high shear granulation process is critical and need to recognize the correlation between the granulation process parameters and the properties of intermediate (granules) and corresponding final product (tablets). The present work examined the influence of water amount (X,) and wet massing time (X2) as independent process variables on the critical quality attributes of granules and corresponding tablets using design of experiment (DoE) technique. A two factor, three level (32) full factorial design was performed; each of these variables was investigated at three levels to characterize their strength and interaction. The dried granules have been analyzed for their size distribution, density and flow pattern. Additionally, the produced tablets have been investigated for weight uniformity, crushing strength, friability and percent capping, disintegration time and drug dissolution. Statistically significant impact (p < 0.05) of water amount was identified for granule growth, percent fines and distribution width and flow behavior. Granule density and compressibility were found to be significantly influenced (p < 0.05) by the two operating conditions. Also, water amount has significant effect (p < 0.05) on tablet weight unifornity, friability and percent capping. Moreover, tablet disintegration time and drug dissolution appears to be significantly influenced (p < 0.05) by the two process variables. On the other hand, the relationship of process parameters with critical quality attributes of granule and final product tablet was identified and correlated. Ultimately, a judicious selection of process parameters in high shear granulation process will allow providing product of desirable quality.

Morphologic Integration of Hilar Ectopic Granule Cells into Dentate Gyrus Circuitry in the Pilocarpine Model of Temporal Lobe Epilepsy

PubMed Central

Cameron, Michael C.; Zhan, Ren-Zhi; Nadler, J. Victor

2014-01-01

After pilocarpine-induced status epilepticus, many granule cells born into the postseizure environment migrate aberrantly into the dentate hilus. Hilar ectopic granule cells (HEGCs) are hyperexcitable and may therefore increase circuit excitability. This study determined the distribution of their axons and dendrites. HEGCs and normotopic granule cells were filled with biocytin during whole-cell patch clamp recording in hippocampal slices from pilocarpine-treated rats. The apical dendrite of 86% of the biocytin-labeled HEGCs extended to the outer edge of the dentate molecular layer. The total length and branching of HEGC apical dendrites that penetrated the molecular layer were significantly reduced compared with apical dendrites of normotopic granule cells. HEGCs were much more likely to have a hilar basal dendrite than normotopic granule cells. They were about as likely as normotopic granule cells to project to CA3 pyramidal cells within the slice, but were much more likely to send at least one recurrent mossy fiber into the molecular layer. HEGCs with burst capability had less well-branched apical dendrites than nonbursting HEGCs, their dendrites were more likely to be confined to the hilus, and some exhibited dendritic features similar to those of immature granule cells. HEGCs thus have many paths along which to receive synchronized activity from normotopic granule cells and to transmit their own hyperactivity to both normotopic granule cells and CA3 pyramidal cells. They may therefore contribute to the highly interconnected granule cell hubs that have been proposed as crucial to development of a hyperexcitable, potentially seizure-prone circuit. PMID:21455997

High-shear granulation as a manufacturing method for cocrystal granules.

PubMed

Rehder, Sönke; Christensen, Niels Peter Aae; Rantanen, Jukka; Rades, Thomas; Leopold, Claudia S

2013-11-01

Cocrystal formation allows the tailoring of physicochemical as well as of mechanical properties of an API. However, there is a lack of large-scale manufacturing methods of cocrystals. Therefore, the objective of this work was to examine the suitability of high-shear wet granulation as a manufacturing method for cocrystal granules on a batch scale. Furthermore, the cocrystal granules were characterized regarding their mechanical properties as well as their dissolution behavior. High-shear wet granulation was found to be a feasible manufacturing method for cocrystal granules. Cocrystal formation depended on the exposure time of the solids to the granulation liquid (water), the amount of liquid, the impeller speed of the granulator, and on the excipients (hydroxyl propylcellulose, microcrystalline cellulose, calcium hydrogenphosphate) used in the formulation. Storage stability was strongly influenced by the excipients, since in presence of calcium hydrogenphosphate, the poorly water-soluble salt calcium tartrate monohydrate was formed at high relative humidity. Interestingly, compactability was increased by cocrystal formation compared to that of the reference granules (piracetam and the respective excipients). The drug release was slightly decreased by cocrystal formation, most likely due to the lower solubility of the cocrystal. In the presence of calcium hydrogenphosphate however, no influence of cocrystal formation on either compactability or on drug release were observed, compared with the reference tablets. It was concluded that high-shear wet granulation is a valuable, however complex, manufacturing method for cocrystals. Cocrystal formation may influence compactability and drug release and thus affect drug performance and should be investigated during pre-formulation. Copyright © 2013 Elsevier B.V. All rights reserved.

Distorted secretory granule composition in mast cells with multiple protease deficiency.

PubMed

Grujic, Mirjana; Calounova, Gabriela; Eriksson, Inger; Feyerabend, Thorsten; Rodewald, Hans-Reimer; Tchougounova, Elena; Kjellén, Lena; Pejler, Gunnar

2013-10-01

Mast cells are characterized by an abundance of secretory granules densely packed with inflammatory mediators such as bioactive amines, cytokines, serglycin proteoglycans with negatively charged glycosaminoglycan side chains of either heparin or chondroitin sulfate type, and large amounts of positively charged proteases. Despite the large biological impact of mast cell granules and their contents on various pathologies, the mechanisms that regulate granule composition are incompletely understood. In this study, we hypothesized that granule composition is dependent on a dynamic electrostatic interrelationship between different granule compounds. As a tool to evaluate this possibility, we generated mice in which mast cells are multideficient in a panel of positively charged proteases: the chymase mouse mast cell protease-4, the tryptase mouse mast cell protease-6, and carboxypeptidase A3. Through a posttranslational effect, mast cells from these mice additionally lack mouse mast cell protease-5 protein. Mast cells from mice deficient in individual proteases showed normal morphology. In contrast, mast cells with combined protease deficiency displayed a profound distortion of granule integrity, as seen both by conventional morphological criteria and by transmission electron microscopy. An assessment of granule content revealed that the distorted granule integrity in multiprotease-deficient mast cells was associated with a profound reduction of highly negatively charged heparin, whereas no reduction in chondroitin sulfate storage was observed. Taken together with previous findings showing that the storage of basic proteases conversely is regulated by anionic proteoglycans, these data suggest that secretory granule composition in mast cells is dependent on a dynamic interrelationship between granule compounds of opposite electrical charge.

Dextrose monohydrate as a non-animal sourced alternative diluent in high shear wet granulation tablet formulations.

PubMed

Mitra, Biplob; Wolfe, Chad; Wu, Sy-Juen

2018-05-01

The feasibility of dextrose monohydrate as a non-animal sourced diluent in high shear wet granulation (HSWG) tablet formulations was determined. Impacts of granulation solution amount and addition time, wet massing time, impeller speed, powder and solution binder, and dry milling speed and screen opening size on granule size, friability and density, and tablet solid fraction (SF) and tensile strength (TS) were evaluated. The stability of theophylline tablets TS, disintegration time (DT) and in vitro dissolution were also studied. Following post-granulation drying at 60 °C, dextrose monohydrate lost 9% water and converted into the anhydrate form. Higher granulation solution amounts and faster addition, faster impeller speeds, and solution binder produced larger, denser and stronger (less friable) granules. All granules were compressed into tablets with acceptable TS. Contrary to what is normally observed, denser and larger granules (at ≥21% water level) produced tablets with a higher TS. The TS of the weakest tablets increased the most after storage at both 25 °C/60% RH and 40 °C/75% RH. Tablet DT was higher for stronger granules and after storage. Tablet dissolution profiles for 21% or less water were comparable and did not change on stability. However, the dissolution profile for tablets prepared with 24% water was slower initially and continued to decrease on stability. The results indicate a granulation water amount of not more than 21% is required to achieve acceptable tablet properties. This study clearly demonstrated the utility of dextrose monohydrate as a non-animal sourced diluent in a HSWG tablet formulation.

Degradation of Glucan Primers in the Absence of Starch Synthase 4 Disrupts Starch Granule Initiation in Arabidopsis*

PubMed Central

Lu, Kuan-Jen; Stettler, Michaela; Streb, Sebastian

2016-01-01

Arabidopsis leaf chloroplasts typically contain five to seven semicrystalline starch granules. It is not understood how the synthesis of each granule is initiated or how starch granule number is determined within each chloroplast. An Arabidopsis mutant lacking the glucosyl-transferase, STARCH SYNTHASE 4 (SS4) is impaired in its ability to initiate starch granules; its chloroplasts rarely contain more than one large granule, and the plants have a pale appearance and reduced growth. Here we report that the chloroplastic α-amylase AMY3, a starch-degrading enzyme, interferes with granule initiation in the ss4 mutant background. The amy3 single mutant is similar in phenotype to the wild type under normal growth conditions, with comparable numbers of starch granules per chloroplast. Interestingly, the ss4 mutant displays a pleiotropic reduction in the activity of AMY3. Remarkably, complete abolition of AMY3 (in the amy3 ss4 double mutant) increases the number of starch granules produced in each chloroplast, suppresses the pale phenotype of ss4, and nearly restores normal growth. The amy3 mutation also restores starch synthesis in the ss3 ss4 double mutant, which lacks STARCH SYNTHASE 3 (SS3) in addition to SS4. The ss3 ss4 line is unable to initiate any starch granules and is thus starchless. We suggest that SS4 plays a key role in granule initiation, allowing it to proceed in a way that avoids premature degradation of primers by starch hydrolases, such as AMY3. PMID:27458017

Effects of Manufacturing Methods on Dissolution and Absorption of Ketoconazole in the Presence of Organic Acid as a pH Modifier.

PubMed

Adachi, Masashi; Hinatsu, Yuta; Kusamori, Kosuke; Katsumi, Hidemasa; Sakane, Toshiyasu; Nakatani, Manabu; Wada, Koichi; Yamamoto, Akira

2017-05-01

Poorly water-soluble compounds have a potential risk of low and variable bioavailability caused by incomplete dissolution. Incorporation of organic acids as pH modifiers is effective method for solubility enhancement of basic compounds and requires no special technique and equipment. The purpose of this study was to evaluate the effect of manufacturing method on the extent of drug solubility enhancement. We successfully prepared the granules and tablets containing ketoconazole (KZ), which is weakly basic, as a model compound and citric acid as a pH modifier using conventional wet and dry granulations. KZ solubility under non-sink condition was enhanced with supersaturation using both wet and dry granulations. High-shear granulation was the most effective method in terms of KZ dissolution enhancement, because both an intimate contact and strong bonding between KZ and incorporated acid were achieved. KZ dissolved amount from the granules prepared by high-shear granulation was about eight times higher than that from the granules without the acid. The granulation involved to suppress a diffusion of acid dissolved, leading to the effectively maintained supersaturation state. The bioavailability of KZ after oral administration to rats was improved by applying high-shear granulation with citric acid independent of gastrointestinal pH. The granules prepared by high-shear granulation showed the bioavailability about 1.7-fold higher than that of the physical mixture in rats with and without neutralization of stomach. As a result, both the dissolution and absorption rates of KZ after oral administration were enhanced using conventional manufacturing technology.

Two Distinct Waxy Alleles Impact the Granule-Bound Starch Synthase in Sorghum

USDA-ARS?s Scientific Manuscript database

The granule-bound starch synthase (GBSS) is the enzyme responsible for amylose synthesis in starch granules. Loss of GBSS activity results in starch granules containing mostly amylopectin and little or no amylose, a phenotype described as waxy. Previously, two phenotypic classes of waxy alleles we...

Tissue classification and segmentation of pressure injuries using convolutional neural networks.

PubMed

Zahia, Sofia; Sierra-Sosa, Daniel; Garcia-Zapirain, Begonya; Elmaghraby, Adel

2018-06-01

This paper presents a new approach for automatic tissue classification in pressure injuries. These wounds are localized skin damages which need frequent diagnosis and treatment. Therefore, a reliable and accurate systems for segmentation and tissue type identification are needed in order to achieve better treatment results. Our proposed system is based on a Convolutional Neural Network (CNN) devoted to performing optimized segmentation of the different tissue types present in pressure injuries (granulation, slough, and necrotic tissues). A preprocessing step removes the flash light and creates a set of 5x5 sub-images which are used as input for the CNN network. The network output will classify every sub-image of the validation set into one of the three classes studied. The metrics used to evaluate our approach show an overall average classification accuracy of 92.01%, an average total weighted Dice Similarity Coefficient of 91.38%, and an average precision per class of 97.31% for granulation tissue, 96.59% for necrotic tissue, and 77.90% for slough tissue. Our system has been proven to make recognition of complicated structures in biomedical images feasible. Copyright © 2018 Elsevier B.V. All rights reserved.

Haemoproteus syrnii in Strix aluco from France: morphology, stages of sporogony in a hippoboscid fly, molecular characterization and discussion on the identification of Haemoproteus species

PubMed Central

Karadjian, Grégory; Puech, Marie-Pierre; Duval, Linda; Chavatte, Jean-Marc; Snounou, Georges; Landau, Irène

2013-01-01

In France, Haemoproteus syrnii is frequently found in the Tawny Owl, Strix aluco. Additional and complementary features of this species, and in particular the characteristics of volutin, are presented. The authors consider the volutin granules as constant in a given species, and discuss their taxonomic value. These cytoplasmic inclusions appear early during the first stages of development of the gametocytes as an initial granule which multiplies as the parasite develops. They were reported in some species of Haemoproteus but are seldom considered as a specific character and described with precision. Sporogony from ookinete to apparently mature sporozoites appears to take place in a pupiparous hippoboscid (Ornithomyia sp.). One specimen was crushed between two slides and stained with Giemsa. Gametocytes of H. syrnii, many ookinetes, an immature oocyst and mature sporozoites were observed spread all over the smear. This would allow classifying this species in the Haemoproteus subgenus. We provide associated molecular data derived from the cyt b and cox 1 gene from this parasite. We discuss the problems of multiple infections and the difficulties in identifying Haemoproteus species and in deriving conclusions from sequences deposited in databases. PMID:24029169

DNA authentication of animal-derived concentrated Chinese medicine granules.

PubMed

Jiang, Li-Li; Lo, Yat-Tung; Chen, Wei-Ting; Shaw, Pang-Chui

2016-09-10

Concentrated Chinese medicine granules (CCMG) offer patients a convenient option for traditional therapy. However with morphological and microscopic characteristics lost, it is difficult to authenticate and control the quality of these medicinal products. This study is the first to examine the feasibility of using DNA techniques to authenticate animal-derived CCMG, which has so far lacking of effective means for authentication. Primers targeting amplicons of different sizes were designed to determine the presence of PCR-amplifiable DNA fragments in two types of CCMG, namely Zaocys and Scorpio. Species-specific primers were designed to differentiate the genuine drugs from their adulterants. The specificity of the designed primers was evaluated in crude drugs (including genuine and adulterant) and CCMG. Results showed that by using species-specific primers, DNA fragments of less than 200bp could be isolated from the CCMG and the concerned source materials. This study demonstrated the presence of small size DNA in animal-derived CCMG and the DNA is effective in species identification. The work has extended the application of DNA techniques in herbal medicine and this approach may be further developed for quality control and regulatory compliance in the CCMG industry. Copyright © 2016 Elsevier B.V. All rights reserved.

Hardened, environmentally disposable composite granules of coal cleaning refuse, coal combustion waste, and other wastes, and method preparing the same

DOEpatents

Burnet, George; Gokhale, Ashok J.

1990-07-10

A hardened, environmentally inert and disposable composite granule of coal cleaning refuse and coal combustion waste, and method for producing the same, wherein the coal combustion waste is first granulated. The coal cleaning refuse is pulverized into fine particles and is then bound, as an outer layer, to the granulated coal combustion waste granules. This combination is then combusted and sintered. After cooling, the combination results in hardened, environmentally inert and disposable composite granules having cores of coal combustion waste, and outer shells of coal cleaning refuse. The composite particles are durable and extremely resistant to environmental and chemical forces.

Hardened, environmentally disposable composite granules of coal cleaning refuse, coal combustion waste, and other wastes, and method preparing the same

DOEpatents

Burnet, G.; Gokhale, A.J.

1990-07-10

A hardened, environmentally inert and disposable composite granule of coal cleaning refuse and coal combustion waste and method for producing the same are disclosed, wherein the coal combustion waste is first granulated. The coal cleaning refuse is pulverized into fine particles and is then bound, as an outer layer, to the granulated coal combustion waste granules. This combination is then combusted and sintered. After cooling, the combination results in hardened, environmentally inert and disposable composite granules having cores of coal combustion waste, and outer shells of coal cleaning refuse. The composite particles are durable and extremely resistant to environmental and chemical forces. 3 figs.

Probing the spatial dependence of the emission spectrum of single human retinal lipofuscin granules using near-field scanning optical microscopy.

PubMed

Haralampus-Grynaviski, N M; Lamb, L E; Simon, J D; Krogmeier, J R; Dunn, R C; Pawlak, A; Rózanowska, M; Sarna, T; Burke, J M

2001-08-01

The emission spectra of single lipofuscin granules are examined using spectrally resolved confocal microscopy and near-field scanning optical microscopy (NSOM). The emission spectrum varies among the granules examined revealing that individual granules are characterized by different distributions of fluorophores. The range of spectra observed is consistent with in vivo spectra of human retinal pigment epithelium cells. NSOM measurements reveal that the shape of the spectrum does not vary with position within the emissive regions of single lipofuscin granules. These results suggest that the relative distribution of fluorophores within the emissive regions of an individual granule is homogeneous on the spatial scale approximately 150 nm.

Aerobic granulation in a modified oxidation ditch with an adjustable volume intraclarifier.

PubMed

Li, Jun; Cai, Ang; Wang, Miao; Ding, Libin; Ni, Yongjiong

2014-04-01

A modified oxidation ditch (MOD) with an adjustable volume intraclarifier was proposed and used to achieve aerobic sludge granulation in continuous flow process. This MOD with working volume of 60L treated onsite wastewater from a town. Excellent aerobic granules with mean diameter of 600μm and sludge volume index (SVI) of 44mL/g were obtained in 120day. Bacterial community analysis revealed that most species from seed sludge were preserved in both MOD and granule SBR (G-SBR) except bacteria (Bacteroidetes) might be easily washed out during granulation. Some different bacterial communities were found in sludges from sequencing batch and continuous flow reactors. Presence of metal ions and inorganics in raw wastewater had positive effect on granule formation, but an adjustable volume intraclarifier for controlling selection pressure and deleting return sludge pump played a key role in aerobic sludge granulation. Copyright © 2014 Elsevier Ltd. All rights reserved.

Preparation and evaluation of gelling granules to improve oral administration.

PubMed

Ito, Ikumi; Ito, Akihiko; Unezaki, Sakae

2015-06-01

We investigated the preparation of oral granules that are solid when stored and that will swell and gel via water absorption, to address problems experienced by patients when taking medication. Important physical properties of gelling granules include elasticity that is normally smooth, quick water absorption and swelling properties that allow easy swallowing. We selected gelatin (GEL), succinylated gelatin (SUC-GEL) and ι-carrageenan (CAR) as matrix polymers that can undergo gelation at room temperature or at cold temperatures. Saccharide and polyethylene glycol (PEG) were added to prepare the experimental granules. The best matrix gelling granule was SUC-GEL. When xylitol (XYL), sorbitol (SOR) and maltitol (MAL) were added, elasticity was improved, and PEG improved the granule's water absorption behavior, which is an important element involved in gelation. The best granules were prepared by selecting SUC-GEL as the matrix and adding a small amount of PEG and XYL in amounts equal to that of SUC-GEL.

Adsorption mechanism for xanthene dyes to cellulose granules.

PubMed

Tabara, Aya; Yamane, Chihiro; Seguchi, Masaharu

2012-01-01

The xanthene dyes, erythrosine, phloxine, and rose bengal, were adsorbed to charred cellulose granules. The charred cellulose granules were preliminarily steeped in ionic (NaOH, NaCl, KOH, KCl, and sodium dodecyl sulfate (SDS)), nonionic (glucose, sucrose, and ethanol), and amphipathic sucrose fatty acid ester (SFAE) solutions, and adsorption tests on the dye to the steeped and charred cellulose granules were conducted. Almost none of the dye was adsorbed when the solutions of ionic and amphipathic molecules were used, but were adsorbed in the case of steeping in the nonionic molecule solutions. Thin-layer chromatography (TLC) and the Fourier transform infra-red (FT-IR) profiles of SFAE which was adsorbed to the charred cellulose granules and extracted by ethyl ether suggested the presence of hydrophobic sites on the surface of the charred cellulose granules. We confirmed that the xanthene dyes could bind to the charred cellulose granules by ionic and hydrophobic bonds.

Microtubules negatively regulate insulin secretion in pancreatic β cells

PubMed Central

Zhu, Xiaodong; Hu, Ruiying; Brissova, Marcela; Stein, Roland W.; Powers, Alvin C.; Gu, Guoqiang; Kaverina, Irina

2015-01-01

Summary For glucose-stimulated insulin secretion (GSIS) insulin granules have to be localized close to the plasma membrane. The role of microtubule-dependent transport in granule positioning and GSIS has been debated. Here, we report that microtubules, counterintuitively, restrict granule availability for secretion. In β cells, microtubules originate at the Golgi and form a dense non-radial meshwork. Non-directional transport along these microtubules limits granule dwelling at the cell periphery, restricting granule availability for secretion. High glucose destabilizes microtubules, decreasing their density; such local microtubule depolymerization is necessary for GSIS, likely because granule withdrawal from the cell periphery becomes inefficient. Consistently, microtubule depolymerization by nocodazole blocks granule withdrawal, increases their concentration at exocytic sites, and dramatically enhances GSIS in vitro and in mice. Furthermore, glucose-driven MT destabilization is balanced by new microtubule formation, which likely prevents over-secretion. Importantly, microtubule density is greater in dysfunctional β cells of diabetic mice. PMID:26418295

Effects of artemisinin sustained-release granules on mixed alga growth and microcystins production and release.

PubMed

Ni, Lixiao; Li, Danye; Hu, Shuzhen; Wang, Peifang; Li, Shiyin; Li, Yiping; Li, Yong; Acharya, Kumud

2015-12-01

To safely and effectively apply artemisinin sustained-release granules to control and prevent algal water-blooms, the effects of artemisinin and its sustained-release granules on freshwater alga (Scenedesmus obliquus (S. obliquus) and Microcystis aeruginosa (M. aeruginosa)), as well as the production and release of microcystins (MCs) were studied. The results showed that artemisinin sustained-release granules inhibited the growth of M. aeruginosa (above 95% IR) and S. obliquus (about 90% IR), with M. aeruginosa more sensitive. The artemisinin sustained-release granules had a longer inhibition effect on growth of pure algae and algal coexistence than direct artemisinin dosing. The artemisinin sustained-release granules could decrease the production and release of algal toxins due to the continued stress of artemisinin released from artemisinin sustained-release granules. There was no increase in the total amount of MC-LR in the algal cell culture medium.

Low-calorie bread baked with charred cellulose granules and wheat flour to eliminate toxic xanthene food dye in the alimentary canal.

PubMed

Tabara, Aya; Yamane, Chihiro; Seguchi, Masaharu

2012-01-01

We baked low-calorie bread by mixing charred cellulose granules with wheat flour, using the charred cellulose granules to eliminate toxic xanthene food dyes contained in processed foods from the alimentary canal. The size of the charred cellulose granules played an important role in determining good breadmaking properties in respect of the bread height (mm) and specific volume (SV, cm3/g). Charred cellulose granules with a diameter above 270 μm were blended with wheat flour at 10% to obtain bread with a lower caloric content (1020 kcal/gram of bread) than the control bread (1126 kcal) made solely from wheat flour. The charred cellulose granules taken out from the bread adsorbed toxic xanthene food dyes at around pH 6.5, such that toxic food dyes taken into the alimentary canal were excreted in the feces with the non-digestible cellulose granules.

Specific lignin accumulation in granulated juice sacs of Citrus maxima.

PubMed

Wu, Jia-Ling; Pan, Teng-Fei; Guo, Zhi-Xiong; Pan, Dong-Ming

2014-12-17

Juice sac granulation occurring in pummelo fruits [Citrus maxima (Burm.) Merr.] is an undesirable trait, and the underlying mechanism remains unresolved. Previous studies have shown that lignin metabolism is closely associated with the process of juice sac granulation. Here, a method suitable for lignin isolation from pummelo tissues is established. Acetylated lignins from different pummelo tissues and cultivars were analyzed by HSQC NMR. The results showed that lignins in granulated juice sacs were characterized by an extremely high abundance of guaiacyl units (91.13-96.82%), in contrast to lignins from other tissues, including leaves, stems, and segment membranes. The abnormally accumulated lignins in granulated juice sacs were specific and mainly polymerized from coniferyl alcohol. No significant difference was found in lignin types among various cultivars. These findings indicated that the mechanism of juice sac granulation might be similar among various cultivars, although very different degrees of juice sac granulation can be observed.

Application of grey system theory on the influencing parameters of aerobic granulation in SBR.

PubMed

Bindhu, B K; Madhu, G

2017-09-01

Aerobic granulation is a promising technology for wastewater treatment. Four operational parameters were selected as influencing factors for this study. Aerobic granulation was experimented with three different values of organic loading rate (3, 6 and 9 kg COD m -3  d -1 ), superficial upflow air velocity (SUAV) (2, 3 and 4 cm s -1 ), settling time (3, 5 and 10 min) and volume exchange ratio (25%, 50% and 75%) in sequencing batch reactor in nine trials for the optimal performance of aerobic granulation. The influence of compared parameters on five reference parameters (sludge volume index (SVI), time taken for the appearance of granules, size and specific gravity of granules and chemical oxygen demand (COD) removal) was analyzed using grey system theory. The grey relational coefficients and grey entropy relational grade of each parameter were calculated. Hydrodynamic shear force in terms of SUAV was found to have the greatest influence on granule appearance, specific gravity of granules and COD removal efficiency. SVI is greatly affected by settling time. The optimal scopes of all the compared parameters were found.

Rapid aerobic granulation in an SBR treating piggery wastewater by seeding sludge from a municipal WWTP.

PubMed

Liu, Jun; Li, Jun; Wang, Xiaodong; Zhang, Qi; Littleton, Helen

2017-01-01

Aerobic sludge granulation was rapidly obtained in the erlenmeyer bottle and sequencing batch reactor (SBR) using piggery wastewater. Aerobic granulation occurred on day 3 and granules with mean diameter of 0.2mm and SVI 30 of 20.3mL/g formed in SBR on day 18. High concentrations of Ca and Fe in the raw piggery wastewater and operating mode accelerated aerobic granulation, even though the seed sludge was from a municipal wastewater treatment plant (WWTP). Alpha diversity analysis revealed Operational Taxonomic Units, Shannon, ACE and Chao 1 indexes in aerobic granules were 2013, 5.51, 4665.5 and 3734.5, which were obviously lower compared to seed sludge. The percentages of major microbial communities, such as Proteobacteria, Bacteroidetes and Firmicutes were obviously higher in aerobic granules than seed sludge. Chloroflexi, Planctomycetes, Actinobacteria, TM7 and Acidobacteria showed much higher abundances in the inoculum. The main reasons might be the characteristics of raw piggery wastewater and granule structure. Copyright © 2016. Published by Elsevier B.V.

Fabrication of dicalcium phosphate dihydrate-coated β-TCP granules and evaluation of their osteoconductivity using experimental rats.

PubMed

Shariff, Khairul Anuar; Tsuru, Kanji; Ishikawa, Kunio

2017-06-01

β-Tricalcium phosphate (β-TCP) has attracted much attention as an artificial bone substitute owing to its biocompatibility and osteoconductivity. In this study, osteoconductivity of β-TCP bone substitute was enhanced without using growth factors or cells. Dicalcium phosphate dihydrate (DCPD), which is known to possess the highest solubility among calcium phosphates, was coated on β-TCP granules by exposing their surface with acidic calcium phosphate solution. The amount of coated DCPD was regulated by changing the reaction time between β-TCP granules and acidic calcium phosphate solution. Histomorphometry analysis obtained from histological results revealed that the approximately 10mol% DCPD-coated β-TCP granules showed the largest new bone formation compared to DCPD-free β-TCP granules, approximately 2.5mol% DCPD-coated β-TCP granules, or approximately 27mol% DCPD-coated β-TCP granules after 2 and 4weeks of implantation. Based on this finding, we demonstrate that the osteoconductivity of β-TCP granules could be improved by coating their surface with an appropriate amount of DCPD. Copyright © 2017 Elsevier B.V. All rights reserved.

Lysine acetyltransferase NuA4 and acetyl-CoA regulate glucose-deprived stress granule formation in Saccharomyces cerevisiae

PubMed Central

Huard, Sylvain; Morettin, Alan; Fullerton, Morgan D.; Côté, Jocelyn

2017-01-01

Eukaryotic cells form stress granules under a variety of stresses, however the signaling pathways regulating their formation remain largely unknown. We have determined that the Saccharomyces cerevisiae lysine acetyltransferase complex NuA4 is required for stress granule formation upon glucose deprivation but not heat stress. Further, the Tip60 complex, the human homolog of the NuA4 complex, is required for stress granule formation in cancer cell lines. Surprisingly, the impact of NuA4 on glucose-deprived stress granule formation is partially mediated through regulation of acetyl-CoA levels, which are elevated in NuA4 mutants. While elevated acetyl-CoA levels suppress the formation of glucose-deprived stress granules, decreased acetyl-CoA levels enhance stress granule formation upon glucose deprivation. Further our work suggests that NuA4 regulates acetyl-CoA levels through the Acetyl-CoA carboxylase Acc1. Altogether this work establishes both NuA4 and the metabolite acetyl-CoA as critical signaling pathways regulating the formation of glucose-deprived stress granules. PMID:28231279

GABAergic excitation after febrile seizures induces ectopic granule cells and adult epilepsy.

PubMed

Koyama, Ryuta; Tao, Kentaro; Sasaki, Takuya; Ichikawa, Junya; Miyamoto, Daisuke; Muramatsu, Rieko; Matsuki, Norio; Ikegaya, Yuji

2012-08-01

Temporal lobe epilepsy (TLE) is accompanied by an abnormal location of granule cells in the dentate gyrus. Using a rat model of complex febrile seizures, which are thought to be a precipitating insult of TLE later in life, we report that aberrant migration of neonatal-generated granule cells results in granule cell ectopia that persists into adulthood. Febrile seizures induced an upregulation of GABA(A) receptors (GABA(A)-Rs) in neonatally generated granule cells, and hyperactivation of excitatory GABA(A)-Rs caused a reversal in the direction of granule cell migration. This abnormal migration was prevented by RNAi-mediated knockdown of the Na(+)K(+)2Cl(-) co-transporter (NKCC1), which regulates the excitatory action of GABA. NKCC1 inhibition with bumetanide after febrile seizures rescued the granule cell ectopia, susceptibility to limbic seizures and development of epilepsy. Thus, this work identifies a previously unknown pathogenic role of excitatory GABA(A)-R signaling and highlights NKCC1 as a potential therapeutic target for preventing granule cell ectopia and the development of epilepsy after febrile seizures.

Optimization for blast furnace slag dry cooling granulation device

NASA Astrophysics Data System (ADS)

Dazhan, Sheng; Yali, Wang; Ruiyun, Wang; Suping, Cui; Xiaoyu, Ma

2017-03-01

Since the large accumulation amount of blast furnace slag (BFS) with recycling value, it has become a hot topic for recovery utilization. Compared with the existing various BFS granulation process, the dry granulation process can promote the use of blast furnace granulated slag as cement substitute and concrete admixtures. Our research group developed a novel dry cooling granulation experiment device to treat BFS. However, there are still some problems to be solved. The purpose of this research is to improve the cooling and granulation efficiency of the existing dry type cooling equipment. This topic uses the FLUENT simulation software to study the impact of the number of air inlet on the cooling effect of the device. The simulation result is that the device possessing eight air inlets can increase the number of hot and cold gas exchanged, resulting in a better cooling effect. According to the power consumption, LCA analysis was carried out on the cooling granulation process. The results show that the device equipped eight air inlets not only improved the original equipment cooling granulation effect, but also increased resource utilization ratio, realized energy-saving and emission reduction.

Composition of the carbohydrate granules of the cyanobacterium, Cyanothece sp. strain ATCC 51142

NASA Technical Reports Server (NTRS)

Schneegurt, M. A.; Sherman, D. M.; Sherman, L. A.; Mitchell, C. A. (Principal Investigator)

1997-01-01

Cyanothece sp. strain ATCC 51142 is an aerobic, unicellular, diazotrophic cyanobacterium that temporally separates O2-sensitive N2 fixation from oxygenic photosynthesis. The energy and reducing power needed for N2 fixation appears to be generated by an active respiratory apparatus that utilizes the contents of large interthylakoidal carbohydrate granules. We report here on the carbohydrate and protein composition of the granules of Cyanothece sp. strain ATCC 51142. The carbohydrate component is a glucose homopolymer with branches every nine residues and is chemically identical to glycogen. Granule-associated protein fractions showed temporal changes in the number of proteins and their abundance during the metabolic oscillations observed under diazotrophic conditions. There also were temporal changes in the protein pattern of the granule-depleted supernatant fractions from diazotrophic cultures. None of the granule-associated proteins crossreacted with antisera directed against several glycogen-metabolizing enzymes or nitrogenase, although these proteins were tentatively identified in supernatant fractions. It is suggested that the granule-associated proteins are structural proteins required to maintain a complex granule architecture.

Bioaugmentation of half-matured granular sludge with special microbial culture promoted establishment of 2,4-dichlorophenoxyacetic acid degrading aerobic granules.

PubMed

Quan, Xiangchun; Ma, Jingyun; Xiong, Weicong; Wang, Xinrui

2015-06-01

Aerobic granular sludge degrading recalcitrant compounds are generally hard to be cultivated. This study investigated the feasibility of cultivating 2,4-dichlorophenoxyacetic acid (2,4-D) degrading aerobic granules using half-matured sludge granules pre-grown on glucose as the seeds and bioaugmentation with a 2,4-D degrading strain Achromobacter sp. QXH. Results showed that bioaugmentation promoted the steady transformation of glucose-grown granules to 2,4-D degrading sludge granules and fast establishment of 2,4-D degradation ability. The 2,4-D degradation rate of the bioaugmented granules was enhanced by 36-62 % compared to the control at 2,4-D concentrations of 144-565 mg/L on Day 18. The inoculated strain was incorporated into the half-matured granules successfully and survived till the end of operation (220 days). Sludge granules at a mean size of 420 µm and capable of utilizing 500 mg/L 2,4-D as the sole carbon source were finally obtained. Sludge microbial community shifted slightly during the whole operation and the dominant bacteria species belonged to Proteobacteria.

Glucokinase is an integral component of the insulin granules in glucose-responsive insulin secretory cells and does not translocate during glucose stimulation.

PubMed

Arden, Catherine; Harbottle, Andrew; Baltrusch, Simone; Tiedge, Markus; Agius, Loranne

2004-09-01

The association of glucokinase with insulin secretory granules has been shown by cell microscopy techniques. We used MIN6 insulin-secretory cells and organelle fractionation to determine the effects of glucose on the subcellular distribution of glucokinase. After permeabilization with digitonin, 50% of total glucokinase remained bound intracellularly, while 30% was associated with the 13,000g particulate fraction. After density gradient fractionation of the organelles, immunoreactive glucokinase was distributed approximately equally between dense insulin granules and low-density organelles that cofractionate with mitochondria. Although MIN6 cells show glucose-responsive insulin secretion, glucokinase association with the granules and low-density organelles was not affected by glucose. Subfractionation of the insulin granule components by hypotonic lysis followed by sucrose gradient centrifugation showed that glucokinase colocalized with the granule membrane marker phogrin and not with insulin. PFK2 (6-phosphofructo-2-kinase-2/fructose-2,6-bisphosphatase)/FDPase-2, a glucokinase-binding protein, and glyceraldehyde phosphate dehydrogenase, which has been implicated in granule fusion, also colocalized with glucokinase after hypotonic lysis or detergent extaction of the granules. The results suggest that glucokinase is an integral component of the granule and does not translocate during glucose stimulation.

Exocyst sec5 regulates exocytosis of newcomer insulin granules underlying biphasic insulin secretion.

PubMed

Xie, Li; Zhu, Dan; Kang, Youhou; Liang, Tao; He, Yu; Gaisano, Herbert Y

2013-01-01

The exocyst complex subunit Sec5 is a downstream effector of RalA-GTPase which promotes RalA-exocyst interactions and exocyst assembly, serving to tether secretory granules to docking sites on the plasma membrane. We recently reported that RalA regulates biphasic insulin secretion in pancreatic islet β cells in part by tethering insulin secretory granules to Ca(2+) channels to assist excitosome assembly. Here, we assessed β cell exocytosis by patch clamp membrane capacitance measurement and total internal reflection fluorescence microscopy to investigate the role of Sec5 in regulating insulin secretion. Sec5 is present in human and rodent islet β cells, localized to insulin granules. Sec5 protein depletion in rat INS-1 cells inhibited depolarization-induced release of primed insulin granules from both readily-releasable pool and mobilization from the reserve pool. This reduction in insulin exocytosis was attributed mainly to reduction in recruitment and exocytosis of newcomer insulin granules that undergo minimal docking time at the plasma membrane, but which encompassed a larger portion of biphasic glucose stimulated insulin secretion. Sec5 protein knockdown had little effect on predocked granules, unless vigorously stimulated by KCl depolarization. Taken together, newcomer insulin granules in β cells are more sensitive than predocked granules to Sec5 regulation.

IoT Big-Data Centred Knowledge Granule Analytic and Cluster Framework for BI Applications: A Case Base Analysis.

PubMed

Chang, Hsien-Tsung; Mishra, Nilamadhab; Lin, Chung-Chih

2015-01-01

The current rapid growth of Internet of Things (IoT) in various commercial and non-commercial sectors has led to the deposition of large-scale IoT data, of which the time-critical analytic and clustering of knowledge granules represent highly thought-provoking application possibilities. The objective of the present work is to inspect the structural analysis and clustering of complex knowledge granules in an IoT big-data environment. In this work, we propose a knowledge granule analytic and clustering (KGAC) framework that explores and assembles knowledge granules from IoT big-data arrays for a business intelligence (BI) application. Our work implements neuro-fuzzy analytic architecture rather than a standard fuzzified approach to discover the complex knowledge granules. Furthermore, we implement an enhanced knowledge granule clustering (e-KGC) mechanism that is more elastic than previous techniques when assembling the tactical and explicit complex knowledge granules from IoT big-data arrays. The analysis and discussion presented here show that the proposed framework and mechanism can be implemented to extract knowledge granules from an IoT big-data array in such a way as to present knowledge of strategic value to executives and enable knowledge users to perform further BI actions.

Granulation for Coking Wastewater Treatment in a Coupled Anaerobic-Aerobic Reactor

NASA Astrophysics Data System (ADS)

Dong, Chunjuan; Lv, Bingnan

2018-06-01

A coupled anaerobic-aerobic granular bio-film reactor was employed with two operation stages: Stage I, granular sludge was formed from digestion sludge using brewery wastewater, and Stage II, granular sludge was acclimatized using coking wastewater. Two oxygenation methods (i.e. A and B) were employed to acclimatize the granules. For method A, dissolved O 2 was supplied through a continuous oxygenation way of 800-15000ml-min-1 . And for method B, dissolved O2 was supplied of 800-15000ml-min-1 18-12 times at 20-60min intervals, 1h each time. The experimental results showed that granules could quickly form in 10d in the EGSB reactor seeded with digestion sludge and little loose granules lack of nutrition, and it was the key factor for granules forming to add little loose granules. It took only about 6 months for granules acclimation using coking wastewater. Both oxygenation methods could run well when acclimatizing the granules. However, method A could have comparatively high and stable operation effect. The actual coking wastewater had distinct inhibition effect on the granules, but the supplement of some oxygen could promote the recovery of SMA, and NaHCO3 supplement could also weaken the inhibition effect of the CWW. Method A had more strongly activity recovery ability than method B.

IoT Big-Data Centred Knowledge Granule Analytic and Cluster Framework for BI Applications: A Case Base Analysis

PubMed Central

Chang, Hsien-Tsung; Mishra, Nilamadhab; Lin, Chung-Chih

2015-01-01

The current rapid growth of Internet of Things (IoT) in various commercial and non-commercial sectors has led to the deposition of large-scale IoT data, of which the time-critical analytic and clustering of knowledge granules represent highly thought-provoking application possibilities. The objective of the present work is to inspect the structural analysis and clustering of complex knowledge granules in an IoT big-data environment. In this work, we propose a knowledge granule analytic and clustering (KGAC) framework that explores and assembles knowledge granules from IoT big-data arrays for a business intelligence (BI) application. Our work implements neuro-fuzzy analytic architecture rather than a standard fuzzified approach to discover the complex knowledge granules. Furthermore, we implement an enhanced knowledge granule clustering (e-KGC) mechanism that is more elastic than previous techniques when assembling the tactical and explicit complex knowledge granules from IoT big-data arrays. The analysis and discussion presented here show that the proposed framework and mechanism can be implemented to extract knowledge granules from an IoT big-data array in such a way as to present knowledge of strategic value to executives and enable knowledge users to perform further BI actions. PMID:26600156

Influence of in line monitored fluid bed granulation process parameters on the stability of Ethinylestradiol.

PubMed

Roßteuscher-Carl, Katrin; Fricke, Sabine; Hacker, Michael C; Schulz-Siegmund, Michaela

2015-12-30

Ethinylestradiol (EE) as a highly active and low dosed compound is prone to oxidative degradation. The stability of the drug substance is therefore a critical parameter that has to be considered during drug formulation. Beside the stability of the drug substance, granule particle size and moisture are critical quality attributes (CQA) of the fluid bed granulation process which influence the tableting ability of the resulting granules. Both CQA should therefore be monitored during the production process by process analytic technology (PAT) according to ICH Q8. This work focusses on the effects of drying conditions on the stability of EE in a fluid-bed granulation process. We quantified EE degradation products 6-alpha-hydroxy-EE, 6-beta-hydroxy-EE, 9(11)-dehydro-EE and 6-oxo-EE during long time storage and accelerated conditions. PAT-tools that monitor granule particle size (Spatial filtering technology) and granule moisture (Microwave resonance technology) were applied and compared with off-line methods. We found a relevant influence of residual granule moisture and thermic stress applied during granulation on the storage stability of EE, whereas no degradation was found immediately after processing. Hence we conclude that drying parameters have a relevant influence on long term EE stability. Copyright © 2015 Elsevier B.V. All rights reserved.

Experimental studies on the effect of moisture content and volume resistivity on electrostatic behaviour of pharmaceutical powders.

PubMed

Choi, Kwangseok; Taghavivand, Milad; Zhang, Lifeng

2017-03-15

Pharmaceutical powders are mainly organic materials and are likely to be charged due to repeated inter-particle and particle-wall contacts during industrial processes. This study experimentally investigated the effect of moisture content (ranging from approximately 1.8 to 30wt.%) on tribocharging behaviour of pharmaceutical granules, as well as their apparent volume resistivity. The tribocharging behaviour of pharmaceutical granules was investigated using a rotating device and apparent volume resistivity was measured in a conventional volume resistivity test cell. Additional measurements were performed on individual ingredients, each having the same moisture content as that of the granules, in order to investigate the effect of each single ingredient on the apparent volume resistivity of granules. In this work, the individual ingredients used for granules were: α-Lactose Monohydrate (α-LMH), Microcrystalline Cellulose (MCC), Hydroxypropyl Methylcellulose (HPMC), and Croscarmellose Sodium (CCS). The results showed that the specific charge of granules began to increase at the moisture contents below 5wt.%, which can be referred as critical moisture content of granules. The apparent volume resistivity showed the same behaviour, indicating that the specific charge could be due to an increase in apparent volume resistivity of granules at reduced moisture content. Finally, it was shown that the apparent volume resistivity measured for granules was mainly affected by that of the α-LMH, the major component of granules accounting for 40wt.%. Copyright © 2017 Elsevier B.V. All rights reserved.

Process Analytical Technology for High Shear Wet Granulation: Wet Mass Consistency Reported by In-Line Drag Flow Force Sensor Is Consistent With Powder Rheology Measured by At-Line FT4 Powder Rheometer.

PubMed

Narang, Ajit S; Sheverev, Valery; Freeman, Tim; Both, Douglas; Stepaniuk, Vadim; Delancy, Michael; Millington-Smith, Doug; Macias, Kevin; Subramanian, Ganeshkumar

2016-01-01

Drag flow force (DFF) sensor that measures the force exerted by wet mass in a granulator on a thin cylindrical probe was shown as a promising process analytical technology for real-time in-line high-resolution monitoring of wet mass consistency during high shear wet granulation. Our previous studies indicated that this process analytical technology tool could be correlated to granulation end point established independently through drug product critical quality attributes. In this study, the measurements of flow force by a DFF sensor, taken during wet granulation of 3 placebo formulations with different binder content, are compared with concurrent at line FT4 Powder Rheometer characterization of wet granules collected at different time points of the processing. The wet mass consistency measured by the DFF sensor correlated well with the granulation's resistance to flow and interparticulate interactions as measured by FT4 Powder Rheometer. This indicated that the force pulse magnitude measured by the DFF sensor was indicative of fundamental material properties (e.g., shear viscosity and granule size/density), as they were changing during the granulation process. These studies indicate that DFF sensor can be a valuable tool for wet granulation formulation and process development and scale up, as well as for routine monitoring and control during manufacturing. Copyright © 2016. Published by Elsevier Inc.

CSR-1 and P granules suppress sperm-specific transcription in the C. elegans germline

PubMed Central

Campbell, Anne C.; Updike, Dustin L.

2015-01-01

Germ granules (P granules) in C. elegans are required for fertility and function to maintain germ cell identity and pluripotency. Sterility in the absence of P granules is often accompanied by the misexpression of soma-specific proteins and the initiation of somatic differentiation in germ cells. To investigate whether this is caused by the accumulation of somatic transcripts, we performed mRNA-seq on dissected germlines with and without P granules. Strikingly, we found that somatic transcripts do not increase in the young adult germline when P granules are impaired. Instead, we found that impairing P granules causes sperm-specific mRNAs to become highly overexpressed. This includes the accumulation of major sperm protein (MSP) transcripts in germ cells, a phenotype that is suppressed by feminization of the germline. A core component of P granules, the endo-siRNA-binding Argonaute protein CSR-1, has recently been ascribed with the ability to license transcripts for germline expression. However, impairing CSR-1 has very little effect on the accumulation of its mRNA targets. Instead, we found that CSR-1 functions with P granules to prevent MSP and sperm-specific mRNAs from being transcribed in the hermaphrodite germline. These findings suggest that P granules protect germline integrity through two different mechanisms, by (1) preventing the inappropriate expression of somatic proteins at the level of translational regulation, and by (2) functioning with CSR-1 to limit the domain of sperm-specific expression at the level of transcription. PMID:25968310

ELLI-1, a novel germline protein, modulates RNAi activity and P-granule accumulation in Caenorhabditis elegans

PubMed Central

Andralojc, Karolina M.; Kelly, Ashley L.; Tanner, Paige C.

2017-01-01

Germ cells contain non-membrane bound cytoplasmic organelles that help maintain germline integrity. In C. elegans they are called P granules; without them, the germline undergoes partial masculinization and aberrant differentiation. One key P-granule component is the Argonaute CSR-1, a small-RNA binding protein that antagonizes accumulation of sperm-specific transcripts in developing oocytes and fine-tunes expression of proteins critical to early embryogenesis. Loss of CSR-1 complex components results in a very specific, enlarged P-granule phenotype. In a forward screen to identify mutants with abnormal P granules, ten alleles were recovered with a csr-1 P-granule phenotype, eight of which contain mutations in known components of the CSR-1 complex (csr-1, ego-1, ekl-1, and drh-3). The remaining two alleles are in a novel gene now called elli-1 (enlarged germline granules). ELLI-1 is first expressed in primordial germ cells during mid-embryogenesis, and continues to be expressed in the adult germline. While ELLI-1 forms cytoplasmic aggregates, they occasionally dock, but do not co-localize with P granules. Instead, the majority of ELLI-1 aggregates accumulate in the shared germline cytoplasm. In elli-1 mutants, several genes that promote RNAi and P-granule accumulation are upregulated, and embryonic lethality, sterility, and RNAi resistance in a hypomorphic drh-3 allele is enhanced, suggesting that ELLI-1 functions with CSR-1 to modulate RNAi activity, P-granule accumulation, and post-transcriptional expression in the germline. PMID:28182654

Ultrastructural and functional characterization of circulating hemocytes from the freshwater crayfish Astacus leptodactylus: cell types and their role after in vivo artificial non-self challenge.

PubMed

Giulianini, Piero Giulio; Bierti, Manuel; Lorenzon, Simonetta; Battistella, Silvia; Ferrero, Enrico Antonio

2007-01-01

The freshwater crayfish Astacus leptodactylus (Eschscholtz, 1823) is an important aquacultured decapod species as well as an invasive species in some European countries. In the current investigation we characterized the different classes of circulating blood cells in A. leptodactylus by means of light and electron microscopy analysis and we explored their reaction to different latex beads particles in vivo by total and differential cell counts at 0.5, 1, 2 and 4h after injections. We identified hemocytes by granule size morphometry as hyaline hemocytes with no or rare tiny granules, small granule hemocytes, unimodal medium diameter granule hemocytes and both small and large granule containing hemocytes. The latter granular hemocytes showed the strongest phenoloxidase l-DOPA reactivity both in granules and cytoplasm. A. leptodactylus respond to foreign particles with strong cellular immune responses. All treatments elicited a total hemocyte increase with a conspicuous recruitment of large granule containing hemocytes. All hemocyte types mounted some phagocytic response but the small granule hemocytes were the only ones involved in phagocytic response to all foreign particles with the highest percentages. These results (1) depict the variability in decapod hemocyte functional morphology; (2) identify the small granule hemocyte as the major phagocytic cell; (3) suggest that the rather rapid recruitment of large granule hemocyte in all treatments plays a relevant role by this hemocyte type in defense against foreign particles, probably in nodule formation.

Reduced starch granule number per chloroplast in the dpe2/phs1 mutant is dependent on initiation of starch degradation

PubMed Central

Malinova, Irina

2017-01-01

An Arabidopsis double knock-out mutant lacking cytosolic disproportionating enzyme 2 (DPE2) and the plastidial phosphorylase (PHS1) revealed a dwarf-growth phenotype, reduced starch content, an uneven distribution of starch within the plant rosette, and a reduced number of starch granules per chloroplast under standard growth conditions. In contrast, the wild type contained 5–7 starch granules per chloroplast. Mature and old leaves of the double mutant were essentially starch free and showed plastidial disintegration. Several analyses revealed that the number of starch granules per chloroplast was affected by the dark phase. So far, it was unclear if it was the dark phase per se or starch degradation in the dark that was connected to the observed decrease in the number of starch granules per chloroplast. Therefore, in the background of the double mutant dpe2/phs1, a triple mutant was generated lacking the initial starch degrading enzyme glucan, water dikinase (GWD). The triple mutant showed improved plant growth, a starch-excess phenotype, and a homogeneous starch distribution. Furthermore, the number of starch granules per chloroplast was increased and was similar to wild type. However, starch granule morphology was only slightly affected by the lack of GWD as in the triple mutant and, like in dpe2/phs1, more spherical starch granules were observed. The characterized triple mutant was discussed in the context of the generation of starch granules and the formation of starch granule morphology. PMID:29155859

Reduced starch granule number per chloroplast in the dpe2/phs1 mutant is dependent on initiation of starch degradation.

PubMed

Malinova, Irina; Fettke, Joerg

2017-01-01

An Arabidopsis double knock-out mutant lacking cytosolic disproportionating enzyme 2 (DPE2) and the plastidial phosphorylase (PHS1) revealed a dwarf-growth phenotype, reduced starch content, an uneven distribution of starch within the plant rosette, and a reduced number of starch granules per chloroplast under standard growth conditions. In contrast, the wild type contained 5-7 starch granules per chloroplast. Mature and old leaves of the double mutant were essentially starch free and showed plastidial disintegration. Several analyses revealed that the number of starch granules per chloroplast was affected by the dark phase. So far, it was unclear if it was the dark phase per se or starch degradation in the dark that was connected to the observed decrease in the number of starch granules per chloroplast. Therefore, in the background of the double mutant dpe2/phs1, a triple mutant was generated lacking the initial starch degrading enzyme glucan, water dikinase (GWD). The triple mutant showed improved plant growth, a starch-excess phenotype, and a homogeneous starch distribution. Furthermore, the number of starch granules per chloroplast was increased and was similar to wild type. However, starch granule morphology was only slightly affected by the lack of GWD as in the triple mutant and, like in dpe2/phs1, more spherical starch granules were observed. The characterized triple mutant was discussed in the context of the generation of starch granules and the formation of starch granule morphology.

Melt Adsorption as a Manufacturing Method for Fine Particles of Wax Matrices without Any Agglomerates.

PubMed

Shiino, Kai; Fujinami, Yukari; Kimura, Shin-Ichiro; Iwao, Yasunori; Noguchi, Shuji; Itai, Shigeru

2017-01-01

We have focused on melt adsorption as manufacture method of wax matrices to control particles size of granules more easily than melt granulation. The purpose of present study was to investigate the possibility of identifying a hydrophobic material with a low melting point, currently used as a meltable binder of melt granulation, to apply as a novel carrier in melt adsorption. Glyceryl monostearate (GM) and stearic acid (SA) were selected as candidate hydrophobic materials with low melting points. Neusilin US2 (US2), with a particle diameter of around 100 µm was selected as a surface adsorbent, while dibasic calcium phosphate dihydrate (DCPD), was used as a non-adsorbent control to prepare melting granules as a standard for comparison. We prepared granules containing ibuprofen (IBU) by melt adsorption or melt granulation and evaluated the particle size, physical properties and crystallinity of granules. Compared with melt granulation using DCPD, melt adsorption can be performed over a wide range of 14 to 70% for the ratio of molten components. Moreover, the particle size; d50 of obtained granules was 100-200 µm, and these physical properties showed good flowability and roundness. The process of melt adsorption did not affect the crystalline form of IBU. Therefore, the present study has demonstrated for the first time that melt adsorption using a hydrophobic material, GM or SA, has the potential capability to control the particle size of granules and offers the possibility of application as a novel controlled release technique.

Continuous melt granulation: Influence of process and formulation parameters upon granule and tablet properties.

PubMed

Monteyne, Tinne; Vancoillie, Jochem; Remon, Jean-Paul; Vervaet, Chris; De Beer, Thomas

2016-10-01

The pharmaceutical industry has a growing interest in alternative manufacturing models allowing automation and continuous production in order to improve process efficiency and reduce costs. Implementing a switch from batch to continuous processing requires fundamental process understanding and the implementation of quality-by-design (QbD) principles. The aim of this study was to examine the relationship between formulation-parameters (type binder, binder concentration, drug-binder miscibility), process-parameters (screw speed, powder feed rate and granulation temperature), granule properties (size, size distribution, shape, friability, true density, flowability) and tablet properties (tensile strength, friability, dissolution rate) of four different drug-binder formulations using Design of experiments (DOE). Two binders (polyethylene glycol (PEG) and Soluplus®) with a different solid state, semi-crystalline vs amorphous respectively, were combined with two model-drugs, metoprolol tartrate (MPT) and caffeine anhydrous (CAF), both having a contrasting miscibility with the binders. This research revealed that the granule properties of miscible drug-binder systems depended on the powder feed rate and barrel filling degree of the granulator whereas the granule properties of immiscible systems were mainly influenced by binder concentration. Using an amorphous binder, the tablet tensile strength depended on the granule size. In contrast, granule friability was more important for tablet quality using a brittle binder. However, this was not the case for caffeine-containing blends, since these phenomena were dominated by the enhanced compression properties of caffeine Form I, which was formed during granulation. Hence, it is important to gain knowledge about formulation behavior during processing since this influences the effect of process parameters onto the granule and tablet properties. Copyright © 2016 Elsevier B.V. All rights reserved.

A Role for Glutamate Transporters in the Regulation of Insulin Secretion

PubMed Central

Gammelsaeter, Runhild; Coppola, Thierry; Marcaggi, Païkan; Storm-Mathisen, Jon; Chaudhry, Farrukh A.; Attwell, David; Regazzi, Romano; Gundersen, Vidar

2011-01-01

In the brain, glutamate is an extracellular transmitter that mediates cell-to-cell communication. Prior to synaptic release it is pumped into vesicles by vesicular glutamate transporters (VGLUTs). To inactivate glutamate receptor responses after release, glutamate is taken up into glial cells or neurons by excitatory amino acid transporters (EAATs). In the pancreatic islets of Langerhans, glutamate is proposed to act as an intracellular messenger, regulating insulin secretion from β-cells, but the mechanisms involved are unknown. By immunogold cytochemistry we show that insulin containing secretory granules express VGLUT3. Despite the fact that they have a VGLUT, the levels of glutamate in these granules are low, indicating the presence of a protein that can transport glutamate out of the granules. Surprisingly, in β-cells the glutamate transporter EAAT2 is located, not in the plasma membrane as it is in brain cells, but exclusively in insulin-containing secretory granules, together with VGLUT3. In EAAT2 knock out mice, the content of glutamate in secretory granules is higher than in wild type mice. These data imply a glutamate cycle in which glutamate is carried into the granules by VGLUT3 and carried out by EAAT2. Perturbing this cycle by knocking down EAAT2 expression with a small interfering RNA, or by over-expressing EAAT2 or a VGLUT in insulin granules, significantly reduced the rate of granule exocytosis. Simulations of granule energetics suggest that VGLUT3 and EAAT2 may regulate the pH and membrane potential of the granules and thereby regulate insulin secretion. These data suggest that insulin secretion from β-cells is modulated by the flux of glutamate through the secretory granules. PMID:21853059

New Starch Phenotypes Produced by TILLING in Barley

PubMed Central

Sparla, Francesca; Falini, Giuseppe; Botticella, Ermelinda; Pirone, Claudia; Talamè, Valentina; Bovina, Riccardo; Salvi, Silvio; Tuberosa, Roberto; Sestili, Francesco; Trost, Paolo

2014-01-01

Barley grain starch is formed by amylose and amylopectin in a 1∶3 ratio, and is packed into granules of different dimensions. The distribution of granule dimension is bimodal, with a majority of small spherical B-granules and a smaller amount of large discoidal A-granules containing the majority of the starch. Starch granules are semi-crystalline structures with characteristic X-ray diffraction patterns. Distinct features of starch granules are controlled by different enzymes and are relevant for nutritional value or industrial applications. Here, the Targeting-Induced Local Lesions IN Genomes (TILLING) approach was applied on the barley TILLMore TILLING population to identify 29 new alleles in five genes related to starch metabolism known to be expressed in the endosperm during grain filling: BMY1 (Beta-amylase 1), GBSSI (Granule Bound Starch Synthase I), LDA1 (Limit Dextrinase 1), SSI (Starch Synthase I), SSIIa (Starch Synthase IIa). Reserve starch of nine M3 mutant lines carrying missense or nonsense mutations was analysed for granule size, crystallinity and amylose/amylopectin content. Seven mutant lines presented starches with different features in respect to the wild-type: (i) a mutant line with a missense mutation in GBSSI showed a 4-fold reduced amylose/amylopectin ratio; (ii) a missense mutations in SSI resulted in 2-fold increase in A:B granule ratio; (iii) a nonsense mutation in SSIIa was associated with shrunken seeds with a 2-fold increased amylose/amylopectin ratio and different type of crystal packing in the granule; (iv) the remaining four missense mutations suggested a role of LDA1 in granule initiation, and of SSIIa in determining the size of A-granules. We demonstrate the feasibility of the TILLING approach to identify new alleles in genes related to starch metabolism in barley. Based on their novel physicochemical properties, some of the identified new mutations may have nutritional and/or industrial applications. PMID:25271438

Downstream processing from melt granulation towards tablets: In-depth analysis of a continuous twin-screw melt granulation process using polymeric binders.

PubMed

Grymonpré, W; Verstraete, G; Vanhoorne, V; Remon, J P; De Beer, T; Vervaet, C

2018-03-01

The concept of twin-screw melt granulation (TSMG) has steadily (re)-gained interest in pharmaceutical formulation development as an intermediate step during tablet manufacturing. However, to be considered as a viable processing option for solid oral dosage forms there is a need to understand all critical sources of variability which could affect this granulation technique. The purpose of this study was to provide an in-depth analysis of the continuous TSMG process in order to expose the critical process parameters (CPP) and elucidate the impact of process and formulation parameters on the critical quality attributes (CQA) of granules and tablets during continuous TSMG. A first part of the study dealt with the screening of various amorphous polymers as binder for producing high-dosed melt granules of two model drug (i.e. acetaminophen and hydrochlorothiazide). The second part of this study described a quality-by-design (QbD) approach for melt granulation of hydrochlorothiazide in order to thoroughly evaluate TSMG, milling and tableting stage of the continuous TSMG line. Using amorphous polymeric binders resulted in melt granules with high milling efficiency due to their brittle behaviour without producing excessive amounts of fines, providing high granule yields with low friability. Therefore, it makes them extremely suitable for further downstream processing. One of the most important CPP during TSMG with polymeric binders was the granulation-torque, which - in case of polymers with high T g - increased during longer granulation runs to critical levels endangering the continuous process flow. However, by optimizing both screw speed and throughput or changing to polymeric binders with lower T g it was possible to significantly reduce this risk. This research paper highlighted that TSMG must be considered as a viable option during formulation development of solid oral dosage forms based on the robustness of the CQA of both melt granules and tablets. Copyright © 2017 Elsevier B.V. All rights reserved.

NEW METHODOLOGY FOR DEVELOPMENT OF ORODISPERSIBLE TABLETS USING HIGH-SHEAR GRANULATION PROCESS.

PubMed

Ali, Bahaa E; Al-Shedfat, Ramadan I; Fayed, Mohamed H; Alanazi, Fars K

2017-05-01

Development of orodispersible delivery system of high mechanical properties and low disintegration time is a big challenge. The aim of the current work was to assess and optimize the high shear granulation process as a new methodology for development of orodispersible tablets of high quality attributes using design of experiment approach. A two factor, three levels (32), full factorial design was carried out to investigate the main and interaction effects of independent variables, water amount (XI) and granulation time (X2) on the characteristics of granules and final product, tablet. The produced granules were analyzed for their granule size, density and flowability. Furthermore, the produced tablets were tested for: weight variation, breaking force/ crushing strength, friability, disintegration time and drug dissolution. Regression analysis results of multiple linear models showed a high correlation between the adjusted R-squared and predicted R-squared for all granules and tablets characteristics, the difference is less than 0.2. All dependent responses of granules and tablets were found to be impacted significantly (p < 0.05) by the two independent variables. However, water amount demonstrated the most dominant effect for all granules and tablet characteristics as shown by higher its coefficient estimate for all selected responses. Numerical optimization using desirability function was performed to optimize the variables under study to provide orodispersible system within the USP limit with respect of mechanical properties and disintegration time. It was found that the higher desirability (0.915) could be attained at the low level pf water (180 g) and short granulation time (1.65 min). Eventually, this study provides the formulator with helpful information in selecting the proper level of water and granulation time to provide an orodispersible system of high crushing strength and very low disintegration time, when high shear granulation methodology was used as a method of manufacture.

New starch phenotypes produced by TILLING in barley.

PubMed

Sparla, Francesca; Falini, Giuseppe; Botticella, Ermelinda; Pirone, Claudia; Talamè, Valentina; Bovina, Riccardo; Salvi, Silvio; Tuberosa, Roberto; Sestili, Francesco; Trost, Paolo

2014-01-01

Barley grain starch is formed by amylose and amylopectin in a 1:3 ratio, and is packed into granules of different dimensions. The distribution of granule dimension is bimodal, with a majority of small spherical B-granules and a smaller amount of large discoidal A-granules containing the majority of the starch. Starch granules are semi-crystalline structures with characteristic X-ray diffraction patterns. Distinct features of starch granules are controlled by different enzymes and are relevant for nutritional value or industrial applications. Here, the Targeting-Induced Local Lesions IN Genomes (TILLING) approach was applied on the barley TILLMore TILLING population to identify 29 new alleles in five genes related to starch metabolism known to be expressed in the endosperm during grain filling: BMY1 (Beta-amylase 1), GBSSI (Granule Bound Starch Synthase I), LDA1 (Limit Dextrinase 1), SSI (Starch Synthase I), SSIIa (Starch Synthase IIa). Reserve starch of nine M3 mutant lines carrying missense or nonsense mutations was analysed for granule size, crystallinity and amylose/amylopectin content. Seven mutant lines presented starches with different features in respect to the wild-type: (i) a mutant line with a missense mutation in GBSSI showed a 4-fold reduced amylose/amylopectin ratio; (ii) a missense mutations in SSI resulted in 2-fold increase in A:B granule ratio; (iii) a nonsense mutation in SSIIa was associated with shrunken seeds with a 2-fold increased amylose/amylopectin ratio and different type of crystal packing in the granule; (iv) the remaining four missense mutations suggested a role of LDA1 in granule initiation, and of SSIIa in determining the size of A-granules. We demonstrate the feasibility of the TILLING approach to identify new alleles in genes related to starch metabolism in barley. Based on their novel physicochemical properties, some of the identified new mutations may have nutritional and/or industrial applications.

Bacterial Selection during the Formation of Early-Stage Aerobic Granules in Wastewater Treatment Systems Operated Under Wash-Out Dynamics

PubMed Central

Weissbrodt, David G.; Lochmatter, Samuel; Ebrahimi, Sirous; Rossi, Pierre; Maillard, Julien; Holliger, Christof

2012-01-01

Aerobic granular sludge is attractive for high-rate biological wastewater treatment. Biomass wash-out conditions stimulate the formation of aerobic granules. Deteriorated performances in biomass settling and nutrient removal during start-up have however often been reported. The effect of wash-out dynamics was investigated on bacterial selection, biomass settling behavior, and metabolic activities during the formation of early-stage granules from activated sludge of two wastewater treatment plants (WWTP) over start-up periods of maximum 60 days. Five bubble-column sequencing batch reactors were operated with feast-famine regimes consisting of rapid pulse or slow anaerobic feeding followed by aerobic starvation. Slow-settling fluffy granules were formed when an insufficient superficial air velocity (SAV; 1.8 cm s−1) was applied, when the inoculation sludge was taken from a WWTP removing organic matter only, or when reactors were operated at 30°C. Fast-settling dense granules were obtained with 4.0 cm s−1 SAV, or when the inoculation sludge was taken from a WWTP removing all nutrients biologically. However, only carbon was aerobically removed during start-up. Fluffy granules and dense granules were displaying distinct predominant phylotypes, namely filamentous Burkholderiales affiliates and Zoogloea relatives, respectively. The latter were predominant in dense granules independently from the feeding regime. A combination of insufficient solid retention time and of leakage of acetate into the aeration phase during intensive biomass wash-out was the cause for the proliferation of Zoogloea spp. in dense granules, and for the deterioration of BNR performances. It is however not certain that Zoogloea-like organisms are essential in granule formation. Optimal operation conditions should be elucidated for maintaining a balance between organisms with granulation propensity and nutrient removing organisms in order to form granules with BNR activities in short start-up periods. PMID:22993513

Bacterial Selection during the Formation of Early-Stage Aerobic Granules in Wastewater Treatment Systems Operated Under Wash-Out Dynamics.

PubMed

Weissbrodt, David G; Lochmatter, Samuel; Ebrahimi, Sirous; Rossi, Pierre; Maillard, Julien; Holliger, Christof

2012-01-01

Aerobic granular sludge is attractive for high-rate biological wastewater treatment. Biomass wash-out conditions stimulate the formation of aerobic granules. Deteriorated performances in biomass settling and nutrient removal during start-up have however often been reported. The effect of wash-out dynamics was investigated on bacterial selection, biomass settling behavior, and metabolic activities during the formation of early-stage granules from activated sludge of two wastewater treatment plants (WWTP) over start-up periods of maximum 60 days. Five bubble-column sequencing batch reactors were operated with feast-famine regimes consisting of rapid pulse or slow anaerobic feeding followed by aerobic starvation. Slow-settling fluffy granules were formed when an insufficient superficial air velocity (SAV; 1.8 cm s(-1)) was applied, when the inoculation sludge was taken from a WWTP removing organic matter only, or when reactors were operated at 30°C. Fast-settling dense granules were obtained with 4.0 cm s(-1) SAV, or when the inoculation sludge was taken from a WWTP removing all nutrients biologically. However, only carbon was aerobically removed during start-up. Fluffy granules and dense granules were displaying distinct predominant phylotypes, namely filamentous Burkholderiales affiliates and Zoogloea relatives, respectively. The latter were predominant in dense granules independently from the feeding regime. A combination of insufficient solid retention time and of leakage of acetate into the aeration phase during intensive biomass wash-out was the cause for the proliferation of Zoogloea spp. in dense granules, and for the deterioration of BNR performances. It is however not certain that Zoogloea-like organisms are essential in granule formation. Optimal operation conditions should be elucidated for maintaining a balance between organisms with granulation propensity and nutrient removing organisms in order to form granules with BNR activities in short start-up periods.

Plastidial Disproportionating Enzyme Participates in Starch Synthesis in Rice Endosperm by Transferring Maltooligosyl Groups from Amylose and Amylopectin to Amylopectin1

PubMed Central

Dong, Xiangbai; Zhang, Du; Liu, Jie; Liu, Hualiang; Tian, Lihong; Jiang, Ling

2015-01-01

Plastidial disproportionating enzyme1 (DPE1), an α-1,4-d-glucanotransferase, has been thought to be involved in storage starch synthesis in cereal crops. However, the precise function of DPE1 remains to be established. We present here the functional identification of DPE1 in storage starch synthesis in rice (Oryza sativa) by endosperm-specific gene overexpression and suppression. DPE1 overexpression decreased amylose content and resulted in small and tightly packed starch granules, whereas DPE1 suppression increased amylose content and formed heterogeneous-sized, spherical, and loosely packed starch granules. Chains with degree of polymerization (DP) of 6 to 10 and 23 to 38 were increased, while chains with DP of 11 to 22 were decreased in amylopectin from DPE1-overexpressing seeds. By contrast, chains with DP of 6 to 8 and 16 to 36 were decreased, while chains with DP of 9 to 15 were increased in amylopectin from DPE1-suppressed seeds. Changes in DPE1 gene expression also resulted in modifications in the thermal and pasting features of endosperm starch granules. In vitro analyses revealed that recombinant DPE1 can break down amylose into maltooligosaccharides in the presence of Glc, while it can transfer maltooligosyl groups from maltooligosaccharide to amylopectin or transfer maltooligosyl groups within and among amylopectin molecules in the absence of Glc. Moreover, a metabolic flow of maltooligosyl groups from amylose to amylopectin was clearly identifiable when comparing DPE1-overexpressing lines with DPE1-suppressed lines. These findings demonstrate that DPE1 participates substantially in starch synthesis in rice endosperm by transferring maltooligosyl groups from amylose and amylopectin to amylopectin. PMID:26471894

Effect of formulation and process variables on lipid based sustained release tablets via continuous twin screw granulation: A comparative study.

PubMed

Kallakunta, Venkata Raman; Tiwari, Roshan; Sarabu, Sandeep; Bandari, Suresh; Repka, Michael A

2018-05-14

The current study's aim is to prepare lipid based sustained release tablets via a twin-screw granulation technique and compare those dosage forms with conventional techniques, namely wet granulation and direct compression. The granules were successfully manufactured in a single-step, continuous twin-screw granulation process with a low proportion of binder (Klucel™ EF, HPC SSL) using Compritol® 888 ATO, Precirol® ATO 5 and Geleol™ as sustained release agents. The granules prepared showed good flow characteristics and compaction properties. DSC and XRD studies were conducted to characterize the granules prepared via a twin-screw granulation method and the results demonstrated the crystalline nature of lipids within the granules. FTIR data indicated that there were no interactions with the formulation components investigated. The formulations developed by all three methods were compressed into tablets with a mechanical strength of 14-16 KP. The tablets formulated were characterized for physicochemical properties, in vitro drug release studies, water uptake and erosion studies. These results showed that the drug was not completely released after 24 h for tablets developed by the wet granulation process using all three lipids. The tablets prepared by the direct compression method demonstrated a burst release within 8 to 10 h from Precirol ATO 5® and Geleol™ formulations compared to Compritol® 888 ATO. However, tablets prepared using twin-screw granulation exhibited sustained release of the drug over 24 h and the water uptake and erosion results were in accordance with dissolution data. Stability data for 45 days at accelerated conditions (40 °C/75% RH) showed similar release profiles with ƒ2 values above 50 for all of the twin screw granulation formulations, indicating the suitability of the process for formulating sustained release tablets. These findings of a single-step, continuous twin-screw granulation process are novel and demonstrate new opportunities for development of sustained release tablets. Copyright © 2017. Published by Elsevier B.V.

DEVELOPMENT OF A FABRICATION PROCESS FOR SOL-GEL/METAL HYDRIDE COMPOSITE GRANULES

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hansen, E; Eric Frickey, E; Leung Heung, L

An external gelation process was developed to produce spherical granules that contain metal hydride particles in a sol-gel matrix. Dimensionally stable granules containing metal hydrides are needed for applications such as hydrogen separation and hydrogen purification that require columns containing metal hydrides. Gases must readily flow through the metal hydride beds in the columns. Metal hydrides reversibly absorb and desorb hydrogen and hydrogen isotopes. This is accompanied by significant volume changes that cause the metal hydride to break apart or decrepitate. Repeated cycling results in very fine metal hydride particles that are difficult to handle and contain. Fine particles tendmore » to settle and pack making it more difficult to flow gases through a metal hydride bed. Furthermore, the metal hydrides can exert a significant force on the containment vessel as they expand. These problems associated with metal hydrides can be eliminated with the granulation process described in this report. Small agglomerates of metal hydride particles and abietic acid (a pore former) were produced and dispersed in a colloidal silica/water suspension to form the feed slurry. Fumed silica was added to increase the viscosity of the feed slurry which helped to keep the agglomerates in suspension. Drops of the feed slurry were injected into a 27-foot tall column of hot ({approx}70 C), medium viscosity ({approx}3000 centistokes) silicone oil. Water was slowly evaporated from the drops as they settled. The drops gelled and eventually solidified to form spherical granules. This process is referred to as external gelation. Testing was completed to optimize the design of the column, the feed system, the feed slurry composition, and the operating parameters of the column. The critical process parameters can be controlled resulting in a reproducible fabrication technique. The residual silicone oil on the surface of the granules was removed by washing in mineral spirits. The granules were dried in air at 40 C. The granules were heated to 230 C for 30 minutes in argon to remove the remaining water and organic materials. The resulting product was spherical composite granules (100 to 2000 micron diameter) with a porous silica matrix containing small agglomerates of metal hydride particles. Open porosity in the silica matrix allows hydrogen to permeate rapidly through the matrix but the pores are small enough to contain the metal hydride particles. Additional porosity around the metal hydride particles, induced using abietic acid as a pore former, allows the particles to freely expand and contract without fracturing the brittle sol-gel matrix. It was demonstrated that the granules readily absorb and desorb hydrogen while remaining integral and dimensionally stable. Microcracking was observed after the granules were cycled in hydrogen five times. The strength of the granules was improved by coating them with a thin layer of a micro-porous polymer sol-gel that would allow hydrogen to freely pass through the coating but would filter out metal hydride poisons such as water and carbon monoxide. It was demonstrated that if a thin sol-gel coating was applied after the granules were cycled, the coating not only improved the strength of the granules but the coated granules retained their strength after additional hydrogen cycling tests. This additional strength is needed to extend the lifetime of the granules and to survive the compressive load in a large column of granules. Additional hydrogen adsorption tests are planned to evaluate the performance of coated granules after one hundred cycles. Tests will also be performed to determine the effects of metal hydride poisons on the granules. The results of these tests will be documented in a separate report. The process that was developed to form these granules could be scaled to a production process. The process to form granules from a mixture of metal hydride particles and pore former such as abietic acid can be scaled up using commercial granulators. The current laboratory-scale external gelation column produces approximately one gram of granules per hour. To increase the production output from a single column, multiple feed injection systems in a larger diameter column could be used.« less

Honey bee drones maintain humoral immune competence throughout all life stages in the absence of vitellogenin production.

PubMed

Gätschenberger, Heike; Gimple, Olaf; Tautz, Jürgen; Beier, Hildburg

2012-04-15

Drones are haploid male individuals whose major social function in honey bee colonies is to produce sperm and mate with a queen. In spite of their limited tasks, the vitality of drones is of utmost importance for the next generation. The immune competence of drones - as compared to worker bees - is largely unexplored. Hence, we studied humoral and cellular immune reactions of in vitro reared drone larvae and adult drones of different age upon artificial bacterial infection. Haemolymph samples were collected after aseptic and septic injury and subsequently employed for (1) the identification of immune-responsive peptides and/or proteins by qualitative proteomic analyses in combination with mass spectrometry and (2) the detection of antimicrobial activity by inhibition-zone assays. Drone larvae and adult drones responded with a strong humoral immune reaction upon bacterial challenge, as validated by the expression of small antimicrobial peptides. Young adult drones exhibited a broader spectrum of defence reactions than drone larvae. Distinct polypeptides including peptidoglycan recognition protein-S2 and lysozyme 2 were upregulated in immunized adult drones. Moreover, a pronounced nodulation reaction was observed in young drones upon bacterial challenge. Prophenoloxidase zymogen is present at an almost constant level in non-infected adult drones throughout the entire lifespan. All observed immune reactions in drones were expressed in the absence of significant amounts of vitellogenin. We conclude that drones - like worker bees - have the potential to activate multiple elements of the innate immune response.

Stimulation of insulin release by glucose is associated with an increase in the number of docked granules in the beta-cells of rat pancreatic islets.

PubMed

Straub, Susanne G; Shanmugam, Geetha; Sharp, Geoffrey W G

2004-12-01

Electron microscopy and quantitative stereological techniques were used to study the dynamics of the docked granule pool in the rat pancreatic beta-cell. The mean number of granules per beta-cell was 11,136. After equilibration in RPMI containing 5.6 mmol/l glucose, 6.4% of the granules (approximately 700) were docked at the plasma membrane (also measured as [means +/- SE] 4.3 +/- 0.6 docked granules per 10 microm of plasma membrane at the perimeter of the cell sections). After a 40-min exposure to 16.7 mmol/l glucose, 10.2% of the granules (approximately 1,060) were docked (6.4 +/- 0.8 granules per 10 microm of plasma membrane). Thus, the docked pool increased by 50% during stimulation with glucose. Islets were also exposed to 16.7 mmol/l glucose in the absence or presence of 10 micromol/l nitrendipine. In the absence and presence of nitrendipine, there were 6.1 +/- 0.7 and 6.3 +/- 0.6 granules per 10 microm of membrane, respectively. Thus, glucose increased granule docking independently of increased [Ca2+]i and exocytosis. The data suggest a limit to the number of docking sites. As the rate of docking exceeded the rate of exocytosis, docking is not rate limiting for insulin release. Only with extremely high release rates, glucose stimulation after a 4-h incubation with a high concentration of fatty acid-free BSA, was the docked granule pool reduced in size.

CSR-1 and P granules suppress sperm-specific transcription in the C. elegans germline.

PubMed

Campbell, Anne C; Updike, Dustin L

2015-05-15

Germ granules (P granules) in C. elegans are required for fertility and function to maintain germ cell identity and pluripotency. Sterility in the absence of P granules is often accompanied by the misexpression of soma-specific proteins and the initiation of somatic differentiation in germ cells. To investigate whether this is caused by the accumulation of somatic transcripts, we performed mRNA-seq on dissected germlines with and without P granules. Strikingly, we found that somatic transcripts do not increase in the young adult germline when P granules are impaired. Instead, we found that impairing P granules causes sperm-specific mRNAs to become highly overexpressed. This includes the accumulation of major sperm protein (MSP) transcripts in germ cells, a phenotype that is suppressed by feminization of the germline. A core component of P granules, the endo-siRNA-binding Argonaute protein CSR-1, has recently been ascribed with the ability to license transcripts for germline expression. However, impairing CSR-1 has very little effect on the accumulation of its mRNA targets. Instead, we found that CSR-1 functions with P granules to prevent MSP and sperm-specific mRNAs from being transcribed in the hermaphrodite germline. These findings suggest that P granules protect germline integrity through two different mechanisms, by (1) preventing the inappropriate expression of somatic proteins at the level of translational regulation, and by (2) functioning with CSR-1 to limit the domain of sperm-specific expression at the level of transcription. © 2015. Published by The Company of Biologists Ltd.

Tracking the Fragile X Mental Retardation Protein in a Highly Ordered Neuronal RiboNucleoParticles Population: A Link between Stalled Polyribosomes and RNA Granules.

PubMed

El Fatimy, Rachid; Davidovic, Laetitia; Tremblay, Sandra; Jaglin, Xavier; Dury, Alain; Robert, Claude; De Koninck, Paul; Khandjian, Edouard W

2016-07-01

Local translation at the synapse plays key roles in neuron development and activity-dependent synaptic plasticity. mRNAs are translocated from the neuronal soma to the distant synapses as compacted ribonucleoparticles referred to as RNA granules. These contain many RNA-binding proteins, including the Fragile X Mental Retardation Protein (FMRP), the absence of which results in Fragile X Syndrome, the most common inherited form of intellectual disability and the leading genetic cause of autism. Using FMRP as a tracer, we purified a specific population of RNA granules from mouse brain homogenates. Protein composition analyses revealed a strong relationship between polyribosomes and RNA granules. However, the latter have distinct architectural and structural properties, since they are detected as close compact structures as observed by electron microscopy, and converging evidence point to the possibility that these structures emerge from stalled polyribosomes. Time-lapse video microscopy indicated that single granules merge to form cargoes that are transported from the soma to distal locations. Transcriptomic analyses showed that a subset of mRNAs involved in cytoskeleton remodelling and neural development is selectively enriched in RNA granules. One third of the putative mRNA targets described for FMRP appear to be transported in granules and FMRP is more abundant in granules than in polyribosomes. This observation supports a primary role for FMRP in granules biology. Our findings open new avenues for the study of RNA granule dysfunctions in animal models of nervous system disorders, such as Fragile X syndrome.

Fluidized Bed Hot-Melt Granulation as a Tool to Improve Curcuminoid Solubility.

PubMed

Teixeira, Cristiane C C; de Paiva Junior, Elias; de Freitas, Luis Alexandre Pedro

2018-04-01

Curcumin is the main bioactive component of Curcuma longa L. and has recently aroused growing interest from the scientific community. Unfortunately, the medicinal properties attributed to curcuminoids are impaired by their low oral bioavailability or low solubility in aqueous solutions. Many strategies have been studied to improve curcumin solubility; however, the preparation of granules using hydrophilic materials has never been attempted. The aim of this work was to develop curcumin granules by fluidized bed hot-melt granulation using the hydrophilic carrier Gelucire® 50:13. A two-level factorial design was used to verify the influence of Gelucire® 50:13 and lactose contents found in the granules on their size, morphology, bulk and tapped densities, flow, moisture content, and water activity. The granules obtained were also evaluated by differential scanning calorimetry, thermogravimetric analysis, X-ray powder diffraction, and infrared spectrometry. The curcumin solubility and dissolution rates in water were determined by liquid chromatography. The best formulation provides an increase of curcumin solubility of 4642-fold and 3.8-fold compared to the physical mixture. The dissolution tests showed a maximum drug release from granules after 45 min of 70% at pH 1.2 and 80% at pH 5.8 and 7.4, while for non-granulated curcumin, the release was below 20% in all pH. The solid-state characterization and solubility measurement showed good stability of granules over 9 months. The results attest that the fluidized bed hot-melt granulation with hydrophilic binders is an attractive and promising alternative to obtain solid forms of curcumin with enhanced bioavailability.

Use of Mixer Torque Rheometer to Clarify the Relationship between the Kneading States of Wet Mass and the Dissolution of Final Product in High Shear Granulation.

PubMed

Otsuka, Tomoko; Kuroiwa, Yosuke; Sato, Kazunari; Yamashita, Kazunari; Hakomori, Tadashi; Kimura, Shin-Ichiro; Iwao, Yasunori; Itai, Shigeru

2018-01-01

The properties of wet mass, which indicate the progress of high shear granulation processes, usually have an effect on final product properties, such as tablet dissolution. The mixer torque rheometer (MTR) is a useful tool for quantitatively measuring the 'kneading state' of wet mass and detecting differences in granules. However, there have been no studies of the relationship between the MTR torque and the final product properties to date. In this study, we measured the MTR torque of wet granules at different kneading states, which were prepared by changing the granulation conditions. We then evaluated the relationship between the MTR torque and the dissolution rate of the final product properties. The amperage of the high shear granulator is usually monitored during granulation, but we could not detect a difference in the kneading state through the amperage. However, using MTR torque we were able to quantify the difference of the wet mass. Moreover, MTR torque showed a high correlation with dissolution, compared with the correlations with other intermediate properties, such as granules particle size and tablet hardness. These other properties are affected by following processes and are not properties that directly relate to the kneading state. Thus, MTR torque is a property of wet mass after granulation, and it can be used to directly evaluate differences of the kneading state, and as a result, dissolution. These results indicate the importance of controlling the kneading state, i.e., the progress of granulation, and the utility of MTR for detecting differences in wet mass.

High frequency pacing of edge localized modes by injection of lithium granules in DIII-D H-mode discharges

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bortolon, A.; Maingi, R.; Mansfield, D. K.

A newly installed Lithium Granule Injector (LGI) was used to pace edge localized modes (ELM) in DIII-D. ELM pacing efficiency was studied injecting lithium granules of nominal diameter 0.3–0.9mm, speed of 50–120 m s -1 and average injection rates up to 100 Hz for 0.9mm granules and up to 700 Hz for 0.3mm granules. The efficiency of ELM triggering was found to depend strongly on size of the injected granules, with triggering efficiency close to 100% obtained with 0.9mm diameter granules, lower with smaller sizes, and weakly depending on granule velocity. Robust ELM pacing was demonstrated in ITER-like plasmas formore » the entire shot length, at ELM frequencies 3–5 times larger than the ‘natural’ ELM frequency observed in reference discharges. Within the range of ELM frequencies obtained, the peak ELM heat flux at the outer strike point was reduced with increasing pacing frequency. The peak heat flux reduction at the inner strike point appears to saturate at high pacing frequency. Lithium was found in the plasma core, with a concurrent reduction of metallic impurities and carbon. Altogether, high frequency ELM pacing using the lithium granule injection appears to be compatible with both H-mode energy confinement and attractive H-mode pedestal characteristics, but further assessment is need« less

Microtubule-dependent association of AKAP350A and CCAR1 with RNA stress granules

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kolobova, Elena; Efimov, Andrey; Kaverina, Irina

Recent investigations have highlighted the importance of subcellular localization of mRNAs to cell function. While AKAP350A, a multifunctional scaffolding protein, localizes to the Golgi apparatus and centrosomes, we have now identified a cytosolic pool of AKAP350A. Analysis of AKAP350A scaffolded complexes revealed two novel interacting proteins, CCAR1 and caprin-1. CCAR1, caprin-1 and AKAP350A along with G3BP, a stress granule marker, relocate to RNA stress granules after arsenite treatment. Stress also caused loss of AKAP350 from the Golgi and fragmentation of the Golgi apparatus. Disruption of microtubules with nocodazole altered stress granule formation and changed their morphology by preventing fusion ofmore » stress granules. In the presence of nocodazole, arsenite induced smaller granules with the vast majority of AKAP350A and CCAR1 separated from G3BP-containing granules. Similar to nocodazole treatment, reduction of AKAP350A or CCAR1 expression also altered the size and number of G3BP-containing stress granules induced by arsenite treatment. A limited set of 69 mRNA transcripts was immunoisolated with AKAP350A even in the absence of stress, suggesting the association of AKAP350A with mRNA transcripts. These results provide the first evidence for the microtubule dependent association of AKAP350A and CCAR1 with RNA stress granules.« less

A Single-Granule-Level Approach Reveals Ecological Heterogeneity in an Upflow Anaerobic Sludge Blanket Reactor

PubMed Central

Mei, Ran; Narihiro, Takashi; Bocher, Benjamin T. W.; Yamaguchi, Takashi; Liu, Wen-Tso

2016-01-01

Upflow anaerobic sludge blanket (UASB) reactor has served as an effective process to treat industrial wastewater such as purified terephthalic acid (PTA) wastewater. For optimal UASB performance, balanced ecological interactions between syntrophs, methanogens, and fermenters are critical. However, much of the interactions remain unclear because UASB have been studied at a “macro”-level perspective of the reactor ecosystem. In reality, such reactors are composed of a suite of granules, each forming individual micro-ecosystems treating wastewater. Thus, typical approaches may be oversimplifying the complexity of the microbial ecology and granular development. To identify critical microbial interactions at both macro- and micro- level ecosystem ecology, we perform community and network analyses on 300 PTA–degrading granules from a lab-scale UASB reactor and two full-scale reactors. Based on MiSeq-based 16S rRNA gene sequencing of individual granules, different granule-types co-exist in both full-scale reactors regardless of granule size and reactor sampling depth, suggesting that distinct microbial interactions occur in different granules throughout the reactor. In addition, we identify novel networks of syntrophic metabolic interactions in different granules, perhaps caused by distinct thermodynamic conditions. Moreover, unseen methanogenic relationships (e.g. “Candidatus Aminicenantes” and Methanosaeta) are observed in UASB reactors. In total, we discover unexpected microbial interactions in granular micro-ecosystems supporting UASB ecology and treatment through a unique single-granule level approach. PMID:27936088

Footprint area analysis of binary imaged Cupriavidus necator cells to study PHB production at balanced, transient, and limited growth conditions in a cascade process.

PubMed

Vadlja, Denis; Koller, Martin; Novak, Mario; Braunegg, Gerhart; Horvat, Predrag

2016-12-01

Statistical distribution of cell and poly[3-(R)-hydroxybutyrate] (PHB) granule size and number of granules per cell are investigated for PHB production in a five-stage cascade (5CSTR). Electron microscopic pictures of cells from individual cascade stages (R1-R5) were converted to binary pictures to visualize footprint areas for polyhydroxyalkanoate (PHA) and non-PHA biomass. Results for each stage were correlated to the corresponding experimentally determined kinetics (specific growth rate μ and specific productivity π). Log-normal distribution describes PHA granule size dissimilarity, whereas for R1 and R4, gamma distribution best reflects the situation. R1, devoted to balanced biomass synthesis, predominately contains cells with rather small granules, whereas with increasing residence time τ, maximum and average granule sizes by trend increase, approaching an upper limit determined by the cell's geometry. Generally, an increase of intracellular PHA content and ratio of granule to cell area slow down along the cascade. Further, the number of granules per cell decreases with increasing τ. Data for μ and π obtained by binary picture analysis correlate well with the experimental results. The work describes long-term continuous PHA production under balanced, transient, and nutrient-deficient conditions, as well as their reflection on the granules size, granule number, and cell structure on the microscopic level.

Odontogenic Differentiation of Human Dental Pulp Stem Cells Stimulated by the Calcium Phosphate Porous Granules

PubMed Central

Nam, Sunyoung; Won, Jong-Eun; Kim, Cheol-Hwan; Kim, Hae-Won

2011-01-01

Effects of three-dimensional (3D) calcium phosphate (CaP) porous granules on the growth and odontogenic differentiation of human dental pulp stem cells (hDPSCs) were examined for dental tissue engineering. hDPSCs isolated from adult human dental pulps were cultured for 3-4 passages, and populated on porous granules. Cell growth on the culture dish showed an ongoing increase for up to 21 days, whereas the growth on the 3D granules decreased after 14 days. This reduction in proliferative potential on the 3D granules was more conspicuous under the osteogenic medium conditions, indicating that the 3D granules may induce the odontogenic differentiation of hDPSCs. Differentiation behavior on the 3D granules was confirmed by the increased alkaline phosphatase activity, up-regulation of odontoblast-specific genes, including dentin sialophosphoprotein (DSPP) and dentin matrix protein 1 (DMP1) by quantitative polymerase chain reaction, and greater level of dentin sialoprotein synthesis by western blot. Moreover, the cellular mineralization, as assessed by Alizarin red S and calcium quantification, was significantly higher in the 3D CaP granules than in the culture dish. Taken all, the 3D CaP porous granules should be useful for dental tissue engineering in combination with hDPSCs by providing favorable 3D substrate conditions for cell growth and odontogenic development. PMID:21772958

High frequency pacing of edge localized modes by injection of lithium granules in DIII-D H-mode discharges

DOE PAGES

Bortolon, A.; Maingi, R.; Mansfield, D. K.; ...

2016-04-08

A newly installed Lithium Granule Injector (LGI) was used to pace edge localized modes (ELM) in DIII-D. ELM pacing efficiency was studied injecting lithium granules of nominal diameter 0.3–0.9mm, speed of 50–120 m s -1 and average injection rates up to 100 Hz for 0.9mm granules and up to 700 Hz for 0.3mm granules. The efficiency of ELM triggering was found to depend strongly on size of the injected granules, with triggering efficiency close to 100% obtained with 0.9mm diameter granules, lower with smaller sizes, and weakly depending on granule velocity. Robust ELM pacing was demonstrated in ITER-like plasmas formore » the entire shot length, at ELM frequencies 3–5 times larger than the ‘natural’ ELM frequency observed in reference discharges. Within the range of ELM frequencies obtained, the peak ELM heat flux at the outer strike point was reduced with increasing pacing frequency. The peak heat flux reduction at the inner strike point appears to saturate at high pacing frequency. Lithium was found in the plasma core, with a concurrent reduction of metallic impurities and carbon. Altogether, high frequency ELM pacing using the lithium granule injection appears to be compatible with both H-mode energy confinement and attractive H-mode pedestal characteristics, but further assessment is need« less

Conceptual framework for model-based analysis of residence time distribution in twin-screw granulation.

PubMed

Kumar, Ashish; Vercruysse, Jurgen; Vanhoorne, Valérie; Toiviainen, Maunu; Panouillot, Pierre-Emmanuel; Juuti, Mikko; Vervaet, Chris; Remon, Jean Paul; Gernaey, Krist V; De Beer, Thomas; Nopens, Ingmar

2015-04-25

Twin-screw granulation is a promising continuous alternative for traditional batchwise wet granulation processes. The twin-screw granulator (TSG) screws consist of transport and kneading element modules. Therefore, the granulation to a large extent is governed by the residence time distribution within each module where different granulation rate processes dominate over others. Currently, experimental data is used to determine the residence time distributions. In this study, a conceptual model based on classical chemical engineering methods is proposed to better understand and simulate the residence time distribution in a TSG. The experimental data were compared with the proposed most suitable conceptual model to estimate the parameters of the model and to analyse and predict the effects of changes in number of kneading discs and their stagger angle, screw speed and powder feed rate on residence time. The study established that the kneading block in the screw configuration acts as a plug-flow zone inside the granulator. Furthermore, it was found that a balance between the throughput force and conveying rate is required to obtain a good axial mixing inside the twin-screw granulator. Although the granulation behaviour is different for other excipients, the experimental data collection and modelling methods applied in this study are generic and can be adapted to other excipients. Copyright © 2015 Elsevier B.V. All rights reserved.

Biomaterials differentially regulate Src kinases and phosphoinositide 3-kinase-γ in polymorphonuclear leukocyte primary and tertiary granule release

PubMed Central

Cohen, Hannah Caitlin; Frost, Dustin C.; Lieberthal, Tyler Jacob; Li, Lingjun; Kao, W. John

2018-01-01

In the foreign body response, infiltrating PMNs exocytose granule subsets to influence subsequent downstream inflammatory and wound healing events. In previous studies, we found that PMNs cultured on poly(ethylene glycol) (PEG)-containing hydrogels (i.e., PEG and gelatin + PEG hydrogels) had enhanced primary granule release, yet similar tertiary granule release compared with PMNs cultured on polydimethylsiloxane or tissue culture polystyrene. PMN primary granules contain microbicidal proteins and proteases, which can potentially injure bystander cells, degrade the extracellular matrix, and promote inflammation. Here, we sought to understand the mechanism of the enhanced primary granule release from PMNs on PEG hydrogels. We found that primary granule release from PMNs on PEG hydrogels was adhesion mediated and involved Src family kinases and PI3K-γ. The addition of gelatin to PEG hydrogels did not further enhance PMN primary granule release. Using stable-isotope dimethyl labeling-based shotgun proteomics, we identified many serum proteins – including Ig gamma constant chain region proteins and alpha-1-acid glycoprotein 1 – that were absorbed/adsorbed in higher quantities on PEG hydrogels than on TCPS, and may be involved in mediating PMN primary granule release. Ultimately, this mechanistic knowledge can be used to direct inflammation and wound healing following biomaterial implantation to promote a more favorable healing response. PMID:25736495

Optimizing granules size distribution for aerobic granular sludge stability: Effect of a novel funnel-shaped internals on hydraulic shear stress.

PubMed

Zhou, Jia-Heng; Zhang, Zhi-Ming; Zhao, Hang; Yu, Hai-Tian; Alvarez, Pedro J J; Xu, Xiang-Yang; Zhu, Liang

2016-09-01

A novel funnel-shaped internals was proposed to enhance the stability and pollutant removal performance of an aerobic granular process by optimizing granule size distribution. Results showed up to 68.3±1.4% of granules in novel reactor (R1) were situated in optimal size range (700-1900μm) compared to less than 29.7±1.1% in conventional reactor (R2), and overgrowth of large granules was effectively suppressed without requiring additional energy. Consequently, higher total nitrogen (TN) removal (81.6±2.1%) achieved in R1 than in R2 (48.1±2.7%). Hydraulic analysis revealed the existence of selectively assigning hydraulic pressure in R1. The total shear rate (τtotal) on large granules was 3.07±0.14 times higher than that of R2, while τtotal of small granules in R1 was 70.7±4.6% in R2. Furthermore, large granules in R1 with intact extracellular polymeric substances (EPS) outer layer structure entrapped hydroxyapatite at center, which formed a core structure and further enhanced the stability of aerobic granules. Copyright © 2016 Elsevier Ltd. All rights reserved.

Exocytosis of Neutrophil Granule Subsets and Activation of Prolyl Isomerase 1 are required for Respiratory Burst Priming

PubMed Central

McLeish, Kenneth R.; Uriarte, Silvia M.; Tandon, Shweta; Creed, Timothy M.; Le, Junyi; Ward, Richard A.

2013-01-01

This study tested the hypothesis that priming the neutrophil respiratory burst requires both granule exocytosis and activation of the prolyl isomerase, Pin1. Fusion proteins containing the TAT cell permeability sequence and either the SNARE domain of syntaxin-4 or the N-terminal SNARE domain of SNAP-23 were used to examine the role of granule subsets in TNF-mediated respiratory burst priming using human neutrophils. Concentration-inhibition curves for exocytosis of individual granule subsets and for priming of fMLF-stimulated superoxide release and phagocytosis-stimulated H2O2 production were generated. Maximal inhibition of priming ranged from 72% to 88%. Linear regression lines for inhibition of priming versus inhibition of exocytosis did not differ from the line of identity for secretory vesicles and gelatinase granules, while the slopes or the y-intercepts were different from the line of identity for specific and azurophilic granules. Inhibition of Pin1 reduced priming by 56%, while exocytosis of secretory vesicles and specific granules was not affected. These findings indicate that exocytosis of secretory vesicles and gelatinase granules and activation of Pin1 are independent events required for TNF-mediated priming of neutrophil respiratory burst. PMID:23363774

Influence of raw material properties upon critical quality attributes of continuously produced granules and tablets.

PubMed

Fonteyne, Margot; Wickström, Henrika; Peeters, Elisabeth; Vercruysse, Jurgen; Ehlers, Henrik; Peters, Björn-Hendrik; Remon, Jean Paul; Vervaet, Chris; Ketolainen, Jarkko; Sandler, Niklas; Rantanen, Jukka; Naelapää, Kaisa; De Beer, Thomas

2014-07-01

Continuous manufacturing gains more and more interest within the pharmaceutical industry. The International Conference of Harmonisation (ICH) states in its Q8 'Pharmaceutical Development' guideline that the manufacturer of pharmaceuticals should have an enhanced knowledge of the product performance over a range of raw material attributes, manufacturing process options and process parameters. This fits further into the Process Analytical Technology (PAT) and Quality by Design (QbD) framework. The present study evaluates the effect of variation in critical raw material properties on the critical quality attributes of granules and tablets, produced by a continuous from-powder-to-tablet wet granulation line. The granulation process parameters were kept constant to examine the differences in the end product quality caused by the variability of the raw materials properties only. Theophylline-Lactose-PVP (30-67.5-2.5%) was used as model formulation. Seven different grades of theophylline were granulated. Afterward, the obtained granules were tableted. Both the characteristics of granules and tablets were determined. The results show that differences in raw material properties both affect their processability and several critical quality attributes of the resulting granules and tablets. Copyright © 2014 Elsevier B.V. All rights reserved.

Mathematical Model of Heat Transfer in the Catalyst Granule with Point Reaction Centers

NASA Astrophysics Data System (ADS)

Derevich, I. V.; Fokina, A. Yu.

2018-01-01

This paper considers a catalyst granule with a porous ceramic chemically inert base and active point centers, at which an exothermic reaction of synthesis takes place. The rate of a chemical reaction depends on temperature by the Arrhenius law. The heat is removed from the catalyst granule surface to the synthesis products by heat transfer. Based on the idea of self-consistent field, a closed system of equations is constructed for calculating the temperatures of the active centers. As an example, a catalyst granule of the Fischer-Tropsch synthesis with active metallic cobalt particles is considered. The stationary temperatures of the active centers are calculated by the timedependent technique by solving a system of ordinary differential equations. The temperature distribution inside the granule has been found for the local centers located on one diameter of the granule and distributed randomly in the granule's volume. The existence of the critical temperature inside the reactor has been established, the excess of which leads to substantial superheating of local centers. The temperature distribution with local reaction centers differs qualitatively from the granule temperature calculated in the homogeneous approximation. The results of calculations are given.

Assessment of intra-granular and extra-granular fracture in the development of tablet tensile strength.

PubMed

Mitra, Biplob; Hilden, Jon; Litster, James D

2018-05-24

When a tablet is compacted from deformable granules and then broken, the fracture plane may cleave granules in two (intra-granular fracture) or separate neighboring granules (extra-granular fracture). In this study, a novel method was developed to quantify the extent of intra- versus extra-granular fracture by compacting tablets from multi-colored ideal granules and evaluating fracture surfaces. The proportions of intra-granular and extra-granular fracture were quantified and modeled in light of a new metric, the deformation potential, Δ, reflecting the solid fraction increase as an initial granule bed is compressed into a final tablet. Results show that a measurable tablet strength is achieved at Δ > 0.18, but intra-granular fracture is not observed until Δ > 0.21. At very large Δ, tablets experience almost exclusively intra-granular fracture, yet the tablet tensile strength is considerably lower than that of a tablet compacted from raw powders versus pre-compacted granules. Thus, secondary compaction of granules appears to weaken the granule matrix, leading to reduced tablet tensile strength even in the presence of strong extra-granular bonding. Copyright © 2018. Published by Elsevier Inc.

Study of quantitative interactions of potato and corn starch granules with ions in diluted solutions of heavy metal salts.

PubMed

Szymońska, Joanna; Molenda, Marcin; Wieczorek, Jerzy

2015-12-10

Interactions of potato and corn starch granules with ions in diluted solutions of silver, lead, copper or iron salts were investigated. It was shown experimentally that granules accumulated the cations in amounts depending on the granule structure and water content as well as a type of both metal and counter-ions present in solution. Potato starch retained almost three times more cations compared to corn starch what was proportional to the total phosphorous content in these starches. Quantity of milligrams of cations bound by 1g of starch was inversely correlated with the cation hydration. Ag(+), Pb(2+) and Cu(2+) were connected in stoichiometric amounts of moles to semicrystalline and amorphous parts of the granules. Fe(3+) ions were accumulated in higher than stoichiometric quantities mainly in granule amorphous regions. Metal ions penetrated into granules together with anions except nitrates which remained on surface of potato starch granules. Cations facilitated the starch thermal decomposition in accordance with values of their standard redox potentials. Nitrates supported this process only in the presence of base metal cations. Copyright © 2015 Elsevier Ltd. All rights reserved.

Control of cerebellar granule cell output by sensory-evoked Golgi cell inhibition

PubMed Central

Duguid, Ian; Branco, Tiago; Chadderton, Paul; Arlt, Charlotte; Powell, Kate; Häusser, Michael

2015-01-01

Classical feed-forward inhibition involves an excitation–inhibition sequence that enhances the temporal precision of neuronal responses by narrowing the window for synaptic integration. In the input layer of the cerebellum, feed-forward inhibition is thought to preserve the temporal fidelity of granule cell spikes during mossy fiber stimulation. Although this classical feed-forward inhibitory circuit has been demonstrated in vitro, the extent to which inhibition shapes granule cell sensory responses in vivo remains unresolved. Here we combined whole-cell patch-clamp recordings in vivo and dynamic clamp recordings in vitro to directly assess the impact of Golgi cell inhibition on sensory information transmission in the granule cell layer of the cerebellum. We show that the majority of granule cells in Crus II of the cerebrocerebellum receive sensory-evoked phasic and spillover inhibition prior to mossy fiber excitation. This preceding inhibition reduces granule cell excitability and sensory-evoked spike precision, but enhances sensory response reproducibility across the granule cell population. Our findings suggest that neighboring granule cells and Golgi cells can receive segregated and functionally distinct mossy fiber inputs, enabling Golgi cells to regulate the size and reproducibility of sensory responses. PMID:26432880

Particle size distribution of wheat starch granules in relation to baking properties of frozen dough.

PubMed

Tao, Han; Wang, Pei; Wu, Fengfeng; Jin, Zhengyu; Xu, Xueming

2016-02-10

The impact of freezing on the wheat starches with different particle size was studied using a range of characterization methods including X-ray diffraction, differential scanning calorimetry, the Rapid Visco Analyser and a reconstitution dough system. Wheat starches were fractionated into A- and B-type granules, and then subjected to freezing/thawing treatment for 3 cycles. The freezing treatment did not cause apparent damage on A-type granular surface but induced cracked structure on B-type granules. It facilitated materials such as amylose, proteins, and lipids leaching from starch granule and an increase in gelatinization temperatures, melting enthalpy, and pasting viscosities. A smaller bread specific volume was obtained from freezing-treated B-granules while the crumb firmness significantly increased (p>0.05). No marked differences were observed in the counterparts of A-granules after freezing treatment. It seemed that the B-type granules were more sensitive to the freezing/thawing treatment, thus facilitating structural transformations from dough to bread. Results indicated that the deterioration in frozen bread quality derived from starch could be minimized by increasing the A-granules content. Copyright © 2015 Elsevier Ltd. All rights reserved.

To determine the end point of wet granulation by measuring powder energies and thermal properties.

PubMed

Dave, Rutesh H; Wu, Stephen H; Contractor, Labdhi D

2012-04-01

Wet granulation has been widely used in pharmaceutical industry as a tablet manufacturing process. However, end-point determination of wet granulation process has always remained a challenge. Many traditional methods are available for end-point determination, yet accuracy and reproducibility still remain a challenge. Microcrystalline cellulose, widely used as an excipient in pharmaceutical industry, was granulated using water. Wet mass was passed through sieve # 12 and dried till constant percentage loss on drying was obtained and dried granules were obtained. Wet and dried granules collected were subjected to basic flow energy, specific energy, bulk density, pressure drop, differential scanning calorimetry and effusivity measurements. Analysis of data revealed various stages of granule growth from initial seed formation by adding 200-400 g of water, granule growth was observed by adding 600-800 g of water and over wetting was observed at 1155 g of water. In this work, we have justified our work to properly identify and utilize this technique for practical purpose to correctly identify the end-point determination of microcrystalline cellulose and explain various principles underlying energies associated with powder and thermal measurements.

Phosphatidylinositol kinase. A component of the chromaffin-granule membrane

PubMed Central

Phillips, John H.

1973-01-01

Phosphorylation of bovine chromaffin granules by ATP leads to the formation of diphosphoinositide in the granule membrane. Both phosphatidylinositol kinase and its substrate are components of this membrane, and triphosphoinositide is not formed under the conditions of the assay. The reaction is Mg2+-dependent and is stimulated by Mn2+ and F− ions. The initial reaction is rapid, with a broad pH profile and a `transition' temperature for its activation energy at 27°C. The apparent Km for ATP is 5μm. ATP, N-ethylmaleimide, Cu2+ ions and NaIO4 are inhibitory. The phospholipids of chromaffin-granule membranes have been analysed: 6.8% of the lipid P is found in phosphatidylinositol, and only 2–3% in phosphatidylserine. Comparison of the rate of phosphorylation of intact and lysed granules suggests that the sites for phosphorylation are on the outer (cytoplasmic) surface of the granules, and diphosphoinositide may therefore make an important contribution to the charge of the chromaffin granule in vivo. PMID:4360713

Kinetic analysis of glucoamylase-catalyzed hydrolysis of starch granules from various botanical sources.

PubMed

Tatsumi, Hirosuke; Katano, Hajime; Ikeda, Tokuji

2007-04-01

The kinetics of glucoamylase-catalyzed hydrolysis of starch granules from six different botanical sources (rice, wheat, maize, cassava, sweet potato, and potato) was studied by the use of an electrochemical glucose sensor. A higher rate of hydrolysis was obtained as a smaller size of starch granules was used. The adsorbed amount of glucoamylase on the granule surface per unit area did not vary very much with the type of starch granules examined, while the catalytic constants of the adsorbed enzyme (k(0)) were determined to be 23.3+/-4.4, 14.8+/-6.0, 6.2+/-1.8, 7.1+/-4.1, 4.6+/-3.0, and 1.6+/-0.6 s(-1) for rice, wheat, maize, cassava, sweet potato, and potato respectively, showing that k(0) was largely influenced by the type of starch granules. A comparison of the k(0)-values in relation to the crystalline structure of the starch granules suggested that k(0) increases as the crystalline structure becomes dense.

Partial nitrification using aerobic granules in continuous-flow reactor: rapid startup.

PubMed

Wan, Chunli; Sun, Supu; Lee, Duu-Jong; Liu, Xiang; Wang, Li; Yang, Xue; Pan, Xiangliang

2013-08-01

This study applied a novel strategy to rapid startup of partial nitrification in continuous-flow reactor using aerobic granules. Mature aerobic granules were first cultivated in a sequencing batch reactor at high chemical oxygen demand in 16 days. The strains including the Pseudoxanthomonas mexicana strain were enriched in cultivated granules to enhance their structural stability. Then the cultivated granules were incubated in a continuous-flow reactor with influent chemical oxygen deamnad being stepped decreased from 1,500 ± 100 (0-19 days) to 750 ± 50 (20-30 days), and then to 350 ± 50 mg l(-1) (31-50 days); while in the final stage 350 mg l(-1) bicarbonate was also supplied. Using this strategy the ammonia-oxidizing bacterium, Nitrosomonas europaea, was enriched in the incubated granules to achieve partial nitrification efficiency of 85-90% since 36 days and onwards. The partial nitrification granules were successfully harvested after 52 days, a period much shorter than those reported in literature. Copyright © 2013 Elsevier Ltd. All rights reserved.

Why do gelatinized starch granules not dissolve completely? Roles for amylose, protein, and lipid in granule "ghost" integrity.

PubMed

Debet, Martine R; Gidley, Michael J

2007-06-13

After gelatinization in water, starch granules persist in swollen hydrated forms known as ghosts. Three potential mechanisms for ghost formation are tested. Proteins and lipids on the granule surface are found to be a determinant of ghost robustness, but not ghost formation. Proteins inside pre-made maize or wheat starch ghosts are degraded extensively by proteases without any apparent change in ghost properties, making an internal protein cross-linking mechanism unlikely. Waxy maize mutants with a range of amylose contents have ghost integrities that correlate with (low) apparent amylose levels. It is hypothesized that ghost formation is due to cross-linking of polysaccharide chains within swollen granules, most likely involving double helices formed from polymer chains that become free to move following heat-induced granule swelling. The size and robustness of granule ghosts is proposed to be determined by the relative rates of swelling and cross-linking, modulated by surface non-polysaccharide components.

Segregation of large granules from close-packed cluster of small granules due to buoyancy.

PubMed

Yang, Xian-qing; Zhou, Kun; Qiu, Kang; Zhao, Yue-min

2006-03-01

Segregation of large granules in a vibrofluidized granular bed with inhomogeneous granular number density distribution is studied by an event-driven algorithm. Simulation results show that the mean vertical position of large granules decreases with the increase of the density ration of the large granules to the small ones. This conclusion is consistent with the explanation that the net pressure due to the small surrounding particle impacts balances the large granular weight, and indict that the upward movement of the large granules is driven by the buoyancy. The values of temperature, density, and pressure of the systems are also computed by changing the conditions such as heating temperature on the bottom and restitution coefficient of particles. These results indicate that the segregation of large granules also happen in the systems with density inversion or even close-packed cluster of particles floating on a low-density fluid, due to the buoyancy. An equation of state is proposed to explain the buoyancy.

Spherical porous hydroxyapatite granules containing composites of magnetic and hydroxyapatite nanoparticles for the hyperthermia treatment of bone tumor.

PubMed

Kamitakahara, Masanobu; Ohtoshi, Naohiro; Kawashita, Masakazu; Ioku, Koji

2016-05-01

Spherical porous granules of hydroxyapatite (HA) containing magnetic nanoparticles would be suitable for the hyperthermia treatment of bone tumor, because porous HA granules act as a scaffold for bone regeneration, and magnetic nanoparticles generate sufficient heat to kill tumor cells under an alternating magnetic field. Although magnetic nanoparticles are promising heat generators, their small size makes them difficult to support in porous HA ceramics. We prepared micrometer-sized composites of magnetic and HA nanoparticles, and then supported them in porous HA granules composed of rod-like particles. The spherical porous HA granules containing the composites of magnetic and HA nanoparticle were successfully prepared using a hydrothermal process without changing the crystalline phase and heat generation properties of the magnetic nanoparticles. The obtained granules generated sufficient heat for killing tumor cells under an alternating magnetic field (300 Oe at 100 kHz). The obtained granules are expected to be useful for the hyperthermia treatment of bone tumors.

Isolation of new polar granule components in Drosophila reveals P body and ER associated proteins

PubMed Central

Thomson, Travis; Liu, Niankun; Arkov, Alexey; Lehmann, Ruth; Lasko, Paul

2008-01-01

Germ plasm, a specialized cytoplasm present at the posterior of the early Drosophila embryo, is necessary and sufficient for germ cell formation. Germ plasm is rich in mitochondria and contains electron dense structures called polar granules. To identify novel polar granule components we isolated proteins that associate in early embryos with Vasa (VAS) and Tudor (TUD), two known polar granule associated molecules. We identified Maternal expression at 31B (ME31B), eIF4A, Aubergine (AUB) and Transitional Endoplasmic Reticulum 94 (TER94) as components of both VAS and TUD complexes and confirmed their localization to polar granules by immuno-electron microscopy. ME31B, eIF4A and AUB are also present in processing (P) bodies, suggesting that polar granules, which are necessary for germ line formation, might be related to P bodies. Our recovery of ER associated proteins TER94 and ME31B confirms that polar granules are closely linked to the translational machinery and to mRNP assembly. PMID:18590813

Drosophila germ granules are structured and contain homotypic mRNA clusters

PubMed Central

Trcek, Tatjana; Grosch, Markus; York, Andrew; Shroff, Hari; Lionnet, Timothée; Lehmann, Ruth

2015-01-01

Germ granules, specialized ribonucleoprotein particles, are a hallmark of all germ cells. In Drosophila, an estimated 200 mRNAs are enriched in the germ plasm, and some of these have important, often conserved roles in germ cell formation, specification, survival and migration. How mRNAs are spatially distributed within a germ granule and whether their position defines functional properties is unclear. Here we show, using single-molecule FISH and structured illumination microscopy, a super-resolution approach, that mRNAs are spatially organized within the granule whereas core germ plasm proteins are distributed evenly throughout the granule. Multiple copies of single mRNAs organize into ‘homotypic clusters' that occupy defined positions within the center or periphery of the granule. This organization, which is maintained during embryogenesis and independent of the translational or degradation activity of mRNAs, reveals new regulatory mechanisms for germ plasm mRNAs that may be applicable to other mRNA granules. PMID:26242323

Tracheal Wall Thickening Is Associated with the Granulation Tissue Formation Around Silicone Stents in Patients with Post-Tuberculosis Tracheal Stenosis

PubMed Central

Eom, Jung Seop; Jeon, Kyeongman; Um, Sang-Won; Koh, Won-Jung; Suh, Gee Young; Chung, Man Pyo; Kwon, O Jung

2013-01-01

Purpose Tracheal restenosis due to excessive granulation tissue around a silicone stent requires repeated bronchoscopic interventions in patients with post-tuberculosis tracheal stenosis (PTTS). The current study was conducted to identify the risk factors for granulation tissue formation after silicone stenting in PTTS patients. Materials and Methods A retrospective study was conducted between January 1998 and December 2010. Forty-two PTTS patients with silicone stenting were selected. Clinical and radiological variables were retrospectively collected and analyzed. Results Tracheal restenosis due to granulation tissue formation were found in 20 patients (47.6%), and repeated bronchoscopic interventions were conducted. In multivariate analysis, tracheal wall thickness, measured on axial computed tomography scan, was independently associated with granulation tissue formation after silicone stenting. Furthermore, the degree of tracheal wall thickness was well correlated with the degree of granulation tissue formation. Conclusion Tracheal wall thickening was associated with granulation tissue formation around silicone stents in patients with post-tuberculosis tracheal stenosis. PMID:23709431

Tracheal wall thickening is associated with the granulation tissue formation around silicone stents in patients with post-tuberculosis tracheal stenosis.

PubMed

Eom, Jung Seop; Kim, Hojoong; Jeon, Kyeongman; Um, Sang-Won; Koh, Won-Jung; Suh, Gee Young; Chung, Man Pyo; Kwon, O Jung

2013-07-01

Tracheal restenosis due to excessive granulation tissue around a silicone stent requires repeated bronchoscopic interventions in patients with post-tuberculosis tracheal stenosis (PTTS). The current study was conducted to identify the risk factors for granulation tissue formation after silicone stenting in PTTS patients. A retrospective study was conducted between January 1998 and December 2010. Forty-two PTTS patients with silicone stenting were selected. Clinical and radiological variables were retrospectively collected and analyzed. Tracheal restenosis due to granulation tissue formation were found in 20 patients (47.6%), and repeated bronchoscopic interventions were conducted. In multivariate analysis, tracheal wall thickness, measured on axial computed tomography scan, was independently associated with granulation tissue formation after silicone stenting. Furthermore, the degree of tracheal wall thickness was well correlated with the degree of granulation tissue formation. Tracheal wall thickening was associated with granulation tissue formation around silicone stents in patients with post-tuberculosis tracheal stenosis.

The Metalloprotease of Listeria monocytogenes Is Regulated by pH▿

PubMed Central

Forster, Brian M.; Bitar, Alan Pavinski; Slepkov, Emily R.; Kota, Karthik J.; Sondermann, Holger; Marquis, Hélène

2011-01-01

Listeria monocytogenes is an intracytosolic bacterial pathogen. Among the factors contributing to escape from vacuoles are a phosphatidylcholine phospholipase C (PC-PLC) and a metalloprotease (Mpl). Both enzymes are translocated across the bacterial membrane as inactive proproteins, whose propeptides serve in part to maintain them in association with the bacterium. We have shown that PC-PLC maturation is regulated by Mpl and pH and that Mpl maturation occurs by autocatalysis. In this study, we tested the hypothesis that Mpl activity is pH regulated. To synchronize the effect of pH on bacteria, the cytosolic pH of infected cells was manipulated immediately after radiolabeling de novo-synthesized bacterial proteins. Immunoprecipitation of secreted Mpl from host cell lysates revealed the presence of the propeptide and catalytic domain in samples treated at pH 6.5 but not at pH 7.3. The zymogen was present in small amounts under all conditions. Since proteases often remain associated with their respective propeptide following autocatalysis, we aimed at determining whether pH regulates autocatalysis or secretion of the processed enzyme. For this purpose, we used an Mpl construct that contains a Flag tag at the N terminus of its catalytic domain and antibodies that can distinguish N-terminal and non-N-terminal Flag. By fluorescence microscopy, we observed the Mpl zymogen associated with the bacterium at physiological pH but not following acidification. Mature Mpl was not detected in association with the bacterium at either pH. Using purified proteins, we determined that processing of the PC-PLC propeptide by mature Mpl is also pH sensitive. These results indicate that pH regulates the activity of Mpl on itself and on PC-PLC. PMID:21803995

Age-related changes in factor VII proteolysis in vivo.

PubMed

Ofosu, F A; Craven, S; Dewar, L; Anvari, N; Andrew, M; Blajchman, M A

1996-08-01

Previous studies have reported that pre-operative plasmas of patients over the age of 40 years who developed post-operative deep vein thrombosis (DVT) had approximately twice the amount of proteolysed factor VII found in plasmas of patients in whom prophylaxis with heparin or low M(r) heparin was successful. These and other studies also reported higher concentrations of thrombin-antithrombin III in pre- and post-operative plasmas of patients who developed post-operative thrombosis than in plasmas of patients in whom prophylaxis was successful. Whether the extent of factor VII proteolysis seen in the patients who developed post-operative DVT is related to the severity of their disease or age is not known. This report investigated age-related changes in the concentrations of total factor VII protein, factor VII zymogen, factor VIIa, tissue factor pathway inhibitor, thrombin-antithrombin III, and prothrombin fragment 1 + 2 in normal plasmas and the relationships between these parameters. With the exception of thrombin-antithrombin III, statistically significant increases in the concentrations of these parameters with age were found. Additionally, the differences between the concentrations of total factor VII protein and factor VII zymogen, an index factor VII proteolysis in vivo, were statistically significant only for individuals over age 40. Using linear regression analysis, a significant correlation was found to exist between the concentrations of plasma factor VIIa and prothrombin fragment 1 + 2. Since factor VIIa-tissue factor probably initiates coagulation in vivo, we hypothesize that the elevated plasma factor VIIa (reflecting a less tightly regulated tissue factor activity and therefore increased thrombin production in vivo) accounts for the high risk for post-operative thrombosis seen in individuals over the age of 40.

Early to Late Endosome Trafficking Controls Secretion and Zymogen Activation in Rodent and Human Pancreatic Acinar Cells.

PubMed

Messenger, Scott W; Thomas, Diana Dh; Cooley, Michelle M; Jones, Elaina K; Falkowski, Michelle A; August, Benjamin K; Fernandez, Luis A; Gorelick, Fred S; Groblewski, Guy E

2015-11-01

Pancreatic acinar cells have an expanded apical endosomal system, the physiological and pathophysiological significance of which is still emerging. Phosphatidylinositol-3,5-bisphosphate (PI(3,5)P 2 ) is an essential phospholipid generated by PIKfyve, which phosphorylates phosphatidylinositol-3-phosphate (PI(3)P). PI(3,5)P 2 is necessary for maturation of early endosomes (EE) to late endosomes (LE). Inhibition of EE to LE trafficking enhances anterograde endosomal trafficking and secretion at the plasma membrane by default through a recycling endosome (RE) intermediate. We assessed the effects of modulating PIKfyve activity on apical trafficking and pancreatitis responses in pancreatic acinar cells. Inhibition of EE to LE trafficking was achieved using pharmacological inhibitors of PIKfyve, expression of dominant negative PIKfyve K1877E, or constitutively active Rab5-GTP Q79L. Anterograde endosomal trafficking was manipulated by expression of constitutively active and dominant negative Rab11a mutants. The effects of these agents on secretion, endolysosomal exocytosis of lysosome associated membrane protein (LAMP1), and trypsinogen activation in response to high-dose CCK-8, bile acids and cigarette toxin was determined. PIKfyve inhibition increased basal and stimulated secretion. Adenoviral overexpression of PIKfyve decreased secretion leading to cellular death. Expression of Rab5-GTP Q79L or Rab11a-GTP Q70L enhanced secretion. Conversely, dominant-negative Rab11a-GDP S25N reduced secretion. High-dose CCK inhibited endolysosomal exocytosis that was reversed by PIKfyve inhibition. PIKfyve inhibition blocked intracellular trypsin accumulation and cellular damage responses to high CCK-8, tobacco toxin, and bile salts in both rodent and human acini. These data demonstrate that EE-LE trafficking acutely controls acinar secretion and the intracellular activation of zymogens leading to the pathogenicity of acute pancreatitis.

The linker connecting the two kringles plays a key role in prothrombin activation

PubMed Central

Pozzi, Nicola; Chen, Zhiwei; Pelc, Leslie A.; Shropshire, Daniel B.; Di Cera, Enrico

2014-01-01

The zymogen prothrombin is proteolytically converted by factor Xa to the active protease thrombin in a reaction that is accelerated >3,000-fold by cofactor Va. This physiologically important effect is paradigmatic of analogous cofactor-dependent reactions in the coagulation and complement cascades, but its structural determinants remain poorly understood. Prothrombin has three linkers connecting the N-terminal Gla domain to kringle-1 (Lnk1), the two kringles (Lnk2), and kringle-2 to the C-terminal protease domain (Lnk3). Recent developments indicate that the linkers, and particularly Lnk2, confer on the zymogen significant flexibility in solution and enable prothrombin to sample alternative conformations. The role of this flexibility in the context of prothrombin activation was tested with several deletions. Removal of Lnk2 in almost its entirety (ProTΔ146–167) drastically reduces the enhancement of thrombin generation by cofactor Va from >3,000-fold to 60-fold because of a significant increase in the rate of activation in the absence of cofactor. Deletion of Lnk2 mimics the action of cofactor Va and offers insights into how prothrombin is activated at the molecular level. The crystal structure of ProTΔ146–167 reveals a contorted architecture where the domains are not vertically stacked, kringle-1 comes within 9 Å of the protease domain, and the Gla-domain primed for membrane binding comes in contact with kringle-2. These findings broaden our molecular understanding of a key reaction of the blood coagulation cascade where cofactor Va enhances activation of prothrombin by factor Xa by compressing Lnk2 and morphing prothrombin into a conformation similar to the structure of ProTΔ146–167. PMID:24821807

Chitosan nanoencapsulated exogenous trypsin biomimics zymogen-like enzyme in fish gastrointestinal tract.

PubMed

Kumari, Rakhi; Gupta, Subodh; Singh, Arvind R; Ferosekhan, S; Kothari, Dushyant C; Pal, Asim Kumar; Jadhao, Sanjay Balkrishna

2013-01-01

Exogenous proteolytic enzyme supplementation is required in certain disease conditions in humans and animals and due to compelling reasons on use of more plant protein ingredients and profitability in animal feed industry. However, limitations on their utility in diet are imposed by their pH specificity, thermolabile nature, inhibition due to a variety of factors and the possibility of intestinal damage. For enhancing the efficacy and safety of exogenous trypsin, an efficient chitosan (0.04%) nanoencapsulation-based controlled delivery system was developed. An experiment was conducted for 45 days to evaluate nanoencapsulated trypsin (0.01% and 0.02%) along with 0.02% bare trypsin and 0.4% chitosan nanoparticles against a control diet on productive efficiency (growth rate, feed conversion and protein efficiency ratio), organo-somatic indices, nutrient digestibility, tissue enzyme activities, hematic parameters and intestinal histology of the fish Labeo rohita. All the synthesized nanoparticles were of desired characteristics. Enhanced fish productive efficiency using nanoencapsulated trypsin over its bare form was noticed, which corresponded with enhanced (P<0.01) nutrient digestibility, activity of intestinal protease, liver and muscle tissue transaminases (alanine and aspartate) and dehydrogenases (lactate and malate), serum blood urea nitrogen and serum protein profile. Intestinal tissues of fish fed with 0.02% bare trypsin showed broadened, marked foamy cells with lipid vacuoles. However, villi were healthier in appearance with improved morphological features in fish fed with nanoencapsulated trypsin than with bare trypsin, and the villi were longer in fish fed with 0.01% nanoencapsulated trypsin than with 0.02% nanoencapsulated trypsin. The result of this premier experiment shows that nanoencapsulated trypsin mimics zymogen-like proteolytic activity via controlled release, and hence the use of 0.01% nanoencapsulated trypsin (in chitosan nanoparticles) over bare trypsin can be favored as a dietary supplement in animals and humans.

Engineered Toxins “Zymoxins” Are Activated by the HCV NS3 Protease by Removal of an Inhibitory Protein Domain

PubMed Central

Shapira, Assaf; Gal-Tanamy, Meital; Nahary, Limor; Litvak-Greenfeld, Dana; Zemel, Romy; Tur-Kaspa, Ran; Benhar, Itai

2011-01-01

The synthesis of inactive enzyme precursors, also known as “zymogens,” serves as a mechanism for regulating the execution of selected catalytic activities in a desirable time and/or site. Zymogens are usually activated by proteolytic cleavage. Many viruses encode proteases that execute key proteolytic steps of the viral life cycle. Here, we describe a proof of concept for a therapeutic approach to fighting viral infections through eradication of virally infected cells exclusively, thus limiting virus production and spread. Using the hepatitis C virus (HCV) as a model, we designed two HCV NS3 protease-activated “zymogenized” chimeric toxins (which we denote “zymoxins”). In these recombinant constructs, the bacterial and plant toxins diphtheria toxin A (DTA) and Ricin A chain (RTA), respectively, were fused to rationally designed inhibitor peptides/domains via an HCV NS3 protease-cleavable linker. The above toxins were then fused to the binding and translocation domains of Pseudomonas exotoxin A in order to enable translocation into the mammalian cells cytoplasm. We show that these toxins exhibit NS3 cleavage dependent increase in enzymatic activity upon NS3 protease cleavage in vitro. Moreover, a higher level of cytotoxicity was observed when zymoxins were applied to NS3 expressing cells or to HCV infected cells, demonstrating a potential therapeutic window. The increase in toxin activity correlated with NS3 protease activity in the treated cells, thus the therapeutic window was larger in cells expressing recombinant NS3 than in HCV infected cells. This suggests that the “zymoxin” approach may be most appropriate for application to life-threatening acute infections where much higher levels of the activating protease would be expected. PMID:21264238

Proteins on exocytic vesicles mediate calcium-triggered fusion.

PubMed Central

Vogel, S S; Zimmerberg, J

1992-01-01

In many exocytic systems, micromolar concentrations of intracellular Ca2+ trigger fusion. We find that aggregates of secretory granules isolated from sea urchin eggs fuse together when perfused with greater than or equal to 10 microM free Ca2+. Mixing of membrane components was demonstrated by transfer of fluorescent lipophilic dye, and melding of granule contents was seen with differential interference microscopy. A technique based upon light scattering was developed to conveniently detect fusion. Two protein modifiers, trypsin and N-ethylmaleimide, inhibit granule-granule fusion at concentrations similar to those that inhibit granule-plasma membrane fusion. We suggest that molecular machinery sufficient for Ca(2+)-triggered fusion resides on secretory granules as purified and that at least some of these essential components are proteinaceous. Images PMID:1584814

Elucidation of the internal physical and chemical microstructure of pharmaceutical granules using X-ray micro-computed tomography, Raman microscopy and infrared spectroscopy.

PubMed

Crean, Barry; Parker, Andrew; Roux, Delphine Le; Perkins, Mark; Luk, Shen Y; Banks, Simon R; Melia, Colin D; Roberts, Clive J

2010-11-01

X-ray micro-computed tomography (XMCT) was used in conjunction with confocal Raman mapping to measure the intra-granular pore size, binder volumes and to provide spatial and chemical maps of internal granular components in α-lactose monohydrate granules formulated with different molecular weights of polyvinyl pyrrolidone (PVP). Infrared spectroscopy was used to understand the molecular association of binder domains. Granules were prepared by high-shear aqueous granulation from α-lactose monohydrate and PVP K29/32 or K90. XMCT was used to visualise the granule microstructure, intra-granular binder distribution and measure intra-granular porosity, which was subsequently related to intrusion porosimetry measurements. Confocal Raman microscopy and infrared microscopy were employed to investigate the distribution of components within the granule and explore the nature of binder substrate interactions. XMCT data sets of internal granule microstructure provided values of residual porosity in the lactose:PVP K29/32 and lactose:PVP K90 granules of 32.41 ± 4.60% and 22.40 ± 0.03%, respectively. The binder volumes of the lactose:PVP K29/32 and lactose:PVP K90 granules were 2.98 ± 0.10% and 3.38 ± 0.07%, respectively, and were attributed to PVP-rich binder domains within the granule. Confocal Raman microscopy revealed anisotropic domains of PVP between 2 μm and 20 μm in size surrounded by larger particles of lactose, in both granule types. Raman data showed that PVP domains contained various amounts of lactose, whilst IR microscopy determined that the PVP was molecularly associated with lactose, rather than residual water. The work shows that XMCT can be applied to investigate granular microstructure and resolve the porosity and the excipient and binder volumes. Combining this technique with vibrational techniques provides further structural information and aids the interpretations of the XMCT images. When used complementarily, these techniques highlighted that porosity and binder volume were the most significant microstructural differences between the α-lactose monohydrate granules formulated with the different grades of PVP. Copyright © 2010 Elsevier B.V. All rights reserved.

The functional morphology of color changing in a spider: development of ommochrome pigment granules.

PubMed

Insausti, Teresita C; Casas, Jérôme

2008-03-01

Studies on the formation of ommochrome pigment granules are very few, despite their generalized occurrence as screening pigments in insect eyes. This is particularly true for ommochrome granules responsible for epidermal coloration. The aims of this study were to characterize the localization of major body pigments in a color changing mimetic spider, Misumena vatia (Thomisidae), and to describe the formation and location of ommochrome pigment granules responsible for the spider's color change from white to yellow. The unpigmented cuticula of this spider is transparent. Both the guanine localized in guanine cells in the opisthosoma and the uric acid localized in epidermis cells in the prosoma are responsible for the white coloration. The bright yellow color is due to the combination of ommochrome pigment granules and the white reflectance from coincident guanine and/or uric acid. The formation of ommochrome pigment granules in epidermis cells proceeds via three distinctive steps. Translucent, UV fluorescent, progranules (type I) are produced by a dense network of endoplasmic reticulum associated with numerous mitochondria and glycogen rosettes. These progranules are present in white spiders only, and regularly distributed in the cytoplasm. The merging of several progranules of type I into a transient state (progranule type II) leads to the formation of granules (type III) characterized by their lack of fluorescence, their spherical sections and their osmophilic-electron-dense contents. They are found in yellow spiders and in the red stripes on the body sides. Their color varies from yellow to red. Thus, white spiders contain only type I granules, yellow tinted spiders contain type II and III granules and bright yellow spiders contain only type III granules. We present a synthetic view of the ontogeny of ommochrome granules. We discuss the physiology of color changing and the nature of the chemical compounds in the different types of granules. Extended studies on the ultrastructural modification and physiological processes associated with color change are required before any statement about the adaptiveness of the color change can be made.

[Comparative analysis between origin of cooked traditional Chinese medicine powder and modern formula granules].

PubMed

Li, Rui; Zhai, Hua-Qiang; Tian, Wei-Lan; Hou, Ji-Ru; Jin, Shi-Yuan; Wang, Yong-Yan

2016-03-01

In this study, the origin and causes of cooked traditional Chinese medicine powder were reviewed, and a comprehensive analysis was made for the time background of modern traditional Chinese medicine formula granules and the future development trend, in order to provide reference for application and promotion of traditional Chinese medicine formula granules. By reference to ancient medical books of previous dynasties, a system review was conducted for infancy, formation, maturity and transition of cooked traditional Chinese medicine powder, and a comprehensive analysis was made for the six factors of cooked traditional Chinese medicine powder's maturity in the Song Dynasty. Efforts were made to collect domestic and foreign research literatures of modern formula granules, understand the detailed development, and conduct an objective analysis of the current clinical application of modern formula granules. According to the comparative analysis for the application characteristics of cooked traditional Chinese medicine powder and modern formula granules, ①the popularity of cooked traditional Chinese medicine powder in the Song Dynasty has six factors: soaring numbers of medical students and medical practitioners, high medical expenses due to huge army, rapid population growth, frequent epidemics and increasing diseases, and insufficient finances of central and local governments. ②On the basis of clinical application characteristics of traditional Chinese medicine formula granules, traditional Chinese medicine formula granules contain extracted and concentrated effective components, which guarantee the curative effect, meet modern people's demands for "quick, simple and convenience" traditional Chinese medicine decoctions, show a relatively high cost performance; however, formula granules are restricted by their varieties and lack unified quality control standards, and single-extract formula granules have not synergy and attenuation effects of combined traditional Chinese medicine decoctions, which also restricts its clinical application and promotion. ③Both have advantages in the process of clinical application, and shall be used based on syndromes. In conclusion, traditional Chinese medicine formula granules do not have disadvantages of "difficult, complicated, turbid and disorderly" cooked traditional Chinese medicine powder, and solve such problems as "inflexibility, expensiveness, restriction, disorder and inefficacy", which is the important basis for promoting traditional Chinese medicine formula granules. Copyright© by the Chinese Pharmaceutical Association.

Plant RNA Regulatory Network and RNA Granules in Virus Infection.

PubMed

Mäkinen, Kristiina; Lõhmus, Andres; Pollari, Maija

2017-01-01

Regulation of post-transcriptional gene expression on mRNA level in eukaryotic cells includes translocation, translation, translational repression, storage, mRNA decay, RNA silencing, and nonsense-mediated decay. These processes are associated with various RNA-binding proteins and cytoplasmic ribonucleoprotein complexes many of which are conserved across eukaryotes. Microscopically visible aggregations formed by ribonucleoprotein complexes are termed RNA granules. Stress granules where the translationally inactive mRNAs are stored and processing bodies where mRNA decay may occur present the most studied RNA granule types. Diverse RNP-granules are increasingly being assigned important roles in viral infections. Although the majority of the molecular level studies on the role of RNA granules in viral translation and replication have been conducted in mammalian systems, some studies link also plant virus infection to RNA granules. An increasing body of evidence indicates that plant viruses require components of stress granules and processing bodies for their replication and translation, but how extensively the cellular mRNA regulatory network is utilized by plant viruses has remained largely enigmatic. Antiviral RNA silencing, which is an important regulator of viral RNA stability and expression in plants, is commonly counteracted by viral suppressors of RNA silencing. Some of the RNA silencing suppressors localize to cellular RNA granules and have been proposed to carry out their suppression functions there. Moreover, plant nucleotide-binding leucine-rich repeat protein-mediated virus resistance has been linked to enhanced processing body formation and translational repression of viral RNA. Many interesting questions relate to how the pathways of antiviral RNA silencing leading to viral RNA degradation and/or repression of translation, suppression of RNA silencing and viral RNA translation converge in plants and how different RNA granules and their individual components contribute to these processes. In this review we discuss the roles of cellular RNA regulatory mechanisms and RNA granules in plant virus infection in the light of current knowledge and compare the findings to those made in animal virus studies.

rab3 mediates cortical granule exocytosis in the sea urchin egg.

PubMed

Conner, S; Wessel, G M

1998-11-15

Egg activation at fertilization in the sea urchin results in the exocytosis of approximately 15,000 cortical granules that are docked at the plasma membrane. Previously, we reported that several integral membrane proteins modeled in the SNARE hypothesis, synaptotagmin, VAMP, and syntaxin, in addition to a small GTPase of the ras superfamily, rab3, were present on cortical granules (Conner, S., Leaf, D., and Wessel, G., Mol. Reprod. Dev. 48, 1-13, 1997). Here we report that rab3 is associated with cortical granules throughout oogenesis, during cortical granule translocation, and while docked at the egg plasma membrane. Following cortical granule exocytosis, however, rab3 reassociates with a different population of vesicles, at least some of which are of endocytic origin. Because of its selective association with cortical granules in eggs and oocytes, we hypothesize that rab3 functions in cortical granule exocytosis. To test this hypothesis, we used a strategy of interfering with rab3 function by peptide competition with its effector domain, a conserved region within specific rab types. We first identified the effector domain sequence in Lytechinus variegatus eggs and find the sequence 94% identical to the effector domain of rab3 in Stronglocentrotus purpuratus. Then, with synthetic peptides to different regions of the rab3 protein, we find that cortical granule exocytosis is inhibited in eggs injected with effector domain peptides, but not with peptides from the hypervariable region or with a scrambled effector peptide. Additionally, effector-peptide-injected eggs injected with IP3 are blocked in their ability to exocytose cortical granules, suggesting that the inhibition is directly on the membrane fusion event and not the result of interference with the signal transduction mechanism leading to calcium release. We interpret these results to mean that rab3 functions in the regulation of cortical granule exocytosis following vesicle docking. Copyright 1998 Academic Press.

Morphologic characterization of specific granules in Greyhound eosinophils.

PubMed

Iazbik, M C; Couto, C G

2005-06-01

"Vacuolated" eosinophils (ie, eosinophils with empty, nonstaining granules) have been described previously in normal Greyhounds. However, to our knowledge, detailed studies of granules in vacuolated and normal eosinophils in this breed have not been performed. The objective of this prospective study was to characterize some of the morphologic, ultrastructural, and cytochemical staining features of specific (primary) granules in both normal and vacuolated eosinophils in Greyhound blood. Morphologic features of eosinophils in Wright's- and Diff-Quik-stained peripheral blood smears from 49 Greyhounds were compared with 200 blood smears from non-Greyhound dogs. Transmission electron microscopy was done on blood from 3 Greyhounds with vacuolated eosinophils and 3 with normal eosinophil granules. Blood smears from 4 of these dogs also were stained cytochemically with alkaline phosphatase (AP), chloracetate esterase (CAE), and alpha naphthyl butyrate esterase (ANBE). The morphologic features and tinctorial properties of vacuolated and normal eosinophils were compared. Twenty-six Greyhounds (53%) had vacuolated eosinophils and 23 (47%) had normal granulated eosinophils in smears stained with Wright's stain. Only 1% of eosinophils were vacuolated in non-Greyhound dogs. Twenty of the 23 (85%) Greyhounds with normal granulated eosinophils on Wright's-stained smears had vacuolated eosinophils in smears stained with Diff-Quik. Ultrastructurally, no morphologic differences were observed between granules of vacuolated and normal eosinophils. Both vacuolated and normal eosinophils in Greyhounds were positive for AP and negative for CAE and ANBE, as expected for normal dogs. Vacuolated eosinophils in Greyhounds likely reflect, at least in part, differential staining properties of the specific granules with different hematologic stains. Ultrastuctural and cytochemical features of eosinophil granules were similar in normal and vacuolated eosinophils from Greyhounds.

The effects of exercise and stress on the survival and maturation of adult-generated granule cells

PubMed Central

Snyder, Jason S.; Glover, Lucas R.; Sanzone, Kaitlin M.; Kamhi, J. Frances; Cameron, Heather A.

2009-01-01

Stress strongly inhibits proliferation of granule cell precursors in the dentate gyrus, while voluntary running has the opposite effect. Few studies, however, have examined the possible effects of these environmental manipulations on the maturation and survival of young granule cells. We examined number of surviving granule cells and the proportion of young neurons that were functionally mature, as defined by seizure-induced immediate-early gene expression, in 14 and 21 day-old granule cells in mice that were given access to a running wheel, restrained daily for 2 hours, or given no treatment during this period. Importantly, treatments began two days after BrdU injection, to isolate effects on survival from those on cell proliferation. We found a large increase in granule cell survival in running mice compared with controls at both time points. In addition, running increased the proportion of granule cells expressing the immediate-early gene Arc in response to seizures, suggesting that it speeds incorporation into circuits, i.e., functional maturation. Stressed mice showed no change in Arc expression, compared to control animals, but, surprisingly, showed a transient increase in survival of 14-day-old granule cells, which was gone 7 days later. Examination of cell proliferation, using the endogenous mitotic marker proliferating cell nuclear antigen (PCNA) showed an increase in cell proliferation after 12 days of running but not after 19 days of running. The number of proliferating cells was unchanged 24 hours after the 12th or 19th episode of daily restraint stress. These findings demonstrate that running has strong effects on survival and maturation of young granule cells as well as their birth and that stress can have positive but short-lived effects on granule cell survival. PMID:19156854

Acoustic emission monitoring from a lab scale high shear granulator--a novel approach.

PubMed

Watson, N J; Povey, M J W; Reynolds, G K; Xu, B H; Ding, Y

2014-04-25

A new approach to the monitoring of granulation processes using passive acoustics together with precise control over the granulation process has highlighted the importance of particle-particle and particle-bowl collisions in acoustic emission. The results have shown that repeatable acoustic results could be obtained but only when a spray nozzle water addition system was used. Acoustic emissions were recorded from a transducer attached to the bowl and an airborne transducer. It was found that the airborne transducer detected very little from the granulation and only experienced small changes throughout the process. The results from the bowl transducer showed that during granulation the frequency content of the acoustic emission shifted towards the lower frequencies. Results from the discrete element model indicate that when larger particles are used the number of collisions the particles experience reduces. This is a result of the volume conservation methodology used in this study, therefore larger particles results in less particles. These simulation results coupled with previous theoretical work on the frequency content of an impacting sphere explain why the frequency content of the acoustic emissions reduces during granule growth. The acoustic system used was also clearly able to identify when large over-wetted granules were present in the system, highlighting its benefit for detecting undesirable operational conditions. High-speed photography was used to study if visual changes in the granule properties could be linked with the changing acoustic emissions. The high speed photography was only possible towards the latter stages of the granulation process and it was found that larger granules produced a higher magnitude of acoustic emission across a broader frequency range. Copyright © 2014 Elsevier B.V. All rights reserved.

Development of an in-line Raman spectroscopic method for continuous API quantification during twin-screw wet granulation.

PubMed

Harting, Julia; Kleinebudde, Peter

2018-04-01

Raman spectroscopy was evaluated as a process analytical technology (PAT) tool for continuous API quantification during twin-screw wet granulation. Therefore, a Raman probe was implemented in front of the granulator barrel. This setup enabled the collection of Raman spectra upon a constant granule flow. To develop an in-line PLS calibration model, eight binary mixtures of the API and lactose monohydrate with API contents between 5 and 50% were pre-blended and granulated in a twin-screw granulator with a screw speed of 150 rpm and a powder feed rate of 40 g/min. Water was used as a granulation liquid with different liquid to solid ratios depending on the API content. Ibuprofen and diclofenac sodium were chosen as model drugs and separated PLS models were built for each API. The predictive performance of the developed PLS models was determined by granulating and monitoring new test samples containing different API concentrations. This evaluation showed that the models were able to predict the API concentration with an RMSEP of 0.59% for ibuprofen and 1.5% for diclofenac sodium. In a second part, the developed in-line Raman spectroscopic method was used to determine the API concentration during a split feeding process. Therefore, the API and lactose monohydrate were added by two independently adjustable feeders into the twin-screw granulator barrel. The in-line spectroscopy analysis which was verified by UV-analysis indicated that the mixing ability of the twin-screw granulator was good for the used settings and all adjusted API concentrations. Copyright © 2018 Elsevier B.V. All rights reserved.

Determination of hydrophobicity of dry-heated wheat starch granules using sucrose fatty acid esters (SFAE).

PubMed

Tabara, Aya; Oneda, Hiroshi; Murayama, Ryuji; Matsui, Yuko; Hirano, Akira; Seguchi, Masaharu

2014-01-01

Sucrose fatty acid esters (SFAE) were adsorbed onto dry-heated (120 °C for 10, 20, 40, 60, and 120 min) wheat starch granules and extracted with ethyl ether in a Soxhlet apparatus without gelatinization of the starch granules. The amount of sucrose in the extracted SFAE was determined by the phenol sulfate method. A gradual increase of the sucrose from 159 to 712 μg, in SFAE per gram of starch, occurred with increasing dry-heating time and demonstrated the increased hydrophobicity of the starch granules. Increase of the SFAE was highly correlated (r = 0.9816) to increase of the oil-binding capacity of the dry-heated wheat starch granules. Non-waxy rice, waxy rice, sweet potato, and potato starch granules also showed higher hydrophobicity after dry-heating by this method.

Obtaining of granular fertilizers based on ashes from combustion of waste residues and ground bones using phosphorous solubilization by bacteria Bacillus megaterium.

PubMed

Rolewicz, M; Rusek, P; Borowik, K

2018-06-15

The article presents research results on obtaining phosphorus granulated fertilizers on the basis of microbiologically activated sewage sludge ashes, ground bones and dried blood from meat industry. Granulation tests were carried out using a laboratory pan granulator as well as on an experimental pilot plant. The aim of the studies was to select the proper composition of the mixture of raw materials and binding agents to obtain granulated fertilizers from waste materials such as MSSA and MBM and bacteria lyophilisate. Obtained fertilizer samples were subjected to physical tests (granulation tests etc.) and quality assessment. The tests confirmed that it was possible to produce granulated phosphate fertilizers using the Bacillus megaterium for solubilization of phosphorus in a simple process. Copyright © 2017 Elsevier Ltd. All rights reserved.

The Impact of Granule Density on Tabletting and Pharmaceutical Product Performance.

PubMed

van den Ban, Sander; Goodwin, Daniel J

2017-05-01

The impact of granule densification in high-shear wet granulation on tabletting and product performance was investigated, at pharmaceutical production scale. Product performance criteria need to be balanced with the need to deliver manufacturability criteria to assure robust industrial scale tablet manufacturing processes. A Quality by Design approach was used to determine in-process control specifications for tabletting, propose a design space for disintegration and dissolution, and to understand the permitted operating limits and required controls for an industrial tabletting process. Granules of varying density (filling density) were made by varying water amount added, spray rate, and wet massing time in a design of experiment (DoE) approach. Granules were compressed into tablets to a range of thicknesses to obtain tablets of varying breaking force. Disintegration and dissolution performance was evaluated for the tablets made. The impact of granule filling density on tabletting was rationalised with compressibility, tabletability and compactibility. Tabletting and product performance criteria provided competing requirements for porosity. An increase in granule filling density impacted tabletability and compactability and limited the ability to achieve tablets of adequate mechanical strength. An increase in tablet solid fraction (decreased porosity) impacted disintegration and dissolution. An attribute-based design space for disintegration and dissolution was specified to achieve both product performance and manufacturability. The method of granulation and resulting granule filling density is a key design consideration to achieve both product performance and manufacturability required for modern industrial scale pharmaceutical product manufacture and distribution.

Granule mobility, fusion frequency and insulin secretion are differentially affected by insulinotropic stimuli.

PubMed

Schumacher, Kirstin; Matz, Magnus; Brüning, Dennis; Baumann, Knut; Rustenbeck, Ingo

2015-05-01

The pre-exocytotic behavior of insulin granules was studied against the background of the entirety of submembrane granules in MIN6 cells, and the characteristics were compared with the macroscopic secretion pattern and the cytosolic Ca(2+) concentration of MIN6 pseudo-islets at 22°C, 32°C and 37°C. The mobility of granules labeled by insulin-EGFP and the fusion events were assessed by TIRF microscopy utilizing an observer-independent algorithm. In the z-dimension, 40 mm K(+) or 30 mm glucose increased the granule turnover. The effect of high K(+) was quickly reversible. The increase by glucose was more sustained and modified the efficacy of a subsequent K(+) stimulus. The effect size of glucose increased with physiological temperature whereas that of high K(+) did not. The mobility in the x/y-dimension and the fusion rates were little affected by the stimuli, in contrast to secretion. Fusion and secretion, however, had the same temperature dependence. Granules that appeared and fused within one image sequence had significantly larger caging diameters than pre-existent granules that underwent fusion. These in turn had a different mobility than residence-matched non-fusing granules. In conclusion, delivery to the membrane, tethering and fusion of granules are differently affected by insulinotropic stimuli. Fusion rates and secretion do not appear to be tightly coupled. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

[Effects of carbon sources changes on the property and morphology of 2,4-D degraded aerobic sludge granules].

PubMed

Ma, Jing-Yun; Quan, Xian-Chun; Xiong, Wei-Cong

2010-11-01

This study investigated the changes of the morphology, structure, and capability of removing the target contamination of the aerobic granules pre-cultured with mixed substrates of glucose and 2,4-dichlorophenoxyacetic acid (2,4-D) in a long-time running sequence batch reactor (SBR), when the carbon source transformed into the sole carbon source of 2,4-D. Results showed that when the substrate turned to the sole carbon source of 2,4-D, the aerobic granules still maintained a strong degradation ability to the target contamination; a 2,4-D removal percentage of 99.2% -100% and an average COD removal rate of 85.6% were achieved at the initial 2,4-D concentration of 361-564 mg/L. Carbon source transformation caused certain damages to the original aerobic granule structure, made some parts of granules disintegrated, and led to granule size decline from 513 microm to 302 microm. However, those granules maintained the main body, re-aggregated and grew after a period of adaptation due to their strong resistance to toxicity. Aerobic granules capable of utilizing 2,4-D as the sole carbon source with a good settling ability (SYI 20-40 mL/g) and a mean diameter of 489 microm were finally obtained in this study. Scanning electron microscope (SEM) observation showed that the diversity of granule microbial species was declined when turned to the sole carbon source.

Development and Optimization of a Wet Granulation Process at Elevated Temperature for a Poorly Compactible Drug Using Twin Screw Extruder for Continuous Manufacturing.

PubMed

Meena, Anuprabha K; Desai, Divyakant; Serajuddin, Abu T M

2017-02-01

The objective of this study was to enhance tabletability of a poorly compactible drug, acetaminophen, by wet granulation using twin screw extruder at high temperature. It was desired that there would be minimum amounts of excipients used and the granules obtained after extrusion would be dry and fall within a size range suitable for tableting without any further processing. Mixtures of acetaminophen (95%) with binders (5% povidone or partially pregelatinized starch) were wet granulated through twin screw extruder at 70°C by adding 7% w/w water. The process had a short granulation time (<1 min), and, on account of the elevated processing temperature used, no drying after extrusion was needed. By optimizing formulation and processing parameters, >90% granules in the size range of 125 to 1000 μm (<3% above 1000 μm and <7% below 125 μm) were obtained without any milling. When the granules were compressed by adding 1% disintegrant and 0.5% lubricant extragranularly, tablets produced (93.6% drug load) had good mechanical strength having hardness >1.7 MPa, which was superior to that of tablets prepared by conventional high shear wet granulation. As the granules could be extruded continuously and did not require drying and milling, the method was amenable to continuous processing. Copyright © 2017 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Evaluation of various processes for simultaneous complexation and granulation to incorporate drug-cyclodextrin complexes into solid dosage forms.

PubMed

Gyanani, Vijay; Siddalingappa, Basavaraj; Betageri, Guru V

2015-01-01

Insoluble drugs often formulated with various excipients to enhance the dissolution. Cyclodextrins (CDs) are widely used excipients to improve dissolution profile of poorly soluble drugs. Drug-CD complexation process is complex and often requires multiple processes to produce solid dosage form. Hence, this study explored commonly used granulation processes for simultaneous complexation and granulation. Poorly soluble drugs ibuprofen and glyburide were selected as experimental drugs. Co-evaporation of drug:CD mixture from a solvent followed by wet granulation with water was considered as standard process for comparison. Spray granulation and fluid bed processing (FBP) using drug:CD solution in ethanol were evaluated as an alternative processes. The dissolution data of glyburide tablets indicated that tablets produced by spray granulation, FBP and co-evaporation-granulation have almost identical dissolution profile in water and 0.1% SLS (>70% in water and >60% in SLS versus 30 and 34%, respectively for plain tablet, in 120 min). Similarly, ibuprofen:CD tablets produced by co-evaporation-granulation and FBP displayed similar dissolution profile in 0.01 M HCl (pH 2.0) and buffer pH 5.5 (>90 and 100% versus 44 and 80% respectively for plain tablets, 120 min). Results of this study demonstrated that spray granulation is simple and cost effective process for low dose poorly soluble drugs to incorporate drug:CD complex into solid dosage form, whereas FBP is suitable for poorly soluble drugs with moderate dose.

Cerebellar granule cells encode the expectation of reward

PubMed Central

Wagner, Mark J; Kim, Tony Hyun; Savall, Joan; Schnitzer, Mark J; Luo, Liqun

2017-01-01

The human brain contains ~60 billion cerebellar granule cells1, which outnumber all other neurons combined. Classical theories posit that a large, diverse population of granule cells allows for highly detailed representations of sensorimotor context, enabling downstream Purkinje cells to sense fine contextual changes2–6. Although evidence suggests a role for cerebellum in cognition7–10, granule cells are known to encode only sensory11–13 and motor14 context. Using two-photon calcium imaging in behaving mice, here we show that granule cells convey information about the expectation of reward. Mice initiated voluntary forelimb movements for delayed water reward. Some granule cells responded preferentially to reward or reward omission, whereas others selectively encoded reward anticipation. Reward responses were not restricted to forelimb movement, as a Pavlovian task evoked similar responses. Compared to predictable rewards, unexpected rewards elicited markedly different granule cell activity despite identical stimuli and licking responses. In both tasks, reward signals were widespread throughout multiple cerebellar lobules. Tracking the same granule cells over several days of learning revealed that cells with reward-anticipating responses emerged from those that responded at the start of learning to reward delivery, whereas reward omission responses grew stronger as learning progressed. The discovery of predictive, non-sensorimotor encoding in granule cells is a major departure from current understanding of these neurons and dramatically enriches contextual information available to postsynaptic Purkinje cells, with important implications for cognitive processing in the cerebellum. PMID:28321129

A Genomewide RNAi Screen for Genes That Affect the Stability, Distribution and Function of P Granules in Caenorhabditis elegans

PubMed Central

Updike, Dustin L.; Strome, Susan

2009-01-01

P granules are non-membrane-bound organelles found in the germ-line cytoplasm throughout Caenorhabditis elegans development. Like their “germ granule” counterparts in other animals, P granules are thought to act as determinants of the identity and special properties of germ cells, properties that include the unique ability to give rise to all tissues of future generations of an organism. Therefore, understanding how P granules work is critical to understanding how cellular immortality and totipotency are retained, gained, and lost. Here we report on a genomewide RNAi screen in C. elegans, which identified 173 genes that affect the stability, localization, and function of P granules. Many of these genes fall into specific classes with shared P-granule phenotypes, allowing us to better understand how cellular processes such as protein degradation, translation, splicing, nuclear transport, and mRNA homeostasis converge on P-granule assembly and function. One of the more striking phenotypes is caused by the depletion of CSR-1, an Argonaute associated with an endogenous siRNA pathway that functions in the germ line. We show that CSR-1 and two other endo-siRNA pathway members, the RNA-dependent RNA polymerase EGO-1 and the helicase DRH-3, act to antagonize RNA and P-granule accumulation in the germ line. Our findings strengthen the emerging view that germ granules are involved in numerous aspects of RNA metabolism, including an endo-siRNA pathway in germ cells. PMID:19805813

Investigating the Use of Polymeric Binders in Twin Screw Melt Granulation Process for Improving Compactibility of Drugs.

PubMed

Batra, Amol; Desai, Dipen; Serajuddin, Abu T M

2017-01-01

Traditionally, the melt granulation for pharmaceutical products was performed at low temperature (<90°C) with high-shear granulators using low-melting waxy binders, and tablets produced using such granules were not amenable to large-scale manufacturing. The situation has changed in recent years by the use of twin screw extruder where the processing temperature could be increased to as high as 180°C and polymers with high T g could be used as binders. In this study, different polymeric binders were screened for their suitability in improving compactibility of 2 drugs, metformin hydrochloride and acetaminophen, by twin screw melt granulation. Processing temperatures for the 2 drugs were set at 180°C and 130°C, respectively. Screw configuration, screw speed, and feed rate were optimized such that all polymeric binders used produced granules. Several hydroxypropyl cellulose, hydroxypropyl methylcellulose, polyvinylpyrrolidone, and methacrylate-based polymers, including Klucel ® EXF, Eudragit ® EPO, and Soluplus ® , demonstrated good tablet tensile strength (>2 MPa) when granules were produced using only 10% wt/wt polymer concentration. Certain polymers provided acceptable compactibility even at 5% wt/wt. Thus, twin screw melt granulation process may be used with different polymers at a wide range of temperature. Due to low excipient concentration, this granulation method is especially suitable for high-dose tablets. Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

Light scattering on PHA granules protects bacterial cells against the harmful effects of UV radiation.

PubMed

Slaninova, Eva; Sedlacek, Petr; Mravec, Filip; Mullerova, Lucie; Samek, Ota; Koller, Martin; Hesko, Ondrej; Kucera, Dan; Marova, Ivana; Obruca, Stanislav

2018-02-01

Numerous prokaryotes accumulate polyhydroxyalkanoates (PHA) in the form of intracellular granules. The primary function of PHA is the storage of carbon and energy. Nevertheless, there are numerous reports that the presence of PHA granules in microbial cells enhances their stress resistance and fitness when exposed to various stress factors. In this work, we studied the protective mechanism of PHA granules against UV irradiation employing Cupriavidus necator as a model bacterial strain. The PHA-accumulating wild type strain showed substantially higher UV radiation resistance than the PHA non-accumulating mutant. Furthermore, the differences in UV-Vis radiation interactions with both cell types were studied using various spectroscopic approaches (turbidimetry, absorption spectroscopy, and nephelometry). Our results clearly demonstrate that intracellular PHA granules efficiently scatter UV radiation, which provides a substantial UV-protective effect for bacterial cells and, moreover, decreases the intracellular level of reactive oxygen species in UV-challenged cells. The protective properties of the PHA granules are enhanced by the fact that granules specifically bind to DNA, which in turn provides shield-like protection of DNA as the most UV-sensitive molecule. To conclude, the UV-protective action of PHA granules adds considerable value to their primary storage function, which can be beneficial in numerous environments.

Process Recovery after CaO Addition Due to Granule Formation in a CSTR Co-Digester—A Tool to Influence the Composition of the Microbial Community and Stabilize the Process?

PubMed Central

Liebrich, Marietta; Kleyböcker, Anne; Kasina, Monika; Miethling-Graff, Rona; Kassahun, Andrea; Würdemann, Hilke

2016-01-01

The composition, structure and function of granules formed during process recovery with calcium oxide in a laboratory-scale fermenter fed with sewage sludge and rapeseed oil were studied. In the course of over-acidification and successful process recovery, only minor changes were observed in the bacterial community of the digestate, while granules appeared during recovery. Fluorescence microscopic analysis of the granules showed a close spatial relationship between calcium and oil and/or long chain fatty acids. This finding further substantiated the hypothesis that calcium precipitated with carbon of organic origin and reduced the negative effects of overloading with oil. Furthermore, the enrichment of phosphate minerals in the granules was shown, and molecular biological analyses detected polyphosphate-accumulating organisms as well as methanogenic archaea in the core. Organisms related to Methanoculleus receptaculi were detected in the inner zones of a granule, whereas they were present in the digestate only after process recovery. This finding indicated more favorable microhabitats inside the granules that supported process recovery. Thus, the granule formation triggered by calcium oxide addition served as a tool to influence the composition of the microbial community and to stabilize the process after overloading with oil. PMID:27681911

Process Recovery after CaO Addition Due to Granule Formation in a CSTR Co-Digester-A Tool to Influence the Composition of the Microbial Community and Stabilize the Process?

PubMed

Liebrich, Marietta; Kleyböcker, Anne; Kasina, Monika; Miethling-Graff, Rona; Kassahun, Andrea; Würdemann, Hilke

2016-03-17

The composition, structure and function of granules formed during process recovery with calcium oxide in a laboratory-scale fermenter fed with sewage sludge and rapeseed oil were studied. In the course of over-acidification and successful process recovery, only minor changes were observed in the bacterial community of the digestate, while granules appeared during recovery. Fluorescence microscopic analysis of the granules showed a close spatial relationship between calcium and oil and/or long chain fatty acids. This finding further substantiated the hypothesis that calcium precipitated with carbon of organic origin and reduced the negative effects of overloading with oil. Furthermore, the enrichment of phosphate minerals in the granules was shown, and molecular biological analyses detected polyphosphate-accumulating organisms as well as methanogenic archaea in the core. Organisms related to Methanoculleus receptaculi were detected in the inner zones of a granule, whereas they were present in the digestate only after process recovery. This finding indicated more favorable microhabitats inside the granules that supported process recovery. Thus, the granule formation triggered by calcium oxide addition served as a tool to influence the composition of the microbial community and to stabilize the process after overloading with oil.

Granule fraction inhomogeneity of calcium carbonate/sorbitol in roller compacted granules.

PubMed

Bacher, C; Olsen, P M; Bertelsen, P; Sonnergaard, J M

2008-02-12

The granule fraction inhomogeneity of roller compacted granules was examined on mixtures of three different morphologic forms of calcium carbonate and three particle sizes of sorbitol. The granule fraction inhomogeneity was determined by the distribution of the calcium carbonate in each of the 10 size fractions between 0 and 2000 microm and by calculating the demixing potential. Significant inhomogeneous occurrence of calcium carbonate in the size fractions was demonstrated, depending mostly on the particles sizes of sorbitol but also on the morphological forms of calcium carbonate. The heterogeneous distribution of calcium carbonate was related to the decrease in compactibility of roller compacted granules in comparison to the ungranulated materials. This phenomenon was explained by a mechanism where fracturing of the ribbon during granulation occurred at the weakest interparticulate bonds (the calcium carbonate: calcium carbonate bonds) and consequently exposed the weakest areas of bond formation on the surface of the granules. Accordingly, the non-uniform allocation of the interparticulate attractive forces in a tablet would cause a lowering of the compactibility. Furthermore, the ability of the powder to agglomerate in the roller compactor was demonstrated to be related to the ability of the powder to be compacted into a tablet, thus the most compactable calcium carbonate and the smallest sized sorbitol improved the homogeneity by decreasing the demixing potential.

Polyhydroxyalkanoate (PHA) Granules Have no Phospholipids.

PubMed

Bresan, Stephanie; Sznajder, Anna; Hauf, Waldemar; Forchhammer, Karl; Pfeiffer, Daniel; Jendrossek, Dieter

2016-05-25

Polyhydroxybutyrate (PHB) granules, also designated as carbonosomes, are supra-molecular complexes in prokaryotes consisting of a PHB polymer core and a surface layer of structural and functional proteins. The presence of suspected phospholipids in the surface layer is based on in vitro data of isolated PHB granules and is often shown in cartoons of the PHB granule structure in reviews on PHB metabolism. However, the in vivo presence of a phospholipid layer has never been demonstrated. We addressed this topic by the expression of fusion proteins of DsRed2EC and other fluorescent proteins with the phospholipid-binding domain (LactC2) of lactadherin in three model organisms. The fusion proteins specifically localized at the cell membrane of Ralstonia eutropha but did not co-localize with PHB granules. The same result was obtained for Pseudomonas putida, a species that accumulates another type of polyhydroxyalkanoate (PHA) granules related to PHB. Notably, DsRed2EC-LactC2 expressed in Magnetospirillum gryphiswaldense was detected at the position of membrane-enclosed magnetosome chains and at the cytoplasmic membrane but not at PHB granules. In conclusion, the carbonosomes of representatives of α-proteobacteria, β-proteobacteria and γ-proteobacteria have no phospholipids in vivo and we postulate that the PHB/PHA granule surface layers in natural producers generally are free of phospholipids and consist of proteins only.

Application of quality by design concepts in the development of fluidized bed granulation and tableting processes.

PubMed

Djuris, Jelena; Medarevic, Djordje; Krstic, Marko; Djuric, Zorica; Ibric, Svetlana

2013-06-01

This study illustrates the application of experimental design and multivariate data analysis in defining design space for granulation and tableting processes. According to the quality by design concepts, critical quality attributes (CQAs) of granules and tablets, as well as critical parameters of granulation and tableting processes, were identified and evaluated. Acetaminophen was used as the model drug, and one of the study aims was to investigate the possibility of the development of immediate- or extended-release acetaminophen tablets. Granulation experiments were performed in the fluid bed processor using polyethylene oxide polymer as a binder in the direct granulation method. Tablets were compressed in the laboratory excenter tablet press. The first set of experiments was organized according to Plackett-Burman design, followed by the full factorial experimental design. Principal component analysis and partial least squares regression were applied as the multivariate analysis techniques. By using these different methods, CQAs and process parameters were identified and quantified. Furthermore, an in-line method was developed to monitor the temperature during the fluidized bed granulation process, to foresee possible defects in granules CQAs. Various control strategies that are based on the process understanding and assure desired quality attributes of the product are proposed. Copyright © 2013 Wiley Periodicals, Inc.

Granulation tissue of chronic pressure ulcers as a predictive indicator of wound closure.

PubMed

Wyffels, Jennifer T; Edsberg, Laura E

2011-10-01

: To describe the temporal relationship between the quantity of granulation tissue in a chronic pressure ulcer (PrU) and its clinical outcome. : Study participants were seen on days 0, 1, 2, 3, 4, 7, 8, 9, 10, 11, 14, 21, 28, 35, and 42. On each visit, the wounds were digitally photographed with a 3-cm calibration target. Images were analyzed using VeV MD (version 1.1.14; VERG Inc, Winnipeg, Manitoba, Canada) and Adobe Photoshop CS3 Extended (version 10.0.1; Adobe Systems Inc, San Jose, California). Granulation tissue was selected from calibrated digital images by 1 of 2 methods: manual selection and automated selection. Granulation tissue area was expressed as a percentage of total wound area. : Academic research laboratory. : Thirty-one chronic PrUs were observed in 27 subjects. : Quantitative measure of granulation tissue area. : There was no relationship between the amount of granulation tissue expressed as a percentage of the total PrU area and wound outcome. : This study is the first to both quantitatively measure the amount of granulation tissue in a chronic PrU and attempt to correlate it to wound outcome. Although counterintuitive, the amount of granulation tissue was not predictive of outcome, and no temporal trends could be described.

Pterin pigment granules are responsible for both broadband light scattering and wavelength selective absorption in the wing scales of pierid butterflies

PubMed Central

Morehouse, Nathan I; Vukusic, Peter; Rutowski, Ron

2006-01-01

A small but growing literature indicates that many animal colours are produced by combinations of structural and pigmentary mechanisms. We investigated one such complex colour phenotype: the highly chromatic wing colours of pierid butterflies including oranges, yellows and patterns which appear white to the human eye, but strongly absorb the ultraviolet (UV) wavelengths visible to butterflies. Pierids produce these bright colours using wing scales that contain collections of minute granules. However, to date, no work has directly characterized the molecular composition or optical properties of these granules. We present results that indicate these granules contain pterin pigments. We also find that pterin granules increase light reflection from single wing scales, such that wing scales containing denser granule arrays reflect more light than those with less dense granule collections. As male wing scales contain more pterin granules than those of females, the sexual dichromatism found in many pierid species can be explained by differences in wing scale pterin deposition. Additionally, the colour pattern elements produced by these pterins are known to be important during mating interactions in a number of pierid species. Therefore, we discuss the potential relevance of our results within the framework of sexual selection and colour signal evolution. PMID:17164199

Optimization of critical quality attributes in continuous twin-screw wet granulation via design space validated with pilot scale experimental data.

PubMed

Liu, Huolong; Galbraith, S C; Ricart, Brendon; Stanton, Courtney; Smith-Goettler, Brandye; Verdi, Luke; O'Connor, Thomas; Lee, Sau; Yoon, Seongkyu

2017-06-15

In this study, the influence of key process variables (screw speed, throughput and liquid to solid (L/S) ratio) of a continuous twin screw wet granulation (TSWG) was investigated using a central composite face-centered (CCF) experimental design method. Regression models were developed to predict the process responses (motor torque, granule residence time), granule properties (size distribution, volume average diameter, yield, relative width, flowability) and tablet properties (tensile strength). The effects of the three key process variables were analyzed via contour and interaction plots. The experimental results have demonstrated that all the process responses, granule properties and tablet properties are influenced by changing the screw speed, throughput and L/S ratio. The TSWG process was optimized to produce granules with specific volume average diameter of 150μm and the yield of 95% based on the developed regression models. A design space (DS) was built based on volume average granule diameter between 90 and 200μm and the granule yield larger than 75% with a failure probability analysis using Monte Carlo simulations. Validation experiments successfully validated the robustness and accuracy of the DS generated using the CCF experimental design in optimizing a continuous TSWG process. Copyright © 2017 Elsevier B.V. All rights reserved.

Atomic force microscopy of pea starch: origins of image contrast.

PubMed

Ridout, Michael J; Parker, Mary L; Hedley, Cliff L; Bogracheva, Tatiana Y; Morris, Victor J

2004-01-01

Atomic force microscopy (AFM) has been used to image the internal structure of pea starch granules. Starch granules were encased in a nonpenetrating matrix of rapid-set Araldite. Images were obtained of the internal structure of starch exposed by cutting the face of the block and of starch in sections collected on water. These images have been obtained without staining, or either chemical or enzymatic treatment of the granule. It has been demonstrated that contrast in the AFM images is due to localized absorption of water within specific regions of the exposed fragments of the starch granules. These regions swell, becoming "softer" and higher than surrounding regions. The images obtained confirm the "blocklet model" of starch granule architecture. By using topographic, error signal and force modulation imaging modes on samples of the wild-type pea starch and the high amylose r near-isogenic mutant, it has been possible to demonstrate differing structures within granules of different origin. These architectural changes provide a basis for explaining the changed appearance and functionality of the r mutant. The growth-ring structure of the granule is suggested to arise from localized "defects" in blocklet distribution within the granule. It is proposed that these defects are partially crystalline regions devoid of amylose.

Carbon granule probe microphone for leak detection. [recovery boilers

NASA Technical Reports Server (NTRS)

Parthasarathy, S. P. (Inventor)

1985-01-01

A microphone which is not subject to corrosion is provided by employing carbon granules to sense sound waves. The granules are packed into a ceramic tube and no diaphragm is used. A pair of electrodes is located in the tube adjacent the carbon granules and are coupled to a sensing circuit. Sound waves cause pressure changes on the carbon granules which results in a change in resistance in the electrical path between the electrodes. This change in resistance is detected by the sensing circuit. The microphone is suitable for use as a leak detection probe in recovery boilers, where it provides reliable operation without corrosion problems associated with conventional microphones.

Preliminary report of the discovery of a new pharmaceutical granulation process using foamed aqueous binders.

PubMed

Keary, Colin M; Sheskey, Paul J

2004-09-01

Spray granulation is commonly used to improve the flow of drug formulation powders by adding liquid binders. We have discovered a new granulation process whereby liquid binders are added as aqueous foam. Initial experiments indicate that foam granulations require less binder than spray granulations, less water is added to the powder mass, rates of addition of foam can be greater than rates of addition of sprayed liquids, and foam can be added in a single batch to the surface of the powder mass for incorporation at some later stage in the process. This new process appears to have no detrimental effects on granulate, tablet, or in vitro drug dissolution properties. In addition, the elimination of spray addition reduces the complexity of the process and avoids the plugging problems associated with spray nozzles. Several formulations were successfully scaled up from laboratory scale (1.5 kg) to pilot scale (15 kg). Process control was good and there was no detrimental effect on tablet and drug dissolution properties. This paper also proposes a working hypothesis of the mechanism by which foam granulation operates.

TIA-1 Self-Multimerization, Phase Separation, and Recruitment into Stress Granules Are Dynamically Regulated by Zn2.

PubMed

Rayman, Joseph B; Karl, Kevin A; Kandel, Eric R

2018-01-02

Stress granules are non-membranous structures that transiently form in the cytoplasm during cellular stress, where they promote translational repression of non-essential RNAs and modulate cell signaling by sequestering key signal transduction proteins. These and other functions of stress granules facilitate an adaptive cellular response to environmental adversity. A key component of stress granules is the prion-related RNA-binding protein, T cell intracellular antigen-1 (TIA-1). Here, we report that recombinant TIA-1 undergoes rapid multimerization and phase separation in the presence of divalent zinc, which can be reversed by the zinc chelator, TPEN. Similarly, the formation and maintenance of TIA-1-positive stress granules in arsenite-treated cells are inhibited by TPEN. In addition, Zn 2+ is released in cells treated with arsenite, before stress granule formation. These findings suggest that Zn 2+ is a physiological ligand of TIA-1, acting as a stress-inducible second messenger to promote multimerization of TIA-1 and subsequent localization into stress granules. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Snapin mediates insulin secretory granule docking, but not trans-SNARE complex formation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Somanath, Sangeeta; Partridge, Christopher J.; Marshall, Catriona

Secretory granule exocytosis is a tightly regulated process requiring granule targeting, tethering, priming, and membrane fusion. At the heart of this process is the SNARE complex, which drives fusion through a coiled-coil zippering effect mediated by the granule v-SNARE protein, VAMP2, and the plasma membrane t-SNAREs, SNAP-25 and syntaxin-1A. Here we demonstrate that in pancreatic β-cells the SNAP-25 accessory protein, snapin, C-terminal H2 domain binds SNAP-25 through its N-terminal Sn-1 domain. Interestingly whilst snapin binds SNAP-25, there is only modest binding of this complex with syntaxin-1A under resting conditions. Instead synataxin-1A appears to be recruited in response to secretory stimulation.more » These results indicate that snapin plays a role in tethering insulin granules to the plasma membrane through coiled coil interaction of snapin with SNAP-25, with full granule fusion competency only resulting after subsequent syntaxin-1A recruitment triggered by secretory stimulation. - Highlights: • Snapin mediates granule docking. • Snapin binds SNAP-25. • SNARE complex forms downstream.« less

Two-compartmental population balance modeling of a pulsed spray fluidized bed granulation based on computational fluid dynamics (CFD) analysis.

PubMed

Liu, Huolong; Li, Mingzhong

2014-11-20

In this work a two-compartmental population balance model (TCPBM) was proposed to model a pulsed top-spray fluidized bed granulation. The proposed TCPBM considered the spatially heterogeneous granulation mechanisms of the granule growth by dividing the granulator into two perfectly mixed zones of the wetting compartment and drying compartment, in which the aggregation mechanism was assumed in the wetting compartment and the breakage mechanism was considered in the drying compartment. The sizes of the wetting and drying compartments were constant in the TCPBM, in which 30% of the bed was the wetting compartment and 70% of the bed was the drying compartment. The exchange rate of particles between the wetting and drying compartments was determined by the details of the flow properties and distribution of particles predicted by the computational fluid dynamics (CFD) simulation. The experimental validation has shown that the proposed TCPBM can predict evolution of the granule size and distribution within the granulator under different binder spray operating conditions accurately. Copyright © 2014 Elsevier B.V. All rights reserved.

Accelerating Aerobic Sludge Granulation by Adding Dry Sewage Sludge Micropowder in Sequencing Batch Reactors

PubMed Central

Li, Jun; Liu, Jun; Wang, Danjun; Chen, Tao; Ma, Ting; Wang, Zhihong; Zhuo, Weilong

2015-01-01

Micropowder (20–250 µm) made from ground dry waste sludge from a municipal sewage treatment plant was added in a sequencing batch reactor (R2), which was fed by synthetic wastewater with acetate as carbon source. Compared with the traditional SBR (R1), aerobic sludge granulation time was shortened 15 days in R2. Furthermore, filamentous bacteria in bulking sludge were controlled to accelerate aerobic granulation and form large granules. Correspondingly, the SVI decreased from 225 mL/g to 37 mL/g. X-ray Fluorescence (XRF) analysis demonstrated that Al and Si from the micropowder were accumulated in granules. A mechanism hypotheses for the acceleration of aerobic granulation by adding dry sludge micropowder is proposed: added micropowder acts as nuclei to induce bacterial attachment; dissolved matters from the micropowder increase abruptly the organic load for starved sludge to control overgrown filamentous bacteria as a framework for aggregation; increased friction from the movement of micropowder forces the filaments which extend outwards to shrink for shaping granules. PMID:26308025

Amyloid-like aggregation of provasopressin in diabetes insipidus and secretory granule sorting.

PubMed

Beuret, Nicole; Hasler, Franziska; Prescianotto-Baschong, Cristina; Birk, Julia; Rutishauser, Jonas; Spiess, Martin

2017-01-26

Aggregation of peptide hormone precursors in the trans-Golgi network is an essential process in the biogenesis of secretory granules in endocrine cells. It has recently been proposed that this aggregation corresponds to the formation of functional amyloids. Our previous finding that dominant mutations in provasopressin, which cause cell degeneration and diabetes insipidus, prevent native folding and produce fibrillar aggregates in the endoplasmic reticulum (ER) might thus reflect mislocalized amyloid formation by sequences that evolved to mediate granule sorting. Here we identified two sequences responsible for fibrillar aggregation of mutant precursors in the ER: the N-terminal vasopressin nonapeptide and the C-terminal glycopeptide. To test their role in granule sorting, the glycopeptide was deleted and/or vasopressin mutated to inactivate ER aggregation while still permitting precursor folding and ER exit. These mutations strongly reduced sorting into granules and regulated secretion in endocrine AtT20 cells. The same sequences - vasopressin and the glycopeptide - mediate physiological aggregation of the wild-type hormone precursor into secretory granules and the pathological fibrillar aggregation of disease mutants in the ER. These findings support the amyloid hypothesis for secretory granule biogenesis.

Regulation of RNA granule dynamics by phosphorylation of serine-rich, intrinsically disordered proteins in C. elegans

PubMed Central

Wang, Jennifer T; Smith, Jarrett; Chen, Bi-Chang; Schmidt, Helen; Rasoloson, Dominique; Paix, Alexandre; Lambrus, Bramwell G; Calidas, Deepika; Betzig, Eric; Seydoux, Geraldine

2014-01-01

RNA granules have been likened to liquid droplets whose dynamics depend on the controlled dissolution and condensation of internal components. The molecules and reactions that drive these dynamics in vivo are not well understood. In this study, we present evidence that a group of intrinsically disordered, serine-rich proteins regulate the dynamics of P granules in C. elegans embryos. The MEG (maternal-effect germline defective) proteins are germ plasm components that are required redundantly for fertility. We demonstrate that MEG-1 and MEG-3 are substrates of the kinase MBK-2/DYRK and the phosphatase PP2APPTR−½. Phosphorylation of the MEGs promotes granule disassembly and dephosphorylation promotes granule assembly. Using lattice light sheet microscopy on live embryos, we show that GFP-tagged MEG-3 localizes to a dynamic domain that surrounds and penetrates each granule. We conclude that, despite their liquid-like behavior, P granules are non-homogeneous structures whose assembly in embryos is regulated by phosphorylation. DOI: http://dx.doi.org/10.7554/eLife.04591.001 PMID:25535836

Proteins with CHADs (Conserved Histidine α-Helical Domains) Are Attached to Polyphosphate Granules In Vivo and Constitute a Novel Family of Polyphosphate-Associated Proteins (Phosins).

PubMed

Tumlirsch, Tony; Jendrossek, Dieter

2017-04-01

On the basis of bioinformatic evidence, we suspected that proteins with a CYTH ( Cy aB th iamine triphosphatase) domain and/or a CHAD ( c onserved h istidine α -helical d omain) motif might represent polyphosphate (polyP) granule-associated proteins. We found no evidence of polyP targeting by proteins with CYTH domains. In contrast, two CHAD motif-containing proteins from Ralstonia eutropha H16 (A0104 and B1017) that were expressed as fusions with enhanced yellow fluorescent protein (eYFP) colocalized with polyP granules. While the expression of B1017 was not detectable, the A0104 protein was specifically identified in an isolated polyP granule fraction by proteome analysis. Moreover, eYFP fusions with the CHAD motif-containing proteins MGMSRV2-1987 from Magnetospirillum gryphiswaldense and PP2307 from Pseudomonas putida also colocalized with polyP granules in a transspecies-specific manner. These data indicated that CHAD-containing proteins are generally attached to polyP granules. Together with the findings from four previously polyP-attached proteins (polyP kinases), the results of this study raised the number of polyP-associated proteins in R. eutropha to six. We suggest designating polyP granule-bound proteins with CHAD motifs as phosins ( pho sphate), analogous to pha sins and oleo sins that are specifically bound to the surface of polyhydroxyalkanoate (PHA) granules in PHA-accumulating bacteria and to oil droplets in oil seed plants, respectively. IMPORTANCE The importance of polyphosphate (polyP) for life is evident from the ubiquitous presence of polyP in all species on earth. In unicellular eukaryotic microorganisms, polyP is located in specific membrane-enclosed organelles, called acidocalcisomes. However, in most prokaryotes, polyP is present as insoluble granules that have been designated previously as volutin granules. Almost nothing is known regarding the macromolecular composition of polyP granules. Particularly, the absence or presence of cellular compounds on the surface of polyP granules has not yet been investigated. In this study, we identified a novel class of proteins that are attached to the surface of polyP granules in three model species of Alphaproteobacteria , Betaproteobacteria , and Gammaproteobacteria These proteins are characterized by the presence of a CHAD ( c onserved h istidine α -helical d omain) motif that functions as a polyP granule-targeting signal. We suggest designating CHAD motif-containing proteins as phosins [analogous to phasins for poly(3-hydroxybutyrate)-associated proteins and to oleosins for oil droplet-associated proteins in oil seed plants]. The expression of phosins in different species confirmed their polyP-targeting function in a transspecies-specific manner. We postulate that polyP granules in prokaryotic species generally have a complex surface structure that consists of one to several polyP kinases and phosin proteins. We suggest differentiating polyP granules from acidocalcisomes by designating them as polyphosphatosomes. Copyright © 2017 American Society for Microbiology.

Purification and Autoactivation Method for Recombinant Coagulation Factor VII.

PubMed

Granovski, Vladimir; Freitas, Marcela C C; Abreu-Neto, Mario Soares; Covas, Dimas T

2018-01-01

Recombinant coagulation factor VII is a very important and complex protein employed for treatment of hemophiliac patients (hemophilia A/B) who develop inhibitors antibodies to conventional treatments (FVIII and FIX). The rFVII is a glycosylated molecule and circulates in plasma as zymogen of 50 kDa. When activated the molecule is cleaved to 20-30 kDa and has a half-life of about 3 h, needing to be processed fast and efficiently until freeze-drying. Here, we describe a very simple and fast purification sequence for rFVII using affinity FVII Select resin and a dialysis system that can be easily scaled up.

Proteases Revisited: Roles and Therapeutic Implications in Fibrosis

PubMed Central

Kryczka, Jakub

2017-01-01

Proteases target many substrates, triggering changes in distinct biological processes correlated with cell migration, EMT/EndMT and fibrosis. Extracellular protease activity, demonstrated by secreted and membrane-bound protease forms, leads to ECM degradation, activation of other proteases (i.e., proteolysis of nonactive zymogens), decomposition of cell-cell junctions, release of sequestered growth factors (TGF-β and VEGF), activation of signal proteins and receptors, degradation of inflammatory inhibitors or inflammation-related proteins, and changes in cell mechanosensing and motility. Intracellular proteases, mainly caspases and cathepsins, modulate lysosome activity and signal transduction pathways. Herein, we discuss the current knowledge on the multidimensional impact of proteases on the development of fibrosis. PMID:28642633

The Low-Threshold Calcium Channel Cav3.2 Mediates Burst Firing of Mature Dentate Granule Cells

PubMed Central

Dumenieu, Mael; Senkov, Oleg; Mironov, Andrey; Bourinet, Emmanuel; Kreutz, Michael R; Dityatev, Alexander; Heine, Martin; Bikbaev, Arthur

2018-01-01

Abstract Mature granule cells are poorly excitable neurons that were recently shown to fire action potentials, preferentially in bursts. It is believed that the particularly pronounced short-term facilitation of mossy fiber synapses makes granule cell bursting a very effective means of properly transferring information to CA3. However, the mechanism underlying the unique bursting behavior of mature granule cells is currently unknown. Here, we show that Cav3.2 T-type channels at the axon initial segment are responsible for burst firing of mature granule cells in rats and mice. Accordingly, Cav3.2 knockout mice fire tonic spikes and exhibit impaired bursting, synaptic plasticity and dentate-to-CA3 communication. The data show that Cav3.2 channels are strong modulators of bursting and can be considered a critical molecular switch that enables effective information transfer from mature granule cells to the CA3 pyramids. PMID:29790938

Massing in high shear wet granulation can simultaneously improve powder flow and deteriorate powder compaction: a double-edged sword.

PubMed

Shi, Limin; Feng, Yushi; Sun, Changquan Calvin

2011-05-18

The influence of massing during high shear wet granulation (HSWG) process on granule properties and performance was investigated using microcrystalline cellulose (MCC). Massing time varied from 0 to 40 min while other factors were fixed. Granule physical properties, including morphology, size, porosity, and specific surface area (SSA), were characterized. Changes in powder properties were profound in the first 10 min of massing but negligible beyond 10 min. With 10 min of massing, granule tabletability decreased by 75% while flowability increased by 75%. The significantly deteriorated tabletability and improved flowability resulted from dramatic changes in granule morphology, porosity, and SSA. The results confirm that massing time is a key process parameter in HSWG, and it must be carefully evaluated and controlled during process development, scale up, and manufacturing. Copyright © 2011 Elsevier B.V. All rights reserved.

Granular cells in the presence of magnetic field

NASA Astrophysics Data System (ADS)

Jurčák, J.; Lemmerer, B.; van Noort, M.

2017-10-01

We present a statistical study of the dependencies of the shapes and sizes of the photospheric convective cells on the magnetic field properties. This analysis is based on a 2.5 hour long SST observations of active region NOAA 11768. We have blue continuum images taken with a cadence of 5.6 sec that are used for segmentation of individual granules and 270 maps of spectropolarimetric CRISP data allowing us to determine the properties of the magnetic field along with the line-of-sight velocities. The sizes and shapes of the granular cells are dependent on the the magnetic field strength, where the granules tend to be smaller in regions with stronger magnetic field. In the presence of highly inclined magnetic fields, the eccentricity of granules is high and we do not observe symmetric granules in these regions. The mean up-flow velocities in granules as well as the granules intensities decrease with increasing magnetic field strength.

Startup and long term operation of enhanced biological phosphorus removal in continuous-flow reactor with granules.

PubMed

Li, Dong; Lv, Yufeng; Zeng, Huiping; Zhang, Jie

2016-07-01

The startup and long term operation of enhanced biological phosphorus removal (EBPR) in a continuous-flow reactor (CFR) with granules were investigated in this study. Through reducing the settling time from 9min to 3min gradually, the startup of EBPR in a CFR with granules was successfully realized in 16days. Under continuous-flow operation, the granules with good phosphorus and COD removal performance were stably operated for more than 6months. And the granules were characterized with particle size of around 960μm, loose structure and good settling ability. During the startup phase, polysaccharides (PS) was secreted excessively by microorganisms to resist the influence from the variation of operational mode. Results of relative quantitative PCR indicated that granules dominated by polyphosphate-accumulating organisms (PAOs) were easier accumulated in the CFR because more excellent settling ability was needed in the system. Copyright © 2016 Elsevier Ltd. All rights reserved.

Starch granule initiation is controlled by a heteromultimeric isoamylase in potato tubers

PubMed Central

Bustos, Regla; Fahy, Brendan; Hylton, Christopher M.; Seale, Robert; Nebane, N. Miranda; Edwards, Anne; Martin, Cathie; Smith, Alison M.

2004-01-01

Starch granule initiation is not understood, but recent evidence implicates a starch debranching enzyme, isoamylase, in the control of this process. Potato tubers contain isoamylase activity attributable to a heteromultimeric protein containing Stisa1 and Stisa2, the products of two of the three isoamylase genes of potato. To discover whether this enzyme is involved in starch granule initiation, activity was reduced by expression of antisense RNA for Stisa1 or Stisa2. Transgenic tubers accumulated a small amount of a soluble glucan, similar in structure to the phytoglycogen of cereal, Arabidopsis, and Chlamydomonas mutants lacking isoamylase. The major effect, however, was on the number of starch granules. Transgenic tubers accumulated large numbers of tiny granules not seen in normal tubers. These data indicate that the heteromultimeric isoamylase functions during starch synthesis to suppress the initiation of glucan molecules in the plastid stroma that would otherwise crystallize to nucleate new starch granules. PMID:14766984

Sub-nm 3D observation of human hair melanin by high-voltage STEM.

PubMed

Imai, Takehito; Higuchi, Kimitaka; Yamamoto, Yuta; Arai, Shigeo; Nakano, Takashi; Tanaka, Nobuo

2016-04-01

The ultrastructure of melanin granules in human hair was studied using 1,000 kV high-voltage scanning transmission electron microscopy to successfully reconstruct three-dimensional images of the whole melanin granule. It was revealed that the melanin granule was composed of a membrane-like outer structure that included many spherical vesicles, and an inner matrix containing a sheet-like structure in the elongated direction of the melanin granule and a sheet-like arrays structure in the cross direction. The outer structure of the melanin granule was maintained even after exposure to hair-bleaching agents to decompose the melanin granule, suggesting that the outer structure was a highly robust structure and composition compared with the inner matrix . © The Author 2015. Published by Oxford University Press on behalf of The Japanese Society of Microscopy. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Crystalline pteridines in the stromal pigment cells of the iris of the great horned owl.

PubMed

Oliphant, L W

1981-01-01

The bright yellow color of the iris of the Great Horned Owl (Bubo virginianus) is due to unusual pigment cells in the iris stroma. These cells are spherical and contain numerous clear lipid droplets. Around the periphery of these cells are ovoid crystalline granules that are highly birefringent and vary in color from yellow to clear gray. Differential extraction of the lipid droplets and peripheral granules with lipid solvents and 2% KOH confirmed the localization of the yellow pigment in these granules. The color, solubility, fluorescence, chromatographic mobility and ultraviolet absorption of the extracted pigment suggest it is primarily xanthopterin. It is proposed that the peripheral granules are crystalline pterinosomes capable of reflecting light. Most of the cells contain yellow reflecting granules and can be considered reflecting xanthophores. Cells lying deeper in the stroma have colorless reflecting granules and can be considered pteridine containing leucophores.

Bioequivalence of the 4-mg Oral Granules and Chewable Tablet Formulations of Montelukast.

PubMed

Knorr, Barbara; Hartford, Alan; Li, Xiujiang Susie; Yang, Amy Yifan; Noonan, Gertrude; Migoya, Elizabeth

2010-06-01

PURPOSE: The primary objective of the studies was to demonstrate bioequivalence between the oral granules formulation and chewable tablet of montelukast in the fasted state. Effect of food on the pharmacokinetics of the oral granules was also evaluated. METHODS: The Formulation Biocomparison Study (Study 1) and the Final Market Image Study (Study 2) each used an open-label, randomized, 3-period crossover design where healthy adult subjects (N = 24 and 30, respectively) received montelukast as a single 4-mg dose of the oral granules formulation and a 4-mg chewable tablet fasted, and a single 4-mg dose of the oral granules formulation with food (on 2 teaspoons of applesauce [Study 1] or after consumption of a high-fat breakfast [Study 2]). The formulations were to be considered bioequivalent if the 90% confidence intervals (CIs) for geometric mean ratios (GMRs) (oral granules/chewable tablet) for the AUC(0-infinity) and C(max) of montelukast were within the prespecified comparability bounds of (0.80, 1.25). For the food-effect assessment in Study 1, comparability bounds were prespecified as (0.50, 2.00) only for the 90% CI of the GMR (oral granules fed/oral granules fasted) for the AUC(0-infinity) of montelukast; the 90% CI of the GMR for the C(max) of montelukast, however, also was computed. In Study 2, 90% CIs of the GMRs (oral granules fed/oral granules fasted) for the AUC(0-infinity) and C(max) of montelukast were computed; comparability bounds were not prespecified. RESULTS: Comparing the exposure of the formulations, the 90% CIs of the GMRs for AUC(0-infinity) and C(max) were within the prespecified bound of (0.80, 1.25). For AUC(0-infinity), the GMRs (90% CI) for Study 1 and Study 2 were 1.01 (0.92, 1.11) and 0.95 (0.91, 0.99), respectively. For C(max), respective values were 0.99 (0.86, 1.13) and 0.92 (0.84, 1.01). When the oral granules formulation was administered with food, 90% CIs of the GMRs for both AUC(0-infinity) and C(max) in both studies were contained within the interval of (0.50, 2.00). CONCLUSIONS: The 4-mg oral granules and 4-mg chewable tablet formulations of montelukast administered in the fasted state are bioequivalent. Single 4-mg doses of the oral granules formulation and the chewable tablet of montelukast are generally well tolerated.

Bioequivalence of the 4-mg Oral Granules and Chewable Tablet Formulations of Montelukast

PubMed Central

Knorr, Barbara; Hartford, Alan; Li, Xiujiang (Susie); Yang, Amy Yifan; Noonan, Gertrude; Migoya, Elizabeth

2010-01-01

Purpose The primary objective of the studies was to demonstrate bioequivalence between the oral granules formulation and chewable tablet of montelukast in the fasted state. Effect of food on the pharmacokinetics of the oral granules was also evaluated. Methods The Formulation Biocomparison Study (Study 1) and the Final Market Image Study (Study 2) each used an open-label, randomized, 3-period crossover design where healthy adult subjects (N = 24 and 30, respectively) received montelukast as a single 4-mg dose of the oral granules formulation and a 4-mg chewable tablet fasted, and a single 4-mg dose of the oral granules formulation with food (on 2 teaspoons of applesauce [Study 1] or after consumption of a high-fat breakfast [Study 2]). The formulations were to be considered bioequivalent if the 90% confidence intervals (CIs) for geometric mean ratios (GMRs) (oral granules/chewable tablet) for the AUC0-∞ and Cmax of montelukast were within the prespecified comparability bounds of (0.80, 1.25). For the food-effect assessment in Study 1, comparability bounds were prespecified as (0.50, 2.00) only for the 90% CI of the GMR (oral granules fed/oral granules fasted) for the AUC0-∞ of montelukast; the 90% CI of the GMR for the Cmax of montelukast, however, also was computed. In Study 2, 90% CIs of the GMRs (oral granules fed/oral granules fasted) for the AUC0-∞ and Cmax of montelukast were computed; comparability bounds were not prespecified. Results Comparing the exposure of the formulations, the 90% CIs of the GMRs for AUC0-∞ and Cmax were within the prespecified bound of (0.80, 1.25). For AUC0-∞, the GMRs (90% CI) for Study 1 and Study 2 were 1.01 (0.92, 1.11) and 0.95 (0.91, 0.99), respectively. For Cmax, respective values were 0.99 (0.86, 1.13) and 0.92 (0.84, 1.01). When the oral granules formulation was administered with food, 90% CIs of the GMRs for both AUC0-∞ and Cmax in both studies were contained within the interval of (0.50, 2.00). Conclusions The 4-mg oral granules and 4-mg chewable tablet formulations of montelukast administered in the fasted state are bioequivalent. Single 4-mg doses of the oral granules formulation and the chewable tablet of montelukast are generally well tolerated. PMID:20686624

Identification of a new-to-science cyanobacterium, Toxifilum mysidocida gen. nov. & sp. nov. (Cyanobacteria, Cyanophyceae).

PubMed

Zimba, Paul V; Huang, I-Shuo; Foley, Jennifer E; Linton, Eric W

2017-02-01

Cyanobacteria occupy many niches within terrestrial, planktonic, and benthic habitats. The diversity of habitats colonized, similarity of morphology, and phenotypic plasticity all contribute to the difficulty of cyanobacterial identification. An unknown marine filamentous cyanobacterium was isolated from an aquatic animal rearing facility having mysid mortality events. The cyanobacterium originated from Corpus Christi Bay, TX. Filaments are rarely solitary, benthic mat forming, unbranched, and narrowing at the ends. Cells are 2.1 × 3.1 μm (width × length). Thylakoids are peripherally arranged on the outer third of the cell; cyanophycin granules and polyphosphate bodies are present. Molecular phylogenetic analysis in addition to morphology (transmission electron microscopy and scanning electron microscopy) and chemical composition all confirm it as a new genus and species we name Toxifilum mysidocida. At least one identified Leptolyngbya appears (based on genetic evidence and TEM) to belong to this new genus. © 2016 Phycological Society of America.

HIGH SHEAR GRANULATION PROCESS: ASSESSING IMPACT OF FORMULATION VARIABLES ON GRANULES AND TABLETS CHARACTERISTICS OF HIGH DRUG LOADING FORMULATION USING DESIGN OF EXPERIMENT METHODOLOGY.

PubMed

Fayed, Mohamed H; Abdel-Rahman, Sayed I; Alanazi, Fars K; Ahmed, Mahrous O; Tawfeek, Hesham M; Ali, Bahaa E

2017-03-01

High shear wet granulation is a significant component procedure in the pharmaceutical industry. The objective of the study was to investigate the influence of two independent formulation variables; polyvinypyrrolidone (PVP) as a binder (X,) and croscarmellose sodium (CCS) as a disintegrant (X2) on the crit- ical quality attributes of acetaminophen granules and their corresponding tablets using design of experiment (DoE) approach. A two factor, three level (32) full factorial design has been applied; each variable was investi- gated at three levels to characterize their strength and interaction. The dried granules have been analyzed for their density, granule size and flowability. Additionally, the produced tablets have been investigated for: break- ing force, friability, disintegration time and t. of drug dissolution. The analysis of variance (ANOVA) showed that the two variables had a significant impact (p < 0.05) on granules and tablets characteristics, while only the binder concentration influenced the tablets friability. Furthermore, significant interactions (p < 0.05) between the two variables, for granules and tablets attributes, were also found. However, variables interaction showed minimal effect for granules flowability as well as tablets friability. Desirability function was carried out to opti- mize the variables under study to obtain product within the USP limit. It was found that the higher desirability (0.985) could be obtained at the medium level of PVP and low level of CCS. Ultimately, this study supplies the formulator with beneficial tools in selecting the proper level of binder and disintegrant to attain product with desired characteristics.

Biphasic adaptation to osmotic stress in the C. elegans germ line.

PubMed

Davis, Michael; Montalbano, Andrea; Wood, Megan P; Schisa, Jennifer A

2017-06-01

Cells respond to environmental stress in multiple ways. In the germ line, heat shock and nutritive stress trigger the assembly of large ribonucleoprotein (RNP) granules via liquid-liquid phase separation (LLPS). The RNP granules are hypothesized to maintain the quality of oocytes during stress. The goal of this study was to investigate the cellular response to glucose in the germ line and determine if it is an osmotic stress response. We found that exposure to 500 mM glucose induces the assembly of RNP granules in the germ line within 1 h. Interestingly, the RNP granules are maintained for up to 3 h; however, they dissociate after longer periods of stress. The RNP granules include processing body and stress granule proteins, suggesting shared functions. Based on several lines of evidence, the germ line response to glucose largely appears to be an osmotic stress response, thus identifying osmotic stress as a trigger of LLPS. Although RNP granules are not maintained beyond 3 h of osmotic stress, the quality of oocytes does not appear to decrease after longer periods of stress, suggesting a secondary adaptation in the germ line. We used an indirect marker of glycerol and observed high levels after 5 and 20 h of glucose exposure. Moreover, in gpdh-1;gpdh-2 germ lines, glycerol levels are reduced concomitant with RNP granules being maintained for an extended period. We speculate that increased glycerol levels may function as a secondary osmoregulatory adaptive response in the germ line, following a primary response of RNP granule assembly. Copyright © 2017 the American Physiological Society.

The Stress Granule RNA-Binding Protein TIAR-1 Protects Female Germ Cells from Heat Shock in Caenorhabditis elegans.

PubMed

Huelgas-Morales, Gabriela; Silva-García, Carlos Giovanni; Salinas, Laura S; Greenstein, David; Navarro, Rosa E

2016-04-07

In response to stressful conditions, eukaryotic cells launch an arsenal of regulatory programs to protect the proteome. One major protective response involves the arrest of protein translation and the formation of stress granules, cytoplasmic ribonucleoprotein complexes containing the conserved RNA-binding proteins TIA-1 and TIAR. The stress granule response is thought to preserve mRNA for translation when conditions improve. For cells of the germline-the immortal cell lineage required for sexual reproduction-protection from stress is critically important for perpetuation of the species, yet how stress granule regulatory mechanisms are deployed in animal reproduction is incompletely understood. Here, we show that the stress granule protein TIAR-1 protects the Caenorhabditis elegans germline from the adverse effects of heat shock. Animals containing strong loss-of-function mutations in tiar-1 exhibit significantly reduced fertility compared to the wild type following heat shock. Analysis of a heat-shock protein promoter indicates that tiar-1 mutants display an impaired heat-shock response. We observed that TIAR-1 was associated with granules in the gonad core and oocytes during several stressful conditions. Both gonad core and oocyte granules are dynamic structures that depend on translation; protein synthesis inhibitors altered their formation. Nonetheless, tiar-1 was required for the formation of gonad core granules only. Interestingly, the gonad core granules did not seem to be needed for the germ cells to develop viable embryos after heat shock. This suggests that TIAR-1 is able to protect the germline from heat stress independently of these structures. Copyright © 2016 Huelgas-Morales et al.

The Stress Granule RNA-Binding Protein TIAR-1 Protects Female Germ Cells from Heat Shock in Caenorhabditis elegans

PubMed Central

Huelgas-Morales, Gabriela; Silva-García, Carlos Giovanni; Salinas, Laura S.; Greenstein, David; Navarro, Rosa E.

2016-01-01

In response to stressful conditions, eukaryotic cells launch an arsenal of regulatory programs to protect the proteome. One major protective response involves the arrest of protein translation and the formation of stress granules, cytoplasmic ribonucleoprotein complexes containing the conserved RNA-binding proteins TIA-1 and TIAR. The stress granule response is thought to preserve mRNA for translation when conditions improve. For cells of the germline—the immortal cell lineage required for sexual reproduction—protection from stress is critically important for perpetuation of the species, yet how stress granule regulatory mechanisms are deployed in animal reproduction is incompletely understood. Here, we show that the stress granule protein TIAR-1 protects the Caenorhabditis elegans germline from the adverse effects of heat shock. Animals containing strong loss-of-function mutations in tiar-1 exhibit significantly reduced fertility compared to the wild type following heat shock. Analysis of a heat-shock protein promoter indicates that tiar-1 mutants display an impaired heat-shock response. We observed that TIAR-1 was associated with granules in the gonad core and oocytes during several stressful conditions. Both gonad core and oocyte granules are dynamic structures that depend on translation; protein synthesis inhibitors altered their formation. Nonetheless, tiar-1 was required for the formation of gonad core granules only. Interestingly, the gonad core granules did not seem to be needed for the germ cells to develop viable embryos after heat shock. This suggests that TIAR-1 is able to protect the germline from heat stress independently of these structures. PMID:26865701

Vigorous convection in a sunspot granular light bridge

NASA Astrophysics Data System (ADS)

Lagg, Andreas; Solanki, Sami K.; van Noort, Michiel; Danilovic, Sanja

2014-08-01

Context. Light bridges are the most prominent manifestation of convection in sunspots. The brightest representatives are granular light bridges composed of features that appear to be similar to granules. Aims: An in-depth study of the convective motions, temperature stratification, and magnetic field vector in and around light bridge granules is presented with the aim of identifying similarities and differences to typical quiet-Sun granules. Methods: Spectropolarimetric data from the Hinode Solar Optical Telescope were analyzed using a spatially coupled inversion technique to retrieve the stratified atmospheric parameters of light bridge and quiet-Sun granules. Results: Central hot upflows surrounded by cooler fast downflows reaching 10 km s-1 clearly establish the convective nature of the light bridge granules. The inner part of these granules in the near surface layers is field free and is covered by a cusp-like magnetic field configuration. We observe hints of field reversals at the location of the fast downflows. The quiet-Sun granules in the vicinity of the sunspot are covered by a low-lying canopy field extending radially outward from the spot. Conclusions: The similarities between quiet-Sun and light bridge granules point to the deep anchoring of granular light bridges in the underlying convection zone. The fast, supersonic downflows are most likely a result of a combination of invigorated convection in the light bridge granule due to radiative cooling into the neighboring umbra and the fact that we sample deeper layers, since the downflows are immediately adjacent to the slanted walls of the Wilson depression. The two movies are available in electronic form at http://www.aanda.org

Localization of DNA and RNA in eosinophil secretory granules.

PubMed

Behzad, Ali R; Walker, David C; Abraham, Thomas; McDonough, John; Mahmudi-Azer, Salahadin; Chu, Fanny; Shaheen, Furquan; Hogg, James C; Paré, Peter D

2010-01-01

Although the accepted paradigm is that the proteins stored in eosinophil crystalloid granules are translated from messenger RNA transcribed in the cell nucleus, recent ultrastructural evidence suggests that protein synthesis may also take place within eosinophilic granules. We used 2 different methods to detect the presence of DNA and RNA in eosinophil secretory granules. Using bromodeoxyuridine, a thymidine analogue, and bromouridine, a uracil analogue, we labeled the DNA and RNA in eosinophils in vivo in rabbits. Immunoelectron microscopy to localize these molecules was performed on ultrathin sections of blood and bone marrow eosinophils using monoclonal anti-bromodeoxyuridine antibody with IgG as a control. The immunogold grain density was measured in each subcellular compartment within the eosinophils and analyzed using image analysis software. A combination of DNA/CD63 immunofluorescence staining and a fluorescently labeled molecular probe that stains RNA was used to examine the presence of DNA and RNA in the secretory granules of human blood eosinophils. The mean density of bromodeoxyuridine-labeled DNA and bromouridine-labeled RNA immunogold grains in the secretory granules of blood and bone marrow eosinophils were significantly higher (p < 0.0005) than cytoplasmic or background staining. We also demonstrated the existence of DNA and RNA in the CD63-positive secretory granules of human peripheral blood eosinophils by means of immunofluorescent staining and a fluorescently labeled molecular probe. These results provide evidence that eosinophil granules are the site of DNA and RNA synthesis and suggest the potential for a new role(s) for eosinophil-secretory granules. Copyright 2009 S. Karger AG, Basel.

Does the performance of wet granulation and tablet hardness affect the drug dissolution profile of carvedilol in matrix tablets?

PubMed

Košir, Darjan; Ojsteršek, Tadej; Vrečer, Franc

2018-06-14

Wet granulation is mostly used process for manufacturing matrix tablets. Compared to the direct compression method, it allows for a better flow and compressibility properties of compression mixtures. Granulation, including process parameters and tableting, can influence critical quality attributes (CQAs) of hydrophilic matrix tablets. One of the most important CQAs is the drug release profile. We studied the influence of granulation process parameters (type of nozzle and water quantity used as granulation liquid) and tablet hardness on the drug release profile. Matrix tablets contained HPMC K4M hydrophilic matrix former and carvedilol as a model drug. The influence of selected HPMC characteristics on the drug release profile was also evaluated using two additional HPMC batches. For statistical evaluation, partial least square (PLS) models were generated for each time point of the drug release profile using the same number of latent factors. In this way, it was possible to evaluate how the importance of factors influencing drug dissolution changes in dependence on time throughout the drug release profile. The results of statistical evaluation show that the granulation process parameters (granulation liquid quantity and type of nozzle) and tablet hardness significantly influence the release profile. On the other hand, the influence of HPMC characteristics is negligible in comparison to the other factors studied. Using a higher granulation liquid quantity and the standard nozzle type results in larger granules with a higher density and lower porosity, which leads to a slower drug release profile. Lower tablet hardness also slows down the release profile.

Microstructure of anammox granules and mechanisms endowing their intensity revealed by microscopic inspection and rheometry.

PubMed

Lin, Ximao; Wang, Yayi

2017-09-01

The anammox process represents a sustainable and cost-effective technique for nitrogen removal from wastewater, where granulation of anammox bacteria could be of great benefit to the system performance. However, knowledge of the specific properties of anammox granules is currently unsatisfactory. In this study, the organization of anammox granules was comprehensively studied from macro to micro scale with a range of microscale techniques. Scanning and transmission electron microscopy and multiple fluorescence labeling combined with confocal laser scanning microscopy were included. Simultaneously, the associated mechanical properties were studied in-depth by rheometry in combination with selective enzymatic hydrolysis. Anammox granules follow a tertiary organization regime, where interactions between individual anammox bacteria made up the primary base, then, the grouping of anammox bacterial cells encapsulated within a thin extracellular polymeric substance (EPS) layer comprised a second arrangement level, and, finally, the cementing of these groups together with other bacteria and polymers gave rise to compact aggregates. α-Polysaccharides and proteins were considered the backbones of anammox granules, contributing greatly to their excellent intensity. β-Polysaccharides concentrated at the outer rims of anammox granules and combined with other macromolecules to form a buffer zone or protective barrier, beneath which anammox bacteria proliferated. Divalent cationic bridging for EPS binding was prevalent and of great significance within the dense anammox granules, while there was also much weak monovalent ionic interaction. The specific organization and composition of anammox granules endows them with excellent intensity and integrity, which can be of importance for full-scale reactor operations where diverse shocks can be expected. Copyright © 2017 Elsevier Ltd. All rights reserved.

Costus root granules improve ulcerative colitis through regulation of TGF-β mediation of the PI3K/AKT signaling pathway

PubMed Central

Wang, Xiaohong; Li, Dan; Zhang, Yong; Wu, Shuang; Tang, Fang

2018-01-01

Ulcerative colitis is a chronic nonspecific inflammatory disease that occurs in the colon and rectum. Costus root is a type of traditional Chinese medicine that exhibits antibacterial properties and serves an inhibitory role in the regeneration of gut bacteria. However, the molecular mechanisms underlying Costus root-mediated improvements in ulcerative colitis remain unclear. A complex formula of Costus root granules was created and investigated in the present study for its therapeutic effects in a rat model of ulcerative colitis. Ingredient dissolution into a traditional water decoction was used as a control. The potential mechanism mediated by Costus root granules was also analyzed in colonic epithelial cells isolated from the experimental rats. The results of the present study demonstrated that Costus root granule treatment inhibited inflammation in colonic tissue. Costus root granule treatment also suppressed the apoptosis of colonic epithelial cells isolated from the rat model of ulcerative colitis. Analyses of the underlying mechanisms of these effects indicated that the administration of Costus root granules increased transforming growth factor β expression, which activated the phosphoinositide 3-kinase/RAC-α serine/threonine-protein kinase signaling pathway in colonic epithelial cells. Notably, the administration of Costus root granules improved stomachache, diarrhea and hematochezia in and increased the body weight of, the ulcerative colitis rats. In conclusion, these results indicate that Costus root granules markedly ameliorate inflammation of the colonic epithelium, decrease the apoptosis of colonic epithelial cells and improve colonic function, which suggests that Costus root granules are an efficient agent for the treatment of ulcerative colitis. PMID:29731832

Stress granules at the intersection of autophagy and ALS

PubMed Central

Monahan, Zachary; Shewmaker, Frank; Pandey, Udai Bhan

2016-01-01

Amyotrophic lateral sclerosis (ALS) is a progressive, fatal disease caused by loss of upper and lower motor neurons. The majority of ALS cases are classified as sporadic (80-90%), with the remaining considered familial based on patient history. The last decade has seen a surge in the identification of ALS-causing genes – including TARDBP (TDP-43), FUS, MATR3 (Matrin-3), C9ORF72 and several others – providing important insights into the molecular pathways involved in pathogenesis. Most of the protein products of ALS-linked genes fall into two functional categories: RNA-binding/homeostasis and protein-quality control (i.e. autophagy and proteasome). The RNA-binding proteins tend to be aggregation-prone with low-complexity domains similar to the prion-forming domains of yeast. Many also incorporate into stress granules (SGs), which are cytoplasmic ribonucleoprotein complexes that form in response to cellular stress. Mutant forms of TDP-43 and FUS perturb SG dynamics, lengthening their cytoplasmic persistence. Recent evidence suggests that SGs are regulated by the autophagy pathway, suggesting a unifying connection between many of the ALS-linked genes. Persistent SGs may give rise to intractable aggregates that disrupt neuronal homeostasis, thus failure to clear SGs by autophagic processes may promote ALS pathogenesis. PMID:27181519

Identification of the antigenic region of Neospora caninum dense granule protein 7 using ELISA.

PubMed

Abdelbaky, Hanan H; Fereig, Ragab M; Nishikawa, Yoshifumi

2018-06-26

Dense granule protein 7 (NcGRA7) is a potent diagnostic antigen of Neospora caninum. Some studies have reported on the difficult expression, low yield, and variable degree of solubility of recombinant NcGRA7. We aimed to unravel the possible causes for these issues and tested NcGRA7 antigenicity in enzyme linked immunosorbent assays (ELISAs). The NcGRA7 coding sequence (217 amino acids) was split into five amino acid regions: NcGRA7m (27-217), NcGRA7m3 (27-160), NcGRA7m4 (27-135), NcGRA7m5 (161-190), and NcGRA7m6 (188-217). Three fragments, NcGRA7m, NcGRA7m3 and NcGRA7m4, exhibited high antigenic properties when tested against experimentally-infected mice and dog sera by ELISA. High levels of IgG2 antibodies against NcGRA7m were observed in field dog sera. In experimentally and naturally-infected cattle, the N. caninum-specific sera only reacted with NcGRA7m, indicating that this antigenic region differs among the three animal species. This study presents valuable information about the antigenic properties and topology of NcGRA7, and highlights its suitability for diagnostic purposes. Copyright © 2018. Published by Elsevier B.V.

Vaccine adjuvants: Tailor-made mast-cell granules

NASA Astrophysics Data System (ADS)

Gunzer, Matthias

2012-03-01

Mast cells induce protective immune responses through secretion of stimulatory granules. Microparticles modelled after mast-cell granules are now shown to replicate and enhance the functions of their natural counterparts and to direct the character of the resulting immunity.

Nucleotide and bivalent cation specificity of the insulin-granule proton translocase.

PubMed Central

Hutton, J C; Peshavaria, M

1983-01-01

1. The nucleotide and bivalent cation specificity of the proton translocase activity of insulin secretory granules was investigated by assessing the inhibitor-sensitive rates of nucleotide hydrolysis by these organelles in relation to their chemiosmotic properties. 2. The relative rates of nucleotide hydrolysis by freeze/thawed granule preparations were: Mg2+ATP (100%) greater than Mg2+GTP (55%) greater than Mg2+UTP (48%) greater than Mg2+ITP (44%) greater than Mg2+CTP (23%) greater than Mg2+TTP (20%), and by intact granules were: Mg2+ATP (100%) greater than Mg2+ITP (74%) greater than Mg2+GTP (60%) greater than Mg2+CTP (35%). Mg2+ATP, Mg2+GTP and Mg2+ITP hydrolyses were inhibited by tributyltin and stimulated, in intact granules, by the protonophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone; Mg2+CTP hydrolysis was not markedly affected by these compounds. Correspondingly, only Mg2+ATP, Mg2+GTP and Mg2+ITP produced large changes in the delta psi and delta mu H+ across the granule membrane. 3. The relative rates of maximal ATPase activity stimulated by bivalent cations in freeze/thawed granule preparations were: Mg2+ (100%) greater than Mn2+ (82%) greater than Ca2+ (40%) greater than Co2+ (36%) greater than Zn2+ (0%), and in intact granules were: Mg2+ (100%) greater than Mn2+ (85%) greater than Co2+ (61%) greater than Ca2+ (42%). Tributyltin and carbonyl cyanide p-trifluoromethoxyphenylhydrazone affected Mg2+-, Mn2+- and Co2+-activated, but not Ca2+-activated, ATP hydrolysis. Correspondingly, only Mg2+, Mn2+ and Co2+ supported the generation of a delta psi and delta mu H+ across granule membranes in the presence of ATP. 4. The results were consistent with a single proton translocase that had its catalytic site exposed on the external face of the granule membrane. The indicated specificity (Mg2+ATP = Mn2+ATP greater than Co2+ATP greater than Mg2+GTP greater than Mg2+ITP) was similar to that of enzymes described in membrane fractions prepared from adenohypophyseal tissue, adrenal chromaffin granules and yeast vacuoles. The insulin-granule activity thus appears to be a type of proton translocase, which is characteristic of intracellular storage vesicles in eukaryotic cells. PMID:6303313

Regulation of platelet granule exocytosis by S-nitrosylation

PubMed Central

Morrell, Craig N.; Matsushita, Kenji; Chiles, Kelly; Scharpf, Robert B.; Yamakuchi, Munekazu; Mason, Rebecca J. A.; Bergmeier, Wolfgang; Mankowski, Joseph L.; Baldwin, William M.; Faraday, Nauder; Lowenstein, Charles J.

2005-01-01

Nitric oxide (NO) regulates platelet activation by cGMP-dependent mechanisms and by mechanisms that are not completely defined. Platelet activation includes exocytosis of platelet granules, releasing mediators that regulate interactions between platelets, leukocytes, and endothelial cells. Exocytosis is mediated in part by N-ethylmaleimide-sensitive factor (NSF), an ATPase that disassembles complexes of soluble NSF attachment protein receptors. We now demonstrate that NO inhibits exocytosis of dense granules, lysosomal granules, and α-granules from human platelets by S-nitrosylation of NSF. Platelets lacking endothelial NO synthase show increased rolling on venules, increased thrombosis in arterioles, and increased exocytosis in vivo. Regulation of exocytosis is thus a mechanism by which NO regulates thrombosis. PMID:15738422

Rab27a mediates the tight docking of insulin granules onto the plasma membrane during glucose stimulation.

PubMed

Kasai, Kazuo; Ohara-Imaizumi, Mica; Takahashi, Noriko; Mizutani, Shin; Zhao, Shengli; Kikuta, Toshiteru; Kasai, Haruo; Nagamatsu, Shinya; Gomi, Hiroshi; Izumi, Tetsuro

2005-02-01

The monomeric small GTPase Rab27a is specifically localized on both secretory granules and lysosome-related organelles. Although natural mutations of the Rab27a gene in human Griscelli syndrome and in ashen mice cause partial albinism and immunodeficiency reflecting the dysfunction of lysosome-related organelles, phenotypes resulting from the defective exocytosis of secretory granules have not been reported. To explore the roles of Rab27a in secretory granules, we analyzed insulin secretion profiles in ashen mice. Ashen mice showed glucose intolerance after a glucose load without signs of insulin resistance in peripheral tissues or insulin deficiency in the pancreas. Insulin secretion from isolated islets was decreased specifically in response to high glucose concentrations but not other nonphysiological secretagogues such as high K+ concentrations, forskolin, or phorbol ester. Neither the intracellular Ca2+ concentration nor the dynamics of fusion pore opening after glucose stimulation were altered. There were, however, marked reductions in the exocytosis from insulin granules predocked on the plasma membrane and in the replenishment of docked granules during glucose stimulation. These results provide the first genetic evidence to our knowledge for the role of Rab27a in the exocytosis of secretory granules and suggest that the Rab27a/effector system mediates glucose-specific signals for the exocytosis of insulin granules in pancreatic beta cells.

Fast insulin secretion reflects exocytosis of docked granules in mouse pancreatic B-cells.

PubMed

Olofsson, Charlotta S; Göpel, Sven O; Barg, Sebastian; Galvanovskis, Juris; Ma, Xiaosong; Salehi, Albert; Rorsman, Patrik; Eliasson, Lena

2002-05-01

A readily releasable pool (RRP) of granules has been proposed to underlie the first phase of insulin secretion. In the present study we combined electron microscopy, insulin secretion measurements and recordings of cell capacitance in an attempt to define this pool ultrastructurally. Mouse pancreatic B-cells contain approximately 9,000 granules, of which 7% are docked below the plasma membrane. The number of docked granules was reduced by 30% (200 granules) during 10 min stimulation with high K+. This stimulus depolarized the cell to -10 mV, elevated cytosolic [Ca2+] ([Ca2+](i)) from a basal concentration of 130 nM to a peak of 1.3 microM and released 0.5 ng insulin/islet, corresponding to 200-300 granules/cell. The Ca2+ transient decayed towards the prestimulatory concentration within approximately 200 s, presumably reflecting Ca2+ channel inactivation. Renewed stimulation with high K+ failed to stimulate insulin secretion when applied in the absence of glucose. The size of the RRP, derived from the insulin measurements, is similar to that estimated from the increase in cell capacitance elicited by photolytic release of caged Ca2+. We propose that the RRP represents a subset of the docked pool of granules and that replenishment of RRP can be accounted for largely by chemical modification of granules already in place or situated close to the plasma membrane.

Ca2+ influx does not trigger glucose-induced traffic of the insulin granules and alteration of their distribution.

PubMed

Niki, Ichiro; Niwa, Tae; Yu, Wei; Budzko, Dorota; Miki, Takashi; Senda, Takao

2003-11-01

This study investigated mechanisms by which glucose increases readily releasable secretory granules via acting on preexocytotic steps, i.e., intracellular granule movement and granule access to the plasma membrane using a pancreatic beta-cell line, MIN6. Glucose-induced activation of the movement occurred at a substimulatory concentration with regard to insulin output. Glucose activation of the movement was inhibited by pretreatment with thapsigargin plus acetylcholine to suppress intracellular Ca2+ mobilization. Inhibitors of calmodulin and myosin light chain kinase also suppressed glucose activation of the movement. Simultaneous addition of glucose with Ca2+ channel blockers or the ATP-sensitive K+ channel opener diazoxide failed to suppress the traffic activation, and addition of these substances on top of glucose stimulation resulted in a further increase. Although stimulatory glucose had minimal changes in the intracellular granule distribution, inhibition of Ca2+ influx revealed increases by glucose of the granules in the cell periphery. In contrast, high K+ depolarization decreased the peripheral granules. Glucose-induced granule margination was abolished when the protein kinase C activity was downregulated. These findings indicate that preexocytotic control of insulin release is regulated by distinct mechanisms from Ca2+ influx, which triggers insulin exocytosis. The nature of the regulation by glucose may explain a part of potentiating effects of the hexose independent of the closure of the ATP-sensitive K+ channel.

Polyhydroxyalkanoate (PHA) Granules Have no Phospholipids

PubMed Central

Bresan, Stephanie; Sznajder, Anna; Hauf, Waldemar; Forchhammer, Karl; Pfeiffer, Daniel; Jendrossek, Dieter

2016-01-01

Polyhydroxybutyrate (PHB) granules, also designated as carbonosomes, are supra-molecular complexes in prokaryotes consisting of a PHB polymer core and a surface layer of structural and functional proteins. The presence of suspected phospholipids in the surface layer is based on in vitro data of isolated PHB granules and is often shown in cartoons of the PHB granule structure in reviews on PHB metabolism. However, the in vivo presence of a phospholipid layer has never been demonstrated. We addressed this topic by the expression of fusion proteins of DsRed2EC and other fluorescent proteins with the phospholipid-binding domain (LactC2) of lactadherin in three model organisms. The fusion proteins specifically localized at the cell membrane of Ralstonia eutropha but did not co-localize with PHB granules. The same result was obtained for Pseudomonas putida, a species that accumulates another type of polyhydroxyalkanoate (PHA) granules related to PHB. Notably, DsRed2EC-LactC2 expressed in Magnetospirillum gryphiswaldense was detected at the position of membrane-enclosed magnetosome chains and at the cytoplasmic membrane but not at PHB granules. In conclusion, the carbonosomes of representatives of α-proteobacteria, β-proteobacteria and γ-proteobacteria have no phospholipids in vivo and we postulate that the PHB/PHA granule surface layers in natural producers generally are free of phospholipids and consist of proteins only. PMID:27222167

Eudragit E as excipient for production of granules and tablets from Phyllanthus niruri L spray-dried extract.

PubMed

Pereira de Souza, Tatiane; Martínez-Pacheco, Ramón; Gómez-Amoza, José Luiz; Petrovick, Pedro Ros

2007-04-27

The aim of this study was to investigate the feasibility of using Eudragit E as a granulating agent for a spray-dried extract from Phyllanthus niruri to obtain tablets containing a high dose of this product. The granules were developed by wet granulation and contained 2.5%, 5.0%, and 10.0% Eudragit E in the final product concentration. The tablets were produced on a single-punch tablet press by direct compression of granules using 0.5% magnesium stearate as a lubricant. The tablets were elaborated following a 2 x 3 factorial design, where Eudragit E concentration and compression force were the independent variables, and tensile strength and the extract release of the tablets were the dependent variables. All granules showed better technological properties than the spray-dried extract, including less moisture sorption. The characteristics of the granules were directly dependent on the proportion of Eudragit E in the formulation. In general, all tablets showed high mechanical resistance with less than 1% friability, less moisture sorption, and a slower extract release profile. The Eudragit E concentration and compression force of the tablets significantly influenced both dependent variables studied. In conclusion, Eudragit E was efficient as a granulating agent for the spray-dried extract, but additional studies are needed to further optimize the formulations in order to achieve less water sorption and improve the release of the extract from the tablets.

Roll Compaction and Tableting of High Loaded Metformin Formulations Using Efficient Binders.

PubMed

Arndt, Oscar-Rupert; Kleinebudde, Peter

2018-04-23

Metformin has a poor tabletability and flowability. Therefore, metformin is typically wet granulated with a binder before tableting. To save production costs, it would be desirable to implement a roll compaction/dry granulation (RCDG) process for metformin instead of using wet granulation. In order to implement RCDG, the efficiency of dry binders is crucial to ensure a high drug load and suitable properties of dry granules and tablets. This study evaluates dry granules manufactured by RCDG and subsequently tableting of high metformin content formulations (≥ 87.5%). Based on previous results, fine particle grades of hydroxypropylcellulose and copovidone in different fractions were compared as dry binders. The formulations are suitable for RCDG and tableting. Furthermore, results can be connected to in-die and out-of-die compressibility analysis. The addition of 7% of dry binder is a good compromise to generate sufficient mechanical properties on the one hand, but also to save resources and ensure a high metformin content on the other hand. Hydroxypropylcellulose was more efficient in terms of granule size, tensile strength and friability. Three percent croscarmellose was added to reach the specifications of the US Pharmacopeia regarding dissolution. The final formulation has a metformin content of 87.5%. A loss in tabletability does not occur for granules compressed at different specific compaction forces, which displays a robust tensile strength of tablets independent of the granulation process.

Nitrous Oxide Production in a Granule-based Partial Nitritation Reactor: A Model-based Evaluation

NASA Astrophysics Data System (ADS)

Peng, Lai; Sun, Jing; Liu, Yiwen; Dai, Xiaohu; Ni, Bing-Jie

2017-04-01

Sustainable wastewater treatment has been attracting increasing attentions over the past decades. However, the production of nitrous oxide (N2O), a potent GHG, from the energy-efficient granule-based autotrophic nitrogen removal is largely unknown. This study applied a previously established N2O model, which incorporated two N2O production pathways by ammonia-oxidizing bacteria (AOB) (AOB denitrification and the hydroxylamine (NH2OH) oxidation). The two-pathway model was used to describe N2O production from a granule-based partial nitritation (PN) reactor and provide insights into the N2O distribution inside granules. The model was evaluated by comparing simulation results with N2O monitoring profiles as well as isotopic measurement data from the PN reactor. The model demonstrated its good predictive ability against N2O dynamics and provided useful information about the shift of N2O production pathways inside granules for the first time. The simulation results indicated that the increase of oxygen concentration and granule size would significantly enhance N2O production. The results further revealed a linear relationship between N2O production and ammonia oxidation rate (AOR) (R2 = 0.99) under the conditions of varying oxygen levels and granule diameters, suggesting that bulk oxygen and granule size may exert an indirect effect on N2O production by causing a change in AOR.

Turnover of pigment granules: cyclic catabolism and anabolism of ommochromes within epidermal cells.

PubMed

Insausti, T C; Casas, J

2009-12-01

Ommochromes are end products of the tryptophan metabolism in arthropods. While the anabolism of ommochromes has been well studied, the catabolism is totally unknown. In order to study it, we used the crab-spider Misumena vatia, which is able to change color reversibly in a few days, from yellow to white and back. Ommochromes is the only pigment class responsible for the body coloration in this animal. The aim of this study was to analyze the fine structure of the epidermal cells in bleaching spiders, in an attempt to correlate morphological changes with the fate of the pigment granules. Central to the process of bleaching is the lysis of the ommochrome granules. In the same cell, intact granules and granules in different degradation stages are found. The degradation begins with granule autolysis. Some components are extruded in the extracellular space and others are recycled via autophagy. Abundant glycogen appears associated to granulolysis. In a later stage of bleaching, ommochrome progranules, typical of white spiders, appear in the distal zone of the same epidermal cell. Catabolism and anabolism of pigment granules thus take place simultaneously in spider epidermal cells. A cyclic pathway of pigment granules formation and degradation, throughout a complete cycle of color change is proposed, together with an explanation for this turnover, involving photoprotection against UV by ommochromes metabolites. The presence of this turnover for melanins is discussed.

Multiple zebrafish atoh1 genes specify a diversity of neuronal types in the zebrafish cerebellum.

PubMed

Kidwell, Chelsea U; Su, Chen-Ying; Hibi, Masahiko; Moens, Cecilia B

2018-06-01

A single Atoh1 basic-helix-loop-helix transcription factor specifies multiple neuron types in the mammalian cerebellum and anterior hindbrain. The zebrafish genome encodes three paralagous atoh1 genes whose functions in cerebellum and anterior hindbrain development we explore here. With use of a transgenic reporter, we report that zebrafish atoh1c-expressing cells are organized in two distinct domains that are separated both by space and developmental time. An early isthmic expression domain gives rise to an extracerebellar population in rhombomere 1 and an upper rhombic lip domain gives rise to granule cell progenitors that migrate to populate all four granule cell territories of the fish cerebellum. Using genetic mutants we find that of the three zebrafish atoh1 paralogs, atoh1c and atoh1a are required for the full complement of granule neurons. Surprisingly, the two genes are expressed in non-overlapping granule cell progenitor populations, indicating that fish use duplicate atoh1 genes to generate granule cell diversity that is not detected in mammals. Finally, live imaging of granule cell migration in wildtype and atoh1c mutant embryos reveals that while atoh1c is not required for granule cell specification per se, it is required for granule cells to delaminate and migrate away from the rhombic lip. Copyright © 2018 Elsevier Inc. All rights reserved.

Nitrous Oxide Production in a Granule-based Partial Nitritation Reactor: A Model-based Evaluation

PubMed Central

Peng, Lai; Sun, Jing; Liu, Yiwen; Dai, Xiaohu; Ni, Bing-Jie

2017-01-01

Sustainable wastewater treatment has been attracting increasing attentions over the past decades. However, the production of nitrous oxide (N2O), a potent GHG, from the energy-efficient granule-based autotrophic nitrogen removal is largely unknown. This study applied a previously established N2O model, which incorporated two N2O production pathways by ammonia-oxidizing bacteria (AOB) (AOB denitrification and the hydroxylamine (NH2OH) oxidation). The two-pathway model was used to describe N2O production from a granule-based partial nitritation (PN) reactor and provide insights into the N2O distribution inside granules. The model was evaluated by comparing simulation results with N2O monitoring profiles as well as isotopic measurement data from the PN reactor. The model demonstrated its good predictive ability against N2O dynamics and provided useful information about the shift of N2O production pathways inside granules for the first time. The simulation results indicated that the increase of oxygen concentration and granule size would significantly enhance N2O production. The results further revealed a linear relationship between N2O production and ammonia oxidation rate (AOR) (R2 = 0.99) under the conditions of varying oxygen levels and granule diameters, suggesting that bulk oxygen and granule size may exert an indirect effect on N2O production by causing a change in AOR. PMID:28367960

Nitrous Oxide Production in a Granule-based Partial Nitritation Reactor: A Model-based Evaluation.

PubMed

Peng, Lai; Sun, Jing; Liu, Yiwen; Dai, Xiaohu; Ni, Bing-Jie

2017-04-03

Sustainable wastewater treatment has been attracting increasing attentions over the past decades. However, the production of nitrous oxide (N 2 O), a potent GHG, from the energy-efficient granule-based autotrophic nitrogen removal is largely unknown. This study applied a previously established N 2 O model, which incorporated two N 2 O production pathways by ammonia-oxidizing bacteria (AOB) (AOB denitrification and the hydroxylamine (NH 2 OH) oxidation). The two-pathway model was used to describe N 2 O production from a granule-based partial nitritation (PN) reactor and provide insights into the N 2 O distribution inside granules. The model was evaluated by comparing simulation results with N 2 O monitoring profiles as well as isotopic measurement data from the PN reactor. The model demonstrated its good predictive ability against N 2 O dynamics and provided useful information about the shift of N 2 O production pathways inside granules for the first time. The simulation results indicated that the increase of oxygen concentration and granule size would significantly enhance N 2 O production. The results further revealed a linear relationship between N 2 O production and ammonia oxidation rate (AOR) (R 2  = 0.99) under the conditions of varying oxygen levels and granule diameters, suggesting that bulk oxygen and granule size may exert an indirect effect on N 2 O production by causing a change in AOR.

Performance of mesophilic anaerobic granules for removal of octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) from aqueous solution.

PubMed

An, Chun-jiang; He, Yan-ling; Huang, Guo-he; Liu, Yong-hong

2010-07-15

The performance of mesophilic anaerobic granules to degrade octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) was investigated under various conditions. The results of batch experiments showed that anaerobic granules were capable of removing HMX from aqueous solution with high efficiency. Both biotic and abiotic mechanisms contributed to the removal of HMX by anaerobic granules under mesophilic conditions. Adsorption appeared to play a significant role in the abiotic process. Furthermore, HMX could be biodegraded by anaerobic granules as the sole substrate. After 16 days of incubation, 99.04% and 96.42% of total HMX could be removed by 1g VSS/L acclimated and unacclimated granules, respectively. Vancomycin, an inhibitor of acetogenic bacteria, caused a significant inhibition of HMX biotransformation, while 2-bromoethanesulfonic acid, an inhibitor of methanogenic bacteria, only resulted in a slight decrease of metabolic activity. The presence of the glucose, as a suitable electron donor and carbon source, was found to enhance the degradation of HMX by anaerobic granules. Our study showed that sulfate had little adverse effects on biotransformation of HMX by anaerobic granules. However, nitrate had significant inhibitory effect on the extent of HMX removal especially in the initial period. This study offered good prospects of using high-rate anaerobic technology in the treatment of munition wastewater. 2010 Elsevier B.V. All rights reserved.

Expression and subcellular localization of the Qa-SNARE syntaxin17 in human eosinophils

DOE Office of Scientific and Technical Information (OSTI.GOV)

Carmo, Lívia A.S.; Dias, Felipe F.; Malta, Kássia K.

Background: SNARE members mediate membrane fusion during intracellular trafficking underlying innate and adaptive immune responses by different cells. However, little is known about the expression and function of these proteins in human eosinophils, cells involved in allergic, inflammatory and immunoregulatory responses. Here, we investigate the expression and distribution of the Qa-SNARE syntaxin17 (STX17) within human eosinophils isolated from the peripheral blood. Methods: Flow cytometry and a pre-embedding immunonanogold electron microscopy (EM) technique that combines optimal epitope preservation and secondary Fab-fragments of antibodies linked to 1.4 nm gold particles for optimal access to microdomains, were used to investigate STX17. Results: STX17more » was detected within unstimulated eosinophils. Immunogold EM revealed STX17 on secretory granules and on granule-derived vesiculotubular transport carriers (Eosinophil Sombrero Vesicles-EoSVs). Quantitative EM analyses showed that 77.7% of the granules were positive for STX17 with a mean±SEM of 3.9±0.2 gold particles/granule. Labeling was present on both granule outer membranes and matrices while EoSVs showed clear membrane-associated labeling. STX17 was also present in secretory granules in eosinophils stimulated with the cytokine tumor necrosis factor alpha (TNF-α) or the CC-chemokine ligand 11 CCL11 (eotaxin-1), stimuli that induce eosinophil degranulation. The number of secretory granules labeled for STX17 was significantly higher in CCL11 compared with the unstimulated group. The level of cell labeling did not change when unstimulated cells were compared with TNF-α-stimulated eosinophils. Conclusions: The present study clearly shows by immunanonogold EM that STX17 is localized in eosinophil secretory granules and transport vesicles and might be involved in the transport of granule-derived cargos. - Highlights: • First demonstration of the Qa-SNARE syntaxin-17 (STX17) in human eosinophils. • High resolution immunogold EM shows STX17 in granules and tubular vesicles. • Unstimulated, TNF-α or CCL11-stimulated eosinophils express STX17. • Our findings suggest a role for STX17 in the transport of granule-derived cargos.« less

DOE Office of Scientific and Technical Information (OSTI.GOV)

Peric, A.D.

Powder and granules of the high density polyethylene (PEHD) were used to prepare mortar based matrices for immobilization of radioactive waste materials containing {sup 137}Cs, as well as containers for solidified radioactive waste form. Seven types of matrices, differ due to the percentage of granules and filler material added, were investigated. PEHD powder and granules were added to mortar matrix preparations with the objective of improving physico-chemical characteristics of the radwaste-mortar matrix mixtures, in particular the leach-rate of the immobilized radionuclide, as well as mechanical characteristics either of mortar matrix and container. In this paper, only mechanical strength aspect ofmore » the investigated mortar and concrete container formulations, is presented. The equivalent diameter of the PEHD granules used was 2.0 mm. PEHD granules were used to replace 100 volume percent of stone granules, sifted size of 2.0 mm, normally used in the matrix preparation, in order to decrease the porosity and density of the mortar matrix and to avoid segregation of the stone particles at the bottom of the immobilized radioactive waste cylindrical form. PEHD powder, particle size of 250 micrometer, was added as filler to the mortar formulation, replacing 5, 8 and 10 wt% of the total cement weight in matrix formulation and 15 and 18 wt% of the total cement weight in container formulation. Cured samples were investigated on mechanical strength, using 150 MPa hydraulic press, in order to determine influence of added polyethylene granules and powder on samples resistance to mechanical forces that solidified waste materials and concrete containers may experience at the disposal site. Results of performed investigations have shown that samples prepared with polyethylene granules, replacing 100 wt% of the stone granules, have almost twice as much mechanical strength than samples prepared with stone aggregate. Samples prepared with PEHD granules and powder have mechanical strength resistance up to 13.5% higher than ones prepared with PEHD granules, solely. Improved Mechanical strength resistance of tested samples accommodates trend that functionally depends on the percentage of PEHD powder added in formulation.« less

Bone regeneration in the presence of a synthetic hydroxyapatite/silica oxide-based and a xenogenic hydroxyapatite-based bone substitute material.

PubMed

Kruse, A; Jung, R E; Nicholls, F; Zwahlen, R A; Hämmerle, C H F; Weber, F E

2011-05-01

A comparison of synthetic hydroxyapatite/silica oxide, xenogenic hydroxyapatite-based bone substitute materials with empty control sites in terms of bone regeneration enhancement in a rabbit calvarial four non-critical-sized defect model. In each of six rabbits, four bicortical calvarial bone defects were generated. The following four treatment modalities were randomly allocated: (1) empty control site, (2) synthetic hydroxyapatite/silica oxide-based (HA/SiO) test granules, (3) xenogenic hydroxyapatite -based granules, (4) synthetic hydroxyapatite/silica oxide -based (HA/SiO) test two granules. The results of the latter granules have not been reported due to their size being three times bigger than the other two granule types. After 4 weeks, the animals were sacrificed and un-decalcified sections were obtained for histological analyses. For statistical analysis, the Kruskal-Wallis test was applied (P<0.05). Histomorphometric analysis showed an average area fraction of newly formed bone of 12.32±10.36% for the empty control, 17.47±6.42% for the xenogenic hydroxyapatite -based granules group, and 21.2±5.32% for the group treated with synthetic hydroxyapatite/silica oxide -based granules. Based on the middle section, newly formed bone bridged the defect to 38.33±37.55% in the empty control group, 54.33±22.12% in the xenogenic hydroxyapatite -based granules group, and to 79±13.31% in the synthetic hydroxyapatite/silica oxide -based granules group. The bone-to-bone substitute contact was 46.38±18.98% for the xenogenic and 59.86±14.92% for the synthetic hydroxyapatite/silica oxide-based granules group. No significant difference in terms of bone formation and defect bridging could be detected between the two bone substitute materials or the empty defect. There is evidence that the synthetic hydroxyapatite/silica oxide granules provide comparable results with a standard xenogenic bovine mineral in terms of bone formation and defect bridging in non-critical size defects. © 2010 John Wiley & Sons A/S.

Effect of anti-vertigo granule on the opening number and blood flow of mouse ear capillary network

NASA Astrophysics Data System (ADS)

Li, Chongxian; Liu, Xiaobin; Li, Jun; Hao, Shaojun; Wang, Xidong; Li, Wenjun; Zhang, Zhengchen

2018-04-01

To observe the effects of anti-glare particles on the open number and blood flow in the auricle of mice with microcirculation disturbance model. Sixty mice, half male and half female, were randomly divided into 6 groups. The mice were given Kangxuan granule suspension, serum brain granule suspension and normal saline of the same volume, respectively, once a day. The mice were anesthetized by intraperitoneal injection of chloral hydrate at 1 hour after the last administration. The mouse was fixed on the observation platform and the auricle was placed on the transmission stage. BZ-2000 microcirculation microscope and microcirculation analysis system were used to observe the changes of blood velocity and capillary opening volume in auricle of mice before administration. The changes of blood velocity and capillaries opening volume of mouse auricle were observed 2 min after epinephrine injection into tail vein of mice. Bear fruit: Compared with those before epinephrine, the opening number of capillary reticulum of auricle in large dose Kangxuan granule group was significantly decreased (P<0.05), and in normal saline group and middle group. In the small dose Kangxuan granule group, the opening number of capillary network of auricle decreased significantly (P<0.01). Compared with the model group, the large dose Kangxuan granule group could significantly increase the opening number of the auricle capillary network in mice (P<0.01). Yangxuannao granule group could significantly increase the opening number of auricle capillary reticulum in mice (P<0.05), compared with the model group by Ridit test. Both Kangxuan granule group and Yangxuannao granule group could significantly improve the auricle hair of mice with microcirculation disorder. The blood flow in fine blood vessels (P<0.01). Kangxuan granule has a good effect on the opening number of capillary network of auricle and blood flow in mice with microcirculation disorder.

Killing effects of Huaier Granule combined with DC-CIK on nude mice transplanted with colon carcinoma cell line.

PubMed

Sun, Wen-Wen; Dou, Jin-Xia; Zhang, Lin; Qiao, Li-Kui; Shen, Na; Zhao, Qiang; Gao, Wen-Yuan

2017-07-11

This study aims to compare the efficacy of different treatments for nude mice transplanted with HT-29 colon carcinoma cell line. BalB/C nude mice were transplanted with HT-29 colon carcinoma cell line and randomly divided into four groups, with 5 mice in each group: blank control group, DC-CIK group, Huaier Granule group, and Huaier Granule group combined with DC-CIK group (combined treatment group). For DC-CIK group and combined treatment group, 1×106 DC-CIK cells were injected via the tail vein 4 days after transplantation. The injection was performed twice weekly for a total of 2 weeks. For Huaier Granule group and combined treatment group, Huaier Granule was administered at the dose of 20 g/60 g, by dissolving 20 g of Huaier granules in 600 ml of pure water. Intragastric administration of 0.2 ml of granules was performed once daily for 3 weeks. For the blank control group, equal volume of normal saline was given. Tumor size and body weight of nude mice were measured every 2 days during the 3-week treatment. The mice were sacrificed at the end of treatment to harvest tumors. Key genes of the signaling pathway were detected by RT-PCR. At the end of treatment, mice in combined treatment group, DC-CIK group and Huaier Granule group remained stable emotionally with normal mobility and water and food intake. However, in the blank control group, the mobility was restricted starting from the third week and the mice were on the verge of dying. The expression of PI3KR1, Akt, Wnt1, CTTNB1, Notch1, Notch2 and Notch3 genes were all downregulated significantly in the combined treatment group compared with DC-CIK group and Huaier Granule group (P<0.05). Therefore, the combined treatment of Huaier Granule combined with DC-CIK achieved the best effect in nude mice transplanted with HT-29 colon carcinoma cell line.

Influence of wet granulation and lubrication on the powder and tableting properties of codried product of microcrystalline cellulose with beta-cyclodextrin.

PubMed

Wu, J; Ho, H; Sheu, M

2001-01-01

The individual influence of wet granulation and lubrication on the powder and tableting properties of codried product of microcrystalline cellulose (MCC) with beta-cyclodextrin (beta-CD) was examined in this study. Avicel PH 101 and 301 were included for comparison. The codried product, Avicel PH 101 and 301 were granulated with water, and the granules were milled to retain three different size fractions: 37-60 microm, 60-150 microm, and 150-420 microm. The original Avicels and codried product were lubricated with magnesium stearate in three different percentages (0.2, 0.5, and 1.0%). The results showed that the powder flowability and disintegration of codried product and Avicels were significantly improved after wet granulation. However, the compactibility of codried product and Avicels decreased with increasing particle size. Nevertheless, the compactibility of the codried excipient after granulation was still better than the non-granulated Avicel PH 101 and 301. On the other hand, codried product and Avicels were sensitive to lubrication and resulted in decreasing compactibility and increasing disintegration. Because of the rounder shape of particles, the codried excipient was more sensitive to magnesium stearate and produced weaker tablets than did Avicels.

Application of image processing on struvite recovery from swine wastewater by using the fluidized bed.

PubMed

Ye, Zhi-Long; Deng, Yujun; Ye, Xin; Lou, Yaoyin; Chen, Shaohua

2018-01-01

Fluidized granulation is one of the common methods used in wastewater treatment and resource recovery with harvesting millimeter-scale large particles. Presently, effective methods are lacking to measure the fluidized granules ranging from micro- to millimeter scales, with the consequence of ineffectively controlling and optimizing the granulation process. In this work, recovering struvite (MgNH 4 PO 4 ·6H 2 O) from swine wastewater by using a fluidized bed was taken as an example. Image processing was applied to analyze the properties of different types of struvite granules, including morphology, particle size distribution, number density and mass concentration. Four stages of the struvite crystal evolution were therefore defined: aggregation, aggregate compaction, cluster-agglomerating and coating growth. These stages could occur simultaneously or sequentially. Up-flow rates of 30-80 mm/s in the fluidized bed sustained 600-876 g/L granular solids. Results revealed that the coating-growth granules were formed with compact aggregates or cluster-agglomerating granules as the nuclei. The growth rates for the different types of particles, including population growth, mass increase and particle size enlargement, were determined. In final, a schematic illustration for struvite granulation process is also presented.

Alkali-silica reactivity of expanded glass granules in structure of lightweight concrete

NASA Astrophysics Data System (ADS)

Bumanis, G.; Bajare, D.; Locs, J.; Korjakins, A.

2013-12-01

Main component in the lightweight concrete, which provides its properties, is aggregate. A lot of investigations on alkali silica reaction (ASR) between cement and lightweight aggregates have been done with their results published in the academic literature. Whereas expanded glass granules, which is relatively new product in the market of building materials, has not been a frequent research object. Therefore lightweight granules made from waste glass and eight types of cement with different chemical and mineralogical composition were examined in this research. Expanded glass granules used in this research is commercially available material produced by Penostek. Lightweight concrete mixtures were prepared by using commercial chemical additives to improve workability of concrete. The aim of the study is to identify effect of cement composition to the ASR reaction which occurs between expanded glass granules and binder. Expanded glass granules mechanical and physical properties were determined. In addition, properties of fresh and hardened concrete were determined. The ASR test was processed according to RILEM AAR-2 testing recommendation. Tests with scanning electron microscope and microstructural investigations were performed for expanded glass granules and hardened concrete specimens before and after exposing them in alkali solution.

Real-time assessment of critical quality attributes of a continuous granulation process.

PubMed

Fonteyne, Margot; Vercruysse, Jurgen; Díaz, Damián Córdoba; Gildemyn, Delphine; Vervaet, Chris; Remon, Jean Paul; De Beer, Thomas

2013-02-01

There exists the intention to shift pharmaceutical manufacturing of solid dosage forms from traditional batch production towards continuous production. The currently applied conventional quality control systems, based on sampling and time-consuming off-line analyses in analytical laboratories, would annul the advantages of continuous processing. It is clear that real-time quality assessment and control is indispensable for continuous production. This manuscript evaluates strengths and weaknesses of several complementary Process Analytical Technology (PAT) tools implemented in a continuous wet granulation process, which is part of a fully continuous from powder-to-tablet production line. The use of Raman and NIR-spectroscopy and a particle size distribution analyzer is evaluated for the real-time monitoring of critical parameters during the continuous wet agglomeration of an anhydrous theophylline- lactose blend. The solid state characteristics and particle size of the granules were analyzed in real-time and the critical process parameters influencing these granule characteristics were identified. The temperature of the granulator barrel, the amount of granulation liquid added and, to a lesser extent, the powder feed rate were the parameters influencing the solid state of the active pharmaceutical ingredient (API). A higher barrel temperature and a higher powder feed rate, resulted in larger granules.

Characterization by X-ray tomography of granulated alumina powder during in situ die compaction

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cottrino, Sandrine; Jorand, Yves, E-mail: yves.jorand@insa-lyon.fr; Maire, Eric

2013-07-15

Compaction process, the aim of which being to obtain green bodies with low porosity and small size, is often used before sintering treatment. Prior to die filling, the ceramic powder is generally granulated to improve flowability. However during compaction, density heterogeneity and critical size defects may appear due to intergranule and granule-die wall frictions. In this work, the influence of granule formulation on the compact morphology has been studied. To do so, a compaction setup was installed inside an X-ray tomography equipment so that the evolution of the compact morphology could be analysed during the whole compaction process. We havemore » demonstrated that high humidity rate and the addition of binder in the granule formulation increase density heterogeneity and generate larger defects. - Highlights: • An original compaction set up was installed inside an X-Ray tomography equipment. • The compaction process of granulated ceramic powder is imaged. • The compact green microstructure is quantified and related to the compaction stages. • The most detrimental defects of dry-pressed parts are caused by hollow granules. • Formulations without binder allow a reduction of the number of large defects.« less

Physicochemical characteristics of insulin secretion granules

PubMed Central

Coore, H. G.; Hellman, B.; Pihl, E.; Täljedal, I.-B.

1969-01-01

β-Granules were prepared from micro-dissected pancreatic islets of obese–hyperglycaemic mice. This fraction contained 60% of the insulin, 30% of the cytochrome oxidase, 16% of the acid phosphatase activity and 20% of the protein present in whole islets. The isolated granules retained a heavy metal during fractionation. Optimum conditions for granule stability were low ionic strength and pH6, the granules being unexpectedly fragile at pH7·4. The stability of the granules was unaffected by sucrose in the concentration range 50–320mm, but 1% (w/v) sodium deoxycholate released all insulin. A solubilizing effect was also noted with ATP and citrate. Spinning through 1·6m-sucrose yielded a further purification in relation to mitochondria and acid-phosphatase-carrying particles but virtually no purification in relation to protein. Electron microscopy revealed that the major contaminants were rough-surfaced vesicles and membranes. A separation of granules from acid phosphatase was achieved by phase distribution in polyethylene glycol and dextran. The location of the enzyme to the interphase was so pronounced in systems buffered with lithium phosphate that the technique may be used for future purification of acid-phosphatase-carrying particles from the β-cells. ImagesPLATE 1 PMID:4887194

A marker of animal-vegetal polarity in the egg of the sea urchin Paracentrotus lividus. The pigment band.

PubMed

Sardet, C; Chang, P

1985-09-01

We have examined the subequatorial accumulation of pigment granules (the so-called 'pigment band') in the egg of the sea urchin Paracentrotus lividus, which constitutes an unambiguous marker of animal-vegetal polarity. Most of the reddish pigment granules are situated at the periphery of the egg. They exhibit occasional saltatory movements and can aggregate into large patches. Pigment granules are retained as a band in the isolated cortex when the egg surface complex is isolated by shearing eggs attached to polylysine-coated surfaces with calcium-free isotonic solutions. Pigment granules remain as the main vesicular component of fertilized egg cortices or of unfertilized egg cortices perfused with calcium to provoke cortical granule exocytosis. They may be anchored to the isolated cortex through associations with the plasma membrane and with an extensive subsurface network of rough endoplasmic reticulum (rough ER). Pigment granules contain antimonate-precipitable calcium and, in this respect and many others, resemble acidic vesicles recently identified in the cortex of unpigmented sea urchin eggs. We discuss the similarities observed between granules and acidic vesicles in various urchin egg species and their possible functions.

Characteristics of aerobic granules grown on glucose a sequential batch shaking reactor.

PubMed

Cai, Chun-guang; Zhu, Nan-wen; Liu, Jun-shen; Wang, Zhen-peng; Cai, Wei-min

2004-01-01

Aerobic heterotrophic granular sludge was cultivated in a sequencing batch shaking reactor (SBSR) in which a synthetic wastewater containing glucose as carbon source was fed. The characteristics of the aerobic granules were investigated. Compared with the conventional activated sludge flocs, the aerobic granules exhibit excellent physical characteristics in terms of settleability, size, shape, biomass density, and physical strength. Scanning electron micrographs revealed that in mature granules little filamentous bacteria could be found, rod-shaped and coccoid bacteria were the dominant microorganisms.

In Vivo Anomalous Diffusion and Weak Ergodicity Breaking of Lipid Granules

NASA Astrophysics Data System (ADS)

Jeon, Jae-Hyung; Tejedor, Vincent; Burov, Stas; Barkai, Eli; Selhuber-Unkel, Christine; Berg-Sørensen, Kirstine; Oddershede, Lene; Metzler, Ralf

2011-01-01

Combining extensive single particle tracking microscopy data of endogenous lipid granules in living fission yeast cells with analytical results we show evidence for anomalous diffusion and weak ergodicity breaking. Namely we demonstrate that at short times the granules perform subdiffusion according to the laws of continuous time random walk theory. The associated violation of ergodicity leads to a characteristic turnover between two scaling regimes of the time averaged mean squared displacement. At longer times the granule motion is consistent with fractional Brownian motion.

Qiwei granules alleviates podocyte lesion in kidney of diabetic KK-Ay mice.

PubMed

Zhou, Jingxin; Sun, Wen; Yoshitomi, Hisae; Li, Linyi; Qin, Lingling; Guo, Xiangyu; Wu, Lili; Zhang, Yan; Wu, Xinli; Xu, Tunhai; Gao, Ming; Liu, Tonghua

2015-03-31

Chinese medicine comprised of all natural herbs is widespread used in the treatment of diabetic nephropathy (DN). Podocyte contributes to the integrity of glomerular filtration barrier whose injury plays an important role in the initiation and progression of DN. Our study aimed to investigate the effect of Qiwei granules on podocyte lesion in diabetic KK-A(y) mice kidney and its underlying mechanism. Twelve-week-old male KK-A(y) mice were randomly divided in vehicle group and Qiwei granules group, while C57BL/6J mice were used as normal control. The mice were gavage with 1.37 g/kg/day Qiwei granules or water for 10 weeks. We measured water, food intake and body weight (BW) and fasting blood glucose (FBG) every 2 weeks, and urine protein every 4 weeks. At the end of the experiment, all surviving mice were sacrificed. The kidney weight and serum renal parameters were measured, and the renal morphology was observed. To search the underlying mechanism, we examined the podocyte positive marker, slit diaphragm protein expression and some involved cell signal pathway. Qiwei granules treatment significantly improved the metabolic parameters, alleviated the urinary protein, and protected renal function in KK-A(y) mice. In addition, the glomerular injuries and podocyte lesions were mitigated with Qiwei granules treatment. Furthermore, Qiwei granules increased expression of nephrin, CD2AP, and integrin alpha3beta1 in the podocytes of KK-A(y) mice. Qiwei granules improved the phosphoration of Akt and inhibited cleaved caspase-3 protein expression. These finding suggest that Qiwei granules protects the podocyte from the development of DN via improving slit diaphragm (SD) molecules expression and likely activating Akt signaling pathway in KK-A(y) mice.

Analysis of carbohydrate storage granules in the diazotrophic cyanobacterium Cyanothece sp. PCC 7822

DOE Office of Scientific and Technical Information (OSTI.GOV)

Welkie, David G.; Sherman, Debra M.; Chrisler, William B.

The unicellular diazotrophic cyanobacteria of the genus Cyanothece demonstrate oscillations in nitrogenase activity and H 2 production when grown under 12h light-12h dark cycles. We established that Cyanothece sp. PCC 7822 allows for the construction of knock-out mutants and our objective was to improve the growth characteristics of this strain and to identify the nature of the intracellular storage granules. We report the physiological and morphological effects of reduction in nitrate and phosphate concentrations in BG-11 media on this strain. We developed a series of BG-11-derived growth media and monitored batch culture growth, nitrogenase activity and nitrogenase-mediated hydrogen production, culturemore » synchronicity, and intracellular storage content. Reduction in NaNO3 and K 2HPO 4 concentrations from 17.6 and 0.23 mM to 4.41 and 0.06 mM, respectively, improved growth characteristics such as cell size and uniformity, and enhanced the rate of cell division. Cells grown in this low NP BG-11 were less complex, a parameter that related to the composition of the intracellular storage granules. Cells grown in low NP BG-11 had less polyphosphate, fewer polyhydroxybutyrate granules and many smaller granules became evident. Biochemical analysis and transmission electron microscopy using the histocytochemical PATO technique demonstrated that these small granules contained glycogen. The glycogen levels and the number of granules per cell correlated nicely with a 2.3 to 3.3-fold change from the minimum at L0 to the maximum at D0. The differences in granule morphology and enzymes between Cyanothece ATCC 51142 and Cyanothece PCC 7822 provide insights into the formation of large starch-like granules in some cyanobacteria.« less

Expression and subcellular localization of the Qa-SNARE syntaxin17 in human eosinophils.

PubMed

Carmo, Lívia A S; Dias, Felipe F; Malta, Kássia K; Amaral, Kátia B; Shamri, Revital; Weller, Peter F; Melo, Rossana C N

2015-10-01

SNARE members mediate membrane fusion during intracellular trafficking underlying innate and adaptive immune responses by different cells. However, little is known about the expression and function of these proteins in human eosinophils, cells involved in allergic, inflammatory and immunoregulatory responses. Here, we investigate the expression and distribution of the Qa-SNARE syntaxin17 (STX17) within human eosinophils isolated from the peripheral blood. Flow cytometry and a pre-embedding immunonanogold electron microscopy (EM) technique that combines optimal epitope preservation and secondary Fab-fragments of antibodies linked to 1.4 nm gold particles for optimal access to microdomains, were used to investigate STX17. STX17 was detected within unstimulated eosinophils. Immunogold EM revealed STX17 on secretory granules and on granule-derived vesiculotubular transport carriers (Eosinophil Sombrero Vesicles-EoSVs). Quantitative EM analyses showed that 77.7% of the granules were positive for STX17 with a mean±SEM of 3.9±0.2 gold particles/granule. Labeling was present on both granule outer membranes and matrices while EoSVs showed clear membrane-associated labeling. STX17 was also present in secretory granules in eosinophils stimulated with the cytokine tumor necrosis factor alpha (TNF-α) or the CC-chemokine ligand 11 CCL11 (eotaxin-1), stimuli that induce eosinophil degranulation. The number of secretory granules labeled for STX17 was significantly higher in CCL11 compared with the unstimulated group. The level of cell labeling did not change when unstimulated cells were compared with TNF-α-stimulated eosinophils. The present study clearly shows by immunanonogold EM that STX17 is localized in eosinophil secretory granules and transport vesicles and might be involved in the transport of granule-derived cargos. Copyright © 2015 Elsevier Inc. All rights reserved.