Posttranscriptional regulation of alpha-amylase II-4 expression by gibberellin in germinating rice seeds.

PubMed

Nanjo, Yohei; Asatsuma, Satoru; Itoh, Kimiko; Hori, Hidetaka; Mitsui, Toshiaki; Fujisawa, Yukiko

2004-06-01

Hormonal regulation of expression of alpha-amylase II-4 that lacks the gibberellin-response cis-element (GARE) in the promoter region of the gene was studied in germinating rice (Oryza sativa L.) seeds. Temporal and spatial expression of alpha-amylase II-4 in the aleurone layer were essentially identical to those of alpha-amylase I-1 whose gene contains GARE, although these were distinguishable in the embryo tissues at the early stage of germination. The gibberellin-responsible expression of alpha-amylase II-4 was also similar to that of alpha-amylase I-1. However, the level of alpha-amylase II-4 mRNA was not increased by gibberellin, indicating that the transcriptional enhancement of alpha-amylase II-4 expression did not occur in the aleurone. Gibberellin stimulated the accumulation of 45Ca2+ into the intracellular secretory membrane system. In addition, several inhibitors for Ca2+ signaling, such as EGTA, neomycin, ruthenium red (RuR), and W-7 prevented the gibberellin-induced expression of alpha-amylase II-4 effectively. While the gibberellin-induced expression of alpha-amylase II-4 occurred normally in the aleurone layer of a rice dwarf mutant d1 which is defective in the alpha subunit of the heterotrimeric G protein. Based on these results, it was concluded that the posttranscriptional regulation of alpha-amylase II-4 expression by gibberellin operates in the aleurone layer of germinating rice seed, which is mediated by Ca2+ but not the G protein.

Psychological stress-induced changes in salivary alpha-amylase and adrenergic activity.

PubMed

Kang, Younhee

2010-12-01

The aim of the study was to examine the relationships among salivary alpha-amylase, plasma catecholamines, blood pressure, and heart rate during psychological stress. This study used a pretest-post-test experimental design with a control group, using repeated measures. A total of 33 participants was divided into the experimental group (n = 16) that underwent a college academic final test as the psychological stress and the control group (n = 17) that did not undergo the test. The levels of salivary alpha-amylase and plasma catecholamines, blood pressure, and heart rate were measured seven times and stress and anxiety were measured once and twice, respectively, as subjective stress markers. Significant changes in the level of salivary alpha-amylase were found in response to psychological stress. However, the correlations of salivary alpha-amylase with the plasma catecholamines, blood pressure, and heart rate were only partially found to be statistically significant. In conclusion, it was shown that salivary alpha-amylase was sensitive to stress throughout this study. Thus, salivary alpha-amylase may be used to measure stress uninvasively in both clinical settings and nursing research where the effects of stress might be scrutinized. Furthermore, the mechanisms of illnesses that are induced by stress could be explored. © 2010 Blackwell Publishing Asia Pty Ltd.

One-step production of immobilized alpha-amylase in recombinant Escherichia coli.

PubMed

Rasiah, Indira A; Rehm, Bernd H A

2009-04-01

Industrial enzymes are often immobilized via chemical cross-linking onto solid supports to enhance stability and facilitate repeated use in bioreactors. For starch-degrading enzymes, immobilization usually places constraints on enzymatic conversion due to the limited diffusion of the macromolecular substrate through available supports. This study describes the one-step immobilization of a highly thermostable alpha-amylase (BLA) from Bacillus licheniformis and its functional display on the surface of polyester beads inside engineered Escherichia coli. An optimized BLA variant (Termamyl) was N-terminally fused to the polyester granule-forming enzyme PhaC of Cupriavidus necator. The fusion protein lacking the signal sequence mediated formation of stable polyester beads exhibiting alpha-amylase activity. The alpha-amylase beads were assessed with respect to alpha-amylase activity, which was demonstrated qualitatively and quantitatively. The immobilized alpha-amylase showed Michaelis-Menten enzyme kinetics exerting a V(max) of about 506 mU/mg of bead protein with a K(m) of about 5 microM, consistent with that of free alpha-amylase. The stability of the enzyme at 85 degrees C and the capacity for repeated usage in a starch liquefaction process were also demonstrated. In addition, structural integrity and functionality of the beads at extremes of pH and temperature, demonstrating their suitability for industrial use, were confirmed by electron microscopy and protein/enzyme analysis. This study proposes a novel, cost-effective method for the production of immobilized alpha-amylase in a single step by using the polyester granules forming protein PhaC as a fusion partner in engineered E. coli.

Improvement of heterologous protein production in Aspergillus oryzae by RNA interference with alpha-amylase genes.

PubMed

Nemoto, Takashi; Maruyama, Jun-ichi; Kitamoto, Katsuhiko

2009-11-01

Aspergillus oryzae RIB40 has three alpha-amylase genes (amyA, amyB, and amyC), and secretes alpha-amylase abundantly. However, large amounts of endogenous secretory proteins such as alpha-amylase can compete with heterologous protein in the secretory pathway and decrease its production yields. In this study, we examined the effects of suppression of alpha-amylase on heterologous protein production in A. oryzae, using the bovine chymosin (CHY) as a reporter heterologous protein. The three alpha-amylase genes in A. oryzae have nearly identical DNA sequences from those promoters to the coding regions. Hence we performed silencing of alpha-amylase genes by RNA interference (RNAi) in the A. oryzae CHY producing strain. The silenced strains exhibited a reduction in alpha-amylase activity and an increase in CHY production in the culture medium. This result suggests that suppression of alpha-amylase is effective in heterologous protein production in A. oryzae.

Molecular cloning and characterization of an alpha-amylase from Pichia burtonii 15-1.

PubMed

Kato, Saemi; Shimizu-Ibuka, Akiko; Mura, Kiyoshi; Takeuchi, Akiko; Tokue, Chiyoko; Arai, Soichi

2007-12-01

An alpha-amylase secreted by Pichia burtonii 15-1 isolated from a traditional starter murcha of Nepal, named Pichia burtonii alpha-amylase (PBA), was studied. The gene was cloned and its nucleotide sequence was determined. PBA was deduced to consist of 494 amino acid residues. It shared certain degrees of amino acid sequence identity with other homologous proteins: 60% with Schwanniomyces occidentalis alpha-amylase, 58% with Saccharomycopsis sp. alpha-amylase, and 47% with Taka-amylase A from Aspergillus oryzae. A three-dimensional structural model of PBA generated using the known three-dimensional structure of Taka-amylase A as a template suggested high structural similarity between them. Kinetic analysis revealed that the K(m) values of PBA were lower than those of Taka-amylase A for the oligosaccharides. Although the k(cat) values of PBA were lower than those of Taka-amylase A for the oligosaccharide substrates, the k(cat)/K(m) values of PBA were higher.

Long-term stability of diurnal salivary cortisol and alpha-amylase secretion patterns.

PubMed

Skoluda, Nadine; La Marca, Roberto; Gollwitzer, Mario; Müller, Andreas; Limm, Heribert; Marten-Mittag, Birgitt; Gündel, Harald; Angerer, Peter; Nater, Urs M

2017-06-01

This study aimed to investigate long-term stability and variability of diurnal cortisol and alpha-amylase patterns. Diurnal cortisol and alpha-amylase secretion patterns were assessed on a single workday with three waves of measurement across a total time period of 24months in 189 participants. Separate hierarchical linear models were analyzed, with and without a number of potential predictor variables (age, BMI, smoking, chronic stress, stress reactivity). While low long-term stability was found in diurnal cortisol, the stability of diurnal alpha-amylase was moderate across the time period of 24months. Several predictor variables had a positive impact on diurnal cortisol and alpha-amylase secretion patterns averaged across waves. Our findings underpin the notion that long-term stability is not necessarily warranted in longitudinal studies. It is important to choose an appropriate study design when attempting to disentangle clinically and biologically relevant changes from naturally occurring variations in diurnal cortisol and alpha-amylase. Copyright © 2017 Elsevier Inc. All rights reserved.

Effect of cellulase, xylanase and α-amylase combinations on the rheological properties of Chinese steamed bread dough enriched in wheat bran.

PubMed

Liu, Wenjun; Brennan, Margaret Anne; Serventi, Luca; Brennan, Charles Stephen

2017-11-01

The present study investigates the effects of α-amylase (6 and 10ppm), xylanase (70 and 120ppm) and cellulase (35 and 60ppm) on the rheological properties of bread dough. The mixing property of dough was measured by using a DoughLAB. The extension and stickiness of dough were analysed using the Texture Analyzer. The results illustrate that the addition of single enzyme and enzyme combinations can increase the extensibility, softening, mixing tolerance index (MTI) and stickiness, whereas decrease the resistance to extension. For water absorption, the addition of single enzyme had no significant effect, while the combination enzyme significantly (p<0.05) decreased the values from 63.9 to 59.6% (wheat flour dough) and 71.4-67.1% (dough incorporated with 15% wheat bran). Compared to the single enzyme with the value of 34.1mm, enzyme combination (6, 120 and 60ppm) increased the extensibility of wheat flour dough by up to 42%. Additionally, combination of α-amylase, xylanase and cellulase had a synergetic effect on the dough rheology. Copyright © 2017 Elsevier Ltd. All rights reserved.

The bean. alpha. -amylase inhibitor is encoded by a lectin gene

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

Moreno, J.; Altabella, T.; Chrispeels, M.J.

The common bean, Phaseolus vulgaris, contains an inhibitor of insect and mammalian {alpha}-amylases that does not inhibit plant {alpha}-amylase. This inhibitor functions as an anti-feedant or seed-defense protein. We purified this inhibitor by affinity chromatography and found that it consists of a series of glycoforms of two polypeptides (Mr 14,000-19,000). Partial amino acid sequencing was carried out, and the sequences obtained are identical with portions of the derived amino acid sequence of a lectin-like gene. This lectin gene encodes a polypeptide of MW 28,000, and the primary in vitro translation product identified by antibodies to the {alpha}-amylase inhibitor has themore » same size. Co- and posttranslational processing of this polypeptide results in glycosylated polypeptides of 14-19 kDa. Our interpretation of these results is that the bean lectins constitute a gene family that encodes diverse plant defense proteins, including phytohemagglutinin, arcelin and {alpha}-amylase inhibitor.« less

Prediction of ingredient quality and the effect of a combination of xylanase, amylase, protease and phytase in the diets of broiler chicks. 2. Energy and nutrient utilisation.

PubMed

Cowieson, A J; Singh, D N; Adeola, O

2006-08-01

1. In order to investigate the effects of xylanase, amylase, protease and phytase in the diets of broiler chickens containing graded concentrations of metabolisable energy (ME), two 42-d experiments were conducted using a total of 2208 broiler chicks (8 treatments with 12 replicate pens in each experiment). 2. Four diets including one positive and three negative control diets were used. Three maize/soybean meal-based negative control (NC) diets were formulated to be identical in available phosphorus (P), calcium (Ca) and amino acids but NC1 contained approximately 0.17 MJ/kg less ME than NC2 and approximately 0.34 MJ/kg less ME than NC3. A positive control (PC) was fed for comparison and was formulated to be adequate in all nutrients, providing approximately 0.63 MJ/kg ME, 0.13% available P, 0.12% Ca and 1 to 2% amino acids more than NC1. 3. The reduction in nutrient density between NC1 and PC was determined using ingredient quality models Avichecktrade mark Corn and Phychecktrade mark that can predict the response to exogenous enzymes in maize/soybean meal-based broiler diets. Supplementation of each diet with or without a cocktail of xylanase, amylase, protease and phytase gave a total of 8 dietary treatments in a 4 x 2 factorial arrangement. The same treatments and diet designs were used in both experiments but conducted in different locations using different batches of maize, soybean meal and minor ingredients. 4. In both experiments, digestibility was improved by the addition of exogenous enzymes, particularly those for P, Ca and certain amino acids. In addition, the supplementation of the PC with enzymes elicited a positive response indicating that over-the-top addition of xylanase, amylase, protease and phytase may offer a nutritionally and economically viable alternative to feed cost reduction. 5. It can be concluded that the digestibility of nutrients by broilers fed on maize/soybean meal-based diets can be improved by the use of a combination of xylanase

Cloning of a yeast alpha-amylase promoter and its regulated heterologous expression

DOEpatents

Gao, Johnway [Richland, WA; Skeen, Rodney S [Pendleton, OR; Hooker, Brian S [Kennewick, WA; Anderson, Daniel B [Pasco, WA

2003-04-01

The present invention provides the promoter clone discovery of an alpha-amylase gene of a starch utilizing yeast strain Schwanniomyces castellii. The isolated alpha-amylase promoter is an inducible promoter, which can regulate strong gene expression in starch culture medium.

Two Strategies for Microbial Production of an Industrial Enzyme-Alpha-Amylase

NASA Technical Reports Server (NTRS)

Bernhardsdotter, Eva C. M. J.; Garriott, Owen; Pusey, Marc L.; Ng, Joseph D.

2003-01-01

Extremophiles are microorganisms that thrive in, from an anthropocentric view, extreme environments including hot springs, soda lakes and arctic water. This ability of survival at extreme conditions has rendered extremophiles to be of interest in astrobiology, evolutionary biology as well as in industrial applications. Of particular interest to the biotechnology industry are the biological catalysts of the extremophiles, the extremozymes, whose unique stabilities at extreme conditions make them potential sources of novel enzymes in industrial applications. There are two major approaches to microbial enzyme production. This entails enzyme isolation directly from the natural host or creating a recombinant expression system whereby the targeted enzyme can be overexpressed in a mesophilic host. We are employing both methods in the effort to produce alpha-amylases from a hyperthermophilic archaeon (Thermococcus) isolated from a hydrothermal vent in the Atlantic Ocean, as well as from alkaliphilic bacteria (Bacillus) isolated from a soda lake in Tanzania. Alpha-amylases catalyze the hydrolysis of internal alpha-1,4-glycosidic linkages in starch to produce smaller sugars. Thermostable alpha-amylases are used in the liquefaction of starch for production of fructose and glucose syrups, whereas alpha-amylases stable at high pH have potential as detergent additives. The alpha-amylase encoding gene from Thermococcus was PCR amplified using carefully designed primers and analyzed using bioinformatics tools such as BLAST and Multiple Sequence Alignment for cloning and expression in E.coli. Four strains of Bacillus were grown in alkaline starch-enriched medium of which the culture supernatant was used as enzyme source. Amylolytic activity was detected using the starch-iodine method.

Patterns of cortisol and alpha-amylase reactivity to psychosocial stress in maltreated women

PubMed Central

Mielock, Alyssa S.; Morris, Matthew C.; Rao, Uma

2016-01-01

Background Childhood maltreatment can trigger enduring changes in major stress response systems, particularly in the context of major depressive disorder (MDD). However, the relative impact of maltreatment versus MDD on hypothalamic-pituitary-adrenal axis and sympathetic-adrenal-medullary system stress reactivity is not well understood. Method This study examined salivary cortisol and alpha-amylase responses to the Trier Social Stress Test (TSST) in 26 maltreated (15 with current MDD) and 26 non-maltreated (17 with current MDD) women. Results Maltreated women showed greater anticipatory cortisol reactivity during the TSST protocol compared to non-maltreated women. Maltreated women also showed rapid deceleration in cortisol levels. Whereas non-maltreated women showed initial declines in alpha-amylase levels but rapidly increasing alpha-amylase levels during the TSST protocol, maltreated women did not exhibit changes in alpha-amylase levels during the TSST protocol. Contrary to expectation, MDD did not impact cortisol or alpha-amylase responses. Limitations The present study is limited by retrospective report of childhood maltreatment, cross-sectional design, and modest sample sizes. Conclusions These findings suggest that childhood maltreatment plays a greater role driving alterations in cortisol and alpha-amylase stress reactivity than MDD. Understanding the biological embedding of maltreatment is critical for elucidating mechanisms linking these experiences to risk for negative mental and physical health outcomes. PMID:27875756

Patterns of cortisol and alpha-amylase reactivity to psychosocial stress in maltreated women.

PubMed

Mielock, Alyssa S; Morris, Matthew C; Rao, Uma

2017-02-01

Childhood maltreatment can trigger enduring changes in major stress response systems, particularly in the context of major depressive disorder (MDD). However, the relative impact of maltreatment versus MDD on hypothalamic-pituitary-adrenal axis and sympathetic-adrenal-medullary system stress reactivity is not well understood. This study examined salivary cortisol and alpha-amylase responses to the Trier Social Stress Test (TSST) in 26 maltreated (15 with current MDD) and 26 non-maltreated (17 with current MDD) women. Maltreated women showed greater anticipatory cortisol reactivity during the TSST protocol compared to non-maltreated women. Maltreated women also showed rapid deceleration in cortisol levels. Whereas non-maltreated women showed initial declines in alpha-amylase levels but rapidly increasing alpha-amylase levels during the TSST protocol, maltreated women did not exhibit changes in alpha-amylase levels during the TSST protocol. Contrary to expectation, MDD did not impact cortisol or alpha-amylase responses. The present study is limited by retrospective report of childhood maltreatment, cross-sectional design, and modest sample sizes. These findings suggest that childhood maltreatment plays a greater role driving alterations in cortisol and alpha-amylase stress reactivity than MDD. Understanding the biological embedding of maltreatment is critical for elucidating mechanisms linking these experiences to risk for negative mental and physical health outcomes. Copyright © 2016 Elsevier B.V. All rights reserved.

Biochemical characterization of the alpha-amylase inhibitor in mungbeans and its application in inhibiting the growth of Callosobruchus maculatus.

PubMed

Wisessing, Anussorn; Engkagul, Arunee; Wongpiyasatid, Arunee; Choowongkomon, Kiattawee

2010-02-24

The insect Callosobruchus maculatus causes considerable damage to harvested mungbean seeds every year, which leads to commercial losses. However, recent studies have revealed that mungbean seeds contain alpha-amylase inhibitors that can inhibit the protein C. maculatus, preventing growth and development of the insect larvae in the seed, thus preventing further damage. For this reason, the use of alpha-amylase inhibitors to interfere with the pest's digestion process has become an interesting alternative biocontrolling agent. In this study, we have isolated and purified the alpha-amylase inhibitor from mungbean seeds (KPS1) using ammonium sulfate precipitation, gel filtration chromatography and reversed phase HPLC. We found that the alpha-amylase inhibitor, isolated as a monomer, had a molecular weight of 27 kDa. The alpha-amylase inhibitor was purified 750-fold with a final yield of 0.4 mg of protein per 30 g of mungbean seeds. Its specific activity was determined at 14.5 U (mg of protein)(-1). Interestingly, we found that the isolated alpha-amylase inhibitor inhibits C. maculatus alpha-amylase but not human salivary alpha-amylase. After preincubation of the enzyme with the inhibitor, the mungbean alpha-amylase inhibitor inhibited C. maculatus alpha-amylase activity by decreasing V(max) while increasing the K(m) constant, indicating that the mungbean alpha-amylase is a mix noncompetitive inhibitor. The in vivo effect of alpha-amylase inhibitor on the mortality of C. maculatus shows that the alpha-amylase inhibitor acts on C. maculatus during the development stage, by reducing carbohydrate digestion necessary for growth and development, rather than during the end laying/hatching stage. Our results suggest that mungbean alpha-amylase inhibitor could be a useful future biocontrolling agent.

Prediction of ingredient quality and the effect of a combination of xylanase, amylase, protease and phytase in the diets of broiler chicks. 1. Growth performance and digestible nutrient intake.

PubMed

Cowieson, A J; Singh, D N; Adeola, O

2006-08-01

1. A total of 2208 broiler chicks were used in two growth experiments (8 treatments and 12 replicate pens in each experiment) to assess the effects of xylanase, amylase, protease and phytase in maize-based diets. 2. A positive control diet was formulated containing adequate nutrient concentrations. A negative control diet was formulated to contain approximately 628 kJ/kg, 0.13%, 0.12% and 1 to 2% less metabolisable energy (ME), phosphorus (P), calcium (Ca) and amino acids, respectively, than the positive control. In addition, two further negative control diets that contained 167 or 334 kJ/kg more ME, respectively, than negative control 1 were formulated. 3. A further 4 dietary treatments were made by supplementing each of the 4 negative control diets with a combination of xylanase, amylase, protease and phytase, resulting in 8 dietary treatments in a 4 by 2 factorial arrangement. 4. The scale of the removal of energy, P, Ca and amino acids from the positive control diet was determined using least square models based on in vivo data for both the xylanase/amylase/protease cocktail and for phytase and it was predicted that performance of birds fed on negative control 1 would be returned by supplemental enzymes to that of those fed on the positive control. 5. In both experiments there was a significantly poorer performance in birds fed on the negative control 1 than in those fed on the positive control. The poorer weight gain and feed conversion ratio could be attributed in part to a reduced intake of digestible energy, P, nitrogen (N) and amino acids associated with birds fed on the negative control diet. 6. Supplementation of the negative control diets with the enzyme combination returned performance to that of the positive control in both experiments. 7. These data indicate that exogenous xylanase, amylase, protease and phytase can be used successfully in a strategically formulated low nutrient density diet to maintain performance to that of birds fed on a

Alpha-amylase inhibitor-1 gene from Phaseolus vulgaris expressed in Coffea arabica plants inhibits alpha-amylases from the coffee berry borer pest.

PubMed

Barbosa, Aulus E A D; Albuquerque, Erika V S; Silva, Maria C M; Souza, Djair S L; Oliveira-Neto, Osmundo B; Valencia, Arnubio; Rocha, Thales L; Grossi-de-Sa, Maria F

2010-06-17

Coffee is an important crop and is crucial to the economy of many developing countries, generating around US$70 billion per year. There are 115 species in the Coffea genus, but only two, C. arabica and C. canephora, are commercially cultivated. Coffee plants are attacked by many pathogens and insect-pests, which affect not only the production of coffee but also its grain quality, reducing the commercial value of the product. The main insect-pest, the coffee berry borer (Hypotheneumus hampei), is responsible for worldwide annual losses of around US$500 million. The coffee berry borer exclusively damages the coffee berries, and it is mainly controlled by organochlorine insecticides that are both toxic and carcinogenic. Unfortunately, natural resistance in the genus Coffea to H. hampei has not been documented. To overcome these problems, biotechnological strategies can be used to introduce an alpha-amylase inhibitor gene (alpha-AI1), which confers resistance against the coffee berry borer insect-pest, into C. arabica plants. We transformed C. arabica with the alpha-amylase inhibitor-1 gene (alpha-AI1) from the common bean, Phaseolus vulgaris, under control of the seed-specific phytohemagglutinin promoter (PHA-L). The presence of the alpha-AI1 gene in six regenerated transgenic T1 coffee plants was identified by PCR and Southern blotting. Immunoblotting and ELISA experiments using antibodies against alpha-AI1 inhibitor showed a maximum alpha-AI1 concentration of 0.29% in crude seed extracts. Inhibitory in vitro assays of the alpha-AI1 protein against H. hampei alpha-amylases in transgenic seed extracts showed up to 88% inhibition of enzyme activity. This is the first report showing the production of transgenic coffee plants with the biotechnological potential to control the coffee berry borer, the most important insect-pest of crop coffee.

Tobacco plants transformed with the bean. alpha. ai gene express an inhibitor of insect. alpha. -amylase in their seeds. [Nicotiana tabacum; Tenebrio molitor

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

Altabella, T.; Chrispeels, M.J.

Bean (Phaseolus vulgaris L.) seeds contain a putative plant defense protein that inhibits insect and mammalian but not plant {alpha}-amylases. We recently presented strong circumstantial evidence that this {alpha}-amylase inhibitor ({alpha}Al) is encoded by an already-identified lectin gene whose product is referred to as lectin-like-protein (LLP). We have now made a chimeric gene consisting of the coding sequence of the lectin gene that encodes LLP and the 5{prime} and 3{prime} flanking sequences of the lectin gene that encodes phytohemagglutinin-L. When this chimeric gene was expressed in transgenic tobacco (Nicotiana tabacum), we observed in the seeds a series of polypeptides (M{submore » r} 10,000-18,000) that cross-react with antibodies to the bean {alpha}-amylase inhibitor. Most of these polypeptides bind to a pig pancreas {alpha}-amylase affinity column. An extract of the seeds of the transformed tobacco plants inhibits pig pancreas {alpha}-amylase activity as well as the {alpha}-amylase present in the midgut of Tenebrio molitor. We suggest that introduction of this lectin gene (to be called {alpha}ai) into other leguminous plants may be a strategy to protect the seeds from the seed-eating larvae of Coleoptera.« less

Pectinase from Trichoderma reesei QM 9414

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

Haltmeier, T.; Leisola, M.; Ulmer, D.

1983-06-01

The present study was undertaken to determine whether T. reesei produces pectinolytic enzymes and, if so, to determine their action pattern and specificity. The aim was also to find out the practical importance of pectinolytic activity in the hydrolysis of cellulosic materials. It was found that the presence of pectinase does not appear to facilitate the hydrolysis of cellulose in plant material through any cell-separating mechanism. It is concluded that cellulase and xylanase activities are the important activities in the practical hydrolysis of cellulosic materials and that lignin is the real limiting factor. (Refs. 12).

Phylogenetic and Comparative Sequence Analysis of Thermostable Alpha Amylases of kingdom Archea, Prokaryotes and Eukaryotes.

PubMed

Huma, Tayyaba; Maryam, Arooma; Rehman, Shahid Ur; Qamar, Muhammad Tahir Ul; Shaheen, Tayyaba; Haque, Asma; Shaheen, Bushra

2014-01-01

Alpha amylase family is generally defined as a group of enzymes that can hydrolyse and transglycosylase α-(1, 4) or α-(1, 6) glycosidic bonds along with the preservation of anomeric configuration. For the comparative analysis of alpha amylase family, nucleotide sequences of seven thermo stable organisms of Kingdom Archea i.e. Pyrococcus furiosus (100-105°C), Kingdom Prokaryotes i.e. Bacillus licheniformis (90-95°C), Geobacillus stearothermophilus (75°C), Bacillus amyloliquefaciens (72°C), Bacillus subtilis (70°C) and Bacillus KSM K38 (55°C) and Eukaryotes i.e. Aspergillus oryzae (60°C) were selected from NCBI. Primary structure composition analysis and Conserved sequence analysis were conducted through Bio Edit tools. Results from BioEdit shown only three conserved regions of base pairs and least similarity in MSA of the above mentioned alpha amylases. In Mega 5.1 Phylogeny of thermo stable alpha amylases of Kingdom Archea, Prokaryotes and Eukaryote was handled by Neighbor-Joining (NJ) algorithm. Mega 5.1 phylogenetic results suggested that alpha amylases of thermo stable organisms i.e. Pyrococcus furiosus (100-105°C), Bacillus licheniformis (90-95°C), Geobacillus stearothermophilus (75°C) and Bacillus amyloliquefaciens (72°C) are more distantly related as compared to less thermo stable organisms. By keeping in mind the characteristics of most thermo stable alpha amylases novel and improved features can be introduced in less thermo stable alpha amylases so that they become more thermo tolerant and productive for industry.

Physical and catalytic properties of alpha-amylase from Tenebrio molitor L. larvae.

PubMed Central

Buonocore, V; Poerio, E; Silano, V; Tomasi, M

1976-01-01

The amylase from Tenebrio molitor L. larvae (yellow mealworm) was characterized according to a number of its molecular and catalytic properties. The insect amylase is a single polypeptide chain with mol.wt. 68000, an isoelectric point of 4.0 and a very low content of sulphur-containing amino acids. The enzyme is a Ca2+-protein and behaves as an alpha-amylase. Removal of Ca2+ by exhaustive dialysis against water causes the irreversible inactivation of the enzyme. Moreover, the enzyme is activated by the presence in the assay mixture of Cl-, or some other inorganic anions that are less effective than Cl-, and is inhibited by F-. Optimal conditions of pH and temperature for the enzymic activity are 5.8 and 37 degrees C. The insect amylase exhibits an identical kinetic behaviour toward starch, amylose and amylopectin; the enzyme hydrolyses glycogen with a higher affinity constant. Compared with the non-insect alpha-amylases described in the literature, Tenebrio molitor amylase has a lower affinity for starch. PMID:942374

Exposure to inhalable dust, wheat flour and alpha-amylase allergens in industrial and traditional bakeries.

PubMed

Bulat, Petar; Myny, Katrien; Braeckman, Lutgart; van Sprundel, Marc; Kusters, Edouard; Doekes, Gert; Pössel, Kerstin; Droste, Jos; Vanhoorne, Michel

2004-01-01

This study was designed to characterize exposure to inhalable dust, wheat flour and alpha-amylase allergens in industrial and traditional bakeries. The study included 70 bakeries from the northern part of Belgium. Based on the degree of automation and a clear division of individual job tasks, four bakeries were identified as industrial and the remaining 66 were identified as traditional ones. Personal, as well as stationary, samples of inhalable dust were collected during full shift periods, usually 5-7 h. The portable pumps aspirated 2 l/min through Teflon personal dust samplers (Millipore, pore size 1.0 microm) mounted in PAS-6 sampling heads. In the collected samples the inhalable dust, wheat flour and alpha-amylase allergens were determined. Wheat flour allergens were measured using enzyme-linked immunosorbent assay inhibition and an antiwheat IgG4 serum pool. The alpha-amylase allergens were measured using a sandwich enzyme immunoassay with affinity-purified polyclonal rabbit IgG antibodies. In total, 440 samples (300 personal and 140 stationary) were processed. The highest inhalable dust exposure was observed in traditional bakeries among bread [geometric mean (GM) 2.10 mg/m3] and bread and pastry workers (GM 1.80 mg/m3). In industrial bakeries the highest dust exposure was measured in bread-producing workers (GM 1.06 mg/m3). Similar relations were observed for wheat flour and alpha-amylase allergens. Bread baking workers in traditional bakeries had the highest exposure to both allergens (wheat flour GM 22.33 microg/m(3), alpha-amylase GM 0.61 ng/m3). The exposure to wheat flour and alpha-amylase allergens in industrial bakeries was higher in bread baking workers (wheat flour GM 6.15 microg/m3, alpha-amylase GM 0.47 ng/m3) than in bread packing workers (wheat flour GM 2.79 microg/m3, alpha-amylase GM 0.15 ng/m3). The data presented suggest that, on average, exposure in the Belgium bakeries studied-industrial as well as traditional-is lower than or similar to

Encapsulation of alpha-amylase into starch-based biomaterials: an enzymatic approach to tailor their degradation rate.

PubMed

Azevedo, Helena S; Reis, Rui L

2009-10-01

This paper reports the effect of alpha-amylase encapsulation on the degradation rate of a starch-based biomaterial. The encapsulation method consisted in mixing a thermostable alpha-amylase with a blend of corn starch and polycaprolactone (SPCL), which were processed by compression moulding to produce circular disks. The presence of water was avoided to keep the water activity low and consequently to minimize the enzyme activity during the encapsulation process. No degradation of the starch matrix occurred during processing and storage (the encapsulated enzyme remained inactive due to the absence of water), since no significant amount of reducing sugars was detected in solution. After the encapsulation process, the released enzyme activity from the SPCL disks after 28days was found to be 40% comparatively to the free enzyme (unprocessed). Degradation studies on SPCL disks, with alpha-amylase encapsulated or free in solution, showed no significant differences on the degradation behaviour between both conditions. This indicates that alpha-amylase enzyme was successfully encapsulated with almost full retention of its enzymatic activity and the encapsulation of alpha-amylase clearly accelerates the degradation rate of the SPCL disks, when compared with the enzyme-free disks. The results obtained in this work show that degradation kinetics of the starch polymer can be controlled by the amount of encapsulated alpha-amylase into the matrix.

A kinetic model to explain the maximum in alpha-amylase activity measurements in the presence of small carbohydrates.

PubMed

Baks, Tim; Janssen, Anja E M; Boom, Remko M

2006-06-20

The effect of the presence of several small carbohydrates on the measurement of the alpha-amylase activity was determined over a broad concentration range. At low carbohydrate concentrations, a distinct maximum in the alpha-amylase activity versus concentration curves was observed in several cases. At higher concentrations, all carbohydrates show a decreasing alpha-amylase activity at increasing carbohydrate concentrations. A general kinetic model has been developed that can be used to describe and explain these phenomena. This model is based on the formation of a carbohydrate-enzyme complex that remains active. It is assumed that this complex is formed when a carbohydrate binds to alpha-amylase without blocking the catalytic site and its surrounding subsites. Furthermore, the kinetic model incorporates substrate inhibition and substrate competition. Depending on the carbohydrate type and concentration, the measured alpha-amylase activity can be 75% lower than the actual alpha-amylase activity. The model that has been developed can be used to correct for these effects in order to obtain the actual amount of active enzyme. 2006 Wiley Periodicals, Inc.

Enzymatic activities in different strains isolated from healthy and brittle leaf disease affected date palm leaves: study of amylase production conditions.

PubMed

Mouna, Jrad; Imen, Fendri; Choba Ines, Ben; Nourredine, Drira; Adel, Kadri; Néji, Gharsallah

2015-02-01

The present study aimed to investigate and compare the enzymatic production of endophytic bacteria isolated from healthy and brittle leaf disease affected date palm leaves (pectinase, cellulase, lipase, and amylase). The findings revealed that the enzymatic products from the bacterial isolates of healthy date palm leaves were primarily 33% amylolytic enzyme, 33 % cellulase, 25 % pectinase, and 25 % lipase. The isolates from brittle leaf disease date palm leaves, on the other hand, were noted to produce 16 % amylolytic enzyme, 20 % cellulose, 50 % pectinase, and 50 % lipase. The effects of temperature and pH on amylase, pectinase, and cellulose activities were investigated. The Bacillus subtilis JN934392 strain isolated from healthy date palm leaves produced higher levels of amylase activity at pH 7. A Box Behnken Design (BBD) was employed to optimize amylase extraction. Maximal activity was observed at pH and temperature ranges of pH 6-6.5 and 37-39 °C, respectively. Under those conditions, amylase activity was noted to be attained 9.37 U/ml. The results showed that the enzyme was able to maintain more than 50 % of its activity over a temperature range of 50-80 °C, with an optimum at 70 °C. This bacterial amylase showed high activity compared to other bacteria, which provides support for its promising candidacy for future industrial application.

LaaA, a novel high-active alkalophilic alpha-amylase from deep-sea bacterium Luteimonas abyssi XH031(T).

PubMed

Song, Qinghao; Wang, Yan; Yin, Chong; Zhang, Xiao-Hua

2016-08-01

Alpha-amylase is a kind of broadly used industrial enzymes, most of which have been exploited from terrestrial organism. Comparatively, alpha-amylase from marine environment was largely undeveloped. In this study, a novel alkalophilic alpha-amylase with high activity, Luteimonas abyssi alpha-amylase (LaaA), was cloned from deep-sea bacterium L. abyssi XH031(T) and expressed in Escherichia coli BL21. The gene has a length of 1428bp and encodes 475 amino acids with a 35-residue signal peptide. The specific activity of LaaA reached 8881U/mg at the optimum pH 9.0, which is obvious higher than other reported alpha-amylase. This enzyme can remain active at pH levels ranging from 6.0 to 11.0 and temperatures below 45°C, retaining high activity even at low temperatures (almost 38% residual activity at 10°C). In addition, 1mM Na(+), K(+), and Mn(2+) enhanced the activity of LaaA. To investigate the function of potential active sites, R227G, D229K, E256Q/H, H327V and D328V mutants were generated, and the results suggested that Arg227, Asp229, Glu256 and Asp328 were total conserved and essential for the activity of alpha-amylase LaaA. This study shows that the alpha-amylase LaaA is an alkali-tolerant and high-active amylase with strong potential for use in detergent industry. Copyright © 2016 Elsevier Inc. All rights reserved.

Validation of an assay for quantification of alpha-amylase in saliva of sheep

PubMed Central

Fuentes-Rubio, Maria; Fuentes, Francisco; Otal, Julio; Quiles, Alberto; Hevia, María Luisa

2016-01-01

The objective of this study was to develop a time-resolved immunofluorometric assay (TR-IFMA) for quantification of salivary alpha-amylase in sheep. For that purpose, after the design of the assay, an analytical and a clinical validation were carried out. The analytical validation of the assay showed intra- and inter-assay coefficients of variation (CVs) of 6.1% and 10.57%, respectively and an analytical limit of detection of 0.09 ng/mL. The assay also demonstrated a high level of accuracy, as determined by linearity under dilution. For clinical validation, a model of acute stress testing was conducted to determine whether expected significant changes in alpha-amylase were picked up in the newly developed assay. In that model, 11 sheep were immobilized and confronted with a sheepdog to induce stress. Saliva samples were obtained before stress induction and 15, 30, and 60 min afterwards. Salivary cortisol was measured as a reference of stress level. The results of TR-IFMA showed a significant increase (P < 0.01) in the concentration of alpha-amylase in saliva after stress induction. The assay developed in this study could be used to measure salivary alpha-amylase in the saliva of sheep and this enzyme could be a possible noninvasive biomarker of stress in sheep. PMID:27408332

Effects of a dietary Aspergillus oryzae extract containing alpha-amylase activity on performance and carcass characteristics of finishing beef cattle.

PubMed

Tricarico, J M; Abney, M D; Galyean, M L; Rivera, J D; Hanson, K C; McLeod, K R; Harmon, D L

2007-03-01

Three experiments were conducted to examine the effects of an Aspergillus oryzae extract containing alpha-amylase activity on performance and carcass characteristics of finishing beef cattle. In Exp. 1, 120 crossbred steers were used in a randomized complete block design to evaluate the effects of roughage source (alfalfa hay vs. cottonseed hulls) and supplemental alpha-amylase at 950 dextrinizing units (DU)/kg of DM. Significant roughage source x alpha-amylase interactions (P < 0.05) were observed for performance. In steers fed cottonseed hulls, supplemental alpha-amylase increased ADG through d 28 and 112 and tended (P < 0.15) to increase ADG in all other periods. The increases in ADG were related to increased DMI and efficiency of gain during the initial 28-d period but were primarily related to increased DMI as the feeding period progressed. Supplemental alpha-amylase increased (P = 0.02) the LM area across both roughage sources. In Exp. 2, 96 crossbred heifers were used in a randomized complete block design with a 2 x 3 factorial arrangement of treatments to evaluate the effects of corn processing (dry cracked vs. high moisture) and supplemental alpha-amylase concentration (0, 580, or 1,160 DU/kg of DM). Alpha-amylase supplementation increased DMI (P = 0.05) and ADG (P = 0.03) during the initial 28 d on feed and carcass-adjusted ADG (P = 0.04) across corn processing methods. Longissimus muscle area was greatest (quadratic effect, P = 0.04), and yield grade was least (quadratic effect, P = 0.02) in heifers fed 580 DU of alpha-amylase/kg of DM across corn processing methods. In Exp. 3, 56 crossbred steers were used in a randomized complete block design to evaluate the effects of supplemental alpha-amylase (930 DU/kg of DM) on performance when DMI was restricted to yield a programmed ADG. Alpha-amylase supplementation did not affect performance when DMI was restricted. We conclude that dietary alpha-amylase supplementation of finishing beef diets may result in

Construction of promoter-probe shuttle vectors for Escherichia coli and corynebacteria on the basis of promoterless alpha-amylase gene.

PubMed

Ugorcáková, J; Bukovská, G; Timko, J

2000-01-01

We constructed new promoter-probe vectors for E. coli and corynebacteria based on the promoterless alpha-amylase gene originating from Bacillus subtilis. Vectors pJUPAE1 and pJUPAE2 are suitable for isolation of transcriptionally active fragments from plasmids, phages or genomic DNA. alpha-Amylase activity can be easily visually detected on agar plates containing a chromogenic substrate, or by direct measurement of alpha-amylase activity.

Alpha-amylase Inhibition and Antioxidant Activity of Marine Green Algae and its Possible Role in Diabetes Management.

PubMed

Unnikrishnan, P S; Suthindhiran, K; Jayasri, M A

2015-10-01

In the continuing search for safe and efficient antidiabetic drug, marine algae become important source which provide several compounds of immense therapeutic potential. Alpha-amylase, alpha-glucosidase inhibitors, and antioxidant compounds are known to manage diabetes and have received much attention recently. In the present study, four green algae (Chaetomorpha aerea, Enteromorpha intestinalis, Chlorodesmis, and Cladophora rupestris) were chosen to evaluate alpha-amylase, alpha-glucosidase inhibitory, and antioxidant activity in vitro. The phytochemical constituents of all the extracts were qualitatively determined. Antidiabetic activity was evaluated by inhibitory potential of extracts against alpha-amylase and alpha-glucosidase by spectrophotometric assays. Antioxidant activity was determined by 2,2-diphenyl-1-picrylhydrazyl, hydrogen peroxide (H2O2), and nitric oxide scavenging assay. Gas chromatography-mass spectrometry (GC-MS) analysis was carried out to determine the major compound responsible for its antidiabetic action. Among the various extracts screened, chloroform extract of C. aerea (IC50 - 408.9 μg/ml) and methanol extract of Chlorodesmis (IC50 - 147.6 μg/ml) showed effective inhibition against alpha-amylase. The extracts were also evaluated for alpha-glucosidase inhibition, and no observed activity was found. Methanol extract of C. rupestris showed notable free radical scavenging activity (IC50 - 666.3 μg/ml), followed by H2O2 (34%) and nitric oxide (49%). Further, chemical profiling by GC-MS revealed the presence of major bioactive compounds. Phenol, 2,4-bis (1,1-dimethylethyl) and z, z-6,28-heptatriactontadien-2-one were predominantly found in the methanol extract of C. rupestris and chloroform extract of C. aerea. Our results demonstrate that the selected algae exhibit notable alpha-amylase inhibition and antioxidant activity. Therefore, characterization of active compounds and its in vivo assays will be noteworthy. Four green algae were

Alpha-amylase Inhibition and Antioxidant Activity of Marine Green Algae and its Possible Role in Diabetes Management

PubMed Central

Unnikrishnan, P. S.; Suthindhiran, K.; Jayasri, M. A.

2015-01-01

Aim: In the continuing search for safe and efficient antidiabetic drug, marine algae become important source which provide several compounds of immense therapeutic potential. Alpha-amylase, alpha-glucosidase inhibitors, and antioxidant compounds are known to manage diabetes and have received much attention recently. In the present study, four green algae (Chaetomorpha aerea, Enteromorpha intestinalis, Chlorodesmis, and Cladophora rupestris) were chosen to evaluate alpha-amylase, alpha-glucosidase inhibitory, and antioxidant activity in vitro. Materials and Methods: The phytochemical constituents of all the extracts were qualitatively determined. Antidiabetic activity was evaluated by inhibitory potential of extracts against alpha-amylase and alpha-glucosidase by spectrophotometric assays. Antioxidant activity was determined by 2,2-diphenyl-1-picrylhydrazyl, hydrogen peroxide (H2O2), and nitric oxide scavenging assay. Gas chromatography-mass spectrometry (GC-MS) analysis was carried out to determine the major compound responsible for its antidiabetic action. Results: Among the various extracts screened, chloroform extract of C. aerea (IC50 − 408.9 μg/ml) and methanol extract of Chlorodesmis (IC50 − 147.6 μg/ml) showed effective inhibition against alpha-amylase. The extracts were also evaluated for alpha-glucosidase inhibition, and no observed activity was found. Methanol extract of C. rupestris showed notable free radical scavenging activity (IC50 – 666.3 μg/ml), followed by H2O2 (34%) and nitric oxide (49%). Further, chemical profiling by GC-MS revealed the presence of major bioactive compounds. Phenol, 2,4-bis (1,1-dimethylethyl) and z, z-6,28-heptatriactontadien-2-one were predominantly found in the methanol extract of C. rupestris and chloroform extract of C. aerea. Conclusion: Our results demonstrate that the selected algae exhibit notable alpha-amylase inhibition and antioxidant activity. Therefore, characterization of active compounds and its in vivo

Progress of pancreatitis disease biomarker alpha amylase enzyme by new nano optical sensor.

PubMed

Attia, M S; Al-Radadi, Najlaa S

2016-12-15

A new nano optical sensor binuclear Pd-(2-aminothiazole) (urea), Pd(atz,ur) complex was prepared and characterized for the assessment of the activity of alpha amylase enzyme in urine and serum samples for early diagnosis of Pancreatitis disease. The assessment of alpha amylase activity is carried out by the quenching of the luminescence intensity of the nano optical sensor binuclear Pd(atz,ur) complex at 457nm by the 2-chloro-4-nitrophenol (2-CNP) which produced from the reaction of the enzyme with 2-chloro-4-nitrophenyl-α-d-maltotrioside (CNPG3) substrate. The remarkable quenching of the luminescence intensity at 457nm of nano Pd(atz,ur) doped in sol-gel matrix by various concentrations of the 2-CNP was successfully used as an optical sensor for the assessment of α-amylase activity. The calibration plot was achieved over the concentration range 8.5×10(-6) to 1.9×10(-9)molL(-1) 2-CNP with a correlation coefficient of (0.999) and a detection limit of (7.4×10(-10)molL(-1)). The method was used satisfactorily for the assessment of the α-amylase activity over activity range (3-321U/L) in different urine and serum samples of pancreatitis patients. The assessment of the alpha amylase biomarker by the proposed method increases its sensitivity (96.88%) and specificity (94.41%) for early diagnosis of pancreatitis diseases. Copyright © 2016 Elsevier B.V. All rights reserved.

Aerobic Fitness Level Affects Cardiovascular and Salivary Alpha Amylase Responses to Acute Psychosocial Stress.

PubMed

Wyss, Thomas; Boesch, Maria; Roos, Lilian; Tschopp, Céline; Frei, Klaus M; Annen, Hubert; La Marca, Roberto

2016-12-01

Good physical fitness seems to help the individual to buffer the potential harmful impact of psychosocial stress on somatic and mental health. The aim of the present study is to investigate the role of physical fitness levels on the autonomic nervous system (ANS; i.e. heart rate and salivary alpha amylase) responses to acute psychosocial stress, while controlling for established factors influencing individual stress reactions. The Trier Social Stress Test for Groups (TSST-G) was executed with 302 male recruits during their first week of Swiss Army basic training. Heart rate was measured continuously, and salivary alpha amylase was measured twice, before and after the stress intervention. In the same week, all volunteers participated in a physical fitness test and they responded to questionnaires on lifestyle factors and personal traits. A multiple linear regression analysis was conducted to determine ANS responses to acute psychosocial stress from physical fitness test performances, controlling for personal traits, behavioural factors, and socioeconomic data. Multiple linear regression revealed three variables predicting 15 % of the variance in heart rate response (area under the individual heart rate response curve during TSST-G) and four variables predicting 12 % of the variance in salivary alpha amylase response (salivary alpha amylase level immediately after the TSST-G) to acute psychosocial stress. A strong performance at the progressive endurance run (high maximal oxygen consumption) was a significant predictor of ANS response in both models: low area under the heart rate response curve during TSST-G as well as low salivary alpha amylase level after TSST-G. Further, high muscle power, non-smoking, high extraversion, and low agreeableness were predictors of a favourable ANS response in either one of the two dependent variables. Good physical fitness, especially good aerobic endurance capacity, is an important protective factor against health

Human Parotid Gland Alpha-Amylase Secretion as a Function of Chronic Hyperbaric Exposure

DTIC Science & Technology

1979-01-01

parotid ...Pullman, WA 99163 Gilman, S. C, G. J. Fischer, R. J. Biersner, R. D. Thornton, and D. A. Miller. 1979. Human parotid gland alpha-amylase secretion...as a function of chronic hyperbaric exposure. Undersea Biomed. Res. 6(3):303-307.—Secretion of a-amylase by the human parotid gland increased

Higher levels of salivary alpha-amylase predict failure of cessation efforts in male smokers.

PubMed

Dušková, M; Simůnková, K; Hill, M; Hruškovičová, H; Hoskovcová, P; Králíková, E; Stárka, L

2010-01-01

The ability to predict the success or failure of smoking cessation efforts will be useful for clinical practice. Stress response is regulated by two primary neuroendocrine systems. Salivary cortisol has been used as a marker for the hypothalamus-pituitary-adrenocortical axis and salivary alpha-amylase as a marker for the sympathetic adrenomedullary system. We studied 62 chronic smokers (34 women and 28 men with an average age of 45.2+/-12.9 years). The levels of salivary cortisol and salivary alpha-amylase were measured during the period of active smoking, and 6 weeks and 24 weeks after quitting. We analyzed the men separately from the women. The men who were unsuccessful in cessation showed significantly higher levels of salivary alpha-amylase over the entire course of the cessation attempt. Before stopping smoking, salivary cortisol levels were higher among the men who were unsuccessful in smoking cessation. After quitting, there were no differences between this group and the men who were successful in cessation. In women we found no differences between groups of successful and unsuccessful ex-smokers during cessation. In conclusions, increased levels of salivary alpha-amylase before and during smoking cessation may predict failure to quit in men. On the other hand, no advantage was found in predicting the failure to quit in women. The results of our study support previously described gender differences in smoking cessation.

Activation of bean (Phaseolus vulgaris) [alpha]-amylase inhibitor requires proteolytic processing of the proprotein

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

Pueyo, J.J.; Hunt, D.C.; Chrispeels, M.J.

Seeds of the common bean (Phaseolus vulgaris) contain a plant defense protein that inhibits the [alpha]-amylases of mammals and insects. This [alpha]-amylase inhibitor ([alpha]Al) is synthesized as a proprotein on the endoplasmic reticulum and is proteolytically processed after arrival in the protein storage vacuoles to polypeptides of relative molecular weight (M[sub r]) 15,000 to 18,000. The authors report two types of evidence that proteolytic processing is linked to activation of the inhibitory activity. First, by surveying seed extracts of wild accessions of P. vulgaris and other species in the genus Phaseolus, they found that antibodies to [alpha]Al recognize large (M[submore » r] 30,000-35,000) polypeptides as well as typical [alpha]Al processing products (M[sub r] 15,000-18,000). [alpha]Al activity was found in all extracts that had the typical [alpha]Al processed polypeptides, but was absent from seed extracts that lacked such polypeptides. Second, they made a mutant [alpha]Al in which asparagine-77 is changed to aspartic acid-77. This mutation slows down the proteolytic processing of pro-[alpha]Al when the gene is expressed in tobacco. When pro-[alpha]Al was separated from mature [alpha]Al by gel filtration, pro-[alpha]Al was found not to have [alpha]-amylase inhibitory activity. The authors interpret these results to mean that formation of the active inhibitor is causally related to proteolytic processing of the proprotein. They suggest that the polypeptide cleavage removes a conformation constraint on the precursor to produce the biochemically active molecule. 43 refs., 5 figs., 1 tab.« less

Effect of neohesperidin dihydrochalcone on the activity and stability of alpha-amylase: a comparative study on bacterial, fungal, and mammalian enzymes.

PubMed

Kashani-Amin, Elaheh; Ebrahim-Habibi, Azadeh; Larijani, Bagher; Moosavi-Movahedi, Ali Akbar

2015-10-01

Neohesperidin dihydrochalcone (NHDC) was recently introduced as an activator of mammalian alpha-amylase. In the current study, the effect of NHDC has been investigated on bacterial and fungal alpha-amylases. Enzyme assays and kinetic analysis demonstrated the capability of NHDC to significantly activate both tested alpha-amylases. The ligand activation pattern was found to be more similar between the fungal and mammalian enzyme in comparison with the bacterial one. Further, thermostability experiments indicated a stability increase in the presence of NHDC for the bacterial enzyme. In silico (docking) test locates a putative binding site for NHDC on alpha-amylase surface in domain B. This domain shows differences in various alpha-amylase types, and the different behavior of the ligand toward the studied enzymes may be attributed to this fact. Copyright © 2015 John Wiley & Sons, Ltd.

Production of alpha-amylase from Aspergillus oryzae for several industrial applications in a single step.

PubMed

Porfirif, María C; Milatich, Esteban J; Farruggia, Beatriz M; Romanini, Diana

2016-06-01

A one-step method as a strategy of alpha-amylase concentration and purification was developed in this work. This methodology requires the use of a very low concentration of biodegradable polyelectrolyte (Eudragit(®) E-PO) and represents a low cost, fast, easy to scale up and non-polluting technology. Besides, this methodology allows recycling the polymer after precipitation. The formation of reversible soluble/insoluble complexes between alpha-amylase and the polymer Eudragit(®) E-PO was studied, and their precipitation in selected conditions was applied with bioseparation purposes. Turbidimetric assays allowed to determine the pH range where the complexes are insoluble (4.50-7.00); pH 5.50 yielded the highest turbidity of the system. The presence of NaCl (0.05M) in the medium totally dissociates the protein-polymer complexes. When the adequate concentration of polymer was added under these conditions to a liquid culture of Aspergillus oryzae, purification factors of alpha-amylase up to 7.43 and recoveries of 88% were obtained in a simple step without previous clarification. These results demonstrate that this methodology is suitable for the concentration and production of alpha-amylase from this source and could be applied at the beginning of downstream processing. Copyright © 2016 Elsevier B.V. All rights reserved.

Molecular cloning of alpha-amylases from cotton boll weevil, Anthonomus grandis and structural relations to plant inhibitors: an approach to insect resistance.

PubMed

Oliveira-Neto, Osmundo B; Batista, João A N; Rigden, Daniel J; Franco, Octávio L; Falcão, Rosana; Fragoso, Rodrigo R; Mello, Luciane V; dos Santos, Roseane C; Grossi-de-Sá, Maria F

2003-01-01

Anthonomus grandis, the cotton boll weevil, causes severe cotton crop losses in North and South America. Here we demonstrate the presence of starch in the cotton pollen grains and young ovules that are the main A. grandis food source. We further demonstrate the presence of alpha-amylase activity, an essential enzyme of carbohydrate metabolism for many crop pests, in A. grandis midgut. Two alpha-amylase cDNAs from A. grandis larvae were isolated using RT-PCR followed by 5' and 3' RACE techniques. These encode proteins with predicted molecular masses of 50.8 and 52.7kDa, respectively, which share 58% amino acid identity. Expression of both genes is induced upon feeding and concentrated in the midgut of adult insects. Several alpha-amylase inhibitors from plants were assayed against A. grandis alpha-amylases but, unexpectedly, only the BIII inhibitor from rye kernels proved highly effective, with inhibitors generally active against other insect amylases lacking effect. Structural modeling of Amylag1 and Amylag2 showed that different factors seem to be responsible for the lack of effect of 0.19 and alpha-AI1 inhibitors on A. grandis alpha-amylase activity. This work suggests that genetic engineering of cotton to express alpha-amylase inhibitors may offer a novel route to A. grandis resistance.

General Subject 1. Report to ICUMSA on the determination of commercial alpha-amylase activity by a spectrophotometric method

USDA-ARS?s Scientific Manuscript database

A report is given on a new industrial method for the determination of the activity or strength of commercial alpha-amylase at a sugarcane factory or refinery, as well as a recommendation. At the present time, the activities or strengths of commercial alpha-amylases cannot be directly compared becau...

The effect of alpha amylase enzyme on quality of sweet sorghum juice for chrystal sugar

NASA Astrophysics Data System (ADS)

Marwati, T.; Cahyaningrum, N.; Widodo, S.; Astiati, U. T.; Budiyanto, A.; Wahyudiono; Arif, A. B.; Richana, N.

2018-01-01

Sweet sorghum juice (Sorghum bicolor L. Moench) has characteristics similar to sugar cane juice and potentially used for sugar substitutes that can support food security. Nevertheless the sweet sorghum juicecontain starch which impede sorghum sugar crystallization. Therefore, research on the enzymatic process is needed to convert starch into reducing sugar. The experimental design used was the Factorial Randomized Design with the first factor was alpha amylase enzyme concentration (0, 20, 40, 60, 80, 100, 120 μL/100 mL) and second factor was incubation time (0, 30, 60, 90 minute) at temperature 100°C. The experiment was conducted on fresh sweet sorghum. The results showed that the addition of the alpha amylase enzyme increased the content of reducing sugar and decreased levels of starch. Elevating concentration of alpha amylase enzyme will increase the reducing sugar content in sweet sorghum juice. The optimum alpha amylase enzyme concentration to produce the highest total sugar was 80 μL/100 mL of sweet sorghum juice with the optimum incubation time was 90 minutes. The results of this study are expected to create a new sweetener for sugar substitution. From the economic prospective aspect, sorghum is a potential crop and can be relied upon to support the success of the food diversification program which further leads to the world food security

Salivary Alpha-Amylase Reactivity in Breast Cancer Survivors

PubMed Central

Wan, Cynthia; Couture-Lalande, Marie-Ève; Narain, Tasha A.; Lebel, Sophie; Bielajew, Catherine

2016-01-01

The two main components of the stress system are the hypothalamic-pituitary-adrenal (HPA) and sympathetic-adrenal-medullary (SAM) axes. While cortisol has been commonly used as a biomarker of HPA functioning, much less attention has been paid to the role of the SAM in this context. Studies have shown that long-term breast cancer survivors display abnormal reactive cortisol patterns, suggesting a dysregulation of their HPA axis. To fully understand the integrity of the stress response in this population, this paper explored the diurnal and acute alpha-amylase profiles of 22 breast cancer survivors and 26 women with no history of cancer. Results revealed that breast cancer survivors displayed identical but elevated patterns of alpha-amylase concentrations in both diurnal and acute profiles relative to that of healthy women, F (1, 39) = 17.95, p < 0.001 and F (1, 37) = 7.29, p = 0.010, respectively. The average area under the curve for the diurnal and reactive profiles was 631.54 ± 66.94 SEM and 1238.78 ± 111.84 SEM, respectively. This is in sharp contrast to their cortisol results, which showed normal diurnal and blunted acute patterns. The complexity of the stress system necessitates further investigation to understand the synergistic relationship of the HPA and SAM axes. PMID:27023572

Salivary Alpha-Amylase Reactivity in Breast Cancer Survivors.

PubMed

Wan, Cynthia; Couture-Lalande, Marie-Ève; Narain, Tasha A; Lebel, Sophie; Bielajew, Catherine

2016-03-23

The two main components of the stress system are the hypothalamic-pituitary-adrenal (HPA) and sympathetic-adrenal-medullary (SAM) axes. While cortisol has been commonly used as a biomarker of HPA functioning, much less attention has been paid to the role of the SAM in this context. Studies have shown that long-term breast cancer survivors display abnormal reactive cortisol patterns, suggesting a dysregulation of their HPA axis. To fully understand the integrity of the stress response in this population, this paper explored the diurnal and acute alpha-amylase profiles of 22 breast cancer survivors and 26 women with no history of cancer. Results revealed that breast cancer survivors displayed identical but elevated patterns of alpha-amylase concentrations in both diurnal and acute profiles relative to that of healthy women, F (1, 39) = 17.95, p < 0.001 and F (1, 37) = 7.29, p = 0.010, respectively. The average area under the curve for the diurnal and reactive profiles was 631.54 ± 66.94 SEM and 1238.78 ± 111.84 SEM, respectively. This is in sharp contrast to their cortisol results, which showed normal diurnal and blunted acute patterns. The complexity of the stress system necessitates further investigation to understand the synergistic relationship of the HPA and SAM axes.

The green tea polyphenol (-)-epigallocatechin gallate precipitates salivary proteins including alpha-amylase: biochemical implications for oral health.

PubMed

Hara, Kumiko; Ohara, Masaru; Hayashi, Ikue; Hino, Takamune; Nishimura, Rumi; Iwasaki, Yoriko; Ogawa, Tetsuji; Ohyama, Yoshihiko; Sugiyama, Masaru; Amano, Hideaki

2012-04-01

Green tea is a popular drink throughout the world, and it contains various components, including the green tea polyphenol (-)-epigallocatechin gallate (EGCG). Tea interacts with saliva upon entering the mouth, so the interaction between saliva and EGCG interested us, especially with respect to EGCG-protein binding. SDS-PAGE revealed that several salivary proteins were precipitated after adding EGCG to saliva. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) peptide mass fingerprinting indicated that the major proteins precipitated by EGCG were alpha-amylase, S100, and cystatins. Surface plasmon resonance revealed that EGCG bound to alpha-amylase at dissociation constant (K(d)) = 2.74 × 10(-6) M, suggesting that EGCG interacts with salivary proteins with a relatively strong affinity. In addition, EGCG inhibited the activity of alpha-amylase by non-competitive inhibition, indicating that EGCG is effective at inhibiting the formation of fermentable carbohydrates involved in caries formation. Interestingly, alpha-amylase reduced the antimicrobial activity of EGCG against the periodontal bacterium Aggregatibacter actinomycetemcomitans. Therefore, we considered that EGCG-salivary protein interactions might have both protective and detrimental effects with respect to oral health. © 2012 Eur J Oral Sci.

Purification and properties of an alpha-amylase protein-inhibitor from Arachis hypogaea seeds.

PubMed

Irshad, M; Sharma, C B

1981-06-15

A protein showing highly specific inhibitory activity towards hog pancreatic and human salivary alpha-amylases (1,4-alpha-D-glucan glucanohydrolase, EC 3.2.1.1), but not towards plant and bacterial alpha-amylases, has been purified 197-fold from an aqueous extract of peanut cotyledons using heat treatment, (NH4)2SO4 precipitation and ion-exchange chromatography on DEAE- and CM-cellulose. The purified inhibitor was homogeneous by polyacrylamide gel electrophoresis. Its molecular weight, as determined by Sephadex G-100 gel-filtration, and its electrophoretic mobility at pH 8 relative to bromophenol blue, were 25 000 and 0.14, respectively. The inhibitory activity was relatively resistant to thermal treatment and markedly increased when the inhibitor was preincubated with the enzyme before the addition of starch. Further, the inhibition was found to be pH-dependent and non-competitive in nature.

Relationship Between Meditation Depth and Waking Salivary Alpha-Amylase Secretion Among Long-Term MBSR Instructors.

PubMed

Haslam, Alyson; Wirth, Michael D; Robb, Sara Wagner

2017-08-01

The purpose of this study was to characterize sympathetic activity by using waking salivary alpha-amylase (sAA) concentrations in a group of long-term meditation instructors and to examine the association between meditation (depth, dose and duration) and the waking alpha-amylase response. Salivary alpha-amylase samples were collected (immediately upon waking and at 15-min, 30-min and 45-min intervals after waking) from mindfulness-based stress reduction instructors to determine both the area under the curve and the awakening slope (difference in alpha-amylase concentrations between waking and 30-min post-waking). It was determined through general linear models that neither years of meditation nor meditation dose were associated with the awakening sAA slope, but higher scores for meditation depth (greater depth) was associated with a more negative (or steeper) awakening slope [Quartile (Q)1: -7 versus Q4: -21 U/mL; p = 0.06], in fully adjusted models. Older age (p = 0.04) and a later time of waking (p < 0.01) also were associated with less negative awakening slope values. Smoking was associated with lower area under the curve values (smokers: 1716 U/mL versus nonsmokers: 2107 U/mL; p = 0.05) in fully adjusted models. The results suggest a 'healthy' sAA waking slope among individuals who meditate more deeply. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

Crystal structures of the psychrophilic alpha-amylase from Alteromonas haloplanctis in its native form and complexed with an inhibitor.

PubMed Central

Aghajari, N.; Feller, G.; Gerday, C.; Haser, R.

1998-01-01

Alteromonas haloplanctis is a bacterium that flourishes in Antarctic sea-water and it is considered as an extreme psychrophile. We have determined the crystal structures of the alpha-amylase (AHA) secreted by this bacterium, in its native state to 2.0 angstroms resolution as well as in complex with Tris to 1.85 angstroms resolution. The structure of AHA, which is the first experimentally determined three-dimensional structure of a psychrophilic enzyme, resembles those of other known alpha-amylases of various origins with a surprisingly greatest similarity to mammalian alpha-amylases. AHA contains a chloride ion which activates the hydrolytic cleavage of substrate alpha-1,4-glycosidic bonds. The chloride binding site is situated approximately 5 angstroms from the active site which is characterized by a triad of acid residues (Asp 174, Glu 200, Asp 264). These are all involved in firm binding of the Tris moiety. A reaction mechanism for substrate hydrolysis is proposed on the basis of the Tris inhibitor binding and the chloride activation. A trio of residues (Ser 303, His 337, Glu 19) having a striking spatial resemblance with serine-protease like catalytic triads was found approximately 22 angstroms from the active site. We found that this triad is equally present in other chloride dependent alpha-amylases, and suggest that it could be responsible for autoproteolytic events observed in solution for this cold adapted alpha-amylase. PMID:9541387

Natural products as alpha-amylase and alpha-glucosidase inhibitors and their hypoglycaemic potential in the treatment of diabetes: an update.

PubMed

Tundis, R; Loizzo, M R; Menichini, F

2010-04-01

The inhibition of alpha-glucosidase and alpha-amylase, enzymes involved in the digestion of carbohydrates, can significantly reduce the post-prandial increase of blood glucose and therefore can be an important strategy in the management of blood glucose level in type 2 diabetic and borderline patients. Currently, there is renewed interest in plant-based medicines and functional foods modulating physiological effects in the prevention and cure of diabetes and obesity. The plant kingdom is a wide field to search for natural effective oral hypoglycaemic agents that have slight or no side effects. More than ca. 1200 plant species have been recorded to be used empirically worldwide for their alleged hypoglycaemic activity. Therefore, natural alpha-glucosidase and alpha-amylase inhibitors from plant sources offer an attractive strategy for the control of hyperglycaemia. This article reviews recent data on plant extracts and isolated natural compounds that are being tested for their hypglycaemic activity, highlights ongoing research and considers the future persepctives.

Screening of traditional antidiabetic medicinal plants of Mauritius for possible alpha-amylase inhibitory effects in vitro.

PubMed

Kotowaroo, M I; Mahomoodally, M F; Gurib-Fakim, A; Subratty, A H

2006-03-01

In this study, seven exotic/indigenous medicinal plants of Mauritius, namely Coix lacryma-jobi (Poaceae), Aegle marmelos (Rutaceae), Artocarpus heterophyllus (Moraceae), Vangueria madagascariensis (Rubiaceae), Azadirachta indica (Meliaceae), Eriobotrya japonica (Rosaceae) and Syzigium cumini (Myrtaceae) were studied for possible effects on starch breakdown by alpha-amylase in vitro. The results showed that only Artocarpus heterophyllus significantly (p < 0.05) inhibited alpha-amylase activity in vitro. To confirm the observed effects, a further biochemical assay was undertaken to investigate the effects of Artocarpus heterophyllus on alpha-amylase activity using rat plasma in vitro. It was found that the aqueous leaf extract significantly (p < 0.05) inhibited alpha-amylase activity in rat plasma. The highest inhibitory activity (27.20 +/- 5.00%) was observed at a concentration of 1000 microg/mL. However, in both cases dose dependency was not observed. Enzyme kinetic studies using the Michaelis-Menten and Lineweaver-Burk equations were performed to establish the type of inhibition involved. In the presence of the plant extract the maximal velocity (Vmax) remained constant (1/150 g / L/s) whereas the Michaelis-Menten constant (Km) increased by 5.79 g / L, indicating that the aqueous leaf extract of Artocarpus heterophyllus behaved as a competitive inhibitor. Results from the present study tend to indicate that Artocarpus heterophyllus could act as a 'starch blocker' thereby reducing post-prandial glucose peaks. Copyright 2006 John Wiley & Sons, Ltd.

Two forms of alpha-amylase in mantle tissue of Mytilus galloprovincialis: purification and molecular properties of form II.

PubMed

Lombraña, M; Suárez, P; San Juan, F

2005-09-01

alpha-Amylase activity has been shown for the first time in a non-digestive tissue from Mytilus galloprovincialis. alpha-amylase from mussel mantle tissue has been purified by affinity chromatography on insoluble starch, followed by gel-filtration chromatography on Superdex-200. The chromatographic and electrophoretic behaviour of M. galloprovincialis alpha-amylase and stability characteristics suggest two forms of this enzyme: one form forming stable aggregates (form I) and a monomeric form (form II) that is more abundant, active and unstable. Both forms show an inverse quantitative variation. Purified form II was highly unstable and the molecular mass was estimated to be 66 kDa by sodium dodecyl sulphate (SDS)-gel electrophoresis. Maximum activity was noted at pH 6.5 and 35 degrees C.

Reflection on design and testing of pancreatic alpha-amylase inhibitors: an in silico comparison between rat and rabbit enzyme models

PubMed Central

2012-01-01

Background Inhibitors of pancreatic alpha-amylase are potential drugs to treat diabetes and obesity. In order to find compounds that would be effective amylase inhibitors, in vitro and in vivo models are usually used. The accuracy of models is limited, but these tools are nonetheless valuable. In vitro models could be used in large screenings involving thousands of chemicals that are tested to find potential lead compounds. In vivo models are still used as preliminary mean of testing compounds behavior in the whole organism. In the case of alpha-amylase inhibitors, both rats and rabbits could be chosen as in vivo models. The question was which animal could present more accuracy with regard to its pancreatic alpha-amylase. Results As there is no crystal structure of these enzymes, a molecular modeling study was done in order to compare the rabbit and rat enzymes with the human one. The overall result is that rabbit enzyme could probably be a better choice in this regard, but in the case of large ligands, which could make putative interactions with the −4 subsite of pancreatic alpha-amylase, interpretation of results should be made cautiously. Conclusion Molecular modeling tools could be used to choose the most suitable model enzyme that would help to identify new enzyme inhibitors. In the case of alpha-amylase, three-dimensional structures of animal enzymes show differences with the human one which should be taken into account when testing potential new drugs. PMID:23352052

Identification and properties of an alpha-amylase from a strain of Eubacterium sp. isolated from the rat intestinal tract.

PubMed

Delahaye, E P; Foglietti, M J; Andrieux, C; Chardon-Loriaux, I; Szylit, O; Raibaud, P

1991-01-01

1. A bacterial amylase was isolated from the intestinal content of monoxenic rats inoculated with Eubacterium sp. B86. 2. Affinity chromatography on cross-linked starch allowed its separation from rat endogenous amylases. 3. The bacterial enzyme was characterized by its pI, molecular weight and action pattern. It behaves as a typical endo-amylase (alpha-amylase).

Isolation and characterisation of a novel alpha-amylase from the extreme haloarchaeon Haloterrigena turkmenica.

PubMed

Santorelli, Marco; Maurelli, Luisa; Pocsfalvi, Gabriella; Fiume, Immacolata; Squillaci, Giuseppe; La Cara, Francesco; Del Monaco, Giovanni; Morana, Alessandra

2016-11-01

An extracellular halophilic alpha-amylase (AmyA) was produced by the haloarchaeon Haloterrigena turkmenica grown in medium enriched with 0.2% (w/v) starch. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and size exclusion chromatography (SEC) analyses showed a major band at 66.0kDa and a peak of 54.0kDa, respectively. Analysis of tryptic fragments of the protein present in the major SDS-PAGE band by nano-LC-ESI-MS/MS led to identification of the alpha-amylase catalytic region, encoded by the htur2110 gene, as the protein possessing the described activity. Optimal values for activity were 55°C, pH 8.5 and 2M NaCl, and high thermostability was showed at 55°C and 3M NaCl. AmyA activity was enhanced by Triton X-100 and was not influenced by n-hexane and chloroform. Starch hydrolysis produced different oligomers with maltose as the smallest end-product. The efficiency of AmyA in degrading starch contained in agronomic residues was tested in grape cane chosen as model substrate. Preliminary results showed that starch was degraded making the enzyme a potential candidate for utilization of agro-industrial waste in fuel and chemicals production. AmyA is one of the few investigated amylases produced by haloarchaea, and the first alpha-amylase described among microorganisms belonging to the genus Haloterrigena. Copyright © 2016 Elsevier B.V. All rights reserved.

Sociodemographic Risk, Parenting, and Effortful Control: Relations to Salivary Alpha-amylase and Cortisol in Early Childhood

PubMed Central

Taylor, Zoe E.; Spinrad, Tracy L.; VanSchyndel, Sarah K.; Eisenberg, Nancy; Huynh, Jacqueline; Sulik, Michael J.; Granger, Douglas A.

2012-01-01

Early sociodemographic risk, parenting, and temperament were examined as predictors of the activity of children’s (N = 148; 81 boys, 67 girls) hypothalamic-pituitary-adrenal axis and autonomic nervous system. Demographic risk was assessed at 18 months (T1), intrusive-overcontrolling parenting and effortful control were assessed at 30 months (T2), and salivary cortisol and alpha-amylase were collected at 72 (T3) months of age. Demographic risk at T1 predicted lower levels of children’s effortful control and higher levels of mothers’ intrusive-overcontrolling parenting at T2. Intrusive-overcontrolling parenting at T2 predicted higher levels of children’s cortisol and alpha-amylase at T3, but effortful control did not uniquely predict children’s cortisol or alpha-amylase. Findings support the open nature of stress responsive physiological systems to influence by features of the early caregiving environment and underscore the utility of including measures of these systems in prevention trials designed to influence child outcomes by modifying parenting behavior. PMID:22949301

Permissive role of the acidification caused by wheat aleurone layers upon. alpha. -amylase induction by GA sub 3

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

Rodriguez-Campos, E.; Bernal-Lugo, I.; Hamabata, A.

1989-04-01

Wheat aleurone has the capacity of acidifying the incubation medium in 1 to 2 pH units. The {alpha}-amylase induction by GA{sub 3} in isolated wheat aleurone layers is strongly dependent on acidic pH of the medium (pH < 5). To examine possible mechanisms {sup 35}-Met incorporation into proteins and {alpha}-amylase, in the presence of GA{sub 3} and Ca{sup 2+} at pH, 4, 5 and 6 was studied. Although {sup 35}-Met uptake decreased markedly ({approx} 90%) at pH 4 in thepresence of GA{sub 3}, incorporation into total protein did not change significantly from other conditions. Auto-radiography of SDS-PAGE showed that mostmore » of the amino acid was in the {alpha}-amylase band, meaning that the effect of acidic pH is specific for GA{sub 3} actions on aleurone tissue. On the other hand, an increase of protonated GA{sub 3} diffusion could be ruled out. Also, there was not {alpha}-amylase inactivation at pH 6. These findings point out to the important physiological role of the acidification caused by the aleurone.« less

Analysis of the extreme diversity of salivary alpha-amylase isoforms generated by physiological proteolysis using liquid chromatography-tandem mass spectrometry.

PubMed

Bailey, Ulla-Maja; Punyadeera, Chamindie; Cooper-White, Justin J; Schulz, Benjamin L

2012-12-12

Saliva is a crucial biofluid for oral health and is also of increasing importance as a non-invasive source of disease biomarkers. Salivary alpha-amylase is an abundant protein in saliva, and changes in amylase expression have been previously associated with a variety of diseases and conditions. Salivary alpha-amylase is subject to a high diversity of post-translational modifications, including physiological proteolysis in the oral cavity. Here we developed methodology for rapid sample preparation and non-targeted LC-ESI-MS/MS analysis of saliva from healthy subjects and observed an extreme diversity of alpha-amylase proteolytic isoforms. Our results emphasize the importance of consideration of post-translational events such as proteolysis in proteomic studies, biomarker discovery and validation, particularly in saliva. Copyright © 2012 Elsevier B.V. All rights reserved.

Cloning, molecular characterization and heterologous expression of AMY1, an alpha-amylase gene from Cryptococcus flavus.

PubMed

Galdino, Alexsandro S; Ulhoa, Cirano J; Moraes, Lídia Maria P; Prates, Maura V; Bloch, Carlos; Torres, Fernando A G

2008-03-01

A Cryptococcus flavus gene (AMY1) encoding an extracellular alpha-amylase has been cloned. The nucleotide sequence of the cDNA revealed an ORF of 1896 bp encoding for a 631 amino acid polypeptide with high sequence identity with a homologous protein isolated from Cryptococcus sp. S-2. The presence of four conserved signature regions, (I) (144)DVVVNH(149), (II) (235)GLRIDSLQQ(243), (III) (263)GEVFN(267), (IV) (327)FLENQD(332), placed the enzyme in the GH13 alpha-amylase family. Furthermore, sequence comparison suggests that the C. flavusalpha-amylase has a C-terminal starch-binding domain characteristic of the CBM20 family. AMY1 was successfully expressed in Saccharomyces cerevisiae. The time course of amylase secretion in S. cerevisiae resulted in a maximal extracellular amylolytic activity (3.93 U mL(-1)) at 60 h of incubation. The recombinant protein had an apparent molecular mass similar to the native enzyme (c. 67 kDa), part of which was due to N-glycosylation.

A Proposed Mechanism for the Thermal Denaturation of a Recombinant Bacillus Halmapalus Alpha-amylase - the Effect of Calcium Ions

NASA Technical Reports Server (NTRS)

Nielsen, Anders D.; Pusey, Marc L.; Fuglsang, Claus C.; Westh, Peter

2003-01-01

The thermal stability of a recombinant alpha-amylase from Bacillus halmapalus alpha-amylase (BHA) has been investigated using circular dichroism spectroscopy (CD) and differential scanning calorimetry (DSC). This alpha-amylase is homologous to other Bacillus alpha-amylases where previous crystallographic studies have identified the existence of 3 calcium binding sites in the structure. Denaturation of BHA is irreversible with a Tm of approximately 89 C, and DSC thermograms can be described using a one-step irreversible model. A 5 C increase in T(sub m) in the presence of 10 fold excess CaCl2 was observed. However, a concomitant increase in the tendency to aggregate was also observed. The presence of 30-40 fold excess calcium chelator (EDTA or EGTA) results in a large destabilization of BHA corresponding to about 40 C lower T(sub m), as determined by both CD and DSC. Ten fold excess EGTA reveals complex DSC thermograms corresponding to both reversible and irreversible transitions, which possibly originate from different populations of BHA:calcium complexes. The observations in the present study have, in combination with structural information of homologous alpha-amylases, provided the basis for the proposal of a simple denaturation mechanism of BHA. The proposed mechanism describes the irreversible thermal denaturation of different BHA:calcium complexes and the calcium binding equilibrium involved. Furthermore, the model accounts for a temperature induced reversible structural change associated with calcium binding.

Determinants of wheat antigen and fungal alpha-amylase exposure in bakeries.

PubMed

Burstyn, I; Teschke, K; Bartlett, K; Kennedy, S M

1998-05-01

The study's objectives were to measure flour antigen exposure in bakeries and define the determinants of exposure. Ninety-six bakery workers, employed in seven different bakeries, participated in the study. Two side-by-side full-shift inhalable dust samples were obtained from each study participant on a single occasion. The flour antigen exposure was measured as wheat antigen and fungal alpha-amylase content of the water-soluble fraction of inhalable dust, assayed via enzyme-linked immunosorbent assays. During the entire sampling period bakers were observed and information on 14 different tasks was recorded at 15-minute intervals. Other production characteristics were also recorded for each sampling day and used in statistical modeling to identify significant predictors of exposure. The mean alpha-amylase antigen exposure was 22.0 ng/m3 (ranging from below the limit of detection of 0.1 ng/m3 to 307.1 ng/m3) and the mean wheat antigen exposure was 109 micrograms/m3 (ranging from below the limit of detection of 1 microgram/m3 to 1018 micrograms/m3). Regression models that explained 74% of variability in wheat antigen and alpha-amylase antigen exposures were constructed. The models indicated that tasks such as weighing, pouring, and operating dough-brakers increased flour antigen exposure, while packing and decorating resulted in lower exposures. Croissant, puff-pastry, and bread/bun production lines were associated with increased exposure, while cake production and substitution of dusting with the use of divider oil were associated with decreased exposure. Exposure levels can be reduced by the automation of forming tasks, alteration of tasks requiring pouring of flour, and changes to the types of products manufactured.

Taxonomy of the Streptomyces strain ZG0656 that produces acarviostatin alpha-amylase inhibitors and analysis of their effects on blood glucose levels in mammalian systems.

PubMed

Geng, P; Bai, G; Shi, Q; Zhang, L; Gao, Z; Zhang, Q

2009-02-01

To clarify the taxonomic status of strain ZG0656 and analyse the effects of its acarviostatin products on blood glucose levels in mammalian systems. Our program to screen for new alpha-amylase inhibitors led to the isolation of strain ZG0656. The polyphasic taxonomic study revealed that strain ZG0656 represents a novel variation of Streptomyces coelicoflavus, for which we propose the name S. coelicoflavus var. nankaiensis. Four chemically distinct alpha-amylase inhibitors, acarviostatins I03, II03, III03 and IV03, were isolated from strain ZG0656. Acarviostatins III03 and IV03 are both novel oligomers. All four acarviostatins are mixed noncompetitive porcine pancreas alpha-amylase inhibitors. Acarviostatin III03 is the most potent alpha-amylase inhibitor known to date. Moreover, in the in vitro and in vivo experiments, acarviostatins III03 showed significant inhibition of starch hydrolysis and glucose transfer to blood. Strain ZG0656 is a novel variation of S. coelicoflavus, whose products are novel effective alpha-amylase inhibitors. Among the products, acarviostatins III03 could significantly depress blood glucose levels in mammalian systems and be developed towards a possible therapeutic agent for diabetes. Acarviostatin III03 is the most potent alpha-amylase inhibitor known to date. The oligomer will benefit the research on the relationship between alpha-amylase and various inhibitors and will offer more choices in diabetes treatments.

Quantitative digital image analysis of chromogenic assays for high throughput screening of alpha-amylase mutant libraries.

PubMed

Shankar, Manoharan; Priyadharshini, Ramachandran; Gunasekaran, Paramasamy

2009-08-01

An image analysis-based method for high throughput screening of an alpha-amylase mutant library using chromogenic assays was developed. Assays were performed in microplates and high resolution images of the assay plates were read using the Virtual Microplate Reader (VMR) script to quantify the concentration of the chromogen. This method is fast and sensitive in quantifying 0.025-0.3 mg starch/ml as well as 0.05-0.75 mg glucose/ml. It was also an effective screening method for improved alpha-amylase activity with a coefficient of variance of 18%.

[Semicontinuous cultivation of fungi of the genus Aspergillus, producers of hydrolases].

PubMed

Blieva, R K

1982-01-01

The production of exohydrolases (alpha-amylase and pectinase) by fungi belonging to the genus Aspergillus was studied in the course of batch cultivation and, if immobilized cells were used, in the semicontinuous regime of growth. The cells were immobilized on a fixed filtering plate and on floating, in the growth medium, polyhedrons. Such a cultivation of immobilized microbial cells in the semicontinuous regime of growth on submerged polyhedrons freely floating in the nutrient medium makes it possible to cultivate the cells for 1.5 months with the active production of exocellular hydrolases. Under these conditions, Aspergillus oryzae 3-9-15 produces more alpha-amylase and A. awamori synthesizes more pectinases.

Alpha-Amylase Inhibition and Antioxidative Capacity of Some Antidiabetic Plants Used by the Traditional Healers in Southeastern Nigeria

PubMed Central

Oyedemi, Blessing O.; Ijeh, Ifeoma I.; Ohanyerem, Princemartins E.; Aiyegoro, Olayinka A.

2017-01-01

Oxidative stress plays a significant role in the pathogenesis of metabolic syndrome including diabetes mellitus (DM). The inhibition of alpha-amylase is an important therapeutic target in the regulation of postprandial increase of blood glucose in diabetic patients. The present study investigated the alpha-amylase inhibitory and antioxidant potential of selected herbal drugs used in the treatment of DM by the traditional healers in Isiala Mbano and Ikwuano regions of southeastern Nigeria. Antioxidant activity was evaluated in terms of free radical scavenging, reducing power, and total phenolic (TPC) and flavonoid content (TFC) in consonance with the TLC profiling. The results showed that methanol crude extracts from Anacardium occidentale (AO) and Ceiba pentandra (CP) recorded higher TPC and TFC, potent free radical scavenging, and efficient reducing power (RP) as compared with other plant samples. All the plant extracts exhibited a relative alpha-amylase inhibition apart from Strophanthus hispidus (SH) extract with a negative effect. We discovered a mild to weak correlation between alpha-amylase inhibition or antioxidative capacity and the total phenol or flavonoid content. At least in part, the results obtained in this work support the traditional use of certain plant species in the treatment of patients with DM. PMID:28367491

Xylanase supplementation of a wheat-based diet improved nutrient digestion and mRNA expression of intestinal nutrient transporters in broiler chickens infected with Clostridium perfringens.

PubMed

Guo, Shuangshuang; Liu, Dan; Zhao, Xu; Li, Changwu; Guo, Yuming

2014-01-01

Necrotic enteritis caused by Clostridium perfringens has become prevalent in the European Union due to the withdrawal of antibiotics in poultry feed. In an experiment with a 2 × 2 factorial arrangement, 336 one-day-old male broiler chicks (Ross 308) were assigned to 4 groups with or without C. perfringens challenge and fed wheat-based diets supplemented with or without xylanase at 5,500 U/kg of diet. The study aimed to investigate effects of xylanase addition on growth performance as well as nutrient digestion and absorption of C. perfringens-infected broilers. Before challenge (d 0-14), xylanase-supplemented birds had greater ADG and lower feed conversion ratio (FCR; P < 0.05). During infection (d 14-21), challenge tended to decrease ADG (P = 0.063) and significantly increased FCR (P < 0.05), whereas xylanase addition greatly reduced FCR (P < 0.05). Clostridium perfringens infection decreased AME values and apparent ileal digestibility of DM of diets (P < 0.05). Xylanase supplementation increased AME values regardless of infection and apparent ileal digestibility of CP in challenged birds (P < 0.05). Activities of duodenal α-amylase and chymotrypsin and pancreatic trypsin were decreased by C. perfringens infection (P < 0.05). Xylanase supplementation elevated pancreatic chymotrypsin activity and reduced duodenal α-amylase and trypsin activities (P < 0.05). It also decreased jejunal α-amylase activity and increased pancreatic α-amylase as well as jejunal sucrase activities in uninfected birds (P < 0.05). The duodenal mRNA expression of sodium glucose cotransporter 1 (SGLT1), H(+)-dependent peptide transporter 1 (PepT1), and liver fatty acid-binding protein (L-FABP) were downregulated (P < 0.05), but ileal SGLT1 gene expression was increased by infection (P < 0.05). Xylanase addition upregulated expression of jejunal SGLT1, PepT1, and L-FABP genes as well as ileal PepT1 and L-FABP genes in challenged broilers (P < 0.05). In conclusion, xylanase supplementation

Production of surfactant and detergent-stable, halophilic, and alkalitolerant alpha-amylase by a moderately halophilic Bacillus sp. Strain TSCVKK.

PubMed

Kiran, Kondepudi Kanthi; Chandra, T S

2008-01-01

A moderately halophilic alkalitolerant Bacillus sp. Strain TSCVKK, with an ability to produce extracellular halophilic, alkalitolerant, surfactant, and detergent-stable alpha-amylase was isolated from soil samples obtained from a salt-manufacturing industry in Chennai. The culture conditions for higher amylase production were optimized with respect to NaCl, substrate, pH, and temperature. Maximum amylase production of 592 mU/ml was achieved in the medium at 48 h with 10% NaCl, 1% dextrin, 0.4% yeast extract, 0.2% tryptone, and 0.2% CaCl(2) at pH 8.0 at 30 degrees C. The enzyme activity in the culture supernatant was highest with 10% NaCl at pH 7.5 and 55 degrees C. The amylase that was partially purified by acetone precipitation was highly stable in various surfactants and detergents. Glucose, maltose, and maltooligosaccharides were the main end products of starch hydrolysis indicating that it is an alpha-amylase.

[Activity of alpha-amylase and concentration of protein in saliva of pregnant women].

PubMed

Ciejak, Magdalena; Olszewska, Maria; Jakubowska, Katarzyna; Zebiełowicz, Dariusz; Safranow, Krzysztof; Chlubek, Dariusz

2007-01-01

One of the hypothetical reasons of the increased incidence of caries in women during the pregnancy may be the increased activity of alpha-amylase, which can be found in their saliva. The enzyme takes part in the process of decomposition of simple sugars, which make basic substrate for caries-causing bacteria. The aim of the paper was the evaluation of the influence of pregnancy and gestational age on the activity of alpha-amylase and the concentration of protein in women's saliva. The examined group consisted of 64 pregnant women at age 17-39, between 21st and 40th week of pregnancy. The control group consisted of 44 healthy women at age 20-35, who were not pregnant. In saliva, which was taken before morning meal, without stimulation, protein concentration was determined by Bradford method and the activity of amylase was determined by kinetic method. The activity of amylase correlated strongly and positively with protein concentration in saliva of both the pregnant (RS = +0.65; p < 0.00001) and the control group (RS = +0.74; p < 0.00001) women. There were no significant differences between examined parameters in the examined and the control group. It has been observed in the examined group, that there is the significant negative correlation between protein concentration in saliva and the week of pregnancy (RS = -0.35; p <0.01). It has been observed, in conducted researches, that there is no relation between the activity of amylase and the pregnancy and gestational age, which proves against the essential role of this enzyme in the increased caries incidence of pregnant women. However, the observed changes of total protein concentration in saliva during pregnancy, suggest that the exact cognition of proteins in pregnant women's saliva may reveal new mechanisms, which lead to an increase of caries risk.

A novel raw starch digesting alpha-amylase from a newly isolated Bacillus sp. YX-1: purification and characterization.

PubMed

Liu, Xu Dong; Xu, Yan

2008-07-01

This study reports the purification and characterization of a novel raw starch digesting alpha-amylase from a newly isolated Bacillus sp. YX-1. Maximum alpha-amylase activity (53 U mL(-1)) was obtained at 45 degrees C after 44 h of incubation. The enzyme was purified using ammonium sulfate precipitation, ion exchange and gel filtration chromatography, and showed a molecular weight of 56 kDa by SDS-PAGE. This enzyme exhibited maximum activity at pH 5.0, performed stability over a broad range of pH 4.5-11.0, and was optimally active at 40-50 degrees C. The enzyme preparation had a strong digesting ability towards various raw starches and efficiently hydrolyzed raw corn starch at a concentration of 20% and pH 5.0, which were normally used in the starch industries, in a period of 12h. By analyzing its partial amino acid sequences, the enzyme was proposed to be a novel alpha-amylase.

Cortisol, salivary alpha-amylase and children's perceptions of their social networks.

PubMed

Ponzi, Davide; Muehlenbein, Michael P; Geary, David C; Flinn, Mark V

2016-01-01

In recent years there has been a growing interest in the use of social network analysis in biobehavioral research. Despite the well-established importance of social relationships in influencing human behavior and health, little is known about how children's perception of their immediate social relationships correlates with biological parameters of stress. In this study we explore the association between two measures of children's personal social networks, perceived network size and perceived network density, with two biomarkers of stress, cortisol and salivary alpha-amylase. Forty children (mean age = 8.30, min age = 5, and max age = 12) were interviewed to collect information about their friendships and three samples of saliva were collected. Our results show that children characterized by a lower pre-interview cortisol concentration and a lower salivary alpha-amylase reactivity to the interview reported the highest density of friendships. We discuss this result in light of the multisystem approach to the study of children's behavioral outcomes, emphasizing that future work of this kind is needed in order to understand the cognitive and biological mechanisms underlying children's and adolescents' social perceptual biases.

Potential of the bean alpha-amylase inhibitor alpha-AI-1 to inhibit alpha-amylase activity in true bugs(Hemiptera)

USDA-ARS?s Scientific Manuscript database

True bugs (Hemiptera) are an important pest complex not controlled by Bt crops. An alternative source of resistance includes inhibitors of digestive enzymes. aAI-1, an a-amylase inhibitor from the common bean, has been shown to inhibit a-amylases of bruchid pests of grain legumes. Here we quantify t...

Amelioration in secretion of hyperthermostable and Ca2+ -independent alpha-amylase of Geobacillus thermoleovorans by some polyamines and their biosynthesis inhibitor methylglyoxal-bis-guanylhydrazone.

PubMed

Uma Maheswar Rao, J L; Satyanarayana, T

2004-01-01

Effect of polyamines and their biosynthesis inhibitors on the production of hyperthermostable and Ca2+ -independent alpha-amylase by Geobacillus thermoleovorans MTCC 4220. The alpha-amylase was produced in starch-yeast extract-tryptone (SYT) broth with different polyamines (PA) and polyamine biosynthesis inhibitors, methylglyoxal-bis-guanylhydrazone (MGBG) and cyclohexylammonium sulphate (CHA) at 70 degrees C. The bacterial pellets were obtained after growing G. thermoleovorans at different temperatures, and used in determining total PA. The cell-free culture filtrates were used in alpha-amylase assays. During growth, total polyamines in biomass increased till 2 h, and thereafter, decreased gradually. The total polyamine content was very high in the biomass cultivated at 55 degrees C when compared with that of higher temperatures. Enzyme titre enhanced up to 70 degrees C, and thereafter declined. Extracellular enzyme and protein levels declined in the presence of exogenously added PA. The intracellular enzyme titres, however, were higher in putrescine (put) and spermidine (spd) than in spermine (spm). Polyamine biosynthesis inhibitor, MGBG enhanced secretion of alpha-amylase in a laboratory fermentor as well as shake flasks, although CHA did not affect it. The intracellular accumulation of put in the presence of MGBG appeared to enhance synthesis and secretion of alpha-amylase. Extracellular enzyme and protein levels were low in the presence of exogenously added PA, but their intracellular levels, however, were higher in put and spd than in spm. A substantial increase in the synthesis and secretion of alpha-amylase was attained in G. thermoleovorans in the presence of polyamine biosynthesis inhibitor MGBG.

The spectrum of low molecular weight alpha-amylase/protease inhibitor genes expressed in the US bread wheat cultivar Butte 86

PubMed Central

2011-01-01

Background Wheat grains accumulate a variety of low molecular weight proteins that are inhibitors of alpha-amylases and proteases and play an important protective role in the grain. These proteins have more balanced amino acid compositions than the major wheat gluten proteins and contribute important reserves for both seedling growth and human nutrition. The alpha-amylase/protease inhibitors also are of interest because they cause IgE-mediated occupational and food allergies and thereby impact human health. Results The complement of genes encoding alpha-amylase/protease inhibitors expressed in the US bread wheat Butte 86 was characterized by analysis of expressed sequence tags (ESTs). Coding sequences for 19 distinct proteins were identified. These included two monomeric (WMAI), four dimeric (WDAI), and six tetrameric (WTAI) inhibitors of exogenous alpha-amylases, two inhibitors of endogenous alpha-amylases (WASI), four putative trypsin inhibitors (CMx and WTI), and one putative chymotrypsin inhibitor (WCI). A number of the encoded proteins were identical or very similar to proteins in the NCBI database. Sequences not reported previously included variants of WTAI-CM3, three CMx inhibitors and WTI. Within the WDAI group, two different genes encoded the same mature protein. Based on numbers of ESTs, transcripts for WTAI-CM3 Bu-1, WMAI Bu-1 and WTAI-CM16 Bu-1 were most abundant in Butte 86 developing grain. Coding sequences for 16 of the inhibitors were unequivocally associated with specific proteins identified by tandem mass spectrometry (MS/MS) in a previous proteomic analysis of milled white flour from Butte 86. Proteins corresponding to WDAI Bu-1/Bu-2, WMAI Bu-1 and the WTAI subunits CM2 Bu-1, CM3 Bu-1 and CM16 Bu-1 were accumulated to the highest levels in flour. Conclusions Information on the spectrum of alpha-amylase/protease inhibitor genes and proteins expressed in a single wheat cultivar is central to understanding the importance of these proteins in both

Rubusuaviins A-F, monomeric and oligomeric ellagitannins from Chinese sweet tea and their alpha-amylase inhibitory activity.

PubMed

Li, Haizhou; Tanaka, Takashi; Zhang, Ying-Jun; Yang, Chong-Ren; Kouno, Isao

2007-09-01

Six new ellagitannins herein, rubusuaviins A-F, were isolated from the aqueous acetone extract of Chinese sweet tea (Tien-cha, dried leaves of Rubus suavissimus S. LEE) together with seven known tannins. Rubusuaviin A was characterized as 1-O-galloyl-2,3-O-(S)-HHDP-4,6-O-(S)-sanguisorboyl-beta-D-glucopyranose. Rubusuaviins B, C, and E are dimeric, trimeric, and tetrameric ellagitannins, respectively, in which the sanguisorboyl groups were connected ellagitannin units. Rubusuaviins D and F were desgalloyl derivatives of rubusuaviins C and E, respectively. The inhibition of alpha-amylase activity by rubusuaviins and related ellagitannins was compared. Ellagitannins with beta-galloyl groups at the glucose C-1 positions showed stronger inhibition compared with the alpha-galloyl and desgalloyl compounds. The molecular weight of these compounds was not important for the inhibition of alpha-amylase activity.

[The primary structure of the alpha-amylase inhibitor Hoe 467A from Streptomyces tendae 4158. A new class of inhibitors].

PubMed

Aschauer, H; Vértesy, L; Nesemann, G; Braunitzer, G

1983-10-01

The native or modified alpha-amylase inhibitor Hoe 467A - isolated from the culture medium of Streptomyces tendae 4158 - and overlapping peptides were degraded by the automatic Edman technique. The oxidized or aminoethylated or oxidized and maleoylated inhibitor was digested with trypsin and the native inhibitor with pepsin. Further digestion with Staphylococcus aureus proteinase was also carried out. After peptic digestion two cystin peptides were isolated, which allowed the establishment of the disulfide bonds. The alpha-amylase inhibitor is a polypeptid consisting of 74 amino-acid residues with a molecular mass of 7958 Da. The inhibitor is composed of all naturally occurring amino acids except methionine and phenylalanine and shows no sequence homology to known inhibitors. The clinical and pharmacological importance in respect to the inhibitors ability for inactivation of human salivary and pancreatic alpha-amylase is discussed. Especially the proteinase resistance of the inhibitor enables a clinical application in human (e.g. Diabetes mellitus) per os.

Development of a solid-state fermentation process for production of an alpha amylase with potentially interesting properties.

PubMed

Hashemi, Maryam; Razavi, Seyed Hadi; Shojaosadati, Seyed Abbas; Mousavi, Seyyed Mohammad; Khajeh, Khosro; Safari, Mohammad

2010-09-01

Ca-independency with potential activity and stability at low pH are among the most interesting characteristics of alpha-amylase in starch industry. In this attempt the synergetic effect of low pH on activity of crude Ca-independent alpha-amylase isolated from a native Bacillus sp. KR-8104 in solid-state fermentation (SSF) was studied using wheat bran (WB) as a substrate. The effects of different parameters including moisturizing agents, solid substrate to moisture ratio, particle size, incubation temperature and period, inoculum (v/w) and supplementation with 1% (w/w) different carbon and nitrogen sources on enzyme production were investigated. Maximum enzyme production of 140U/g dry fermented substrate was obtained from wheat bran moistened with tap water at a ratio of 1:1.5 and supplemented with 1% (w/w) NH(4)NO(3) and 1% (w/w) lactose after 48h incubation at 37 degrees C. Even though the production of alpha-amylase was lower at 40 and 45 degrees C, the viable cell count was higher. In addition response surface methodology (RSM) was applied to find optimum conditions of temperature and pH on crude amylase activity. Using central composite design (CCD) a quadratic mathematical model equation was derived for the prediction of enzyme activity. The results showed that the model was in good agreement with experimental results, with R(2)=0.90 (p<0.0001) and the low pH has a synergetic effect on enzyme activity at higher temperature. Copyright 2010 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Purification and characterization of alpha-amylase from Bacillus licheniformis CUMC305

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

Krishnan, T.; Chandra, A.K.

Alpha-amylase produced by Bacillus licheniformis CUMC305 was purified 212-fold with a 42% yield through a series of four steps. The purified enzyme was homogeneous as shown by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and discontinuous gel electrophoresis. The purified enzyme showed maximal activity at 90 degrees C and pH 9.0, and 91% of this activity remained at 100 degrees C. In the presence of substrate (soluble starch), the alpha-amylase enzyme was fully stable after a 4-hour incubation at 100 degrees C. The enzyme showed 100% stability in the pH range 7 to 9; 95% stability at pH 10; and 84, 74,more » 68, and 50% stability at pH values of 6, 5, 4, and 3, respectively, after 18 hours of treatment. The activation energy for this enzyme was calculated as 5.1 x 10 to the power of 5 J/mol. The molecular weight was estimated to be 28,000 by sodium dodecyl sulfate-gel electrophoresis. The relative rates of hydrolysis of soluble starch, amylose, amylopectin, and glycogen were 1.27, 1.8, 1.94, and 2.28 mg/ml, respectively. Vmax values for hydrolysis of these substrates were calculated as 0.738, 1.08, 0.8, and 0.5 mg of maltose/ml per min, respectively. Of the cations, Na+, Ca(2+), and Mg(2+), showed stimulatory effect, wheras Hg(2+), Cu(2+), Ni(2+), Zn(2+), Ag+, Fe(2+), Co(2+), Cd(2+), Al(3+), and Mn(2+) were inhibitory. Of the anions, azide, F-, SO/sub 3/(2-), SO/sub 4/(3-), S/sub 2/O/sub 3/(2-), MoO/sub 4/(2-), and Wo/sub 4/(2-) showed an excitant effect. p-Chloromercuribenzoic acid and sodium iodoacetate were inhibitory, whereas cysteine, reduced glutathione, thiourea, beta-mercaptoethanol, and sodium glycerophosphate afforded protection to enzyme activity. Alpha-amylase was fairly resistant to EDTA treatment at 30 degrees C, but heating at 90 degrees C in presence of EDTA resulted in the complete loss of enzyme activity. (Refs. 32).« less

Daily Social Interactions and the Biological Stress Response: Are There Age Differences in Links Between Social Interactions and Alpha-Amylase?

PubMed

Birditt, Kira S; Tighe, Lauren A; Nevitt, Michael R; Zarit, Steven H

2017-12-12

According to the strength and vulnerability integration (SAVI) model, older people are better able to avoid negative social interactions than younger people, but when they do experience negative interactions, they are equally or more emotionally and physiologically reactive than younger people. Less is known about the links between daily negative and positive social encounters and the sympathetic adrenal medullary system (a key stress pathway) and whether there are age differences in these links. This study considers whether negative and positive social interactions are associated with diurnal alpha-amylase (a measure of the sympathetic adrenal medullary system) and whether there are differences in these links by age. Participants were from the Daily Health, Stress, and Relationship Study, which includes a random sample of 89 individuals (aged 40-95) who completed 14 days of daily diary interviews and provided saliva samples four times a day (wake, 30 min after wake, lunch, and bedtime) for four of those days that were assayed for alpha-amylase. Days in which people reported more negative interactions were associated with flatter morning declines in alpha-amylase, indicating greater stress. Links between positive interactions and diurnal alpha-amylase varied by age group. Findings are consistent with the SAVI model indicating that older adults respond differently to social stimuli than younger people. © The Author(s) 2017. Published by Oxford University Press on behalf of The Gerontological Society of America. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Purification and characterization of an alpha-amylase of Pichia burtonii isolated from the traditional starter "murcha" in Nepal.

PubMed

Takeuchi, Akiko; Shimizu-Ibuka, Akiko; Nishiyama, Yoshitaka; Mura, Kiyoshi; Okada, Sanae; Tokue, Chiyoko; Arai, Soichi

2006-12-01

Among more than 20 yeast strains isolated from the traditional starter "murcha" in Nepal, we characterized a yeast that might be involved in saccharification. This strain, identified as Pichia burtonii, produced an extracellular amylolytic enzyme when cultured in the presence of starch in the medium. Since no amylase secreted by P. burtonii has yet been reported, we purified the enzyme and determined its N-terminal amino acid sequence. Together with the results of a hydrolyzing activity assay toward various substrates, it was found to be an alpha-amylase. The purified enzyme, named Pichia burtonii alpha-amylase (PBA), was a glycoprotein with an apparent molecular mass of 51 kDa. Enzyme activity was optimal at pH 5.0 at 40 degrees C. The enzyme retained 80% of its original activity after incubation under the optimal pH condition at 50 degrees C for 30 min. The activity was inhibited by metal ions such as Cd(2+), Cu(2+), Hg(2+), Al(3+), and Zn(2+).

Production and characterization of a thermostable alpha-amylase from Nocardiopsis sp. endophyte of yam bean.

PubMed

Stamford, T L; Stamford, N P; Coelho, L C; Araújo, J M

2001-01-01

Thermostable amylolytic enzymes have been currently investigated to improve industrial processes of starch degradation. Studies on production of alpha-amylase by Nocardiopsis sp., an endophytic actinomycete isolated from yam bean (Pachyrhizus erosus L. Urban), showed that higher enzyme levels were obtained at the end of the logarithmic growth phase after incubation for 72 h at pH 8.6. Maximum activity of alpha-amylase was obtained at pH 5.0 and 70 degrees C. The isolated enzyme exhibited thermostable properties as indicated by retention of 100% of residual activity at 70 degrees C, and 50% of residual activity at 90 degrees C for 10 min. Extracellular enzyme from Nocardiopsis sp. was purified by fractional precipitation with ammonium sulphate. After 60% saturation produced 1130 U mg-1 protein and yield was 28% with purification 2.7-fold. The enzyme produced by Nocardiopsis sp. has potential for industrial applications.

Peer Victimization and Aggression: Moderation by Individual Differences in Salivary Cortisol and Alpha-Amylase

ERIC Educational Resources Information Center

Rudolph, Karen D.; Troop-Gordon, Wendy; Granger, Douglas A.

2010-01-01

This research examined whether variations in salivary measures of the hypothalamic-pituitary-adrenal axis (cortisol) and autonomic nervous system (alpha amylase [sAA]) contribute to individual differences in the association between peer victimization and aggression. Children (N = 132; M age = 9.46 years, SD = 0.33) completed a measure of peer…

Enzyme activity screening of thermophilic bacteria isolated from Dusun Tua Hot Spring, Malaysia

NASA Astrophysics Data System (ADS)

Msarah, Marwan; Ibrahim, Izyanti; Aqma, Wan Syaidatul

2018-04-01

Thermophilic bacteria have biotechnological importance due to the availability of unique enzymes which are stable in extreme circumstances. The aim of this study includes to isolate thermophilic bacteria from hot spring and screen for important enzyme activities. Water samples from the Dusun Tua Hot Spring were collected and the physiochemical characterisation of water was measured. Eight thermophilic bacteria were isolated and determined to have at least three strong enzyme activity including protease, lipase, amylase, cellulase, pectinase and xylanase. The results showed that HuluC2 displayed all the enzyme activities and can be further studied.

Immobilization of alpha-amylase from Bacillus circulans GRS 313 on coconut fiber.

PubMed

Dey, Gargi; Nagpal, Varima; Banerjee, Rintu

2002-01-01

A simple and inexpensive method for immobilizing alpha-amylase from Bacillus circulans GRS 313 on coconut fiber was developed. The immobilization conditions for highest efficiency were optimized with respect to immobilization pH of 5.5, 30 degrees C, contact time of 4 h, and enzyme to support a ratio of 1:1 containing 0.12 mg/mL of protein. The catalytic properties of the immobilized enzyme were compared with that of the free enzyme. The activity of amylase adsorbed on coconut fiber was 38.7 U/g of fiber at its optimum pH of 5.7 and 48 degrees C, compared with the maximum activity of 40.2 U/mL of free enzyme at the optimum pH of 4.9 and 48 degrees C. The reutilization capacity of the immobilized enzyme was up to three cycles.

Evaluation of the alpha-amylase activity as an indicator of pasteurization efficiency and microbiological quality of liquid whole eggs.

PubMed

Silva, Guilherme Resende da; Menezes, Liliane Denize Miranda; Lanza, Isabela Pereira; Oliveira, Daniela Duarte de; Silva, Carla Aparecida; Klein, Roger Wilker Tavares; Assis, Débora Cristina Sampaio de; Cançado, Silvana de Vasconcelos

2017-09-01

In order to evaluate the efficiency of the pasteurization process in liquid whole eggs, an UV/visible spectrophotometric method was developed and validated for the assessment of alpha-amylase activity. Samples were collected from 30 lots of raw eggs (n = 30) and divided into three groups: one was reserved for analysis of the raw eggs, the second group was pasteurized at 61.1°C for 3.5 minutes (n = 30), and the third group was pasteurized at 64.4°C for 2.5 minutes (n = 30). In addition to assessing alpha-amylase activity, the microbiological quality of the samples was also evaluated by counting total and thermotolerant coliforms, mesophilic aerobic microorganisms, Staphylococcus spp., and Salmonella spp. The validated spectrophotometric method demonstrated linearity, with a coefficient of determination (R2) greater than 0.99, limits of detection (LOD) and quantification (LOQ) of 0.48 mg kg-1 and 1.16 mg kg-1, respectively, and acceptable precision and accuracy with relative standard deviation (RSD) values of less than 10% and recovery rates between 98.81% and 105.40%. The results for alpha-amylase activity in the raw egg samples showed high enzyme activity due to near-complete hydrolysis of the starch, while in the eggs pasteurized at 61.1°C, partial inactivation of the enzyme was observed. In the samples of whole eggs pasteurized at 64.4°C, starch hydrolysis did not occur due to enzyme inactivation. The results of the microbiological analyses showed a decrease (P < 0.0001) in the counts for all the studied microorganisms and in the frequency of Salmonella spp. in the pasteurized egg samples according to the two binomials under investigation, compared to the raw egg samples, which showed high rates of contamination (P < 0.0001). After pasteurization, only one sample (3.33%) was positive for Salmonella spp., indicating failure in the pasteurization process, which was confirmed by the alpha-amylase test. It was concluded that the validated methodology for

Symptoms of prenatal depression are associated with raised salivary alpha-amylase levels.

PubMed

Braithwaite, Elizabeth C; Ramchandani, Paul G; Lane, Tracy A; Murphy, Susannah E

2015-10-01

Prenatal depression increases risk for a number of adverse offspring outcomes, however the biological mechanisms underlying this association remain unclear. It has been suggested that maternal glucocorticoids may mediate this link, though supporting evidence has been mixed. An alternative mechanism of effect may be via depression-induced changes in maternal sympathetic nervous system (SNS) function. We examined this hypothesis by determining the relationship between symptoms of maternal prenatal depression and diurnal salivary alpha-amylase (sAA) levels. 76 pregnant women were recruited during either the second or third trimester of pregnancy. Participants self-reported depressive symptoms using the Edinburgh postnatal depression scale. Saliva samples, to be assayed for alpha-amylase activity, were collected at home over two working days. Participants with depressive symptoms in later pregnancy had elevated awakening sAA levels compared with non-depressed controls (t(73) = -2.737, p = 0.008), and continued to have raised sAA throughout the day (F(1) = 10.924, p = 0.002). Our findings highlight that symptoms of depression during late pregnancy are associated with increased maternal SNS activity. Thus, changes in maternal SNS function, which may include increased vasoconstriction and reduced foetal blood flow, could, in part, mediate associations between prenatal depression and adverse offspring outcomes. Copyright © 2015 Elsevier Ltd. All rights reserved.

Assessing Agreement Between Salivary Alpha Amylase Levels Collected by Passive Drool and Eluted Filter Paper in Adolescents With Cancer

PubMed Central

Ameringer, Suzanne; Munro, Cindy; Elswick, R.K.

2014-01-01

Purpose/Objectives To assess the validity of filter paper (FP) against the gold standard of passive drool (PD) for collecting salivary alpha amylase as a surrogate biomarker of psychological stress in adolescents with cancer. Design Part of a longitudinal, descriptive study of symptoms in adolescents with cancer during chemotherapy. Setting A pediatric hematology/oncology treatment center. Sample 33 saliva sample pairs from nine adolescents with cancer, aged 13–18 years. Methods Salivary alpha amylase was collected by PD and FP at four time points during a cycle of chemotherapy: days 1 (time 1) and 2 (time 2) of chemotherapy, day 7–10 (time 3), and day 1 of the next cycle (time 4). A random effects regression was used to assess the correlation between PD and FP values, and a Bland Altman analysis was conducted to assess agreement between the values. Main Research Variables Salivary alpha amylase. Findings The estimated correlation between PD and FP values was r = 0.91, p < 0.001. Regression results were also used to rescale FP values to the levels of the PD values because the FP values were on a different scale than the PD values. The Bland Altman analysis revealed that the agreement between the rescaled FP values and PD values was not satisfactory. Conclusions Eluted FP may not be a valid method for collecting salivary alpha amylase in adolescents with cancer. Implications for Nursing Psychological stress in adolescents with cancer may be linked to negative outcomes, such as greater symptom severity and post-traumatic stress disorder. Nurses need valid, efficient, biobehavioral measures to assess psychological stress in the clinical setting. PMID:22750901

The impact of single nucleotide polymorphism in monomeric alpha-amylase inhibitor genes from wild emmer wheat, primarily from Israel and Golan

PubMed Central

2010-01-01

Background Various enzyme inhibitors act on key insect gut digestive hydrolases, including alpha-amylases and proteinases. Alpha-amylase inhibitors have been widely investigated for their possible use in strengthening a plant's defense against insects that are highly dependent on starch as an energy source. We attempted to unravel the diversity of monomeric alpha-amylase inhibitor genes of Israeli and Golan Heights' wild emmer wheat with different ecological factors (e.g., geography, water, and temperature). Population methods that analyze the nature and frequency of allele diversity within a species and the codon analysis method (comparing patterns of synonymous and non-synonymous changes in protein coding sequences) were used to detect natural selection. Results Three hundred and forty-eight sequences encoding monomeric alpha-amylase inhibitors (WMAI) were obtained from 14 populations of wild emmer wheat. The frequency of SNPs in WMAI genes was 1 out of 16.3 bases, where 28 SNPs were detected in the coding sequence. The results of purifying and the positive selection hypothesis (p < 0.05) showed that the sequences of WMAI were contributed by both natural selection and co-evolution, which ensured conservation of protein function and inhibition against diverse insect amylases. The majority of amino acid substitutions occurred at the C-terminal (positive selection domain), which ensured the stability of WMAI. SNPs in this gene could be classified into several categories associated with water, temperature, and geographic factors, respectively. Conclusions Great diversity at the WMAI locus, both between and within populations, was detected in the populations of wild emmer wheat. It was revealed that WMAI were naturally selected for across populations by a ratio of dN/dS as expected. Ecological factors, singly or in combination, explained a significant proportion of the variations in the SNPs. A sharp genetic divergence over very short geographic distances compared to

Comparing Dental Stress in New Child Patients and Returning Patients Using Salivary Cortisol, Immunoglobulin-A and Alpha- Amylase.

PubMed

Alaki, Sumer M; Safi, Ayman; Ouda, Soliman; Nadhreen, Alaa

this study was aimed at comparing dental stress in children having their first dental visit to those returning for dental treatment using salivary biomarkers of stress including salivary cortisol (s-cortisol), Immunoglobulin-A (s-IgA) and alpha-amylase (s-α-amylase). Additionally, the study was aimed at monitoring the change in stress in new patients as they progressed from the waiting to the clinical areas. salivary samples were collected from 40 children who had not been to a dentist before and similar samples were collected from 40 children who were returning for completion of dental treatment. Salivary cortisol, s-IgA and s-α-amylase concentrations were obtained by Enzyme-linked Immunosorbent Assay (ELISA). salivary cortisol levels were higher for new patients at the waiting area compared to that at the dental chair (p=0.05). Salivary alpha-amylase significantly increased in new patients while being seated in the dental chair. Returning patients had higher s-α-amylase (p=0.001) and s-IgA (p=0.016) compared to new patients. Returning patients had the lowest level of s-cortisol when providers were faculty pediatric dentists than with students and interns (p=0.035). children coming in for their first dental visit may experience dental stress at the waiting area before being seated for dental examination. Returning children may experience higher levels of stress compared to new child patients possibly due to previous dental exposure.

Serum trypsin, alpha-amylase and lipase during bombesin stimulation in normal subjects and patients with pancreatic insufficiency.

PubMed

Hafkenscheid, J C; Hessels, M; Jansen, J B; Lamers, C B

1984-01-31

The effect of infusion of bombesin (60 pmol/kg 20 min) on pancreatic enzymes in serum was studied in 13 normal subjects and 12 patients with pancreatic insufficiency. In normal subjects administration of bombesin induced large increases in serum trypsin (p less than 0.01), while serum total alpha-amylase and pancreatic alpha-amylase did not change and serum lipase showed only a modest rise (0.01 less than p less than 0.05). Patients with pancreatic insufficiency had significantly lower serum concentrations of all enzymes studied (p less than 0.01) and in such patients bombesin did not change the concentrations of pancreatic enzymes in serum. It is concluded that determination of the serum trypsin response to bombesin may be of help in the diagnosis of pancreatic insufficiency.

The 2.0-A resolution structure of soybean beta-amylase complexed with alpha-cyclodextrin.

PubMed

Mikami, B; Hehre, E J; Sato, M; Katsube, Y; Hirose, M; Morita, Y; Sacchettini, J C

1993-07-13

New crystallographic findings are presented which offer a deeper understanding of the structure and functioning of beta-amylase, the first known exo-type starch-hydrolyzing enzyme. A refined three-dimensional structure of soybean beta-amylase, complexed with the inhibitor alpha-cyclodextrin, has been determined at 2.0-A resolution with a conventional R-value of 17.5%. The model contains 491 amino acid residues, 319 water molecules, 1 sulfate ion, and 1 alpha-cyclodextrin molecule. The protein consists of a core with an (alpha/beta)8 supersecondary structure, plus a smaller globular region formed by long loops (L3, L4, and L5) extending from beta-strands beta 3, beta 4, and beta 5. Between the two regions is a cleft that opens into a pocket whose floor contains the postulated catalytic center near the carboxyl group of Glu 186. The annular alpha-cyclodextrin binds in (and partly projects from) the cleft with its glucosyl O-2/O-3 face abutting the (alpha/beta)8 side and with its alpha-D(1 --> 4) glucosidic linkage progression running clockwise as viewed from that side. The ligand does not bind deeply enough to interact with the carboxyl group of Glu 186. Rather, it occupies most of the cleft entrance, strongly suggesting that alpha-cyclodextrin inhibits catalysis by blocking substrate access to the more deeply located reaction center. Of the various alpha-cyclodextrin interactions with protein residues in loops L4, L5, L6, and L7, most notable is the shallow inclusion complex formed with Leu 383 (in L7, on the core side of the cleft) through contacts of its methyl groups with the C-3 atoms of four of the ligand's D-glucopyranosyl residues. All six residues of the bound alpha-cyclodextrin are of 4C1 conformation and are joined by alpha-1,4 linkages with similar torsional angles to form a nearly symmetrical torus as reported for crystalline inclusion complexes with alpha-cyclodextrin. We envision a significant role for the methyl groups of Leu 383 at the cleft entrance

Stilbene Glucoside, a Putative Sleep Promoting Constituent from Polygonum multiflorum Affects Sleep Homeostasis by Affecting the Activities of Lactate Dehydrogenase and Salivary Alpha Amylase.

PubMed

Wei, Qian; Ta, Guang; He, Wenjing; Wang, Wei; Wu, Qiucheng

2017-01-01

Chinese herbal medicine (CHM) has been used for treating insomnia for centuries. The most used CHM for insomnia was Polygonum multiflorum. However, the molecular mechanism for CHM preventing insomnia is unknown. Stilbene glucoside (THSG), an important active component of P. multiflorum, may play an important role for treating insomnia. To test the hypothesis, Kunming mice were treated with different dosages of THSG. To examine the sleep duration, a computer-controlled sleep-wake detection system was implemented. Electroencephalogram (EEG) and electromyogram (EMG) electrodes were implanted to determine sleep-wake state. RT-PCR and Western blot was used to measure the levels of lactate dehydrogenase (LDH) and saliva alpha amylase. Spearman's rank correlation coefficient was used to identify the strength of correlation between the variables. The results showed that THSG significantly prolonged the sleep time of the mice (p<0.01). THSG changed sleep profile by reducing wake and rapid eye movement (REM) period, and increasing non-REM period. RT-PCR and Western blot analysis showed that THSG could down-regulate the levels of LDH and saliva alpha amylase (p<0.05). The level of lactate and glucose was positively related with the activity of LDH and saliva alpha amylase (p<0.05), respectively. On the other hand, the activities of LDH and amylase were negatively associated with sleep duration (p<0.05). The levels of lactate and glucose affect sleep homeostasis. Thus, THSG may prevent insomnia by regulating sleep duration via LDH and salivary alpha amylase.

Longitudinal and Immediate Effect of Kundalini Yoga on Salivary Levels of Cortisol and Activity of Alpha-Amylase and Its Effect on Perceived Stress.

PubMed

García-Sesnich, Jocelyn N; Flores, Mauricio Garrido; Ríos, Marcela Hernández; Aravena, Jorge Gamonal

2017-01-01

Stress is defined as an alteration of an organism's balance in response to a demand perceived from the environment. Diverse methods exist to evaluate physiological response. A noninvasive method is salivary measurement of cortisol and alpha-amylase. A growing body of evidence suggests that the regular practice of Yoga would be an effective treatment for stress. To determine the Kundalini Yoga (KY) effect, immediate and after 3 months of regular practice, on the perception of psychological stress and the salivary levels of cortisol and alpha-amylase activity. To determine the psychological perceived stress, levels of cortisol and alpha-amylase activity in saliva, and compare between the participants to KY classes performed for 3 months and a group that does not practice any type of yoga. The total sample consisted of 26 people between 18 and 45-year-old; 13 taking part in KY classes given at the Faculty of Dentistry, University of Chile and 13 controls. Salivary samples were collected, enzyme-linked immunosorbent assay was performed to quantify cortisol and kinetic reaction test was made to determine alpha-amylase activity. Perceived Stress Scale was applied at the beginning and at the end of the intervention. Statistical analysis was applied using Stata v11.1 software. Shapiro-Wilk test was used to determine data distribution. The paired analysis was fulfilled by t -test or Wilcoxon signed-rank test. T -test or Mann-Whitney's test was applied to compare longitudinal data. A statistical significance was considered when P < 0.05. KY practice had an immediate effect on salivary cortisol. The activity of alpha-amylase did not show significant changes. A significant decrease of perceived stress in the study group was found. KY practice shows an immediate effect on salivary cortisol levels and on perceived stress after 3 months of practice.

Cortisol and alpha-amylase as stress response indicators during pre-hospital emergency medicine training with repetitive high-fidelity simulation and scenarios with standardized patients.

PubMed

Valentin, Bernd; Grottke, Oliver; Skorning, Max; Bergrath, Sebastian; Fischermann, Harold; Rörtgen, Daniel; Mennig, Marie-Therese; Fitzner, Christina; Müller, Michael P; Kirschbaum, Clemens; Rossaint, Rolf; Beckers, Stefan K

2015-04-08

In emergency medicine, the benefits of high-fidelity simulation (SIM) are widely accepted and standardized patients (SP) are known to mimic real patients accurately. However, only limited data are available concerning physicians' stress markers within these training environments. The aim of this pilot study was to investigate repetitive stress among healthcare professionals in simulated pre-hospital emergency scenarios using either SIM or SPs. Teams with one emergency medical services (EMS) physician and two paramedics completed three SIM scenarios and two SP scenarios consecutively. To evaluate stress, salivary cortisol and alpha-amylase were measured in saliva samples taken before, during and after the scenarios. A total of 14 EMS physicians (29% female; mean age: 36.8 ± 5.0 years; mean duration of EMS-experience: 9.1 ± 5.8 years) and 27 paramedics (11% female; age: 30.9 ± 6.9 years; EMS experience: 8.1 ± 6.0 years) completed the study. Alpha-amylase and cortisol levels did not differ significantly between the two professions. Cortisol values showed a gradual and statistically significant reduction over time but little change was observed in response to each scenario. In contrast, alpha-amylase activity increased significantly in response to every SIM and SP scenario, but there was no clear trend towards an overall increase or decrease over time. Increases in salivary alpha-amylase activity suggest that both SIM and SP training produce stress among emergency healthcare professionals. Corresponding increases in salivary cortisol levels were not observed. Among physicians in the emergency setting, it appears that alpha-amylase provides a more sensitive measure of stress levels than cortisol.

Cognitive performance and morning levels of salivary cortisol and alpha-amylase in children reporting high vs. low daily stress perception.

PubMed

Maldonado, Enrique F; Fernandez, Francisco J; Trianes, M Victoria; Wesnes, Keith; Petrini, Orlando; Zangara, Andrea; Enguix, Alfredo; Ambrosetti, Lara

2008-05-01

The aim of the present study was to assess the effects of daily stress perception on cognitive performance and morning basal salivary cortisol and alpha-amylase levels in healthy children aged 9-12. Participants were classified by whether they had low daily perceived stress (LPS, n = 27) or a high daily perceived stress (HPS, n = 26) using the Children Daily Stress Inventory (CDSI). Salivary cortisol and alpha-amylase were measured at awakening and 30 minutes later. Cognitive performance was assessed using the Cognitive Drug Research assessment system. The HPS group exhibited significantly poorer scores on speed of memory (p < .05) and continuity of attention (p < .05) relative to the LPS group. The HPS group also showed significantly lower morning cortisol levels at awakening and at +30 minutes measures in comparison with the LPS group (p < .05), and mean morning cortisol levels were negatively correlated with speed of memory (p < .05) in the 53 participants. No significant differences were observed between both groups in alpha-amylase levels. These findings suggest that daily perceived stress in children may impoverish cognitive performance via its modulating effects on the HPA axis activity.

Adolescents' increasing stress response to social evaluation: pubertal effects on cortisol and alpha-amylase during public speaking.

PubMed

van den Bos, Esther; de Rooij, Mark; Miers, Anne C; Bokhorst, Caroline L; Westenberg, P Michiel

2014-01-01

Stress responses to social evaluation are thought to increase during adolescence, which may be due to pubertal maturation. However, empirical evidence is scarce. This study is the first to investigate the relation between pubertal development and biological responses to a social-evaluative stressor longitudinally. Participants performed the Leiden Public Speaking Task twice, with a 2-year interval (N = 217; age at Time 1: 8-17 years). The results support an increase in sensitivity to social evaluation during adolescence. The overall cortisol and alpha-amylase responses increased-both between and within participants-and were more strongly related to self-reported pubertal development than to age. The cortisol response shifted from speech delivery toward anticipation. The alpha-amylase response increased in both phases. © 2013 The Authors. Child Development © 2013 Society for Research in Child Development, Inc.

Bean alpha-amylase inhibitors in transgenic peas inhibit development of pea weevil larvae.

PubMed

de Sousa-Majer, Maria José; Hardie, Darryl C; Turner, Neil C; Higgins, Thomas J V

2007-08-01

This glasshouse study used an improved larval measurement procedure to evaluate the impact of transgenic pea, Pisum sativum L., seeds expressing a-amylase inhibitor (AI)-1 or -2 proteins on pea weevil, Bruchus pisorum L. Seeds of transgenic 'Laura' and 'Greenfeast' peas expressing alpha-(AI)-1 reduced pea weevil survival by 93-98%. Larval mortality occurred at an early instar. Conversely, in nontransgenic cultivars, approximately 98-99% of the pea weevils emerged as adults. By measuring the head capsule size, we determined that larvae died at the first to early third instar in alpha-(AI)-1 transgenic peas, indicating that this inhibitor is highly effective in controlling this insect. By contrast, transgenic Laura and 'Dundale' expressing alpha-(AI)-2 did not affect pea weevil survival, but they did delay larval development. After 77 d of development, the head capsule size indicated that the larvae were still at the third instar stage in transgenic alpha-(AI)-2 peas, whereas adult bruchids had developed in the nontransgenic peas.

The effects of autonomy support on salivary alpha-amylase: The role of individual differences.

PubMed

Sieber, Vanda; Schüler, Julia; Wegner, Mirko

2016-12-01

The empirical evidence for the relationship between autonomy-supportive environments and physiological stress is inconsistent. Whereas some studies report a decrease in stress in autonomy-supportive environments, other studies show a negative effect of autonomy on physiological stress. As previous research has not considered individual differences within this relationship, the present research aims to close this empirical gap by proposing that an implicit autonomy disposition, which is defined as a dispositional preference for self-determination, serves as a moderator. In an experiment, we tested whether the autonomy disposition moderates the effect of different teaching styles (controlling, autonomy-supportive, and neutral) on the acute physiological stress response (salivary alpha-amylase) in adolescents (N=69). The study revealed that participants with a high implicit autonomy disposition displayed lower salivary alpha-amylase responses when exposed to autonomy-supportive vignettes compared to when they were exposed to controlling or neutral teaching styles. The opposite pattern was found in students with a low implicit autonomy disposition. The results illustrate that experimentally induced variations in autonomy support lead to different physiological stress responses, depending on individual differences in the implicit autonomy disposition. Copyright © 2016 Elsevier Ltd. All rights reserved.

Combined Expression of Aspergillus nidulans Endoxylanase X24 and Aspergillus oryzae (alpha)-Amylase in Industrial Baker's Yeasts and Their Use in Bread Making.

PubMed

Monfort, A; Blasco, A; Prieto, J A; Sanz, P

1996-10-01

The Aspergillus nidulans endoxylanase X24 and the Aspergillus oryzae (alpha)-amylase cDNAs were placed under the control of the Saccharomyces cerevisiae actin promoter (pACT1) and introduced into baker's yeast. Bread made with transformants expressing both enzymes (YEpACT-AMY-ACT-X24) showed a 30% increase in volume and reduced firmness in comparison with that produced with a commercial strain. Endoxylanase X24 and (alpha)-amylase seem to act synergistically to improve the quality of bread in terms of volume and density.

Feeding value of field beans (Vicia faba L. var. minor) with and without enzyme containing tannase, pectinase and xylanase activities for broilers.

PubMed

Abdulla, Jalil Mahmwd; Rose, Stephen Paul; Mackenzie, Alexander Mackay; Pirgozliev, Vasil Radoslavov

2017-04-01

Effects of field beans with various tannin content and exogenous enzyme mixture containing tannase, pectinase and xylanase activities on N-corrected dietary apparent metabolisable energy (AMEn), coefficients of dry matter (DMR) and nitrogen retention (NR), fat digestibility, gastrointestinal tract (GIT) development, jejunal villus morphometry, ileal digesta viscosity and sialic acid were examined. Birds' growth performance and energy conversion ratio (ECR) were also measured. Birds were fed one of eight mash diets. The Control diet contained as major ingredients wheat (400 g/kg) and soybean meal (SBM) (127 g/kg and 221 g crude protein/kg and 12.83 MJ AMEn/kg. To reduce nutrient density, the Control diet also contained washed sand at 119 g/kg. Another three diets containing 300 g/kg of each of three experimental field bean cultivar samples in replacement for SBM and sand were also mixed. Each diet was fed to nine pens with two male Ross 308 broilers. Diets high in tannin had low AMEn, ECR, DMR and NR (p < 0.001). Feeding field beans increased (p < 0.001) the weights of the pancreas and the proventriculus and gizzard (PG) of the birds. Supplementing diets with the enzyme mixture improved (p < 0.001) feed conversion efficiency, AMEn and all nutrient utilisation coefficients despite the tannins in diets. The enzyme mixture reduced ileal digesta viscosity (p < 0.001) and the weight of pancreas, total GIT and PG (p < 0.05) of the birds. It can be concluded that the feeding value of field beans with different tannin contents may vary when fed to broilers. The supplementation of the enzyme mixture improved the feeding value of diets for broilers. The beneficial effect of the addition of the enzyme mixture seems to be mediated through reduced ileal digesta viscosity and improved nutrient availability.

Dynamic rheological properties of dough as affected by amylases from various sources.

PubMed

Doğan, Ismail S

2002-12-01

The effect of alpha-amylases from cereal, fungal, and bacterial sources on dough dynamic rheological properties was investigated. Dynamic rheological study of flour-and-water doughs during resting period showed significant changes in dough rheological properties as a function of alpha-amylases. Addition of alpha-amylases caused a time-dependent decrease in G', storage modulus. The enzyme action on starch during baking increased viscous flow properties. These changes were temperature-dependent. The thermal inactivation temperature of alpha-amylase plays an important role in modification of starch. Rheological changes in dough will alter the machinability of the dough and the quality of end products.

Combined Expression of Aspergillus nidulans Endoxylanase X24 and Aspergillus oryzae (alpha)-Amylase in Industrial Baker's Yeasts and Their Use in Bread Making

PubMed Central

Monfort, A.; Blasco, A.; Prieto, J. A.; Sanz, P.

1996-01-01

The Aspergillus nidulans endoxylanase X24 and the Aspergillus oryzae (alpha)-amylase cDNAs were placed under the control of the Saccharomyces cerevisiae actin promoter (pACT1) and introduced into baker's yeast. Bread made with transformants expressing both enzymes (YEpACT-AMY-ACT-X24) showed a 30% increase in volume and reduced firmness in comparison with that produced with a commercial strain. Endoxylanase X24 and (alpha)-amylase seem to act synergistically to improve the quality of bread in terms of volume and density. PMID:16535419

Application of a statistical design to the optimization of parameters and culture medium for alpha-amylase production by Aspergillus oryzae CBS 819.72 grown on gruel (wheat grinding by-product).

PubMed

Kammoun, Radhouane; Naili, Belgacem; Bejar, Samir

2008-09-01

The production optimization of alpha-amylase (E.C.3.2.1.1) from Aspergillus oryzae CBS 819.72 fungus, using a by-product of wheat grinding (gruel) as sole carbon source, was performed with statistical methodology based on three experimental designs. The optimisation of temperature, agitation and inoculum size was attempted using a Box-Behnken design under the response surface methodology. The screening of nineteen nutrients for their influence on alpha-amylase production was achieved using a Plackett-Burman design. KH(2)PO(4), urea, glycerol, (NH(4))(2)SO(4), CoCl(2), casein hydrolysate, soybean meal hydrolysate, MgSO(4) were selected based on their positive influence on enzyme formation. The optimized nutrients concentration was obtained using a Taguchi experimental design and the analysis of the data predicts a theoretical increase in the alpha-amylase expression of 73.2% (from 40.1 to 151.1 U/ml). These conditions were validated experimentally and revealed an enhanced alpha-amylase yield of 72.7%.

Potentized Mercuric chloride and Mercuric iodide enhance alpha-amylase activity in vitro.

PubMed

Sukul, N C; De, A; Sukul, A; Sinhababu, S P

2002-10-01

Mercuric chloride 30c and Mercuric iodide 30c were prepared by successive dilution in 30 steps of 1:100 followed by sonication at 20KHz for 30s at each step. Both were prepared in two media: 90% ethanol and distilled water. Three preparations of Mercuric chloride 30 in water were used: 12-month old, 1-month old and 4-day old. The controls for the water and ethanol-water preparations were pure water 30c and 90% ethanol 30c, respectively. For the three water preparations there were three matched controls of water 30c of the same ages. Each potentized substance or its control was mixed with distilled water 1:100 before testing. Hydrolysis of starch by alpha-amylase was measured by the standard procedure after incubation for 15 min at 27 degrees C. Mercuric chloride 30c and Mercuric iodide 30c in both water and aqueous ethanol media, enhanced enzyme activity significantly, compared to their respective controls. Mercuric chloride 30c, prepared in water 12 months previously, produced no significant change in the enzyme activity compared to its control. We hypothesize that the structure of the active molecule imprinted on water polymers during the process of dynamization. The specifically structured water interacts with the active sites of alpha-amylase, modifying its activity. Ethanol molecules have large non-polar part stabilizing the water structure and thus retaining activity for a longer time.

Longitudinal and Immediate Effect of Kundalini Yoga on Salivary Levels of Cortisol and Activity of Alpha-Amylase and Its Effect on Perceived Stress

PubMed Central

García-Sesnich, Jocelyn N; Flores, Mauricio Garrido; Ríos, Marcela Hernández; Aravena, Jorge Gamonal

2017-01-01

Context: Stress is defined as an alteration of an organism's balance in response to a demand perceived from the environment. Diverse methods exist to evaluate physiological response. A noninvasive method is salivary measurement of cortisol and alpha-amylase. A growing body of evidence suggests that the regular practice of Yoga would be an effective treatment for stress. Aims: To determine the Kundalini Yoga (KY) effect, immediate and after 3 months of regular practice, on the perception of psychological stress and the salivary levels of cortisol and alpha-amylase activity. Settings and Design: To determine the psychological perceived stress, levels of cortisol and alpha-amylase activity in saliva, and compare between the participants to KY classes performed for 3 months and a group that does not practice any type of yoga. Subjects and Methods: The total sample consisted of 26 people between 18 and 45-year-old; 13 taking part in KY classes given at the Faculty of Dentistry, University of Chile and 13 controls. Salivary samples were collected, enzyme-linked immunosorbent assay was performed to quantify cortisol and kinetic reaction test was made to determine alpha-amylase activity. Perceived Stress Scale was applied at the beginning and at the end of the intervention. Statistical Analysis Used: Statistical analysis was applied using Stata v11.1 software. Shapiro–Wilk test was used to determine data distribution. The paired analysis was fulfilled by t-test or Wilcoxon signed-rank test. T-test or Mann–Whitney's test was applied to compare longitudinal data. A statistical significance was considered when P < 0.05. Results: KY practice had an immediate effect on salivary cortisol. The activity of alpha-amylase did not show significant changes. A significant decrease of perceived stress in the study group was found. Conclusions: KY practice shows an immediate effect on salivary cortisol levels and on perceived stress after 3 months of practice. PMID:28546677

Three camelid VHH domains in complex with porcine pancreatic alpha-amylase. Inhibition and versatility of binding topology.

PubMed

Desmyter, Aline; Spinelli, Silvia; Payan, Francoise; Lauwereys, Marc; Wyns, Lode; Muyldermans, Serge; Cambillau, Christian

2002-06-28

Camelids produce functional antibodies devoid of light chains and CH1 domains. The antigen-binding fragment of such heavy chain antibodies is therefore comprised in one single domain, the camelid heavy chain antibody VH (VHH). Here we report on the structures of three dromedary VHH domains in complex with porcine pancreatic alpha-amylase. Two VHHs bound outside the catalytic site and did not inhibit or inhibited only partially the amylase activity. The third one, AMD9, interacted with the active site crevice and was a strong amylase inhibitor (K(i) = 10 nm). In contrast with complexes of other proteinaceous amylase inhibitors, amylase kept its native structure. The water-accessible surface areas of VHHs covered by amylase ranged between 850 and 1150 A(2), values similar to or even larger than those observed in the complexes between proteins and classical antibodies. These values could certainly be reached because a surprisingly high extent of framework residues are involved in the interactions of VHHs with amylase. The framework residues that participate in the antigen recognition represented 25-40% of the buried surface. The inhibitory interaction of AMD9 involved mainly its complementarity-determining region (CDR) 2 loop, whereas the CDR3 loop was small and certainly did not protrude as it does in cAb-Lys3, a VHH-inhibiting lysozyme. AMD9 inhibited amylase, although it was outside the direct reach of the catalytic residues; therefore it is to be expected that inhibiting VHHs might also be elicited against proteases. These results illustrate the versatility and efficiency of VHH domains as protein binders and enzyme inhibitors and are arguments in favor of their use as drugs against diabetes.

Cloning and Characterization of an alpha-amylase Gene from the Hyperthermophilic Archaeon Thermococcus Thioreducens

NASA Technical Reports Server (NTRS)

Bernhardsdotter, Eva C. M. J.; Pusey, Mark L.; Ng, Joseph D.; Garriott, Owen K.

2004-01-01

The gene encoding an extracellular alpha-amylase, TTA, from the hyperthermophilic archaeon Thermococcus thioreducens was cloned and expressed in Escherichia coli. Primary structural analysis revealed high similarity with other a-amylases from the Thermococcus and Pyrococcus genera, as well as the four highly conserved regions typical for a-amylases. The 1374 bp gene encodes a protein of 457 amino acids, of which 435 constitute the mature protein preceded by a 22 amino acid signal peptide. The molecular weight of the purified recombinant enzyme was estimated to be 43 kDa by denaturing gel electrophoresis. Maximal enzymatic activity of recombinant TTA was observed at 90 C and pH 5.5 in the absence of exogenous Ca(2+), and the enzyme was considerably stable even after incubation at 90 C for 2 hours. The thermostability at 90 and 102 C was enhanced in the presence of 5 mM Ca(2+). The extraordinarily high specific activity (about 7.4 x 10(exp 3) U/mg protein at 90 C, pH 5.5 with soluble starch as substrate) together with its low pH optimum makes this enzyme an interesting candidate for starch processing applications.

Norepinephrine transporter blocker atomoxetine increases salivary alpha amylase.

PubMed

Warren, Christopher M; van den Brink, Ruud L; Nieuwenhuis, Sander; Bosch, Jos A

2017-04-01

It has been suggested that central norepinephrine (NE) activity may be inferred from increases in salivary alpha-amylase (SAA), but data in favor of this proposition are limited. We administered 40mg of atomoxetine, a selective NE transporter blocker that increases central NE levels, to 24 healthy adult participants in a double-blind, placebo-controlled cross-over design. Atomoxetine administration significantly increased SAA secretion and concentrations at 75-180min after treatment (more than doubling baseline levels). Consistent with evidence that elevation in central NE is a co-determinant of hypothalamic-pituitary-adrenal axis activity, salivary cortisol also approximately doubled at the same time points. Moreover, changes in salivary cortisol positively correlated with SAA (0.44

Variability in Arabinoxylan, Xylanase activity and Xylanase inhibitor levels in hard spring wheat

USDA-ARS?s Scientific Manuscript database

Arabinoxylans (AX), xylanase, and xylanase inhibitors have an important role in many cereal food processing applications. The effect of genotype (G), growing location (L), and their interaction (G*L) on AX, apparent xylanase and apparent xylanase inhibition activities of Triticum aestivum xylanase i...

Diurnal patterns of salivary alpha-amylase and cortisol secretion in female adolescent tennis players after 16 weeks of training.

PubMed

Filaire, Edith; Ferreira, Jose Pedro; Oliveira, Miguel; Massart, Alain

2013-07-01

We examined the effects of 16 weeks of training on diurnal pattern of salivary alpha-amylase (sAA), cortisol, and the ratio of sAA over cortisol (AOC) in 12 national adolescent female tennis players. Stress and recovery were also evaluated using the Recovery-Stress-Questionnaire for Athletes-RESTQ-Sport. Data were collected after a 2-week rest (January, W0), and 4 months after W0 (W16). Subjects collected five saliva samples throughout a day. While all participants displayed the previously shown decrease after awakening in adolescents at W0, they showed a rise in the alpha-amylase awakening response and a higher alpha-amylase activity output (p<0.01) at W16 compared to W0. For the daily rhythm of cortisol we found subjects having a low overall output of salivary cortisol (p<0.01) and a blunted response to awakening at W16. Furthermore, an increase in the ratio AOC at W16, and a negative correlation between this ratio and Sport-specific recovery score. Our findings offer support for the hypothesis that increase of training load during the study period induced asymmetry activation between the two stress systems, in relation to psychological alterations and performance decrease. These results provide encouragement to continue exploring the impact of training program using a psychobiological approach among young athletes in order to prevent fatigue and preserve the health of these athletes. Copyright © 2012 Elsevier Ltd. All rights reserved.

SusG: A Unique Cell-Membrane-Associated [alpha]-Amylase from a Prominent Human Gut Symbiont Targets Complex Starch Molecules

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

Koropatkin, Nicole M.; Smith, Thomas J.

SusG is an {alpha}-amylase and part of a large protein complex on the outer surface of the bacterial cell and plays a major role in carbohydrate acquisition by the animal gut microbiota. Presented here, the atomic structure of SusG has an unusual extended, bilobed structure composed of amylase at one end and an unprecedented internal carbohydrate-binding motif at the other. Structural studies further demonstrate that the carbohydrate-binding motif binds maltooligosaccharide distal to, and on the opposite side of, the amylase catalytic site. SusG has an additional starch-binding site on the amylase domain immediately adjacent to the active cleft. Mutagenesis analysismore » demonstrates that these two additional starch-binding sites appear to play a role in catabolism of insoluble starch. However, elimination of these sites has only a limited effect, suggesting that they may have a more important role in product exchange with other Sus components.« less

Xylanases of marine fungi of potential use for biobleaching of paper pulp.

PubMed

Raghukumar, Chandralata; Muraleedharan, Usha; Gaud, V R; Mishra, R

2004-10-01

Microbial xylanases that are thermostable, active at alkaline pH and cellulase-free are generally preferred for biobleaching of paper pulp. We screened obligate and facultative marine fungi for xylanase activity with these desirable traits. Several fungal isolates obtained from marine habitats showed alkaline xylanase activity. The crude enzyme from NIOCC isolate 3 (Aspergillus niger), with high xylanase activity, cellulase-free and unique properties containing 580 U l(-1) xylanase, could bring about bleaching of sugarcane bagasse pulp by a 60 min treatment at 55 degrees C, resulting in a decrease of ten kappa numbers and a 30% reduction in consumption of chlorine during bleaching. The culture filtrate showed peaks of xylanase activity at pH 3.5 and pH 8.5. When assayed at pH 3.5, optimum activity was detected at 50 degrees C, with a second peak of activity at 90 degrees C. When assayed at pH 8.5, optimum activity was seen at 80 degrees C. The crude enzyme was thermostable at 55 degrees C for at least 4 h and retained about 60% activity. Gel filtration of the 50-80% ammonium sulphate-precipitated fraction of the crude culture filtrate separated into two peaks of xylanase with specific activities of 393 and 2,457 U (mg protein)(-1). The two peaks showing xylanase activity had molecular masses of 13 and 18 kDa. Zymogram analysis of xylanase of crude culture filtrate as well as the 50-80% ammonium sulphate-precipitated fraction showed two distinct xylanase activity bands on native PAGE. The crude culture filtrate also showed moderate activities of beta-xylosidase and alpha- l-arabinofuranosidase, which could act synergistically with xylanase in attacking xylan. This is the first report showing the potential application of crude culture filtrate of a marine fungal isolate possessing thermostable, cellulase-free alkaline xylanase activity in biobleaching of paper pulp.

Interaction between wheat alpha-amylase/trypsin bi-functional inhibitor and mammalian digestive enzymes: Kinetic, equilibrium and structural characterization of binding.

PubMed

Cuccioloni, Massimiliano; Mozzicafreddo, Matteo; Ali, Ishtiaq; Bonfili, Laura; Cecarini, Valentina; Eleuteri, Anna Maria; Angeletti, Mauro

2016-12-15

Alpha-amylase/trypsin bi-functional inhibitors (ATIs) are non-gluten protein components of wheat and other cereals that can hypersensitise the human gastrointestinal tract, eventually causing enteropathies in predisposed individuals. These inhibitory proteins can act both directly by targeting specific pro-inflammatory receptors, and indirectly by impairing the activity of digestive enzymes, the latter event causing the accumulation of undigested peptides with potential immunogenic properties. Herein, according to a concerted approach based on in vitro and in silico methods we characterized kinetics, equilibrium parameters and modes of binding of the complexes formed between wheat ATI and two representative mammalian digestive enzymes, namely trypsin and alpha-amylase. Interestingly, we demonstrated ATI to target both enzymes with independent binding sites and with moderately high affinity. Copyright © 2016 Elsevier Ltd. All rights reserved.

Assessment of salivary amylase as a stress biomarker in pregnant patients.

PubMed

Guglielminotti, J; Dehoux, M; Mentré, F; Bedairia, E; Montravers, P; Desmonts, J-M; Longrois, D

2012-01-01

Chronic stress during pregnancy has been associated with worsened maternal and fetal outcomes. Acute stress immediately before spinal anaesthesia for caesarean section may contribute to hypotension. Therefore objective measures of acute stress may help identify women at risk of adverse outcomes. Salivary alpha-amylase is a stress biomarker that has so far been poorly investigated during pregnancy. The reference change value is the difference between two sequential results that must be exceeded for a change to be considered clinically relevant. Our first aim was to determine if salivary alpha-amylase increased in pregnant patients when subjected to the stress of transfer to the operating room. Our second aim was to determine if changes in salivary alpha-amylase were likely to be clinically significant by measuring reference change value in healthy volunteers. In 15 pregnant patients undergoing planned caesarean section under spinal anaesthesia, salivary alpha-amylase, systolic blood pressure, heart rate, and immediate anxiety were measured on the morning of surgery on the ward and again in the operating room. The reference change value was calculated from 18 healthy volunteers. A median 220% increase in salivary alpha-amylase activity (P=0.0015) and a 17% increase in systolic blood pressure (P=0.0006) were observed between the ward and operating room. No changes of immediate anxiety or heart rate were observed. Reference change value was ±76% in volunteers and 13 of the 15 pregnant patients had a salivary alpha-amylase increase greater than the reference change value. When pregnant women are taken to the operating room, a clinically and statistically significant increase in salivary alpha-amylase was observed. Further studies are required to define its clinical usefulness. Copyright © 2011 Elsevier Ltd. All rights reserved.

Study of starch fermentation at low pH by Lactobacillus fermentum Ogi E1 reveals uncoupling between growth and alpha-amylase production at pH 4.0.

PubMed

Calderon Santoyo, M; Loiseau, G; Rodriguez Sanoja, R; Guyot, J P

2003-01-15

Lactobacillus fermentum Ogi E1 is an amylolytic heterofermentative lactic acid bacterium previously isolated from ogi, a Benin maize sourdough. In the present study, the effect of different pH between 3.5 and 6.0 on starch fermentation products and alpha-amylase production was investigated. Whereas a pH of 5.0 was optimum for specific growth rate and lactic acid production, growth was only slightly affected at suboptimal pH of 4.0 and 6.0. Over a pH range of 6.0 to 3.5, yields of product formation from substrate and of biomass relative to ATP were constant. These results showed that L. fermentum Ogi E1 was particularly acid tolerant, and well adapted to the acid conditions that develop during natural fermentation of cereal doughs. This acid tolerance may partly explain the dominance of L. fermentum in various traditional African sourdoughs. Surprisingly, alpha-amylase production, unlike growth, dropped dramatically when the strain was cultivated at pH 4.0 with starch. With maltose as substrate, the yield of alpha-amylase relative to biomass remained unchanged at pH 4.0 and 5.0, unlike that observed with starch. Based on the distribution of enzyme activity between extra- and intracellular fractions and fermentation kinetics, it appears that starch was first hydrolyzed into dextrins by alpha-amylase activity, and maltose was produced from dextrins by extracellular enzyme activity, transferred into the cell and then hydrolyzed into glucose by intracellular alpha-glucosidase.

Norepinephrine infusion with and without alpha-adrenergic blockade by phentolamine increases salivary alpha amylase in healthy men.

PubMed

Kuebler, Ulrike; von Känel, Roland; Heimgartner, Nadja; Zuccarella-Hackl, Claudia; Stirnimann, Guido; Ehlert, Ulrike; Wirtz, Petra H

2014-11-01

Mental stress reliably induces increases in salivary alpha amylase (sAA), a suggested surrogate marker for sympathetic nervous system (SNS) reactivity. While stress-induced sAA increases correlate with norepinephrine (NE) secretion, a potential mediating role of noradrenergic mechanisms remains unclear. In this study, we investigated for the first time in humans whether a NE-stress-reactivity mimicking NE-infusion with and without alpha-adrenergic blockade by phentolamine would induce changes in sAA. In a single-blind placebo-controlled within-subjects design, 21 healthy men (29-66 years) took part in three different experimental trials varying in terms of substance infusion with a 1-min first infusion followed by a 15-min second infusion: saline-infusion (trial-1), NE-infusion (5 μg/min) without alpha-adrenergic blockade (trial-2), and with phentolamine-induced non-selective blockade of alpha1- and alpha2-adrenergic receptors (trial-3). Saliva samples were collected immediately before, during, and several times after substance infusion in addition to blood pressure and heart rate readings. Experimental trials significantly differed in sAA reactivity to substance-infusion (p=.001) with higher sAA reactivity following NE-infusion with (trial-3; p=.001) and without alpha-adrenergic-blockade (trial-2; p=.004) as compared to placebo-infusion (trial-1); sAA infusion reactivity did not differ between trial-2 and trial-3 (p=.29). Effective phentolamine application was verified by blood pressure and heart rate infusion reactivity. Salivary cortisol was not affected by NE, either with or without alpha-adrenergic-blockade. We found that NE-infusion stimulates sAA secretion, regardless of co-administered non-selective alpha-adrenergic blockade by phentolamine, suggesting that the mechanism underlying stress-induced sAA increases may involve NE. Copyright © 2014 Elsevier Ltd. All rights reserved.

Production and characterization of alpha-amylase from mango kernel by Fusarium solani NAIMCC-F-02956 using submerged fermentation.

PubMed

Kumar, Devendra; Yadav, Kaushlesh K; Muthukumar, M; Garg, Neelima

2013-11-01

Microbial production of enzymes using low valued agro industrial wastes is gaining importance globally. Mango is one of the major fruit processed into a variety of products. During processing 40-50% of solid waste is generated in form of peel and stones. After decortications of mango stone, kernel is obtained which is a rich source of starch (upto 60%). It was utilized as a substrate for alpha-amylase production using Fusarium soloni. Maximum alpha-amylase production (0.889 U g(-1)) was recorded using a substrate concentration of 5% (w/v), pH-4 and temperature 30 degrees C on 9th day of incubation. Supplementation of production medium with micronutrients viz., Ca2+, Fe2+ or Mg2+ improved the enzyme production while, Zn2+, B3+ or Mn2+ ions exhibited inhibitory effect. The extracellular protein was precipitated by ammonium sulphate up to 70% saturation, dialyzed and purified (27.84 fold) by gel-exclusion (Sephadex G-75) chromatography. Protein profiling on 12% SDS-PAGE revealed three bands corresponding to 26, 27 and 30 kDa molecular sizes. The optimum amylase activity was achieved at pH 5.0 at 40 degrees C. The Michaelis constant (KM), Vmax and activation energy (-Ea) were found to be 3.7 mg ml(-1), 0.24 U mg(-1) and 42.39 kJ mole(-1), respectively.

Salivary Alpha Amylase as a Noninvasive Biomarker for Dental Fear and Its Correlation with Behavior of Children during Dental Treatment

PubMed Central

Noorani, Hina; Shivaprakash, PK

2014-01-01

ABSTrACT Objective: Objectives of our studies were to predict dental fear in a child patient depending on salivary alpha amylase (sAA) level before and after dental treatment and to evaluate correla­tion of later with behavior of child patient during dental treatment. Materials and methods: Seventy-seven children between age of 5 and 12 years were divided in three groups. Group 1 consisted of 25 school children who did not undergo any dental treatment. Groups 2 and 3 underwent dental treatment without and with local anesthesia respectively. Groups 2 and 3 were administered child fear survey schedule-dental subscale (CFSS-DS) questionnaire before treatment. Salivary samples were collected for sAA estimation in groups 2 and 3 children before and after completion of dental treatment and behavior during treatment was noted using Frankel behavior rating scale. Group 1 acted as control in which salivary sample was collected in absence of dental stress. Results: When groups 2 and 3 were combined, pretreatment sAA level had a statistically significant (p = 0.0094) correlation with CFSS-DS scores. Conclusion: Alpha amylase can be used as a screening tool to predict level of dental fear in a child patient. How to cite this article: Noorani H, Joshi HV, Shivaprakash PK. Salivary Alpha Amylase as a Noninvasive Biomarker for Dental Fear and Its Correlation with Behavior of Children during Dental Treatment. Int J Clin Pediatr Dent 2014;7(1):19-23. PMID:25206232

Purification of an alpha amylase from Aspergillus flavus NSH9 and molecular characterization of its nucleotide gene sequence.

PubMed

Karim, Kazi Muhammad Rezaul; Husaini, Ahmad; Sing, Ngieng Ngui; Sinang, Fazia Mohd; Roslan, Hairul Azman; Hussain, Hasnain

2018-04-01

In this study, an alpha-amylase enzyme from a locally isolated Aspergillus flavus NSH9 was purified and characterized. The extracellular α-amylase was purified by ammonium sulfate precipitation and anion-exchange chromatography at a final yield of 2.55-fold and recovery of 11.73%. The molecular mass of the purified α-amylase was estimated to be 54 kDa using SDS-PAGE and the enzyme exhibited optimal catalytic activity at pH 5.0 and temperature of 50 °C. The enzyme was also thermally stable at 50 °C, with 87% residual activity after 60 min. As a metalloenzymes containing calcium, the purified α-amylase showed significantly increased enzyme activity in the presence of Ca 2+ ions. Further gene isolation and characterization shows that the α-amylase gene of A. flavus NSH9 contained eight introns and an open reading frame that encodes for 499 amino acids with the first 21 amino acids presumed to be a signal peptide. Analysis of the deduced peptide sequence showed the presence of three conserved catalytic residues of α-amylase, two Ca 2+ -binding sites, seven conserved peptide sequences, and several other properties that indicates the protein belongs to glycosyl hydrolase family 13 capable of acting on α-1,4-bonds only. Based on sequence similarity, the deduced peptide sequence of A. flavus NSH9 α-amylase was also found to carry two potential surface/secondary-binding site (SBS) residues (Trp 237 and Tyr 409) that might be playing crucial roles in both the enzyme activity and also the binding of starch granules.

[Are amylases in bakery products and flour potential food allergens?].

PubMed

Baur, X; Sander, I; Jansen, A; Czuppon, A B

1994-05-21

The enzyme alpha-amylase from the mould Aspergillus oryzae (Asp o II) routinely used for the production of bread, cakes and pastries has in recent years been identified as an inhalative allergen for occupational diseases (bakers' asthma). It is doubtful whether this amylase in the final product, i.e. after the baking procedure, can still be regarded as an allergen. To clarify this question, detailed case histories on 138 subjects were recorded (98 allergics, 20 patients suffering form chronic intestinal diseases, 20 healthy controls). The clinical examinations included prick skin test and IgE antibody determination using one of the customary enzyme preparations. EAST showed a few of these 138 bread consumers to be weakly sensitized to the enzyme. One of the subjects displayed a significant reaction to alpha-amylase heated to 200 degrees C. As expected, eleven bakers sensitized to alpha-amylase by inhaling it in the workplace (positive prick test, positive case history) predominantly exhibited specific IgE antibodies to the native enzyme. Apart from one weakly positive finding, heated alpha-amylase yielded negative results in this collective. Baking conditions vary widely, especially with regard to single components, temperature and duration. Thus, further investigations as to residual allergenicity or the feasible occurrence of new antigenic determinants during the production of bread, cake and pastries are required. 27% of bakers examined and 9% of atopics showed antibodies to a flour inherent enzyme, a beta-amylase. On the whole, the selected conditions hinted at a weakly sensitizing potential inherent in baking flour and in added amylase.

Effects of beta-glucanase and xylanase supplementation on gastrointestinal digestive enzyme activities of weaned piglets fed a barley-based diet.

PubMed

Fan, C L; Han, X Y; Xu, Z R; Wang, L J; Shi, L R

2009-04-01

The effects of supplementing a barley-based diet for weaned piglets withexogenous beta-glucanase and xylanase on gastrointestinal digestiveenzyme activities were investigated. Thirty-six cross-bred weaned pigletswere randomly assigned to two groups with three pens based on sexand mass. Each group was fed on the diet based on barley with or withoutadded beta-glucanase and xylanase (0.15%) for a 4-week period. Theresults showed that enzyme supplementation improved growth performanceof piglets significantly (p < 0.05), but had no effect (p = 0.091)on average daily feed intake. The results also showed that supplementationof beta-glucanase and xylanase had no effect on pepsin activity in gastriccontents but slightly decreased (p = 0.092) the pepsin activity ingastric mucosa. Meanwhile, no effect of enzyme supplementation ontrypsin activity in duodenal contents was observed. However, the activitiesof amylase and lipase in duodenal contents were significantly(p < 0.05) decreased, whereas the activities of maltase, sucrase andgamma-glutamyl transpeptidase (gamma-GT) in jejunal and ileal mucosa wereenhanced significantly (p < 0.05). The improvement of disaccharidaseand gamma-GT activity may be attributed to the positive impacts of exogenousenzymes on digestion and absorption of the nutrients. In conclusion,the current results indicated that supplementation with enzymes in barley-based diets could improve the growth performance of piglets,decrease the activities of amylase and lipase in duodenal contents andincrease the activities of disaccharidase and gamma-GT in jejunal and ilealmucosa.

Daytime Secretion of Salivary Cortisol and Alpha-Amylase in Preschool-Aged Children with Autism and Typically Developing Children

ERIC Educational Resources Information Center

Kidd, Sharon A.; Corbett, Blythe A.; Granger, Douglas A.; Boyce, W. Thomas; Anders, Thomas F.; Tager, Ira B.

2012-01-01

We examined daytime salivary cortisol and salivary alpha-amylase (sAA) secretion levels and variability in preschool-aged children with autism (AUT) and typically developing children (TYP). Fifty-two subjects (26 AUT and 26 TYP) were enrolled. Salivary samples were obtained at waking, midday, and bedtime on two consecutive days at three phases…

Maltose effects on barley malt diastatic power enzyme activity and thermostability at high isothermal mashing temperature: II. Alpha-amylase

USDA-ARS?s Scientific Manuscript database

Maltose, the primary product of starch degradation during mashing, has the potential as a compatible solute to affect the activity of and increase the thermostability of barley malt alpha-amylase activity at high temperatures used in mashing and temperatures above those normally used in mashing. To ...

Protoplast transformation in coryneform bacteria and introduction of an alpha-amylase gene from Bacillus amyloliquefaciens into Brevibacterium lactofermentum.

PubMed Central

Smith, M D; Flickinger, J L; Lineberger, D W; Schmidt, B

1986-01-01

The goal of this study was to investigate the likelihood of developing useful transformation systems for coryneform bacteria. Two species of coryneform bacteria, Brevibacterium lactofermentum and Corynebacterium lilium, were transformed with chimeras constructed from pUB110 and a cryptic coryneform plasmid (pGX1901). C. lilium protoplasts were also efficiently transfected with phage CS1 DNA. High transformation and transfection frequencies were obtained after only 2 min of lysozyme treatment of lysozyme-sensitive mutants. A series of experiments was also conducted to determine whether DNA from other species of important industrial microbes from the genus Bacillus could be expressed in coryneform bacteria. Evidence of restriction of Bacillus subtilis DNA by B. lactofermentum was observed but could be overcome. A Bacillus amyloliquefaciens alpha-amylase gene (amyEBamP) was subcloned onto a plasmid able to replicate in B. lactofermentum. B. lactofermentum transformants for this plasmid expressed amylase activity and produced material cross-reactive to amylase antibody. Images PMID:3008649

Accuracy of alpha amylase in diagnosing microaspiration in intubated critically-ill patients.

PubMed

Dewavrin, Florent; Zerimech, Farid; Boyer, Alexandre; Maboudou, Patrice; Balduyck, Malika; Duhamel, Alain; Nseir, Saad

2014-01-01

Amylase concentration in respiratory secretions was reported to be a potentially useful marker for aspiration and pneumonia. The aim of this study was to determine accuracy of α-amylase in diagnosing microaspiration in critically ill patients. Retrospective analysis of prospectively collected data collected in a medical ICU. All patients requiring mechanical ventilation for at least 48 h, and included in a previous randomized controlled trial were eligible for this study, provided that at least one tracheal aspirate was available for α-amylase measurement. As part of the initial trial, pepsin was quantitatively measured in all tracheal aspirates during a 48-h period. All tracheal aspirates were frozen, allowing subsequent measurement of α-amylase for the purpose of the current study. Microaspiration was defined as the presence of at least one positive tracheal aspirate for pepsin (>200 ng.mL-1). Abundant microaspiration was defined as the presence of pepsin at significant level in >74% of tracheal aspirates. Amylase was measured in 1055 tracheal aspirates, collected from 109 patients. Using mean α-amylase level per patient, accuracy of α-amylase in diagnosing microaspiration was moderate (area under the receiver operator curve 0.72±0.05 [95%CI 0.61-0.83], for an α-amylase value of 1685 UI.L-1). However, when α-amylase levels, coming from all samples, were taken into account, area under the receiver operator curve was 0.56±0.05 [0.53-0.60]. Mean α-amylase level, and percentage of tracheal aspirates positive for α-amylase were significantly higher in patients with microaspiration, and in patients with abundant microaspiration compared with those with no microaspiration; and similar in patients with microaspiration compared with those with abundant microaspiration. α-amylase and pepsin were significantly correlated (r2 = 0.305, p = 0.001). Accuracy of mean α-amylase in diagnosing microaspiration is moderate. Further, when all α-amylase levels

Influence of pectinase treatment on the physicochemical properties of potato flours.

PubMed

Kim, Eun-Jung; Kim, Hyun-Seok

2015-01-15

Untreated and pectinase-treated potato flours from Atlantic and Superior cultivars were characterised to identify the effects of pectinase treatment on their physicochemical properties. Steam-cooked potato whole-tissues were treated with and without pectinase to prepare the dehydrated potato flours. Untreated and pectinase-treated potato flours were investigated with respect to morphology, chemical composition, starch leaching, swelling power, gelatinization, and pasting viscosity. Upon viewing with scanning electron microscopy and light microscopy, the pectinase-treated (relative to untreated) potato flours revealed that the retrograded starch materials were present in intact parenchyma cells, apparently exhibiting granular structures. Their protein and ash contents were reduced through pectinase treatment. While starch leachate contents were lower for the pectinase-treated potato flours, the opposite trend in swelling powers was observed. Pectinase-treated potato flours exhibited higher melting temperatures and pasting viscosities than untreated counterparts. Overall, the modification of potato flour morphology by pectinase treatment may result in alteration of physicochemical properties of potato flours. Copyright © 2014 Elsevier Ltd. All rights reserved.

Solid State Fermentation of a Raw Starch Digesting Alkaline Alpha-Amylase from Bacillus licheniformis RT7PE1 and Its Characteristics.

PubMed

Tabassum, Romana; Khaliq, Shazia; Rajoka, Muhammad Ibrahim; Agblevor, Foster

2014-01-01

The thermodynamic and kinetic properties of solids state raw starch digesting alpha amylase from newly isolated Bacillus licheniformis RT7PE1 strain were studied. The kinetic values Q p , Y p/s , Y p/X , and q p were proved to be best with 15% wheat bran. The molecular weight of purified enzyme was 112 kDa. The apparent K m and V max values for starch were 3.4 mg mL(-1) and 19.5 IU mg(-1) protein, respectively. The optimum temperature and pH for α -amylase were 55°C, 9.8. The half-life of enzyme at 95°C was 17h. The activation and denaturation activation energies were 45.2 and 41.2 kJ mol(-1), respectively. Both enthalpies (ΔH (∗)) and entropies of activation (ΔS (∗)) for denaturation of α -amylase were lower than those reported for other thermostable α -amylases.

Solid State Fermentation of a Raw Starch Digesting Alkaline Alpha-Amylase from Bacillus licheniformis RT7PE1 and Its Characteristics

PubMed Central

Tabassum, Romana; Khaliq, Shazia; Rajoka, Muhammad Ibrahim; Agblevor, Foster

2014-01-01

The thermodynamic and kinetic properties of solids state raw starch digesting alpha amylase from newly isolated Bacillus licheniformis RT7PE1 strain were studied. The kinetic values Q p, Y p/s, Y p/X, and q p were proved to be best with 15% wheat bran. The molecular weight of purified enzyme was 112 kDa. The apparent K m and V max values for starch were 3.4 mg mL−1 and 19.5 IU mg−1 protein, respectively. The optimum temperature and pH for α-amylase were 55°C, 9.8. The half-life of enzyme at 95°C was 17h. The activation and denaturation activation energies were 45.2 and 41.2 kJ mol−1, respectively. Both enthalpies (ΔH ∗) and entropies of activation (ΔS ∗) for denaturation of α-amylase were lower than those reported for other thermostable α-amylases. PMID:24587909

Blunted cortisol response to psychosocial stress in atopic patients is associated with decrease in salivary alpha-amylase and aldosterone: Focus on sex and menstrual cycle phase.

PubMed

Hlavacova, N; Solarikova, P; Marko, M; Brezina, I; Jezova, D

2017-04-01

A decreased responsiveness of the hypothalamic-pituitary-adrenocortical axis to stress stimuli in patients with atopy is well documented. The aim of this study was to investigate personality traits, salivary alpha-amylase activity and the aldosterone response to psychosocial stress procedure based on public speech in atopic patients with respect to sex and the menstrual cycle (MC) phase. The study was performed in 106 subjects of both sexes, 53 atopic patients suffering from allergic rhinitis, allergic asthma or atopic dermatitis and 53 age-, sex-, the MC phase- and BMI- matched healthy controls. Substantially attenuated activity of alpha-amylase and reduced secretion of aldosterone during the psychosocial stress were observed in the whole sample of patients with atopy. Higher activity of alpha-amylase observed in the follicular compared to the luteal phase in healthy women was not present in atopic patients. In both males and females, atopy was associated with blunted cortisol response but no changes in the heart rate. Psychological characterization revealed a significantly higher trait anxiety and higher preference for avoidance-oriented coping strategy in female but not male atopic patients. These findings provide evidence that patients with atopy exhibit insufficient alpha-amylase and aldosterone responsiveness to psychosocial stress, thus suggesting decreased sympathetic activity. Potential disturbances in sex hormone status during the MC in female patients with atopy have to be considered in future research. Changes in personality traits were demonstrated in female atopic patients, but not in male patients. Copyright © 2017. Published by Elsevier Ltd.

Preparation and characterization of a dextran-amylase conjugate.

PubMed

Marshall, J J

1976-07-01

Bacillus amyloliquefaciens alpha-amylase was attached to dextran after activation of the polysaccharide by using a modification of the cyanogen bromide method. The soluble dextran-amylase conjugate was purified by molecular-sieve chromatography. The conjugated enzyme has greater stability than the unmodified enzyme at low pH values, during heat treatment, and on removal of calcium ions with a chelating agent. Attachment of dextran to alpha-amylase did not alter the Michaelis constant of the enzyme acting on starch. The polysaccharide-enzyme conjugate probably consists of a cross-linked aggregate of many dextran and many enzyme molecules, in which a proportion of the enzyme molecules, although not inactivated, are unable to express their activity, except after dextranase treatment.

Immunohistochemistry of active gibberellins and gibberellin-inducible alpha-amylase in developing seeds of morning glory.

PubMed

Nakayama, Akira; Park, Seijin; Zheng-Jun, Xu; Nakajima, Masatoshi; Yamaguchi, Isomaro

2002-07-01

Gibberellins (GAs) in developing seeds of morning glory (Pharbitis nil) were quantified and localized by immunostaining. The starch grains began to be digested after the GA contents had increased and reached a plateau. Immunohistochemical staining with the antigibberellin A(1)-methyl ester-antiserum, which has high affinity to biologically active GAs, showed that GA(1) and/or GA(3) were localized around starch grains in the integument of developing young seeds, suggesting the participation of GA-inducible alpha-amylase in this digestion. We isolated an alpha-amylase cDNA (PnAmy1) that was expressed in the immature seeds, and using an antibody raised against recombinant protein, it was shown that PnAmy1 was expressed in the immature seeds. GA responsiveness of PnAmy1 was shown by treating the young fruits 9 d after anthesis with GA(3). RNA-blot and immunoblot analyses showed that PnAmy1 emerged soon after the rapid increase of GA(1/3). An immunohistochemical analysis of PnAmy1 showed that it, like the seed GA(1/3), was also localized around starch grains in the integument of developing young seeds. The localization of GA(1/3) in the integument coincident with the expression of PnAmy1 suggests that both function as part of a process to release sugars for translocation or for the further development of the seeds.

Effects of dietary xylanase supplementation on performance and functional digestive parameters in broilers fed wheat-based diets.

PubMed

Liu, Wen-Chao; Kim, In-Ho

2017-03-01

This experiment was conducted to evaluate dietary xylanase supplementation in broilers wheat-based diets on performance and functional digestive parameters including ileal digesta viscosity, apparent ileal digestibility, intestinal morphology and microflora, digestive enzyme activities, and excreta odor content. A total of 600 1-day-old Ross 308 male broilers with an initial average BW of 45 ± 0.6 g were randomly allotted into 4 treatments with 10 replicate pens per treatment and 15 broilers in each pen for 35 d. The 4 dietary treatments were wheat-based diets and supplemented with 0, 1,875, 3,750, and 5,625 XU/kg xylanase. Xylanase supplementation improved (linear, P < 0.05) the body weight gain and decreased (linear, P < 0.05) the feed conversion ratio during d 1 to 18 and for the duration of the experiment. Dietary supplementation of xylanase led to a decrease (linear, P < 0.01) in ileal digesta viscosity. The apparent ileal digestibility of dry matter (DM), crude protein (CP), gross energy, and most amino acids (with the exception of Ile, Phe, Asp, Glu, and Pro) were increased (linear, P < 0.05) by xylanase supplementation. Increasing the dietary xylanase levels improved (linear, P < 0.05) the villus height and the ratio of villus height to crypt depth of the duodenum, jejunum, and ileum. In addition, inclusion of xylanase increased (linear, P < 0.05) the Lactobacillus numbers in the ileum and cecum, while decreased the ileal E. coli counts (linear, P < 0.01) and cecal E. coli populations (linear, P < 0.01; quadratic, P < 0.05). There were no significant differences (P > 0.05) in trypsin, amylase, and protease activities of small intestine among dietary treatments. Furthermore, xylanase supplementation reduced excreta NH3 (linear, P < 0.05; quadratic, P < 0.05) and total mercaptan (R.SH) (linear, P < 0.01) concentration. Taken together, dietary xylanase supplementation in broilers wheat-based diets had beneficial effects on growth

THESIS-ABSTRACT Supplementation levels of exogenous alpha-amylase in broilers diets.

PubMed

Oliveira, H B; Silva, M I A; Mesquita, F R

2017-08-17

This study aimed to evaluate the supplementation levels of an exogenous alpha-amylase in broilers diets and compare two indicators in determining the diets energy. The experiment was divided into two parallel evaluations, being one of performance and the other of metabolism. In performance assay, 1,700 one-day-old Cobb-500 male chicks were used. The animals were distributed in 50 experimental plots and evaluated five treatments with ten replicates in a completely randomized design (CRD). The treatments were: a positive control (PC), a negative control (NC) and three alpha-amylase supplementation levels 200, 400 and 600 g/t, and the NC was formulated with 50 and 90 kcal of energy reduction in relation to the PC to the phases from 1 to 21 days and from 22 to 42 days, respectively. In the metabolism assay were used 240 animals, 150 birds for stage from 14 to 21 days and 90 birds to stage from 35 to 42 days of age and the treatments were the same as the performance assay, with six replicates per treatment in CRD. All diets of metabolism test contained the digestibility indicators Lipe ® (eucalyptus purified lignin) and chromic oxide (Cr 2 O 3 ), in concentrations of 0.05 and 1.0%, respectively. In the period from 1 to 21 days old, no significant differences were observed in weight gain (WG) (P > 0.05), however, feed intake (FI) was found higher by using 200 ppm of enzyme (P < 0.05) and better feed conversion (FC) with the PC (P < 0.05). From 22 to 42 days, no significant differences were observed on the WG (P > 0.05), but were observed lower FI and better FC to PC treatment (P < 0.05). In the period from 1 to 42 days of age, significant differences were also not observed on the WG (P > 0.05), but there was lower FI and better FC for the PC treatment (P < 0.05). The AMEn (apparent metabolizable energy corrected for nitrogen balance), determined using the total collection, reaffirmed the values ​​calculated for the PC and NC with intermediate data obtained from the

RNA interference-based silencing of the alpha-amylase (amy1) gene in Aspergillus flavus decreases fungal growth and aflatoxin production in maize kernels.

PubMed

Gilbert, Matthew K; Majumdar, Rajtilak; Rajasekaran, Kanniah; Chen, Zhi-Yuan; Wei, Qijian; Sickler, Christine M; Lebar, Matthew D; Cary, Jeffrey W; Frame, Bronwyn R; Wang, Kan

2018-06-01

Expressing an RNAi construct in maize kernels that targets the gene for alpha-amylase in Aspergillus flavus resulted in suppression of alpha-amylase (amy1) gene expression and decreased fungal growth during in situ infection resulting in decreased aflatoxin production. Aspergillus flavus is a saprophytic fungus and pathogen to several important food and feed crops, including maize. Once the fungus colonizes lipid-rich seed tissues, it has the potential to produce toxic secondary metabolites, the most dangerous of which is aflatoxin. The pre-harvest control of A. flavus contamination and aflatoxin production is an area of intense research, which includes breeding strategies, biological control, and the use of genetically-modified crops. Host-induced gene silencing, whereby the host crop produces siRNA molecules targeting crucial genes in the invading fungus and targeting the gene for degradation, has shown to be promising in its ability to inhibit fungal growth and decrease aflatoxin contamination. Here, we demonstrate that maize inbred B104 expressing an RNAi construct targeting the A. flavus alpha-amylase gene amy1 effectively reduces amy1 gene expression resulting in decreased fungal colonization and aflatoxin accumulation in kernels. This work contributes to the development of a promising technology for reducing the negative economic and health impacts of A. flavus growth and aflatoxin contamination in food and feed crops.

Immobilization of pectinase onto chitosan magnetic nanoparticles by macromolecular cross-linker.

PubMed

Sojitra, Uttam V; Nadar, Shamraja S; Rathod, Virendra K

2017-02-10

Pectinase was immobilized onto chitosan magnetic nanoparticles (CMNPs) by dextran polyaldehyde as a macromolecular cross-linking agent. The parameters like cross-linking concentration, time and CMNPs to enzyme ratio were optimized. Further, prepared magnetic pectinase nanobiocatalyst was characterized by FT-IR and XRD. The thermal kinetic studies for immobilized pectinase showed two folds improved thermal stability in the range of 55-75°C as compared to free form. The V max and K m values of immobilized pectinase were found to be nearly equal to native form which indicated that conformational flexibility of pectinase was retained even after immobilization. The residual activity of immobilized pectinase was 85% after seven successive cycles of reuse, while it retained upto 89% residual activity on storage of fifteen days which exhibited excellent stability and durability. The conformational changes in pectinase after immobilization were evaluated by FT-IR spectroscopy data analysis tools. Finally, magnetic pectinase nanobiocatalyst was employed for apple juice clarification which showed turbidity reduction upto 74% after 150min treatment. Copyright © 2016 Elsevier Ltd. All rights reserved.

Alternative method for quantification of alfa-amylase activity.

PubMed

Farias, D F; Carvalho, A F U; Oliveira, C C; Sousa, N M; Rocha-Bezerrra, L C B; Ferreira, P M P; Lima, G P G; Hissa, D C

2010-05-01

A modification of the sensitive agar diffusion method was developed for macro-scale determination of alfa-amylase. The proposed modifications lower costs with the utilisation of starch as substrate and agar as supporting medium. Thus, a standard curve was built using alfa-amylase solution from Aspergillus oryzae, with concentrations ranging from 2.4 to 7,500 U.mL-1. Clear radial diffusion zones were measured after 4 hours of incubation at 20 A degrees C. A linear relationship between the logarithm of enzyme activities and the area of clear zones was obtained. The method was validated by testing alpha-amylase from barley at the concentrations of 2.4; 60; 300 and 1,500 U.mL-1. The proposed method turned out to be simpler, faster, less expensive and able to determine on a macro-scale alpha-amylase over a wide range (2.4 to 7,500 U.mL-1) in scientific investigation as well as in teaching laboratory activities.

Structure-Specificity Relationships of an Intracellular Xylanase from Geobacillus stearothermophilus

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

Solomon,V.; Teplitsky, A.; Shulami, S.

2007-01-01

Geobacillus stearothermophilus T-6 is a thermophilic Gram-positive bacterium that produces two selective family 10 xylanases which both take part in the complete degradation and utilization of the xylan polymer. The two xylanases exhibit significantly different substrate specificities. While the extracellular xylanase (XT6; MW 43.8 kDa) hydrolyzes the long and branched native xylan polymer, the intracellular xylanase (IXT6; MW 38.6 kDa) preferentially hydrolyzes only short xylo-oligosaccharides. In this study, the detailed three-dimensional structure of IXT6 is reported, as determined by X-ray crystallography. It was initially solved by molecular replacement and then refined at 1.45 {angstrom} resolution to a final R factormore » of 15.0% and an R{sub free} of 19.0%. As expected, the structure forms the classical ({alpha}/{beta}){sub 8} fold, in which the two catalytic residues (Glu134 and Glu241) are located on the inner surface of the central cavity. The structure of IXT6 was compared with the highly homologous extracellular xylanase XT6, revealing a number of structural differences between the active sites of the two enzymes. In particular, structural differences derived from the unique subdomain in the carboxy-terminal region of XT6, which is completely absent in IXT6. These structural modifications may account for the significant differences in the substrate specificities of these otherwise very similar enzymes.« less

Characterization, kinetic, and thermodynamic studies of marine pectinase from Bacillus subtilis.

PubMed

Joshi, Manasi; Nerurkar, Madhura; Adivarekar, Ravindra

2015-01-01

Characterization, kinetic and thermodynamic parameters of purified pectinase from Bacillus subtilis, isolated from a marine sediment sample collected from Chinchani beach at Tarapore, India, were studied. Marine pectinase produced under submerged growth conditions was purified by ammonium sulfate precipitation followed by gel filtration chromatography using DEAE cellulose. Partial characterization of the marine pectinase was carried out in terms of effect of pH, temperature, substrate concentration, and metal ions. It was found that pectinase from marine B. subtilis showed maximal activity in alkaline buffer at pH 9.0 and at 40°C. It was also found that metal ions, namely, Mn(2+) and Fe(2+), stimulate pectinase activity. Marine pectinase followed Michaelis-Menten kinetics. The kinetics and thermodynamic parameters of the purified marine pectinase from B. subtilis were studied as the characterization of the enzyme is vital for its use in industrial processes.

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.

A proprietary alpha-amylase inhibitor from white bean (Phaseolus vulgaris): A review of clinical studies on weight loss and glycemic control

PubMed Central

2011-01-01

Obesity, and resultant health hazards which include diabetes, cardiovascular disease and metabolic syndrome, are worldwide medical problems. Control of diet and exercise are cornerstones of the management of excess weight. Foods with a low glycemic index may reduce the risk of diabetes and heart disease as well as their complications. As an alternative to a low glycemic index diet, there is a growing body of research into products that slow the absorption of carbohydrates through the inhibition of enzymes responsible for their digestion. These products include alpha-amylase and glucosidase inhibitors. The common white bean (Phaseolus vulgaris) produces an alpha-amylase inhibitor, which has been characterized and tested in numerous clinical studies. A specific and proprietary product named Phase 2® Carb Controller (Pharmachem Laboratories, Kearny, NJ) has demonstrated the ability to cause weight loss with doses of 500 to 3000 mg per day, in either a single dose or in divided doses. Clinical studies also show that Phase 2 has the ability to reduce the post-prandial spike in blood glucose levels. Experiments conducted incorporating Phase 2 into food and beverage products have found that it can be integrated into various products without losing activity or altering the appearance, texture or taste of the food. There have been no serious side effects reported following consumption of Phase 2. Gastro-intestinal side effects are rare and diminish upon extended use of the product. In summary, Phase 2 has the potential to induce weight loss and reduce spikes in blood sugar caused by carbohydrates through its alpha-amylase inhibiting activity. PMID:21414227

A proprietary alpha-amylase inhibitor from white bean (Phaseolus vulgaris): a review of clinical studies on weight loss and glycemic control.

PubMed

Barrett, Marilyn L; Udani, Jay K

2011-03-17

Obesity, and resultant health hazards which include diabetes, cardiovascular disease and metabolic syndrome, are worldwide medical problems. Control of diet and exercise are cornerstones of the management of excess weight. Foods with a low glycemic index may reduce the risk of diabetes and heart disease as well as their complications. As an alternative to a low glycemic index diet, there is a growing body of research into products that slow the absorption of carbohydrates through the inhibition of enzymes responsible for their digestion. These products include alpha-amylase and glucosidase inhibitors. The common white bean (Phaseolus vulgaris) produces an alpha-amylase inhibitor, which has been characterized and tested in numerous clinical studies. A specific and proprietary product named Phase 2® Carb Controller (Pharmachem Laboratories, Kearny, NJ) has demonstrated the ability to cause weight loss with doses of 500 to 3000 mg per day, in either a single dose or in divided doses. Clinical studies also show that Phase 2 has the ability to reduce the post-prandial spike in blood glucose levels. Experiments conducted incorporating Phase 2 into food and beverage products have found that it can be integrated into various products without losing activity or altering the appearance, texture or taste of the food. There have been no serious side effects reported following consumption of Phase 2. Gastro-intestinal side effects are rare and diminish upon extended use of the product. In summary, Phase 2 has the potential to induce weight loss and reduce spikes in blood sugar caused by carbohydrates through its alpha-amylase inhibiting activity.

Applications of pectinases in the commercial sector: a review.

PubMed

Kashyap, D R; Vohra, P K; Chopra, S; Tewari, R

2001-05-01

Pectinases are one of the upcoming enzymes of fruit and textile industries. These enzymes break down complex polysaccharides of plant tissues into simpler molecules like galacturonic acids. The role of acidic pectinases in bringing down the cloudiness and bitterness of fruit juices is well established. Recently, there has been a good number of reports on the application of alkaline pectinases in the textile industry for the retting and degumming of fiber crops, production of good quality paper, fermentation of coffee and tea, oil extractions and treatment of pectic waste water. This review discusses various types of pectinases and their applications in the commercial sector.

Discovering an Accessible Enzyme: Salivary [alpha]-Amylase--"Prima Digestio Fit in Ore"--A Didactic Approach for High School Students

ERIC Educational Resources Information Center

Marini, Isabella

2005-01-01

Human salivary [alpha]-amylase is used in this experimental approach to introduce biology high school students to the concept of enzyme activity in a dynamic way. Through a series of five easy, rapid, and inexpensive laboratory experiments students learn what the activity of an enzyme consists of: first in a qualitative then in a semi-quantitative…

Chestnut astringent skin extract, an alpha-amylase inhibitor, retards carbohydrate absorption in rats and humans.

PubMed

Tsujita, Takahiro; Takaku, Takeshi; Suzuki, Tsuneo

2008-02-01

Inhibitors of carbohydrate-hydrolyzing enzyme play an important role to control postprandial blood glucose levels. In this paper, we investigated the effect of an ethanol extract from chestnut astringent skin (CAS) on alpha-amylase. Chestnut astringent skin extract strongly inhibited human and porcine pancreatic alpha-amylase. We also investigated the effect of CAS extract on carbohydrate absorption in rats and humans. Oral administration of CAS extract to normal rats fed corn starch (2 g/kg body weight), significantly suppressed the increase of blood glucose levels after starch loading in a dose-dependent manner. The effective dose of CAS extract required to achieve 20 and 40% suppression of the rise in blood glucose level was estimated to be 40 and 155 mg/kg body weight, respectively. Chestnut astringent skin extract also suppressed the rise in plasma insulin level and the fall in plasma non-esterified fatty acid level. In the type 2 diabetic rat model, CAS extract significantly suppressed the rise in blood glucose level after starch loading in a dose-dependent manner. Chestnut astringent skin extract also suppressed the rise in plasma glucose level after boiled rice loading in a dose-dependent manner in humans. The amount of CAS extract required to achieve 11 and 23% suppression in the rise in plasma glucose level was 300 and 600 mg/person, respectively. These results suggest that CAS extract retards absorption of carbohydrate and reduces post-prandial hyperglycemia.

Secretome analysis of the thermophilic xylanase hyper-producer Thermomyces lanuginosus SSBP cultivated on corn cobs.

PubMed

Winger, A M; Heazlewood, J L; Chan, L J G; Petzold, C J; Permaul, K; Singh, S

2014-11-01

Thermomyces lanuginosus is a thermophilic fungus known for its ability to produce industrially important enzymes including large amounts of xylanase, the key enzyme in hemicellulose hydrolysis. The secretome of T. lanuginosus SSBP was profiled by shotgun proteomics to elucidate important enzymes involved in hemicellulose saccharification and to characterise the presence of other industrially interesting enzymes. This study reproducibly identified a total of 74 proteins in the supernatant following growth on corn cobs. An analysis of proteins revealed nine glycoside hydrolase (GH) enzymes including xylanase GH11, β-xylosidase GH43, β-glucosidase GH3, α-galactosidase GH36 and trehalose hydrolase GH65. Two commercially produced Thermomyces enzymes, lipase and amylase, were also identified. In addition, other industrially relevant enzymes not currently explored in Thermomyces were identified including glutaminase, fructose-bisphosphate aldolase and cyanate hydratase. Overall, these data provide insight into the novel ability of a cellulase-free fungus to utilise lignocellulosic material, ultimately producing a number of enzymes important to various industrial processes.

Salivary alpha amylase in on-call from home fire and emergency service personnel.

PubMed

Hall, Sarah J; Aisbett, Brad; Robertson, Samuel J; Ferguson, Sally A; Turner, Anne I

2017-11-01

The effect of working on-call from home on the sympatho-adrenal medullary system activity is currently unknown. This study had two aims, Aim 1: examine salivary alpha amylase awakening response (AAR) and diurnal salivary alpha amylase (sAA) profile in fire and emergency service workers who operate on-call from home following a night on-call with a call (NIGHT-CALL), a night on-call without a call (NO-CALL) and an off-call night (OFF-CALL), and Aim 2: explore whether there was an anticipatory effect of working on-call from home (ON) compared to when there was an off-call (OFF) on the diurnal sAA profile. Participants wore activity monitors, completed sleep and work diaries and collected seven saliva samples a day for one week. AAR area under the curve with respect to ground (AUC G ), AAR area under the curve with respect to increase (AUC I ), AAR reactivity, diurnal sAA slope, diurnal sAA AUC G and mean 12-h sAA concentrations were calculated. Separate generalised estimating equation models were constructed for each variable of interest for each aim. For Aim 1, there were no differences between NIGHT-CALL or NO-CALL and OFF-CALL for any response variable. For Aim 2, there was no difference between any response variable of interest when ON the following night compared to when OFF the following night ( n  = 14). These findings suggest that there is no effect of working on-call from home on sAA, but should be interpreted with caution, as overnight data were not collected. Future research, using overnight heart rate monitoring, could help confirm these findings. © 2017 The authors.

Evaluation of alpha- amylase inhibition by Urtica dioica and Juglans regia extracts.

PubMed

Rahimzadeh, Mahsa; Jahanshahi, Samaneh; Moein, Soheila; Moein, Mahmood Reza

2014-06-01

One strategy for the treatment of diabetes is inhibition of pancreatic α- amylase. Plants contains different chemical constituents with potential for inhibition of α-amylase and hence maybe used as therapeutic. Urtica dioica and Juglans regia Linn were tested for α-amylase inhibition. Different concentrations of leaf aqueous extracts were incubated with enzyme substrate solution and the activity of enzyme was measured. For determination of the type of inhibition, Dixon plot was depicted. Acarbose was used as the standard inhibitor. Both plant extracts showed time and concentration dependent inhibition of α-amylase. 60% inhibition was seen with 2 mg/ml of U. dioica and 0.4 mg/ml of J. regia aqueous extract. Dixon plots revealed the type of α-amylase inhibition by these two extracts as competitive inhibition. Determination of the type of α-amylase inhibition by these plant extracts could provide by successful use of plant chemicals as drug targets.

Evaluation of alpha- amylase inhibition by Urtica dioica and Juglans regia extracts

PubMed Central

Rahimzadeh, Mahsa; Jahanshahi, Samaneh; Moein, Soheila; Moein, Mahmood Reza

2014-01-01

Objective(s): One strategy for the treatment of diabetes is inhibition of pancreatic α- amylase. Plants contains different chemical constituents with potential for inhibition of α-amylase and hence maybe used as therapeutic. Materials and Methods: Urtica dioica and Juglans regia Linn were tested for α-amylase inhibition. Different concentrations of leaf aqueous extracts were incubated with enzyme substrate solution and the activity of enzyme was measured. For determination of the type of inhibition, Dixon plot was depicted. Acarbose was used as the standard inhibitor. Results: Both plant extracts showed time and concentration dependent inhibition of α-amylase. 60% inhibition was seen with 2 mg/ml of U. dioica and 0.4 mg/ml of J. regia aqueous extract. Dixon plots revealed the type of α-amylase inhibition by these two extracts as competitive inhibition. Conclusion: Determination of the type of α-amylase inhibition by these plant extracts could provide by successful use of plant chemicals as drug targets. PMID:25140210

Substrate binding ability of chemically inactivated pectinase for the substrate pectic acid.

PubMed

Chiba, Y; Kobayashi, M

1995-07-01

Pectinase (polygalacturonase) was purified from a commercial pectinase preparation from a mold. Substrate binding of pectinase was measured by centrifugal affinity chromatography using an immobilized substrate, pectic acid. Desorption of pectinase from the affinity matrix with the substrate pectin and pectic acid gave Kd values of 5.3 and 8.5 mg/ml, respectively. Chemical modification of pectinase by 1-ethyl-3-(3-dimethyl-aminopropyl)carbodiimide (EDC) and diethyl pyrocarbonate (DEP) caused a loss of most of the enzyme activity, but the substrate binding ability was not impaired. Thus, the pectinase preparation was digested with lysyl endopeptidase and the resulting peptides were treated with pectic acid-affinity gel. Three peptide fragments, which were recovered from the affinity column and sequenced, were identical to sequences in the second pectinase gene from Aspergillus niger. The first peptide contained 17 amino acids, Asp101-Ser117, and the second and third peptides corresponded to 18 amino acids of Asn152-Asp169. These results indicate that the inactivated pectinase retained substrate binding ability and would function as an acidic polysaccharide recognizing protein.

Alpha-amylase inhibitor, CS-1036 binds to serum amylase in a concentration-dependent and saturable manner.

PubMed

Honda, Tomohiro; Kaneno-Urasaki, Yoko; Ito, Takashi; Kimura, Takako; Matsushima, Nobuko; Okabe, Hiromi; Yamasaki, Atsushi; Izumi, Takashi

2014-03-01

(2R,3R,4R)-4-hydroxy-2-(hydroxymethyl)pyrrolidin-3-yl 4-O-(6-deoxy-β-D-glucopyranosyl)-α-D-glucopyranoside (CS-1036), which is an α-amylase inhibitor, exhibited biphasic and sustained elimination with a long t1/2 (18.4-30.0 hours) in rats and monkeys, but exhibited a short t1/2 (3.7-7.9 hours) in humans. To clarify the species differences in the t1/2, the plasma protein binding of CS-1036 was evaluated by ultrafiltration. A concentration-dependent and saturable plasma protein binding of CS-1036 was observed in rats and monkeys with the dissociation rate constant (KD) of 8.95 and 27.2 nM, and maximal binding capacity (Bmax) of 52.8 and 22.1 nM, respectively. By the assessments of the recombinant amylase and immunoprecipitation, the major binding protein of CS-1036 in rats was identified as salivary amylase (KD 5.64 nM). CS-1036 also showed concentration-dependent and saturable binding to human salivary and pancreatic amylase, with similar binding affinity in rats. However, the protein binding of CS-1036 was constant in human plasma (≤10.2%) due to the lower serum amylase level compared with rats and monkeys. From the calculation of the unbound fraction (fu) in plasma based on in vitro KD and Bmax, the dose-dependent increase in fu after oral administration is speculated to lead to a dose-dependent increase in total body clearance and a high area under the curve/dose at lower doses, such as 0.3 mg/kg in rats.

Efficient production of lignocellulolytic enzymes xylanase, β-xylosidase, ferulic acid esterase and β-glucosidase by the mutant strain Aspergillus awamori 2B.361 U2/1

PubMed Central

Gottschalk, Leda Maria Fortes; de Sousa Paredes, Raquel; Teixeira, Ricardo Sposina Sobral; da Silva, Ayla Sant’Ana; da Silva Bon, Elba Pinto

2013-01-01

The production of xylanase, β-xylosidase, ferulic acid esterase and β-glucosidase by Aspergillus awamori 2B.361 U2/1, a hyper producer of glucoamylase and pectinase, was evaluated using selected conditions regarding nitrogen nutrition. Submerged cultivations were carried out at 30 °C and 200 rpm in growth media containing 30 g wheat bran/L as main carbon source and either yeast extract, ammonium sulfate, sodium nitrate or urea, as nitrogen sources; in all cases it was used a fixed molar carbon to molar nitrogen concentration of 10.3. The use of poor nitrogen sources favored the accumulation of xylanase, β-xylosidase and ferulic acid esterase to a peak concentrations of 44,880; 640 and 118 U/L, respectively, for sodium nitrate and of 34,580, 685 and 170 U/L, respectively, for urea. However, the highest β-glucosidase accumulation of 10,470 U/L was observed when the rich organic nitrogen source yeast extract was used. The maxima accumulation of filter paper activity, xylanase, β-xylosidase, ferulic acid esterase and β-glucosidase by A. awamori 2B.361 U2/1 was compared to that produced by Trichoderma reesei Rut-C30. The level of β-glucosidase was over 17-fold higher for the Aspergillus strain, whereas the levels of xylanase and β-xylosidase were over 2-fold higher. This strain also produced ferulic acid esterase (170 U/L), which was not detected in the T. reesei culture. PMID:24294256

Wax removal for accelerated cotton scouring with alkaline pectinase.

PubMed

Agrawal, Pramod B; Nierstrasz, Vincent A; Klug-Santner, Barbara G; Gübitz, Georg M; Lenting, Herman B M; Warmoeskerken, Marijn M C G

2007-03-01

A rational approach has been applied to design a new environmentally acceptable and industrially viable enzymatic scouring process. Owing to the substrate specificity, the selection of enzymes depends on the structure and composition of the substrate, i.e. cotton fibre. The structure and composition of the outer layers of cotton fibre has been established on the basis of thorough literature study, which identifies wax and pectin removal to be the key steps for successful scouring process. Three main issues are discussed here, i.e. benchmarking of the existing alkaline scouring process, an evaluation of several selected acidic and alkaline pectinases for scouring, and the effect of wax removal treatment on pectinase performance. It has been found that the pectinolytic capability of alkaline pectinases on cotton pectin is nearly 75% higher than that of acidic pectinases. It is concluded that an efficient wax removal prior to pectinase treatment indeed results in improved performance in terms of hydrophilicity and pectin removal. To evaluate the hydrophilicity, the structural contact angle (theta) was measured using an auto-porosimeter.

Extracellular Xylanopectinolytic Enzymes by Bacillus subtilis ADI1 from EFB's Compost

PubMed Central

Nawawi, Muhammad Hariadi; Mohamad, Rosfarizan; Tahir, Paridah Md.

2017-01-01

Microbial xylanase and pectinase are two extremely valuable enzymes, which have captivated much attention. This can be seen from the increased demand for these enzymes by many industrial sectors. This study investigates the isolation and screening of extracellular xylanopectinolytic enzymes-producing bacteria in a submerged fermentation (SmF). Samples are collected from the compost of empty fruit bunch (EFB) at Biocompost Pilot Plant, located at Biorefinery Plant, Universiti Putra Malaysia. From the experiment, out of 20 isolates, 11 isolates show xylanase or/and pectinase activity, and only one isolate (EFB-11) shows the concurrent activities of xylanase and pectinase. These activities are selected for enzyme production under submerged fermentation (quantitative screening). At the 72nd hour of incubation, xylanase and pectinase show the highest production, which ranges about 42.33 U/mL and 62.17 U/mL (with low amount of cellulase present), supplemented with 2% (w/v) of rice bran as carbon source at incubation temperature level, which is 30°C. Meanwhile, the pH of media is shifted to 8.42, which indicates that EFB-11 isolate is alkalotolerant bacteria and identified as Bacillus subtilis ADI1. This strain proves to have potential in agroindustrial bioconversion and has a promising ability to scale up to an industrial scale. PMID:28523288

Xylanase II from an alkaliphilic thermophilic Bacillus with a distinctly different structure from other xylanases: evolutionary relationship to alkaliphilic xylanases.

PubMed

Kulkarni, N; Lakshmikumaran, M; Rao, M

1999-10-05

A 1.0 kilobase gene fragment from the genomic DNA of an alkaliphilic thermophilic Bacillus was found to code for a functional xylanase (XynII). The complete nucleotide sequence including the structural gene and the 5' and 3' flanking sequences of the xylanase gene have been determined. An open reading frame starting from ATG initiator codon comprising 402 nucleotides gave a preprotein of 133 amino acids of calculated molecular mass 14.090 kDa. The occurrence of three potential N-glycosylation sites in XynII gene is a unique feature for a gene of bacterial origin. The stop codon was followed by hairpin loop structures indicating the presence of transcription termination signals. The secondary structure analysis of XynII predicted that the polypeptide was primarily formed of beta-sheets. XynII appeared to be a member of family G/11 of xylanases based on its molecular weight and basic pI (8.0). However, sequence homology revealed similar identity with families 10 and 11 of xylanases. The conserved triad (Val-Val-Xaa, where Xaa is Asn or Asp) was identified only in the xylanases from alkaliphilic organisms. Our results implicate for the first time the concept of convergent evolution for XynII and provide a basis for research in evolutionary relationship among the xylanases from alkaliphilic and neutrophilic organisms. Copyright 1999 Academic Press.

Biosilica-Immobilized Enzymes for Biocatalysis (Preprint)

DTIC Science & Technology

2007-08-01

strengthening of dough Lipases, Proteases Accelerates cheese ripening Glucose Isomerase Production of high fructose corn syrup Pectinases Fruit juice...Manufacture of glucose syrups and starch modification Maltogenic alpha-amylase Improves shelf life of bread Lipoxygenase Used for bleaching and...such as pH, ionic strength or temperature. In any approach to immobilize enzymes, the goal is to achieve a high specific activity without

Enhanced enzymatic hydrolysis of lignocellulose by optimizing enzyme complexes.

PubMed

Zhang, Mingjia; Su, Rongxin; Qi, Wei; He, Zhimin

2010-03-01

To enhance the conversion of the cellulose and hemicellulose, the corncob pretreated by aqueous ammonia soaking was hydrolyzed by enzyme complexes. The saturation limit for cellulase (Spezyme CP) was determined as 15 mg protein/g glucan (50 filter paper unit (FPU)/g glucan). The accessory enzymes (beta-glucosidase, xylanase, and pectinase) were supplemented to hydrolyze cellobiose (cellulase-inhibiting product), hemicellulose, and pectin (the component covering the fiber surfaces), respectively. It was found that beta-glucosidase (Novozyme 188) loading of 1.45 mg protein/g glucan [30 cellobiase units (CBU)/g glucan] was enough to eliminate the cellobiose inhibitor, and 2.9 mg protein/g glucan (60 CBU/g glucan) was the saturation limit. The supplementation of xylanase and pectinase can increase the conversion of cellulose and hemicellulose significantly. The yields of glucose and xylose enhanced with the increasing enzyme loading, but the increasing trend became low at high loading. Compared with xylanase, pectinase was more effective to promote the hydrolysis of cellulose and hemicellulose. The supplementation of pectinase with 0.12 mg protein/g glucan could increase the yields of glucose and xylose by 7.5% and 29.3%, respectively.

Effects of sorghum (Sorghum bicolor (L.) Moench) tannins on alpha-amylase activity and in vitro digestibility of starch in raw and processed flours

USDA-ARS?s Scientific Manuscript database

The effect of condensed tannins (CT) on in vitro starch digestibility in cooked, wholegrain sorghum flours and on corn starch was investigated. CT extracts were also tested for their inhibitory effect on alpha-amylases. Rapidly digestible starch, slowly digestible starch, and resistant starch were n...

Effect of exogenous xylanase, amylase, and protease as single or combined activities on nutrient digestibility and growth performance of broilers fed corn/soy diets

PubMed Central

Amerah, A. M.; Romero, L. F.; Awati, A.; Ravindran, V.

2017-01-01

Abstract Two trials (a 42-d performance and a 21-d cohort digestibility) were conducted to evaluate the performance and nutrient digestibility of broilers fed corn diets supplemented with exogenous xylanase, amylase, and protease as single or combined activities. A nutritionally adequate, positive control (PC) diet was formulated. The negative control (NC) diet was formulated to be lower in metabolizable energy (∼86 kcal/kg diet) and digestible amino acids (1 to 2%) compared to PC. The other 4 treatments were based on the NC and they were either supplemented with xylanase (X), amylase (A), protease (P), or a combination of X, A, and P (XAP; to provide 2,000 U of X, 200 U of A, and 4,000 U of P/kg diet). All diets were marginal in AvP and Ca and contained a background of phytase (1,000 FTU/kg). In each trial, male broiler (Ross 308) chicks were allocated to the 5 treatments (10 replicates of 20 birds/pen and 9 replicates of 8 birds/cage for the performance and digestibility trials, respectively). In the digestibility trial, ileal digesta was collected on d21 for the determination of nutrient utilization. Data were subjected to one-way ANOVA and means were separated by Tukey's HSD test. Only the XAP improved (P < 0.05) AMEn compared to NC. X, A or XAP improved (P < 0.05) N digestibility and apparent ileal digestible energy (AIDE). Both P and XAP improved N retention. The relative improvement in energy digestibility due to enzyme supplementation was greater at the ileal level than that measured in the excreta. The measured changes on AIDE due to supplemental enzymes were much higher than the sum of calculated contributions from starch, fat, and protein. Supplementation of all enzymes reduced (P < 0.05) ileal flow of soluble rhamnose and mannose relative to NC. In the performance trial, both X and XAP improved (P < 0.05) weight gain (WG) and only XAP improved (P < 0.05) FCR compared to NC during the starter phase (1-21d). Over the entire period (1–42d), WG

Further Experiments on Gibberellin-Stimulated Amylase Production in Cereal Grains

ERIC Educational Resources Information Center

Coppage, Jo; Hill, T. A.

1973-01-01

Experiments conducted on wheat and barley grains to analyze activities of alpha- and beta-amylase enzymes. Gibberellins were used exogenously. Techniques are described in detail. Results on different cultivars revealed that beta-amylase was not an invariable result of imbibition. Techniques employed can be used by school students. (PS)

Inhibitory effects of rosmarinic acid extracts on porcine pancreatic amylase in vitro.

PubMed

McCue, Patrick P; Shetty, Kalidas

2004-01-01

Porcine pancreatic alpha-amylase (PPA) was allowed to react with herbal extracts containing rosmarinic acid (RA) and purified RA. The derivatized enzyme-phytochemical mixtures obtained were characterized for residual amylase activity. These in vitro experiments showed that the amylase activity was inhibited in the presence of these phytochemicals. The extent of amylase inhibition correlated with increased concentration of RA. RA-containing oregano extracts yielded higher than expected amylase inhibition than similar amount of purified RA, suggesting that other phenolic compounds or phenolic synergies may contribute to additional amylase inhibitory activity. The significance of food-grade, plant-based amylase inhibitors for modulation of diabetes mellitus and other oxidation-linked diseases is hypothesized and discussed.

[POLYMORPHISM OF ALFA-AMYLASE AND CONJUGATION IN COMMON WHEAT ENZYME TYPES WITH QUANTITATIVE TRAITS OF PLANTS].

PubMed

Netsvetaev, V P; Bondarenko, L S; Motorina, I P

2015-01-01

Using polymorphism of alpha-amylase in the winter common wheat studied inheritance isoenzymes and its conjugation enzyme types with germinating grain on the "vine", grain productivity, plant height and time of ear formation. It is shown that the polymorphism isoenzyme of alpha-amylase wheat is limited by the presence of different loci whose products are similar in electrophoretic parameters. In this regard, one component of the enzyme can be controlling at one or two or three genes. Identification of a locus controlling alpha-amylase isoenzyme in the fast moving part of the electrophoretogram, designated as α-Amy-B7. Determine the distance of the locus to factor α-Amy-B6.

Hyperexpression of the gene for a Bacillus alpha-amylase in Bacillus subtilis cells: enzymatic properties and crystallization of the recombinant enzyme.

PubMed

Ikawa, K; Araki, H; Tsujino, Y; Hayashi, Y; Igarashi, K; Hatada, Y; Hagihara, H; Ozawa, T; Ozaki, K; Kobayashi, T; Ito, S

1998-09-01

We have constructed a new excretion vector, pHSP64, to develop a hyperexcretion system for Bacillus subtilis [Sumitomo et al., Biosci. Biotech. Biochem., 59, 2172-2175 (1995)]. The structural gene for a novel liquefying semi-alkaline alpha-amylase from the alkaliphilic Bacillus sp. KSM-1378 was amplified by PCR. It was cloned into a SalI-SmaI site of pHSP64 and the recombinant plasmid obtained was introduced into B. subtilis. The transformed B. subtilis hyperproduced the alpha-amylase activity extracellularly, corresponding to approximately 1.0 g (5 x 10(6) units) per liter of an optimized liquid culture. The recombinant enzyme was purified to homogeneity by a simple purification procedure with very high yield. No significant differences in physiochemical and catalytic properties were observed between the recombinant enzyme and the native enzyme produced by Bacillus sp. KSM-1378. The enzymatic properties of the recombinant enzyme were further examined with respect to the responses to various metal ions. The recombinant enzyme could easily be crystallized at room temperature within one day in a buffered solution of 10% (w/v) ammonium sulfate (pH 6.5).

Mold Pectinase Modified with Dialdehyde Derivatives of Dextran and Cellulose.

PubMed

Kobayashi, M; Chiba, Y; Funane, K; Ohya, S; Kato, Y

1996-01-01

Chemical modification of mold pectinase with dextran- and cellulose-dialdehydes was examined to improve the enzyme characteristics. The modified pectinase with dextran-dialdehyde retained about 50% of the original activity, and more than 80% of the total amino groups were modified. HPLC gel filtration analysis showed an increase in molecular weight of the reaction product. Reaction with cellulose-dialdehyde provided an immobilized form of pectinase. The immobilized pectinase was resistant to both acidic and alkaline pHs, and also acquired heat stability at 60°C. The optimum pH of the modified enzyme shifted from pH 4.5 to 5.0-5.5, and this enzyme had higher activity at neutral pH regions than the native enzyme. A rather low recovery of immobilized enzyme (14.5%) should be improved by the combination with various methods hitherto established.

Preparation, characterization and catalytic behavior of pectinase covalently immobilized onto sodium alginate/graphene oxide composite beads.

PubMed

Dai, Xiao-Yan; Kong, Li-Min; Wang, Xiao-Ling; Zhu, Qing; Chen, Kai; Zhou, Tao

2018-07-01

Pectinase was immobilized onto sodium alginate/graphene oxide beads via amide bonds by using N,N'-dicyclohexylcarbodiimide/N-hydroxysuccinimide as the activating agent. The immobilized pectinase was characterized by Fourier transform infrared spectra and scanning electron microscopy analyses. Immobilization conditions were optimized by Box-Behnken design and the response surface method. The activity of the immobilized pectinase prepared under optimal conditions reached 1236.86 ± 40.21 U/g, with an enzyme activity recovery of 83.5%. The optimal pH of free pectinase was 4.5, while that of immobilized pectinase was shifted to 4.0. The optimal temperature of immobilized pectinase was increased to 60 °C, which was 10 °C higher than that of free form. Furthermore, the immobilized pectinase possessed a superior thermal stability and storage stability to those of free pectinase. Reusability studies indicated that the immobilized pectinase retained 73% of initial activity after six times cycles. Due to these good properties, such immobilized pectinase may find application in food industry. Copyright © 2018 Elsevier Ltd. All rights reserved.

Improved α-amylase production by Aspergillus oryzae after a double deletion of genes involved in carbon catabolite repression.

PubMed

Ichinose, Sakurako; Tanaka, Mizuki; Shintani, Takahiro; Gomi, Katsuya

2014-01-01

In filamentous fungi, the expression of secretory glycoside hydrolase encoding genes, such as those for amylases, cellulases, and xylanases, is generally repressed in the presence of glucose. CreA and CreB have been observed to be regulating factors for carbon catabolite repression. In this study, we generated single and double deletion creA and/or creB mutants in Aspergillus oryzae. The α-amylase activities of each strain were compared under various culture conditions. For the wild-type strain, mRNA levels of α-amylase were markedly decreased in the later stage of submerged culture under inducing conditions, whereas this reduced expression was not observed for single creA and double creA/creB deletion mutants. In addition, α-amylase activity of the wild-type strain was reduced in submerged culture containing high concentrations of inducing sugars, whereas all constructed mutants showed higher α-amylase activities. In particular, the α-amylase activity of the double deletion mutant in a medium containing 5% starch was >10-fold higher than that of the wild-type strain under the same culture conditions. In solid-state cultures using wheat bran as a substrate, the α-amylase activities of single creA and double deletion mutants were >2-fold higher than that of the wild-type strain. These results suggested that deleting both creA and creB resulted in dramatic improvements in the production of secretory glycoside hydrolases in filamentous fungi.

Kinetic studies of amylase and biomass production by Calvatia gigantea

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

Kekos, D.; Macris, B.J.

1987-01-01

Production of alpha-amylase (alpha-4, glucan 4-glucanohydrolase, EC 3.2.1.1) by microorganisms has been practiced for many years in small and large scale operations and the literature on this enzyme is voluminous. Aspergillus niger and Aspergillus oryzae have been reported as the main fungal species used for commercial production of the enzyme. On the other hand, large volumes of low-cost agricultural products such as acorn (the perisperm-free dry seed contains approximately 60% starch) are wasted in many countries and provide a challenge to biotechnology to efficiently utilize these rich sources of starch for the production of high added value products like enzymes.more » C. gigantea is an edible puffball excreting high levels of alpha-amylase when cultivated on different sources of starch containing elevated quantities of toxic tannic compounds. This fungus has been employed for the production of microbial protein from wastes and acorns containing high levels of toxic tannic compounds. The same fungus was also reported to grow on both hydrolyzable and condensed tannins as sole carbon sources. The present work was undertaken to investigate certain kinetic characteristics of alpha-amylase and biomass production by C. gigantea grown on soluble and acorn starch in a lab fermenter. (Refs. 18).« less

Engineering Thermostable Microbial Xylanases Toward its Industrial Applications.

PubMed

Kumar, Vishal; Dangi, Arun Kumar; Shukla, Pratyoosh

2018-03-01

Xylanases are one of the important hydrolytic enzymes which hydrolyze the β-1, 4 xylosidic linkage of the backbone of the xylan polymeric chain which consists of xylose subunits. Xylanases are mainly found in plant cell walls and are produced by several kinds of microorganisms such as fungi, bacteria, yeast, and some protozoans. The fungi are considered as most potent xylanase producers than that of yeast and bacteria. There is a broad series of industrial applications for the thermostable xylanase as an industrial enzyme. Thermostable xylanases have been used in a number of industries such as paper and pulp industry, biofuel industry, food and feed industry, textile industry, etc. The present review explores xylanase-substrate interactions using gene-editing tools toward the comprehension in improvement in industrial stability of xylanases. The various protein-engineering and metabolic-engineering methods have also been explored to improve operational stability of xylanase. Thermostable xylanases have also been used for improvement in animal feed nutritional value. Furthermore, they have been used directly in bakery and breweries, including a major use in paper and pulp industry as a biobleaching agent. This present review envisages some of such applications of thermostable xylanases for their bioengineering.

Effect of exogenous xylanase, amylase, and protease as single or combined activities on nutrient digestibility and growth performance of broilers fed corn/soy diets.

PubMed

Amerah, A M; Romero, L F; Awati, A; Ravindran, V

2017-04-01

Two trials (a 42-d performance and a 21-d cohort digestibility) were conducted to evaluate the performance and nutrient digestibility of broilers fed corn diets supplemented with exogenous xylanase, amylase, and protease as single or combined activities. A nutritionally adequate, positive control (PC) diet was formulated. The negative control (NC) diet was formulated to be lower in metabolizable energy (∼86 kcal/kg diet) and digestible amino acids (1 to 2%) compared to PC. The other 4 treatments were based on the NC and they were either supplemented with xylanase (X), amylase (A), protease (P), or a combination of X, A, and P (XAP; to provide 2,000 U of X, 200 U of A, and 4,000 U of P/kg diet). All diets were marginal in AvP and Ca and contained a background of phytase (1,000 FTU/kg). In each trial, male broiler (Ross 308) chicks were allocated to the 5 treatments (10 replicates of 20 birds/pen and 9 replicates of 8 birds/cage for the performance and digestibility trials, respectively). In the digestibility trial, ileal digesta was collected on d21 for the determination of nutrient utilization. Data were subjected to one-way ANOVA and means were separated by Tukey's HSD test. Only the XAP improved (P < 0.05) AMEn compared to NC. X, A or XAP improved (P < 0.05) N digestibility and apparent ileal digestible energy (AIDE). Both P and XAP improved N retention. The relative improvement in energy digestibility due to enzyme supplementation was greater at the ileal level than that measured in the excreta. The measured changes on AIDE due to supplemental enzymes were much higher than the sum of calculated contributions from starch, fat, and protein. Supplementation of all enzymes reduced (P < 0.05) ileal flow of soluble rhamnose and mannose relative to NC. In the performance trial, both X and XAP improved (P < 0.05) weight gain (WG) and only XAP improved (P < 0.05) FCR compared to NC during the starter phase (1-21d). Over the entire period (1-42d), WG and FI

Characteristics of pectinase treated with ultrasound both during and after the immobilization process.

PubMed

Ma, Xiaobin; Wang, Danli; Yin, Michelle; Lucente, Juliet; Wang, Wenjun; Ding, Tian; Ye, Xingqian; Liu, Donghong

2017-05-01

In this study, ultrasound was applied both during and after the immobilization process and characteristics of different immobilized pectinase samples were studied. When introduced during the immobilization process, ultrasound at an intensity of 9WmL -1 for 20min increased the immobilization yield 92.28% more than the control. When introduced to the already immobilized pectinase, ultrasound at an intensity of 4.5WmL -1 for 10min increased the pectinase activity by 30.05%. Results of scanning electron microscope demonstrated that ultrasound increased surface area and loosened structures of immobilized enzymes. Higher V max and lower K m were obtained after ultrasound treatment, indicating the increased catalytic efficiency and enhanced affinity of immobilized pectinase. Furthermore, the optimum temperature and pH for free and immobilized pectinase remained unchanged at 50°C and pH 4. Thermostability, reaction stability and reusability of two ultrasound-treated pectinase enzymes slightly decreased due to structural matrix changes. Copyright © 2016 Elsevier B.V. All rights reserved.

Extremozymes: A Potential Source for Industrial Applications.

PubMed

Dumorné, Kelly; Córdova, David Camacho; Astorga-Eló, Marcia; Renganathan, Prabhaharan

2017-04-28

Extremophilic microorganisms have established a diversity of molecular strategies in order to survive in extreme conditions. Biocatalysts isolated by these organisms are termed extremozymes, and possess extraordinary properties of salt allowance, thermostability, and cold adaptivity. Extremozymes are very resistant to extreme conditions owing to their great solidity, and they pose new opportunities for biocatalysis and biotransformations, as well as for the development of the economy and new line of research, through their application. Thermophilic proteins, piezophilic proteins, acidophilic proteins, and halophilic proteins have been studied during the last few years. Amylases, proteases, lipases, pullulanases, cellulases, chitinases, xylanases, pectinases, isomerases, esterases, and dehydrogenases have great potential application for biotechnology, such as in agricultural, chemical, biomedical, and biotechnological processes. The study of extremozymes and their main applications have emerged during recent years.

Salivary Alpha-Amylase Activity Levels in Catatonic Schizophrenia Decrease after Electroconvulsive Therapy.

PubMed

Kanayama, Misako; Miyaoka, Tsuyoshi; Araki, Tomoko; Hayashida, Maiko; Hashioka, Sadayuki; Horiguchi, Jun

2018-01-01

Dysfunction of the autonomic nervous system (ANS) in schizophrenia has been detected by electrophysiological methods, but the underlying mechanisms remain unknown. Several studies have suggested that measuring salivary alpha-amylase activity levels is useful for evaluating the ANS activity and that sAA levels increase in schizophrenia and correlate with Brief Psychiatric Rating Scale (BPRS) scores. However, no study has examined the relationship between sAA activity levels and symptoms of schizophrenia with catatonic state. We present the case of a 59-year-old female with persistent catatonic schizophrenia treated by electroconvulsive therapy. We evaluated the ANS activity by measuring sAA activity levels before and after ECT, and we evaluated her symptoms using the BPRS and Bush-Francis Catatonia Rating Scale (BFCRS). ECT was highly effective and BPRS and BFCRS scores substantially decreased. sAA activity levels decreased from 125 kU/l to 33 kU/l. sAA activity levels could be a potential biomarker of schizophrenia with catatonic state.

Effects of early life adversity on cortisol/salivary alpha-amylase symmetry in free-ranging juvenile rhesus macaques.

PubMed

Petrullo, Lauren A; Mandalaywala, Tara M; Parker, Karen J; Maestripieri, Dario; Higham, James P

2016-11-01

Early life adversity (ELA) affects physiological and behavioral development. One key component is the relationship between the developing Hypothalamic-Pituitary-Adrenal (HPA) axis and the Sympathetic Nervous System (SNS). Recent studies suggest a relationship between early life adversity and asymmetry in cortisol (a measure of HPA activation) and salivary alpha-amylase (sAA: a correlate of SNS activation) responses to stress among human children, but to our knowledge there have been no comparable studies in nonhumans. Here, we investigate the responses of these two analytes in "low stress" and "high stress" situations in free-ranging juvenile rhesus macaques (Macaca mulatta) on Cayo Santiago, Puerto Rico. Behavioral data on maternal maltreatment were collected during the first 3months of life to determine individual rates of ELA, and saliva samples were collected from subjects noninvasively during juvenility. Irrespective of ELA, salivary alpha-amylase levels were lower in low stress situations and higher in high stress situations. For cortisol however, high ELA subjects exhibited higher low stress concentrations and blunted acute responses during high stress situations compared to moderate and low ELA subjects. Cortisol and sAA values were positively correlated among low ELA subjects, suggesting symmetry, but were uncorrelated or negatively correlated among moderate and high ELA subjects, suggesting asymmetry in these individuals. These findings indicate dysregulation of the stress response among juveniles maltreated during infancy: specifically, attenuated cortisol reactivity coupled with typical sAA reactivity characterize the stress response profiles of juveniles exposed to higher rates of ELA during the first 3months of life. Copyright © 2016 Elsevier Inc. All rights reserved.

General Subject 2. Report to ICUMSA on the determination of carry-over alpha-amylase activity in white and refined sugars by a spectrophotometric method

USDA-ARS?s Scientific Manuscript database

A report is given on a new industrial method for the determination of carry-over alpha-amylase activity in raw and refined sugars, as well as a recommendation. In recent years, there has been increased concern over carry-over activity of mostly high temperature (HT) and very high temperature (VHT) s...

Characterization of the Activity and Stability of Amylase from Saliva and Detergent: Laboratory Practicals for Studying the Activity and Stability of Amylase from Saliva and Various Commercial Detergents

ERIC Educational Resources Information Center

Valls, Cristina; Rojas, Cristina; Pujadas, Gerard; Garcia-Vallve, Santi; Mulero, Miquel

2012-01-01

This article presents two integrated laboratory exercises intended to show students the role of [alpha]-amylases (AAMYs) in saliva and detergents. These laboratory practicals are based on the determination of the enzymatic activity of amylase from saliva and different detergents using the Phadebas test (quantitative) and the Lugol test…

Pectinase activity of Nectria coccinea (Pers ex Fries) fries in relation to beech bark disease

Treesearch

R. Perrin

1983-01-01

The pectinase activity of Nectria coccinea was studied in vitro and in vivo in relation to the pectinases of Cryptococcus fagisuga and the nature of the bark. Any pectinases necessary for degradation of pectic material were secreted by the fungus in vitro. Some pectinases produced by the insect are of great significance in the...

Economic Analysis of the Production of Amylases and Other Hydrolases by Aspergillus awamori in Solid-State Fermentation of Babassu Cake

PubMed Central

de Castro, Aline Machado; Carvalho, Daniele Fernandes; Freire, Denise Maria Guimarães; Castilho, Leda dos Reis

2010-01-01

Amylases are one of the most important industrial enzymes produced worldwide, with their major application being in ethanol manufacturing. This work investigated the production of amylases by solid-state fermentation of babassu cake, using the filamentous fungus Aspergillus awamori IOC-3914. Lab-scale experiments were carried out to generate input data for simulations of an industrial plant for amylase production. Additionally to the target enzymes, other hydrolases (cellulases, xylanases, and proteases) were also produced, enriching the final product. The most suitable fermentation time was 144 hours, when exoamylase and endoamylase activities of 40.5 and 42.7 U g−1 were achieved, respectively. A first evaluation showed a large impact of the inoculum propagation medium on production costs. Therefore, five propagation media were compared, and PDA medium presented the best cost-benefit ratio. The credits obtained from sales of fermented cake as a coproduct enabled a significant decrease in the production cost of the enzyme product, down to 10.40 USD kg−1. PMID:21048867

Enhancement of Penicillium echinulatum glycoside hydrolase enzyme complex.

PubMed

dos Santos Costa, Patrícia; Büchli, Fernanda; Robl, Diogo; Delabona, Priscila da Silva; Rabelo, Sarita Candida; Pradella, José Geraldo da Cruz

2016-05-01

The enhancement of enzyme complex produced by Penicillium echinulatum grown in several culture media components (bagasse sugarcane pretreated by various methods, soybean meal, wheat bran, sucrose, and yeast extract) was studied to increment FPase, xylanase, pectinase, and β-glucosidase enzyme activities. The present results indicated that culture media composed with 10 g/L of the various bagasse pretreatment methods did not have any substantial influence with respect to the FPase, xylanase, and β-glucosidase attained maximum values of, respectively, 2.68 FPU/mL, 2.04, and 115.4 IU/mL. On the other hand, proposed culture media to enhance β-glucosidase production composed of 10 g/L steam-exploded bagasse supplemented with soybean flour 5.0 g/L, yeast extract 1.0 g/L, and sucrose 10.0 g/L attained, respectively, 3.19 FPU/mL and 3.06 IU/mL while xylanase was maintained at the same level. The proteomes obtained from the optimized culture media for enhanced FPase, xylanase, pectinase, and β-glucosidase production were analyzed using mass spectrometry and a panel of GH enzyme activities against 16 different substrates. Culture medium designed to enhance β-glucosidase activity achieved higher enzymatic activities values (13 measured activities), compared to the culture media for FPase/pectinase (9 measured activities) and xylanase (7 measured activities), when tested against the 16 substrates. Mass spectrometry analyses of secretome showed a consistent result and the greatest number of spectral counts of Cazy family enzymes was found in designed β-glucosidase culture medium, followed by FPase/pectinase and xylanase. Most of the Cazy identified protein was cellobiohydrolase (GH6 and GH7), endoglucanase (GH5), and endo-1,4-β-xylanase (GH10). Enzymatic hydrolysis of hydrothermally pretreated sugarcane bagasse performed with β-glucosidase enhanced cocktail achieved 51.4 % glucose yield with 10 % w/v insoluble solids at enzyme load of 15 FPU/g material. Collectively the

Optimization of Freeze Drying Conditions for Purified Pectinase from Mango (Mangifera indica cv. Chokanan) Peel

PubMed Central

Mehrnoush, Amid; Mustafa, Shuhaimi; Yazid, Abdul Manap Mohd

2012-01-01

Response surface methodology (RSM) along with central composite design (CCD) was applied to optimize the freeze drying conditions for purified pectinase from mango (Mangifera indica cv. Chokanan) peel. The effect of pectinase content (−2.66, 62.66 mg/mL), Arabic gum (−1.21, 10.21%, w/v), and maltodextrin (0.73, 7.26%, w/v) as independent variables on activity, yield, and storage stability of freeze-dried enzyme was evaluated. Storage stability of pectinase was investigated after one week at 4 °C and yield percentage of the enzyme after encapsulation was also determined. The independent variables had the most significant (p < 0.05) effect on pectinase activity and yield of the enzyme. It was observed that the interaction effect of Arabic gum and maltodextrin improved the enzymatic properties of freeze-dried pectinase. The optimal conditions for freeze-dried pectinase from mango peel were obtained using 30 mg/mL of pectinase content, 4.5 (%, w/v) of Arabic gum, and 4 (%, w/v) of maltodextrin. Under these conditions, the maximum activity (11.12 U/mL), yield (86.4%) and storage stability (84.2%) of encapsulated pectinase were achieved. PMID:22489134

Optimization of freeze drying conditions for purified pectinase from mango (Mangifera indica cv. Chokanan) peel.

PubMed

Mehrnoush, Amid; Mustafa, Shuhaimi; Yazid, Abdul Manap Mohd

2012-01-01

Response surface methodology (RSM) along with central composite design (CCD) was applied to optimize the freeze drying conditions for purified pectinase from mango (Mangifera indica cv. Chokanan) peel. The effect of pectinase content (-2.66, 62.66 mg/mL), Arabic gum (-1.21, 10.21%, w/v), and maltodextrin (0.73, 7.26%, w/v) as independent variables on activity, yield, and storage stability of freeze-dried enzyme was evaluated. Storage stability of pectinase was investigated after one week at 4 °C and yield percentage of the enzyme after encapsulation was also determined. The independent variables had the most significant (p < 0.05) effect on pectinase activity and yield of the enzyme. It was observed that the interaction effect of Arabic gum and maltodextrin improved the enzymatic properties of freeze-dried pectinase. The optimal conditions for freeze-dried pectinase from mango peel were obtained using 30 mg/mL of pectinase content, 4.5 (%, w/v) of Arabic gum, and 4 (%, w/v) of maltodextrin. Under these conditions, the maximum activity (11.12 U/mL), yield (86.4%) and storage stability (84.2%) of encapsulated pectinase were achieved.

Evening salivary cortisol and alpha-amylase at 14months and neurodevelopment at 4years: Sex differences.

PubMed

Andiarena, Ainara; Balluerka, Nekane; Murcia, Mario; Ibarluzea, Jesús; Glover, Vivette; Vegas, Oscar

2017-08-01

Stress system activity in early life can have long-term effects on neurodevelopment. The main aim of this study was to assess the association of child evening salivary cortisol and alpha-amylase basal levels at 14months of age with longer-term neuropsychological development at 4years in a low-risk population-based birth cohort derived from the INMA (Environment and Childhood) project in Spain. We included 186 parent-children pairs with information on both stress system activity and neurodevelopment. Both stress markers at 14months of age showed an association with neuropsychological development at 4years. Salivary cortisol showed a sex-specific pattern of association. In girls, cortisol levels at 14months were negatively associated with cognitive development [long-term declarative memory (β=-17.8, p=0.028; 95% CI=-33.2 to -2.5); executive function (β=-9.8, p=0.08; 95% CI=-21 to 1)] and gross motor development (β=-13; p=0.022; 95% CI=-24 to -2), whereas in boys cortisol levels were negatively associated with socioemotional development [autistic-like behaviours: Incidence Rate Ratio (IRR)=1.6, p=0.039; 95% CI=1.01 to 2.41]. Salivary alpha-amylase was positively associated with socioemotional development in boys only [social competence (β=2.11, p=0.013; 95% CI=0.47 to 3.72), autistic-like behaviours (IRR=0.93, p=0.042; 95% CI=0.87 to 0.99) and hyperactivity symptoms (IRR=0.81, p=0.021; 95% CI=0.69 to 0.97)]. These results suggest that stress system activity in early life is associated with longer-term neurodevelopment and that sex is an important factor in this relationship. Copyright © 2017 Elsevier Inc. All rights reserved.

Salivary Cortisol, Salivary Alpha Amylase, and the Dental Anxiety Scale

PubMed Central

Sadi, Hana; Finkelman, Matthew; Rosenberg, Morton

2013-01-01

The aim of this study was to investigate the correlation between dental anxiety, salivary cortisol, and salivary alpha amylase (sAA) levels. Furthermore, the aim was to look into individual differences such as age, race, gender, any existing pain, or traumatic dental experience and their effect on dental anxiety. This study followed a cross-sectional design and included a convenience sample of 46. Every patient was asked to complete the Dental Anxiety Scale (DAS) and a basic demographic/dental history questionnaire. A saliva sample, utilizing the method of passive drooling, was then collected in 2-mL cryovials. Samples were analyzed for salivary cortisol and sAA levels by Salimetrics. Significant associations were observed between DAS scores and presence of pain and history of traumatic dental experience. However, no significant correlations were observed between DAS, cortisol, and sAA levels. Our study reconfirms that dental anxiety is associated with presence of pain and a history of traumatic dental experience. On the other hand, our study was the first to our knowledge to test the correlation between the DAS and sAA; nevertheless, our results failed to show any significant correlation between dental anxiety, cortisol, and sAA levels. PMID:23763559

Self-compassionate young adults show lower salivary alpha-amylase responses to repeated psychosocial stress.

PubMed

Breines, Juliana G; McInnis, Christine M; Kuras, Yuliya I; Thoma, Myriam V; Gianferante, Danielle; Hanlin, Luke; Chen, Xuejie; Rohleder, Nicolas

2015-10-01

In this study we tested the hypothesis that participants higher in dispositional self-compassion would show lower stress-induced reactivity of salivary alpha-amylase (sAA), a marker of sympathetic nervous system activation. Thirty-three healthy participants (18-34 years old) were exposed to a standardized laboratory stressor on two consecutive days. Self-compassion, self-esteem, and demographic factors were assessed by questionnaire and sAA was assessed at baseline and at 1, 10, 30, and 60 minutes following each stressor. Self-compassion was a significant negative predictor of sAA responses on both days. This relationship remained significant when controlling for self-esteem, subjective distress, age, gender, ethnicity, and Body Mass Index (BMI). These results suggest that self-compassion may serve as a protective factor against stress-induced physiological changes that have implications for health.

Self-compassionate young adults show lower salivary alpha-amylase responses to repeated psychosocial stress

PubMed Central

Breines, Juliana G.; McInnis, Christine M.; Kuras, Yuliya I.; Thoma, Myriam V.; Gianferante, Danielle; Hanlin, Luke; Chen, Xuejie; Rohleder, Nicolas

2015-01-01

In this study we tested the hypothesis that participants higher in dispositional self-compassion would show lower stress-induced reactivity of salivary alpha-amylase (sAA), a marker of sympathetic nervous system activation. Thirty-three healthy participants (18–34 years old) were exposed to a standardized laboratory stressor on two consecutive days. Self-compassion, self-esteem, and demographic factors were assessed by questionnaire and sAA was assessed at baseline and at 1, 10, 30, and 60 minutes following each stressor. Self-compassion was a significant negative predictor of sAA responses on both days. This relationship remained significant when controlling for self-esteem, subjective distress, age, gender, ethnicity, and Body Mass Index (BMI). These results suggest that self-compassion may serve as a protective factor against stress-induced physiological changes that have implications for health. PMID:26005394

Purification and Properties of Acid Stable Xylanases from Aspergillus kawachii.

PubMed

Ito, K; Ogasawara, H; Sugimoto, T; Ishikawa, T

1992-01-01

Five extracellular endo-xylanases were recognized in the culture broth of shochu koji mold (Aspergillus kawachii, IFO 4308), and three major xylanases (XylA, XylB, and XylC) were purified and characterized. The molecular masses of XylA, XylB, and XylC were 35,000, 26,000, and 29,000, and isoelectric points were pH 6.7, 4.4, and 3.5, respectively. Amino acid compositions and other properties were studied and these three xylanases were found to be greatly different in their properties. These three xylanases, XylA, XylB, and XylC, were stable between pH 3-10, 3-10, and 1-9 and the optimum pHs were 5.5, 4.5, and 2.0, respectively. Consequently, these xylanases were acid stable xylanases, especially XylC was an acidophilic xylanase (acid xylanase). These xylanases produced various xylooligosaccharides including xylose from xylan and the main product was xylobiose in all xylanases. The production of acid xylanase (XylC) was enhanced with a low initial pH of the medium.

[Growth inhibition effect of immobilized pectinase on Microcystis aeruginosa].

PubMed

Shen, Qing-Qing; Peng, Qian; Lai, Yong-Hong; Ji, Kai-Yan; Han, Xiu-Lin

2012-12-01

To confirm the growth inhibition effect of immobilized pectinase on algae, co-cultivation method was used to investigate the effect of immobilized pectinase on the growth of Microcystis aeruginosa. After co-cultivation, the damage status of the algae was observed through electron microscope, and the effect of immobilized pectase on the physiological and biochemical characteristics of the algae was also measured. The results showed that the algae and immobilized pectase co-cultivated solution etiolated distinctly on the third day and there was a significantly positive correlation between the extent of etiolation and the dosage as well as the treating time of the immobilized pectinase. Under electron microscope, plasmolysis was found in the slightly damaged cells, and the cell surface of these cells was rough, uneven and irregular; the severely damaged cells were collapsed or disintegrated completely. The algal yield and the chlorophyll a content decreased significantly with the increase of the treating time. The measurement of the malondiadehyde (MDA) value showed that the antioxidation system of the treated algal cells was destroyed, and their membrane lipid was severely peroxidated. The study indicated that the immobilized pectinase could efficiently inhibit the growth of M. aeruginosa, and the inhibitory rate reached up to 96%.

Alpha-amylase serum levels in professional soccer players are not related with physical fitness.

PubMed

Sanchis-Gomar, Fabian; Alis, Rafael; Rampinini, Ermanno; Bosio, Andrea; Romagnoli, Marco; Lombardi, Giovanni; Lippi, Giuseppe

2017-03-01

Recent evidence has showed that serum or salivary values of α-amylase predict endurance running performance. In this study we investigate whether serum α-amylase concentration may be associated with training status during a competitive season and after a detraining period in professional soccer players. The study population consisted in 15 male professional soccer players from an Italian major league team (age [mean±SD] 27±5 years, weight 76.9±4.1 kg, height 1.82±0.05 m). Serum α-amylase levels were measured 3 times during the last part of a competitive season (January, March and May) and just before preseason training (July). Metabolic and cardiovascular fitness of soccer players was improved during the last part of the season. The levels of α-amylase did not change significantly throughout the study period (χ2=7.331, P=0.062), nor they were found to be associated with variation of physical fitness and training status. The α-amylase fluctuations throughout a competitive season and after vacation time were meaningless in professional soccer players. No significant associations with physical fitness variations could be observed. These results suggest that α-amylase concentration may be a useful parameter for identifying individual inclination to endurance exercise, but not for predicting actual training status.

Induction of Pectinase Hyper Production by Multistep Mutagenesis Using a Fungal Isolate--Aspergillus flavipes.

PubMed

Akbar, Sabika; Prasuna, R Gyana; Khanam, Rasheeda

2014-04-01

Aspergillus flavipes, a slow growing pectinase producing ascomycete, was isolated from soil identified and characterised in the previously done preliminary studies. Optimisation studies revealed that Citrus peel--groundnut oil cake [CG] production media is the best media for production of high levels of pectinase up to 39 U/ml using wild strain of A. flavipes. Strain improvement of this isolated strain for enhancement of pectinase production using multistep mutagenesis procedure is the endeavour of this project. For this, the wild strain of A. flavipes was treated with both physical (UV irradiation) and chemical [Colchicine, Ethidium bromide, H2O2] mutagens to obtain Ist generation mutants. The obtained mutants were assayed and differentiated basing on pectinase productivity. The better pectinase producing strains were further subjected to multistep mutagenesis to attain stability in mutants. The goal of this project was achieved by obtaining the best pectinase secreting mutant, UV80 of 45 U/ml compared to wild strain and sister mutants. This fact was confirmed by quantitatively analysing 3rd generation mutants obtained after multistep mutagenesis.

Immobilization of pectinase on silica-based supports: Impacts of particle size and spacer arm on the activity.

PubMed

Alagöz, Dilek; Tükel, S Seyhan; Yildirim, Deniz

2016-06-01

The pectinase was separately immobilized onto Florisil and nano silica supports through both glutaraldehyde and 3-glyoxypropyltrietoxysilane spacer arms. The effects of spacer arm, particle size of support and ionic liquids on the activities of pectinase preparations were investigated. The immobilization of pectinase onto Florisil and nano silica through 3-glyoxypropyltrietoxysilane spacer arm completely led to inactivation of enzyme; however, 10 and 75% pectinase activity were retained when it was immobilized through glutaraldehyde spacer arm onto Florisil and nano silica, respectively. The pectinase immobilized onto nano silica through glutaraldehyde spacer arm showed 6.3-fold higher catalytic efficiency than that of the pectinase immobilized onto Florisil through same spacer arm. A 2.3-fold increase in thermal stability of pectinase was provided upon immobilization onto nano silica at 35°C. The effects of IL/buffer mixture and volume ratio of IL/buffer mixture on the catalytic activities of free and immobilized pectinase preparations were also tested. All the pectinase preparations showed highest activity in 10% (v/v) 1-butyl-3-methylimidazolium hexafluorophosphate containing medium and their activities significantly affected from the concentration of 1-butyl-3-methylimidazolium hexafluorophosphate. Copyright © 2016 Elsevier B.V. All rights reserved.

Thermophilic xylanases: from bench to bottle.

PubMed

Basit, Abdul; Liu, Junquan; Rahim, Kashif; Jiang, Wei; Lou, Huiqiang

2018-01-17

Lignocellulosic biomass is a valuable raw material. As technology has evolved, industrial interest in new ways to take advantage of this raw material has grown. Biomass is treated with different microbial cells or enzymes under ideal industrial conditions to produce the desired products. Xylanases are the key enzymes that degrade the xylosidic linkages in the xylan backbone of the biomass, and commercial enzymes are categorized into different glycoside hydrolase families. Thermophilic microorganisms are excellent sources of industrially relevant thermostable enzymes that can withstand the harsh conditions of industrial processing. Thermostable xylanases display high-specific activity at elevated temperatures and distinguish themselves in biochemical properties, structures, and modes of action from their mesophilic counterparts. Natural xylanases can be further improved through genetic engineering. Rapid progress with genome editing, writing, and synthetic biological techniques have provided unlimited potential to produce thermophilic xylanases in their natural hosts or cell factories including bacteria, yeasts, and filamentous fungi. This review will discuss the biotechnological potential of xylanases from thermophilic microorganisms and the ways they are being optimized and produced for various industrial applications.

Microbial xylanases: engineering, production and industrial applications.

PubMed

Juturu, Veeresh; Wu, Jin Chuan

2012-01-01

Enzymatic depolymerization of hemicellulose to monomer sugars needs the synergistic action of multiple enzymes, among them endo-xylanases (EC 3.2.1.8) and β-xylosidases (EC 3.2.1.37) (collectively xylanases) play a vital role in depolymerizing xylan, the major component of hemicellulose. Recent developments in recombinant protein engineering have paved the way for engineering and expressing xylanases in both heterologous and homologous hosts. Functional expression of endo-xylanases has been successful in many hosts including bacteria, yeasts, fungi and plants with yeasts being the most promising expression systems. Functional expression of β-xylosidases is more challenging possibly due to their more complicated structures. The structures of endo-xylanases of glycoside hydrolase families 10 and 11 have been well elucidated. Family F/10 endo-xylanases are composed of a cellulose-binding domain and a catalytic domain connected by a linker peptide with a (β/α)8 fold TIM barrel. Family G/11 endo-xylanases have a β-jelly roll structure and are thought to be able to pass through the pores of hemicellulose network owing to their smaller molecular sizes. The structure of a β-D-xylosidase belonging to family 39 glycoside hydrolase has been elucidated as a tetramer with each monomer being composed of three distinct regions: a catalytic domain of the canonical (β/α)8--TIM barrel fold, a β-sandwich domain and a small α-helical domain with the enzyme active site that binds to D-xylooligomers being present on the upper side of the barrel. Glycosylation is generally considered as one of the most important post-translational modifications of xylanases, but a few examples showed functional expression of eukaryotic xylanases in bacteria. The optimal ratio of these synergistic enzymes is very important in improving hydrolysis efficiency and reducing enzyme dosage but has hardly been addressed in literature. Xylanases have been used in traditional fields such as food, feed

Optimization of extraction of novel pectinase enzyme discovered in red pitaya (Hylocereus polyrhizus) peel.

PubMed

Zohdi, Nor Khanani; Amid, Mehrnoush

2013-11-20

Plant peels could be a potential source of novel pectinases for use in various industrial applications due to their broad substrate specificity with high stability under extreme conditions. Therefore, the extraction conditions of a novel pectinase enzyme from pitaya peel was optimized in this study. The effect of extraction variables, namely buffer to sample ratio (2:1 to 8:1, X₁), extraction temperature (-15 to +25 °C, X₂) and buffer pH (4.0 to 12.0, X₃) on specific activity, temperature stability, storage stability and surfactant agent stability of pectinase from pitaya peel was investigated. The study demonstrated that the optimum conditions for the extraction of pectinase from pitaya sources could improve the enzymatic characteristics of the enzyme and protect its activity and stability during the extraction procedure. The optimum extraction conditions cause the pectinase to achieve high specific activity (15.31 U/mg), temperature stability (78%), storage stability (88%) and surfactant agent stability (83%). The most desirable conditions to achieve the highest activity and stability of pectinase enzyme from pitaya peel were the use of 5:1 buffer to sample ratio at 5 °C and pH 8.0.

Effects of exposure to GSM mobile phone base station signals on salivary cortisol, alpha-amylase, and immunoglobulin A.

PubMed

Augner, Christoph; Hacker, Gerhard W; Oberfeld, Gerd; Florian, Matthias; Hitzl, Wolfgang; Hutter, Jörg; Pauser, Gernot

2010-06-01

The present study aimed to test whether exposure to radiofrequency electromagnetic fields (RF-EMF) emitted by mobile phone base stations may have effects on salivary alpha-amylase, immunoglobulin A (IgA), and cortisol levels. Fifty seven participants were randomly allocated to one of three different experimental scenarios (22 participants to scenario 1, 26 to scenario 2, and 9 to scenario 3). Each participant went through five 50-minute exposure sessions. The main RF-EMF source was a GSM-900-MHz antenna located at the outer wall of the building. In scenarios 1 and 2, the first, third, and fifth sessions were "low" (median power flux density 5.2 microW/m(2)) exposure. The second session was "high" (2126.8 microW/m(2)), and the fourth session was "medium" (153.6 microW/m(2)) in scenario 1, and vice versa in scenario 2. Scenario 3 had four "low" exposure conditions, followed by a "high" exposure condition. Biomedical parameters were collected by saliva samples three times a session. Exposure levels were created by shielding curtains. In scenario 3 from session 4 to session 5 (from "low" to "high" exposure), an increase of cortisol was detected, while in scenarios 1 and 2, a higher concentration of alpha-amylase related to the baseline was identified as compared to that in scenario 3. IgA concentration was not significantly related to the exposure. RF-EMF in considerably lower field densities than ICNIRP-guidelines may influence certain psychobiological stress markers. Copyright © 2010 The Editorial Board of Biomedical and Environmental Sciences. Published by Elsevier B.V. All rights reserved.

Alkaline thermostable pectinase enzyme from Aspergillus niger strain MCAS2 isolated from Manaslu Conservation Area, Gorkha, Nepal.

PubMed

Khatri, Bhim Prakash; Bhattarai, Tribikram; Shrestha, Sangita; Maharjan, Jyoti

2015-01-01

Pectinase enzymes are one of the commercially important enzymes having great potential in various industries especially in food industry. Pectinases accounts for 25 % of global food enzymes produced and their market is increasing day by day. Therefore, the exploration of microorganism with novel characteristics has always been the focus of the research. Microorganism dwelling in unique habitat may possess unique characteristics. As such, a pectinase producing fungus Aspergillus niger strain MCAS2 was isolated from soil of Manaslu Conservation Area (MCA), Gorkha, Nepal. The optimum production of pectinase enzyme was observed at 48 h of fermentation. The pectinase enzyme was partially purified by cold acetone treatment followed by Sephadex G-75 gel filtration chromatography. The partially purified enzyme exhibited maximum activity 60 U/mg which was almost 8.5-fold higher than the crude pectinase. The approximate molecular weight of the enzyme was found to be 66 kDa as observed from SDS-PAGE. The pectinase enzyme was active at broad range of temperature (30-70 °C) and pH (6.2-9.2). Optimum temperature and pH of the pectinase enzyme were 50 °C and 8.2 respectively. The enzyme was stable up to 70 °C and about 82 % of pectinase activity was still observed at 100 °C. The thermostable and alkaline nature of this pectinase can meet the demand of various industrial processes like paper and pulp industry, in textile industry, fruit juice industry, plant tissue maceration and wastewater treatment. In addition, the effect of different metal ions on pectinase activity was also studied.

Immobilization of pectinase on oxidized pulp fiber and its application in whitewater treatment.

PubMed

Wu, Rina; He, Bei-Hai; Zhao, Guang-Lei; Qian, Li-Ying; Li, Xiao-Feng

2013-09-12

Modified pulp fiber was originally used as a new type of carrier for pectinase immobilization. Pulp fiber was oxidized by sodium periodate to produce aldehyde groups for covalently binding with amino groups of pectinase. Results showed that the enzymatic activity of immobilized pectinase on pulp fiber reached 65 μgg(-1)min(-1) when immobilization pH value, temperature and time were of 7.0, 20 °C and 15 min, respectively. The immobilized pectinase showed higher thermo stability in a wider temperature range of 40-70 °C than its free type and its optimal pH shifted from 8.0 to 8.8. Furthermore, the immobilized pectinase exhibited good operational stability. When employed in whitewater treatment of papermaking industry, it still efficiently decreased the cationic demand after operating repeatedly for six batches. The results obtained demonstrate a promising route to prepare available, cheap and biodegradable carrier for immobilizing enzymes with potential application in wastewater treatment in papermaking industry. Copyright © 2013 Elsevier Ltd. All rights reserved.

Starch digestibility, energy utilization, and growth performance of broilers fed corn-soybean basal diets supplemented with enzymes.

PubMed

Stefanello, C; Vieira, S L; Santiago, G O; Kindlein, L; Sorbara, J O B; Cowieson, A J

2015-10-01

A study was conducted to evaluate the effects of dietary α-amylase and β-xylanase supplementation of corn-soy diets, formulated with or without supplemental phytase, on growth performance, energy utilization, and starch digestibility in broiler chickens. A total of 336 slow-feathering, Cobb × Cobb 500 male broilers were randomly distributed to 6 treatments having 8 replicates of 7 birds each. Birds were fed a common starter diet to d 14 post-hatch (3,050 kcal/kg AMEn, 21.7% CP, 1.05% Ca, and 0.53% nPP). The experimental diets were provided afterwards until d 25. A 2 × 3 factorial arrangement of 2 control diets (basal = corn-soy diet without added phytase or PHY = corn-soy diet formulated with 1,000 phytase units/kg) and 3 carbohydrase supplementations (0, 80 kilo-Novo α-amylase units/kg, or 80 kilo-Novo α-amylase units/kg + 100 fungal β-xylanase units/kg) was used from d 14 to 25. Excreta were collected from 21 to 24 d and all birds were euthanized at 25 d for jejunum and ileum content collection. Samples of feed, excreta, and jejunal and ileal digesta were analyzed for determination of total tract retention and ileal apparent digestibility. No interactions between diet and carbohydrase were observed. Broilers fed diets formulated with phytase or supplemented with amylase + xylanase had higher BW gain (BWG) and lower FCR (P < 0.05) when compared with birds fed diets without carbohydrases. Relative to the basal diet, AMEn was increased (P < 0.01) by 70 kcal/kg and 99 kcal/kg when birds were fed the diet supplemented with amylase and amylase + xylanase, respectively. Starch digestibility in the jejunum and ileum was increased (P < 0.05) by 3.5% and 2.4%, respectively, when birds were fed the diet supplemented with amylase + xylanase. Results from this experiment show that corn-soy diets having phytase and supplemented with amylase and xylanase led to increased growth performance, AMEn, and starch digestibility in broilers. Furthermore, the efficacy of

Continuous production of pectinase by immobilized yeast cells on spent grains.

PubMed

Almeida, Catarina; Brányik, Tomás; Moradas-Ferreira, Pedro; Teixeira, José

2003-01-01

A yeast strain secreting endopolygalacturonase was used in this work to study the possibility of continuous production of this enzyme. It is a feasible and interesting alternative to fungal batch production essentially due to the specificity of the type of pectinase excreted by Kluyveromyces marxianus CCT 3172, to the lower broth viscosity and to the easier downstream operations. In order to increase the reactors' productivity, a cellulosic carrier obtained from barley spent grains was tested as an immobilization support. Two types of reactors were studied for pectinase production using glucose as a carbon and energy source--a continuous stirred tank reactor (CSTR) and a packed bed reactor (PBR) with recycled flow. The highest value for pectinase volumetric productivity (P(V)=0.98 U ml(-1) h(-1)) was achieved in the PBR for D=0.40 h(-1), a glucose concentration on the inlet of S(in)=20 g l(-1), and a biomass load in the support of X(i)=0.225 g g(-1). The results demonstrate the attractiveness of the packed bed system for pectinase production.

Diurnal Salivary Alpha-amylase Dynamics among Dementia Family Caregivers

PubMed Central

Liu, Yin; Granger, Douglas A.; Kim, Kyungmin; Klein, Laura C.; Almeida, David M.; Zarit, Steven H.

2016-01-01

Objective The study examined diurnal regulation of salivary alpha-amylase (sAA) in association with daily stressors, adult day services (ADS) use, and other caregiving characteristics. Methods A sample of 165 family caregivers of individuals with dementia (IWD) completed an 8-day diary study. Caregivers provided 5 saliva samples across the 8 days. On some days, caregivers provided all or most of the care. On other days, their relative attended ADS for part of the day. A 3-level unconditional linear spline model was fit to describe the typical sAA diurnal rhythms. Predictors were then added to the unconditional model to test the hypotheses on ADS use and daily stressors. Results Daily ADS use did not have an effect on diurnal sAA regulation. However, controlling for daily ADS use, greater ADS use over the 8 days was associated with a more prominent rise between 30 minutes after wake-up and before lunch, and a more prominent decline between before lunch and late afternoon. Fewer ADS days were associated with a more flattened sAA diurnal rhythm. Additionally, greater daily care-related stressor exposures had a within-person association with lower sAA levels in the late afternoon. Care-related stressor exposures had significant within- and between-person associations with sAA diurnal slopes. Furthermore, daily positive experiences had a significant between-person association with sAA diurnal slopes. Conclusions Caring for a disabled family member may heighten the vulnerability to potential physiological conditions. Respite from care stressors from ADS use may have some biobehavioral benefits on sAA regulations. PMID:27786517

Salivary Alpha Amylase and Cortisol Levels in Children with Global Developmental Delay and Their Relation with the Expectation of Dental Care and Behavior during the Intervention

ERIC Educational Resources Information Center

dos Santos, Marcio Jose Possari; Bernabe, Daniel Galera; Nakamune, Ana Claudia de Melo Stevanato; Perri, Silvia Helena Venturoli; de Aguiar, Sandra Maria Herondina Coelho Avila; de Oliveira, Sandra Helena Penha

2012-01-01

The purpose of this study was to analyze the alpha-amylase (sAA) and cortisol levels in children with Global developmental delay (GDD) before and after dental treatment and its association with the children's behavior during treatment. The morning salivary cortisol levels and activity of sAA of 33 children with GDD were evaluated before and after…

Some distinguishable properties between acid-stable and neutral types of alpha-amylases from acid-producing koji.

PubMed

Suganuma, Toshihiko; Fujita, Kiyotaka; Kitahara, Kanefumi

2007-11-01

The highly humid climate of Japan facilitates the growth of various molds. Among these molds, Aspergillus oryzae is the most important and popular in Japan, and has been used as yellow-koji in producing many traditional fermented beverages and foods, such as Japanese sake, and soy sauce. Taka-amylase A (TAA), a major enzyme produced by the mold, is well known worldwide to be a leading enzyme for industrial utilization and academic study, since many extensive studies have been carried out with TAA. In southern Kyushu, the other koji's of citric acid-producing molds have often been used, such as in the production of a traditional distilled liquor of shochu. The koji molds black-koji and white-koji produce two types of alpha-amylase, namely, acid-stable (AA) and common neutral (NA). The latter enzyme is enzymatically and genetically similar to TAA. In this review, we investigate AA from three molds, Aspergillus niger, A. kawachii and A. awamori, and the yeast Cryptococcus sp. regarding the distinguishable properties between AA and NA. (i) The N-terminus amino acid sequences of AA determined by molecular cloning started with the sequence of L-S-A-, whereas those of NA started with A-T-P-. (ii) Most of the full sequences of AA were composed of, besides a core catalytic domain, an extra domain of a hinge region and a carbohydrate binding domain, which could be responsible for raw-starch-digestibility. The AA from A. niger has no exceptionally extra domain, similarly to NA. (iii) Simple methods for distinguishing AA from NA using CNP-alpha-G3 and G5 as substrates were developed by our group. (iv) The number of subsite in AA on the basis of its cleavage pattern of maltooligosaccharides was estimated to be five, which differs from that of TAA, 7-9. AA has many advantages in industrial applications, such as its acid-stability, thermostability, and raw-starch digesting properties.

The xylanase inhibitor TAXI-III counteracts the necrotic activity of a Fusarium graminearum xylanase in vitro and in durum wheat transgenic plants.

PubMed

Moscetti, Ilaria; Faoro, Franco; Moro, Stefano; Sabbadin, Davide; Sella, Luca; Favaron, Francesco; D'Ovidio, Renato

2015-08-01

The xylanase inhibitor TAXI-III has been proven to delay Fusarium head blight (FHB) symptoms caused by Fusarium graminearum in transgenic durum wheat plants. To elucidate the molecular mechanism underlying the capacity of the TAXI-III transgenic plants to limit FHB symptoms, we treated wheat tissues with the xylanase FGSG_03624, hitherto shown to induce cell death and hydrogen peroxide accumulation. Experiments performed on lemmas of flowering wheat spikes and wheat cell suspension cultures demonstrated that pre-incubation of xylanase FGSG_03624 with TAXI-III significantly decreased cell death. Most interestingly, a reduced cell death relative to control non-transgenic plants was also obtained by treating, with the same xylanase, lemmas of TAXI-III transgenic plants. Molecular modelling studies predicted an interaction between the TAXI-III residue H395 and residues E122 and E214 belonging to the active site of xylanase FGSG_03624. These results provide, for the first time, clear indications in vitro and in planta that a xylanase inhibitor can prevent the necrotic activity of a xylanase, and suggest that the reduced FHB symptoms on transgenic TAXI-III plants may be a result not only of the direct inhibition of xylanase activity secreted by the pathogen, but also of the capacity of TAXI-III to avoid host cell death. © 2014 BSPP AND JOHN WILEY & SONS LTD.

The 53-kDa proteolytic product of precursor starch-hydrolyzing enzyme of Aspergillus niger has Taka-amylase-like activity.

PubMed

Ravi-Kumar, K; Venkatesh, K S; Umesh-Kumar, S

2007-04-01

The 53-kDa amylase secreted by Aspergillus niger due to proteolytic processing of the precursor starch-hydrolyzing enzyme was resistant to acarbose, a potent alpha-glucosidase inhibitor. The enzyme production was induced when A. niger was grown in starch medium containing the inhibitor. Antibodies against the precursor enzyme cross-reacted with the 54-kDa Taka-amylase protein of A. oryzae. It resembled Taka-amylase in most of its properties and also hydrolyzed starch to maltose of alpha-anomeric configuration. However, it did not degrade maltotriose formed during the reaction and was not inhibited by zinc ions.

Harsh discipline and behavior problems: the moderating effects of cortisol and alpha-amylase.

PubMed

Chen, Frances R; Raine, Adrian; Rudo-Hutt, Anna S; Glenn, Andrea L; Soyfer, Liana; Granger, Douglas A

2015-01-01

Numerous studies link harsh discipline to adjustment problems in youth, yet not all individuals exposed to harsh discipline develop behavior problems. Contemporary theory suggests that this relationship could be moderated by individual differences in environmentally sensitive biological systems. This study investigated whether the interaction between hypothalamic-pituitary-adrenal (HPA) activity and autonomic nervous system (ANS) arousal moderated the link between harsh discipline and behavior problems. Three saliva samples were collected on a single day from 425 inner city youth (50% male, age 11-12 years, 80% African American) and were later assayed for cortisol (HPA) and alpha-amylase (ANS). Problem behavior was assessed by self- and parent-report using the Child Behavior Checklist. Youth also reported the level of harsh discipline that they experienced. Harsh discipline was positively associated with externalizing and internalizing problems only when there were asymmetrical profiles of HPA activity and ANS arousal. This pattern was evident for boys but not girls. Findings are discussed in relation to prevailing theories suggesting that biological susceptibility translates adversity into risk for behavior problems. Copyright © 2014 Elsevier B.V. All rights reserved.

Enhancement of pectinase production by ultraviolet irradiation and diethyl sulfate mutagenesis of a Fusarium oxysporum isolate.

PubMed

Yin, L B; Zhang, C F; Xia, Q L; Yang, Y; Xiao, K; Zhao, L Z

2016-09-23

Fusarium oxysporum strain BM-201 was treated with ultraviolet (UV) radiation to obtain a high pectinase-producing strain. Mutant UV-10-41 was obtained and then treated by diethyl sulfate. Next, the mutant UV-diethyl sulfate-43 derived from UV-10-41 was selected as high pectinase-producing strain. Mutant UV-diethyl sulfate-43 was incubated on slant for 10 generations, demonstrating that the pectinase-producing genes were stable. Pectinase activity reached 391.2 U/mL, which is 73.6% higher than that of the original strain.

Unintended changes in protein expression revealed by proteomic analysis of seeds from transgenic pea expressing a bean alpha-amylase inhibitor gene.

PubMed

Chen, Hancai; Bodulovic, Greg; Hall, Prudence J; Moore, Andy; Higgins, Thomas J V; Djordjevic, Michael A; Rolfe, Barry G

2009-09-01

Seeds of genetically modified (GM) peas (Pisum sativum L.) expressing the gene for alpha-amylase inhibitor-1 (alphaAI1) from the common bean (Phaseolus vulgaris L. cv. Tendergreen) exhibit resistance to the pea weevil (Bruchus pisorum). A proteomic analysis was carried out to compare seeds from GM pea lines expressing the bean alphaAI1 protein and the corresponding alphaAI1-free segregating lines and non-GM parental line to identify unintended alterations to the proteome of GM peas due to the introduction of the gene for alphaAI1. Proteomic analysis showed that in addition to the presence of alphaAI1, 33 other proteins were differentially accumulated in the alphaAI1-expressing GM lines compared with their non-GM parental line and these were grouped into five expression classes. Among these 33 proteins, only three were found to be associated with the expression of alphaAI1 in the GM pea lines. The accumulation of the remaining 30 proteins appears to be associated with Agrobacterium-mediated transformation events. Sixteen proteins were identified after MALDI-TOF-TOF analysis. About 56% of the identified proteins with altered accumulation in the GM pea were storage proteins including legumin, vicilin or convicilin, phaseolin, cupin and valosin-containing protein. Two proteins were uniquely expressed in the alphaAI1-expressing GM lines and one new protein was present in both the alphaAI1-expressing GM lines and their alphaAI1-free segregating lines, suggesting that both transgenesis and transformation events led to demonstrable changes in the proteomes of the GM lines tested.

Analysis of functional xylanases in xylan degradation by Aspergillus niger E-1 and characterization of the GH family 10 xylanase XynVII.

PubMed

Takahashi, Yui; Kawabata, Hiroaki; Murakami, Shuichiro

2013-01-01

Xylanases produced by Aspergillus niger are industrially important and many types of xylanases have been reported. Individual xylanases have been well studied for their enzymatic properties, gene cloning, and heterologous expression. However, less attention has been paid to the relationship between xylanase genes carried on the A. niger genome and xylanases produced by A. niger strains. Therefore, we examined xylanase genes encoded on the genome of A. niger E-1 and xylanases produced in culture. Seven putative xylanase genes, xynI-VII (named in ascending order of the molecular masses of the deduced amino acid sequences), were amplified from the strain E-1 genome using primers designed from the genome sequence of A. niger CBS 513.88 by PCR and phylogenetically classified into three clusters. Additionally, culture supernatant analysis by DE52 anion-exchange column chromatography revealed that this strain produced three xylanases, XynII, XynIII, and XynVII, which were identified by N-terminal amino acid sequencing and MALDI-TOF-MS analyses, in culture when gown in 0.5% xylan medium supplemented with 50 mM succinate. Furthermore, XynVII, the only GH family 10 xylanase in A. niger E-1, was purified and characterized. The purified enzyme showed a single band with a molecular mass of 35 kDa by SDS-PAGE. The highest activity of purified XynVII was observed at 55°C and pH 5.5. The enzyme was stable in the broad pH range of 3-10 and up to 60°C and was resistant to most metal ions and modifying regents. XynVII showed high specificity against beechwood xylan with K m and V max values of 2.8 mg mL(-1) and 127 μmol min(-1)mg(-1), respectively. TLC and MALDI-TOF-MS analyses showed that the final hydrolyzed products of the enzyme from beechwood xylan were xylose, xylobiose, and xylotriose substituted with a 4-o-metylglucuronic acid residue.

Immobilization and stabilization of pectinase by multipoint attachment onto an activated agar-gel support.

PubMed

Li, Tuoping; Li, Suhong; Wang, Na; Tain, Lirui

2008-08-15

Pectinase was immobilized on an activated agar-gel support by multipoint attachment. The maximal activity of immobilized pectinase was obtained at 5°C, pH 3.6, with a 24h reaction time at an enzyme dose of 0.52mg protein/g gel, and the gel was activated with 1.0M glycidol. These conditions increased the thermal stability of the immobilized pectinase 19-fold compared with the free enzyme at 65°C. The optimal temperature for pectinase activity changed from 40 to 50°C after immobilization; however, the optimal pH remained unchanged. The immobilized enzyme also exhibited great operational stability, and an 81% residual activity was observed in the immobilized enzyme after 10 batch reactions. Copyright © 2008 Elsevier Ltd. All rights reserved.

Diurnal salivary alpha-amylase dynamics among dementia family caregivers.

PubMed

Liu, Yin; Granger, Douglas A; Kim, Kyungmin; Klein, Laura C; Almeida, David M; Zarit, Steven H

2017-02-01

The study examined diurnal regulation of salivary alpha-amylase (sAA) in association with daily stressors, adult day services (ADS) use, and other caregiving characteristics. A sample of 165 family caregivers of individuals with dementia (IWD) completed an 8-day diary study. Caregivers provided 5 saliva samples across the 8 days. On some days, caregivers provided all or most of the care. On other days, their relative attended ADS for part of the day. A 3-level unconditional linear spline model was fit to describe the typical sAA diurnal rhythms. Predictors were then added to the unconditional model to test the hypotheses on ADS use and daily stressors. Daily ADS use did not have an effect on diurnal sAA regulation. However, controlling for daily ADS use, greater ADS use over the 8 days was associated with a more prominent rise between 30 min after wake-up and before lunch, and a more prominent decline between before lunch and late afternoon. Fewer ADS days were associated with a more flattened sAA diurnal rhythm. Additionally, greater daily care-related stressor exposures had a within-person association with lower sAA levels in the late afternoon. Care-related stressor exposures had significant within- and between-person associations with sAA diurnal slopes. Furthermore, daily positive experiences had a significant between-person association with sAA diurnal slopes. Caring for a disabled family member may heighten the vulnerability to potential physiological conditions. Respite from care stressors from ADS use may have some biobehavioral benefits on sAA regulations. (PsycINFO Database Record (c) 2017 APA, all rights reserved).

Activity and cellular localization of amylases of rabbit cecal bacteria.

PubMed

Sirotek, K; Marounek, M; Suchorská, O

2006-01-01

Five 11-week-old rabbits, fed a commercial granulated feed, were slaughtered and cecal starch-degrading bacteria enumerated; total concentration of cultivable bacteria utilizing starch averaged 5.5 x 10(10) CFU/g. The activity and cellular localization of amylases was determined in 9 bacteria identified as Actinomyces israeli (strains AA2 and AD4), Bacteroides spp. (strain AA3), Dichelobacter nodosus (strain AA4), Mitsuokella multiacidus (strain AA6), Eubacterium spp. (strains AA7 and AB2), Clostridium spp. (strains AD1 and AA5). Four strains (AA3, AA4, AA5, AD4) produced extracellular amylases with an activity of 26-35 micromol of reducing sugars per h per mg of protein; in five strains (AA2, AA6, AA7, AB2, AD1) amylases were membrane-bound with an activity of 14-18 micromol of reducing sugars per h per mg of protein. All strains exhibited a low intracellular amylolytic activity. The pH optimum of amylases was 6.8-7.0. In strains producing extracellular amylases a substantial loss of viscosity was observed during incubations of cultivation supernatant with starch, similar to viscosity reduction in starch solutions treated with alpha-amylase; this indicates an endo-type (random cleavage) of extracellular amylase reaction in the bacteria under study. No strain possessed glucoamylase activity.

Hypoglycemic activity of Pyrus biossieriana Buhse leaf extract and arbutin: Inhibitory effects on alpha amylase and alpha glucosidase

PubMed Central

Yousefi, Fatemeh; Mahjoub, Soleiman; Pouramir, Mahdi; Khadir, Fatemeh

2013-01-01

Background: The mechanism of hypoglycemic and hypolipidemic activities of Pyrus biossieriana Buhse leaf extract (PbBLE) and its phytochemical component arbutin, have not been well determined. The present study was performed to understand the hypoglycemic activity mechanisms of pbBLE and arbutin more clearly. Methods: In vitro enzymatic carbohydrate digestion with PbBLE and arbutin was assessed using α-amylase and α-glucosidase powders. The enzyme solutions were premixed with PbBLE and arbutin at different concentrations (0.1, 1, 10 and 100 mg/ml). Substrate solutions and colorimetric reagents were added to the reaction. The release of glucose was determined by spectrophotometric method. Acarbose was used as the positive control. Results: The extract (10, 100 mg/ ml) completely inhibit α- amylase and α- glucosidase activities. The extract produced higher reduction of α-amylase and α-glucosidase activity than arbutin. Inhibition at various concentrations (0.1, 1, 10, 100 mg/ml) were significantly different (p<0.05). Conclusion: Our results exhibited that both the extract and arbutin were able to suppress the enzymes strongly. PMID:24294470

Highly cold-active pectinases under wine-like conditions from non-Saccharomyces yeasts for enzymatic production during winemaking.

PubMed

Merín, M G; Morata de Ambrosini, V I

2015-05-01

The influence of oenological factors on cold-active pectinases from 15 preselected indigenous yeasts belonging to Aureobasidium pullulans, Filobasidium capsuligenum, Rhodotorula dairenensis, Cryptococcus saitoi and Saccharomyces cerevisiae was investigated. Pectinolytic enzymes were constitutive or partially constitutive; and high glucose concentration (200 g l(-1) ) did not affect or increased pectinase production at 12°C and pH 3·5 (up to 113·9 U mg(-1) ) only in A. pullulans strains. SO2 (120 mg l(-1) ) slightly affected the growth of A. pullulans strains but did not affect pectinase production levels. Ethanol (15%) barely affected pectinase activity of A. pullulans strains but diminished relative activity to 12-79% of basidiomycetous yeasts. Moreover, non-Saccharomyces strains showed promising properties of oenological interest. This study demonstrates that cold-active pectinases from some A. pullulans strains were able to remain active at glucose, ethanol and SO2 concentrations usually found in vinification, and suggests their potential use as processing aids for low-temperature winemaking. Nowadays, there is increasing interest in low-temperature winemaking. Nevertheless, commercial oenological pectinases, produced by fungi, are rarely active at low temperatures. Cold-active pectinases that are stable under vinification conditions are needed. This study indicated that cold-active and acid-tolerant pectinases from non-Saccharomcyes yeasts were able to remain active at glucose, ethanol and SO2 concentrations usually found in winemaking. Furthermore, not only are these yeasts a source of cold-active pectinases, but the yeasts themselves are also potential adjunct cultures for oenology to produce these enzymes during cold-winemaking. © 2015 The Society for Applied Microbiology.

Effects of the culture media optimization on pectinase production by Enterobacter sp. PSTB-1.

PubMed

Reddy, M Purna Chandra; Saritha, K V

2016-12-01

In the present study, media composition for high production of pectinase by Enterobacter sp. PSTB-1 in submerged fermentation was optimized using response surface methodology (RSM). Mango fruit processing industrial waste (MIW) was used as substrate (carbon source) as it contains high amount of pectin. Enterobacter sp. PSTB-1 used in present study has given pectin clear zone (PCZ) of 34 mm is higher than other isolates. The experimental results made by statistical design for high pectinase production revealed that the suitable media components: NaNO 3 2.0 g/l, KCl 0.50 g/l, KH 2 PO 4 1.0 g/l, MgSO 4 ·7H 2 O 0.50 g/l, Yeast extract 1.0 g/l, mango industrial waste powder 5.0 g/l. The actual pectinase activity was 75.23 % correlated with the predicted pectinase activity where the model (CCD) was significant. Response surface modelling applied effectively to optimize the production of pectinase in submerged fermentation to make the process low cost-effective by using powdered mango industrial waste as substrate.

Potential application of alkaline pectinase from Bacillus subtilis SS in pulp and paper industry.

PubMed

Ahlawat, Sonia; Mandhan, R P; Dhiman, Saurabh Sudha; Kumar, Rakesh; Sharma, Jitender

2008-06-01

Pectinase production from Bacillus subtilis SS was optimized under solid-state fermentation (5,943 U/g of dry bacterial bran). The pectinase produced was stable in neutral to alkaline pH range at 70 degrees C; therefore, the suitability of this pectinase in pulp and paper industry was investigated. The enzyme pretreatment process was optimized, and a pectinase dose of 5 IU/g of oven-dried pulp (10% consistency) at pH 9.5 temperature 70 degrees C after 150 min of treatment gave the best pretreatment to the pulp. An increase of 4.3% in brightness along with an increase of 14.8 and 65.3% in whiteness and fluorescence, respectively, whereas a 15% decrease in the yellowness of the pretreated pulp were observed. There was a 5.85% reduction in kappa number and 6.1% reduction in permanganate number along with a reduction in the chemical oxygen demand value. Significant characteristics showed by pectinase open new possibilities of application of this cellulase-free enzyme in the pulp and paper industry by reducing the negative environmental impact of chemicals apart from improving the properties of paper.

A hyper-thermostable α-amylase from Pyrococcus furiosus accumulates in Nicotiana tabacum as functional aggregates.

PubMed

Zhu, Hong; Reynolds, L Bruce; Menassa, Rima

2017-06-19

Alpha amylase hydrolyzes α-bonds of polysaccharides such as starch and produces malto-oligosaccharides. Its starch saccharification applications make it an essential enzyme in the textile, food and brewing industries. Commercially available α-amylase is mostly produced from Bacillus or Aspergillus. A hyper-thermostable and Ca 2++ independent α-amylase from Pyrococcus furiosus (PFA) expressed in E.coli forms insoluble inclusion bodies and thus is not feasible for industrial applications. We expressed PFA in Nicotiana tabacum and found that plant-produced PFA forms functional aggregates with an accumulation level up to 3.4 g/kg FW (fresh weight) in field conditions. The aggregates are functional without requiring refolding and therefore have potential to be applied as homogenized plant tissue without extraction or purification. PFA can also be extracted from plant tissue upon dissolution in a mild reducing buffer containing SDS. Like the enzyme produced in P. furiosus and in E. coli, plant produced PFA preserves hyper-thermophilicity and hyper-thermostability and has a long shelf life when stored in lyophilized leaf tissue. With tobacco's large biomass and high yield, hyper-thermostable α-amylase was produced at a scale of 42 kg per hectare. Tobacco may be a suitable bioreactor for industrial production of active hyperthermostable alpha amylase.

Effect of domestic cooking on the starch digestibility, predicted glycemic indices, polyphenol contents and alpha amylase inhibitory properties of beans (Phaseolis vulgaris) and breadfruit (Treculia africana).

PubMed

Chinedum, E; Sanni, S; Theressa, N; Ebere, A

2018-01-01

The effect of processing on starch digestibility, predicted glycemic indices (pGI), polyphenol contents and alpha amylase inhibitory properties of beans (Phaseolis vulgaris) and breadfruit (Treculia africana) was studied. Total starch ranged from 4.3 to 68.3g/100g, digestible starch ranged from 4.3 to 59.2 to 65.7g/100g for the raw and processed legumes; Resistance starch was not detected in most of the legumes except in fried breadfruit and the starches in both the raw and processed breadfruit were more rapidly digested than those from raw and cooked beans. Raw and processed breadfruit had higher hydrolysis curves than raw and processed beans with the amylolysis level in raw breadfruit close to that of white bread. Raw beans had a low glycemic index (GI); boiled beans and breadfruit had intermediate glycemic indices respectively while raw and fried breadfruit had high glycemic indices. Aqueous extracts of the food samples had weak α-amylase inhibition compared to acarbose. The raw and processed legumes contained considerable amounts of dietary phenols and flavonoids. The significant correlation (r=0.626) between α-amylase inhibitory actions of the legumes versus their total phenolic contents suggests the contribution of the phenolic compounds in these legumes to their α-amylase inhibitory properties. Copyright © 2017 Elsevier B.V. All rights reserved.

Use of fruit residues for pectinase production by Aspergillus flavipes FP-500 and Aspergillus terreus FP-370.

PubMed

Martínez-Trujillo, A; Arreguín-Rangel, L; García-Rivero, M; Aguilar-Osorio, G

2011-08-01

Utilization of fruit residues for pectinase production by two Aspergillus strains for recognizing the effects of some factors during fermentation and describing enzyme production kinetics. Pectinase production on several fruit residues was compared. The effects of three factors on the production of several pectinases were evaluated by a full factorial 2(k) experimental design. Higher activities were obtained on lemon peel. In both strains, acidic pH values and high carbon source concentration favoured exopectinase and endopectinase production, while higher pH values and low carbon source concentration promoted pectin lyase and rhamnogalacturonase production. Unstructured mathematical modelling provided a good description of pectinase production in a submerged batch culture. Fruit residues were very good substrates for pectinase production, and Aspergillus strains used showed a promising performance in submerged fermentation. Mathematical modelling was useful to describe growth and pectinase production. Lemon peel can be used as a substrate to obtain high pectinase titres by Aspergillus flavipes FP-500 and Aspergillus terreus FP-370. The factors that contributed to improve the yield were identified, which supports the possibility of using this substrate in the industrial production of these enzymes. © 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.

Stress exacerbates pain in the everyday lives of women with fibromyalgia syndrome--The role of cortisol and alpha-amylase.

PubMed

Fischer, Susanne; Doerr, Johanna M; Strahler, Jana; Mewes, Ricarda; Thieme, Kati; Nater, Urs M

2016-01-01

Although fibromyalgia syndrome (FMS) is a chronic condition, its cardinal symptom pain is known to fluctuate over the day. Stress has often been claimed to exacerbate pain; however, there is barely any evidence on whether or not this is true on a day-to-day basis (and, alternatively, on whether pain leads to increased stress levels). Using an ecologically valid measurement design, we tested whether and how stress and pain are intertwined in participants with FMS. We additionally examined the role of the two major stress-responsive systems, the hypothalamic-pituitary-adrenal axis and the autonomic nervous system, as potential mediators of this relationship. An ambulatory assessment study was conducted over the course of 14 days. On each day, 32 females with FMS provided six diary entries on momentary stress and pain levels. Saliva samples were collected at the same time points to determine cortisol and alpha-amylase as indicators of stress-responsive systems. Higher stress at a given measurement time point was associated with higher reported pain levels at the subsequent time point (UC=1.47, p<0.001), but not vice versa (UC<0.01, p=0.179). The stress-pain relationship was neither mediated by momentary cortisol nor by alpha-amylase; however, momentary cortisol was independently associated with momentary pain (UC=0.27, p=0.009). Stress seems to be a powerful exacerbating factor for pain as experienced by patients with FMS in their everyday lives. Cortisol may be involved in the diurnal fluctuation of pain levels in patients with FMS. Future studies should identify relevant daily stressors in persons with FMS and scrutinize the mechanisms underlying the cortisol-pain relationship. Copyright © 2015 Elsevier Ltd. All rights reserved.

Structure of waxy maize starch hydrolyzed by maltogenic alpha-amylase in relation to its retrogradation

USDA-ARS?s Scientific Manuscript database

Maltogenic a-amylase is widely used as an antistaling agent in bakery foods. The objective of this study was to determine the degree of hydrolysis (DH) and starch structure after maltogenic amylase treatments in relation to its retrogradation. Waxy maize starch was cooked and hydrolyzed to different...

Daytime Secretion of Salivary Cortisol and Alpha-Amylase in Preschool-Aged Children with Autism and Typically Developing Children

PubMed Central

Corbett, Blythe A.; Granger, Douglas A.; Boyce, W. Thomas; Anders, Thomas F.; Tager, Ira B.

2013-01-01

We examined daytime salivary cortisol and salivary alpha-amylase (sAA) secretion levels and variability in preschool-aged children with autism (AUT) and typically developing children (TYP). Fifty-two subjects (26 AUT and 26 TYP) were enrolled. Salivary samples were obtained at waking, midday, and bedtime on two consecutive days at three phases (baseline, 3 months later, 6 months later). There were modest increases in waking cortisol and sAA levels in AUT relative to TYP, but the increases were not statistically significant. Important differences were observed in cortisol and sAA variability between AUT and TYP. There was also a graded response among AUT by functional status—cortisol and sAA secretion levels were higher when IQ was lower. PMID:22477468

Enhanced production of pectinase by Aspergillus terreus NCFT 4269.10 using banana peels as substrate.

PubMed

Sethi, Bijay Kumar; Nanda, Prativa Kumari; Sahoo, Santilata

2016-06-01

Aspergillus terreus NCFT4269.10 was implemented in solid-state (SSF) and liquid static surface fermentation (LSSF) for biosynthesis of pectinase. Amongst various substrates, like, mustard oil cake, neem oil cake, groundnut oil cake, black gram peels, green gram peels, chickling vetch peels/grass pea peels wheat bran, pearl millet residues, finger millet waste, broken rice, banana peels (BP), apple pomace (AP) and orange peels, banana peel (Musa paradisiaca L.; Family: Musaceae) was most suitable for pectinase biosynthesis (LSSF: 400 ± 21.45 Uml -1 ; SSF: 6500 ± 1116.21 Ug -1 ). Optimization of process parameters using one-variable-at-a-time method revealed that an initial medium pH of 5.0 at 30 °C and 96 h of incubation along with mannitol, urea, ammonium persulfate and isoleucine have positive influence on pectinase production. Further, K + (1 mM), Riboflavin (10 mg 100 ml -1 ) and gibberellic acid (0.025 %, w/v) supported in enhanced pectinase production. Banana peels and AP at a ratio of 9:1, moisture content of 90 % with 2 % inoculum size were suitable combinations for production of pectinase. Similarly, 96 h of soaking time with 0.1 M phosphate buffer (pH 6.5) is essential for pectinase recovery. Purification to electrophoretic homogeneity revealed 1.42 fold purification with 8.08 % yield and a molecular weight of 24.6 kDa. Scaling up of various fermentation parameters and supplementing BP as the substrate for pectinase production with better recovery could make it promising for different industrial exploitation.

Alpha-amylase reactivity in relation to psychopathic traits in adults.

PubMed

Glenn, Andrea L; Remmel, Rheanna J; Raine, Adrian; Schug, Robert A; Gao, Yu; Granger, Douglas A

2015-04-01

Recent investigations of the psychobiology of stress in antisocial youth have benefited from a multi-system measurement model. The inclusion of salivary alpha-amylase (sAA), a surrogate marker of autonomic/sympathetic nervous system (ANS) activity, in addition to salivary cortisol, a biomarker of the hypothalamic-pituitary-adrenal (HPA) axis functioning, has helped define a more complete picture of individual differences and potential dysfunction in the stress response system of these individuals. To the authors' knowledge, no studies have examined sAA in relation to antisocial behavior in adults or in relation to psychopathic traits specifically. In the present study, we examined sAA, in addition to salivary cortisol, in a relatively large sample (n=158) of adult males (M age=36.81, range=22-67 years; 44% African-American, 34% Caucasian, 16% Hispanic) recruited from temporary employment agencies with varying levels of psychopathic traits. Males scoring highest in psychopathy were found to have attenuated sAA reactivity to social stress compared to those scoring lower in psychopathy. No differential relationships with the different factors of psychopathy were observed. In contrast to studies of antisocial youth, there were no interactions between sAA and cortisol levels in relation to psychopathy, but there was a significant interaction between pre-stressor levels of sAA and cortisol. Findings reveal potential regulatory deficits in the fast-acting, 'fight or flight', component of the stress response in adult males with psychopathic traits, as well as abnormalities in how this system may interact with the HPA axis. Copyright © 2015 Elsevier Ltd. All rights reserved.

Cortisol and Alpha-amylase changes during an Ultra-Running Event.

PubMed

Deneen, Whitney P; Jones, Alexis B

2017-01-01

Elevated stress hormone concentrations can positively affect an athlete's overall performance during a competition, and in many cases, are necessary to be able to perform exercise. During extreme exercise, the body's ability to utilize energy efficiently can affect an athlete's performance. Elevated hormonal concentrations can have many benefits in regards to an athlete's overall performance during a competition. The purpose of this study was to examine the effects of long distance running, such as seen during an ultra-running event (distances beyond 26.2 miles), on the activity of the hypothalamic-pituitary-adrenocortical (HPA) axis production of cortisol (CORT) as compared to autonomic nervous system production of salivary alpha-amylase (AA). Despite the well-known effects of exercise on CORT and AA response, it is unclear what effect running beyond the marathon distance has on these levels. This study investigates what effect long duration cardio exercise, such as running up to 100K (kilometers) distance, has on the neuroendocrine system, by means of saliva samples provided by participants signed up for an ultra-marathon event. The findings of this study show that the autonomic nervous system may present a response signal during physical stress that is independent of the HPA axis response. At distances beyond the marathon length, the production of CORT and AA was found to be suppressed for athletes, which could help them in their continued performance. Furthermore, this study recognizes a difference in the overall male and female response to stress in regards to CORT and AA production.

Alpha-Amylase Reactivity in Relation to Psychopathic Traits in Adults

PubMed Central

Glenn, Andrea L.; Remmel, Rheanna J.; Raine, Adrian; Schug, Robert A.; Gao, Yu; Granger, Douglas A.

2015-01-01

Recent investigations of the psychobiology of stress in antisocial youth have benefited from a multi-system measurement model. The inclusion of salivary alpha-amylase (sAA), a surrogate marker of autonomic/sympathetic nervous system (ANS) activity, in addition to salivary cortisol, a biomarker of the hypothalamic-pituitary-adrenal (HPA) axis functioning, has helped define a more complete picture of individual differences and potential dysfunction in the stress response system of these individuals. To the authors' knowledge, no studies have examined sAA in relation to antisocial behavior in adults or in relation to psychopathic traits specifically. In the present study, we examined sAA, in addition to salivary cortisol, in a relatively large sample (n = 158) of adult males (M age = 36.81, range = 22-67 years; 44% African-American, 34% Caucasian, 16% Hispanic) recruited from temporary employment agencies with varying levels of psychopathic traits. Males scoring highest in psychopathy were found to have attenuated sAA reactivity to social stress compared to those scoring lower in psychopathy. No differential relationships with the different factors of psychopathy were observed. In contrast to studies of antisocial youth, there were no interactions between sAA and cortisol levels in relation to psychopathy, but there was a significant interaction between pre-stressor levels of sAA and cortisol. Findings reveal potential regulatory deficits in the fast-acting, ‘fight or flight’, component of the stress response in adult males with psychopathic traits, as well as abnormalities in how this system may interact with the HPA axis. PMID:25662339

Gibberellin induces alpha-amylase gene in seed coat of Ipomoea nil immature seeds.

PubMed

Nakajima, Masatoshi; Nakayama, Akira; Xu, Zheng-Jun; Yamaguchi, Isomaro

2004-03-01

Two full-length cDNAs encoding gibberellin 3-oxidases, InGA3ox1 and InGA3ox2, were cloned from developing seeds of morning glory (Ipomoea nil (Pharbitis nil) Choisy cv. Violet) with degenerate-PCR and RACEs. The RNA-blot analysis for these clones revealed that the InGA3ox2 gene was organ-specifically expressed in the developing seeds at 6-18 days after anthesis. In situ hybridization showed the signals of InGA3ox2 mRNA in the seed coat, suggesting that active gibberellins (GAs) were synthesized in the tissue, although no active GA was detected there by immunohistochemistry. In situ hybridization analysis for InAmy1 (former PnAmy1) mRNA showed that InAmy1 was also synthesized in the seed coat. Both InGA3ox2 and InAmy1 genes were expressed spatially overlapped without a clear time lag, suggesting that both active GAs and InAmy1 were synthesized almost simultaneously in seed coat and secreted to the integument. These observations support the idea that GAs play an important role in seed development by inducing alpha-amylase.

Alpha-amylase inhibitory activity and sterol composition of the marine algae, Sargassum glaucescens

PubMed Central

Payghami, Nasrin; Jamili, Shahla; Rustaiyan, Abdolhossein; Saeidnia, Soodabeh; Nikan, Marjan; Gohari, Ahmad Reza

2015-01-01

Background: Sargassum species (phaeophyceae) are economically important brown algae in southern parts of Iran. Sargassum is mainly harvested as a row material in alginate production industries and is a source of plant foods or plant bio-stimulants even a component of animal foods. Objective: In this study, Sargassum glaucescens, collected from the seashore of Chabahar, was employed for phytochemical and biological evaluations. Materials and Methods: For that purpose, the dried algae was extracted by methanol and subjected to different chromatographic separation methods. Results: Six sterols, fucosterol (1), 24(S)-hydroxy-24-vinylcholesterol (2), 24(R)-hydroxy-24-vinylcholesterol (3), stigmasterol (4), β-sitosterol (5) and cholesterol (6) were identified by spectroscopic methods including 1H-NMR, 13C-NMR and mass spectroscopy. In vitro alpha-amylase inhibitory test was performed on the methanolic extract and the results revealed a potent inhibition (IC50 = 8.9 ± 2.4 mg/mL) of the enzyme compared to acarbose as a positive control. Conclusion: Various biological activities and distribution of sterols in Sargassum genus have been critically reviewed here. The results concluded that these algae are a good candidate for further anti-diabetic investigations in animals and human. PMID:26692744

Synthesis and properties of immobilized pectinase onto the macroporous polyacrylamide microspheres.

PubMed

Lei, Zhongli; Jiang, Qin

2011-03-23

Pectinase was covalently immobilized onto the macroporous polyacrylamide (PAM) microspheres synthesized via an inverse suspension polymerization approach, resulting in 81.7% immobilization yield. The stability of the macroporous PAM support, which has a large surface area, is not impeded by the adsorbed proteins despite the fact that up to 296.3 mg of enzyme is immobilized per gram of the carrier particles. The immobilized enzyme retained more than 75% of its initial activity over 30 days, and the optimum temperature/pH also increased to the range of 50-60 °C/3.0-5.0. The immobilized enzyme also exhibited great operational stability, and more than 75% residual activity was observed after 10 batch reactions. The kinetics of a model reaction catalyzed by the immobilized pectinase was finally investigated. Moreover, the immobilized pectinase could be recovered by centrifuging and showed durable activity at the process of recycle.

Pectinases From Sphenophorus levis Vaurie, 1978 (Coleoptera: Curculionidae): Putative Accessory Digestive Enzymes

PubMed Central

Evangelista, Danilo Elton; de Paula, Fernando Fonseca Pereira; Rodrigues, André; Henrique-Silva, Flávio

2015-01-01

The cell wall in plants offers protection against invading organisms and is mainly composed of the polysaccharides pectin, cellulose, and hemicellulose, which can be degraded by plant cell wall degrading enzymes (PCWDEs). Such enzymes are often synthesized by free living microorganisms or endosymbionts that live in the gut of some animals, including certain phytophagous insects. Thus, the ability of an insect to degrade the cell wall was once thought to be related to endosymbiont enzyme activity. However, recent studies have revealed that some phytophagous insects are able to synthesize their own PCWDEs by endogenous genes, although questions regarding the origin of these genes remain unclear. This study describes two pectinases from the sugarcane weevil, Sphenophorus levis Vaurie, 1978 (Sl-pectinases), which is considered one of the most serious agricultural pests in Brazil. Two cDNA sequences identified in a cDNA library of the insect larvae coding for a pectin methylesterase (PME) and an endo-polygalacturonase (endo-PG)—denominated Sl-PME and Sl-endoPG, respectively—were isolated and characterized. The quantitative real-time reverse transcriptase polymerase chain reaction expression profile for both Sl-pectinases showed mRNA production mainly in the insect feeding stages and exclusively in midgut tissue of the larvae. This analysis, together Western blotting data, suggests that Sl-pectinases have a digestive role. Phylogenetic analyses indicate that Sl-PME and Sl-endoPG sequences are closely related to bacteria and fungi, respectively. Moreover, the partial genomic sequences of the pectinases were amplified from insect fat body DNA, which was certified to be free of endosymbiotic DNA. The analysis of genomic sequences revealed the existence of two small introns with 53 and 166 bp in Sl-endoPG, which is similar to the common pattern in fungal introns. In contrast, no intron was identified in the Sl-PME genomic sequence, as generally observed in bacteria

Development of reproducible assays for polygalacturonase and pectinase.

PubMed

Li, Qian; Coffman, Anthony M; Ju, Lu-Kwang

2015-05-01

Polygalacturonase and pectinase activities reported in the literature were measured by several different procedures. These procedures do not give comparable results, partly owing to the complexity of the substrates involved. This work was aimed at developing consistent and efficient assays for polygalacturonase and pectinase activities, using polygalacturonic acid and citrus pectin, respectively, as the substrate. Different enzyme mixtures produced by Aspergillus niger and Trichoderma reesei with different inducing carbon sources were used for the method development. A series of experiments were conducted to evaluate the incubation time, substrate concentration, and enzyme dilution. Accordingly, for both assays the recommended (optimal) hydrolysis time is 30min and substrate concentration is 5g/L. For polygalacturonase, the sample should be adjusted to have 0.3-0.8U/mL polygalacturonase activity, because in this range the assay outcomes were consistent (independent of dilution factors). Such a range did not exist for the pectinase assay. The recommended procedure is to assay the sample at multiple (at least 2) dilution factors and determine, by linear interpolation, the dilution factor that would release reducing sugar equivalent to 0.4g/L d-galacturonic acid, and then calculate the activity of the sample accordingly (dilution factor×0.687U/mL). Validation experiments showed consistent results using these assays. Effects of substrate preparation methods were also examined. Copyright © 2015 Elsevier Inc. All rights reserved.

Temperature effect in the production of multiple xylanases by Aspergillus fumigatus.

PubMed

Lenartovicz, Veridiana; Marques de Souza, Cristina Giatti; Moreira, Fabiana Guillen; Peralta, Rosane Marina

2002-01-01

This work has evaluated the temperature effect in the production of multiple xylanases by a locally isolated strain of Aspergillus fumigatus Fresenius. Three isoenzymes, identified as xylanases I, II, and III with apparent molecular weight of 45.7 KDa, 39.8 KDa and 18.2 KDa, respectively, were produced in cultures developed at 30 degrees C and at 42 degrees C. The pattern of distribution of xylanase activity among the three isoenzymes was greatly affected by the growth temperature: at 30 degrees C, the total xylanase activity was distributed homogeneously among the three enzymes, while at 42 degrees C, the total xylanase activity was mainly due to the fractions with the highest MW (I and II) and the xylanase III was a minor component.

Rapid development of xylanase assay conditions using Taguchi methodology.

PubMed

Prasad Uday, Uma Shankar; Bandyopadhyay, Tarun Kanti; Bhunia, Biswanath

2016-11-01

The present investigation is mainly concerned with the rapid development of extracellular xylanase assay conditions by using Taguchi methodology. The extracellular xylanase was produced from Aspergillus niger (KP874102.1), a new strain isolated from a soil sample of the Baramura forest, Tripura West, India. Four physical parameters including temperature, pH, buffer concentration and incubation time were considered as key factors for xylanase activity and were optimized using Taguchi robust design methodology for enhanced xylanase activity. The main effect, interaction effects and optimal levels of the process factors were determined using signal-to-noise (S/N) ratio. The Taguchi method recommends the use of S/N ratio to measure quality characteristics. Based on analysis of the S/N ratio, optimal levels of the process factors were determined. Analysis of variance (ANOVA) was performed to evaluate statistically significant process factors. ANOVA results showed that temperature contributed the maximum impact (62.58%) on xylanase activity, followed by pH (22.69%), buffer concentration (9.55%) and incubation time (5.16%). Predicted results showed that enhanced xylanase activity (81.47%) can be achieved with pH 2, temperature 50°C, buffer concentration 50 Mm and incubation time 10 min.

Amylase - urine

MedlinePlus

... page: //medlineplus.gov/ency/article/003607.htm Amylase - urine To use the sharing features on this page, ... test that measures the amount of amylase in urine. Amylase is an enzyme that helps digest carbohydrates. ...

Synthesis and in vitro study of benzofuran hydrazone derivatives as novel alpha-amylase inhibitor.

PubMed

Taha, Muhammad; Shah, Syed Adnan Ali; Imran, Syahrul; Afifi, Muhammad; Chigurupati, Sridevi; Selvaraj, Manikandan; Rahim, Fazal; Ullah, Hayat; Zaman, Khalid; Vijayabalan, Shantini

2017-12-01

The α-amylase acts as attractive target to treat type-2 diabetes mellitus. Therefore in discovering a small molecule as α-amylase inhibitor, we have synthesized benzofuran carbohydrazide analogs (1-25), characterized through different spectroscopic techniques such as 1 HNMR and EI-MS. All screened analog shows good α-amylase inhibitory potentials with IC 50 value ranging between 1.078±0.19 and 2.926±0.05µM when compared with acarbose having IC 50 =0.62±0.22µM. Only nine analogs among the series such as analogs 3, 5, 7, 8, 10, 12, 21, 23 and 24 exhibit good inhibitory potential with IC 50 values 1.644±0.128, 1.078±0.19, 1.245±0.25, 1.843±0.19, 1.350±0.24, 1.629±0.015, 1.353±0.232, 1.359±0.119 and 1.488±0.07µM when compare with standard drug acarbose. All other analogs showed good to moderate α-amylase inhibitory potentials. The SAR study was conducted on the basis of substituent difference at the phenyl ring. The binding interaction between analogs and active site of enzyme was confirmed by docking studies. Copyright © 2017 Elsevier Inc. All rights reserved.

Potency of Amylase-producing Bacteria and Optimization Amylase Activities

NASA Astrophysics Data System (ADS)

Indriati, G.; Megahati, R. R. P.; Rosba, E.

2018-04-01

Enzymes are capable to act as biocatalyst for a wide variety of chemical reactions. Amylase have potential biotechnological applications in a wide range of industrial processes and account for nearly 30% of the world’s enzyme market. Amylase are extracellular enzymes that catalyze the hydrolysis of internal α-1,4-glycosidic linkages in starch to dextrin, and other small carbohydrate molecules constituted of glucose units. Although enzymes are produced from animal and plant sources, the microbial sources are generally the most suitable for commercial applications. Bacteria from hot springs is widely used as a source of various enzymes, such as amylase. But the amount of amylase-producing bacteria is still very limited. Therefore it is necessary to search sources of amylase-producing bacteria new, such as from hot springs Pariangan. The purpose of this study was to isolation of amylase-producing bacteria from Pariangan hot spring, West Sumatera and amylase activity optimization. The results were obtained 12 isolates of thermophilic bacteria and 5 isolates of amyalse-producing bacteria with the largest amylolytic index of 3.38 mm. The highest amylase activity was obtained at 50°C and pH 7.5.

Optimisation of low temperature extraction of banana juice using commercial pectinase.

PubMed

Sagu, Sorel Tchewonpi; Nso, Emmanuel Jong; Karmakar, Sankha; De, Sirshendu

2014-05-15

The objective of this work was to develop a process with optimum conditions for banana juice. The procedure involves hydrolyzing the banana pulp by commercial pectinase followed by cloth filtration. Response surface methodology with Doehlert design was utilised to optimize the process parameters. The temperature of incubation (30-60 °C), time of reaction (20-120 min) and concentration of pectinase (0.01-0.05% v/w) were the independent variables and viscosity, clarity, alcohol insoluble solids (AIS), total polyphenol and protein concentration were the responses. Total soluble sugar, pH, conductivity, calcium, sodium and potassium concentration in the juice were also evaluated. The results showed reduction of AIS and viscosity with reaction time and pectinase concentration and reduction of polyphenol and protein concentration with temperature. Using numerical optimization, the optimum conditions for the enzymatic extraction of banana juice were estimated. Depectinization kinetics was also studied at optimum temperature and variation of kinetic constants with enzyme dose was evaluated. Copyright © 2013 Elsevier Ltd. All rights reserved.

Environmental conditions affect transcription of the pectinase genes of Erwinia chrysanthemi 3937.

PubMed Central

Hugouvieux-Cotte-Pattat, N; Dominguez, H; Robert-Baudouy, J

1992-01-01

To depolymerize plant pectin, the phytopathogenic enterobacterium Erwinia chrysanthemi produces a series of enzymes which include a pectin-methyl-esterase encoded by the pem gene and five isoenzymes of pectate lyases encoded by the five genes pelA, pelB, pelC, pelD, and pelE. We have constructed transcriptional fusions between the pectinase gene promoters and the uidA gene, encoding beta-glucuronidase, to study the regulation of these E. chrysanthemi pectinase genes individually. The transcription of the pectinase genes is dependent on many environmental conditions. All the fusions were induced by pectic catabolic products and responded, to different degrees, to growth phase, catabolite repression, temperature, and nitrogen starvation. Transcription of pelA, pelD, and pelE was also increased in anaerobic growth conditions. High osmolarity of the culture medium increased expression of pelE but decreased that of pelD; the other pectinase genes were not affected. The level of expression of each gene was different. Transcription of pelA was very low under all growth conditions. The expression of the pelB, pelC, and pem genes was intermediate. The pelE gene had a high basal level of expression. Expression of pelD was generally the most affected by changes in culture conditions and showed a low basal level but very high induced levels. These differences in the expression of the pectinase genes of E. chrysanthemi 3937 presumably reflect their role during infection of plants, because the degradation of pectic polymers of the plant cell walls is the main determinant of tissue maceration caused by soft rot erwiniae. PMID:1447147

Preparation and properties of immobilized pectinase onto the amphiphilic PS-b-PAA diblock copolymers.

PubMed

Lei, Zhongli; Bi, Shuxian

2007-01-30

Well-defined amphiphilic block copolymers poly(styrene-b-acrylic acid) (PS-b-PAA) with controlled block length were synthesized using atom transfer radical polymerization (ATRP). Pectinase enzyme was immobilized on the well-defined amphiphilic block copolymers PS-b-PAA. The carboxyl groups on the amphiphilic PS-b-PAA diblock copolymers present a very simple, mild, and time-saving process for enzyme immobilization. Various characteristics of immobilized pectinase such as the pH and temperature stability, thermal stability, and storage stability were valuated. Among them the pH optimum and temperature optimum of free and immobilized pectinase were found to be pH 6.0 and 65 degrees C.

Molecular Cloning and Characterizations of Xylanase Inhibitor Protein from Wheat (Triticum Aestivum).

PubMed

Liu, Xinyu; Zhang, Yakun; Wei, Zhaohui; Chen, Hongge; Jia, Xincheng

2017-07-01

Xylanase inhibitor proteins (XIPs) were regarded to inhibit the activity of xylanases during baking and gluten-starch separation processes. To avoid the inhibition to xylanases, it is necessary to define the conditions under which the inhibition takes place. In this study, we cloned the XIP gene from 2 different variety of Triticum aestivum, that is, Zhengmai 9023 and Zhengmai 366, and investigated the properties of XIP protein expressed by Pichia pastoris. The results showed that the 2 XIP genes (xip-9023 and xip-366) were highly homologous with only 3 nucleotide differences. XIP-9023 showed the optimal inhibition pH and temperature were 7 °C and 40 °C, respectively. Inhibition of xylanase by XIP-9023 reached the maximum in 40 min. At 50% inhibition of xylanase, the molar ratio of inhibitor: xylanase was 26:1. XIP-9023 was active to various fungal xylanases tested as well as to a bacterial xylanase produced by Paenibacillus sp. isolated from cow rumen. © 2017 Institute of Food Technologists®.

Horizontal Gene Transfer of Pectinases from Bacteria Preceded the Diversification of Stick and Leaf Insects

PubMed Central

Shelomi, Matan; Danchin, Etienne G. J.; Heckel, David; Wipfler, Benjamin; Bradler, Sven; Zhou, Xin; Pauchet, Yannick

2016-01-01

Genes acquired by horizontal transfer are increasingly being found in animal genomes. Understanding their origin and evolution requires knowledge about the phylogenetic relationships from both source and recipient organisms. We used RNASeq data and respective assembled transcript libraries to trace the evolutionary history of polygalacturonase (pectinase) genes in stick insects (Phasmatodea). By mapping the distribution of pectinase genes on a Polyneoptera phylogeny, we identified the transfer of pectinase genes from known phasmatodean gut microbes into the genome of an early euphasmatodean ancestor that took place between 60 and 100 million years ago. This transfer preceded the rapid diversification of the suborder, enabling symbiont-free pectinase production that would increase the insects’ digestive efficiency and reduce dependence on microbes. Bacteria-to-insect gene transfer was thought to be uncommon, however the increasing availability of large-scale genomic data may change this prevailing notion. PMID:27210832

Psychosocial determinants of diurnal alpha-amylase among healthy Quebec workers.

PubMed

Marchand, Alain; Juster, Robert-Paul; Lupien, Sonia J; Durand, Pierre

2016-04-01

Salivary alpha-amylase (sAA) is a stress-sensitive biomarker the shows promise as an indirect proxy of sympathetic-adrenal-medullary axis activities that are otherwise difficult to discern non-invasively. This comprehensive study investigated diurnal sAA in association with numerous psychosocial characteristics related to mental health, work stress, and non-work stress. Participants included 395 workers (56.1% women, age: M=41.3, SD=10.81) from across 34 distinct workplaces. Diurnal sAA was sampled over two non-consecutive work days at awakening, 30 min after awakening, 14h00, 16h00, and bedtime. Well-validated psychometrics and survey items were used to measure mental health (psychological distress, depression, burnout, work characteristics) (task design, demands, social relations, gratifications), and non-work characteristics (marital/parental status, economic statuses, marital and parental stress, work-family conflicts). Preliminary results revealed that men showed occasionally higher sAA concentrations than women. Multilevel regressions were used to analyze sAA concentrations nested according to levels (i) for each time-point, (ii) between workers, and (iii) across workplaces while covarying for time of awakening, sex, age, cigarette smoking, alcohol consumption, regular physical activity, psychotropic drug use, and body mass index. Main results revealed that psychological demands, support from colleagues, interpersonal conflicts, job recognition and job insecurity appear to be associated with diurnal sAA, while non-work factors did not. Our findings showing a distinct diurnal profile for sAA replicate and expand those of Nater et al. (2007, Psychoneuroendocrinology 32, 392-401), providing further evidence that sAA is associated to subjective psychosocial factors. Copyright © 2016 Elsevier Ltd. All rights reserved.

Four rice seed cDNA clones belonging to the alpha-amylase/trypsin inhibitor gene family encode potential rice allergens.

PubMed

Alvarez, A M; Fukuhara, E; Nakase, M; Adachi, T; Aoki, N; Nakamura, R; Matsuda, T

1995-07-01

Four rice seed proteins encoded by cDNAs belonging to the alpha-amylase/trypsin inhibitor gene family were overexpressed as TrpE-fusion proteins in E. coli. The expressed rice proteins were detected by SDS-PAGE as major proteins in bacterial cell lysates. Western blot analyses showed that all the recombinant proteins were immunologically reactive to rabbit polyclonal antibodies and to a mouse monoclonal antibody (25B9) specific for a previously isolated rice allergen of 16 kDa. Some truncated proteins from deletion mutants of the cDNAs retained their reactivity to the specific antibodies. These results suggest that the cDNAs encode potential rice allergens and that some epitopes of the recombinant proteins are still immunoreactive when they are expressed as their fragments.

Salivary flow and alpha-amylase: collection technique, duration, and oral fluid type.

PubMed

Beltzer, Emilie K; Fortunato, Christine K; Guaderrama, Melissa M; Peckins, Melissa K; Garramone, Bianca M; Granger, Douglas A

2010-09-01

There has been renewed interest in salivary alpha-amylase (sAA), a surrogate marker of autonomic/sympathetic activity, in biosocial research on stress vulnerability, reactivity, and recovery. This study explored the impact of saliva flow rate on sAA measurement by examining the influence of (1) the technique used to collect oral fluid-synthetic swab, cotton pledget, hydrocellulose microsponge, or passive drool; (2) collection point duration--the length of time the technique is employed (1-5min); and (3) oral fluid type--whole unstimulated saliva (not absorbed by any material) or oral fluid sampled from areas near the parotid, submandibular, or sublingual salivary glands. sAA activity (U/mL) was the highest in oral fluid collected from the parotid and submandibular gland areas. The volume (mL) of oral fluid collected increased, and the activity of sAA (U/mL) decreased, as collection point duration lengthened. The magnitude of these effects varied according to collection technique and oral fluid type. Across all conditions, there were positive correlations (range .70-.88) between sAA activity (U/mL) and sAA output (U/min). Management of these potential sources of measurement error will be essential to ensuring the success of future research on the correlates and concomitants of sAA activity, stress-related reactivity and recovery, and diurnal variation. Copyright 2010 Elsevier Inc. All rights reserved.

Mechanism of removal of undesirable residual amylase, insoluble starch, and select colorants from refinery streams by powdered activated carbons

USDA-ARS?s Scientific Manuscript database

There is a need in the world-wide sugar industry to find a practical and economical solution to remove or inactivate residual alpha-amylases that are high temperature stable from factory or refinery streams. A survey of refineries that used amylase and had activated carbon systems for decolorization...

Structural analysis of xylanase inhibitor protein I (XIP-I), a proteinaceous xylanase inhibitor from wheat (Triticum aestivum, var. Soisson).

PubMed Central

Payan, Françoise; Flatman, Ruth; Porciero, Sophie; Williamson, Gary; Juge, Nathalie; Roussel, Alain

2003-01-01

A novel class of proteinaceous inhibitors exhibiting specificity towards microbial xylanases has recently been discovered in cereals. The three-dimensional structure of xylanase inhibitor protein I (XIP-I) from wheat (Triticum aestivum, var. Soisson) was determined by X-ray crystallography at 1.8 A (1 A=0.1 nm) resolution. The inhibitor possesses a (beta/alpha)(8) barrel fold and has structural features typical of glycoside hydrolase family 18, namely two consensus regions, approximately corresponding to the third and fourth barrel strands, and two non-proline cis -peptide bonds, Ser(36)-Phe and Trp(256)-Asp (in XIP-I numbering). However, detailed structural analysis of XIP-I revealed several differences in the region homologous with the active site of chitinases. The catalytic glutamic acid residue of family 18 chitinases [Glu(127) in hevamine, a chitinase/lysozyme from the rubber tree (Hevea brasiliensis)] is conserved in the structure of the inhibitor (Glu(128)), but its side chain is fully engaged in salt bridges with two neighbouring arginine residues. Gly(81), located in subsite -1 of hevamine, where the reaction intermediate is formed, is replaced by Tyr(80) in XIP-I. The tyrosine side chain fills the subsite area and makes a strong hydrogen bond with the side chain of Glu(190) located at the opposite side of the cleft, preventing access of the substrate to the catalytic glutamic acid. The structural differences in the inhibitor cleft structure probably account for the lack of activity of XIP-I towards chitin. PMID:12617724

Production of plant cell wall degrading enzymes by monoculture and co-culture of Aspergillus niger and Aspergillus terreus under SSF of banana peels.

PubMed

Rehman, Shazia; Aslam, Hina; Ahmad, Aqeel; Khan, Shakeel Ahmed; Sohail, Muhammad

2014-01-01

Filamentous fungi are considered to be the most important group of microorganisms for the production of plant cell wall degrading enzymes (CWDE), in solid state fermentations. In this study, two fungal strains Aspergillus niger MS23 and Aspergillus terreus MS105 were screened for plant CWDE such as amylase, pectinase, xylanase and cellulases (β-glucosidase, endoglucanase and filterpaperase) using a novel substrate, Banana Peels (BP) for SSF process. This is the first study, to the best of our knowledge, to use BP as SSF substrate for plant CWDE production by co-culture of fungal strains. The titers of pectinase were significantly improved in co-culture compared to mono-culture. Furthermore, the enzyme preparations obtained from monoculture and co-culture were used to study the hydrolysis of BP along with some crude and purified substrates. It was observed that the enzymatic hydrolysis of different crude and purified substrates accomplished after 26 h of incubation, where pectin was maximally hydrolyzed by the enzyme preparations of mono and co-culture. Along with purified substrates, crude materials were also proved to be efficiently degraded by the cocktail of the CWDE. These results demonstrated that banana peels may be a potential substrate in solid-state fermentation for the production of plant cell wall degrading enzymes to be used for improving various biotechnological and industrial processes.

Production of plant cell wall degrading enzymes by monoculture and co-culture of Aspergillus niger and Aspergillus terreus under SSF of banana peels

PubMed Central

Rehman, Shazia; Aslam, Hina; Ahmad, Aqeel; Khan, Shakeel Ahmed; Sohail, Muhammad

2014-01-01

Filamentous fungi are considered to be the most important group of microorganisms for the production of plant cell wall degrading enzymes (CWDE), in solid state fermentations. In this study, two fungal strains Aspergillus niger MS23 and Aspergillus terreus MS105 were screened for plant CWDE such as amylase, pectinase, xylanase and cellulases (β-glucosidase, endoglucanase and filterpaperase) using a novel substrate, Banana Peels (BP) for SSF process. This is the first study, to the best of our knowledge, to use BP as SSF substrate for plant CWDE production by co-culture of fungal strains. The titers of pectinase were significantly improved in co-culture compared to mono-culture. Furthermore, the enzyme preparations obtained from monoculture and co-culture were used to study the hydrolysis of BP along with some crude and purified substrates. It was observed that the enzymatic hydrolysis of different crude and purified substrates accomplished after 26 h of incubation, where pectin was maximally hydrolyzed by the enzyme preparations of mono and co-culture. Along with purified substrates, crude materials were also proved to be efficiently degraded by the cocktail of the CWDE. These results demonstrated that banana peels may be a potential substrate in solid-state fermentation for the production of plant cell wall degrading enzymes to be used for improving various biotechnological and industrial processes. PMID:25763058

Extractive fermentation of xylanase from Aspergillus tamarii URM 4634 in a bioreactor.

PubMed

da Silva, Anna Carolina; Soares de França Queiroz, Alana Emília; Evaristo dos Santos Nascimento, Talita Camila; Rodrigues, Cristine; Gomes, José Erick Galindo; Souza-Motta, Cristina Maria; Porto de Souza Vandenberghe, Luciana; Valente de Medeiros, Erika; Moreira, Keila Aparecida; Herculano, Polyanna Nunes

2014-08-01

Of the many reported applications for xylanase, its use as a food supplement has played an important role for monogastric animals, because it can improve the utilisation of nutrients. The aim of this work was to produce xylanase by extractive fermentation in an aqueous two-phase system using Aspergillus tamarii URM 4634, increasing the scale of production in a bioreactor, partially characterising the xylanase and evaluating its influence on monogastric digestion in vitro. Through extractive fermentation in a bioreactor, xylanase was obtained with an activity of 331.4 U mL(-1) and 72% yield. The xylanase was stable under variable pH and temperature conditions, and it was optimally active at pH 3.6 and 90 °C. Xylanase activity potentiated the simulation of complete monogastric digestion by 6%, and only Mg2+ inhibited its activity. This process provides a system for efficient xylanase production by A. tamarii URM 4634 that has great potential for industrial use.

Close evolutionary relatedness among functionally distantly related members of the (alpha/beta)8-barrel glycosyl hydrolases suggested by the similarity of their fifth conserved sequence region.

PubMed

Janecek, S

1995-12-11

A short conserved sequence equivalent to the fifth conserved sequence region of alpha-amylases (173_LPDLD, Aspergillus oryzae alpha-amylase) comprising the calcium-ligand aspartate, Asp-175, was identified in the amino acid sequences of several members of the family of (alpha/beta)8-barrel glycosyl hydrolases. Despite the fact that the aspartate is not invariantly conserved, the stretch can be easily recognised in all sequences to be positioned 26-28 amino acid residues in front of the well-known catalytic aspartate (Asp-206, A. oryzae alpha-amylase) located in the beta 4-strand of the barrel. The identification of this region revealed remarkable similarities between some alpha-amylases (those from Bacillus megaterium, Bacillus subtilis and Dictyoglomus thermophilum) on the one hand and several different enzyme specificities (such as oligo-1,6-glucosidase, amylomaltase and neopullulanase, respectively) on the other hand. The most interesting example was offered by B. subtilis alpha-amylase and potato amylomaltase with the regions LYDWN and LYDWK, respectively. These observations support the idea that all members of the family of glycosyl hydrolases adopting the structure of the alpha-amylase-type (alpha/beta)8-barrel are mutually closely related and the strict evolutionary borders separating the individual enzyme specificities can be hardly defined.

Evaluation of certain food additives and contaminants.

PubMed

2004-01-01

This report represents the conclusions of a Joint FAO/WHO Expert Committee convened to evaluate the safety of various food additives, with a view to recommending acceptable daily intakes (ADIs) and to prepare specifications for the identity and purity of food additives. The first part of the report contains a general discussion of the principles governing the toxicological evaluation of food additives (including flavouring agents) and contaminants, assessments of intake, and the establishment and revision of specifications for food additives. A summary follows of the Committee's evaluations of toxicological and intake data on various specific food additives (alpha-amylase from Bacillus lichenformis containing a genetically engineered alpha-amylase gene from B. licheniformis, annatto extracts, curcumin, diacetyl and fatty acid esters of glycerol, D-tagatose, laccase from Myceliophthora thermophila expressed in Aspergillus oryzae, mixed xylanase, beta-glucanase enzyme preparation produced by a strain of Humicola insolens, neotame, polyvinyl alcohol, quillaia extracts and xylanase from Thermomyces lanuginosus expressed in Fusarium venenatum), flavouring agents, a nutritional source of iron (ferrous glycinate, processed with citric acid), a disinfectant for drinking-water (sodium dichloroisocyanurate) and contaminants (cadmium and methylmercury). Annexed to the report are tables summarizing the Committee's recommendations for ADIs of the food additives, recommendations on the flavouring agents considered, and tolerable intakes of the contaminants considered, changes in the status of specifications and further information requested or desired.

Subsite mapping of enzymes. Application of the depolymerase computer model to two alpha-amylases.

PubMed Central

Allen, J D; Thoma, J A

1976-01-01

In the preceding paper (Allen and Thoma, 1976) we developed a depolymerase computer model, which uses a minimization routine to establish a subsite map for a depolymerase. In the present paper we show how the model is applied to experimental data for two alpha-amylases. Michaelis parameters and bond-cleavage frequencies for substrates of chain lengths up to twelve glucosyl units have been reported for Bacillus amyloliquefaciens, and a subsite map has been proposed for this enzyme [Thoma et al. (1971) J. Biol. Chem. 246, 5621-5635]. By applying the computer model to the experimental data, we have arrived at a ten-subsite map. We find that a significant improvement in this map is achieved by allowing the hydrolytic rate coefficient to vary as a function of the number of occupied subsites comprising the enzyme-binding region. The bond-cleavage frequencies, the enzyme is found to have eight subsites. A partial subsite map is arrived at, but the entire binding region cannot be mapped because Michaelis parameters are complicated by transglycosylation reactions. The hydrolytic rate coefficients for this enzyme are not constant. PMID:999630

Production optimization of a heat-tolerant alkaline pectinase from Bacillus subtilis ZGL14 and its purification and characterization.

PubMed

Yu, Ping; Zhang, Yishu; Gu, Donglu

2017-09-03

Alkaline pectinase has important applications in the pretreatment of waste water from food processing and in both the fabric and paper industries. In this study, a 2-level factorial design was used to screen significant factors that affect the activity of alkaline pectinase, and the response surface methodology (RSM) with a Box-Behnken design (BBD) was used to optimize their concentrations. Starch, peptone, KH 2 PO 4 and K 2 HPO 4 ·3H 2 O were found to significantly affect the activity of alkaline pectinase. Their optimal concentrations were as follows: 4.68% starch, 1.6% peptone, 0.26% KH 2 PO 4 and 0.68% K 2 HPO 4 ·3H 2 O. Under the above conditions, the activity of alkaline pectinase was significantly improved to 734.11 U/mL. Alkaline pectinase was purified to homogeneity with a recovery rate of 9.6% and a specific activity of 52372.52 U/mg. Its optimal temperature and pH were 50°C and 8.6, respectively. The purified enzyme showed strong thermo-stability and good alkali resistance. In addition, the activity of alkaline pectinase was improved in the presence of Mg 2+ . Cu 2+ , Mn 2+ , and Co 2+ significantly inhibited its activity. This study provides an important basis for the future development and use of a heat-tolerant alkaline pectinase from B. subtilis ZGL14.

Production optimization of a heat-tolerant alkaline pectinase from Bacillus subtilis ZGL14 and its purification and characterization

PubMed Central

Yu, Ping; Zhang, Yishu; Gu, Donglu

2017-01-01

ABSTRACT Alkaline pectinase has important applications in the pretreatment of waste water from food processing and in both the fabric and paper industries. In this study, a 2-level factorial design was used to screen significant factors that affect the activity of alkaline pectinase, and the response surface methodology (RSM) with a Box-Behnken design (BBD) was used to optimize their concentrations. Starch, peptone, KH2PO4 and K2HPO4·3H2O were found to significantly affect the activity of alkaline pectinase. Their optimal concentrations were as follows: 4.68% starch, 1.6% peptone, 0.26% KH2PO4 and 0.68% K2HPO4·3H2O. Under the above conditions, the activity of alkaline pectinase was significantly improved to 734.11 U/mL. Alkaline pectinase was purified to homogeneity with a recovery rate of 9.6% and a specific activity of 52372.52 U/mg. Its optimal temperature and pH were 50°C and 8.6, respectively. The purified enzyme showed strong thermo-stability and good alkali resistance. In addition, the activity of alkaline pectinase was improved in the presence of Mg2+. Cu2+, Mn2+, and Co2+ significantly inhibited its activity. This study provides an important basis for the future development and use of a heat-tolerant alkaline pectinase from B. subtilis ZGL14. PMID:28282260

Pectinase production by Aspergillus niger using banana (Musa balbisiana) peel as substrate and its effect on clarification of banana juice.

PubMed

Barman, Sumi; Sit, Nandan; Badwaik, Laxmikant S; Deka, Sankar C

2015-06-01

Optimization of substrate concentration, time of incubation and temperature for crude pectinase production from A. niger was carried out using Bhimkol banana (Musa balbisiana) peel as substrate. The crude pectinase produced was partially purified using ethanol and effectiveness of crude and partially purified pectinase was studied for banana juice clarification. The optimum substrate concentration, incubation time and temperature of incubation were 8.07 %, 65.82 h and 32.37 °C respectively, and the polygalacturonase (PG) activity achieved was 6.6 U/ml for crude pectinase. The partially purified enzyme showed more than 3 times of polygalacturonase activity as compared to the crude enzyme. The SDS-PAGE profile showed that the molecular weight of proteins present in the different pectinases varied from 34 to 42 kDa. The study further revealed that highest clarification was achieved when raw banana juice was incubated for 60 min with 2 % concentration of partially purified pectinase and the absorbance obtained was 0.10.

Salivary cortisol, alpha-amylase and immunoglobulin a responses to a morning session of basketball or volleyball training in boys aged 14-18 years.

PubMed

Bruzda-Zwiech, A; Konieczka, M; Hilt, A; Daszkowska, M; Grzegorczyk, J; Szczepańska, J

2017-01-01

This study investigates whether a single session of routine morning basketball or volleyball training affects saliva levels of cortisol, alpha-amylase (sAA) and secretory immunoglobulin A (sIgA) in boys aged 14–18 years. Twenty-nine boys who participate in basketball or volleyball training, recruited from the Marcin Gortat’s Athletic Championship School in Lodz, were enrolled in the study. The 90-minute routine exercise program included 15 minutes of warm-up followed by basketball or volleyball practice. Unstimulated saliva samples were collected prior to and immediately after the exercise, and were analysed using ELISA. One training session resulted in a significant increase of sAA concentration in all participants, as well as in the volleyball and basketball subgroups (p=0.00022; p=0.0029; p=0.0011; respectively). Post-exercise cortisol levels were significantly lower than pre-exercise levels (p=0.00002) throughout the group, as well as in the volleyball and basketball subgroups (p=0.0048; p=0.0019; p=0.0048; respectively). The exercise protocol did not significantly affect sIgA level, either in the whole examined group or the volleyball subgroup, however a weak significant increase of sIgA was observed in the basketball subgroup (p=0.046). The routine morning training session comprising a warm-up followed by basketball or volleyball practice seems to activate the sympatho-adrenal-medullary system, with a subsequent increase of alpha-amylase, but does not affect oral immunity in 14-18-year-old boys.

Purification and biochemical characterization of a thermostable and acid-stable alpha-amylase from Bacillus licheniformis B4-423.

PubMed

Wu, Xiangrong; Wang, Yuxia; Tong, Bending; Chen, Xianghua; Chen, Jianhua

2018-04-01

Novel thermostable amylase need to be continuously explored with the improvement of industrial requirements. A new acidophilic and thermostable amylase producing bacterium isolated from spring was identified as Bacillus strain on the basis of 16S rDNA. The amylase was purified by ammonium sulphate precipitation, gel chromatography and anion exchange chromatography. SDS-PAGE revealed that the enzyme was monomeric with a molecular weight of 58 kDa. The amylase exhibited optimal activity at pH 5.0 and temperature 100 °C. Then the enzyme showed high stability in pH ranges 4.0-10.0 and more than 90% of maximal activity was found from 20 °C to 80 °C. Apart from good stability toward SDS and non-ionic detergent, the purified enzyme exhibited high compatibility with some inhibitors such as urea and EDTA. The results demonstrated the stability of the enzyme in different organic solvents. Moreover, we determined the amylase gene, compared the structure with α-amylase BAA and BLA and found some thermostability determinants in our enzyme. Overall, presenting various properties were including high thermostability, Ca 2+ -independency, broad temperature and pH profiles, organic-solvent tolerance as well as excellent stability with detergents. Such characteristics have not been reported for this type of enzyme, and the α-amylase will be a suitable candidate in industrial fields. Copyright © 2017 Elsevier B.V. All rights reserved.

Self-compassion training modulates alpha-amylase, heart rate variability, and subjective responses to social evaluative threat in women

PubMed Central

Arch, Joanna J.; Brown, Kirk Warren; Dean, Derek J.; Landy, Lauren N.; Brown, Kimberley; Laudenslager, Mark L.

2014-01-01

A growing body of research has revealed that social evaluative stressors trigger biological and psychological responses that in chronic forms have been linked to aging and disease. Recent research suggests that self-compassion may protect the self from typical defensive responses to evaluation. We investigated whether brief training in self-compassion moderated biopsychological responses to the Trier Social Stress Test (TSST) in women. Compared to attention (placebo) and no-training control conditions, brief self-compassion training diminished sympathetic (salivary alpha-amylase), cardiac parasympathetic, and subjective anxiety responses, though not HPA-axis (salivary cortisol) responses to the TSST. Self-compassion training also led to greater self-compassion under threat relative to the control groups. In that social stress pervades modern life, self-compassion represents a promising approach to diminishing its potentially negative psychological and biological effects. PMID:24636501

Screening and production study of microbial xylanase producers from Brazilian Cerrado.

PubMed

Alves-Prado, Heloiza Ferreira; Pavezzi, Fabiana Carina; Leite, Rodrigo Simões Ribeiro; de Oliveira, Valéria Maia; Sette, Lara Durães; Dasilva, Roberto

2010-05-01

Hemicelluloses are polysaccharides of low molecular weight containing 100 to 200 glycosidic residues. In plants, the xylans or the hemicelluloses are situated between the lignin and the collection of cellulose fibers underneath. The xylan is the most common hemicellulosic polysaccharide in cell walls of land plants, comprising a backbone of xylose residues linked by beta-1,4-glycosidic bonds. So, xylanolytic enzymes from microorganism have attracted a great deal of attention in the last decade, particularly because of their biotechnological characteristics in various industrial processes, related to food, feed, ethanol, pulp, and paper industries. A microbial screening of xylanase producer was carried out in Brazilian Cerrado area in Selviria city, Mato Grosso do Sul State, Brazil. About 50 bacterial strains and 15 fungal strains were isolated from soil sample at 35 degrees C. Between these isolated microorganisms, a bacterium Lysinibacillus sp. and a fungus Neosartorya spinosa as good xylanase producers were identified. Based on identification processes, Lysinibacillus sp. is a new species and the xylanase production by this bacterial genus was not reported yet. Similarly, it has not reported about xylanase production from N. spinosa. The bacterial strain P5B1 identified as Lysinibacillus sp. was cultivated on submerged fermentation using as substrate xylan, wheat bran, corn straw, corncob, and sugar cane bagasse. Corn straw and wheat bran show a good xylanase activity after 72 h of fermentation. A fungus identified as N. spinosa (strain P2D16) was cultivated on solid-state fermentation using as substrate source wheat bran, wheat bran plus sawdust, corn straw, corncob, cassava bran, and sugar cane bagasse. Wheat bran and corncobs show the better xylanase production after 72 h of fermentation. Both crude xylanases were characterized and a bacterial xylanase shows optimum pH for enzyme activity at 6.0, whereas a fungal xylanase has optimum pH at 5.0-5.5. They were

Salivary cortisol and alpha-amylase levels during an assessment procedure correlate differently with risk-taking measures in male and female police recruits

PubMed Central

van den Bos, Ruud; Taris, Ruben; Scheppink, Bianca; de Haan, Lydia; Verster, Joris C.

2013-01-01

Recent laboratory studies have shown that men display more risk-taking behavior in decision-making tasks following stress, whilst women are more risk-aversive or become more task-focused. In addition, these studies have shown that sex differences are related to levels of the stress hormone cortisol (indicative of activation of the hypothalamus-pituitary-adrenocortical-axis): the higher the levels of cortisol the more risk-taking behavior is shown by men, whereas women generally display more risk-aversive or task-focused behavior following higher levels of cortisol. Here, we assessed whether such relationships hold outside the laboratory, correlating levels of cortisol obtained during a job-related assessment procedure with decision-making parameters in the Cambridge Gambling Task (CGT) in male and female police recruits. The CGT allows for discriminating different aspects of reward-based decision-making. In addition, we correlated levels of alpha-amylase [indicative of activation of the sympatho-adrenomedullary-axis (SAM)] and decision-making parameters. In line with earlier studies men and women only differed in risk-adjustment in the CGT. Salivary cortisol levels correlated positively and strongly with risk-taking measures in men, which was significantly different from the weak negative correlation in women. In contrast, and less strongly so, salivary alpha-amylase levels correlated positively with risk-taking in women, which was significantly different from the weak negative correlation with risk-taking in men. Collectively, these data support and extend data of earlier studies indicating that risky decision-making in men and women is differently affected by stress hormones. The data are briefly discussed in relation to the effects of stress on gambling. PMID:24474909

Xylanase production by a newly isolated Aspergillus niger SS7 in submerged culture.

PubMed

Bakri, Yasser; Al-Jazairi, Manal; Al-Kayat, Ghassan

2008-01-01

Xylanase production by a newly isolated Aspergillus niger SS7 was studied in submerged culture. The optimum initial pH for xylanase production was found to be 7.0. Different agricultural and industrial wastes were evaluated for their ability to induce xylanase production by this isolate. The best xylanase production (293.82 IU/ml) was recorded at 3% (w/v) corn cob hulls after 120 h of incubation. The Aspergillus niger SS7 isolate grown in a simple medium, proved to be a promising microorganism for xylanase production.

Gender differences in automatic thoughts and cortisol and alpha-amylase responses to acute psychosocial stress in patients with obsessive-compulsive personality disorder.

PubMed

Kanehisa, Masayuki; Kawashima, Chiwa; Nakanishi, Mari; Okamoto, Kana; Oshita, Harumi; Masuda, Koji; Takita, Fuku; Izumi, Toshihiko; Inoue, Ayako; Ishitobi, Yoshinobu; Higuma, Haruka; Ninomiya, Taiga; Akiyoshi, Jotaro

2017-08-01

Obsessive-compulsive personality disorder (OCPD) has a pervasive pattern of preoccupation with orderliness, perfection, and mental and interpersonal control at the expense of flexibility, openness, and efficiency. The aims of the present study were to explore the relationship between OCPD and psychological stress and psychological tests. We evaluated 63 OCPD patients and 107 healthy controls (HCs). We collected saliva samples from patients and controls before and after a social stress procedure, the Trier Social Stress Test (TSST), to measure the concentrations of salivary alpha-amylase (sAA) and salivary cortisol. The Childhood Trauma Questionnaire (CTQ), Profile of Mood State (POMS), State-Trait Anxiety Inventory (STAI), Beck Depression Inventory (BDI), Social Adaptation Self-Evaluation Scale (SASS), and Depression and Anxiety Cognition Scale (DACS) were administered to patients and HCs. Following TSST exposure, the salivary amylase and cortisol levels were significantly decreased in male patients compared with controls. Additionally, OCPD patients had higher CTQ, POMS, STAI, and BDI scores than HCs and exhibited significantly higher anxiety and depressive states. OCPD patients scored higher on future denial and threat prediction as per the DACS tool. According to a stepwise regression analysis, STAI, POMS, and salivary cortisol responses were independent predictors of OCPD. Our results suggested that attenuated sympathetic and parasympathetic reactivity in male OCPD patients occurs along with attenuated salivary amylase and cortisol responses to the TSST. In addition, there was a significant difference between OCPD patients and HCs in child trauma, mood, anxiety, and cognition. The finding support the modeling role of cortisol (20min) on the relationships between STAI trait and depression among OCPD. Copyright © 2017 Elsevier B.V. All rights reserved.

An alkaline active xylanase: insights into mechanisms of high pH catalytic adaptation.

PubMed

Mamo, Gashaw; Thunnissen, Marjolein; Hatti-Kaul, Rajni; Mattiasson, Bo

2009-09-01

The alkaliphilic bacterium, Bacillus halodurans S7, produces an alkaline active xylanase (EC 3.2.1.8), which differs from many other xylanases in being operationally stable under alkaline conditions as well as at elevated temperature. Compared to non-alkaline active xylanases, this enzyme has a high percent composition of acidic amino acids which results in high ratio of negatively to positively charged residues. A positive correlation was observed between the charge ratio and the pH optima of xylanases. The recombinant xylanase was crystallized using a hanging drop diffusion method. The crystals belong to the space group P2(1)2(1)2(1) and the structure was determined at a resolution of 2.1 A. The enzyme has the common eight-fold TIM-barrel structure of family 10 xylanases; however, unlike non-alkaline active xylanases, it has a highly negatively charged surface and a deeper active site cleft. Mutational analysis of non-conserved amino acids which are close to the acid/base residue has shown that Val169, Ile170 and Asp171 are important to hydrolyze xylan at high pH. Unlike the wild type xylanase which has optimum pH at 9-9.5, the triple mutant xylanase (V169A, I170F and D171N), which was constructed using sequence information of alkaline sensitive xylanses was optimally active around pH 7. Compared to non-alkaline active xylanases, the alkaline active xylanases have highly acidic surfaces and fewer solvent exposed alkali labile residues. Based on these results obtained from sequence, structural and mutational analysis, the possible mechanisms of high pH stability and catalysis are discussed. This will provide useful information to understand the mechanism of high pH adaptation and engineering of enzymes that can be operationally stable at high pH.

Cortisol and alpha amylase reactivity and timing of puberty: Vulnerabilities for antisocial behaviour in young adolescents

PubMed Central

Susman, Elizabeth J.; Granger, Douglas A; Blades, Keeva T.; Randazzo, William; Heaton, Jodi A.; Dorn, Lorah D.

2009-01-01

The theoretical framework proposed that cortisol and saliva alpha amylase (sAA) reactivitiy are vulnerabilities for antisocial behaviour. These indices of hypothalamic-pituitary-adrenal (HPA) and sympathetic-adrenal-medulary (SAM) components of the stress system, respectively, were considered vulnerabilities that also interact with the putative stressful transition of timing of puberty to predispose adolescents toward antisocial behaviour. The sample consisted of 8- to-13-year-old boys and girls (N=135) and a parent. For boys, timing of puberty moderated the association between cortisol and sAA reactivity and antisocial behaviour. Higher cortisol reactivity in later timing boys was related to a composite index of antisocial behaviour and rule-breaking behaviour problems. In contrast, lower sAA reactivity and earlier timing of puberty in boys was related to rule breaking and conduct disorder symptoms. The interaction between timing of puberty and HPA or SAM regulation and timing of puberty in boys suggests that reproductive, neuroendocrine mechanisms may be involved in the extensively documented adverse consequences of off-time pubertal development. PMID:19819639

Cortisol and alpha amylase reactivity and timing of puberty: vulnerabilities for antisocial behaviour in young adolescents.

PubMed

Susman, Elizabeth J; Dockray, Samantha; Granger, Douglas A; Blades, Keeva T; Randazzo, William; Heaton, Jodi A; Dorn, Lorah D

2010-05-01

The theoretical framework proposed that cortisol and saliva alpha amylase (sAA) reactivitiy are vulnerabilities for antisocial behaviour. These indices of hypothalamic-pituitary-adrenal (HPA) and sympathetic-adrenal-medulary (SAM) components of the stress system, respectively, were considered vulnerabilities that also interact with the putative stressful transition of timing of puberty to predispose adolescents toward antisocial behaviour. The sample consisted of 8- to-13-year-old boys and girls (N=135) and a parent. For boys, timing of puberty moderated the association between cortisol and sAA reactivity and antisocial behaviour. Higher cortisol reactivity in later timing boys was related to a composite index of antisocial behaviour and rule-breaking behaviour problems. In contrast, lower sAA reactivity and earlier timing of puberty in boys was related to rule breaking and conduct disorder symptoms. The interaction between timing of puberty and HPA or SAM regulation and timing of puberty in boys suggests that reproductive, neuroendocrine mechanisms may be involved in the extensively documented adverse consequences of off-time pubertal development. Copyright 2009 Elsevier Ltd. All rights reserved.

FT-Raman spectroscopic analysis of enhanced activity of supercritical carbon dioxide treated bacterial alpha-amylase.

PubMed

Paul, Kaninika; Dutta, Sayantani; Bhattacharjee, Paramita

2017-09-01

Our previous investigation on high pressure supercritical carbon dioxide treatment of a bacterial α-amylase had revealed enhanced activity of the same. 1 H NMR analysis of the activity enhanced enzyme led the authors to hypothesize that the enhancement was possibly owing to alterations in the active site of the enzyme. In the present study, the changes in the active site of the treated enzyme was analysed by Fourier-transform Raman (FT-Raman) spectroscopy. The spectra obtained revealed shifting of bands in the active site of α-amylase indicating a nudging effect of the bonds in this region consequent to high pressure treatment. Also, shifts in bands in the OH stretching vibration of water were observed in the enzyme spectra. These variations in the spectra confirmed changes in the active site as well as in the water associated with the same that perhaps had a concerted effect on the increased activity of α-amylase. Copyright © 2017 Elsevier Inc. All rights reserved.

Enhancement of heterogeneous alkaline xylanase production in Pichia pastoris GS115

NASA Astrophysics Data System (ADS)

Zheng, Wei

2017-08-01

A series of strategies were applied to improve expression level of the recombinant alkaline xylanase from Bacillus pumilus G1-3 in Pichia pastoris GS115. Codon optimization of xylanase gene xynG1-3 from B. pumilus G1-3 were carried out for its heterogeneous expression in P. pastoris. The activity of xylanase encoded by optimized gene (xynG1-3-opt) was up to 33641 U/mL, which was 37% higher than that by wild-type (xynG1-3) gene. The results will greatly contribute to increasing the production of recombinant proteins in P. pastoris and improving the industrial production of the alkaline xylanase.

Structural features of immunostimulatory polysaccharide purified from pectinase hydrolysate of barley leaf.

PubMed

Kim, Hoon; Kwak, Bong-Shin; Hong, Hee-Do; Suh, Hyung-Joo; Shin, Kwang-Soon

2016-06-01

Four polysaccharide fractions were isolated from young barley leaves treated with or without pectinase followed by ethanol fractionation. Among the polysaccharide fractions, BLE-P isolated from pectinase digested with a high molecular weight had the most enhanced macrophage stimulatory activity, indicating that pectinase digestion of barley leaf is a useful method for enhancement of its activity. BLE-P was further purified by column chromatography to identify the chemical and structural properties. BLE-P-I eluted in void volume fraction showed potent macrophage stimulatory activity. Monosaccharide composition and linkage analysis indicated that at least three kinds of polysaccharide, that is, glucuronoarabinoxylan (GAX; 40-45%), rhamnogalacturonan-I (RG-I) with branching mainly involving a type II arabinogalactan (AG-II) side chain (30-35%), and linear glucan such as starch and cellulose (less than 10%) coexisted in BLE-P-I. Given the association with macrophage stimulatory activity, it is likely that the GAX and to the RG-I polysaccharide branched with an AG-II side chain may be important for expression of the activity in barley leaf. Copyright © 2016 Elsevier B.V. All rights reserved.

Gene cloning, overexpression, and characterization of a xylanase from Penicillium sp. CGMCC 1669.

PubMed

Liu, Wanli; Shi, Pengjun; Chen, Qiang; Yang, Peilong; Wang, Guozeng; Wang, Yaru; Luo, Huiying; Yao, Bin

2010-09-01

A xylanase-encoding gene, xyn11F63, was isolated from Penicillium sp. F63 CGMCC1669 using degenerated polymerase chain reaction (PCR) and thermal asymmetric interlaced (TAIL)-PCR techniques. The full-length chromosomal gene consists of 724 bp, including a 73-bp intron, and encodes a 217 amino acid polypeptide. The deduced amino acid sequence of xyn11F63 shows the highest identity of 70% to the xylanase from Penicillium sp. strain 40, which belongs to glycosyl hydrolases family 11. The gene was overexpressed in Pichia pastoris, and its activity in the culture medium reached 516 U ml(-1). After purification to electrophoretic homogeneity, the enzyme showed maximal activity at pH 4.5 and 40 degrees C, was stable at acidic buffers of pH 4.5-9.0, and was resistant to proteases (proteinase K, trypsin, subtilisin A, and alpha-chymotrypsin). The specific activity, K (m), and V (max) for oat spelt xylan substrate was 7,988 U mg(-1), 22.2 mg ml(-1), and 15,105.7 micromol min(-1) mg(-1), respectively. These properties make XYN11F63 a potential economical candidate for use in feed and food industrial applications.

Detergent-compatible bacterial amylases.

PubMed

Niyonzima, Francois N; More, Sunil S

2014-10-01

Proteases, lipases, amylases, and cellulases are enzymes used in detergent formulation to improve the detergency. The amylases are specifically supplemented to the detergent to digest starchy stains. Most of the solid and liquid detergents that are currently manufactured contain alkaline enzymes. The advantages of using alkaline enzymes in the detergent formulation are that they aid in removing tough stains and the process is environmentally friendly since they reduce the use of toxic detergent ingredients. Amylases active at low temperature are preferred as the energy consumption gets reduced, and the whole process becomes cost-effective. Most microbial alkaline amylases are used as detergent ingredients. Various reviews report on the production, purification, characterization, and application of amylases in different industry sectors, but there is no specific review on bacterial or fungal alkaline amylases or detergent-compatible amylases. In this mini-review, an overview on the production and property studies of the detergent bacterial amylases is given, and the stability and compatibility of the alkaline bacterial amylases in the presence of the detergents and the detergent components are highlighted.

Exo-pectinase production by Bacillus pumilus using different agricultural wastes and optimizing of medium components using response surface methodology.

PubMed

Tepe, Ozlem; Dursun, Arzu Y

2014-01-01

In this research, the production of exo-pectinase by Bacillus pumilus using different agricultural wastes was studied. Agricultural wastes containing pectin such as wheat bran, sugar beet pulp, sunflower plate, orange peel, banana peel, apple pomace and grape pomace were tested as substrates, and activity of exo-pectinase was determined only in the mediums containing sugar beet pulp and wheat bran. Then, effects of parameters such as concentrations of solid substrate (wheat bran and sugar beet pulp) (A), ammonium sulphate (B) and yeast extract (C) on the production of exo-pectinase were investigated by response surface methodology. First, wheat bran was used as solid substrate, and it was determined that exo-pectinase activity increased when relatively low concentrations of ammonium sulphate (0.12-0.21% w/v) and yeast extract (0.12-0.3% w/v) and relatively high wheat bran (~5-6% w/v) were used. Then, exo-pectinase production was optimized by response surface methodology using sugar beet pulp as a solid substrate. In comparison to P values of the coefficients, values of not greater than 0.05 of A and B (2) showed that the effect of these process variables in exo-pectinase production was important and that changes done in these variables will alter the enzyme activity.

Cloning and Characterization of an Alpha-amylase Gene from the Hyperthermophilic Archaeon Thermococcus Thioreducens

NASA Technical Reports Server (NTRS)

Bernhardsdotter, Eva C. M. J.; Pusey, Marc L.; Ng, Joseph D.; Garriott, Owen K.

2004-01-01

The gene encoding an extracellular a-amylase, TTA, from the hyperthermophilic archaeon Thermococcus thioreducens was cloned and expressed in Escherichia coli. Primary structural analysis revealed high similarity with other a-amylases from the Thermococcus and Pyrococcus genera, as well as the four highly conserved regions typical for a-amylases. The 1374 bp gene encodes a protein of 457 amino acids, of which 435 constitute the mature protein preceded by a 22 amino acid signal peptide. The molecular weight of the purified recombinant enzyme was estimated to be 43 kDa by denaturing gel electrophoresis. Maximal enzymatic activity of recombinant TTA was observed at 90 C and pH 5.5 in the absence of exogenous Ca(2+), and the enzyme was considerably stable even after incubation at 90 C for 2 hours. The thermostability at 90 and 102 C was enhanced in the presence of 5 mM Ca(2+). The extraordinarily high specific activity (about 7.4 x 10(exp 3) U/mg protein at 90 C, pH 5.5 with soluble starch as substrate) together with its low pH optimum makes this enzyme an interesting candidate for starch processing applications.

Individual Differences in Preschoolers' Salivary Cortisol and Alpha-Amylase Reactivity: Relations to Temperament and Maladjustment

PubMed Central

Spinrad, Tracy L.; Eisenberg, Nancy; Granger, Douglas A.; Eggum, Natalie D.; Sallquist, Julie; Haugen, RG; Kupfer, Anne; Hofer, Claire

2009-01-01

We examined the relations of 84 preschoolers' (43 boys; mean age = 54 months) situational stress reactivity to their observed emotions and mothers' reports of temperament and adjustment. Salivary cortisol and salivary alpha-amylase (sAA) were collected prior to, and following, a frustrating task. Children's anger, sadness, and positive affect were measured, and mothers reported on preschoolers' dispositional emotionality, regulation, impulsivity, and problem behaviors. Forty-seven percent of children had an increase in sAA and 52% had an increase in cortisol following the challenging task. On average, sAA levels showed the predicted pattern of rise following the frustrating task, followed by return to baseline. For cortisol, there was a mean increase from pre-task to 40 minutes post-test. sAA reactivity was associated with relatively low levels of dispositional anger and impulsivity and relatively high regulation, particularly for girls. sAA reactivity also was related to low externalizing problems for girls, but not boys. Although cortisol reactivity was unrelated to children's emotions and maladjustment, it was positively related to mothers' reports of regulation. The findings suggest that sAA reactivity in response to a frustrating social task may reflect girls' constrained behavior. PMID:19348808

Individual differences in preschoolers' salivary cortisol and alpha-amylase reactivity: relations to temperament and maladjustment.

PubMed

Spinrad, Tracy L; Eisenberg, Nancy; Granger, Douglas A; Eggum, Natalie D; Sallquist, Julie; Haugen, R G; Kupfer, Anne; Hofer, Claire

2009-06-01

We examined the relations of 84 preschoolers' (43 boys; mean age=54 months) situational stress reactivity to their observed emotions and mothers' reports of temperament and adjustment. Salivary cortisol and salivary alpha-amylase (sAA) were collected prior to, and following, a frustrating task. Children's anger, sadness, and positive affect were measured, and mothers reported on preschoolers' dispositional emotionality, regulation, impulsivity, and problem behaviors. Forty-seven percent of children had an increase in sAA and 52% had an increase in cortisol following the challenging task. On average, sAA levels showed the predicted pattern of rise following the frustrating task, followed by return to baseline. For cortisol, there was a mean increase from pre-task to 40 min post-test. sAA reactivity was associated with relatively low levels of dispositional anger and impulsivity and relatively high regulation, particularly for girls. sAA reactivity also was related to low externalizing problems for girls, but not boys. Although cortisol reactivity was unrelated to children's emotions and maladjustment, it was positively related to mothers' reports of regulation. The findings suggest that sAA reactivity in response to a frustrating social task may reflect girls' constrained behavior.

Invariant glycines and prolines flanking in loops the strand beta 2 of various (alpha/beta)8-barrel enzymes: a hidden homology?

PubMed Central

Janecek, S.

1996-01-01

The question of parallel (alpha/beta)8-barrel fold evolution remains unclear, owing mainly to the lack of sequence homology throughout the amino acid sequences of (alpha/beta)8-barrel enzymes. The "classical" approaches used in the search for homologies among (alpha/beta)8-barrels (e.g., production of structurally based alignments) have yielded alignments perfect from the structural point of view, but the approaches have been unable to reveal the homologies. These are proposed to be "hidden" in (alpha/beta)8-barrel enzymes. The term "hidden homology" means that the alignment of sequence stretches proposed to be homologous need not be structurally fully satisfactory. This is due to the very long evolutionary history of all (alpha/beta)8-barrels. This work identifies so-called hidden homology around the strand beta 2 that is flanked by loops containing invariant glycines and prolines in 17 different (alpha/beta)8-barrel enzymes, i.e., roughly in half of all currently known (alpha/beta)8-barrel proteins. The search was based on the idea that a conserved sequence region of an (alpha/beta)8-barrel enzyme should be more or less conserved also in the equivalent part of the structure of the other enzymes with this folding motif, given their mutual evolutionary relatedness. For this purpose, the sequence region around the well-conserved second beta-strand of alpha-amylase flanked by the invariant glycine and proline (56_GFTAIWITP, Aspergillus oryzae alpha-amylase numbering), was used as the sequence-structural template. The proposal that the second beta-strand of (alpha/beta)8-barrel fold is important from the evolutionary point of view is strongly supported by the increasing trend of the observed beta 2-strand structural similarity for the pairs of (alpha/beta)8-barrel enzymes: alpha-amylase and the alpha-subunit of tryptophan synthase, alpha-amylase and mandelate racemase, and alpha-amylase and cyclodextrin glycosyltransferase. This trend is also in agreement with the

Invariant glycines and prolines flanking in loops the strand beta 2 of various (alpha/beta)8-barrel enzymes: a hidden homology?

PubMed

Janecek, S

1996-06-01

The question of parallel (alpha/beta)8-barrel fold evolution remains unclear, owing mainly to the lack of sequence homology throughout the amino acid sequences of (alpha/beta)8-barrel enzymes. The "classical" approaches used in the search for homologies among (alpha/beta)8-barrels (e.g., production of structurally based alignments) have yielded alignments perfect from the structural point of view, but the approaches have been unable to reveal the homologies. These are proposed to be "hidden" in (alpha/beta)8-barrel enzymes. The term "hidden homology" means that the alignment of sequence stretches proposed to be homologous need not be structurally fully satisfactory. This is due to the very long evolutionary history of all (alpha/beta)8-barrels. This work identifies so-called hidden homology around the strand beta 2 that is flanked by loops containing invariant glycines and prolines in 17 different (alpha/beta)8-barrel enzymes, i.e., roughly in half of all currently known (alpha/beta)8-barrel proteins. The search was based on the idea that a conserved sequence region of an (alpha/beta)8-barrel enzyme should be more or less conserved also in the equivalent part of the structure of the other enzymes with this folding motif, given their mutual evolutionary relatedness. For this purpose, the sequence region around the well-conserved second beta-strand of alpha-amylase flanked by the invariant glycine and proline (56_GFTAIWITP, Aspergillus oryzae alpha-amylase numbering), was used as the sequence-structural template. The proposal that the second beta-strand of (alpha/beta)8-barrel fold is important from the evolutionary point of view is strongly supported by the increasing trend of the observed beta 2-strand structural similarity for the pairs of (alpha/beta)8-barrel enzymes: alpha-amylase and the alpha-subunit of tryptophan synthase, alpha-amylase and mandelate racemase, and alpha-amylase and cyclodextrin glycosyltransferase. This trend is also in agreement with the

Synergistic effect of cellulase and xylanase during hydrolysis of natural lignocellulosic substrates.

PubMed

Song, Hui-Ting; Gao, Yuan; Yang, Yi-Min; Xiao, Wen-Jing; Liu, Shi-Hui; Xia, Wu-Cheng; Liu, Zi-Lu; Yi, Li; Jiang, Zheng-Bing

2016-11-01

Synergistic combination of cellulase and xylanase has been performed on pre-treated substrates in many previous studies, while few on natural substrates. In this study, three unpretreated lignocellulosic substrates were studied, including corncob, corn stover, and rice straw. The results indicated that when the mixed cellulase and xylanase were applied, reducing sugar concentrations were calculated as 19.53, 15.56, and 17.35mg/ml, respectively, based on the 3,5 dinitrosalicylic acid (DNS) method. Compared to the treatment with only cellulose, the hydrolysis yields caused by mixed cellulase and xylanase were improved by 133%, 164%, and 545%, respectively. In addition, the conversion yield of corncob, corn stover, and rice straw by cellulase-xylanase co-treatment reached 43.9%, 48.5%, and 40.2%, respectively, based on HPLC analysis, which confirmed the synergistic effect of cellulase-xylanase that was much higher than either of the single enzyme treatment. The substrate morphology was also evaluated to explore the synergistic mechanism of cellulase-xylanase. Copyright © 2016 Elsevier Ltd. All rights reserved.

Heart rate, salivary α-amylase activity, and cooperative behavior in previously naïve children receiving dental local anesthesia.

PubMed

Arhakis, Aristidis; Menexes, George; Coolidge, Trilby; Kalfas, Sotirios

2012-01-01

Psychosomatic indicators, such as heart rate (HR), salivary alpha amylase (sAA) activity, and behavior, can be used to determine stress. This study's aim was to assess the pattern of changes of salivary alpha amylase, heart rate, and cooperative behavior in previously naïve children receiving dental treatment under local anesthesia. Included were 30 children with no prior dental experience who needed 4 or more sessions of dental treatment involving local anesthesia. In each session, sAA, HR, and behavior were assessed before and during the application of local anesthesia and at the end of the treatment. The highest sAA value was always observed at the end of each session; overall, the value was lower in the fourth session. HR always increased during the local anesthesia, and did not vary across sessions. No significant relationship was found between child cooperation and either sAA or HR. In this sample, child cooperation may not be an accurate indicator of stress. Based on salivary alpha amylase activity changes, dental treatment involving local anesthesia in naïve children appeared to be less stressful after 3 sessions.

The pectinases from Sphenophorus levis: Potential for biotechnological applications.

PubMed

Habrylo, Olivier; Evangelista, Danilo Elton; Castilho, Priscila Vasques; Pelloux, Jérôme; Henrique-Silva, Flávio

2018-06-01

Pectinases represent about one fifth of the enzyme worldwide market due their wide range of biotechnological applications. Current commercial pectinases are exclusively obtained from microbial sources, but here we report a pectin methylesterase (Sl-PME) and an endo-polygalacturonase (Sl-EPG) bioprospected from the sugarcane weevil, Sphenophorus levis, which revealed good potential for industrial applications. Sl-PME and Sl-EPG were overexpressed in Pichia pastoris, purified and enzymatically characterized. Sl-EPG presents optimal activity at pH 4-5 and 50 °C, showing that it can be used for juice extraction and clarification. On the other hand, Sl-PME presents optimal activity at pH 6-8 and 40 °C, and thus, suitable for both acidic and alkaline processing, such as coffee and tea fermentation. Sl-EPG shows V max  = 3.23 mM/min, K M  = 2.4 g/L and k cat  = 418.6 s -1 . While Sl-PME shows V max  = 0.14 mM/min, K M  = 4.1 g/L and k cat  = 1.7 s -1 . A PG inhibitor (PGIP2) weakly interfered in the Sl-EPG activity and Sl-PME was not affected by a usual PME inhibitor. Moreover, these enzymes manifested synergistic action towards methylesterified pectin. Here, we propose these enzymes as novel alternative tools for the current commercial pectinases. Copyright © 2018 Elsevier B.V. All rights reserved.

Characterization, analysis, and application of fabricated Fe3O4-chitosan-pectinase nanobiocatalyst.

PubMed

Seenuvasan, Muthulingam; Kumar, Kannaiyan Sathish; Malar, Carlin Geor; Preethi, Sridhar; Kumar, Madhava Anil; Balaji, Nagarajan

2014-03-01

The investigation on fabrication of Fe3O4-chitosan-pectinase nanobiocatalyst was performed by covalently binding the pectinase onto carboxyl group activated chitosan-coated magnetic nanoparticles (CMNPs). The morphological and size distribution analysis of the different magnetic nanoparticles (MNPs) was done using transmission electron microscopy (TEM), and the average diameter was 11.07 ± 3.04, 11.55 ± 3.16, and 11.59 ± 3.16 nm for MNPs, CMNPs, and fabricated nanobiocatalyst, respectively, suggesting that there was no significant change in the size of MNPs after coating and binding. The characteristic peaks occurred at 2θ of 30.39, 35.43, 43.37, 57.22, and 62.9, and their corresponding indices 220, 311, 400, 520, and 441 for different MNPs from the X-ray diffraction (XRD) studies confirmed the presence of Fe3O4 with the spinel structure, and there was no phase change even after coating and binding. The various required characteristic absorption peaks (575, 585, 1,563, 1,614, 1,651, and 1,653 cm(-1)) from Fourier transform infrared (FT-IR) spectroscopy confirmed the surface modifications and binding of pectinase onto the MNPs. At the weight ratio of about 19.8 × 10(-3) mg bound pectinase/mg activated CMNPs, the activity of fabricated nanobiocatalyst was found to be maximum. In order to monitor their improved activity, the pH, temperature, reusability, storage ability, and kinetic studies were established.

Site-directed mutagenesis at aspartate and glutamate residues of xylanase from Bacillus pumilus.

PubMed Central

Ko, E P; Akatsuka, H; Moriyama, H; Shinmyo, A; Hata, Y; Katsube, Y; Urabe, I; Okada, H

1992-01-01

To elucidate the reaction mechanism of xylanase, the identification of amino acids essential for its catalysis is of importance. Studies have indicated the possibility that the reaction mechanism of xylanase is similar to that of hen's egg lysozyme, which involves acidic amino acid residues. On the basis of this assumption, together with the three-dimensional structure of Bacillus pumilus xylanase and its amino acid sequence similarity to other xylanases of different origins, three acidic amino acids, namely Asp-21, Glu-93 and Glu-182, were selected for site-directed mutagenesis. The Asp residue was altered to either Ser or Glu, and the Glu residues to Ser or Asp. The purified mutant xylanases D21E, D21S, E93D, E93S, E182D and E182S showed single protein bands of about 26 kDa on SDS/PAGE. C.d. spectra of these mutant enzymes show no effect on the secondary structure of xylanase, except that of D21E, which shows a little variation. Furthermore, mutations of Glu-93 and Glu-182 resulted in a drastic decrease in the specific activity of xylanase as compared with mutation of Asp-21. On the basis of these results we propose that Glu-93 and Glu-182 are the best candidates for the essential catalytic residues of xylanase. Images Fig. 1. Fig. 4 Fig. 5 PMID:1359880

Cloning, expression and characterization of a metagenome derived thermoactive/thermostable pectinase.

PubMed

Singh, Rajvinder; Dhawan, Samriti; Singh, Kashmir; Kaur, Jagdeep

2012-08-01

The gene encoding a thermostable pectinase was isolated from a soil metagenome sample. The gene sequence corresponded to an open reading frame of 1,311 bp encoding a translation product of 47.9 kDa. It showed maximum (93 %) identity to a Bacillus licheniformis glycoside hydrolase. Deduced amino acid analysis showed an absence of highly conserved cysteine residues in the N-terminal region at positions 24 and 42, and in the C-terminal region at positions 389, 394, 413 and 424. pQpecJKR01 (pQE30 expression vector containing the pectinase gene) was expressed in Escherichia coli strain M15 as a recombinant fusion protein containing an N-terminal 6× His tag. Biochemical properties of this pectinase were novel. The enzyme had temperature and pH optima of 70 °C and 7.0, respectively, but was active over a broad temperature and pH range. The enzyme was stable at 60 °C with a half-life of 5 h and the enzyme activity was inhibited by 0.1 % diethyl pyrocarbonate and 5 mM dicyclohexyl carbodiimide. The enzyme could be of great use in industrial processes due to its activity over a broad pH range and at high temperature.

Immunochemical Studies on α-Amylase III. Immunochemical Relationships Among Amylases from Various Microorganisms1

PubMed Central

Sirisinha, Stitaya; Allen, Peter Z.

1965-01-01

Sirishinha, Stitaya (University of Rochester School of Medicine and Dentistry, Rochester, N.Y.), and Peter Z. Allen. Immunochemical studies on α-amylase. III. Immunochemical relationships among amylases from various microorganisms. J. Bacteriol. 90:1120–1128. 1965.—Immunochemical relationships among amylases obtained from a selected group of microorganisms were examined, and a cross-reaction was detected between the α-amylases of Bacillus stearothermophilus and B. subtilis. Immunodiffusion and quantitative precipitin studies, as well as cross-neutralization tests, indicate that B. stearothermophilus α-amylase reacts with a portion of antibody present in antisera to crystalline B. subtilis α-amylase. Amylases from these two species thus have some aspects of structure in common. Limited data obtained by immunodiffusion suggest that groupings which confer cross-reactivity to the B. stearothermophilus enzyme are lost after exposure to mercaptoethanol in the presence of ethylenediamine-tetraacetate, followed by treatment with iodoacetamide. With the antisera employed and within the concentration range examined, no immunochemical cross-reaction was observed among amylases from Aspergillus oryzae, B. subtilis, B. polymyxa, B. macerans, Pseudomonas saccharophila, and Euglena sanguinis. Immunoelectrophoresis of partially purified B. stearothermophilus α-amylase by use of antiserum to the crude enzyme, together with localization of amylase activity in immunoelectrophoretic plates, suggests that B. stearothermophilus α-amylase is antigenic in the rabbit. Images PMID:5847799

Immobilization of pectinase from Leucoagaricus gongylophorus on magnetic particles.

PubMed

Adalberto, Paulo Roberto; José dos Santos, Francisco; Golfeto, Camilla Calemi; Costa Iemma, Mônica Rosas; Ferreira de Souza, Dulce Helena; Cass, Quezia Bezerra

2012-10-21

Polygalacturonases (EC 3.2.1.15) hydrolyze the α-1,4-glycosidic linkages in polygalacturonic acid chains. The interest on specific inhibitors of pectinase and the versatility of magnetic support for enzyme immobilization endorsed the preparation of an immobilized enzyme reactor (IMER). This work presents the synthesis of CoFe(2)O(4) amino-derivatives, which was employed as the support for the immobilization of pectinases from Leucoagaricus gongylophorus. Amino-functionalized CoFe(2)O(4) was obtained from glyceryl-derivatized CoFe(2)O(4) and was characterized by infrared spectroscopy and electronic microscopy. The immobilized enzyme maintained the same thermal, chemical and kinetic behaviour of the free enzyme (T(opt) 60 °C; pH(opt) 5.0; K(app)(M) = 0.5 mg min(-1); V(app)(M) ≈ 5.0 μmol min(-1) mL(-1)). The straightforward synthesis of CoFe(2)O(4) derivatives and the efficiency of immobilization offer wide perspectives for the use of the developed new IMER.

Optimization of Enzyme Co-Immobilization with Sodium Alginate and Glutaraldehyde-Activated Chitosan Beads.

PubMed

Gür, Sinem Diken; İdil, Neslihan; Aksöz, Nilüfer

2018-02-01

In this study, two different materials-alginate and glutaraldehyde-activated chitosan beads-were used for the co-immobilization of α-amylase, protease, and pectinase. Firstly, optimization of multienzyme immobilization with Na alginate beads was carried out. Optimum Na alginate and CaCl 2 concentration were found to be 2.5% and 0.1 M, respectively, and optimal enzyme loading ratio was determined as 2:1:0.02 for pectinase, protease, and α-amylase, respectively. Next, the immobilization of multiple enzymes on glutaraldehyde-activated chitosan beads was optimized (3% chitosan concentration, 0.25% glutaraldehyde with 3 h of activation and 3 h of coupling time). While co-immobilization was successfully performed with both materials, the specific activities of enzymes were found to be higher for the enzymes co-immobilized with glutaraldehyde-activated chitosan beads. In this process, glutaraldehyde was acting as a spacer arm. SEM and FTIR were used for the characterization of activated chitosan beads. Moreover, pectinase and α-amylase enzymes immobilized with chitosan beads were also found to have higher activity than their free forms. Three different enzymes were co-immobilized with these two materials for the first time in this study.

[Comparative characteristics of microbial proteases by the level of hydrolysis of protein substrates].

PubMed

Rimareva, L V; Overchenko, M B; Serba, E M; Trifonova, V V

1997-01-01

Screening of enzyme preparations displaying a maximum proteolytic activity at pH 4.0-5.5 and effecting deep proteolysis of plant proteins was performed. Amyloprotooryzin prepared from Aspergillus oryzae 387 containing a complex of proteolytic enzymes was the most effective. The amino acid composition of the hydrolysates obtained was studied. Amyloprotooryzin increased the contents of amino acids by 108-227%, depending on the substrate used. The enzymatic complex of amyloprotooryzin was studied; in addition, proteases, alpha-amylase, exo-beta-glucanase, and xylanase were detected in the complex.

Similarity of different beta-strands flanked in loops by glycines and prolines from distinct (alpha/beta)8-barrel enzymes: chance or a homology?

PubMed Central

Janecek, S.

1995-01-01

Many (alpha/beta)8-barrel enzymes contain their conserved sequence regions at or around the beta-strand segments that are often preceded and succeeded by glycines and prolines, respectively. alpha-Amylase is one of these enzymes. Its sequences exhibit a very low degree of similarity, but strong conservation is seen around its beta-strands. These conserved regions were used in the search for similarities with beta-strands of other (alpha/beta)8-barrel enzymes. The analysis revealed an interesting similarity between the segment around the beta 2-strand of alpha-amylase and the one around the beta 4-strand of glycolate oxidase that are flanked in loops by glycines and prolines. The similarity can be further extended on other members of the alpha-amylase and glycolate oxidase subfamilies, i.e., cyclodextrin glycosyltransferase and oligo-1,6-glucosidase, and flavocytochrome b2, respectively. Moreover, the alpha-subunit of tryptophan synthase, the (alpha/beta)8-barrel enzyme belonging to the other subfamily of (alpha/beta)8-barrels, has both investigated strands, beta 2 and beta 4, similar to beta 2 of alpha-amylase and beta 4 of glycolate oxidase. The possibilities of whether this similarity exists only by chance or is a consequence of some processes during the evolution of (alpha/beta)8-barrel proteins are briefly discussed. PMID:7549888

Similarity of different beta-strands flanked in loops by glycines and prolines from distinct (alpha/beta)8-barrel enzymes: chance or a homology?

PubMed

Janecek, S

1995-06-01

Many (alpha/beta)8-barrel enzymes contain their conserved sequence regions at or around the beta-strand segments that are often preceded and succeeded by glycines and prolines, respectively. alpha-Amylase is one of these enzymes. Its sequences exhibit a very low degree of similarity, but strong conservation is seen around its beta-strands. These conserved regions were used in the search for similarities with beta-strands of other (alpha/beta)8-barrel enzymes. The analysis revealed an interesting similarity between the segment around the beta 2-strand of alpha-amylase and the one around the beta 4-strand of glycolate oxidase that are flanked in loops by glycines and prolines. The similarity can be further extended on other members of the alpha-amylase and glycolate oxidase subfamilies, i.e., cyclodextrin glycosyltransferase and oligo-1,6-glucosidase, and flavocytochrome b2, respectively. Moreover, the alpha-subunit of tryptophan synthase, the (alpha/beta)8-barrel enzyme belonging to the other subfamily of (alpha/beta)8-barrels, has both investigated strands, beta 2 and beta 4, similar to beta 2 of alpha-amylase and beta 4 of glycolate oxidase. The possibilities of whether this similarity exists only by chance or is a consequence of some processes during the evolution of (alpha/beta)8-barrel proteins are briefly discussed.

Direct purification of pectinase from mango (Mangifera Indica Cv. Chokanan) peel using a PEG/salt-based Aqueous Two Phase System.

PubMed

Mehrnoush, Amid; Sarker, Md Zaidul Islam; Mustafa, Shuhaimi; Yazid, Abdul Manap Mohd

2011-10-10

An Aqueous Two-Phase System (ATPS) was employed for the first time for the separation and purification of pectinase from mango (Mangifera Indica Cv. Chokanan) peel. The effects of different parameters such as molecular weight of the polymer (polyethylene glycol, 2,000-10,000), potassium phosphate composition (12-20%, w/w), system pH (6-9), and addition of different concentrations of neutral salts (0-8%, w/w) on partition behavior of pectinase were investigated. The partition coefficient of the enzyme was decreased by increasing the PEG molecular weight. Additionally, the phase composition showed a significant effect on purification factor and yield of the enzyme. Optimum conditions for purification of pectinase from mango peel were achieved in a 14% PEG 4000-14% potassium phosphate system using 3% (w/w) NaCl addition at pH 7.0. Based on this system, the purification factor of pectinase was increased to 13.2 with a high yield of (97.6%). Thus, this study proves that ATPS can be an inexpensive and effective method for partitioning of pectinase from mango peel.

Endocrine and emotional response to exclusion among women and men; cortisol, salivary alpha amylase, and mood.

PubMed

Helpman, Liat; Penso, Julia; Zagoory-Sharon, Orna; Feldman, Ruth; Gilboa-Schechtman, Eva

2017-05-01

Social exclusion is ubiquitous and painful. Evolutionary models indicate sex differences in coping with social stress. Recent empirical data suggest different sex patterns in hypothalamic-pituitary-adrenal (HPA) and sympathetic-adrenal-medullary (SAM) reactivity. The present study sought to test this hypothesis. We examined differences in endocrine and emotional response to exclusion by using a virtual ball tossing paradigm (Cyberball). Saliva samples and mood ratings were collected to reflect levels before, and repeatedly following, exclusion. The sample included 21 women and 23 men. Cortisol and salivary alpha amylase (sAA), biomarkers of the HPA and SAM systems, respectively, were used as indices of two arms of stress response. Following exclusion, all participants experienced mood worsening followed by mood improvement, with men reporting less distress than women. Women evinced decline in cortisol following the Cyberball task, whereas men's cortisol levels showed a non-significant rise, and then decline, following exclusion. Our results concur with previous findings showing SAM reactivity to be gender-neutral and HPA reactivity to be gender-divergent. Additional studies are needed to examine sex-specific response to social exclusion. Implications for individual differences in recovery from stress are discussed.

Xylanase XYN IV from Trichoderma reesei showing exo- and endo-xylanase activity

USDA-ARS?s Scientific Manuscript database

A novel xylanase from Trichoderma reesei Rut C30, named XYN IV, was purified from the cellulolytic system of the fungus. The enzyme was discovered on its ability to attack aldotetraohexenuronic acid (HexA-2Xyl-4Xyl-4Xyl, HexA3Xyl3), releasing the reducing-end xylose residue. XYN IV exhibited catalyt...

Children’s cortisol and salivary alpha-amylase interact to predict attention bias to threatening stimuli

PubMed Central

Ursache, Alexandra; Blair, Clancy

2017-01-01

Physiological responses to threat occur through both the autonomic nervous system (ANS) and the hypothalamic pituitary adrenal (HPA) axis. Activity in these systems can be measured through salivary alpha-amylase (sAA) and salivary cortisol, respectively. Theoretical work and empirical studies have suggested the importance of examining the coordination of these systems in relation to cognitive functioning and behavior problems. Less is known, however, about whether these systems interactively predict more automatic aspects of attention processing such as attention toward emotionally salient threatening stimuli. We used a dot probe task to assess attention bias toward threatening stimuli in 347 kindergarten children. Cortisol and sAA were assayed from saliva samples collected prior to children’s participation in assessments on a subsequent day. Using regression analyses, we examined relations of sAA and cortisol to attention bias. Results indicate that cortisol and sAA interact in predicting attention bias. Higher levels of cortisol predicted greater bias toward threat for children who had high levels of sAA, but predicted greater bias away from threat for children who had low levels of sAA. These results suggest that greater symmetry in HPA and ANS functioning is associated with greater reliance on automatic attention processes in the face of threat. PMID:25455863

Preparation using pectinase and characterization of nanofibers from orange peel waste in juice factories.

PubMed

Hideno, Akihiro; Abe, Kentaro; Yano, Hiroyuki

2014-06-01

This study reports the preparation and characterization of nanofibers consisting mainly of cellulose microfibrils from orange peel (OP), which is a significant byproduct of orange juice production. Three treatments (boiling, alkaline, and pectinase) were investigated with and without subsequent grinding treatment. It was possible to prepare the cellulose nanofibers (CNFs) using these methods, except for the boiling treatment with grinding. Interestingly, only pectinase and a mild-physical blender treatment without grinding produced nanofibers. The width of the nanofibers from OP was approximately 10 to 50 nm. The microfibril bundles of OP were considered to be thinner than those of commercial CNFs. Our data indicated that the removal of pectic polysaccharides and hemicelluloses covering the cellulose microfibrils was important for the preparation of nanofibers from OP. These nanofibers from OP using pectinase are proposed to be applicable as food materials, pharmaceuticals, and filters for the tractive characteristics of the sheet. This study demonstrates: (1) it was possible to prepare the nanofibers from orange peel using pectinase and (2) the width of the nanofibers from orange peel was approximately 10 to 50 nm. (3) Removal of polysaccharides such as pectin and hemicelluloses covering cellulose microfibrils was very important for preparation of nanofibers from OP. Considering the tractive characteristics of the sheets from nanofibers and the origin of orange peel, they are suitable for application of food materials, pharmaceuticals, and filters. © 2014 Institute of Food Technologists®

Effect of the lectins wheat germ agglutinin (WGA) and Ulex europaeus agglutinin (UEA-I) on the alpha-amylase secretion of rat pancreas in vitro and in vivo.

PubMed

Mikkat, U; Damm, I; Schröder, G; Schmidt, K; Wirth, C; Weber, H; Jonas, L

1998-05-01

Lectins are able to bind to cholecystokinin (CCK) receptors and other glycosylated membrane proteins. The lectins wheat germ agglutinin (WGA) and Ulex europaeus agglutinin (UEA-I) are used for affinity chromatography to isolate the highly glycosylated CCK-A receptor of pancreatic acinar cells. According to the working hypothesis that lectin binding to the CCK receptor should alter the ligand-receptor interaction, the effect of WGA and UEA-I on CCK-8-induced enzyme secretion was studied on isolated rat pancreatic acini in vitro. In vitro both lectins showed a dosage-dependent inhibition of CCK-8-induced alpha-amylase secretion of acini over 60 min. WGA showed a strong inhibitory effect on amylase secretion, approximately 40%, in vitro. UEA-I caused a smaller, but significant decrease, approximately 20%, in enzyme secretion of isolated acini. Additionally, both lectins inhibited cerulein/secretin- or cerulein-induced pancreatic secretion of rats in vivo, but not after secretin alone. The results are discussed with respect to a possible influence of both lectins on the interaction of CCK or cerulein with the CCK-A receptor.

Influences of AMY1 gene copy number and protein expression on salivary alpha-amylase activity before and after citric acid stimulation in splenic asthenia children.

PubMed

Yang, Zemin; Lin, Jing; Chen, Longhui; Zhang, Min; Yang, Xiaorong; Chen, Weiwen

2015-06-01

To compare the correlations between salivary alpha-amylase (sAA) activity and amylase, alpha 1 (salivary) gene (AMYl) copy number or its gene expression between splenic asthenia and healthy children, and investigate the reasons of attenuated sAA activity ratio before and after citric acid stimulation in splenic asthenia children. Saliva samples from 20 splenic asthenia children and 29 healthy children were collected before and after citric acid stimulation. AMYl copy number, sAA activity, and total sAA and glycosylated sAA contents were determined, and their correlations were analyzed. Although splenic asthenia and healthy children had no differences in AMY1 copy number, splenic asthenia children had positive correlations between AMY1 copy number and sAA activity before or after citric acid stimulation. Splenic asthenia children had a higher sAA glycosylated proportion ratio and glycosylated sAA content ratio, while their total sAA content ratio and sAA activity ratio were lower compared with healthy children. The glycosylated sAA content ratio was higher than the total sAA content ratio in both groups. Splenic asthenia and healthy children had positive correlations between total sAA or glycosylated sAA content and sAA activity. However, the role played by glycosylated sAA content in sAA activity in healthy children increased after citric acid stimulation, while it decreased in splenic asthenia children. Genetic factors like AMY1 copy number variations, and more importantly, sAA glycosylation abnormalities leading to attenuated sAA activity after citric acid stimulation, which were the main reasons of the attenuated sAA activity ratio in splenic asthenia children compared with healthy children.

Two novel, putatively cell wall-associated and glycosylphosphatidylinositol-anchored alpha-glucanotransferase enzymes of Aspergillus niger.

PubMed

van der Kaaij, R M; Yuan, X-L; Franken, A; Ram, A F J; Punt, P J; van der Maarel, M J E C; Dijkhuizen, L

2007-07-01

In the genome sequence of Aspergillus niger CBS 513.88, three genes were identified with high similarity to fungal alpha-amylases. The protein sequences derived from these genes were different in two ways from all described fungal alpha-amylases: they were predicted to be glycosylphosphatidylinositol anchored, and some highly conserved amino acids of enzymes in the alpha-amylase family were absent. We expressed two of these enzymes in a suitable A. niger strain and characterized the purified proteins. Both enzymes showed transglycosylation activity on donor substrates with alpha-(1,4)-glycosidic bonds and at least five anhydroglucose units. The enzymes, designated AgtA and AgtB, produced new alpha-(1,4)-glycosidic bonds and therefore belong to the group of the 4-alpha-glucanotransferases (EC 2.4.1.25). Their reaction products reached a degree of polymerization of at least 30. Maltose and larger maltooligosaccharides were the most efficient acceptor substrates, although AgtA also used small nigerooligosaccharides containing alpha-(1,3)-glycosidic bonds as acceptor substrate. An agtA knockout of A. niger showed an increased susceptibility towards the cell wall-disrupting compound calcofluor white, indicating a cell wall integrity defect in this strain. Homologues of AgtA and AgtB are present in other fungal species with alpha-glucans in their cell walls, but not in yeast species lacking cell wall alpha-glucan. Possible roles for these enzymes in the synthesis and/or maintenance of the fungal cell wall are discussed.

Effects of alpha-amylase reaction mechanisms on analysis of resistant-starch contents.

PubMed

Moore, Samuel A; Ai, Yongfeng; Chang, Fengdan; Jane, Jay-lin

2015-01-22

This study aimed to understand differences in the resistant starch (RS) contents of native and modified starches obtained using two standard methods of RS content analysis: AOAC Method 991.43 and 2002.02. The largest differences were observed in native potato starch, cross-linked wheat distarch phosphate, and high-amylose corn starch stearic-acid complex (RS5) between using AOAC Method 991.43 with Bacillus licheniformis α-amylase (BL) and AOAC Method 2002.02 with porcine pancreatic α-amylase (PPA). To determine possible reasons for these differences, we hydrolyzed raw-starch granules with BL and PPA with equal activity at pH 6.9 and 37°C for up to 84 h and observed the starch granules displayed distinct morphological differences after the hydrolysis. Starches hydrolyzed by BL showed erosion on the surface of the granules; those hydrolyzed by PPA showed pitting on granule surfaces. These results suggested that enzyme reaction mechanisms, including the sizes of the binding sites and the reaction patterns of the two enzymes, contributed to the differences in the RS contents obtained using different methods of RS analysis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Molecular, Biochemical, and Dietary Regulation Features of α-Amylase in a Carnivorous Crustacean, the Spiny Lobster Panulirus argus.

PubMed

Rodríguez-Viera, Leandro; Perera, Erick; Martos-Sitcha, Juan Antonio; Perdomo-Morales, Rolando; Casuso, Antonio; Montero-Alejo, Vivian; García-Galano, Tsai; Martínez-Rodríguez, Gonzalo; Mancera, Juan Miguel

2016-01-01

Alpha-amylases are ubiquitously distributed throughout microbials, plants and animals. It is widely accepted that omnivorous crustaceans have higher α-amylase activity and number of isoforms than carnivorous, but contradictory results have been obtained in some species, and carnivorous crustaceans have been less studied. In addition, the physiological meaning of α-amylase polymorphism in crustaceans is not well understood. In this work we studied α-amylase in a carnivorous lobster at the gene, transcript, and protein levels. It was showed that α-amylase isoenzyme composition (i.e., phenotype) in lobster determines carbohydrate digestion efficiency. Most frequent α-amylase phenotype has the lowest digestion efficiency, suggesting this is a favoured trait. We revealed that gene and intron loss have occurred in lobster α-amylase, thus lobsters express a single 1830 bp cDNA encoding a highly conserved protein with 513 amino acids. This protein gives rise to two isoenzymes in some individuals by glycosylation but not by limited proteolysis. Only the glycosylated isoenzyme could be purified by chromatography, with biochemical features similar to other animal amylases. High carbohydrate content in diet down-regulates α-amylase gene expression in lobster. However, high α-amylase activity occurs in lobster gastric juice irrespective of diet and was proposed to function as an early sensor of the carbohydrate content of diet to regulate further gene expression. We concluded that gene/isoenzyme simplicity, post-translational modifications and low Km, coupled with a tight regulation of gene expression, have arose during evolution of α-amylase in the carnivorous lobster to control excessive carbohydrate digestion in the presence of an active α-amylase.

Molecular, Biochemical, and Dietary Regulation Features of α-Amylase in a Carnivorous Crustacean, the Spiny Lobster Panulirus argus

PubMed Central

Martos-Sitcha, Juan Antonio; Perdomo-Morales, Rolando; Casuso, Antonio; Montero-Alejo, Vivian; García-Galano, Tsai; Martínez-Rodríguez, Gonzalo; Mancera, Juan Miguel

2016-01-01

Alpha-amylases are ubiquitously distributed throughout microbials, plants and animals. It is widely accepted that omnivorous crustaceans have higher α-amylase activity and number of isoforms than carnivorous, but contradictory results have been obtained in some species, and carnivorous crustaceans have been less studied. In addition, the physiological meaning of α-amylase polymorphism in crustaceans is not well understood. In this work we studied α-amylase in a carnivorous lobster at the gene, transcript, and protein levels. It was showed that α-amylase isoenzyme composition (i.e., phenotype) in lobster determines carbohydrate digestion efficiency. Most frequent α-amylase phenotype has the lowest digestion efficiency, suggesting this is a favoured trait. We revealed that gene and intron loss have occurred in lobster α-amylase, thus lobsters express a single 1830 bp cDNA encoding a highly conserved protein with 513 amino acids. This protein gives rise to two isoenzymes in some individuals by glycosylation but not by limited proteolysis. Only the glycosylated isoenzyme could be purified by chromatography, with biochemical features similar to other animal amylases. High carbohydrate content in diet down-regulates α-amylase gene expression in lobster. However, high α-amylase activity occurs in lobster gastric juice irrespective of diet and was proposed to function as an early sensor of the carbohydrate content of diet to regulate further gene expression. We concluded that gene/isoenzyme simplicity, post-translational modifications and low Km, coupled with a tight regulation of gene expression, have arose during evolution of α-amylase in the carnivorous lobster to control excessive carbohydrate digestion in the presence of an active α-amylase. PMID:27391425

Thermostable microbial xylanases for pulp and paper industries: trends, applications and further perspectives.

PubMed

Kumar, Vishal; Marín-Navarro, Julia; Shukla, Pratyoosh

2016-02-01

Xylanases are enzymes with biotechnological relevance in a number of fields, including food, feed, biofuel, and textile industries. Their most significant application is in the paper and pulp industry, where they are used as a biobleaching agent, showing clear economic and environmental advantages over chemical alternatives. Since this process requires high temperatures and alkali media, the identification of thermostable and alkali stable xylanases represents a major biotechnological goal in this field. Moreover, thermostability is a desirable property for many other applications of xylanases. The review makes an overview of xylanase producing microorganisms and their current implementation in paper biobleaching. Future perspectives are analyzed focusing in the efforts carried out to generate thermostable enzymes by means of modern biotechnological tools, including metagenomic analysis, enzyme molecular engineering and nanotechnology. Furthermore, structural and mutagenesis studies have revealed critical sites for stability of xylanases from glycoside hydrolase families GH10 and GH11, which constitute the main classes of these enzymes. The overall conclusions of these works are summarized here and provide relevant information about putative weak spots within xylanase structures to be targeted in future protein engineering approaches.

Gedunin and Azadiradione: Human Pancreatic Alpha-Amylase Inhibiting Limonoids from Neem (Azadirachta indica) as Anti-Diabetic Agents.

PubMed

Ponnusamy, Sudha; Haldar, Saikat; Mulani, Fayaj; Zinjarde, Smita; Thulasiram, Hirekodathakallu; RaviKumar, Ameeta

2015-01-01

Human pancreatic α-amylase (HPA) inhibitors offer an effective strategy to lower postprandial hyperglycemia via control of starch breakdown. Limonoids from Azadirachta indica known for their therapeutic potential were screened for pancreatic α-amylase inhibition, a known anti-diabetic target. Studies were carried out to reveal their mode of action so as to justify their hypoglycemic potential. Of the nine limonoids isolated/semi-synthesized from A.indica and screened for α-amylase inhibition, azadiradione and exhibited potential inhibition with an IC50 value of 74.17 and 68.38 μM, respectively against HPA under in vitro conditions. Further screening on AR42J α-amylase secretory cell line for cytotoxicity and bioactivity revealed that azadiradione and gedunin exhibited cytotoxicity with IC50 of 11.1 and 13.4μM. Maximal secreted α-amylase inhibition of 41.8% and 53.4% was seen at 3.5 and 3.3μM, respectively. Michaelis-Menten kinetics suggested a mixed mode of inhibition with maltopentaose (Ki 42.2, 18.6 μM) and starch (Ki' 75.8, 37.4 μM) as substrate with a stiochiometry of 1:1 for both azadiradione and gedunin, respectively. The molecular docking simulation indicated plausible π-alkyl and alkyl-alkyl interactions between the aromatic amino acids and inhibitors. Fluorescence and CD confirmed the involvement of tryptophan and tyrosine in ligand binding to HPA. Thermodynamic parameters suggested that binding is enthalpically and entropically driven with ΔG° of -21.25 kJ mol-1 and -21.16 kJ mol-1 for azadiradione and gedunin, respectively. Thus, the limonoids azadiradione and gedunin could bind and inactivate HPA (anti-diabetic target) and may prove to be lead drug candidates to reduce/control post-prandial hyperglycemia.

Effect of starch and amylase on the expression of amylase-binding protein A in Streptococcus gordonii

PubMed Central

Nikitkova, A.E.; Haase, E.M.; Scannapieco, F.A.

2012-01-01

SUMMARY Streptococcus gordonii is a common oral commensal bacterial species in tooth biofilm (dental plaque) and specifically binds to salivary amylase through the surface exposed amylase-binding protein A (AbpA). When S. gordonii cells are pretreated with amylase, amylase bound to AbpA facilitates growth with starch as a primary nutrition source. The goal of this study was to explore possible regulatory effects of starch, starch metabolites and amylase on the expression of S. gordonii AbpA. An amylase ligand-binding assay was used to assess the expression of AbpA in culture supernatants and on bacterial cells from S. gordonii grown in defined medium supplemented with 1% starch, 0.5 mg ml−1 amylase, with starch and amylase together, or with various linear malto-oligosaccharides. Transcription of abpA was determined by reverse transcription quantitative polymerase chain reaction. AbpA was not detectable in culture supernatants containing either starch alone or amylase alone. In contrast, the amount of AbpA was notably increased when starch and amylase were both present in the medium. The expression of abpA was significantly increased (P < 0.05) following 40 min of incubation in defined medium supplemented with starch and amylase. Similar results were obtained in the presence of maltose and other short-chain malto-oligosacchrides. These results suggest that the products of starch hydrolysis produced from the action of salivary α-amylase, particularly maltose and maltotriose, regulate AbpA expression in S. gordonii. PMID:22759313

The diurnal course of salivary alpha-amylase in nurses: an investigation of potential confounders and associations with stress.

PubMed

Wingenfeld, Katja; Schulz, Michael; Damkroeger, Annika; Philippsen, Christine; Rose, Matthias; Driessen, Martin

2010-09-01

In psychoneuroendocrinology research, salivary measures have become increasingly important. While several studies focus on determinants of salivary cortisol such as age, gender, and gynaecological variables, less research has focused on confounding variables of salivary alpha-amylase (sAA). In a large sample of nurses (N=215) we analyzed the impact of age, gender, intake of oral contraceptives, smoking, coffee consumption as well as psychological parameters, such as work stress and burnout, on basal diurnal sAA release. Saliva was collected at 07:00 h, 11:30 h, 17:30 h, and 20:00 h on a working day during early shift. Only gender could be identified to have an impact on sAA, with females having a more pronounced sAA increase over the course of the day. Whereas depression, anxiety, work stress and burnout were not associated with sAA, a small negative correlation between social difficulties, measured with the Chronic Stress Screening Scale, and sAA could be identified. Copyright (c) 2010 Elsevier B.V. All rights reserved.

Maize pollen coat xylanase facilitates pollen tube penetration into silk during sexual reproduction.

PubMed

Suen, Der Fen; Huang, Anthony H C

2007-01-05

Cell wall hydrolases are well documented to be present on pollen, but their roles on the stigma during sexual reproduction have not been previously demonstrated. We explored the function of the tapetum-synthesized xylanase, ZmXYN1, on maize (Zea mays L.) pollen. Transgenic lines (xyl-less) containing little or no xylanase in the pollen coat were generated with use of an antisense construct of the xylanase gene-coding region driven by the XYN1 gene promoter. Xyl-less and wild-type plants had similar vegetative growth. Electron microscopy revealed no appreciable morphological difference in anther cells and pollen between xyl-less lines and the wild type, whereas immunofluorescence microscopy and biochemical analyses indicated an absence of xylanase on xyl-less pollen. Xyl-less pollen germinated as efficiently as wild-type pollen in vitro in a liquid medium but less so on gel media of increasing solidity or on silk, which is indicative of partial impaired water uptake. Once germinated in vitro or on silk, the xyl-less and wild-type pollen tubes elongated at comparable rates. Tubes of germinated xyl-less pollen on silk did not penetrate into the silk as efficiently as tubes of wild-type pollen, and this lower efficiency could be overcome by the addition of xylanase to the silk. For wild-type pollen, coat xylanase activity on oat spelled xylan in vitro and tube penetration into silk were inhibited by xylose but not glucose. The overall findings indicate that maize pollen coat xylanase facilitates pollen tube penetration into silk via enzymatic xylan hydrolysis.

Functional characterization of Penicillium occitanis Pol6 and Penicillium funiculosum GH11 xylanases.

PubMed

Driss, Dorra; Berrin, Jean Guy; Juge, Nathalie; Bhiri, Fatma; Ghorbel, Raoudha; Chaabouni, Semia Ellouz

2013-08-01

Xylanases are hemicellulolytic enzymes, which are responsible for the degradation of heteroxylans constituting the lignocellulosic plant cell wall. Xylanases from the GH11 family are considered as true xylanases because of their high substrate specificity. In order to study in depth a crucial difference in the thumb region between two closely related xylanases from Penicillium in terms of kinetic parameters and inhibition sensitivity, the GH11 xylanases from Penicillium occitanis Pol6 (PoXyn3) and from Penicillium funiculosum (PfXynC) were heterologously expressed in Pichia pastoris. The PoXyn3 and PfXynC cDNAs encoding mature xylanases were cloned into pGAPZαA vectors and integrated into the genome of P. pastoris X-33 under the control of the glyceraldehyde 3-phosphate dehydrogenase constitutive promoter. PfXynC was expressed as a His-tagged recombinant protein and purified from the supernatant homogeneity by a one-step purification protocol using immobilized metal affinity chromatography. The recombinant PoXyn3 was purified using a single anion-exchange chromatography. The purified recombinant enzymes were optimally active at 45°C and pH 4.0 for PoXyn3 and 40°C and pH 3.0 for PfXynC. The measured kinetic parameters (k(cat) and Vmax) showed that PfXynC was five times more active than PoXyn3 irrespective of the substrate whereas the apparent affinity (K(m)) was similar. The recombinant enzymes showed distinct sensitivity to the Triticum aestivum xylanase inhibitor TAXI-I. Copyright © 2013 Elsevier Inc. All rights reserved.

Aspergillus oryzae S2 alpha-amylase production under solid state fermentation: optimization of culture conditions.

PubMed

Sahnoun, Mouna; Kriaa, Mouna; Elgharbi, Fatma; Ayadi, Dorra-Zouari; Bejar, Samir; Kammoun, Radhouane

2015-04-01

Aspergillus oryzae S2 was assayed for alpha-amylase production under solid state fermentation (SSF). In addition to AmyA and AmyB already produced in monitored submerged culture, the strain was noted to produce new AmyB oligomeric forms, in particular a dominant tetrameric form named AmyC. The latter was purified to homogeneity through fractional acetone precipitation and size exclusion chromatography. SDS-PAGE and native PAGE analyses revealed that, purified AmyC was an approximately 172 kDa tetramer of four 42 kDa subunits. AmyC was also noted to display the same NH2-terminal amino acid sequence residues and approximately the same physico-chemical properties of AmyA and AmyB, to exhibit maximum activity at pH 5.6 and 60 °C, and to produce maltose and maltotriose as major starch hydrolysis end-products. Soyabean meal was the best substitute to yeast extract compared to fish powder waste and wheat gluten waste. AmyC production was optimized under SSF using statistical design methodology. Moisture content of 76.25%, C/N substrate ratio of 0.62, and inoculum size of 10(6.87) spores allowed maximum activity of 22118.34 U/g of dried substrate, which was 33 times higher than the one obtained before the application of the central composite design (CCD). Copyright © 2015 Elsevier B.V. All rights reserved.

Pectinase hydrolysis of Dendrobium huoshanense polysaccharide and its effect on protein nonenzymatic glycation.

PubMed

Zha, Xue-Qiang; Li, Xiao-Long; Zhang, Hai-Lin; Cui, Shao-Hua; Liu, Jian; Wang, Jun-Hui; Pan, Li-Hua; Luo, Jian-Ping

2013-10-01

The aim of this study was to investigate the inhibitory effects of molecular weight alteration of Dendrobium huoshanense polysaccharide on protein nonenzymatic glycation. For this purpose, one homogeneous active polysaccharide DHPD1 with molecular weight 3.2 kDa was extracted from D. huoshanense. GC analysis showed that DHPD1 was mainly composed of glucose, arabinose, galactose in a molar ratio of 0.023:1.023:0.021 with a trace of mannose and xylose. In order to get DHPD1-derived fragments with different molecular weight, response surface methodology was employed to optimize the enzymatic degradation conditions. The maximum reducing sugar production (0.399 mg/mL) was obtained under an optimal condition including pectinase dosage 126 U/mL, reaction pH 4.46 and reaction temperature 48 °C. By applying this condition, three DHPD1-derived fragments with different molecular weights were obtained through changing the hydrolysis time. Infrared spectroscopy analysis indicated that the backbone structure of DHPD1 was not destroyed by pectinase hydrolysis. Monosaccharide composition analysis showed that pectinase preferred to liberate glucose from DHPD1. The inhibitory action of DHPD1 on protein nonenzymatic glycation reduced with the decrease of molecular weight. Copyright © 2013 Elsevier B.V. All rights reserved.

A Simple Method for the Determination of Xylanase Activity on Insoluble Substrates

USDA-ARS?s Scientific Manuscript database

The propensity for a xylanase to convert insoluble (arabino)xylan into soluble oligosaccharides is an important parameter in the baking, pulp and paper, prebiotics, and biofuel industries. Current methods for determining xylanase activity on insoluble substrates are labor intensive, non-specific, or...

Effect of the glass transition temperature on alpha-amylase activity in a starch matrix.

PubMed

Chaudhary, Vinita; Panyoyai, Naksit; Small, Darryl M; Shanks, Robert A; Kasapis, Stefan

2017-02-10

This study optimises a protocol for the estimation of α-amylase activity in a condensed starch matrix in the vicinity of the glass transition region. Enzymatic activity on the vitrified starch system was compared with that of a reference substrate, maltodextrin. The activity was assayed as the rate of release of reducing sugar using a dinitrosalicylic acid procedure. The condensed carbohydrate matrices served the dual purpose of acting as a substrate as well as producing a pronounced effect on the ability to enzymatic hydrolysis. Activation energies were estimated throughout the glass transition region of condensed carbohydrate preparations based on the concept of the spectroscopic shift factor. Results were used to demonstrate a considerable moderation by the mechanical glass transition temperature, beyond the expected linear effect of the temperature dependence, on the reaction rate of starch hydrolysis by α-amylase in comparison with the low-molecular weight chain of maltodextrin. Copyright © 2016. Published by Elsevier Ltd.

Evaluation of free and immobilized Aspergillus niger NRC1ami pectinase applicable in industrial processes.

PubMed

Esawy, Mona A; Gamal, Amira A; Kamel, Zeinat; Ismail, Abdel-Mohsen S; Abdel-Fattah, Ahmed F

2013-02-15

The Aspergillus niger NRC1ami pectinase was evaluated according to its hydrolysis efficiency of dry untreated orange peels (UOP), HCl-treated orange peels and NaOH-treated orange peels (HOP and NOP). Pectinase was entrapped in polyvinyl alcohol (PVA) sponge and the optimum pH and temperature of the free and immobilized enzymes were shifted from 4, 40 °C to 6, 50 °C respectively. The study of pH stability of free and immobilized pectinase showed that the immobilization process protected the enzyme strongly from severe alkaline pHs. The immobilization process improved the enzyme thermal stability to great instant. The unique feature of the immobilization process is its ability to solve the orange juice haze problem completely. Immobilized enzyme was reused 12 times in orange juice clarification with 9% activity loss from the original activity. Maximum reaction rate (V(max)) and Michaelis-Menten constant (K(m)) of the partially purified form were significantly changed after immobilization. Copyright © 2012 Elsevier Ltd. All rights reserved.

Industrially Important Carbohydrate Degrading Enzymes from Yeasts: Pectinases, Chitinases, and β-1,3-Glucanases

NASA Astrophysics Data System (ADS)

Gummadi, Sathyanarayana N.; Kumar, D. Sunil; Dash, Swati S.; Sahu, Santosh Kumar

Polysaccharide degrading enzymes are hydrolytic enzymes, which have a lot of industrial potential and also play a crucial role in carbon recycling. Pectinases, chitinases and glucanases are the three major polysaccharide degrading enzymes found abundantly in nature and these enzymes are mainly produced by fungal strains. Production of these enzymes by yeasts is advantageous over fungi, because the former are easily amenable to genetic manipulations and time required for growth and production is less than that of the latter. Several yeasts belonging to Saccharomyces, Pichia, Rhodotorula and Cryptococcus produce extracellular pectinases, glucanases and chitinases. This chapter emphasizes on the biological significance of these enzymes, their production and their industrial applications.

Xylanase production with xylan rich lignocellulosic wastes by a local soil isolate of Trichoderma viride

PubMed Central

Goyal, Meenakshi; Kalra, K.L.; Sareen, V.K.; Soni, G.

2008-01-01

In the present study, cultural and nutritional conditions for enhanced production of xylanase by a local soil isolate of Trichoderma viride, using various lignocellulosic substrates in submerged culture fermentation have been optimized. Of the lignocellulosics used, maize straw was the best inducer followed by jowar straw for xylanase production. The highest activity achieved was between 14 to 17 days of fermentation. A continuous increase in xylanase production was observed with increasing level of lignocellulosics in the medium and highest activity was observed with maize straw at 5% level. Xylanase production with higher levels of lignocellulosics (3 to 5%) of maize, jowar and barseem was found to be higher as compared to that with commercial xylan as carbon source. Sodium nitrate was the best nitrogen source among the six sources used. Maximum xylanase production was achieved with initial medium pH of 3.5–4.0 and incubation temperature of 25ºC.The enzyme preparation was effective in bringing about saccharification of different lignocellulosics. The xylanase production could be further improved by using alkali treated straw as carbon source. PMID:24031262

A novel cellulase free alkaliphilic xylanase from alkali tolerant Penicillium citrinum: production, purification and characterization.

PubMed

Dutta, T; Sengupta, R; Sahoo, R; Sinha Ray, S; Bhattacharjee, A; Ghosh, S

2007-02-01

The enzymatic hydrolysis of xylan has potential economic and environment-friendly applications. Therefore, attention is focused here on the discovery of new extremophilic xylanase in order to meet the requirements of industry. An extracellular xylanase was purified from the culture filtrate of P. citrinum grown on wheat bran bed in solid substrate fermentation. Single step purification was achieved using hydrophobic interaction chromatography. The purified enzyme showed a single band on SDS-PAGE with an apparent molecular weight of c. 25 kDa and pI of 3.6. Stimulation of the activity by beta mercaptoethanol, dithiotheritol (DTT) and cysteine was observed. Moderately thermostable xylanase showed optimum activity at 50 degrees C at pH 8.5. Xylanase purified from P. citrinum was alkaliphilic and moderately thermostable in nature. The present work reports for the first time the purification and characterization of a novel endoglucanase free alkaliphilic xylanase from the alkali tolerant fungus Penicillium citrinum. The alkaliphilicity and moderate thermostability of this xylanase may have potential implications in paper and pulp industries.

Benefits from additives and xylanase during enzymatic hydrolysis of bamboo shoot and mature bamboo.

PubMed

Li, Kena; Wang, Xiao; Wang, Jingfeng; Zhang, Junhua

2015-09-01

Effects of additives (BSA, PEG 6000, and Tween 80) on enzymatic hydrolysis of bamboo shoot and mature bamboo fractions (bamboo green, bamboo timber, bamboo yellow, bamboo node, and bamboo branches) by cellulases and/or xylanase were evaluated. The addition of additives was comparable to the increase of cellulase loadings in the conversion of cellulose and xylan in bamboo fractions. Supplementation of xylanase (1 mg/g DM) with cellulases (10 FPU/g DM) in the hydrolysis of bamboo fractions was more efficient than addition of additives in the production of glucose and xylose. Moreover, addition of additives could further increase the glucose release from different bamboo fractions by cellulases and xylanase. Bamboo green exhibited the lowest hydrolyzability. Almost all of the polysaccharides in pretreated bamboo shoot fractions were hydrolyzed by cellulases with the addition of additives or xylanase. Additives and xylanase showed great potential for reducing cellulase requirement in the hydrolysis of bamboo. Copyright © 2015 Elsevier Ltd. All rights reserved.

Structural Insight into and Mutational Analysis of Family 11 Xylanases: Implications for Mechanisms of Higher pH Catalytic Adaptation.

PubMed

Bai, Wenqin; Zhou, Cheng; Zhao, Yueju; Wang, Qinhong; Ma, Yanhe

2015-01-01

To understand the molecular basis of higher pH catalytic adaptation of family 11 xylanases, we compared the structures of alkaline, neutral, and acidic active xylanases and analyzed mutants of xylanase Xyn11A-LC from alkalophilic Bacillus sp. SN5. It was revealed that alkaline active xylanases have increased charged residue content, an increased ratio of negatively to positively charged residues, and decreased Ser, Thr, and Tyr residue content relative to non-alkaline active counterparts. Between strands β6 and β7, alkaline xylanases substitute an α-helix for a coil or turn found in their non-alkaline counterparts. Compared with non-alkaline xylanases, alkaline active enzymes have an inserted stretch of seven amino acids rich in charged residues, which may be beneficial for xylanase function in alkaline conditions. Positively charged residues on the molecular surface and ionic bonds may play important roles in higher pH catalytic adaptation of family 11 xylanases. By structure comparison, sequence alignment and mutational analysis, six amino acids (Glu16, Trp18, Asn44, Leu46, Arg48, and Ser187, numbering based on Xyn11A-LC) adjacent to the acid/base catalyst were found to be responsible for xylanase function in higher pH conditions. Our results will contribute to understanding the molecular mechanisms of higher pH catalytic adaptation in family 11 xylanases and engineering xylanases to suit industrial applications.

Mind your thoughts: associations between self-generated thoughts and stress-induced and baseline levels of cortisol and alpha-amylase.

PubMed

Engert, Veronika; Smallwood, Jonathan; Singer, Tania

2014-12-01

Stress is a major health burden in today's society. Research shows that negative cognitive styles are associated with increased stress reactivity, low mood and accelerated cellular aging. Our study sought to unravel the relationship between the content of self-generated thoughts and psychosocial stress measured in terms of hypothalamic-pituitary-adrenal axis and sympathetic activity. Features of self-generated thoughts were assessed using thought sampling while participants performed cognitive tasks following a stress induction or in a baseline condition. More negatively toned emotional thoughts and more social temporal thoughts with a past focus were associated with increased cortisol and alpha-amylase levels, both after stress and at baseline. More social temporal thoughts with a future focus, on the other hand, had an overall attenuating effect on the levels of both stress markers. Our results indicate a fundamental link between the thoughts and stress levels we experience. Understanding the mechanisms governing this mind-body association may have important implications for understanding and counteracting the high incidence of stress-related disorders in today's society. Copyright © 2014 Elsevier B.V. All rights reserved.

A reusable multipurpose magnetic nanobiocatalyst for industrial applications.

PubMed

Perwez, Mohammad; Ahmad, Razi; Sardar, Meryam

2017-10-01

A multipurpose magnetic nanobiocatalyst is developed by conjugating Pectinex 3XL (a commercial enzyme containing pectinase, xylanase and cellulase activities) on 3-aminopropyl triethoxysilane activated magnetic nanoparticles. The nanobiocatalyst retained 87% of pectinase, 69% of xylanase and 58% of cellulase activity after conjugation on modified nanoparticles as compared to their soluble counterparts. Thermal stability data at 70°C showed increase in enzyme stability after conjugation to nanoparticles and the kinetic parameters (K m and V max ) remain unaltered after immobilization. The immobilized enzyme system can be successfully used upto 5th cycle after that slight decrease in enzyme activities was observed. The nanobiocatalyst retained high pectinase activities in organic solvents and chemical reagents as compared to free enzymes. DLS data shows that the nanoparticles size increases from 63nm to 86nm after immobilization. Atomic Force Microscopy data confirms the deposition of enzymes on the nanoparticles. The nanobiocatalyst was used for the clarification of pine apple and orange juice and was also used for the production of bioethanol. Hydrolysis of pretreated wheat straw produced 1.39g/l and 1.59g/l after treatment with free Pectinex 3xL and nanobiocatalyst respectively. The concentration of bioethanol also increases by 1.4 fold as compared to the free enzyme. Copyright © 2017 Elsevier B.V. All rights reserved.

The Use of Xylanases from Different Microbial Origin in Bread Baking and Their Effects on Bread Qualities

NASA Astrophysics Data System (ADS)

Al-Widyan, Omar; Khataibeh, Moayad H.; Abu-Alruz, Khaled

Effects of xylanases on bread quality were examined. Enzymes used were endo-xylanase (EC 3.2.1.8) from different sources of microorganisms. Baked loaves were assessed for Loaves volume, colour and staling rate. Xylanases produced from rumen microorganisms M6 had clearly positive effects on loaf volume of bread as well as anti-firming potential. M3 (produced from Trichoderma longibrachiatum) improved crumb softness. The use of xylanase for breadmaking lowered firmness of bread crumb effectively compared with control loaf. It can be summarized that xylanases had significant positive effects on bread characteristics. In particular, they had advantage in retarding the staling rate of bread. It is recommended that the optimum dosage of enzymes, method of application in industrial scale especially with xylanase should be studied further in order to gain the great advantages of enzyme addition in breadmaking.

Structural Insight into and Mutational Analysis of Family 11 Xylanases: Implications for Mechanisms of Higher pH Catalytic Adaptation

PubMed Central

Bai, Wenqin; Zhou, Cheng; Zhao, Yueju; Wang, Qinhong; Ma, Yanhe

2015-01-01

To understand the molecular basis of higher pH catalytic adaptation of family 11 xylanases, we compared the structures of alkaline, neutral, and acidic active xylanases and analyzed mutants of xylanase Xyn11A-LC from alkalophilic Bacillus sp. SN5. It was revealed that alkaline active xylanases have increased charged residue content, an increased ratio of negatively to positively charged residues, and decreased Ser, Thr, and Tyr residue content relative to non-alkaline active counterparts. Between strands β6 and β7, alkaline xylanases substitute an α-helix for a coil or turn found in their non-alkaline counterparts. Compared with non-alkaline xylanases, alkaline active enzymes have an inserted stretch of seven amino acids rich in charged residues, which may be beneficial for xylanase function in alkaline conditions. Positively charged residues on the molecular surface and ionic bonds may play important roles in higher pH catalytic adaptation of family 11 xylanases. By structure comparison, sequence alignment and mutational analysis, six amino acids (Glu16, Trp18, Asn44, Leu46, Arg48, and Ser187, numbering based on Xyn11A-LC) adjacent to the acid/base catalyst were found to be responsible for xylanase function in higher pH conditions. Our results will contribute to understanding the molecular mechanisms of higher pH catalytic adaptation in family 11 xylanases and engineering xylanases to suit industrial applications. PMID:26161643

A dispersion model for predicting the extent of starch liquefaction by Bacillus licheniformis alpha-amylase during reactive extrusion.

PubMed

Komolprasert, V; Ofoli, R Y

1991-03-25

A Baker-Perkins corotating twin screw extruder was used as a bioreactor to hydrolyze pregelantinized corn starch by themophilic Bacillus licheniformis alpha-amylase. The extruder was modeled as a tube, and characterized as a closed system. This characterization is not in the thermodynamic sense; rather, it relates to the profile of a tracer fluid upon entry to and exit from the reaction zone. The reaction kinetics were modeled by a modified first-order equation, which allowed the dispersion equation to be solved analytically with the Danckwerts boundary condition. Data from several extrusion runs were super-imposed to obtain a profile to evaluate the model. The dispersion number, determined from the first and second moments of the RTD curve, was primarily a function of the length of the reaction zone. There was good agreement between predictions and experimental data, especially at low dispersion numbers. In general, the axial dispersion model appears to be suitable for analysis of enzymatic reactions of up to 30% conversion. At a fixed flow rate and constant temperature, the extent of starch conversion depends significantly on moisture content, residence time and enzyme dosage, but not on screw speed.

Characterization of pectinase activity for enology from yeasts occurring in Argentine Bonarda grape.

PubMed

Merín, María Gabriela; Martín, María Carolina; Rantsiou, Kalliopi; Cocolin, Luca; de Ambrosini, Vilma Inés Morata

2015-01-01

Pectinolytic enzymes are greatly important in winemaking due to their ability to degrade pectic polymers from grape, contributing to enhance process efficiency and wine quality. This study aimed to analyze the occurrence of pectinolytic yeasts during spontaneous fermentation of Argentine Bonarda grape, to select yeasts that produce extracellular pectinases and to characterize their pectinolytic activity under wine-like conditions. Isolated yeasts were grouped using PCR-DGGE and identified by partial sequencing of 26S rRNA gene. Isolates comprised 7 genera, with Aureobasidium pullulans as the most predominant pectinolytic species, followed by Rhodotorula dairenensis and Cryptococcus saitoi. No pectinolytic activity was detected among ascomycetous yeasts isolated on grapes and during fermentation, suggesting a low occurrence of pectinolytic yeast species in wine fermentation ecosystem. This is the first study reporting R. dairenensis and Cr. saitoi species with pectinolytic activity. R. dairenensis GM-15 produced pectinases that proved to be highly active at grape pH, at 12 °C, and under ethanol and SO2 concentrations usually found in vinifications (pectinase activity around 1.1 U/mL). This strain also produced cellulase activity at 12 °C and pH 3.5, but did not produce β-glucosidase activity under these conditions. The strain showed encouraging enological properties for its potential use in low-temperature winemaking.

Characterization of pectinase activity for enology from yeasts occurring in Argentine Bonarda grape

PubMed Central

Merín, María Gabriela; Martín, María Carolina; Rantsiou, Kalliopi; Cocolin, Luca; de Ambrosini, Vilma Inés Morata

2015-01-01

Pectinolytic enzymes are greatly important in winemaking due to their ability to degrade pectic polymers from grape, contributing to enhance process efficiency and wine quality. This study aimed to analyze the occurrence of pectinolytic yeasts during spontaneous fermentation of Argentine Bonarda grape, to select yeasts that produce extracellular pectinases and to characterize their pectinolytic activity under wine-like conditions. Isolated yeasts were grouped using PCR-DGGE and identified by partial sequencing of 26S rRNA gene. Isolates comprised 7 genera, with Aureobasidium pullulans as the most predominant pectinolytic species, followed by Rhodotorula dairenensis and Cryptococcus saitoi. No pectinolytic activity was detected among ascomycetous yeasts isolated on grapes and during fermentation, suggesting a low occurrence of pectinolytic yeast species in wine fermentation ecosystem. This is the first study reporting R. dairenensis and Cr. saitoi species with pectinolytic activity. R. dairenensis GM-15 produced pectinases that proved to be highly active at grape pH, at 12 °C, and under ethanol and SO2 concentrations usually found in vinifications (pectinase activity around 1.1 U/mL). This strain also produced cellulase activity at 12 °C and pH 3.5, but did not produce β-glucosidase activity under these conditions. The strain showed encouraging enological properties for its potential use in low-temperature winemaking. PMID:26413065

Amylase Test: MedlinePlus Lab Test Information

MedlinePlus

... page: https://medlineplus.gov/labtests/amylasetest.html Amylase Test To use the sharing features on this page, please enable JavaScript. What is an Amylase Test? An amylase test measures the amount of amylase ...

Autonomic markers associated with generalized social phobia symptoms: heart rate variability and salivary alpha-amylase.

PubMed

García-Rubio, María J; Espín, Laura; Hidalgo, Vanesa; Salvador, Alicia; Gómez-Amor, Jesús

2017-01-01

The study of autonomic nervous system changes associated with generalized social phobia (GSP) disorder has increased in recent years, showing contradictory results. The present study aimed to evaluate how young people with GSP reacted before, during, and after exposure to the Trier Stress Social Test (TSST), focusing on their autonomic changes (heart rate variability (HRV) and salivary alpha-amylase (sAA)) compared to a control group (non-GSP). Some psychological variables were also considered. Sex was specifically studied as a possible modulator of autonomic fluctuations and psychological state. Eighty young people were randomly distributed into two counterbalanced situations: stress condition (N = 18 and 21 for GSP and non-GSP, respectively) and control condition (N = 21 and 20 for GSP and non-GSP, respectively), where cardiovascular variables were continuously recorded. Psychological questionnaires about mood and perceived stress were filled out, and five saliva samples were collected to analyze sAA. GSP participants showed higher values on low- and high-frequency ratios (HR domains), compared to non-GSP people, during exposure to the TSST, but no differences were observed after the stressor. Furthermore, the two groups did not differ in sAA. Importantly, positive affect in GSP participants was modulated by sex. The present study suggests that the balance between high- and low-frequency domains of HRV is a key cardiovascular marker reflecting the stress response of GSP people, as well the importance of sex in positive affect when facing a stressful situation.

Kinetic stabilization of Bacillus licheniformis alpha-amylase through introduction of hydrophobic residues at the surface.

PubMed

Machius, Mischa; Declerck, Nathalie; Huber, Robert; Wiegand, Georg

2003-03-28

It is generally assumed that in proteins hydrophobic residues are not favorable at solvent-exposed sites, and that amino acid substitutions on the surface have little effect on protein thermostability. Contrary to these assumptions, we have identified hyperthermostable variants of Bacillus licheniformis alpha-amylase (BLA) that result from the incorporation of hydrophobic residues at the surface. Under highly destabilizing conditions, a variant combining five stabilizing mutations unfolds 32 times more slowly and at a temperature 13 degrees C higher than the wild-type. Crystal structure analysis at 1.7 A resolution suggests that stabilization is achieved through (a) extension of the concept of increased hydrophobic packing, usually applied to cavities, to surface indentations, (b) introduction of favorable aromatic-aromatic interactions on the surface, (c) specific stabilization of intrinsic metal binding sites, and (d) stabilization of a beta-sheet by introducing a residue with high beta-sheet forming propensity. All mutated residues are involved in forming complex, cooperative interaction networks that extend from the interior of the protein to its surface and which may therefore constitute "weak points" where BLA unfolding is initiated. This might explain the unexpectedly large effect induced by some of the substitutions on the kinetic stability of BLA. Our study shows that substantial protein stabilization can be achieved by stabilizing surface positions that participate in underlying cooperatively formed substructures. At such positions, even the apparently thermodynamically unfavorable introduction of hydrophobic residues should be explored.

Unexpected high digestion rate of cooked starch by the Ct-Maltase-Glucoamylase small intestine mucosal alpha-glucosidase subunit

USDA-ARS?s Scientific Manuscript database

For starch digestion to glucose, two luminal alpha-amylases and four gut mucosal alpha-glucosidase subunits are employed. The aim of this research was to investigate, for the first time, direct digestion capability of individual mucosal alpha-glucosidases on cooked (gelatinized) starch. Gelatinized ...

Purification and Characterization of Haloalkaline, Organic Solvent Stable Xylanase from Newly Isolated Halophilic Bacterium-OKH

PubMed Central

Sanghvi, Gaurav; Jivrajani, Mehul; Patel, Nirav; Jivrajani, Heta; Bhaskara, Govinal Badiger; Patel, Shivani

2014-01-01

A novel, alkali-tolerant halophilic bacterium-OKH with an ability to produce extracellular halophilic, alkali-tolerant, organic solvent stable, and moderately thermostable xylanase was isolated from salt salterns of Mithapur region, Gujarat, India. Identification of the bacterium was done based upon biochemical tests and 16S rRNA sequence. Maximum xylanase production was achieved at pH 9.0 and 37°C temperature in the medium containing 15% NaCl and 1% (w/v) corn cobs. Sugarcane bagasse and wheat straw also induce xylanase production when used as carbon source. The enzyme was active over a range of 0–25% sodium chloride examined in culture broth. The optimum xylanase activity was observed at 5% sodium chloride. Xylanase was purified with 25.81%-fold purification and 17.1% yield. Kinetic properties such as Km and Vmax were 4.2 mg/mL and 0.31 μmol/min/mL, respectively. The enzyme was stable at pH 6.0 and 50°C with 60% activity after 8 hours of incubation. Enzyme activity was enhanced by Ca2+, Mn2+, and Mg2+ but strongly inhibited by heavy metals such as Hg2+, Fe3+, Ni2+, and Zn2+. Xylanase was found to be stable in organic solvents like glutaraldehyde and isopropanol. The purified enzyme hydrolysed lignocellulosic substrates. Xylanase, purified from the halophilic bacterium-OKH, has potential biotechnological applications. PMID:27350996

Susceptibility to corrosion of laser welding composite arch wire in artificial saliva of salivary amylase and pancreatic amylase.

PubMed

Zhang, Chao; Liu, Jiming; Yu, Wenwen; Sun, Daqian; Sun, Xinhua

2015-10-01

In this study, laser-welded composite arch wire (CAW) with a copper interlayer was exposed to artificial saliva containing salivary amylase or pancreatic amylase, and the resultant corrosion behavior was studied. The purpose was to determine the mechanisms by which salivary amylase and pancreatic amylase contribute to corrosion. The effects of amylase on the electrochemical resistance of CAW were tested by potentiodynamic polarization measurements. The dissolved corrosion products were determined by ICP-OES, and the surfaces were analyzed by SEM, AFM and EDS. The results showed that both exposure to salivary amylase and pancreatic amylase significantly improved the corrosion resistance of CAW. Even isozyme could have different influences on the alloy surface. When performing in vitro research of materials to be used in oral cavity, the effect of α-amylase should be taken into account since a simple saline solution does not entirely simulate the physiological situation. Copyright © 2015 Elsevier B.V. All rights reserved.

Enhancement of thermoalkaliphilic xylanase production by Pichia pastoris through novel fed-batch strategy in high cell-density fermentation.

PubMed

Shang, Tingting; Si, Dayong; Zhang, Dongyan; Liu, Xuhui; Zhao, Longmei; Hu, Cong; Fu, Yu; Zhang, Rijun

2017-06-21

Xylanase degrades xylan into monomers of various sizes by catalyzing the endohydrolysis of the 1,4-β-D-xylosidic linkage randomly, possessing potential in wide industrial applications. Most of xylanases are susceptible to be inactive when suffering high temperature and high alkaline process. Therefore, it is necessary to develop a high amount of effective thermoalkaliphilic xylanases. This study aims to enhance thermoalkaliphilic xylanase production in Pichia pastoris through fermentation parameters optimization and novel efficient fed-batch strategy in high cell-density fermentation. Recombinant xylanase activity increased 12.2%, 7.4%, 12.0% and 9.9% by supplementing the Pichia pastoris culture with 20 g/L wheat bran, 5 mg/L L-histidine, 10 mg/L L-tryptophan and 10 mg/L L-methionine in shake flasks, respectively. Investigation of nutritional fermentation parameters, non-nutritional fermentation parameters and feeding strategies in 1 L bioreactor and 1 L shake flask revealed that glycerol and methanol feeding strategies were the critical factors for high cell density and xylanase activity. In 50 L bioreactor, a novel glycerol feeding strategy and a four-stage methanol feeding strategy with a stepwise increase in feeding rate were developed to enhance recombinant xylanase production. In the initial 72 h of methanol induction, the linear dependence of xylanase activity on methanol intake was observed (R 2  = 0.9726). The maximum xylanase activity was predicted to be 591.2 U/mL, while the actual maximum xylanase activity was 560.7 U/mL, which was 7.05 times of that in shake flask. Recombinant xylanase retained 82.5% of its initial activity after pre-incubation at 80 °C for 50 min (pH 8.0), and it exhibited excellent stability in the broad temperature (60-80 °C) and pH (pH 8.0-11.0) ranges. Efficient glycerol and methanol fed-batch strategies resulting in desired cell density and xylanase activity should be applied in other P. pastoris

Astilbe thunbergii reduces postprandial hyperglycemia in a type 2 diabetes rat model via pancreatic alpha-amylase inhibition by highly condensed procyanidins.

PubMed

Kato, Eisuke; Kushibiki, Natsuka; Inagaki, Yosuke; Kurokawa, Mihoko; Kawabata, Jun

2017-09-01

Type 2 diabetes mellitus (T2DM) is a common global health problem. Prevention of this disease is an important task, and functional food supplements are considered an effective method. We found potent pancreatic α-amylase inhibition in Astilbe thunbergii root extract (AT) and confirmed that AT treatment in a T2DM rat model reduces post-starch administration blood glucose levels. Activity-guided isolation revealed procyanidin (AT-P) as the α-amylase inhibitory component with IC 50  = 1.7 μg/mL against porcine pancreatic α-amylase. Structure analysis of AT-P revealed it is a B-type procyanidin comprised of four types of flavan-3-ols, some with a galloyl group, and catechin attached as the terminal unit. The abundant AT-P content and its comparable α-amylase inhibition to acarbose, the anti-diabetic medicine, suggest that AT is a promising food supplement for diabetes prevention.

Cellulase and Xylanase Production from Three Isolates of Indigenous Endophytic Fungi

NASA Astrophysics Data System (ADS)

Yopi; Tasia, W.; Melliawati, R.

2017-12-01

Cellulases and hemicellulases have good potential to be used in energy production, in pulp, paper, textile industries, as well as in animal feed industries. Moreover, its utilization in food industries also cannot be ignored, among others, cellulase and xylanase roles in bakery, wine, and fruit and vegetables juice production. One of the potential enzyme source is endophytic fungi. Object of this study is to explore the potency of endophytic fungi isolated from medicinal plants as source of cellulolytic and xylanolytic enzymes. HL.47F.216 is endophytic fungi isolated from traditional medicinal plants ironwood tree was determined as xylanase producer. HL.51F.235 from pin-flower tree is cellulase producer, while CBN.6F.29 which produces both xylanase and cellulase is originated from Madagascar periwinkle. HL.47F.216 showed 2.5 cm in clear zone diameter and its xylanase activity was 0.262 U/mL with optimum condition pH 7 at 50°C. HL.51F.235 showed 2.4 cm clear zone diameter and 0.239 U/mL of cellulase activity at pH 5 and 70°C. CBN.6F.29 showed 2.8 cm and 0.394 U/mL (pH 5, 40°C) for its cellulase activity, while 2.3 cm and 0.439 U/mL (pH 8, 70°C) for its xylanase activity. Xylanase from HL.47F.216 and CBN.6F.29 showed low molecular masses of 20 kDa and 37-50 kDa, respectively. Molecular masses for cellulases from HL.51F.235 and CBN.6F.29 were 25 and 50 kDa for HL.51F.235 and 100 kDa for CBN.6F.29. Based on macroscopic and microscopic identification, fungal isolate CBN.6F.29 is a member of Class Coelomycetes, while HL.47F.216 was Acremonium sp. and HL.51F.235 was Aspergillus nigri.

Utilization of deoiled Jatropha curcas seed cake for production of xylanase from thermophilic Scytalidium thermophilum.

PubMed

Joshi, Chetna; Khare, S K

2011-01-01

Jatropha curcas is a major biodiesel crop. Large amount of deoiled cake is generated as by-product during biodiesel production from its seeds. Deoiled J. curcas seed cake was assessed as substrate for the production of xylanase from thermophilic fungus Scytalidium thermophilum by solid-state fermentation. The seed cake was efficiently utilized by S. thermophilum for its growth during which it produced good amount of heat stable extracellular xylanase. The solid-state fermentation conditions were optimized for maximum xylanase production. Under the optimized conditions viz. deoiled seed cake supplemented with 1% oat-spelt xylan, adjusted to pH 9.0, moisture content 1:3 w/v, inoculated with 1×10(6) spores per 5 g cake and incubated at 45 °C, 1455 U xylanase/g deoiled seed cake was obtained. The xylanase was useful in biobleaching of paper pulp. Solid-state fermentation of deoiled cake appears a potentially viable approach for its effective utilization. Copyright © 2010 Elsevier Ltd. All rights reserved.

Purification and characterization of a thermostable hypothetical xylanase from Aspergillus oryzae HML366.

PubMed

He, Haiyan; Qin, Yongling; Li, Nan; Chen, Guiguang; Liang, Zhiqun

2015-03-01

In the current study, fermentation broth of Aspergillus oryzae HML366 in sugar cane bagasse was subjected to ultrafiltration and ion exchange chromatography, and two xylanases, XynH1 and XynH2, were purified. Time-of-flight mass spectrometry coupled with SDS-PAGE analysis revealed that XynH1 is identical to the hypothetical A. oryzae RIB40 protein XP_001826985.1, with a molecular weight of 33.671 kDa. Likewise, XynH2 was identified as xylanase XynF1 with a molecular weight of 35.402 kDa. Sequence analysis indicated that XynH1 belongs to glycosyl hydrolases family 10. The specific activity of XynH1 was measured at 476.9 U/mg. Optimal xylanase activity was observed at pH 6.0, and enzyme remained active within pH 4.0-10.0 and at a temperature below 70 °C. Mg(2+), Mn(2+), Ca(2+), and K(+) enhanced the XynH1 xylanase activity to 146, 122, 114, and 108%, respectively. XynH1 hydrolyzed Birchwood xylan and Larchwood xylan effectively. The K m and V max of XynH1 values determined were 1.16 mM and 336 μmol/min/mg with Birchwood xylan as the substrate. A. oryzae HML366 xylanase XynH1 showed superior heat and pH tolerance, therefore may have significant applications in paper and biofuel industries. These studies constitute the first investigation of the xylanase activities of the hypothetical protein XP_001826985.1 form A. oryzae.

Structural features, substrate specificity, kinetic properties of insect α-amylase and specificity of plant α-amylase inhibitors.

PubMed

Kaur, Rimaljeet; Kaur, Narinder; Gupta, Anil Kumar

2014-11-01

α-Amylase is an important digestive enzyme required for the optimal growth and development of insects. Several insect α-amylases had been purified and their physical and chemical properties were characterized. Insect α-amylases of different orders display variability in structure, properties and substrate specificity. Such diverse properties of amylases could be due to different feeding habits and gut environment of insects. In this review, structural features and properties of several insect α-amylases were compared. This could be helpful in exploring the diversity in characteristics of α-amylase between the members of the same class (insecta). Properties like pH optima are reflected in enzyme structural features. In plants, α-amylase inhibitors (α-AIs) occur as part of natural defense mechanisms against pests by interfering in their digestion process and thus could also provide access to new pest management strategies. AIs are quite specific in their action; therefore, these could be employed according to their effectiveness against target amylases. Potential of transgenics with α-AIs has also been discussed for insect resistance and controlling infestation. The differences in structural features of insect α-amylases provided reasons for their efficient functioning at different pH and the specificity towards various substrates. Various proteinaceous and non-proteinaceous inhibitors discussed could be helpful in controlling pest infestation. In depth detailed studies are required on proteinaceous α-AI-α-amylase interaction at different pH's as well as the insect proteinase action on these inhibitors before selecting the α-AI for making transgenics resistant to particular insect. Copyright © 2014 Elsevier Inc. All rights reserved.

Xylanase supplementation on enzymatic saccharification of dilute acid pretreated poplars at different severities

Treesearch

Chao Zhang; Xinshu Zhuang; Zhao Jiang Wang; Fred Matt; Franz St. John; J.Y. Zhu

2013-01-01

Three pairs of solid substrates from dilute acid pretreatment of two poplar wood samples were enzymatically hydrolyzed by cellulase preparations supplemented with xylanase. Supplementation of xylanase improved cellulose saccharification perhaps due to improved cellulose accessibility by xylan hydrolysis. Total xylan removal directly affected enzymatic cellulose...

Pectinase activity determination: an early deceleration in the release of reducing sugars throws a spanner in the works!

PubMed

Biz, Alessandra; Farias, Fernanda Cardoso; Motter, Francine Aline; de Paula, Diogo Henrique; Richard, Peter; Krieger, Nadia; Mitchell, David Alexander

2014-01-01

Recently, it has been suggested that pectinases could be used to hydrolyze pectin in biorefineries based on pectin-rich agro-industrial wastes. However, for this to be viable, the cost of their production would need to be lowered significantly. In fact, over the last few decades, there have been many attempts to improve pectinase production by existing strains or to screen for new strains from environmental isolates. In these studies, it is necessary to measure pectinase activities. Many researchers use single-time-point assays that involve incubation of pectinolytic extracts with pectic substrates for a fixed time, followed by determination of the liberated reducing sugars. However, different researchers use quite different conditions for this assay. Furthermore, no attention has been given to the reaction profile during the assay. In the current work, we show, for the first time, that a significant deceleration of the rate of liberation of reducing sugars occurs over the first ten minutes of the reaction. As a consequence, the incubation time used in a single-time-point assay has a large effect on the value obtained for the activity. In fact, we demonstrate that, depending on the particular combination of incubation time, pectin concentration and reaction temperature, the same extract could be reported to have activities that differ by an order of magnitude. In addition, we show that the relative activities obtained with polygalacturonic acid do not correlate with those obtained with pectin. We conclude that it is currently impossible to make meaningful comparisons between pectinase activities reported in the literature by workers who have used different assay conditions. Therefore there is an urgent need for the development of a standardized assay for evaluating the saccharification potential of pectinase complexes.

Pectinase Activity Determination: An Early Deceleration in the Release of Reducing Sugars Throws a Spanner in the Works!

PubMed Central

Biz, Alessandra; Farias, Fernanda Cardoso; Motter, Francine Aline; de Paula, Diogo Henrique; Richard, Peter; Krieger, Nadia; Mitchell, David Alexander

2014-01-01

Recently, it has been suggested that pectinases could be used to hydrolyze pectin in biorefineries based on pectin-rich agro-industrial wastes. However, for this to be viable, the cost of their production would need to be lowered significantly. In fact, over the last few decades, there have been many attempts to improve pectinase production by existing strains or to screen for new strains from environmental isolates. In these studies, it is necessary to measure pectinase activities. Many researchers use single-time-point assays that involve incubation of pectinolytic extracts with pectic substrates for a fixed time, followed by determination of the liberated reducing sugars. However, different researchers use quite different conditions for this assay. Furthermore, no attention has been given to the reaction profile during the assay. In the current work, we show, for the first time, that a significant deceleration of the rate of liberation of reducing sugars occurs over the first ten minutes of the reaction. As a consequence, the incubation time used in a single-time-point assay has a large effect on the value obtained for the activity. In fact, we demonstrate that, depending on the particular combination of incubation time, pectin concentration and reaction temperature, the same extract could be reported to have activities that differ by an order of magnitude. In addition, we show that the relative activities obtained with polygalacturonic acid do not correlate with those obtained with pectin. We conclude that it is currently impossible to make meaningful comparisons between pectinase activities reported in the literature by workers who have used different assay conditions. Therefore there is an urgent need for the development of a standardized assay for evaluating the saccharification potential of pectinase complexes. PMID:25337712

Production, Purification, and Characterization of a Major Penicillium glabrum Xylanase Using Brewer's Spent Grain as Substrate

PubMed Central

Beitel, Susan Michelz; Fortkamp, Diana; Terrasan, César Rafael Fanchini; de Almeida, Alex Fernando

2013-01-01

In recent decades, xylanases have been used in many processing industries. This study describes the xylanase production by Penicillium glabrum using brewer's spent grain as substrate. Additionally, this is the first work that reports the purification and characterization of a xylanase using this agroindustrial waste. Optimal production was obtained when P. glabrum was grown in liquid medium in pH 5.5, at 25 °C, under stationary condition for six days. The xylanase from P. glabrum was purified to homogeneity by a rapid and inexpensive procedure, using ammonium sulfate fractionation and molecular exclusion chromatography. SDS-PAGE analysis revealed one band with estimated molecular mass of 18.36 kDa. The optimum activity was observed at 60 °C, in pH 3.0. The enzyme was very stable at 50 °C, and high pH stability was verified from pH 2.5 to 5.0. The ion Mn2+ and the reducing agents β-mercaptoethanol and DTT enhanced xylanase activity, while the ions Hg2+, Zn2+, and Cu2+ as well as the detergent SDS were strong inhibitors of the enzyme. The use of brewer's spent grain as substrate for xylanase production cannot only add value and decrease the amount of this waste but also reduce the xylanase production cost. PMID:23762855

Purification and Characterization of a Highly Efficient Calcium-Independent α-Amylase from Talaromyces pinophilus 1-95

PubMed Central

Xian, Liang; Wang, Fei; Luo, Xiang; Feng, Yu-Liang; Feng, Jia-Xun

2015-01-01

Alpha-amylase is a very important enzyme in the starch conversion process. Most of the α-amylases are calcium-dependent and exhibit poor performance in the simultaneous saccharification and fermentation process of industrial bioethanol production that uses starch as feedstock. In this study, an extracellular amylolytic enzyme was purified from the culture broth of newly isolated Talaromyces pinophilus strain 1-95. The purified amylolytic enzyme, with an apparent molecular weight of 58 kDa on SDS-PAGE, hydrolyzed maltopentaose, maltohexaose, and maltoheptaose into mainly maltose and maltotriose and minor amount of glucose, confirming the endo-acting mode of the enzyme, and hence, was named Talaromyces pinophilus α-amylase (TpAA). TpAA was most active at pH 4.0–5.0 (with the temperature held at 37°C) and 55°C (at pH 5.0), and stable within the pH range of 5.0–9.5 (at 4°C) and below 45°C (at pH 5.0). Interestingly, the Ca2+ did not improve its enzymatic activity, optimal temperature, or thermostability of the enzyme, indicating that the TpAA was Ca2+-independent. TpAA displayed higher enzyme activity toward malto-oligosaccharides and dextrin than other previously reported α-amylases. This highly active Ca2+-independent α-amylase may have potential applications in starch-to-ethanol conversion process. PMID:25811759

Effects of structure and xylanase treatment of brewers' spent grain on performance and nutrient availability in broiler chickens.

PubMed

Denstadli, V; Westereng, B; Biniyam, H G; Ballance, S; Knutsen, S H; Svihus, B

2010-06-01

1. A factorial (2 x 3) feeding trial was set up to investigate the effects of coarse or finely ground brewers' spent grain (BSG) and xylanase treatment, either with no xylanase, top-dressed with xylanase or pre-treated with xylanase. 2. The experimental diets shared the same basal formulation and were fed to male broiler chickens (Ross 308) housed in individual cages from 12 to 29 d of age. 3. Xylanase pre-treatment reduced the dietary concentration of arabinoxylan by 15-30%. Pellet durability increased when BSG was ground. 4. Feed utilisation was significantly higher (6%) when the birds were given coarse BSG rather than ground BSG, whereas there was no significant effect of enzyme treatment. Apparent metabolisable energy was unaffected by the dietary treatments. 5. The overall starch digestibility was high (99%), with no dietary differences, whereas ileal protein digestibility was low (57%). Xylanase top-dressing tended to improve ileal protein digestibility but, in general, xylanase treatment had no major effect on overall performance in male broilers given diets with BSG.

Mapping the intestinal alpha-glucogenic enzyme specificities of starch digesting maltase-glucoamylase and sucrase-isomaltase

USDA-ARS?s Scientific Manuscript database

Inhibition of intestinal alpha-glucosidases and pancreatic alpha-amylases is an approach to controlling blood glucose and serum insulin levels in individuals with Type II diabetes. The two human intestinal glucosidases are maltase-glucoamylase and sucrase-isomaltase. Each incorporates two family 31 ...

Effect of exogenous xylanase on rumen in vitro gas production and degradability of wheat straw.

PubMed

Togtokhbayar, Norovsambuu; Cerrillo, María A; Rodríguez, Germán Buendía; Elghandour, Mona M M Y; Salem, Abdelfattah Z M; Urankhaich, Chuluunbaatar; Jigjidpurev, Sukhbaatar; Odongo, Nicholas E; Kholif, Ahmed E

2015-08-01

The objective of this study was to determine effects of xylanase on in vitro gas production (GP) and in sacco degradability of wheat straw. Rumen fluid was obtained from three Mongolian native goats fitted with permanent rumen cannulas. The trial consisted of five doses (0, 0.5, 1.0, 1.5, 2.0 μL/g of substrate) of a commercial xylanase (Dyadic® xylanase PLUS, Dyadic International, Inc., Jupiter, FL, USA). For the in sacco degradability, different levels of xylanase enzyme were added directly onto 2 g of wheat straw in nylon bags and incubated in the rumen for 3, 6, 12, 24 and 48 h to estimate degradability of wheat straw. Total GP increased (P < 0.001) at all times of incubation at intermediate levels of xylanase. Methane production had a similar pattern at 3 and 12 h of incubation; increased linearly at 24 h of incubation, and was unaffected at 6 and 48 h of incubation. Rumen NH3 -N concentration increased linearly at 3 h and the highest values were observed with intermediate enzyme levels. All ruminal volatile fatty acids increased linearly with intermediate levels of the fibrolytic enzyme. The in sacco rate of dry matter degradation decreased linearly (P = 0.020) with increasing enzymes. Intermediate levels of xylanase improved rumen kinetic fermentation and degradability. The outcome of this research indicated that the application of xylanase enzyme could improve in vitro GP fermentation of wheat straw. © 2015 Japanese Society of Animal Science.

Xylanase production from marine derived Trichoderma pleuroticola 08ÇK001 strain isolated from Mediterranean coastal sediments.

PubMed

Korkmaz, Melih N; Ozdemir, Sennur C; Uzel, Ataç

2017-10-01

Xylanases constitutes one the most important enzymes with diverse applications in different industries such as bioethanol production, animal feedstock production, production of xylo-oligosaccharides, baking industry, paper and pulp industry, xylitol production, fruit juice, and beer finishing, degumming, and agriculture. Currently, industrial xylanases are mainly produced by Aspergillus and Trichoderma members. Since the marine environments are less studied compared to terrestrial environments and harbors great microbial diversity we aimed to investigate the xylanase production of 88 marine-derived filamentous fungal strains. These strains are semi-quantitatively screened for their extracellular xylanase production and Trichoderma pleuroticola 08ÇK001 xylanase activity was further characterized. Optimum pH and temperature was determined as 5.0 and 50 °C, respectively. The enzyme preparation retained 53% of its activity at pH 5.0 after 1 h and have found resistant against several ions and compounds such as K + , Ba 2+ , Na + , β-mercaptoethanol, Triton X-100 and toluene. This study demonstrates that marine-derived fungal strains are prolific sources for xylanase production and presents the first report about the production and characterization of xylanase from a marine derived T. pleuroticola strain. The characteristics of T. pleuroticola 08ÇK001 xylanase activity indicate possible employment in some industrial processes such as animal feed, juice and wine industries or paper pulping applications. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Gene Expression and Molecular Characterization of a Xylanase from Chicken Cecum Metagenome

PubMed Central

AL-Darkazali, Hind; Meevootisom, Vithaya

2017-01-01

A xylanase gene xynAMG1 with a 1,116-bp open reading frame, encoding an endo-β-1,4-xylanase, was cloned from a chicken cecum metagenome. The translated XynAMG1 protein consisted of 372 amino acids including a putative signal peptide of 23 amino acids. The calculated molecular mass of the mature XynAMG1 was 40,013 Da, with a theoretical pI value of 5.76. The amino acid sequence of XynAMG1 showed 59% identity to endo-β-1,4-xylanase from Prevotella bryantii and Prevotella ruminicola and 58% identity to that from Prevotella copri. XynAMG1 has two conserved motifs, DVVNE and TEXD, containing two active site glutamates and an invariant asparagine, characteristic of GH10 family xylanase. The xynAMG1 gene without signal peptide sequence was cloned and fused with thioredoxin protein (Trx.Tag) in pET-32a plasmid and overexpressed in Escherichia coli Tuner™(DE3)pLysS. The purified mature XynAMG1 was highly salt-tolerant and stable and displayed higher than 96% of its catalytic activity in the reaction containing 1 to 4 M NaCl. It was only slightly affected by common organic solvents added in aqueous solution to up to 5 M. This chicken cecum metagenome-derived xylanase has potential applications in animal feed additives and industrial enzymatic processes requiring exposure to high concentrations of salt and organic solvents. PMID:28751915

The constitutive production of pectinase by the CT1 mutant of Penicillium occitainis is modulated by pH.

PubMed

Romdhane, Zamen Ben; Tounsi, Hajer; Hadj-Sassi, Azza; Hadj-Taieb, Noomen; Gargouri, Ali

2013-01-01

The aim of the present study was to investigate pectinases production by CT1 mutant of Penicillium occitanis on glucose based media. Two main groups of pectinases were followed: lyases (pectin and pectate lyases) and hydrolases (polygalacturonases and polymethylgalacturonases). When cultivated in different liquid media, where either the starting glucose concentration or the nature of nitrogen sources used was varied, the CT1 mutant secreted either lyases or hydrolases. In fact, the pH of these various media seemed to correlate with the activity produced: The lyases were highly and exclusively produced at neutral or alkaline ambient pH, whereas hydrolases were highly produced on acidic ambient pH. Such conclusion was confirmed by following pectinase production in the same culture medium (with the same glucose concentration and the same nitrogen source) set at two initial pH of 4 and 7. Altogether, these results suggest that the pectinases control by PacC signaling pathway of P. occitanis should resemble to that of Aspergillus and its ability to "activate the expression of alkaline-expressed genes and repress acid-expressed genes" remains intact in the CT1 over-producing and constitutive strain. Enzymes produced at acidic pH (hydrolases) and at neutral pH (lyases) were applied in the hydrolysis of orange peel and gave results comparable to commercial enzymes.

Immobilization of pectinase onto Fe3O4@SiO2-NH2 and its activity and stability.

PubMed

Fang, Gang; Chen, Honggao; Zhang, Yunpeng; Chen, Anqing

2016-07-01

Fe3O4 magnetic nanometer particles (MNPS) with a diameter of 30-40nm and coated by ammoniated silicon dioxides were developed by sol-gel method. The X-ray diffraction (XRD) pattern indicates that the Fe3O4 is wrapped in the amorphous SiO2, and the Fourier-transform infrared spectroscopy (FT-IR) analysis confirmed the NH2 group had been successfully introduced onto the surface of Fe3O4@SiO2 nanoparticles. The main factors related to the immobilization process of pectinase onto the magnetic support were optimized by multi-factor orthogonal experiments, and the recommended combination is that 3% glutaraldehyde (coupling agent), pH=8.0, 50°C, and cross-linking time 8h. Scanning electron microscope (SEM) and FT-IR analyses confirmed that the pectinase had been immobilized onto the surface of the amino modified nanoparticles. The storage stability and reusability of the pectinase were enhanced by the immobilization that 64.4% of the total starting activity was retrieved after 7 recycling batches and only about 21.1% of the total activity was lost after 30-day storage. For the application of the immobilized pectinase, the optimal pH and temperature were 8.5 and 55°C, respectively. Copyright © 2016 Elsevier B.V. All rights reserved.

Biotransformation of catechin and extraction of active polysaccharide from green tea leaves via simultaneous treatment with tannase and pectinase.

PubMed

Baik, Joo Hyun; Shin, Kwang-Soon; Park, Yooheon; Yu, Kwang-Won; Suh, Hyung Joo; Choi, Hyeon-Son

2015-08-30

Green tea is a dietary source of bioactive compounds for human health. Enzymatic treatments induce the bioconversion of bioactive components, which can improve biological activities. In this study, we investigated the effect of simultaneous treatment with tannase and Rapidase on biotransformation of catechins and extraction of polysaccharide from green tea extract (GTE). Tannase and pectinase treatments induced the biotransformation of catechins and altered tea polysaccharide () content. The addition of GTE to the enzyme reaction resulted in a significant increase in degallated catechins, including gallic acid, a product of the tannase reaction (314.5-4076.0 µg mL(-1)) and a reduction in epigallocatechin gallate (EGCG). Biotransformation of catechins improved the radical scavenging activity of GTE. Pectinase treatment led to change of TPS composition in GTE by hydrolyzing polysaccharides. In addition, pectinase-driven hydrolysis in polysaccharides significantly increased TPS-induced Interleukin 6 (IL-6) production in macrophages. In particular, treatment of Rapidase (TPS-Ra) led to the highest IL-6 production among TPS samples, similar to treatment of highly purified pectinase (TPS-GTE), a positive control. Simultaneous processing with tannase and Rapidase can be an efficient method for the extraction of bioactive polysaccharides and biotransformation of catechins with enhanced radical scavenging activity from green tea. © 2014 Society of Chemical Industry.

Degradation of complex carbohydrate: immobilization of pectinase from Bacillus licheniformis KIBGE-IB21 using calcium alginate as a support.

PubMed

Rehman, Haneef Ur; Aman, Afsheen; Silipo, Alba; Qader, Shah Ali Ul; Molinaro, Antonio; Ansari, Asma

2013-08-15

Pectinases are heterogeneous group of enzymes that catalyse the hydrolysis of pectin substances which is responsible for the turbidity and undesirable cloudiness in fruits juices. In current study, partially purified pectinase from Bacillus licheniformis KIBGE-IB21 was immobilized in calcium alginate beads. The effect of sodium alginate and calcium chloride concentration on immobilization was studied and it was found that the optimal sodium alginate and calcium chloride concentration was 3.0% and 0.2 M, respectively. It was found that immobilization increases the optimal reaction time for pectin degradation from 5 to 10 min and temperature from 45 to 55°C, whereas, the optimal pH remained same with reference to free enzyme. Thermal stability of enzyme increased after immobilization and immobilized pectinase retained more than 80% of its initial activity after 5 days at 30°C as compared with free enzyme which showed only 30% of residual activity. The immobilized enzyme also exhibited good operational stability and 65% of its initial activity was observed during third cycle. In term of pectinase immobilization efficiency and stability, this calcium alginate beads approach seemed to permit good results and can be used to make a bioreactor for various applications in food industries. Copyright © 2013 Elsevier Ltd. All rights reserved.

In-vitro alpha amylase inhibitory activity of the leaf extracts of Adenanthera pavonina.

PubMed

Wickramaratne, M Nirmali; Punchihewa, J C; Wickramaratne, D B M

2016-11-15

Diabetes has caused a major burden to the health sector in the developing countries and has shown an increasing trend among the urban population. It is estimated that most patients are with type II diabetes which could be easily treated with dietary changes, exercise, and medication. Sri Lanka carries a long history ayurvedic medicine where it uses the plant for treating many diseases. Therefore it is important to screen medicinal plants scientifically so they could be used safely and effectively in the traditional medical system and also be used for further investigations. Adenanthera pavonina is a plant used in the Ayurvedic medical system in Sri Lanka for treating many diseases including diabetics. We evaluated the anti-diabetic properties and the antioxidant properties of Adenanthera pavonina leaves. The methanol extract of the leaves was sequentially extracted with petroleum ether and thereafter was partitioned between EtOAc, and water. The α-amylase inhibition assay was performed using the 3,5- dinitrosalicylic acid method. The antioxidant activities were measured using the DPPH free radical scavenging activity and the total phenolic content using Folin-Ciocalteu's reagent. The cytotoxicity of the extract was evaluated using the Brine shrimp bioassay. The IC 50 values of α amylase inhibitory activity of MeOH, EtOAc, petroleum ether, and water were 16.16 ± 2.23, 59.93 ± 0.25, 145.49 ± 4.86 and 214.85 ± 9.72 μg/ml respectively and was similar to that of Acarbose (18.63 ± 1.21 (μg/ml). Antioxidant activities were also determined and the EtOAc fraction showed the highest total phenolic content (34. 62 ± 1.14 mg/g extract) and the highest DPPH scavenging activity with an IC 50 of 249.92 ± 3.35 μg/ml. The leaf extracts of Adenanthera pavonina exhibit remarkable α-amylase inhibitory activity in the crude methanolic extract. Hence leaves of Adenanthera pavonina has a potential to be used as a regular green vegetable and

Enzymatic production of maltodextrins derived from sago flour using heat-stable alpha-amylase and pullulanase

NASA Astrophysics Data System (ADS)

Laga, A.; Syarifuddin, A.; Dirpan, A.

2018-05-01

Maltodextrins are produced by starch modification in a partial hydrolysis thus altered physical sago properties. Sago as one of starch resources has characteristic with high amylopectin that influences high viscosity during cooking. Partial hydrolysis or liquefaction will influences starch hydrolysis and the size of maltodextrin produced. The aim of this study was to analyze the degree of sago starch hydrolysis during the enzymatic process using single α-amylase and combination with pullulanase The starting solids content was 20% (w/v), with adjusted pH to 6.5, and calcium (Ca2+ ions) addition as high as 50 ppm. The majority of starches used in the study contain 0.2 % (w/v), to combination of 0.2 % (w/w) and 0, 3 gram per kg of sago. The sago suspension temperatures were started from 105 °C lowered to 60 °C for 30 minutes, respectively. Optimum liquefied starch yields, which accounted for virtually all of the starch present, were obtained at temperatures of 80°C and above, for 120 minutes, with each sampling every 20 minutes. Observed parameters were levels of reducing sugars, degree of hydrolysis, and refined sago starch. The result showed that there was a significant increase in reducing sugars, degree of hydrolysis during 120 minutes until liquefaction process for both enzymatic treatments. The amount of reducing sugars was 95.76 g/L at 120 min for the single α-amylase and 98.84 g/L combination with pullulanase. The degree of hydrolysis was 37.93 % at 120 minutes for the single α-amylase and 37.32 % combination with pullulanase, whereas 0.035 % and 0.038 % for refined sago starch value respectively.

Developing a xylanase XYNZG from Plectosphaerella cucumerina for baking by heterologously expressed in Kluyveromyces lactis.

PubMed

Zhan, Fei Xiang; Wang, Qin Hong; Jiang, Si Jing; Zhou, Yu Ling; Zhang, Gui Min; Ma, Yan He

2014-12-16

Xylanase can replace chemical additives to improve the volume and sensory properties of bread in the baking. Suitable baking xylanase with improved yield will promote the application of xylanase in baking industry. The xylanase XYNZG from the Plectosphaerella cucumerina has been previously characterized by heterologous expression in Pichia pastoris. However, P. pastoris is not a suitable host for xylanase to be used in the baking process since P. pastoris does not have GRAS (Generally Regarded As Safe) status and requires large methanol supplement during the fermentation in most conditions, which is not allowed to be used in the food industry. Kluyveromyces lactis, as another yeast expression host, has a GRAS status, which has been successfully used in food and feed applications. No previous work has been reported concerning the heterologous expression of xylanase gene xynZG in K. lactis with an aim for application in baking. The xylanase gene xynZG from the P. cucumerina was heterologously expressed in K. lactis. The recombinant protein XYNZG in K. lactis presented an approximately 19 kDa band on SDS-PAGE and zymograms analysis. Transformant with the highest halo on the plate containing the RBB-xylan (Remazol Brilliant Blue-xylan) was selected for the flask fermentation in different media. The results indicated that the highest activity of 115 U/ml at 72 h was obtained with the YLPU medium. The mass spectrometry analysis suggested that the hydrolytic products of xylan by XYNZG were mainly xylobiose and xylotriose. The results of baking trials indicated that the addition of XYNZG could reduce the kneading time of dough, increase the volume of bread, improve the texture, and have more positive effects on the sensory properties of bread. Xylanase XYNZG is successfully expressed in K. lactis, which exhibits the highest activity among the published reports of the xylanase expression in K. lactis. The recombinant XYNZG can be used to improve the volume and sensory

Optimization of covalent immobilization of pectinase on sodium alginate support.

PubMed

Li, Tuoping; Wang, Na; Li, Suhong; Zhao, Qiancheng; Guo, Mei; Zhang, Cheyun

2007-09-01

Pectinase was immobilized on a sodium alginate support using glutaraldehyde and retained 66% activity. The optimal pH for activity shifted from 3.0 to 3.5 after immobilization; however, the optimum temperature remained unchanged at 40 degrees C. The immobilized enzyme also had a higher thermal stability and reusability than the free enzyme, and retained 80% of initial activity after 11 batch reactions.

Xylanases, nucleic acids encoding them and methods for making and using them

DOEpatents

Gray, Kevin A; Dirmeier, Reinhard

2013-07-16

The invention relates to enzymes having xylanase, mannanase and/or glucanase activity, e.g., catalyzing hydrolysis of internal .beta.-1,4-xylosidic linkages or endo-.beta.-1,4-glucanase linkages; and/or degrading a linear polysaccharide beta-1,4-xylan into xylose. Thus, the invention provides methods and processes for breaking down hemicellulose, which is a major component of the cell wall of plants, including methods and processes for hydrolyzing hemicelluloses in any plant or wood or wood product, wood waste, paper pulp, paper product or paper waste or byproduct. In addition, methods of designing new xylanases, mannanases and/or glucanases and methods of use thereof are also provided. The xylanases, mannanases and/or glucanases have increased activity and stability at increased pH and temperature.

Irradiation of industrial enzyme preparations. II. Characterization of fungal pectinase by thin-layer isoelectric focusing and gel filtration

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

Delincee, H.

1978-01-01

Industrial dry fungal pectinase from A. niger was irradiated with doses (up to 1 Mrad) of /sup 60/Co-..gamma..rays effective in reducing microbial contamination. The pectinase was characterized by thin-layer isoelectric focusing and gel filtration in order to detect possible radiation-induced structural alterations. Thin-layer isoelectric focusing revealed at least fifteen multiple forms with pectin-depolymerizing activity, with isoelectric points in the range pH 4.5 to 7. Heterogeneity of pectinesterase was also demonstrated, the main band occurring around pH 4. By thin-layer gel filtration the molecular weight of the pectin-depolymerase was estimated as being about 36,000, and that of pectinesterase as about 33,000.more » Radiation-induced changes of the charge properties or molecular size of the irradiated pectinase preparation were not observed. The feasibility of using ionizing radiation for the reduction of microbial contamination of industrial enzyme preparations looks promising.« less

Aureobasidium pullulans xylanase, gene and signal sequence

DOEpatents

Xin-Liang, Li; Ljungdahl, Lars G.

1997-01-01

A xylanase from Aureobasidium pullulans having a high specific activity is provided as well as a signal protein for controlling excretion into cell culture medium of proteins to which it is attached. DNA encoding these proteins is also provided.

Dry-grind processing using amylase corn and superior yeast to reduce the exogenous enzyme requirements in bioethanol production.

PubMed

Kumar, Deepak; Singh, Vijay

2016-01-01

Conventional corn dry-grind ethanol production process requires exogenous alpha and glucoamylases enzymes to breakdown starch into glucose, which is fermented to ethanol by yeast. This study evaluates the potential use of new genetically engineered corn and yeast, which can eliminate or minimize the use of these external enzymes, improve the economics and process efficiencies, and simplify the process. An approach of in situ ethanol removal during fermentation was also investigated for its potential to improve the efficiency of high-solid fermentation, which can significantly reduce the downstream ethanol and co-product recovery cost. The fermentation of amylase corn (producing endogenous α-amylase) using conventional yeast and no addition of exogenous α-amylase resulted in ethanol concentration of 4.1 % higher compared to control treatment (conventional corn using exogenous α-amylase). Conventional corn processed with exogenous α-amylase and superior yeast (producing glucoamylase or GA) with no exogenous glucoamylase addition resulted in ethanol concentration similar to control treatment (conventional yeast with exogenous glucoamylase addition). Combination of amylase corn and superior yeast required only 25 % of recommended glucoamylase dose to complete fermentation and achieve ethanol concentration and yield similar to control treatment (conventional corn with exogenous α-amylase, conventional yeast with exogenous glucoamylase). Use of superior yeast with 50 % GA addition resulted in similar increases in yield for conventional or amylase corn of approximately 7 % compared to that of control treatment. Combination of amylase corn, superior yeast, and in situ ethanol removal resulted in a process that allowed complete fermentation of 40 % slurry solids with only 50 % of exogenous GA enzyme requirements and 64.6 % higher ethanol yield compared to that of conventional process. Use of amylase corn and superior yeast in the dry-grind processing industry

Aureobasidium pullulans xylanase, gene and signal sequence

DOEpatents

Li Xinliang; Ljungdahl, L.G.

1997-01-07

A xylanase from Aureobasidium pullulans having a high specific activity is provided, as well as a signal protein for controlling excretion into cell culture medium of proteins to which it is attached. DNA encoding these proteins is also provided. 4 figs.

Response of broiler chickens fed wheat-based diets to xylanase supplementation.

PubMed

Gonzalez-Ortiz, G; Sola-Oriol, D; Martinez-Mora, M; Perez, J F; Bedford, M R

2017-08-01

The objective of this study was to evaluate the effect of xylanase supplementation on performance, footpad score (FPS), nutrient digestibility, and intestinal morphometry in broiler chickens. Two-hundred-eighty-eight Ross 308 broiler chicks (one d old) were placed in one of 3 experimental treatments: positive control (PC), negative control (NC) (-150 kcal/kg), and XYL (NC supplemented with xylanase). Each treatment had 8 replicates with 12 animals each. Starter (zero to 21 d) and grower (21 to 42 d) diets, based on wheat and soy-bean meal, were available ad libitum. Body weight gain (BWG) and feed intake were measured, and mortality corrected feed conversion ratio (FCR) was calculated. The relative weights of the empty gastrointestinal tract compartments and FPS were recorded on d 21 and d 42. On d 42, ileal contents were collected for nutrient digestibility determination. Statistical comparisons were performed using one-way ANOVA (JMP Pro 12). The reduction of energy resulted in lighter birds at the end of the study (PC: 2,710 vs. NC: 2,546 g; P = 0.030) whereas xylanase supplementation numerically increased BWG by 84 g (P = 0.229) and improved FCR by 12 points (P = 0.145) compared with the NC. Significant differences in FPS were observed on d 21. Score 0 (no lessions) was predominant in PC and XYL treatments, while score 1 (mild lesions) had a higher frequency in NC birds. Xylanase supplementation numerically increased organic matter (5.9%) and energy (4.7%) utilization with values above those observed with the NC. No treatment effects were observed in any of the morphometric measurements, with the exception of the gizzard (P = 0.036) and the ileum (P = 0.088) on d 42. Xylanase resulted in higher relative gizzard weights compared to NC (P = 0.102). Supplementation of broiler diets with xylanase tended to influence performance, which may be due to a better utilization of nutrients. The increase in gizzard and ileum weights in birds 42 d but not 21 d of age

Immobilization of Aspergillus niger xylanase on magnetic latex beads.

PubMed

Tyagi, R; Gupta, M N

1995-04-01

Xylanase from Pectinex 3XL was purified 70-fold by precipitation with an enteric polymer, Eudragit S-100. The purified xylanase was immobilized on magnetic latex beads via carbodi-imide coupling. The immobilized preparation showed 80% of the total activity bound to the beads. The pH optimum remained unchanged at 6.0 and the Km increased from 0.25 g/100 ml (free enzyme) to 0.39 g/100 ml on immobilization. Immobilization resulted in significant thermal stability at 60 degrees C. The time course of hydrolysis of xylan at 60 degrees C by free enzyme as well as immobilized enzyme was also studied.

High-resolution α-amylase assay combined with high-performance liquid chromatography-solid-phase extraction-nuclear magnetic resonance spectroscopy for expedited identification of α-amylase inhibitors: proof of concept and α-amylase inhibitor in cinnamon.

PubMed

Okutan, Leyla; Kongstad, Kenneth T; Jäger, Anna K; Staerk, Dan

2014-11-26

Type 2 diabetes affects millions of people worldwide, and new improved drugs or functional foods containing selective α-amylase inhibitors are needed for improved management of blood glucose. In this article the development of a microplate-based high-resolution α-amylase inhibition assay with direct photometric measurement of α-amylase activity is described. The inhibition assay is based on porcine pancreatic α-amylase with 2-chloro-4-nitrophenyl-α-D-maltotriose as substrate, which this gives a stable, sensitive, and cheap inhibition assay as requested for high-resolution purposes. In combination with HPLC-HRMS-SPE-NMR, this provides an analytical platform that allows simultaneous chemical and biological profiling of α-amylase inhibitors in plant extracts. Proof-of-concept with an artificial mixture of six compounds-of which three are known α-amylase inhibitors-showed that the high-resolution α-amylase inhibition profiles allowed detection of sub-microgram amounts of the α-amylase inhibitors. Furthermore, the high-resolution α-amylase inhibition assay/HPLC-HRMS-SPE-NMR platform allowed identification of cinnamaldehyde as the α-amylase inhibitor in cinnamon (Cinnamomum verum Presl.).

Effect of additives on adsorption and desorption behavior of xylanase on acid-insoluble lignin from corn stover and wheat straw.

PubMed

Li, Yanfei; Ge, Xiaoyan; Sun, Zongping; Zhang, Junhua

2015-06-01

The competitive adsorption between cellulases and additives on lignin in the hydrolysis of lignocelluloses has been confirmed, whereas the effect of additives on the interaction between xylanase and lignin is not clear. In this work, the effects of additives, poly(ethylene glycol) 2000, poly(ethylene glycol) 6000, Tween 20, and Tween 80, on the xylanase adsorption/desorption onto/from acid-insoluble lignin from corn stover (CS-lignin) and wheat straw (WS-lignin) were investigated. The results indicated that the additives could adsorb onto isolated lignin and reduce the xylanase adsorption onto lignin. Compared to CS-lignin, more additives could adsorb onto WS-lignin, making less xylanase adsorbed onto WS-lignin. In addition, the additives could enhance desorption of xylanase from lignin, which might be due to the competitive adsorption between xylanase and additives on lignin. The released xylanase from lignin still exhibited hydrolytic capacity in the hydrolysis of isolated xylan and xylan in corn stover. Copyright © 2015 Elsevier Ltd. All rights reserved.

Modulation of starch digestion for slow glucose release through "toggling" of activities of mucosal "alpha"-glucosidases

USDA-ARS?s Scientific Manuscript database

Starch digestion involves the breakdown by alpha-amylase to small linear and branched malto-oligosaccharides, which are in turn hydrolyzed to glucose by the mucosal alpha-glucosidases, maltase-glucoamylase (MGAM) and sucrase-isomaltase (SI). MGAM and SI are anchored to the small intestinal brush-bor...

Xylanases, nucleic acids encoding them and methods for making and using them

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

Gray, Kevin A.; Dirmeier, Richard

The invention relates to enzymes having xylanase, mannanase and/or glucanase activity, e.g., catalyzing hydrolysis of internal .beta.-1,4-xylosidic linkages or endo-.beta.-1,4-glucanase linkages; and/or degrading a linear polysaccharide beta-1,4-xylan into xylose. Thus, the invention provides methods and processes for breaking down hemicellulose, which is a major component of the cell wall of plants, including methods and processes for hydrolyzing hemicelluloses in any plant or wood or wood product, wood waste, paper pulp, paper product or paper waste or byproduct. In addition, methods of designing new xylanases, mannanases and/or glucanases and methods of use thereof are also provided. The xylanases, mannanases and/or glucanasesmore » have increased activity and stability at increased pH and temperature.« less

Enzyme-synthesized highly branched maltodextrins have slow glucose generation at the mucosal alpha-glucosidase level and are slowly digestible "in vivo"

USDA-ARS?s Scientific Manuscript database

For digestion of starch in humans, alpha-amylase first hydrolyzes starch molecules to produce alpha-limit dextrins, followed by complete hydrolysis to glucose by the mucosal alpha-glucosidases in the small intestine. It is known that alpha-1,6 linkages in starch are hydrolyzed at a lower rate than a...

Sequence-structural features and evolutionary relationships of family GH57 α-amylases and their putative α-amylase-like homologues.

PubMed

Janeček, Stefan; Blesák, Karol

2011-08-01

The glycoside hydrolase family 57 (GH57) contains α-amylase and a few other amylolytic specificities. It counts ~400 members from Archaea (1/4) and Bacteria (3/4), mostly of extremophilic prokaryotes. Only 17 GH57 enzymes have been biochemically characterized. The main goal of the present bioinformatics study was to analyze sequences having the clear GH57 α-amylase features. Of the 107 GH57 sequences, 59 were evaluated as α-amylases (containing both GH57 catalytic residues), whereas 48 were assigned as GH57 α-amylase-like proteins (having a substitution in one or both catalytic residues). Forty-eight of 59 α-amylases were from Archaea, but 42 of 48 α-amylase-like proteins were of bacterial origin. The catalytic residues were substituted in most cases in Bacteroides and Prevotella by serine (instead of catalytic nucleophile glutamate) and glutamate (instead of proton donor aspartate). The GH57 α-amylase specificity has thus been evolved and kept enzymatically active mainly in Archaea.

Contribution of protein, starch, and fat to the apparent ileal digestible energy of corn- and wheat-based broiler diets in response to exogenous xylanase and amylase without or with protease.

PubMed

Romero, L F; Sands, J S; Indrakumar, S E; Plumstead, P W; Dalsgaard, S; Ravindran, V

2014-10-01

The ileal energy contribution of protein, starch, and fat in response to 2 exogenous enzyme combinations was studied in 2 digestibility assays with 21- (experiment 1; 432 birds) and 42-d-old (experiment 2; 288 birds) Ross 308 broiler chickens. A 2 × 2 × 3 factorial arrangement of treatments with 2 base grains (corn or wheat), without or with high fiber ingredients (corn distillers dried grains with solubles and canola meal), and 3 enzyme treatments was implemented. Enzyme treatments, fed from 12 to 21 d or 32 to 42 d, were 1) without enzymes, 2) with xylanase from Trichoderma ressei (2,000 U/kg) and amylase from Bacillus licheniformis (200 U/kg; XA), or 3) with XA plus protease from Bacillus subtilis (4,000 U/kg; XAP). All diets contained Escherichia coli phytase (500 FTU/kg). Apparent ileal digestibility (AID) of protein, starch, and fat, as well as the apparent ileal digestible energy, were determined using titanium dioxide as inert marker. A generalized mixed model was used to test main effects and 2-way interactions at P < 0.05. An enzyme × grain interaction was detected for AID of starch at 21 and 42 d, and AID of fat at 21 d, with greater effects of enzymes in wheat-based compared with corn-based diets, but significant increments due to enzymes compared with controls in both diet types. Apparent ileal digestibility of fat at 42 d increased with enzyme supplementation compared with the control treatments. The XA and XAP treatments gradually (P < 0.05) increased AID of protein at 21 d, but only XAP increased AID of protein compared with the control at 42 d. Compared with the controls, XA increased AID energy by 52 or 87 kcal, and XAP by 104 or 152 kcal/kg of DM at 21 or 42 d, respectively. The caloric contribution of starch, fat, and protein were affected differentially by base grain and the presence of fibrous ingredients at 21 and 42 d of age. ©2014 Poultry Science Association Inc.

Thermodynamic and kinetic studies on pectinase extracted from Aspergillus aculeatus: Free and immobilized enzyme entrapped in alginate beads.

PubMed

de Oliveira, Rodrigo Lira; da Silva, Osmar Soares; Converti, Attilio; Porto, Tatiana Souza

2018-05-01

The kinetics and thermodynamics of Aspergillus aculeatus pectinase, either free or immobilized in alginate beads, were investigated. Pectinase immobilization ensured an enzyme immobilization yield of 59.71%. The irreversible denaturation of pectinase in both preparations was evaluated at temperatures ranging from 30 to 60 °C. When temperature was raised, the first-order thermal denaturation constant increased from 0.0011 to 0.0231 min -1 for the free enzyme and from 0.0017 to 0.0700 min -1 for the immobilized one, respectively. The results of residual activity tests enabled us to estimate, for denaturation of both free and immobilized pectinase, the activation energy (E ⁎ d  = 85.1 and 101.6 kJ·mol -1 ), enthalpy (82.59 ≤ ΔH ⁎ d  ≤ 82.34 kJ·mol -1 and 99.11 ≤ ΔH ⁎ d  ≤ 98.86 kJ·mol -1 ), entropy (-63.26 ≤ ΔS ⁎ d  ≤ -63.85 J·mol -1 ·K -1 and -5.50 ≤ ΔS ⁎ d  ≤ -5.23 J·mol -1 ·K -1 ) and Gibbs free energy (101.8 ≤ ΔG ⁎ d  ≤ 104.7 kJ·mol -1 and 100.6 ≤ ΔG ⁎ d  ≤ 102.0 kJ·mol -1 ). The integral activity of a continuous system using the free and immobilized enzyme was also predicted, whose results indicated a satisfactory enzyme long-term thermostability in both preparations at temperatures commonly used to clarify juice. These results suggest that both free and immobilized pectinase from A. aculeatus may be profitably exploited in future food industrial applications, with special concern to the immobilized enzyme because of its reusability. Copyright © 2018 Elsevier B.V. All rights reserved.

Solid-state Fermentation of Xylanase from Penicillium canescens 10-10c in a Multi-layer-packed Bed Reactor

NASA Astrophysics Data System (ADS)

Assamoi, Antoine A.; Destain, Jacqueline; Delvigne, Frank; Lognay, Georges; Thonart, Philippe

Xylanase is produced by Penicillium canescens 10-10c from soya oil cake in static conditions using solid-state fermentation. The impact of several parameters such as the nature and the size of inoculum, bed-loading, and aeration is evaluated during the fermentation process. Mycelial inoculum gives more production than conidial inoculum. Increasing the quantity of inoculum enhances slightly xylanase production. Forced aeration induces more sporulation of strain and reduces xylanase production. However, forced moistened air improves the production compared to production obtained with forced dry air. In addition, increasing bed-loading reduces the specific xylanase production likely due to the incapacity of the Penicillium strain to grow deeply in the fermented soya oil cake mass. Thus, the best cultivation conditions involve mycelial inoculum form, a bed loading of 1-cm height and passive aeration. The maximum xylanase activity is obtained after 7 days of fermentation and attains 10,200 U/g of soya oil cake. These levels are higher than those presented in the literature and, therefore, show all the potentialities of this stock and this technique for the production of xylanase.

Occurrence of serum antibodies against wheat alpha-amylase inhibitor 0.19 in celiac disease.

PubMed

Sánchez, D; Štěpánová-Honzová, S; Hospodková, M; Hoffmanová, I; Hábová, V; Halada, P; Tlaskalová-Hogenová, H; Tučková, L

2018-05-10

The alcohol-soluble fraction of wheat gluten (gliadins) induces in genetically susceptible individuals immunologically mediated celiac disease (CLD). However, gliadins and related cereal proteins are not unique foodstuff targets of CLD patients´ immune system. Non-gluten wheat alpha-amylase inhibitor 0.19 (AAI 0.19) has been found to be capable of activating human monocyte-derived dendritic cells and inducing pro-inflammatory status in intestinal mucosa of patients with celiac disease (CLD). The possible contribution of this reactivity in incomplete remission of CLD patients on a gluten-free diet (GFD) is matter of contention. In an attempt to characterize the antigenicity of AAI 0.19 in patients with active CLD, patients on a GFD and healthy controls we developed ELISA employing wheat recombinant AAI 0.19. Using this test we revealed a significant (P<0.001) elevation of IgA anti-AAI 0.19 antibodies (Ab) in patients with active CLD (12 out of 30 patients were seropositive) but also in CLD patients on a GFD (15/46), in contrast to healthy controls (2/59). Anti-AAI 0.19 IgG Ab levels were increased (P<0.001) only in patients with active CLD (14/30) in contrast to the controls. Interestingly, the levels of anti-AAI 0.19 IgG Ab were decreased in CLD patients on a GFD (P<0.001, 1/46) compared to the controls (1/59). Notably, 20 out of 30 of patients with active CLD were positive either for IgA or for IgG anti-AAI 0.19 Ab. Thus, the majority of CLD patients developed a robust IgA and IgG Ab response against AAI 0.19. These findings may contribute to the broadening of the knowledge about CLD pathogenesis.

Temperature influence on fluorescence intensity and enzyme activity of the fusion protein of GFP and hyperthermophilic xylanase.

PubMed

Zhang, Chong; Liu, Min-Sheng; Xing, Xin-Hui

2009-09-01

By constructing the expression system for fusion protein of GFPmut1 (a green fluorescent protein mutant) with the hyperthermophilic xylanase obtained from Dictyoglomus thermophilum Rt46B.1, the effects of temperature on the fluorescence of GFP and its relationship with the activities of GFP-fused xylanase have been studied. The fluorescence intensities of both GFP and GFP-xylanase have proved to be thermally sensitive, with the thermal sensitivity of the fluorescence intensity of GFP-xylanase being 15% higher than that of GFP. The lost fluorescence intensity of GFP inactivated at high temperature of below 60 degrees C in either single or fusion form can be completely recovered by treatment at 0 degrees C. By the fluorescence recovery of GFP domain at low temperature, the ratios of fluorescence intensity to xylanase activity (Rgfp/Axyl) at 15 degrees C and 37 degrees C have been compared. Even though the numbers of molecules of GFP and xylanase are equivalent, the Rgfp/Axyl ratio at 15 degrees C is ten times of that at 37 degrees C. This is mainly due to the fact that lower temperature is more conducive to the correct folding of GFP than the hyperthermophilic xylanase during the expression. This study has indicated that the ratio of GFP fluorescence to the thermophilic enzyme activity for the fusion proteins expressed at different temperatures could be helpful in understanding the folding properties of the two fusion partners and in design of the fusion proteins.

Pie waste - A component of food waste and a renewable substrate for producing ethanol.

PubMed

Magyar, Margaret; da Costa Sousa, Leonardo; Jayanthi, Singaram; Balan, Venkatesh

2017-04-01

Sugar-rich food waste is a sustainable feedstock that can be converted into ethanol without an expensive thermochemical pretreatment that is commonly used in first and second generation processes. In this manuscript we have outlined the pie waste conversion to ethanol through a two-step process, namely, enzyme hydrolysis using commercial enzyme products mixtures and microbial fermentation using yeast. Optimized enzyme cocktail was found to be 45% alpha amylase, 45% gamma amylase, and 10% pectinase at 2.5mg enzyme protein/g glucan produced a hydrolysate with high glucose concentration. All three solid loadings (20%, 30%, and 40%) produced sugar-rich hydrolysates and ethanol with little to no enzyme or yeast inhibition. Enzymatic hydrolysis and fermentation process mass balance was carried out using pie waste on a 1000g dry weight basis that produced 329g ethanol at 20% solids loading. This process clearly demonstrate how food waste could be efficiently converted to ethanol that could be used for making biodiesel by reacting with waste cooking oil. Copyright © 2017 Elsevier Ltd. All rights reserved.

Cloning and heterologous expression of cellulose free thermostable xylanase from Bacillus brevis.

PubMed

Goswami, Girish K; Krishnamohan, Medichtrla; Nain, Vikrant; Aggarwal, Chetana; Ramesh, Bandarupalli

2014-01-01

Xylanase gene isolated from Bacillus brevis was expressed in E. coli BL21. Sequencing of the gene (Gen Bank accession number: HQ179986) showed that it belongs to family 11 xylanases. The recombinant xylanase was predominantly secreted to culture medium and showed mesophilic nature (optimum activity at 55°C and pH 7.0). The cell free culture medium exhibited 30 IU/ml xylanse activity. The enzyme did not show any cellulose activity and was active under wide range of temperature (40°C to 80°C) and pH (4 to 9). The enzyme showed considerable thermo stability and regained over 90% of activity, when returned to 55°C after boiling for 5 min. These physiochemical properties of B. brevis xylanse show high potential of its applications in paper and pulp industry.

Inhibition of Sunn pest, Eurygaster integriceps, α-amylases by α-amylase inhibitors (T-αAI) from Triticale.

PubMed

Mehrabadi, Mohammad; Bandani, Ali R; Saadati, Fatemeh

2010-01-01

The effect of triticale α-amylases inhibitors on starch hydrolysis catalyzed by the Sunn pest, Eurygaster integriceps Puton (Hemiptera: Scutelleridae) midgut amylases was examined. Biochemical studgawies showed that inhibitors from Triticale (a hybrid of wheat and rye) had inhibitiory effects on E. integriceps α-amylases. The effects of the triticale α-amylase inhibitor (T-αAI) on α-amylase of E. integriceps showed a dose dependent manner of inhibition, e.g. less inhibition of enzyme activity (around 10%) with a lower dose (0.25 mg protein) and high inhibition of enzyme activity (around 80%) when a high dose of inhibitor was used (1.5 mg protein). The enzyme kinetic studies using Michaelis-Menten and Lineweaver-Burk equations showed the K(m) remained constant (0.58%) but the maximum velocity (V(max)) decreased in the presence of a crude extract of Triticale inhibitors, indicating mixed inhibition. The temperature giving 50% inactivation of enzyme (T(50)) during a 30-min incubation at pH 7.0 was 73° C. The maximum inhibitory activity was achieved at 35° C and pH 5.0. Gel assays showed the meaningful inhibition of E. integriceps α-amylases by various concentrations of Triticale inhibitors. Based on the data presented in this study, it could be said that the T-αAI has good inhibitory activity on E. integriceps gut α-amylase.

Inhibition of Sunn Pest, Eurygaster integriceps, α-Amylases by α-Amylase Inhibitors (T-αAI) from Triticale

PubMed Central

Mehrabadi, Mohammad; Bandani, Ali R.; Saadati, Fatemeh

2010-01-01

The effect of triticale α-amylases inhibitors on starch hydrolysis catalyzed by the Sunn pest, Eurygaster integriceps Puton (Hemiptera: Scutelleridae) midgut amylases was examined. Biochemical studgawies showed that inhibitors from Triticale (a hybrid of wheat and rye) had inhibitiory effects on E. integriceps α-amylases. The effects of the triticale α-amylase inhibitor (T-αAI) on α-amylase of E. integriceps showed a dose dependent manner of inhibition, e.g. less inhibition of enzyme activity (around 10%) with a lower dose (0.25 mg protein) and high inhibition of enzyme activity (around 80%) when a high dose of inhibitor was used (1.5 mg protein). The enzyme kinetic studies using Michaelis-Menten and Lineweaver-Burk equations showed the Km remained constant (0.58%) but the maximum velocity (Vmax) decreased in the presence of a crude extract of Triticale inhibitors, indicating mixed inhibition. The temperature giving 50% inactivation of enzyme (T50) during a 30-min incubation at pH 7.0 was 73° C. The maximum inhibitory activity was achieved at 35° C and pH 5.0. Gel assays showed the meaningful inhibition of E. integriceps α-amylases by various concentrations of Triticale inhibitors. Based on the data presented in this study, it could be said that the T-αAI has good inhibitory activity on E. integriceps gut α-amylase. PMID:21062146

Agricultural waste from the tequila industry as substrate for the production of commercially important enzymes.

PubMed

Huitron, C; Perez, R; Sanchez, A E; Lappe, P; Rocha Zavaleta, L

2008-01-01

Approximately 1 million tons of Agave tequilana plants are processed annually by the Mexican Tequila industry generating vast amounts of agricultural waste. The aim of this study was to investigate the potential use of Agave tequilana waste as substrate for the production of commercially important enzymes. Two strains of Aspergillus niger (CH-A-2010 and CH-A-2016), isolated from agave fields, were found to grow and propagate in submerged cultures using Agave tequilana waste as substrate. Isolates showed simultaneous extracellular inulinase, xylanase, pectinase, and cellulase activities. Aspergillus CH-A-2010 showed the highest production of inulinase activity (1.48 U/ml), whereas Aspergillus niger CH-A-2016 produced the highest xylanase (1.52 U/ml) and endo-pectinase (2.7U/ml) activities. In both cases production of enzyme activities was significantly higher on Agave tequilana waste than that observed on lemon peel and specific polymeric carbohydrates. Enzymatic hydrolysis of raw A. tequilana stems and leaves, by enzymes secreted by the isolates yielded maximum concentrations of reducing sugars of 28.2 g/l, and 9.9 g/l respectively. In conclusion, Agave tequilana waste can be utilized as substrate for the production of important biotechnological enzymes.

Salivary alpha-amylase, salivary cortisol, and anxiety during a youth taekwondo championship

PubMed Central

Capranica, Laura; Condello, Giancarlo; Tornello, Francesco; Iona, Teresa; Chiodo, Salvatore; Valenzano, Anna; De Rosas, Mario; Messina, Giovanni; Tessitore, Antonio; Cibelli, Giuseppe

2017-01-01

Abstract The aim of this study was to assess the stress-related responses and the coach's capability to match perceived efforts of youth athletes during a taekwondo championship. Using a cross-sectional study design, salivary cortisol (sC) and alpha-amylase (sAA) were measured in 6 males and 3 females young (11.0 ± 0.9 years) athletes at awakening, 5 minutes before, and 1 minute and 30 minutes after official combats. State anxiety was recorded 60 minutes before the first competition, whereas coach's and athletes’ ratings of perceived exertion (RPE) were obtained at the end of the combats. Time-matched (awakening and pre-competition) salivary samples and trait anxiety were collected 7-day postcompetition during a resting day. No effect for match outcome emerged. No difference emerged between athletes and coach RPEs. Higher (P = .03) state anxiety (41.6 ± 10.9 points) was shown than trait anxiety (34.8 ± 7.1 points). Time-matched sAA were similar. Peak sAA observed at the end of the combat (114.2 ± 108.1 U/mL) was higher (P < .01) than the other samples (range: 20.6–48.1 U/mL), whereas sC increased (P < .05) from awakening (8.0 ± 1.5 nmol/L), with peak levels observed at 30 minutes into the recovery phase (19.3 ± 4.3 nmol/L). Furthermore, pre-competition sC (16.5 ± 4.5 nmol/L) values were higher (P < .01) with respect to time-matched samples during the resting day (4.6 ± 1.0 nmol/L). The 3 athletes engaged in consecutive matches showed a tendency toward increasing sAA and sC. Taekwondo combats pose a high stress on young athletes, eliciting a fast reactivity of the sympathetic-adreno-medullary system relative to the hypothalamic-pituitary-adrenocortical system. Understanding the athlete's efforts during combats, coaches are recommended to apply effective recovery strategies between matches. PMID:28700470

Thermoresistant xylanases from Trichoderma stromaticum: Application in bread making and manufacturing xylo-oligosaccharides.

PubMed

Carvalho, Elck Almeida; Dos Santos Góes, Laís Mendes; Uetanabaro, Ana Paula T; da Silva, Erik Galvão Paranhos; Rodrigues, Luciano Brito; Pirovani, Carlos Priminho; da Costa, Andréa Miura

2017-04-15

The enzymes Xyl1 and Xyl2 from T. stromaticum were purified and identified by mass spectrometry (MALDI-TOF/MS). Xyl1 contained three proteins with similarity to xylanase family 10, 62 and anarabinofuranosidase of the Trichoderma genus and Xyl2 contained a protein with similarity to endo-1,4-β-xylanase. High xylanase activity was found at 50°C for Xyl1 and 60°C for Xyl2 and pH 5.0 for both, retaining more than 80% of activities for one hour at 60°C and pH 5-8. Ag 2+ and β-mercaptoethanol increased while SDS and EDTA inhibited the xylanase activity of both Xyl1 and Xyl2 extracts. The Km and V max values for purified Xyl2 were 9.6mg/mL and 28.57μmol/min/mg, respectively. In application tests, both Xyl1 and Xyl2 were effective in degrading beechwood xylan to produce xylo-oligosaccharides. In baking, adding Xyl1 increased the softness and volume of wheat bread and whole grain bread, qualities increasingly desired by consumers in this segment. Copyright © 2016 Elsevier Ltd. All rights reserved.

The potato amylase inhibitor gene SbAI regulates cold-induced sweetening in potato tubers by modulating amylase activity.

PubMed

Zhang, Huiling; Liu, Jun; Hou, Juan; Yao, Ying; Lin, Yuan; Ou, Yongbin; Song, Botao; Xie, Conghua

2014-09-01

Potato cold-induced sweetening (CIS) is critical for the postharvest quality of potato tubers. Starch degradation is considered to be one of the key pathways in the CIS process. However, the functions of the genes that encode enzymes related to starch degradation in CIS and the activity regulation of these enzymes have received less attention. A potato amylase inhibitor gene known as SbAI was cloned from the wild potato species Solanum berthaultii. This genetic transformation confirmed that in contrast to the SbAI suppression in CIS-resistant potatoes, overexpressing SbAI in CIS-sensitive potatoes resulted in less amylase activity and a lower rate of starch degradation accompanied by a lower reducing sugar (RS) content in cold-stored tubers. This finding suggested that the SbAI gene may play crucial roles in potato CIS by modulating the amylase activity. Further investigations indicated that pairwise protein-protein interactions occurred between SbAI and α-amylase StAmy23, β-amylases StBAM1 and StBAM9. SbAI could inhibit the activities of both α-amylase and β-amylase in potato tubers primarily by repressing StAmy23 and StBAM1, respectively. These findings provide the first evidence that SbAI is a key regulator of the amylases that confer starch degradation and RS accumulation in cold-stored potato tubers. © 2014 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Effects of enzymes to improve sensory quality of frozen dough bread and analysis on its mechanism.

PubMed

Wang, Xuan; Pei, Dudu; Teng, Yuefei; Liang, Jianfen

2018-01-01

Baking quality of frozen dough is negatively affected by dough weakening and by a reduction in both yeast viability and activity during freezing and frozen storage. The objective of this study was to investigate effects of different enzymes, such as α-amylase, xylanase, celluase, glucose oxidase, and lipase on the texture and sensory quality of bread after frozen storage, as well as on dough properties, in terms of fermentation characteristics, freezable water contents and microstructure. Except for α-amylase, other enzymes improved the bread sensory quality and got higher overall acceptability, especially xylanase. Dough fermentative behavior showed that the maximum heights of frozen dough were increased by 33.2, 19.7 and 7.4%, respectively with xylanase, cellulase and lipase. Cellulase lowered gas holding ability of dough. Thermodynamic properties indicated that addition of enzyme decreased the freezable water contents in frozen dough. Scanning electronic microscopy revealed that freezing and frozen storage disrupted dough gluten network causing separation of starch granules from the gluten matrix. Inclusion of cellulase, xylanase and lipase made the frozen dough having a more continuous gluten network and smoother surface, and glucose oxidase increased the stability of the gluten work.

Purification of pectinase from mango (Mangifera indica L. cv. Chokanan) waste using an aqueous organic phase system: a potential low cost source of the enzyme.

PubMed

Amid, Mehrnoush; Abdul Manap, Mohd Yazid; Mustafa, Shuhaimi

2013-07-15

As a novel method of purification, an aqueous organic phase system (AOPS) was employed to purify pectinase from mango waste. The effect of different parameters, such as the alcohol concentration (ethanol, 1-propanol, and 2-propanol), the salt type and concentration (ammonium sulfate, potassium phosphate and sodium citrate), the feed stock crude load, the aqueous phase pH and NaCl concentration, were investigated in the recovery of pectinase from mango peel. The partition coefficient (K), selectivity (S), purification factor (PF) and yield (Y, %) were investigated in this study as important parameters for the evaluation of enzyme recovery. The desirable partition efficiency for pectinase purification was achieved in an AOPS of 19% (w/w) ethanol and 22% (w/w) potassium phosphate in the presence of 5% (w/w) NaCl at pH 7.0. Based on the system, the purification factor of pectinase was enhanced 11.7, with a high yield of 97.1%. Copyright © 2013 Elsevier B.V. All rights reserved.

Effects of a high-pressure treatment on the wheat alpha-amylase inhibitor and its relationship to elimination of allergenicity

NASA Astrophysics Data System (ADS)

Yamamoto, S.; Takanohashi, K.; Hara, T.; Odani, S.; Suzuki, A.; Nishiumi, T.

2010-03-01

In this study, the effects of high-pressure treatment on structure and allergeincity of alpha amylase inhibitor (a-AI) were investigated. The pressure-induced structural changes of α-AI were estimated by fluorescence spectra and by fourth derivative UV-spectroscopy for probed tyrosine residues and by circular dichroism (CD) spectroscopy. The changes in the tertiary structure detected by fluorescence spectra and by fourth derivative UV-spectroscopy under high pressure were indicated at over 300 MPa. Measurements of CD spectroscopy suggested that the effects of a high-pressure treatment on changes in the secondary structure of α-AI were little. From our results, pressure-induced changes of the α-AI structure were not apparent. On the other hands, the IgE-specific binding activities of pressurized α-AI to sera from allergic patients against wheat, which is estimated by observations of dot-blotting, were decreased by high-pressure treatment. It is known that the pressure-induced elimination of allergenicity is related to the tertiary structural changes of allergen molecules. This study are suspected that the epitopes of α-AI do not contain tyrosine residues, and thus the decrease of IgE-specific binding activities is probably caused by the tertiary structural changes of these parts of α-AI.

Gender determines cortisol and alpha-amylase responses to acute physical and psychosocial stress in patients with borderline personality disorder.

PubMed

Inoue, Ayako; Oshita, Harumi; Maruyama, Yoshihiro; Tanaka, Yoshihiro; Ishitobi, Yoshinobu; Kawano, Aimi; Ikeda, Rie; Ando, Tomoko; Aizawa, Saeko; Masuda, Koji; Higuma, Haruka; Kanehisa, Masayuki; Ninomiya, Taiga; Akiyoshi, Jotaro

2015-07-30

Borderline personality disorder (BPD) is characterized by affective instability, unstable relationships, and identity disturbance. We measured salivary alpha-amylase (sAA) and salivary cortisol levels in all participants during exposure to the Trier Social Stress Test (TSST) and an electric stimulation stress. Seventy-two BPD patients were compared with 377 age- and gender- matched controls. The State and Trait versions of the Spielberger Anxiety Inventory test (STAI-S and STAI-T, respectively), the Profile of Mood State (POMS) tests, and the Beck Depression Inventory (BDI), the Depression and Anxiety Cognition Scale (DACS) were administered to participants before electrical stimulation. Following TSST exposure, salivary cortisol levels significantly decreased in female patients and significantly increased in male patients compared with controls. POMS tension-anxiety, depression-dejection, anger-hostility, fatigue, and confusion scores were significantly increased in BPD patients compared with controls. In contrast, vigor scores were significantly decreased in BPD patients relative to controls. Furthermore, STAI-T and STAI-S anxiety scores and BDI scores were significantly increased in BPD patient compared with controls. DACS scores were significantly increased in BPD patient compared with controls. Different stressors (e.g., psychological or physical) induced different responses in the HPA and SAM systems in female or male BPD patients. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

A fluid response: Alpha-amylase reactions to acute laboratory stress are related to sample timing and saliva flow rate.

PubMed

Nagy, Tamás; van Lien, René; Willemsen, Gonneke; Proctor, Gordon; Efting, Marieke; Fülöp, Márta; Bárdos, György; Veerman, Enno C I; Bosch, Jos A

2015-07-01

Salivary alpha-amylase (sAA) is used as a sympathetic (SNS) stress marker, though its release is likely co-determined by SNS and parasympathetic (PNS) activation. The SNS and PNS show asynchronous changes during acute stressors, and sAA responses may thus vary with sample timing. Thirty-four participants underwent an eight-minute memory task (MT) and cold pressor task (CPT). Cardiovascular SNS (pre-ejection period, blood pressure) and PNS (heart rate variability) activity were monitored continuously. Unstimulated saliva was collected repeatedly during and after each laboratory stressor, and sAA concentration (U/ml) and secretion (U/minute) determined. Both stressors increased anxiety. The MT caused an immediate and continued cardiac SNS activation, but sAA concentration increased at task cessation only (+54%); i.e., when there was SNS-PNS co-activation. During the MT sAA secretion even decreased (-35%) in conjunction with flow rate and vagal tone. The CPT robustly increased blood pressure but not sAA. In summary, sAA fluctuations did not parallel changes in cardiac SNS activity or anxiety. sAA responses seem contingent on sample timing and flow rate, likely involving both SNS and PNS influences. Verification using other stressors and contexts seems warranted. Copyright © 2015 Elsevier B.V. All rights reserved.

Contribution of protein, starch, and fat to the apparent ileal digestible energy of corn- and wheat-based broiler diets in response to exogenous xylanase and amylase without or with protease1

PubMed Central

Romero, L. F.; Sands, J. S.; Indrakumar, S. E.; Plumstead, P. W.; Dalsgaard, S.; Ravindran, V.

2014-01-01

The ileal energy contribution of protein, starch, and fat in response to 2 exogenous enzyme combinations was studied in 2 digestibility assays with 21- (experiment 1; 432 birds) and 42-d-old (experiment 2; 288 birds) Ross 308 broiler chickens. A 2 × 2 × 3 factorial arrangement of treatments with 2 base grains (corn or wheat), without or with high fiber ingredients (corn distillers dried grains with solubles and canola meal), and 3 enzyme treatments was implemented. Enzyme treatments, fed from 12 to 21 d or 32 to 42 d, were 1) without enzymes, 2) with xylanase from Trichoderma ressei (2,000 U/kg) and amylase from Bacillus licheniformis (200 U/kg; XA), or 3) with XA plus protease from Bacillus subtilis (4,000 U/kg; XAP). All diets contained Escherichia coli phytase (500 FTU/kg). Apparent ileal digestibility (AID) of protein, starch, and fat, as well as the apparent ileal digestible energy, were determined using titanium dioxide as inert marker. A generalized mixed model was used to test main effects and 2-way interactions at P < 0.05. An enzyme × grain interaction was detected for AID of starch at 21 and 42 d, and AID of fat at 21 d, with greater effects of enzymes in wheat-based compared with corn-based diets, but significant increments due to enzymes compared with controls in both diet types. Apparent ileal digestibility of fat at 42 d increased with enzyme supplementation compared with the control treatments. The XA and XAP treatments gradually (P < 0.05) increased AID of protein at 21 d, but only XAP increased AID of protein compared with the control at 42 d. Compared with the controls, XA increased AID energy by 52 or 87 kcal, and XAP by 104 or 152 kcal/kg of DM at 21 or 42 d, respectively. The caloric contribution of starch, fat, and protein were affected differentially by base grain and the presence of fibrous ingredients at 21 and 42 d of age. PMID:25071229

Salivary alpha-amylase during pregnancy: diurnal course and associations with obstetric history, maternal demographics, and mood.

PubMed

Giesbrecht, Gerald F; Granger, Douglas A; Campbell, Tavis; Kaplan, Bonnie

2013-03-01

Diurnal patterns of salivary alpha amylase (sAA) in pregnant women have not previously been described. The current study employed ecological momentary assessment to examine the association between the diurnal sAA, obstetric history, maternal demographics, and mood during pregnancy. Saliva was self-collected by 83 pregnant women (89% White, age 25.3-43.0 years; mean gestational age 21.9 weeks, range 6-37 weeks; gravida 1-6) at home over three days. Results indicated that current pregnancy (gestational age and fetal sex) and maternal demographics were not related to diurnal sAA. In contrast, a history of previous miscarriage (Parameter = -.17; SE = .05; p < .05) was associated with an atypical diurnal pattern. Even after accounting for obstetric history, trait anxiety (Parameter = .16; SE = .04; p < .001) was associated with increased sAA over the day while chronic levels of fatigue (Parameter = -.06; SE = .03; p < .05) were associated with decreased sAA. In a separate model, we also tested the time varying covariation of sAA and mood. The effects of momentary mood were in contrast to those for trait mood. Both momentary depression (Parameter = .22; SE = .09; p < .01) and vigour/positive mood (Parameter = .12; SE = .04; p < .001) were associated with momentary increases in sAA while momentary anxiety and fatigue were not related to sAA. The findings suggest that basal sAA during pregnancy is sensitive to emotional arousal. Evaluating diurnal patterns of sAA holds promise for advancing understanding of how emotional arousal during pregnancy may affect fetal development. Copyright © 2012 Wiley Periodicals, Inc.

Increased alpha-Amylase Response to an Acute Psychosocial Stress Challenge in Healthy Adults with Childhood Adversity

PubMed Central

Kuras, Yuliya I.; McInnis, Christine M.; Thoma, Myriam V.; Chen, Xuejie; Hanlin, Luke; Gianferante, Danielle; Rohleder, Nicolas

2017-01-01

Childhood adversity is highly prevalent and linked to lasting psychological and physiological consequences. A potential mechanism for negative health outcomes is altered stress reactivity. While previous research has addressed associations of childhood adversity with stress system reactivity, sympathetic nervous system (SNS) stress reactivity is understudied. We therefore set out here to examing salivary alpha-amylase (sAA) reactivity in relation with childhood adversity. Forty-one healthy adult subjects (n=24 male; n=17 female) aged 18-34 years underwent the Trier Social Stress Test (TSST) and completed the Childhood Trauma Questionnaire (CTQ). Saliva for measurement of sAA was collected at three time points; before the TSST, immediately after, and 10 minutes post-TSST. We found that those with childhood trauma had a higher overall sAA response to the TSST, as seen in a repeated measures ANOVA (CTQ by time interaction: F(1.8,71.5)=6.46, p=.01) and an independent samples t-test indicating higher sAA baseline to peak response (t=3.22, p=.003). There was also a positive correlation between sAA reactivity and the CTQ subscales of childhood physical abuse (r=.46, p=.005) and emotional abuse (r=.37, p=.024). Healthy adults with low-to-moderate childhood adversity had a heightened sAA response immediately following the stressor. Higher SNS reactivity could be a link to negative health outcomes in adults with early adversity. Future research should address whether altered sAA reactivity is predictive of negative health outcomes in those with childhood adversity. PMID:27577885

Alkalistable endo-β-1,4-xylanase production from a newly isolated alkalitolerant Penicillium sp. SS1 using agro-residues.

PubMed

Bajaj, Bijender Kumar; Sharma, Mukul; Sharma, Sunny

2011-09-01

Thermostable and alkalitolerant xylanases have got intense research focus due to their vast applications in various industries including pulp and paper, food, feed, textile, biofuel, etc. In the present investigation, a Penicillum sp. SS1 isolated from degrading woody material was found to produce moderately thermoactive and alkalistable endo-β-1,4-xylanase (xylanase). Maximum xylanase production was observed after fourth day of fermentation (43.84 IU/ml). The organism produced substantial quantities of xylanase using agricultural residues like wheat bran (20.6 IU/ml), rice bran (21.8 IU/ml) and sawdust (10.7 IU/ml) as carbon sources. The enzyme preparation was totally free of filter paper activity (FPase) and possessed negligible carboxymethyl cellulase (CMCase) activity; this could be an important feature of enzyme if the intended application of enzyme is in pulp and paper industries. Among nitrogen sources examined, yeast extract supported maximum xylanase production (45.74 IU/ml), and was followed by soybean meal (22.2 IU/ml) and ammonium sulphate (20 IU/ml). Maximum xylanase production was observed at initial medium pH 9 (25.6 IU/ml); however, at pH 8 and 10 also significantly high enzyme titre was observed (24 and 21.2 IU/ml, respectively). Thus, Penicillium sp. SS1 displayed capability of growing and producing xylanase at high alkaline pH (8-10). Maximum xylanase activity was reported at 50 °C, however, significantly high activity was observed at 60 °C (65.4%), however, at 70-80 °C activity was lost considerably. At 50-60 °C the enzyme retained very high activity up to 30-60 min (91-100%), however, prolonged incubation (90 min) caused considerable activity reduction (residual activity 63-68%).

Effects of exogenous xylanase on performance, nutrient digestibility and caecal thermal profiles of broilers given wheat-based diets.

PubMed

Cowieson, A J; Masey O'Neill, H V

2013-06-01

1. Five dietary treatments were used in a 49 d broiler trial to assess the effect of xylanase on performance, nutrient digestibility and thermal profiles of the caeca and head. Treatments included an industry-standard control diet and four further diets where xylanase was introduced with or without a metabolisable energy density dilution either from day one or the introduction was delayed until d 28. 2. The addition of xylanase with no associated energy dilution from day one resulted in the most consistent beneficial effects on performance, with significant improvements in weight gain compared with the industry-standard to d 28 and at d 49. Addition of xylanase from d 28 (with no energy dilution) was the second most successful strategy and resulted in a significant improvement in feed conversion ratio (FCR) from d 29 to 49 and overall. 3. Addition of xylanase improved ileal digestible energy values at d 28 by around 0.35 MJ/kg and ileal nitrogen digestibility coefficients by around 3%. On d 49 xylanase improved ileal digestible energy values by around 0.9 MJ/kg and ileal nitrogen digestibility coefficients by around 4.6%. 4. Thermal imaging of the head and caeca of three birds per replicate on d 49 revealed a significant increase in caecal surface temperature following xylanase addition with no effect on head temperature profile. These increases were particularly large (around 1.4ºC, or 3.9%) when xylanase was added from day one with no corresponding energy dilution in feed formulation. 5. It can be concluded that supplemental xylanase is effective in improving performance and nutrient digestibility in broilers given wheat-based diets. The correlation between the magnitude of this effect and the increased temperature in the caeca presents additional evidence that the hind-gut microflora may play an important, if yet unquantified, role in the outworking of these mechanisms.

Optimization of pectinase immobilization on grafted alginate-agar gel beads by 24 full factorial CCD and thermodynamic profiling for evaluating of operational covalent immobilization.

PubMed

Abdel Wahab, Walaa A; Karam, Eman A; Hassan, Mohamed E; Kansoh, Amany L; Esawy, Mona A; Awad, Ghada E A

2018-07-01

Pectinase produced by a honey derived from the fungus Aspergillus awamori KX943614 was covalently immobilized onto gel beads made of alginate and agar. Polyethyleneimine, glutaraldehyde, loading time and enzyme's units were optimized by 2 4 full factorial central composite design (CCD). The immobilization process increased the optimal working pH for the free pectinase from 5 to a broader range of pH4.5-5.5 and the optimum operational temperature from 55°C to a higher temperature, of 60°C, which is favored to reduce the enzyme's microbial contamination. The thermodynamics studies showed a thermal stability enhancement against high temperature for the immobilized formula. Moreover, an increase in half-lives and D-values was achieved. The thermodynamic studies proved that immobilization of pectinase made a remarkable increase in enthalpy and free energy because of enzyme stability enhancement. The reusability test revealed that 60% of pectinase's original activity was retained after 8 successive cycles. This gel formula may be convenient for immobilization of other industrial enzymes. Copyright © 2018 Elsevier B.V. All rights reserved.

Polypeptides having xylanase activity and polynucleotides encoding same

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

Spodsberg, Nikolaj

The present invention relates to isolated polypeptides having xylanase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

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

Lopez de Leon, Alfredo; Rey, Michael

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Lopez de Leon, Alfredo; Rey, Michael

2010-12-14

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Lopez de Leon, Alfredo; Rey, Michael

2016-05-31

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Tang, Lan; Liu, Ye; Duan, Junxin; Ding, Hanshu

2013-04-30

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Tang, Lan; Liu, Ye; Duan, Junxin; Hanshu, Ding

2012-10-30

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Lopez de Leon, Alfredo; Rey, Michael

2015-01-27

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Spodsberg, Nikolaj

2014-10-21

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Spodsberg, Nikolaj

2017-05-02

The present invention relates to isolated polypeptides having xylanase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Polypeptides having xylanase activity and polynucleotides encoding same

DOEpatents

Spodsberg, Nikolaj

2014-10-14

The present invention relates to isolated polypeptides having xylanase activity and polynucleotides encoding the polypeptides. The invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Sequential and simultaneous strategies for biorefining of wheat straw using room temperature ionic liquids, xylanases and cellulases.

PubMed

Husson, Eric; Auxenfans, Thomas; Herbaut, Mickael; Baralle, Manon; Lambertyn, Virginie; Rakotoarivonina, Harivoni; Rémond, Caroline; Sarazin, Catherine

2018-03-01

Sequential and simultaneous strategies for fractioning wheat straw were developed in combining 1-ethyl-3-methyl imidazolium acetate [C2mim][OAc], endo-xylanases from Thermobacillus xylanilyticus and commercial cellulases. After [C2mim][OAc]-pretreatment, hydrolysis catalyzed by endo-xylanases of wheat straw led to efficient xylose production with very competitive yield (97.6 ± 1.3%). Subsequent enzymatic saccharification allowed achieving a total degradation of cellulosic fraction (>99%). These high performances revealed an interesting complementarity of [C2mim][OAc]- and xylanase-pretreatments for increasing enzymatic digestibility of cellulosic fraction in agreement with the structural and morphological changes of wheat straw induced by each of these pretreatment steps. In addition a higher tolerance of endo-xylanases from T. xylaniliticus to [C2mim][AcO] until 30% v/v than cellulases from T. reesei was observed. Based on this property, a simultaneous strategy combining [C2mim][OAc]- and endo-xylanases as pretreatment in a one-batch produced xylose with similar yield than those obtained by the sequential strategy. Copyright © 2017 Elsevier Ltd. All rights reserved.

Halophilic Amylase from a Moderately Halophilic Micrococcus

PubMed Central

Onishi, Hiroshi

1972-01-01

A moderately halophilic Micrococcus sp., isolated from unrefined solar salt, produced a considerable amount of extracellular dextrinogenic amylase when cultivated aerobically in media containing 1 to 3 m NaCl. The Micrococcus amylase had maximal activity at pH 6 to 7 in 1.4 to 2 m NaCl or KCl at 50 C. Calcium ion and a high concentration of NaCl or KCl were essential for activity and stability of the amylase. The salt response of the amylase depended greatly on the pH and temperature of the enzyme assay. PMID:5058445

Production of xylanase from an alkali tolerant Streptomyces sp. 7b under solid-state fermentation, its purification, and characterization.

PubMed

Bajaj, Bijender Kumar; Singh, Narendera Pratap

2010-11-01

Streptomyces sp. 7b showed highest xylanase activity among 41 bacterial isolates screened under submerged fermentation. The organism grew over broad pH (5-11) and temperatures range (25-55 degrees C) and displayed maximum xylanase production on wheat bran (1230 U/g) under solid-state fermentation. Xylanase production was enhanced substantially (76%-77%) by inclusion of trypton (2180 U/g) or beef extract (2170 U/g) and moderately (36%-46%) by yeast extract (1800 U/g) or soybean meal (1670 U/g). Inclusion of readily utilizable sugars such as glucose, maltose, fructose, lactose or xylose in the substrate repressed the xylanase production. The optimum initial pH of the medium for maximum enzyme production was 7 to 8; however, appreciable level of activity was obtained at pH 6 (1,680 U/g) and 9 (1,900 U/g). Most appropriate solid to liquid ratio for maximum xylanase production in solid-state fermentation was found to be 1:2.5. The organism produced a single xylanase of molecular weight of approximately 30 kDa as analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis after purification with ammonium sulfate precipitation, and carboxy methyl sephadex chromatography. The enzyme was purified to the extent of 5.68-fold by salt precipitation and ion-exchange chromatography. Optimum temperature and pH for maximum xylanase activity were 50 degrees C and 6, respectively.

Biomimetic Mineralization of the Alginate/Gelatin/Calcium Oxalate Matrix for Immobilization of Pectinase: Influence of Matrix on the Pectinolytic Activity.

PubMed

Bustamante-Vargas, Cindy Elena; de Oliveira, Débora; Valduga, Eunice; Venquiaruto, Luciana Dornelles; Paroul, Natalia; Backes, Geciane Toniazzo; Dallago, Rogério Marcos

2016-07-01

Pectinases catalyze the degradation of pectic substances and are used in several processes, mainly in food and textile industries. In this study, a biomimetic matrix of alginate/gelatin/calcium oxalate (AGOCa) was synthesized for the in situ immobilization via encapsulation of crude pectinase from Aspergillus niger ATCC 9642, obtaining an immobilization efficiency of about 61.7 %. To determine the performance of AGOCa matrix, this was compared to control matrices of alginate/calcium oxalate (AOxal) and alginate/water (ACa). By the evaluation of pH and temperature effects on the enzyme activity, it was observed an increase on pectinolytic activity for both three tested matrices with an increase on pH and temperature. The kinetic parameters for pectinase immobilized in the three matrices were determined using citric pectin as substrate. Values of K m of 0.003, 0.0013, and 0.0022 g mL(-1) and V max of 3.85, 4.32, and 3.17 μmol min(-1) g(-1) for AGOCa, AOxal, and ACa matrices were obtained, respectively. After 33 days of storage, the pectinase immobilized in the three different matrices kept its initial activity, but that immobilized in AGOCa presented high stability to the storage with a relative activity of about 160 %. The enzyme immobilized in AGOCa, AOxal, and ACa could be used in 10, 8, and 7 cycles, respectively, keeping 40 % of its initial activity.

Production of xylanases by mangrove fungi from the Philippines and their application in enzymatic pretreatment of recycled paper pulps.

PubMed

Torres, Jeremy Martin O; Dela Cruz, Thomas Edison E

2013-04-01

Mangrove fungi are vastly unexplored for enzymes with industrial application. This study aimed to assess the biocatalytic activity of mangrove fungal xylanases on recycled paper pulp. Forty-four mangrove fungal (MF) isolates were initially screened for xylanolytic activity in minimal medium with corn cob xylan as the sole carbon source. Eight MF were further cultivated under submerged fermentation for the production of crude xylanases. These crude enzymes were then characterized and tested for the pretreatment of recycled paper pulps. Results showed that 93 % of the tested MF isolates exhibited xylanolytic activity in solid medium. In submerged fermentation, salinity improved the growth of the fungal isolates but did not influence xylanase production. The crude xylanases were mostly optimally active at 50 °C and pH 7. Changes in pH had a greater effect on xylanase stability than temperature. More than half of the activity was lost at pH 9 for majority of the crude enzymes. However, two thermophilic xylanases from Fusarium sp. KAWIT-A and Aureobasidium sp. 2LIPA-M and one alkaliphilic xylanase from Phomopsis sp. MACA-J were also produced. All crude enzymes exhibited cellulase activities ranging from 4 to 21 U/ml. Enzymatic pretreatment of recycled paper pulps with 5 % consistency produced 70-650 mg of reducing sugars per gram of pulp at 50 °C after 60 min. The release of high amounts of reducing sugars showed the potential of mangrove fungal crude xylanases in the local paper and pulp industry. The diverse properties shown by the tested crude enzymes also indicate its potential applications to other enzyme-requiring industries.

Refining the multisystem view of the stress response: coordination among cortisol, alpha-amylase, and subjective stress in response to relationship conflict.

PubMed

Laurent, Heidemarie K; Powers, Sally I; Granger, Douglas A

2013-07-02

This study investigated associations among young adults' hypothalamic-pituitary-adrenal axis activity, autonomic nervous system activity, and subjective stress in response to interpersonal conflict to better characterize coordination across stress systems. Seven saliva samples were collected from 199 young adult opposite-sex couples before, during, and after they discussed an unresolved relationship conflict. Samples were later assayed for cortisol and alpha-amylase (sAA). Couples rated anticipatory stress prior to the conflict and perceived stress immediately following the task. Growth curve modeling was used to examine two possible levels of within-person coordination across physiological systems: alignment between cortisol and sAA responses throughout the sampling period ("matched phase coordination"), and association between overall levels of cortisol and sAA in response to conflict ("average level coordination"). Whereas both partners showed the former type of coordination, only women showed the latter type. Positive anticipation of the stressor predicted stronger cortisol-sAA matched phase coordination for women. Pre-task ratings related to women's sAA, and post-task ratings related to both partners' cortisol responses. Implications for a multisystem interpretation of normal and pathological responses to daily stress are discussed. Copyright © 2013 Elsevier Inc. All rights reserved.

Refining the multisystem view of the stress response: Coordination among cortisol, alpha-amylase, and subjective stress in response to relationship conflict

PubMed Central

Powers, Sally I.; Granger, Douglas A.

2013-01-01

This study investigated associations among young adults' hypothalamic-pituitary-adrenal axis activity, autonomic nervous system activity, and subjective stress in response to interpersonal conflict to better characterize coordination across stress systems. Seven saliva samples were collected from 199 young adult opposite-sex couples before, during, and after they discussed an unresolved relationship conflict. Samples were later assayed for cortisol and alpha-amylase (sAA). Couples rated anticipatory stress prior to the conflict and perceived stress immediately following the task. Growth curve modeling was used to examine two possible levels of within-person coordination across physiological systems: alignment between cortisol and sAA responses throughout the sampling period (“matched phase coordination”), and association between overall levels of cortisol and sAA in response to conflict (“average level coordination”). Whereas both partners showed the former type of coordination, only women showed the latter type. Positive anticipation of the stressor predicted stronger cortisol-sAA matched phase coordination for women. Pre-task ratings related to women's sAA, and post-task ratings related to both partners' cortisol responses. Implications for a multisystem interpretation of normal and pathological responses to daily stress are discussed. PMID:23684904

A novel method for simultaneous purification and immobilization of a xylanase-lichenase chimera via SpyTag/SpyCatcher spontaneous reaction.

PubMed

Lin, Yuanqing; Jin, Wenhui; Wang, Jindan; Cai, Zhengwen; Wu, Shuyu; Zhang, Guangya

2018-08-01

We generated a bifunctional enzyme chimera containing the xylanase and lichenase coupled with SpyTag between them. Meanwhile, we generated another chimera containing SpyCatcher and elastin-like polypeptides (ELPs). As ELPs could bond to the xylanase-lichenase chimera through SpyTag/SpyCatcher spontaneous reaction in mild condition, which would lead to the formation of a 3-arm star multifunctional chimera. We purified the xylanase-lichenase by the non-chromatographic purification tag of ELPs. Interestingly, 57.5% of the xylanase and 47.2% of the lichenase in chimera self-assembled into insoluble active particles during the process of purification, which could serve as immobilized bifunctional enzymes. Notably, the immobilized chimera xylanase-lichenase showed a remarkable stability even after 10 reaction cycles, which retained around 56% (lichenase) and 44% (xylanase) of their initial activities, respectively. Moreover, the enhanced thermostability of the immobilized enzymes was also achieved. After incubating at 60 °C for 60 min, the residual activity of the immobilized lichenase was 35%, while the free one was only 24%. Unexpectedly, the free xylanase almost lost its activity when incubated at 55 °C for 60 min, whereas the immobilized xylanase retained 10% of its activity. However, the catalytic efficiency (k cat /K m ) of the free xylanase was 1.7-fold higher than the immobilized one, while the free lichenase was 1.1-fold higher than the immobilized one. This is among the first known reports that two enzymes are purified and immobilized in one-step. This novel strategy is easy to scale up and may meet the demands of biofuel industry. It would have great potentials in other biotechnological fields, such as the multifunctional biomaterials systems. Copyright © 2018 Elsevier Inc. All rights reserved.

Free cortisol and salivary alpha-amylase levels during a six-hour-water immersion in healthy young men

NASA Astrophysics Data System (ADS)

Rohleder, N.; Wirth, D.; Fraßl, W.; Kowoll, R.; Schlemmer, M.; Vogler, S.; Kirsch, K. A.; Kirschbaum, C.; Gunga, H.-C.

2005-08-01

Limited data are available on the response of stress systems to microgravity. Increased activity of stress systems is reported during space flight, but unchanged or decreased activity during simulated microgravity. We here investigated the impact of head-out water immersion on the activity of the hypothalamus-pituitary-adrenal (HPA) axis and the sympathetic-adrenal-medullary (SAM) system.Eight healthy young men were exposed to a six-hour water immersion in a thermo neutral bath and a control condition. Saliva samples were taken before, during, and after interventions to assess cortisol as an index for HPA axis activity, and salivary α-amylase as an index for SAM system activity.Cortisol levels uniformly decreased during both conditions. Amylase levels increased during both conditions, but were significantly lower during the first half of water immersion compared to the control condition.In conclusion, the HPA axis is not influenced by simulated microgravity, while SAM system activity shows initial decreases during water immersion.

Investigation into the cross-correlation of salivary cortisol and alpha-amylase responses to psychological stress.

PubMed

Engert, Veronika; Vogel, Susanne; Efanov, Simona I; Duchesne, Annie; Corbo, Vincent; Ali, Nida; Pruessner, Jens C

2011-10-01

Stress is a multidimensional construct. To accurately represent stress physiology, multiple stress measures across multiple stress-related systems should be assessed. However, associations may be masked given that different systems underlie different time courses. Salivary cortisol and alpha-amylase (sAA) are reliable biological stress markers of the sympathetic nervous system (SNS) and the hypothalamus pituitary adrenal (HPA) axis, respectively. Studies examining the link between sAA and cortisol levels in response to stress have produced inconsistent results. Here, we investigated whether the covariance of stress-induced sAA and cortisol release is dependent on the distinct temporal dynamics of the two stress markers. A total of 50 male participants were exposed to a psychological laboratory stressor with high frequency (2-min interval) saliva sampling in two independent studies. Synchronized time series of sAA and cortisol measures before, during and after stress induction were obtained. Cross-correlation analysis was applied to test for the association of sAA and cortisol levels at various stages relative to the onset of the stressor. Positive and negative cross-correlations between lagged pairs of sAA and cortisol measures were found in both studies. The strongest correlation was found for sAA preceding cortisol release by 13.5 min (r = .27, p < .001). With a smaller effect size cortisol also significantly preceded sAA by 13.5 min (r = -.16, p < .001). We suggest that sAA and cortisol stress responses are reliably associated at various time lags throughout a stressful situation. As a possible connection site between HPA axis and SNS that may underlie sAA-cortisol associations, we discuss CRF neurons of the hypothalamus involved in sympathetic regulation. Copyright © 2011 Elsevier Ltd. All rights reserved.

Carbodiimide for Covalent α-Amylase Immobilization onto Magnetic Nanoparticles

NASA Astrophysics Data System (ADS)

Milani, Zeinab Mortazavi; Jalal, Razieh; Goharshadi, Elaheh K.

Covalent cross-linking of enzymes to magnetite (Fe3O4) nanoparticles (MNPs) is one of the useful enzyme immobilization methods which provides repeated use of the catalyst, facilitates enzyme separation from the reaction mixture, and sometimes improves biocatalysts stability. The aim of this study was to immobilize α-amylase onto MNPs via covalent attachment using carbodiimide (CDI) molecules. MNPs were synthesized by the co-precipitation method. The size and the structure of the particles were characterized by X-ray diffraction and transmission electron microscopy. The effects of different operational conditions of direct α-amylase binding on MNPs in the presence of CDI were investigated by using the shaking method. Fourier transform infrared spectroscopy was used to confirm the success of immobilization. The optimum conditions and catalytic properties of immobilized α-amylase were also evaluated. The efficiency of immobilization and the residual activity of the immobilized α-amylase were dependent on the mass ratio of MNPs: CDI: α-amylase and the immobilization temperature. The optimum pH for the free and immobilized amylase was 6. The free and immobilized α-amylase showed maximum activity at 20∘C and 35∘C, respectively. The immobilized α-amylase was more thermostable than the free one. The retained activity for free α-amylase after 19 storage days was 57.7% whereas it was 100% for the immobilized α-amylase. In repeated batch experiments, the immobilized α-amylase retained a residual activity of 45% after 11 repeated uses. The Km and Vmax values for the immobilized enzyme were larger than those of the free enzyme. The immobilization of α-amylase on MNPs using CDI improves its stability and reusability.

Influence of media composition on the production of alkaline α-amylase from Bacillus subtilis CB-18.

PubMed

Ogbonnaya, Nwokoro; Odiase, Anthonia

2012-01-01

Starch, a homopolysaccharide is an important and an abundant food reserve and energy source. Starches are processed to yield different products which find many industrial applications. Alpha-amylases hydrolyze starch by cleaving α-1,4-glucosidic bonds and have been used in food, textile and pharmaceutical industries [Sun et al. 2010]. Enzymatic conversion of starch with amylase presents an economically superior alternative to the conventional method of starch gelatinization. Alkaline α-amylase has an important position in the global enzyme market as a constituent of detergent. In this paper, we screened soil bacteria and an isolate, alkalophilic Bacillus subtilis CB-18 was found to produce an alkaline α-amylase in different media. MATERIAL AND METHODS. Screening of the isolates for amylolytic activity was carried out by growing bacteria isolated from the soil in starch agar plates and subsequently staining the plates with iodine solution to reveal zones of hydrolysis of starch. The selected isolate, Bacillus subtlis CB-18 was grown in different media at alkaline pH to evaluate the influence of media composition on alkaline α-amylase production. Enzyme assay was carried out by growing the culture in a broth medium and obtaining cell - free culture supernatant after centrifugation at 2515 × g for 15 minutes Amylase activity was determined by incubating 0.5 ml of crude enzyme solution in 0.1M Tris/HCl buffer (pH 8.5) with 0.5 ml of 1% soluble starch solution. The reaction was terminated by the addition of DNS reagent and reducing sugar produced from the amylolytic reaction was determined. Bacillus subtilis CB-18 used for this work was selected because it produced 7 mm zone diameter on starch agar plate. This organism was cultured in different alkaline broth media containing 2% soluble starch as inducer carbohydrate for α-amylase production. Among the carbon sources used for enzyme production, sorbitol was the best to stimulate enzyme production with α-amylase

Acarviosine-simmondsin, a novel compound obtained from acarviosine-glucose and simmondsin by Thermus maltogenic amylase and its in vivo effect on food intake and hyperglycemia.

PubMed

Baek, Jin-Sook; Kim, Hye-Young; Abbott, Thomas P; Moon, Tae-Wha; Lee, Soo-Bok; Park, Cheon-Seok; Park, Kwan-Hwa

2003-03-01

Simmondsin was modified with acarviosine-glucose using the transglycosylation activity of Thermus maltogenic amylase to synthesize a novel compound with both antiobesity and hypoglycemic activity. The LC/MS and 13C NMR analyses confirmed that the structure of the major transglycosylation product was acarviosine-simmondsin (Acv-simmondsin), in which acarviosine was attached to the glucose moiety of simmondsin by an alpha-(1,6)-glycosidic linkage. It was found that Acv-simmondsin was a potent competitive inhibitor of alpha-glucosidase with the Ki value of 0.69 microM and a mixed type inhibitor of alpha-amylase with the Ki and KI of 20.78 microM and 26.31 microM, respectively. The administration of Acv-simmondsin (0.1 g/100 g diet/day) to mice for 5 days significantly reduced food intake by 35%, compared to 25% with simmondsin in control obese mice. Acv-simmondsin (50 mg/kg BW) suppressed the postprandial blood glucose response to sucrose (1 g/kg BW) by 74%, compared to 71% with acarbose, in normal rats.

Evaluation of salivary catalase, vitamin C, and alpha-amylase in smokers and non-smokers: a retrospective cohort study.

PubMed

Ahmadi-Motamayel, Fatemeh; Falsafi, Parisa; Goodarzi, Mohammad Taghi; Poorolajal, Jalal

2017-05-01

Saliva and its defence systems such as antioxidants and minerals are very important in the pathogenesis of different diseases. Cigarette smoking has many destructive effects. Oxidative stresses play an important role in the side effects of smoking. This study assessed the effect of cigarette smoking on salivary levels of catalase, vitamin C, and α-amylase. This retrospective cohort study was carried out in Hamadan, Iran, on 510 subjects; 259 subjects were smokers (the exposed group) and 251 were non-smokers (the unexposed group). Five microliters of unstimulated saliva was collected by spitting method. Catalase, vitamin C, and α-amylase salivary levels were determined by spectrophotometric assay. Data were analyzed with t-test using STATA 12. Vitamin C level in smokers was significantly lower than that in non-smokers. The salivary catalase levels were lower and α-amylase levels were higher in smokers, but the differences were not statistically significant (P = 0.416 and P = 0.265, respectively). Smokers were younger than non-smokers. Smoking resulted in a change in salivary antioxidant levels. Changes in antioxidant levels can influence the deleterious effects of smoking on oral mucosa; it might also indicate systemic changes and changes in the serum levels of oxidative agents. Further studies are necessary to understand the mechanisms and real effects of smoking, to determine the benefits of supplementary antioxidants for treatment and to reduce the dangerous side effects of smoking. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Serum Amylase in Bulimia Nervosa and Purging Disorder: Differentiating the Association with Binge Eating versus Purging Behavior

PubMed Central

Wolfe, Barbara E.; Jimerson, David C.; Smith, Adrian; Keel, Pamela K.

2011-01-01

Objective Elevated serum amylase levels in bulimia nervosa (BN), associated with increased salivary gland size and self-induced vomiting in some patients, provide a possible marker of symptom severity. The goal of this study was to assess whether serum hyperamylasemia in BN is more closely associated with binge eating episodes involving consumption of large amounts of food or with purging behavior. Method Participants included women with BN (n=26); women with “purging disorder” (PD), a subtype of EDNOS characterized by recurrent purging in the absence of objectively large binge eating episodes (n=14); and healthy non-eating disorder female controls (n=32). There were no significant differences in age or body mass index (BMI) across groups. The clinical groups reported similar frequency of self-induced vomiting behavior and were free of psychotropic medications. Serum samples were obtained after overnight fast and were assayed for alpha-amylase by enzymatic method. Results Serum amylase levels were significantly elevated in BN (60.7 ± 25.4 international units [IU]/liter, mean ± sd) in comparison to PD (44.7 ± 17.1 IU/L, p < 02) and to Controls (49.3 ± 15.8, p < .05). Conclusion These findings provide evidence to suggest that it is recurrent binge eating involving large amounts of food, rather than self-induced vomiting, which contributes to elevated serum amylase values in BN. PMID:21781981

Salivary alpha-amylase and cortisol responsiveness following electrical stimulation stress in patients with the generalized type of social anxiety disorder.

PubMed

Tamura, A; Maruyama, Y; Ishitobi, Y; Kawano, A; Ando, T; Ikeda, R; Inoue, A; Imanaga, J; Okamoto, S; Kanehisa, M; Ninomiya, T; Tanaka, Y; Tsuru, J; Akiyoshi, J

2013-11-01

Social anxiety disorder is believed to be a stress-induced disease. Although it can be inferred from the symptoms during attacks that there exists some abnormality of autonomic nervous system in any of the stress systems in social anxiety disorder, little evidence has been reported. This study focused on comparing the reactivity of 2 stress systems, the autonomic nervous system (ANS) and the hypothalamic-pituitary-adrenal (HPA) axis in patients with social anxiety disorder. 32 patients with the generalized type of social anxiety disorder were compared with 80 age- and gender-matched controls. We collected saliva samples from patients and controls before and after electrical stimulation to measure the concentrations of salivary alpha-amylase (sAA) and salivary cortisol. Profile of Mood State (POMS) and State-Trait Anxiety Inventory (STAI) scores and Heart Rate Variability (HRV) were also determined following stimulation. SAA in patients displayed a significantly higher level at baseline and a significantly larger response to electrical stimulation as compared to controls, whereas no group differences were seen in any HRV. Neither within-subject nor group differences were seen in salivary cortisol levels. These results suggest that SAD patients displayed enhanced ANS (but not HPA axis) activity vs. healthy controls. © Georg Thieme Verlag KG Stuttgart · New York.

Combination of Xylanase and Debranching Enzymes Specific to Wheat Arabinoxylan Improve the Growth Performance and Gut Health of Broilers.

PubMed

Lei, Zhao; Shao, Yuxin; Yin, Xiaonan; Yin, Dafei; Guo, Yuming; Yuan, Jianmin

2016-06-22

Arabinoxylan (AX) is the major antinutritional factor of wheat. This study evaluated the synergistic effects of xylanase and debranching enzymes (arabinofuranosidase [ABF] and feruloyl esterase [FAE]) on AX. During in vitro tests, the addition of ABF or FAE accelerated the hydrolysis of water-soluble AX (WE-AX) and water-insoluble AX (WU-AX) and produced more xylan oligosaccharides (XOS) than xylanase alone. XOS obtained from WE-AX stimulated greater proliferation of Lactobacillus brevis and Bacillus subtilis than did fructo-oligosaccharides (FOS) and glucose. During in vivo trials, xylanase increased the average daily growth (ADG), decreased the feed-conversion ratio (FCR), and reduced the digesta viscosity of jejunum and intestinal lesions of broilers fed a wheat-based diet on day 36. ABF or FAE additions further improved these effects. Broilers fed a combination of xylanase, ABF, and FAE exhibited the best growth. In conclusion, the synergistic effects among xylanase, ABF, and FAE increased AX degradation, which improve the growth performance and gut health of broilers.

Increased alpha-amylase response to an acute psychosocial stress challenge in healthy adults with childhood adversity.

PubMed

Kuras, Yuliya I; McInnis, Christine M; Thoma, Myriam V; Chen, Xuejie; Hanlin, Luke; Gianferante, Danielle; Rohleder, Nicolas

2017-01-01

Childhood adversity is highly prevalent and linked to lasting psychological and physiological consequences. A potential mechanism for negative health outcomes is altered stress reactivity. While previous research has addressed associations of childhood adversity with stress system reactivity, sympathetic nervous system (SNS) stress reactivity is understudied. We therefore set out here to examining salivary alpha-amylase (sAA) reactivity in relation with childhood adversity. Forty-one healthy adult subjects (n = 24 male; n = 17 female) aged 18-34 years underwent the Trier Social Stress Test (TSST) and completed the Childhood Trauma Questionnaire (CTQ). Saliva for measurement of sAA was collected at three time points; before the TSST, immediately after, and 10 min post-TSST. We found that those with childhood trauma had a higher overall sAA response to the TSST, as seen in a repeated measures ANOVA (CTQ by time interaction: F(1.8,71.5) = 6.46, p = .01) and an independent samples t-test indicating higher sAA baseline to peak response (t = 3.22, p = .003). There was also a positive correlation between sAA reactivity and the CTQ subscales of childhood physical abuse (r = .46, p = .005) and emotional abuse (r = .37, p = .024). Healthy adults with low-to-moderate childhood adversity had a heightened sAA response immediately following the stressor. Higher SNS reactivity could be a link to negative health outcomes in adults with early adversity. Future research should address whether altered sAA reactivity is predictive of negative health outcomes in those with childhood adversity. © 2016 Wiley Periodicals, Inc.

Potent α-glucosidase and α-amylase inhibitory activities of standardized 50% ethanolic extracts and sinensetin from Orthosiphon stamineus Benth as anti-diabetic mechanism.

PubMed

Mohamed, Elsnoussi Ali Hussin; Siddiqui, Mohammad Jamshed Ahmad; Ang, Lee Fung; Sadikun, Amirin; Chan, Sue Hay; Tan, Soo Choon; Asmawi, Mohd Zaini; Yam, Mun Fei

2012-10-08

In the present study, we tested a 50% ethanolic extract of Orthosiphon stamineus plants and its isolated bioactive compound with respect to their α-glucosidase and α-amylase inhibitory activities. Bioactive flavonoid sinensetin was isolated from 50% ethanolic extract of Orthosiphon stamineus. The structure of this pure compound was determined on the NMR data and the α-glucosidase and α-amylase inhibitory activities of isolated sinensetin and 50% ethanolic extract of Orthosiphon stamineus were evaluated. In vitro studies of a 50% ethanolic extract of O. stamineus and the isolated sinensetin compound showed inhibitory activity on α-glucosidase (IC50: 4.63 and 0.66 mg/ml, respectively) and α-amylase (IC50: 36.70 mg/ml and 1.13 mg/ml, respectively). Inhibition of these enzymes provides a strong biochemical basis for the management of type 2 diabetes via the control of glucose absorption. Alpha-glucosidase and α-amylase inhibition could the mechanisms through which the 50% ethanolic extract of O. stamineus and sinensetin exert their antidiabetic activity, indicating that it could have potential use in the management of non-insulin-dependent diabetes.

Interactions between xylanase and glucanase in maize-soy-based diets for broilers.

PubMed

Cowieson, A J; Bedford, M R; Ravindran, V

2010-04-01

1. The effect of various doses of xylanase and glucanase on the performance and ileal nutrient digestibility of broiler chickens fed on maize/soy-based diets was evaluated. 2. A total of 960 male broilers were used in separate growth and digestibility trials with each involving 10 treatments and 6 replicates. The 10 treatments included a positive control reference diet, a negative control diet with a lower energy density and 8 further diets where xylanase and glucanase were added to the negative control, individually and in combination. 3. Birds which received the negative control diet returned poorer (6 points; P < 0.05) feed conversion ratios compared with those fed on the positive control, confirming the lower energy density of the negative control diet. Ileal digestibility of energy determined at 21 and 42 d was also significantly lower for the negative control compared with the positive control. At d 21 birds that received the negative control diet returned lower ileal amino acid digestibility (for most amino acids) compared with their counterparts fed on the positive control whereas at d 42 this effect was not apparent. 4. Supplementation of the negative control with both glucanase and xylanase improved feed conversion ratio (FCR) and ileal nutrient digestibility. When both enzymes were added simultaneously a sub-additive effect (i.e. simultaneous use resulted in benefits greater than either enzyme independently but less than the sum of the individual effects) was observed, with the best performance achieved with the combination of xylanase (16 000 BXU/kg) and glucanase (30 000 BU/kg). 5. It can be concluded that the removal of oil to accommodate the anticipated energy digestibility improvement with enzymes can have deleterious effects on FCR and ileal amino acid digestibility in young broilers which enzymes may not adequately mitigate. Thus, in order to maximise the response to non-starch polysaccharide degrading enzymes in maize/soy-based broiler diets, it

Production of xylanase and β-xylosidase from autohydrolysis liquor of corncob using two fungal strains.

PubMed

Michelin, Michele; Polizeli, Maria de Lourdes T M; Ruzene, Denise S; Silva, Daniel P; Ruiz, Héctor A; Vicente, António A; Jorge, João A; Terenzi, Héctor F; Teixeira, José A

2012-09-01

Agroindustrial residues are materials often rich in cellulose and hemicellulose. The use of these substrates for the microbial production of enzymes of industrial interest is mainly due to their high availability associated with their low cost. In this work, corncob (CCs) particles decomposed to soluble compounds (liquor) were incorporated in the microbial growth medium through autohydrolysis, as a strategy to increase and undervalue xylanase and β-xylosidase production by Aspergillus terricola and Aspergillus ochraceus. The CCs autohydrolysis liquor produced at 200 °C for 5, 15, 30 or 50 min was used as the sole carbon source or associated with untreated CC. The best condition for enzyme synthesis was observed with CCs submitted to 30 min of autohydrolysis. The enzymatic production with untreated CCs plus CC liquor was higher than with birchwood xylan for both microorganisms. A. terricola produced 750 total U of xylanase (144 h cultivation) and 30 total U of β-xylosidase (96-168 h) with 0.75% untreated CCs and 6% CCs liquor, against 650 total U of xylanase and 2 total U of β-xylosidase in xylan; A. ochraceus produced 605 total U of xylanase and 56 total U of β-xylosidase (168 h cultivation) with 1% untreated CCs and 10% CCs liquor against 400 total U of xylanase and 38 total U of β-xylosidase in xylan. These results indicate that the treatment of agroindustrial wastes through autohydrolysis can be a viable strategy in the production of high levels of xylanolytic enzymes.

Genetic and Biochemical Characterization of Monokaryotic Progeny Strains of Button Mushroom (Agaricus bisporus)

PubMed Central

Kwon, Hyuk Woo; Choi, Min Ah; Yun, Yeo Hong; Oh, Youn-Lee; Kong, Won-Sik

2015-01-01

To promote the selection of promising monokaryotic strains of button mushroom (Agaricus bisporus) during breeding, 61 progeny strains derived from basidiospores of two different lines of dikaryotic parental strains, ASI1038 and ASI1346, were analyzed by nucleotide sequencing of the intergenic spacer I (IGS I) region in their rDNA and by extracellular enzyme assays. Nineteen different sizes of IGS I, which ranged from 1,301 to 1,348 bp, were present among twenty ASI1346-derived progeny strains, while 15 different sizes of IGS I, which ranged from 700 to 1,347 bp, were present among twenty ASI1038-derived progeny strains. Phylogenetic analysis of the IGS sequences revealed that different clades were present in both the ASI10388- and ASI1346-derived progeny strains. Plating assays of seven kinds of extracellular enzymes (β-glucosidase, avicelase, CM-cellulase, amylase, pectinase, xylanase, and protease) also revealed apparent variation in the ability to produce extracellular enzymes among the 40 tested progeny strains from both parental A. bisporus strains. Overall, this study demonstrates that characterization of IGS I regions and extracellular enzymes is useful for the assessment of the substrate-degrading ability and heterogenicity of A. bisporus monokaryotic strains. PMID:25892920

Cortisol and salivary alpha-amylase trajectories following a group social-evaluative stressor with adolescents.

PubMed

Katz, Deirdre A; Peckins, Melissa K

2017-12-01

Intraindividual variability in stress responsivity and the interrelationship of multiple neuroendocrine systems make a multisystem analytic approach to examining the human stress response challenging. The present study makes use of an efficient social-evaluative stress paradigm - the Group Public Speaking Task for Adolescents (GPST-A) - to examine the hypothalamic-pituitary-adrenocortical (HPA)-axis and Autonomic Nervous System (ANS) reactivity profiles of 54 adolescents with salivary cortisol and salivary alpha-amylase (sAA). First, we account for individuals' time latency of hormone concentrations between individuals. Second, we use a two-piece multilevel growth curve model with landmark registration to examine the reactivity and recovery periods of the stress response separately. This analytic approach increases the models' sensitivity to detecting trajectory differences in the reactivity and recovery phases of the stress response and allows for interindividual variation in the timing of participants' peak response following a social-evaluative stressor. The GPST-A evoked typical cortisol and sAA responses in both males and females. Males' cortisol concentrations were significantly higher than females' during each phase of the response. We found no gender difference in the sAA response. However, the rate of increase in sAA as well as overall sAA secretion across the study were associated with steeper rates of cortisol reactivity and recovery. This study demonstrates a way to model the response trajectories of salivary biomarkers of the HPA-axis and ANS when taking a multisystem approach to neuroendocrine research that enables researchers to make conclusions about the reactivity and recovery phases of the HPA-axis and ANS responses. As the study of the human stress response progresses toward a multisystem analytic approach, it is critical that individual variability in peak latency be taken into consideration and that accurate modeling techniques capture

Development of a bifunctional xylanase-cellulase chimera with enhanced activity on rice and barley straws using a modular xylanase and an endoglucanase procured from camel rumen metagenome.

PubMed

Khalili Ghadikolaei, Kamran; Akbari Noghabi, Kambiz; Shahbani Zahiri, Hossein

2017-09-01

The camel rumen metagenome is an untapped source of glycoside hydrolases. In this study, novel genes encoding for a modular xylanase (XylC) and a cellulase (CelC) were isolated from a camel rumen metagenome and expressed in Escherichia coli BL21 (DE3). XylC with xylanase (Xyn), CBM, and carbohydrate esterase (CE) domains was characterized as a β-1,4-endoxylanase with remarkable catalytic activity on oat-spelt xylan (K cat  = 2919 ± 57 s -1 ). The implication of XylC's modular structure in its high catalytic activity was analyzed by truncation and fusion construction with CelC. The resulting fusions including Cel-CBM, Cel-CBM-CE, and Xyn-CBM-Cel showed remarkable enhancement in CMCase activity with K cat values of 742 ± 12, 1289 ± 34.5, and 2799 ± 51 s -1 compared to CelC with a K cat of 422 ± 3.5 s -1 . It was also shown that the bifunctional Xyn-CBM-Cel with synergistic xylanase/cellulase activities was more efficient than XylC and CelC in hydrolysis of rice and barley straws.

Growth performance, nutrient utilization, and digesta characteristics in broiler chickens fed corn or wheat diets without or with supplemental xylanase.

PubMed

Kiarie, E; Romero, L F; Ravindran, V

2014-05-01

Efficacy of supplemental xylanase on growth performance, nutrient utilization, and digesta characteristics in broiler chickens fed corn- or wheat-based diets was investigated. In experiment 1, 192 male broilers (8 birds/pen; n = 6) were fed 4 diets (corn or wheat without or with 1,250 xylanase units/kg) in 2 phases (starter, d 0-21 and grower, d 22-42). There was no interaction (P > 0.05) between diet and xylanase on performance (d 0-42). Wheat diets resulted (P < 0.01) in better performance than corn diets, whereas xylanase-fed birds had improved (P < 0.01) BW gain (2,457 vs. 2,275 g) and feed per gain (1.677 vs. 1.762) relative to birds not fed xylanase. In experiment 2, TiO2 (0.3%) was added in starter diets used in experiment 1, allocated to 13-d-old broiler chicks (n = 6) housed in cages (6 birds/cage) and fed up to d 21. Excreta samples were obtained from d 17 to 20 and birds were euthanized on d 21 for digesta. Corn diets had a greater concentration (10.7 vs. 9.8%) of insoluble nonstarch polysaccharides (NSP) than wheat diets, which in turn had more than twice the concentration of soluble NSP. There was an interaction (P < 0.03) between diet type and xylanase on jejunal digesta viscosity but not (P > 0.10) on apparent ileal digestibilities of nutrients, cecal volatile fatty acids, and AMEn. In this context, diet type influenced (P < 0.05) cecal volatile fatty acids and retention of nutrients and fiber but did not affect (P = 0.45) AMEn. In contrast, xylanase-fed birds showed higher (P < 0.05) ceca digesta acetic acid, apparent ileal digestibilities of nutrients, and retention of components. As a result, birds fed xylanase had higher AMEn (3,059 vs. 2,995 kcal/kg; P < 0.01) compared with birds not fed xylanase. Although wheat diets had superior growth performance, the AMEn was similar in both diets. Xylanase improved growth performance and AMEn independent of diet type, suggesting hydrolysis of both soluble and insoluble NSP.

Decreased salivary alpha-amylase levels are associated with performance deficits during sleep loss.

PubMed

Pajcin, Maja; Banks, Siobhan; White, Jason M; Dorrian, Jill; Paech, Gemma M; Grant, Crystal; Johnson, Kayla; Tooley, Katie; Fidock, Justin; Kamimori, Gary H; Della Vedova, Chris B

2017-04-01

During sleep deprivation, neurobehavioral functions requiring sustained levels of attention and alertness are significantly impaired. Discrepancies between subjective measures of sleepiness and objective performance during sustained operations have led to interest in physiological monitoring of operator performance. Alertness, vigilance, and arousal are modulated by the wake-promoting actions of the central noradrenergic system. Salivary alpha-amylase (sAA) has been proposed as a sensitive peripheral measure of noradrenergic activity, but limited research has investigated the relationship between sAA and performance. In a laboratory-controlled environment, we investigated the relationship between sAA levels, subjective sleepiness, and performance during two days (50h) of total sleep deprivation. Beginning at 09:00, twelve healthy participants (5 females) aged 22.5±2.5years (mean±SD) provided saliva samples, recorded ratings of subjective sleepiness, completed a brief 3-min psychomotor vigilance task (PVT-B) and performed a 40-min simulated driving task, at regular 3h intervals during wakefulness. Ratings of subjective sleepiness exhibited a constant linear increase (p<0.001) during sleep deprivation. In contrast, sAA levels showed a marked diurnal profile, with levels increasing during the day (p<0.001) and steadily declining in the evening and early-morning (p<0.001). PVT-B (mean reaction time and mean slowest 10% reaction time) and simulated driving performance (speed deviation and lane deviation) also exhibited diurnal profiles across the two days of sleep deprivation. Performance peaked in the afternoon (p<0.001) and then steadily worsened as wakefulness continued into the evening and early-morning (p<0.001). Further analysis revealed that higher sAA levels in the hour preceding each performance assessment were associated with better PVT-B and driving performance (p<0.001). These findings suggest that sAA measures may be suitable indicators of performance

[Amylase production by Aureobasidium pullulans in liquid and solid media].

PubMed

Lodato, P B; Forchiassin, F; Segovia de Huergo, M B

1997-01-01

Amylase production by a strain of Aureobasidium pullulans isolated in the laboratory was evaluated in liquid media (complex and synthetic) and in solid medium (wheat bran). There was an inhibitory effect in amylase production or amylase secretion by glucose. Asparagine was the best nitrogen source for amylase production (4-6 g/l). Only chlamidospores and melanin but not, amylase activity, were obtained with ammonium sulfate. Amylase production in solid culture was higher than the production obtained in the liquid media assayed. Optimum initial moisture content in solid culture ranged between 57 and 74%. No difference was observed in amylase production between solid media inoculated with cells grown in liquid or solid media.

Production of beta-xylanase by a Thermomyces lanuginosus MC 134 mutant on corn cobs and its application in biobleaching of bagasse pulp.

PubMed

Kumar, Kuttanpillai Santhosh; Manimaran, Ayyachamy; Permaul, Kugen; Singh, Suren

2009-05-01

The production of hemicellulases by Thermomyces lanuginosus SK using oatspelts xylan was examined during submerged cultivation. A high level of extracellular xylanase (346+/-10 U ml(-1)) production was observed on the fifth day; however, accessory enzyme levels were low. T. lanuginosus SK was further subjected to UV and N-methyl-N-nitro-N-nitrosoguanidine mutagenesis. The T. lanuginosus MC 134 mutant showed a 1.5 fold increase in xylanase production on oatspelts xylan, compared to the wild type strain. Xylanase production was further enhanced to 3299+/-95 U ml(-1) by using corn cobs under optimized growth conditions. A reduction in xylanase production was observed in a 5 L fermenter. Also, the biobleaching efficiency of crude xylanase was evaluated on bagasse pulp, and a brightness of 46.07+/-0.05% was observed with the use of 50 U of crude xylanase per gram of pulp. This brightness was 3.6 points higher than that of the untreated samples. Reducing sugars (25.78+/-0.14 mg g(-1)) and UV-absorbing lignin-derived compound values were considerably higher in xylanase-treated samples. T. lanuginosus MC 134 has a potential application in the pulp and paper industries.

Production of a cellulase-free alkaline xylanase from Bacillus pumilus MTCC 5015 by submerged fermentation and its application in biobleaching.

PubMed

Thomas, Leya; Sindhu, Raveendran; Binod, Parameswaran; Pandey, Ashok

2015-06-01

Here, we described the production of a cellulase-free alkaline xylanase from Bacillus pumilus MTCC 5015 by submerged fermentation and its application in biobleaching. Various process parameters affecting xylanase production by B. pumilus were optimized by adopting a Plackett-Burman design (PBD) as well as Response surface methodology (RSM). These statistical methods aid in improving the enzyme yield by analysing the individual crucial components of the medium. Maximum production was obtained with 4% yeast extract, 0.08% magnesium sulphate, 30 h of inoculum age, incubation temperature of 33.5 degrees C and pH 9.0. Under optimized conditions, the xylanase activity was 372 IU/ml. Media engineering improved a 5-fold increase in the enzyme production. Scanning electron microscopy (SEM) showed significant changes on the surface of xylanase treated pulps as a result of xylan hydrolysis. Increased roughness of paper carton fibres was apparent in scanning electron micrograph due to opening of the micro fibrils present on the surface by xylanase action. The untreated pulp did not show any such change. These results demonstrated that the B. pumilus MTCC 5015 xylanase was effective in bio-bleaching of paper carton.

Cloning, Expression and Characteristics of a Novel Alkalistable and Thermostable Xylanase Encoding Gene (Mxyl) Retrieved from Compost-Soil Metagenome

PubMed Central

Verma, Digvijay; Kawarabayasi, Yutaka; Miyazaki, Kentaro; Satyanarayana, Tulasi

2013-01-01

Background The alkalistable and thermostable xylanases are in high demand for pulp bleaching in paper industry and generating xylooligosaccharides by hydrolyzing xylan component of agro-residues. The compost-soil samples, one of the hot environments, are expected to be a rich source of microbes with thermostable enzymes. Methodology/Principal Findings Metagenomic DNA from hot environmental samples could be a rich source of novel biocatalysts. While screening metagenomic library constructed from DNA extracted from the compost-soil in the p18GFP vector, a clone (TSDV-MX1) was detected that exhibited clear zone of xylan hydrolysis on RBB xylan plate. The sequencing of 6.321 kb DNA insert and its BLAST analysis detected the presence of xylanase gene that comprised 1077 bp. The deduced protein sequence (358 amino acids) displayed homology with glycosyl hydrolase (GH) family 11 xylanases. The gene was subcloned into pET28a vector and expressed in E. coli BL21 (DE3). The recombinant xylanase (rMxyl) exhibited activity over a broad range of pH and temperature with optima at pH 9.0 and 80°C. The recombinant xylanase is highly thermostable having T1/2 of 2 h at 80°C and 15 min at 90°C. Conclusion/Significance This is the first report on the retrieval of xylanase gene through metagenomic approach that encodes an enzyme with alkalistability and thermostability. The recombinant xylanase has a potential application in paper and pulp industry in pulp bleaching and generating xylooligosaccharides from the abundantly available agro-residues. PMID:23382818

Screening of Thermophilic Bacteria Produce Xylanase from Sapan Sungai Aro Hot Spring South Solok

NASA Astrophysics Data System (ADS)

Irdawati, I.; Syamsuardi, S.; Agustien, A.; Rilda, Y.

2018-04-01

xylanase is one of the enzymes with great prospects as hemicellulose hydrolyzing enzyme. Global annual market demand for this enzyme reach US 200 million. This enzyme catalyzes the xylan (hemicellulose) reactions breaking into xilooligosakarida and xylose. Xylanase can be applied to various industrial sectors such as bread, sugar xylose, biofuels, especially in bleaching paper (bleaching) pulp. Xylanase Isable to replace conventional chemical bleaching using chlorine that is not friendly for the environment. Currently xylanase production is extracted from the thermophilic bacteria for enzyme stability at high temperatures that are suitable for industrial applications. Thermophilic bacteria can be isolated from a hot spring, one of the which is a source of Sapan Sungai Aro Hot Spring, located in the district South Solok. The aim of this study was to select and identification of thermophilic bacteria can produce xylanase.This roomates is a descriptive study, which was Carried out in the Laboratory of Microbiology, Mathematic and Science Faculty of Padang State University, and Laboratory of Bacteriology, BasoVeterinary Research Center. The research procedure consisted of the preparation and sterilization of materials and tools, medium manufacturing, regeneration, selection and identification. Selection is performed by using a semiquantitative screening plate that contains xylan substrate. Identification is based on microscopic and biochemical characteristics until the genus level.Selection results Showed 12 out of 16 isolates had xilanolitik activity, with the highest activity is SSA2 with xilanolitik index of 0.74. The top five index producehigestxilanolitik isolates that are SSA2, SSA3 and SSA4 identified as Bacillus sp. 1., and SSAS6 and SSA7 is Bacillus sp. 2.

[Identification and polymorphism of pectinase genes PGU in the Saccharomyces bayanus complex].

PubMed

Shalamitskiy, M Yu; Naumov, G I

2016-05-01

Pectinase (endo-polygalacturonase) is the key enzyme splitting plant pectin. The corresponding single gene PGU1 is documented for the yeast S. cerevisiae. On the basis of phylogenetic analysis of the PGU nucleotide sequence available in the GenBank, a family of divergent PGU genes is found in the species complex S. bayanus: S. bayanus var. uvarum, S. eubayanus, and hybrid taxon S. pastorianus. The PGU genes have different chromosome localization.

Production, partial purification and characterization of xylanase using Nicotiana tabacum leaf dust as substrate.

PubMed

Acharya, Komal P; Shilpkar, Prateek

2016-03-01

Isolated Bacillus sp. was used in the present study for production of xylanase from Nicotiana tabacum leaf dust. The strain was able to give a maximum of 1.77 Uml⁻¹ xylanase activity under optimized fermentation conditions which was further increased upto 2.77 Uml⁻¹ after extraction and partial purification of enzyme. After partial purification, the enzyme was characterized and it gave the highest xylanase activity at pH 7.0, when 0.2 ml enzyme was incubated with 2.0% substrate (Nicotiana tabacum leaf dust) for 60 min at 60°C. Saccharification study of Nicotiana tabacum leaf dust with partially purified enzyme revealed that 18.4% reducing sugar was released in 20 hrs incubation, and TLC and HPTLC analysis showed that xylose and glucose sugars were obtained after hydrolysis of substrate. FTIR analysis confirmed decomposition of substrate.

Polypeptides having xylanase activity and polynucleotides encoding the same

DOEpatents

Spodsberg, Nikolaj [Bagsvaed, DK

2014-01-07

The present invention relates to isolated polypeptides having xylanase activity and isolated polynucleotides encoding the polypeptides. The inventino also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides.

Bottleneck in secretion of α-amylase in Bacillus subtilis.

PubMed

Yan, Shaomin; Wu, Guang

2017-07-19

Amylase plays an important role in biotechnology industries, and Gram-positive bacterium Bacillus subtilis is a major host to produce heterogeneous α-amylases. However, the secretion stress limits the high yield of α-amylase in B. subtilis although huge efforts have been made to address this secretion bottleneck. In this question-oriented review, every effort is made to answer the following questions, which look simple but are long-standing, through reviewing of literature: (1) Does α-amylase need a specific and dedicated chaperone? (2) What signal sequence does CsaA recognize? (3) Does CsaA require ATP for its operation? (4) Does an unfolded α-amylase is less soluble than a folded one? (5) Does α-amylase aggregate before transporting through Sec secretion system? (6) Is α-amylase sufficient stable to prevent itself from misfolding? (7) Does α-amylase need more disulfide bonds to be stabilized? (8) Which secretion system does PrsA pass through? (9) Is PrsA ATP-dependent? (10) Is PrsA reused after folding of α-amylase? (11) What is the fate of PrsA? (12) Is trigger factor (TF) ATP-dependent? The literature review suggests that not only the most of those questions are still open to answers but also it is necessary to calculate ATP budget in order to better understand how B. subtilis uses its energy for production and secretion.

Properties of Thermosensitive Extracellular α-Amylases of Bacillus subtilis

PubMed Central

Yamane, Kunio; Maruo, Bunji

1974-01-01

Enzymological properties of four thermosensitive α-amylases (M3, M9, M18, and M20) brought by different mutation sites in α-amylase structural gene of Bacillus subtilis were compared with those of the parental α-amylase NA64. Two thermosensitive α-amylases (M9 and M20) were altered not only in their thermosensitivity but also in their immunological properties, catalytic properties, molecular weights determined by the gel filtration on a Bio-Gel P-100 column, and others. The other two thermosensitive α-amylases (M3 and M18) were altered only in their thermosensitivity. Images PMID:4218234

Analysis on evolutionary relationship of amylases from archaea, bacteria and eukaryota.

PubMed

Yan, Shaomin; Wu, Guang

2016-02-01

Amylase is one of the earliest characterized enzymes and has many applications in clinical and industrial settings. In biotechnological industries, the amylase activity is enhanced through modifying amylase structure and through cloning and expressing targeted amylases in different species. It is important to understand how engineered amylases can survive from generation to generation. This study used phylogenetic and statistical approaches to explore general patterns of amylases evolution, including 3118 α-amylases and 280 β-amylases from archaea, eukaryota and bacteria with fully documented taxonomic lineage. First, the phylogenetic tree was created to analyze the evolution of amylases with focus on individual amylases used in biofuel industry. Second, the average pairwise p-distance was computed for each kingdom, phylum, class, order, family and genus, and its diversity implies multi-time and multi-clan evolution. Finally, the variance was further partitioned into inter-clan variance and intra-clan variance for each taxonomic group, and they represent horizontal and vertical gene transfer. Theoretically, the results show a full picture on the evolution of amylases in manners of vertical and horizontal gene transfer, and multi-time and multi-clan evolution as well. Practically, this study provides the information on the surviving chance of desired amylase in a given taxonomic group, which may potentially enhance the successful rate of cloning and expression of amylase gene in different species.

Oligomerization triggered by foldon: a simple method to enhance the catalytic efficiency of lichenase and xylanase.

PubMed

Wang, Xinzhe; Ge, Huihua; Zhang, Dandan; Wu, Shuyu; Zhang, Guangya

2017-07-03

Effective and simple methods that lead to higher enzymatic efficiencies are highly sough. Here we proposed a foldon-triggered trimerization of the target enzymes with significantly improved catalytic performances by fusing a foldon domain at the C-terminus of the enzymes via elastin-like polypeptides (ELPs). The foldon domain comprises 27 residues and can forms trimers with high stability. Lichenase and xylanase can hydrolyze lichenan and xylan to produce value added products and biofuels, and they have great potentials as biotechnological tools in various industrial applications. We took them as the examples and compared the kinetic parameters of the engineered trimeric enzymes to those of the monomeric and wild type ones. When compared with the monomeric ones, the catalytic efficiency (k cat /K m ) of the trimeric lichenase and xylanase increased 4.2- and 3.9- fold. The catalytic constant (k cat ) of the trimeric lichenase and xylanase increased 1.8- fold and 5.0- fold than their corresponding wild-type counterparts. Also, the specific activities of trimeric lichenase and xylanase increased by 149% and 94% than those of the monomeric ones. Besides, the recovery of the lichenase and xylanase activities increased by 12.4% and 6.1% during the purification process using ELPs as the non-chromatographic tag. The possible reason is the foldon domain can reduce the transition temperature of the ELPs. The trimeric lichenase and xylanase induced by foldon have advantages in the catalytic performances. Besides, they were easier to purify with increased purification fold and decreased the loss of activities compared to their corresponding monomeric ones. Trimerizing of the target enzymes triggered by the foldon domain could improve their activities and facilitate the purification, which represents a simple and effective enzyme-engineering tool. It should have exciting potentials both in industrial and laboratory scales.

Biotechnological Processes in Microbial Amylase Production

PubMed Central

Arshad, M. K. Md; Lakshmipriya, Thangavel; Hashim, Uda; Chinni, Suresh V.

2017-01-01

Amylase is an important and indispensable enzyme that plays a pivotal role in the field of biotechnology. It is produced mainly from microbial sources and is used in many industries. Industrial sectors with top-down and bottom-up approaches are currently focusing on improving microbial amylase production levels by implementing bioengineering technologies. The further support of energy consumption studies, such as those on thermodynamics, pinch technology, and environment-friendly technologies, has hastened the large-scale production of the enzyme. Herein, the importance of microbial (bacteria and fungi) amylase is discussed along with its production methods from the laboratory to industrial scales. PMID:28280725

Biotechnological Processes in Microbial Amylase Production.

PubMed

Gopinath, Subash C B; Anbu, Periasamy; Arshad, M K Md; Lakshmipriya, Thangavel; Voon, Chun Hong; Hashim, Uda; Chinni, Suresh V

2017-01-01

Amylase is an important and indispensable enzyme that plays a pivotal role in the field of biotechnology. It is produced mainly from microbial sources and is used in many industries. Industrial sectors with top-down and bottom-up approaches are currently focusing on improving microbial amylase production levels by implementing bioengineering technologies. The further support of energy consumption studies, such as those on thermodynamics, pinch technology, and environment-friendly technologies, has hastened the large-scale production of the enzyme. Herein, the importance of microbial (bacteria and fungi) amylase is discussed along with its production methods from the laboratory to industrial scales.

Production of enzymes by a newly isolated Bacillus sp. TMF-1 in solid state fermentation on agricultural by-products: The evaluation of substrate pretreatment methods.

PubMed

Salim, Abdalla Ali; Grbavčić, Sanja; Šekuljica, Nataša; Stefanović, Andrea; Jakovetić Tanasković, Sonja; Luković, Nevena; Knežević-Jugović, Zorica

2017-03-01

Study on potential of different agro-industrial waste residues for supporting the growth of newly isolated Bacillus sp. TMF-1 strain under solid-state fermentation (SSF) was conducted aiming to produce several industrially valuable enzymes. Since the feasibility of the initial study was confirmed, further objectives included evaluation of several pretreatments of the studied agricultural by-products (soybean meal, sunflower meal, maize bran, maize pericarp, olive oil cake and wheat bran) on the microbial productivity as means of enhancing the yields of produced proteases, α-amylases, cellulases and pectinases. Among acid/alkaline treatment, ultrasound and microwave assisted methods, chemical treatments superiorly affected most of the studied substrates. Highest yields of produced proteases (50.5IUg -1 ) and α-amylases (50.75IUg -1 ) were achieved on alkaline treated corn pericarp. Alkaline treatment also promoted the secretion of cellulases on maize bran (1.19IUg -1 ). Highest yield of pectinases was obtained on untreated soybean meal (64.90IUg -1 ). Copyright © 2016 Elsevier Ltd. All rights reserved.

Enhanced production of xylanase from locally isolated fungal strain using agro-industrial residues under solid-state fermentation.

PubMed

Abdullah, Roheena; Nisar, Kinza; Aslam, Aafia; Iqtedar, Mehwish; Naz, Shagufta

2015-01-01

This study is related to the isolation of fungal strain for xylanase production using agro-industrial residues. Forty fungal strains with xylanolytic potential were isolated by using xylan agar plates and quantitatively screened in solid-state fermentation. Of all the tested isolates, the strain showing highest ability to produce xylanase was assigned the code Aspergillus niger LCBT-14. For the enhanced production of the enzyme, five different fermentation media were evaluated. Out of all media, M4 containing wheat bran gave maximum enzyme production. Effect of different variables including incubation time, temperature, pH, carbon and nitrogen sources has been investigated. The optimum enzyme production was obtained after 72 h at 30°C and pH 4. Glucose as a carbon source while ammonium sulphate and yeast extract as nitrogen sources gave maximum xylanase production (946 U/mL/min). This study was successful in producing xylanase by A. niger LCBT-14 economically by utilising cheap indigenous substrate.

A Novel Aqueous Micellar Two-Phase System Composed of Surfactant and Sorbitol for Purification of Pectinase Enzyme from Psidium guajava and Recycling Phase Components

PubMed Central

Murshid, Fara Syazana; Manap, Mohd Yazid; Hussin, Muhaini

2015-01-01

A novel aqueous two-phase system composed of a surfactant and sorbitol was employed for the first time to purify pectinase from Psidium guajava. The influences of different parameters, including the type and concentration of the surfactant and the concentration and composition of the surfactant/sorbitol ratio, on the partitioning behavior and recovery of pectinase were investigated. Moreover, the effects of system pH and the crude load on purification fold and the yield of purified pectinase were studied. The experimental results indicated that the pectinase was partitioned into surfactant-rich top phase, and the impurities were partitioned into the sorbitol-rich bottom phase with the novel method involving an ATPS composed of 26% (w/w) Triton X-100 and 23% (w/w) sorbitol at 54.2% of the TLL crude load of 20% (w/w) at pH 6.0. The enzyme was successfully recovered by this method with a high purification factor of 15.2 and a yield of 98.3%, whereas the phase components were also recovered and recycled at rates above 96%. This study demonstrated that this novel ATPS method can be used as an efficient and economical alternative to the traditional ATPS for the purification and recovery of the valuable enzyme. PMID:25756051

A novel aqueous micellar two-phase system composed of surfactant and sorbitol for purification of pectinase enzyme from Psidium guajava and recycling phase components.

PubMed

Amid, Mehrnoush; Murshid, Fara Syazana; Manap, Mohd Yazid; Hussin, Muhaini

2015-01-01

A novel aqueous two-phase system composed of a surfactant and sorbitol was employed for the first time to purify pectinase from Psidium guajava. The influences of different parameters, including the type and concentration of the surfactant and the concentration and composition of the surfactant/sorbitol ratio, on the partitioning behavior and recovery of pectinase were investigated. Moreover, the effects of system pH and the crude load on purification fold and the yield of purified pectinase were studied. The experimental results indicated that the pectinase was partitioned into surfactant-rich top phase, and the impurities were partitioned into the sorbitol-rich bottom phase with the novel method involving an ATPS composed of 26% (w/w) Triton X-100 and 23% (w/w) sorbitol at 54.2% of the TLL crude load of 20% (w/w) at pH 6.0. The enzyme was successfully recovered by this method with a high purification factor of 15.2 and a yield of 98.3%, whereas the phase components were also recovered and recycled at rates above 96%. This study demonstrated that this novel ATPS method can be used as an efficient and economical alternative to the traditional ATPS for the purification and recovery of the valuable enzyme.

A xylose-stimulated xylanase-xylose binding protein chimera created by random nonhomologous recombination.

PubMed

Ribeiro, Lucas Ferreira; Tullman, Jennifer; Nicholes, Nathan; Silva, Sérgio Ruschi Bergamachi; Vieira, Davi Serradella; Ostermeier, Marc; Ward, Richard John

2016-01-01

Saccharification of lignocellulosic material by xylanases and other glycoside hydrolases is generally conducted at high concentrations of the final reaction products, which frequently inhibit the enzymes used in the saccharification process. Using a random nonhomologous recombination strategy, we have fused the GH11 xylanase from Bacillus subtilis (XynA) with the xylose binding protein from Escherichia coli (XBP) to produce an enzyme that is allosterically stimulated by xylose. The pT7T3GFP_XBP plasmid containing the XBP coding sequence was randomly linearized with DNase I, and ligated with the XynA coding sequence to create a random XynA-XBP insertion library, which was used to transform E. coli strain JW3538-1 lacking the XBP gene. Screening for active XBP was based on the expression of GFP from the pT7T3GFP_XBP plasmid under the control of a xylose inducible promoter. In the presence of xylose, cells harboring a functional XBP domain in the fusion protein (XBP+) showed increased GFP fluorescence and were selected using FACS. The XBP+ cells were further screened for xylanase activity by halo formation around xylanase producing colonies (XynA+) on LB-agar-xylan media after staining with Congo red. The xylanase activity ratio with xylose/without xylose in supernatants from the XBP+/XynA+ clones was measured against remazol brilliant blue xylan. A clone showing an activity ratio higher than 1.3 was selected where the XynA was inserted after the asparagine 271 in the XBP, and this chimera was denominated as XynA-XBP271. The XynA-XBP271 was more stable than XynA at 55 °C, and in the presence of xylose the catalytic efficiency was ~3-fold greater than the parental xylanase. Molecular dynamics simulations predicted the formation of an extended protein-protein interface with coupled movements between the XynA and XBP domains. In the XynA-XBP271 with xylose bound to the XBP domain, the mobility of a β-loop in the XynA domain results in an increased access to the

Specific starch digestion of maize alpha-limit dextrins by recombinant mucosal glucosidase enzymes

USDA-ARS?s Scientific Manuscript database

Starch digestion requires two luminal enzymes, salivary and pancreatic alpha-amylase (AMY), and four small intestinal mucosal enzyme activities from the N- and C-terminals of maltase-glucoamylase (MGAM) and sucrose-isomaltase (SI) complexes. AMY is not a requirement for starch digestion to glucose b...

Contribution of mucosal maltase-glucoamylase activities to mouse small intestinal starch alpha-glucogenesis

USDA-ARS?s Scientific Manuscript database

Digestion of starch requires activities provided by 6 interactive small intestinal enzymes. Two of these are luminal endo-glucosidases named alpha-amylases. Four are exo-glucosidases bound to the luminal surface of enterocytes. These mucosal activities were identified as 4 different maltases. Two ma...

Characterization of glucoamylase and α-amylase from Monascus anka: enhanced production of α-amylase in red koji.

PubMed

Yoshizaki, Yumiko; Susuki, Tomoka; Takamine, Kazunori; Tamaki, Hisanori; Ito, Kiyoshi; Sameshima, Yoshihiro

2010-12-01

To enhance glucoamylase and α-amylase production from Monascus anka, we investigated the influence of different culture conditions on enzyme production and purified and characterized these enzymes. The effect of different raw materials was investigated by using solid-state plates of raw materials such as barley and non-waxy or waxy rice. The barley plate was the most suitable for mycelial growth, but glucoamylase and α-amylase production per growth area did not differ according to the raw material. Investigation of the effect of temperature showed that incubation at 37 °C promoted maximal cell growth, while incubation at 25 °C and at 40 °C resulted in enhanced α-amylase and glucoamylase production, respectively. Characterization of the purified enzymes revealed that α-amylase was unstable at acidic pH and less resistant to heat (stable at < 40 °C) than glucoamylase. When these culture conditions were applied to enzyme production in red koji, reducing the temperature from 35 °C to 25 °C for 48 h in the late stages of growth resulted in higher glucoamylase and α-amylase production (1.4 and 18 times, respectively) with a concomitant increase in protein stability. Copyright © 2010 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Energy and nutrient utilization of broiler chickens fed corn-soybean meal and corn-based diets supplemented with xylanase.

PubMed

Stefanello, C; Vieira, S L; Carvalho, P S; Sorbara, J O B; Cowieson, A J

2016-08-01

A study was conducted to evaluate the effects of increased levels of a β-xylanase on energy and nutrient utilization of broiler chickens fed corn-soy diets. A total of 480 slow feathering Cobb × Cobb 500 male broilers were randomly distributed to 10 treatments having 8 replicates of 6 birds each. Birds were fed a common starter diet to d 14 post hatch (3,050 kcal/kg AMEn, 21.7% CP, 1.05% Ca, and 0.53% nPP). The experimental diets were provided afterwards until 25 d. Two experimental diets, a conventional corn/soy-based basal diet (CS) and the basal diet in which 40% of the diet was displaced by corn (CN), were fed as-is or supplemented with 50, 100, 150, or 200 fungal β-xylanase units (FXU)/kg. Dietary treatments were distributed factorially as a 2 × 5 arrangement. Samples of feed, excreta, and ileal digesta were analyzed for determination of ileal digestible energy (IDE), metabolizable energy, and total tract retention of protein and lipid. No interactions between diet and xylanase were observed. The CS diets had higher (P < 0.05) energy utilization and nutrient digestibility when compared to the CN diets. AMEn and IDE were improved (P < 0.05) by 192 and 145 kcal/kg, respectively, when diets were supplemented with 100 FXU/kg xylanase. The xylanase added to the CN diet led to quadratic increases (P < 0.05) in IDE (Y = - 0.014x(2) + 2.570x + 3,155; r(2) = 0.60) and in AMEn (Y = - 0.016x(2) + 3.982x + 3,155; r(2) = 0.68). Crude protein digestibility and AMEn were linearly increased (P < 0.05) when xylanase was added to the CN diet. In conclusion, energy utilization and digestibility of crude protein and dry matter increased with xylanase supplementation in corn/soy-based diets. When xylanase was tested in the CS diet, 92 and 124 FXU/kg maximized the energy release effect; however, the maximum energy response in the CN diet or corn was not achieved until 200 FXU/kg. © 2016 Poultry Science Association Inc.

Production of Sporotrichum thermophile xylanase by solid state fermentation utilizing deoiled Jatropha curcas seed cake and its application in xylooligosachharide synthesis.

PubMed

Sadaf, Ayesha; Khare, S K

2014-02-01

De-oiled Jatropha curcas seed cake, a plentiful by-product of biodiesel industry was used as substrate for the production of a useful xylanase from Sporotrichum thermophile in solid state fermentation. Under the optimized conditions, 1025U xylanase/g (deoiled seed cake) was produced. The xylanase exhibited half life of 4h at 45°C and 71.44min at 50°C respectively. It was stable in a broad pH range of 7.0-11.0. Km and Vmax were 12.54mg/ml and 454.5U/ml/min respectively. S. thermophile xylanase is an endoxylanase free of exoxylanase activity, hence advantageous for xylan hydrolysis to produce xylooligosachharides. Hydrolysis of oat spelt xylan by S. thermophile xylanase yielded 73% xylotetraose, 15.4% xylotriose and 10% xylobiose. The S. thermophile endoxylanase thus seem potentially useful in the food industries. Copyright © 2013 Elsevier Ltd. All rights reserved.

Amylases and Their Importance during Glycan Degradation: Genome Sequence Release of Salmonella Amylase Knockout Strains.

PubMed

Arabyan, Narine; Huang, Bihua C; Weimer, Bart C

2017-05-18

Amylases catalyze the cleavage of α-d-1,4 and α-d-1,6-glycosidic bonds in starch and related carbohydrates. Amylases are widely distributed in nature and are important in carbohydrate metabolism. This is the release of four single and two double deletions in Salmonella enterica serovar Typhimurium LT2 that are important for glycan degradation during infection. Copyright © 2017 Arabyan et al.

[Determination of exogenous gamma-amylase residue in honey].

PubMed

Fei, Xiaoqing; Wu, Bin; Shen, Chongyu; Zhang, Rui; Ding, Tao; Li, Lihua

2012-08-01

A novel method for the determination of exogenous gamma-amylase residue in honey using liquid chromatography-isotope ratio mass spectrometry (LC-IRMS) was established. After pre-separation by gel column chromatography, the gamma-amylase in honey samples was separated from the sugars. The gamma-amylase was then used to catalyze maltose into glucose. This enzymatic reaction was under the conditions of 55 degrees C and 0.03 mol/L phosphate buffer solution (pH 4.5) for 48 h. The maltose and glucose in the above enzymatic reaction solution were separated using liquid chromatography. By measuring the content of glucose with isotope ratio mass spectrometry, the gamma-amylase in honey can be determined. The linear range of gamma-amylase was 5 - 200 U/kg with the quantification limit of 5 U/kg. The recoveries were between 89.6% and 108.2% with the relative standard deviations from 3.3% to 4.9%. This method was used to analyze 38 honey and rice syrup samples, and the detection rate of gamma-amylase was 76.3%. To further verify the detection capability of this method, an authentic honey was adulterated with 15% (mass fraction) rice syrup. The gamma-amylase content in this sample was 10.2 U/kg. This method can effectively identify honey adulteration with rice syrups from the perspective of enzymology.

Phylogenetic Distribution of Intron Positions in Alpha-Amylase Genes of Bilateria Suggests Numerous Gains and Losses

PubMed Central

Da Lage, Jean-Luc; Maczkowiak, Frédérique; Cariou, Marie-Louise

2011-01-01

Most eukaryotes have at least some genes interrupted by introns. While it is well accepted that introns were already present at moderate density in the last eukaryote common ancestor, the conspicuous diversity of intron density among genomes suggests a complex evolutionary history, with marked differences between phyla. The question of the rates of intron gains and loss in the course of evolution and factors influencing them remains controversial. We have investigated a single gene family, alpha-amylase, in 55 species covering a variety of animal phyla. Comparison of intron positions across phyla suggests a complex history, with a likely ancestral intronless gene undergoing frequent intron loss and gain, leading to extant intron/exon structures that are highly variable, even among species from the same phylum. Because introns are known to play no regulatory role in this gene and there is no alternative splicing, the structural differences may be interpreted more easily: intron positions, sizes, losses or gains may be more likely related to factors linked to splicing mechanisms and requirements, and to recognition of introns and exons, or to more extrinsic factors, such as life cycle and population size. We have shown that intron losses outnumbered gains in recent periods, but that “resets” of intron positions occurred at the origin of several phyla, including vertebrates. Rates of gain and loss appear to be positively correlated. No phase preference was found. We also found evidence for parallel gains and for intron sliding. Presence of introns at given positions was correlated to a strong protosplice consensus sequence AG/G, which was much weaker in the absence of intron. In contrast, recent intron insertions were not associated with a specific sequence. In animal Amy genes, population size and generation time seem to have played only minor roles in shaping gene structures. PMID:21611157

Characterization of two coleopteran α-amylases and molecular insights into their differential inhibition by synthetic α-amylase inhibitor, acarbose.

PubMed

Channale, Sonal M; Bhide, Amey J; Yadav, Yashpal; Kashyap, Garima; Pawar, Pankaj K; Maheshwari, V L; Ramasamy, Sureshkumar; Giri, Ashok P

2016-07-01

Post-harvest insect infestation of stored grains makes them unfit for human consumption and leads to severe economic loss. Here, we report functional and structural characterization of two coleopteran α-amylases viz. Callosobruchus chinensis α-amylase (CcAmy) and Tribolium castaneum α-amylase (TcAmy) along with their interactions with proteinaceous and non-proteinaceous α-amylase inhibitors. Secondary structural alignment of CcAmy and TcAmy with other coleopteran α-amylases revealed conserved motifs, active sites, di-sulfide bonds and two point mutations at spatially conserved substrate or inhibitor-binding sites. Homology modeling and molecular docking showed structural differences between these two enzymes. Both the enzymes had similar optimum pH values but differed in their optimum temperature. Overall, pattern of enzyme stabilities were similar under various temperature and pH conditions. Further, CcAmy and TcAmy differed in their substrate affinity and catalytic efficiency towards starch and amylopectin. HPLC analysis detected common amylolytic products like maltose and malto-triose while glucose and malto-tetrose were unique in CcAmy and TcAmy catalyzed reactions respectively. At very low concentrations, wheat α-amylase inhibitor was found to be superior over the acarbose as far as complete inhibition of amylolytic activities of CcAmy and TcAmy was concerned. Mechanism underlying differential amylolytic reaction inhibition by acarbose was discussed. Copyright © 2016 Elsevier Ltd. All rights reserved.

Depolymerization of starch and pectin using superporous matrix supported enzymes.

PubMed

Lali, Arvind; Manudhane, Kushal; Motlekar, Nuzhat; Karandikar, Priti

2002-08-01

Immobilized enzyme catalyzed biotransformations involving macromolecular substrates and/or products are greatly retarded due to slow diffusion of large substrate molecules in and out of the typical enzyme supports. Slow diffusion of macromolecules into the matrix pores can be speeded up by use of macroporous supports as enzyme carriers. Depolymerization reactions of polysaccharides like starch, pectin, and dextran to their respective low molecular weight products are some of the reactions that can benefit from use of such superporous matrices. In the present work, an indigenously prepared rigid cross-linked cellulose matrix (called CELBEADS) has been used as support for immobilizing alpha amylase (1,4-alpha-D-glucan glucanohydrolase, EC 3.2.1.1.) and pectinase (endo-PG: poly(1,4-alpha-galactouronide) glycanohydrolase, EC 3.2.1.15). The immobilized enzymes were used for starch and pectin hydrolysis respectively, in batch, packed bed and expanded bed modes. The macroporosity of CELBEADS was found to permit through-flow and easy diffusion of substrates pectin and starch to enzyme sites in the porous supports and gave reaction rates comparable to the rates obtained using soluble enzymes.

Application of microbial α-amylase in industry - A review.

PubMed

de Souza, Paula Monteiro; de Oliveira Magalhães, Pérola

2010-10-01

Amylases are one of the main enzymes used in industry. Such enzymes hydrolyze the starch molecules into polymers composed of glucose units. Amylases have potential application in a wide number of industrial processes such as food, fermentation and pharmaceutical industries. α-Amylases can be obtained from plants, animals and microorganisms. However, enzymes from fungal and bacterial sources have dominated applications in industrial sectors. The production of α-amylase is essential for conversion of starches into oligosaccharides. Starch is an important constituent of the human diet and is a major storage product of many economically important crops such as wheat, rice, maize, tapioca, and potato. Starch-converting enzymes are used in the production of maltodextrin, modified starches, or glucose and fructose syrups. A large number of microbial α-amylases has applications in different industrial sectors such as food, textile, paper and detergent industries. The production of α-amylases has generally been carried out using submerged fermentation, but solid state fermentation systems appear as a promising technology. The properties of each α-amylase such as thermostability, pH profile, pH stability, and Ca-independency are important in the development of fermentation process. This review focuses on the production of bacterial and fungal α-amylases, their distribution, structural-functional aspects, physical and chemical parameters, and the use of these enzymes in industrial applications.

Purification and characterization of α-Amylase from Miswak Salvadora persica.

PubMed

Mohamed, Saleh A; Almulaiky, Yaaser Q; Ahmed, Youssri M; Al-Bar, Omar A M; Ibrahim, Ibrahim H

2014-04-01

The miswak (Salvadora persica) is a natural toothbrush. It is well known that very little information has been reported on enzymes in miswak as medicinal plant. Recently, we study peroxidase in miswak. In the present study, the main goal of this work is to purify and characterize α-amylase from miswak. The second goal is to study the storage stability of α-amylase in toothpaste. The purification method included chromatography of miswak α-amylase on DEAE-Sepharose column and Sephacryl S-200 column. Molecular weight was determined by gel filtration and SDS-PAGE. Five α-amylases A1, A4a, A4b, A5a and A5b from miswak were purified and they had molecular weights of 14, 74, 16, 30 and 20 kDa, respectively. α-Amylases had optimum pH from 6 to 8. Affinity of the substrates toward all enzymes was studied. Miswak α-amylases A1, A4a, A4b, A5a and A5b had Km values for starch and glycogen of 3.7, 3.7, 7.1, 0.52, 4.3 mg/ml and 5.95, 5.9 4.16, 6.3, 6.49 mg/ml, respectively. The optimum temperature for five enzymes ranged 40°C- 60°C. Miswak α-amylases were stable up to 40°C- 60°C after incubation for 30 min. Ca+2 activated all the miswak α-amylases, while Ni2+, Co+2 and Zn+2 activated or inhibited some of these enzymes. The metal chelators, EDTA, sodium citrate and sodium oxalate had inhibitory effects on miswak α-amylases. PMSF, p-HMB, DTNB and 1,10 phenanthroline caused inhibitory effect on α-amylases. The analysis of hydrolytic products after starch hydrolysis by miswak α-amylases on paper chromatography revealed that glucose, maltose, maltotriose and oligosaccharide were the major products. Crude miswak α-amylase in the toothpaste retained 55% of its original activity after 10 months of storage at room temperature. From these findings, α-amylases from miswak can be considered as beneficial enzymes for pharmaceuticals. Therefore, we study the storage stability of the crude α-amylase of miswak, which contained the five α-amylases, in toothpaste. The enzyme in

Purification and characterization of α-Amylase from Miswak Salvadora persica

PubMed Central

2014-01-01

Background The miswak (Salvadora persica) is a natural toothbrush. It is well known that very little information has been reported on enzymes in miswak as medicinal plant. Recently, we study peroxidase in miswak. In the present study, the main goal of this work is to purify and characterize α-amylase from miswak. The second goal is to study the storage stability of α-amylase in toothpaste. Method The purification method included chromatographaphy of miswak α-amylase on DEAE-Sepharose column and Sephacryl S-200 column. Molecular weight was determined by gel filtration and SDS-PAGE. Results Five α-amylases A1, A4a, A4b, A5a and A5b from miswak were purified and they had molecular weights of 14, 74, 16, 30 and 20 kDa, respectively. α-Amylases had optimum pH from 6 to 8. Affinity of the substrates toward all enzymes was studied. Miswak α-amylases A1, A4a, A4b, A5a and A5b had Km values for starch and glycogen of 3.7, 3.7, 7.1, 0.52, 4.3 mg/ml and 5.95, 5.9 4.16, 6.3, 6.49 mg/ml, respectively. The optimum temperature for five enzymes ranged 40°C- 60°C. Miswak α-amylases were stable up to 40°C- 60°C after incubation for 30 min. Ca+2 activated all the miswak α-amylases, while Ni2+, Co+2 and Zn+2 activated or inhibited some of these enzymes. The metal chelators, EDTA, sodium citrate and sodium oxalate had inhibitory effects on miswak α-amylases. PMSF, p-HMB, DTNB and 1,10 phenanthroline caused inhibitory effect on α-amylases. The analysis of hydrolytic products after starch hydrolysis by miswak α-amylases on paper chromatography revealed that glucose, maltose, maltotriose and oligosaccharide were the major products. Crude miswak α-amylase in the toothpaste retained 55% of its original activity after 10 months of storage at room temperature. Conclusions From these findings, α-amylases from miswak can be considered as beneficial enzymes for pharmaceuticals. Therefore, we study the storage stability of the crude α-amylase of miswak, which contained the five

A Pilot Study of Psychotherapist Trainees’ Alpha-Amylase and Cortisol Levels During Treatment of Recently Suicidal Clients With Borderline Traits

PubMed Central

Miller, Grant D.; Iverson, Katherine M.; Kemmelmeier, Markus; MacLane, Chelsea; Pistorello, Jacqueline; Fruzzetti, Alan E.; Crenshaw, Katrina Y.; Erikson, Karen M.; Katrichak, Barrie M.; Oser, Megan; Pruitt, Larry D.; Watkins, Melanie M.

2010-01-01

Psychotherapists often experience stress while providing psychotherapy, in particular when working with difficult presentations such as suicidality. As part of a larger study on the treatment of recently suicidal college students with borderline traits, 6 therapists in training collected their own salivary samples for alpha-amylase (AA) and cortisol (C) analyses immediately before and after sessions with 2 selected clients. On average, samples were collected for the same therapist–patient dyad throughout the year-long study to ensure that data reflected therapist responses across stages of treatment. Therapists also completed a working alliance questionnaire and rated perceived session difficulty immediately after each selected session. Contrary to expectations, therapists demonstrated elevated levels of stress as measured by AA and C at presession relative to postsession levels. Greater session difficulty was related to more pronounced declines in AA, whereas a stronger working alliance was linked to more pronounced reductions in C. Results suggest that physiological stress responses while working with recently suicidal clients with borderline traits occur primarily in terms of session anticipatory anxiety, whereas AA and C changes may be affected differently by factors such as session difficulty and working alliance. This is a pilot study, limited by its sample size, but the design, findings, and inclusion of physiological measures present an initial step in an essential line of research. PMID:21709772

Potent α-glucosidase and α-amylase inhibitory activities of standardized 50% ethanolic extracts and sinensetin from Orthosiphon stamineus Benth as anti-diabetic mechanism

PubMed Central

2012-01-01

Background In the present study, we tested a 50% ethanolic extract of Orthosiphon stamineus plants and its isolated bioactive compound with respect to their α-glucosidase and α-amylase inhibitory activities. Methods Bioactive flavonoid sinensetin was isolated from 50% ethanolic extract of Orthosiphon stamineus. The structure of this pure compound was determined on the NMR data and the α-glucosidase and α-amylase inhibitory activities of isolated sinensetin and 50% ethanolic extract of Orthosiphon stamineus were evaluated. Results In vitro studies of a 50% ethanolic extract of O. stamineus and the isolated sinensetin compound showed inhibitory activity on α-glucosidase (IC50: 4.63 and 0.66 mg/ml, respectively) and α-amylase (IC50: 36.70 mg/ml and 1.13 mg/ml, respectively). Inhibition of these enzymes provides a strong biochemical basis for the management of type 2 diabetes via the control of glucose absorption. Conclusion Alpha-glucosidase and α-amylase inhibition could the mechanisms through which the 50% ethanolic extract of O. stamineus and sinensetin exert their antidiabetic activity, indicating that it could have potential use in the management of non-insulin-dependent diabetes. PMID:23039079

Polypeptides having xylanase activity and polynucleotides encoding same

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

Spodsberg, Nikolaj; Shaghasi, Tarana

The present invention relates to polypeptides having xylanase activity, catalytic domains, and carbohydrate binding domains, and polynucleotides encoding the polypeptides, catalytic domains, and carbohydrate binding domains. The present invention also relates to nucleic acid constructs, vectors, and host cells comprising the polynucleotides as well as methods of producing and using the polypeptides, catalytic domains, and carbohydrate binding domains.