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Sample records for protein hydrolysate preparation

  1. Antioxidant Properties of Fish Protein Hydrolysates Prepared from Cod Protein Hydrolysate by Bacillus sp.

    PubMed

    Godinho, I; Pires, C; Pedro, S; Teixeira, B; Mendes, R; Nunes, M L; Batista, I

    2016-03-01

    Fermentative protein hydrolysates (FPH) were prepared with a proteolytic bacterium, Bacillus strain exhibiting high proteolytic activity. Three FPH with 1, 2, and 4 % of cod protein hydrolysate (CPH) and 0.5 % of yeast extract in the culture were prepared. The yields achieved varied between 30 and 58 % based on protein content. A general decrease of leucine, isoleucine, valine, alanine, arginine, threonine, proline, and glutamic acid was observed. All FPHs showed higher reducing power and DPPH radical scavenging activity than CPH, but similar ABTS radical scavenging activity. However, FPHs exhibited lower Cu(+2)-chelating activity than CPH. The ACE inhibitory activity of FPHs was not improved relatively to that recorded in CPH. The fermentative process seems to have potential to obtaining hydrolysates with improved biological activities or even to produce protein hydrolysates from native fish proteins. PMID:26590847

  2. Optimization of the Preparation of Fish Protein Anti-Obesity Hydrolysates Using Response Surface Methodology

    PubMed Central

    Liu, Liyuan; Wang, Yanping; Peng, Chen; Wang, Jinju

    2013-01-01

    The enzymatic condition for producing the anti-obesity hydrolysates from fish water-soluble protein was optimized with the aid of response surface methodology, which also derived a statistical model for experimental validation. Compared with neutral protease, papain and protamex, the porcine pancreas lipase inhibitory rate of hydrolysates from fish water-soluble protein was higher with alkaline protease. Results showed that the model terms were significant, the terms of lack of fit were not significant, and the optimal conditions for the hydrolysis by alkaline protease were initial pH 11, temperature 39 °C, enzyme dosage 122 U/mL and 10 h of hydrolysis time. Under these conditions, the porcine pancreas lipase and the α-amylase inhibitory rate could reach 53.04% ± 1.32% and 20.03 ± 0.89%, while predicted value were 54.63% ± 1.75%, 21.22% ± 0.70%, respectively. In addition, Lineweaver-Burk plots showed noncompetitive inhibition. The Ki value calculated was 84.13 mg/mL. These results demonstrated that fish water-soluble protein could be used for obtaining anti-obesity hydrolysates. PMID:23377020

  3. Abalone Protein Hydrolysates: Preparation, Angiotensin I Converting Enzyme Inhibition and Cellular Antioxidant Activity

    PubMed Central

    Park, Soo Yeon; Je, Jae-Young; Hwang, Joung-Youl; Ahn, Chang-Bum

    2015-01-01

    Abalone protein was hydrolyzed by enzymatic hydrolysis and the optimal enzyme/substrate (E/S) ratios were determined. Abalone protein hydrolysates (APH) produced by Protamex at E/S ratio of 1:100 showed angiotensin I converting enzyme inhibitory activity with IC50 of 0.46 mg/mL, and APH obtained by Flavourzyme at E/S ratio of 1:100 possessed the oxygen radical absorbance capacity value of 457.6 μM trolox equivalent/mg sample. Flavourzyme abalone protein hydrolysates (FAPH) also exhibited H2O2 scavenging activity with IC50 of 0.48 mg/mL and Fe2+ chelating activity with IC50 of 2.26 mg/mL as well as high reducing power. FAPH significantly (P<0.05) protected H2O2-induced hepatic cell damage in cultured hepatocytes, and the cell viability was restored to 90.27% in the presence of FAPH. FAPH exhibited 46.20% intracellular ROS scavenging activity and 57.89% lipid peroxidation inhibition activity in cultured hepatocytes. Overall, APH may be useful as an ingredient for functional foods. PMID:26451354

  4. Abalone Protein Hydrolysates: Preparation, Angiotensin I Converting Enzyme Inhibition and Cellular Antioxidant Activity.

    PubMed

    Park, Soo Yeon; Je, Jae-Young; Hwang, Joung-Youl; Ahn, Chang-Bum

    2015-09-01

    Abalone protein was hydrolyzed by enzymatic hydrolysis and the optimal enzyme/substrate (E/S) ratios were determined. Abalone protein hydrolysates (APH) produced by Protamex at E/S ratio of 1:100 showed angiotensin I converting enzyme inhibitory activity with IC50 of 0.46 mg/mL, and APH obtained by Flavourzyme at E/S ratio of 1:100 possessed the oxygen radical absorbance capacity value of 457.6 μM trolox equivalent/mg sample. Flavourzyme abalone protein hydrolysates (FAPH) also exhibited H2O2 scavenging activity with IC50 of 0.48 mg/mL and Fe(2+) chelating activity with IC50 of 2.26 mg/mL as well as high reducing power. FAPH significantly (P<0.05) protected H2O2-induced hepatic cell damage in cultured hepatocytes, and the cell viability was restored to 90.27% in the presence of FAPH. FAPH exhibited 46.20% intracellular ROS scavenging activity and 57.89% lipid peroxidation inhibition activity in cultured hepatocytes. Overall, APH may be useful as an ingredient for functional foods. PMID:26451354

  5. Preparation and antioxidative properties of a rapeseed ( Brassica napus ) protein hydrolysate and three peptide fractions.

    PubMed

    Xue, Zhaohui; Yu, Wancong; Liu, Zhiwei; Wu, Moucheng; Kou, Xiaohong; Wang, Jiehua

    2009-06-24

    This study investigated the possibility of converting the insoluble rapeseed meal protein into functionally active ingredients for food applications. The rapeseed ( Brassica napus ) meal protein isolates were first digested by Alcalase and Flavourzyme, and the resultant rapeseed crude hydrolysate (RSCH) exhibited a dose-dependent reducing antioxidant power and hydroxyl radical scavenging ability. RSCH could also inhibit the malonyldialdehyde (MDA) generation by 50% in blood serum at 150 mg/mL. RSCH was further separated into three fractions (RSP1, RSP2, and RSP3) by Sephadex gel filtration according to their different molecular weights. The amino acid compositions and antioxidant potentials were assessed for RSP1-3 fractions. All three fractions showed inhibiting effects on superoxide anion generation to various extents. They could also inhibit the autohemolysis of rat red blood cells and MDA formation in rat liver tissue homogenate. The results suggested that rapeseed peptide hydrolysate may be useful as a human food addition as a source of bioactive peptides with antioxidant properties.

  6. 21 CFR 102.22 - Protein hydrolysates.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... derived. (a) “Hydrolyzed wheat gluten,” “hydrolyzed soy protein,” and “autolyzed yeast extract” are... “hydrolyzed milk protein” is not an acceptable name for this ingredient because it is not specific to the ingredient (hydrolysates can be prepared from other milk proteins). The names “hydrolyzed vegetable...

  7. 21 CFR 102.22 - Protein hydrolysates.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... derived. (a) “Hydrolyzed wheat gluten,” “hydrolyzed soy protein,” and “autolyzed yeast extract” are... “hydrolyzed milk protein” is not an acceptable name for this ingredient because it is not specific to the ingredient (hydrolysates can be prepared from other milk proteins). The names “hydrolyzed vegetable...

  8. 21 CFR 102.22 - Protein hydrolysates.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... derived. (a) “Hydrolyzed wheat gluten,” “hydrolyzed soy protein,” and “autolyzed yeast extract” are... “hydrolyzed milk protein” is not an acceptable name for this ingredient because it is not specific to the ingredient (hydrolysates can be prepared from other milk proteins). The names “hydrolyzed vegetable...

  9. 21 CFR 102.22 - Protein hydrolysates.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... derived. (a) “Hydrolyzed wheat gluten,” “hydrolyzed soy protein,” and “autolyzed yeast extract” are... “hydrolyzed milk protein” is not an acceptable name for this ingredient because it is not specific to the ingredient (hydrolysates can be prepared from other milk proteins). The names “hydrolyzed vegetable...

  10. 21 CFR 102.22 - Protein hydrolysates.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... derived. (a) “Hydrolyzed wheat gluten,” “hydrolyzed soy protein,” and “autolyzed yeast extract” are... “hydrolyzed milk protein” is not an acceptable name for this ingredient because it is not specific to the ingredient (hydrolysates can be prepared from other milk proteins). The names “hydrolyzed vegetable...

  11. Preparation by enzymolysis and bioactivity of iron complex of fish protein hydrolysate (Fe-FPH) from low value fish

    NASA Astrophysics Data System (ADS)

    Deng, Shanggui; Huo, Jiancong; Xie, Chao

    2008-08-01

    Preparation of Fe2+ chelate of fish protein hydrolysate (Fe-FPH) obtained from low value fish proteins was introduced and its bioactivity was studied by compound enzymolysis. The optimum conditions for hydrolysate chelating Fe2+ are DH (degree of hydrolysis) at 5%, pH 7.0, 20°C and 15 min chelating time for FM (material not being defatted). Four types of Fe-FPH including CA (deposit after chelating), CB (deposit in 50% of absolute ethanol solution), CC (suspended deposit in 80% of absolute ethanol solution), and CD (bottom deposit in 80% of absolute ethanol solution) were fractionated with absolute ethanol from FM. Structural analysis through infra-red spectrum revealed that Fe2+ was combined strongly with amino-group and carboxyl-group in each chelate and each Fe2+ could form two five-member ring structures. All of the four chelates were shown more significant antioxidative activity and can be used as natural hydrophobic and hydrophilic antioxidant. Among all the chelates, the CB possesses the most effective antioxidative activity at 92% as high as that of a-tocopherol. Among all Fe-FPHs, only CD showed the most effective antibacterial activity against Escherichia coli, Staphylococcus aureus, Salmonella typhi, and Bacillus subtilis and can be used as natural antibacterial. It provides a more effective way for utilization of low value fish proteins and key information of Fe-FPH as additive in food industry.

  12. Preparation of ferrous chelate of hairtail (Trichiurus haumela) protein hydrolysate (Fe(II)-HPH) and its antibacterial activity

    NASA Astrophysics Data System (ADS)

    Lin, Huimin; Zhang, Bin; Yu, Tian; Deng, Shanggui

    The preparation of a ferrous chelate of hairtail (Trichiurus haumela) protein hydrolysate (Fe(II)-HPH) and its antibacterial activity were studied. The optimal conditions of hydrolysis by papain and ferrous chelation were obtained by single-factor experiments and orthogonal test, with the antibacterial activities as the index. In addition, the antibacterial activity of Fe(II)-HPH was evaluated using the Plackett-Burman design. The orthogonal test results showed that Fe(II)-HPH had an antibacterial activity of 98.3% under a temperature of 40 °C at pH 6.5 for an enzymolysis duration of eight hours in the presence of 20,000 U/g of enzyme. The Plackett-Burman design analysis showed that the three most significant factors (P < 0.05) influencing the antibacterial activity of Fe(II)-HPH were pH, the concentration (mg/mL), and presence of magnesium sulfate.

  13. Protein Hydrolysates/Peptides in Animal Nutrition

    NASA Astrophysics Data System (ADS)

    McCalla, Jeff; Waugh, Terry; Lohry, Eric

    The use of protein hydrolysates as an important nutrient for growth and maintenance has been increasing in animal nutrition. Although animal proteins and protein hydrolysates are widely used however, recently vegetable protein hydrolysates are gaining importance. This chapter reviews the use of protein hydrolysates developed by enzyme hydrolysis and by solid state fermentation process in animal nutrition especially for piglets and compares it with the standard products such as plasma and fishmeal.

  14. Effects of Soy Protein Hydrolysates Prepared by Varying Subcritical Media on the Physicochemical Properties of Pork Patties

    PubMed Central

    Davaatseren, Munkhtugs

    2016-01-01

    This study investigated the effect of soy protein hydrolysates (SPH) prepared by varying subcritical media on the physicochemical properties of pork patties. For resource of SPH, two different soybean species (Glycine max Merr.) of Daewonkong (DWK) and Saedanbaek (SDB) were selected. SPH was prepared by subcritical processing at 190℃ and 25 MPa under three different of media (water, 20% ethanol and 50% ethanol). Solubility and free amino group content revealed that water was better to yield larger amount of SPH than ethanol/water mixtures, regardless of species. Molecular weight (Mw) distribution of SPH was also similar between two species, while slightly different Mw distribution was obtained by subcritical media. For pork patty application, 50% ethanol treatment showed clear red color comparing to control after 14 d of storage. In addition, ethanol treatment had better oxidative stability than control and water treatment based on thiobarbituric acid-reactive substances (TBARS) analysis. For eating quality, although 20% ethanol treatment in SDB showed slightly higher cooking loss than control, generally addition of SPH did not affect the water-binding properties and hardness of pork patties. Consequently, the present study indicated that 50% ethanol was the best subcritical media to produce SPH possessing antioxidant activity, and the SPH produced from DWK exhibited better antioxidant activity than that produced SDB. PMID:27499657

  15. Effects of Soy Protein Hydrolysates Prepared by Varying Subcritical Media on the Physicochemical Properties of Pork Patties.

    PubMed

    Lee, Yun-Kyung; Ko, Bo-Bae; Davaatseren, Munkhtugs; Hong, Geun-Pyo

    2016-01-01

    This study investigated the effect of soy protein hydrolysates (SPH) prepared by varying subcritical media on the physicochemical properties of pork patties. For resource of SPH, two different soybean species (Glycine max Merr.) of Daewonkong (DWK) and Saedanbaek (SDB) were selected. SPH was prepared by subcritical processing at 190℃ and 25 MPa under three different of media (water, 20% ethanol and 50% ethanol). Solubility and free amino group content revealed that water was better to yield larger amount of SPH than ethanol/water mixtures, regardless of species. Molecular weight (Mw) distribution of SPH was also similar between two species, while slightly different Mw distribution was obtained by subcritical media. For pork patty application, 50% ethanol treatment showed clear red color comparing to control after 14 d of storage. In addition, ethanol treatment had better oxidative stability than control and water treatment based on thiobarbituric acid-reactive substances (TBARS) analysis. For eating quality, although 20% ethanol treatment in SDB showed slightly higher cooking loss than control, generally addition of SPH did not affect the water-binding properties and hardness of pork patties. Consequently, the present study indicated that 50% ethanol was the best subcritical media to produce SPH possessing antioxidant activity, and the SPH produced from DWK exhibited better antioxidant activity than that produced SDB. PMID:27499657

  16. Characterisation of novel fungal and bacterial protease preparations and evaluation of their ability to hydrolyse meat myofibrillar and connective tissue proteins.

    PubMed

    Ryder, Kate; Ha, Minh; Bekhit, Alaa El-Din; Carne, Alan

    2015-04-01

    The catalytic capability of four commercially available food-grade fungal and bacterial protease preparations (AFP, FPII, F60K and HT) was evaluated over a range of pH, temperature and substrate conditions using esterase and caseinolytic activity assays and time course hydrolysis over 120 and 60 min of myofibrillar and connective tissue proteins, respectively. The protease preparations displayed similar casein hydrolysis kinetics and were active in hydrolysing BODIPY-FL casein to varying extents at postmortem aging meat pH (5.0-6.0). All of the four proteases exhibited selective hydrolytic activity towards meat myofibrillar proteins including myosin and actin. Significant hydrolysis of two meat tenderisation protein markers troponin T and desmin by the four proteases was detected by western blot. The results obtained indicate that the new fungal protease preparations AFP and FPII, bacterial protease preparation HT and the new source of fungal protease preparation F60K have potential for use in meat tenderising applications.

  17. Effect of Soy Protein Hydrolysates Prepared by Subcritical Water Processing on the Physicochemical Properties of Pork Patty during Chilled Storage

    PubMed Central

    Min, Sang-Gi

    2015-01-01

    The present study was carried out to investigate the effects of soy protein hydrolysates (SPHs) addition on the quality characteristics of pork patties. The SPHs was prepared by subcritical water process (SWP) at 180℃ without holding time and mixed with the pork patty components at varying concentrations (0-3%), and the patties were stored at 4℃ for 14 d. As quality parameters, instrumental color, thiobarbituric acid-reactive substances (TBARS), pH, water holding capacity (WHC) and shear force were measured at the end of storage. Regardless of SPHs concentration, the addition of SPHs significantly manifested low L* and high a* values compared to those of untreated control (p<0.05). For b* value, addition of SPHs in the 0.5-1.5% was unaffected, while >2.0% of SPHs caused significantly lower b* than control (p<0.05). The color changes in pork patties with and without SPHs were also identified in visual appearance where the pork patties containing 0.5-2.0% showed bright red color which was comparable to brownish color of control and patties containing >2.5% SPHs. Lipid oxidation was delayed by the addition of 0.5-1.5% SPHs, while it was accelerated by the addition of 3% SPHs. The pH of patties increased with increasing concentration of SPHs, whereas there were no significant differences in WHC and shear force of patties. Consequently, the results indicated that the addition of 0.5-1.5% SPHs had a potential advantage in suppressing oxidative deterioration of fat-containing meat products during chilled storage. PMID:26761879

  18. Preparation and Evaluation of the Chelating Nanocomposite Fabricated with Marine Algae Schizochytrium sp. Protein Hydrolysate and Calcium.

    PubMed

    Lin, Jiaping; Cai, Xixi; Tang, Mengru; Wang, Shaoyun

    2015-11-11

    Marine algae have been becoming a popular research topic because of their biological implication. The algae peptide-based metal-chelating complex was investigated in this study. Schizochytrium sp. protein hydrolysate (SPH) possessing high Ca-binding capacity was prepared through stepwise enzymatic hydrolysis to a degree of hydrolysis of 22.46%. The nanocomposites of SPH chelated with calcium ions were fabricated in aqueous solution at pH 6 and 30 °C for 20 min, with the ratio of SPH to calcium 3:1 (w/w). The size distribution showed that the nanocomposite had compact structure with a radius of 68.16 ± 0.50 nm. SPH was rich in acidic amino acids, accounting for 33.55%, which are liable to bind with calcium ions. The molecular mass distribution demonstrated that the molecular mass of SPH was principally concentrated at 180-2000 Da. UV scanning spectroscopy and Fourier transform infrared spectroscopy suggested that the primary sites of calcium-binding corresponded to the carboxyl groups, carbonyl groups, and amino groups of SPH. The results of fluorescent spectroscopy, size distribution, atomic force microscope, and (1)H nuclear magnetic resonance spectroscopy suggested that calcium ions chelated with SPH would cause intramolecular and intermolecular folding and aggregating. The SPH-calcium chelate exerted remarkable stability and absorbability under either acidic or basic conditions, which was in favor of calcium absorption in the gastrointestinal tracts of humans. The investigation suggests that SPH-calcium chelate has the potential prospect to be utilized as a nutraceutical supplement to improve bone health in the human body. PMID:26499390

  19. Preparation and Evaluation of the Chelating Nanocomposite Fabricated with Marine Algae Schizochytrium sp. Protein Hydrolysate and Calcium.

    PubMed

    Lin, Jiaping; Cai, Xixi; Tang, Mengru; Wang, Shaoyun

    2015-11-11

    Marine algae have been becoming a popular research topic because of their biological implication. The algae peptide-based metal-chelating complex was investigated in this study. Schizochytrium sp. protein hydrolysate (SPH) possessing high Ca-binding capacity was prepared through stepwise enzymatic hydrolysis to a degree of hydrolysis of 22.46%. The nanocomposites of SPH chelated with calcium ions were fabricated in aqueous solution at pH 6 and 30 °C for 20 min, with the ratio of SPH to calcium 3:1 (w/w). The size distribution showed that the nanocomposite had compact structure with a radius of 68.16 ± 0.50 nm. SPH was rich in acidic amino acids, accounting for 33.55%, which are liable to bind with calcium ions. The molecular mass distribution demonstrated that the molecular mass of SPH was principally concentrated at 180-2000 Da. UV scanning spectroscopy and Fourier transform infrared spectroscopy suggested that the primary sites of calcium-binding corresponded to the carboxyl groups, carbonyl groups, and amino groups of SPH. The results of fluorescent spectroscopy, size distribution, atomic force microscope, and (1)H nuclear magnetic resonance spectroscopy suggested that calcium ions chelated with SPH would cause intramolecular and intermolecular folding and aggregating. The SPH-calcium chelate exerted remarkable stability and absorbability under either acidic or basic conditions, which was in favor of calcium absorption in the gastrointestinal tracts of humans. The investigation suggests that SPH-calcium chelate has the potential prospect to be utilized as a nutraceutical supplement to improve bone health in the human body.

  20. Protein and protein hydrolysates in sports nutrition.

    PubMed

    van Loon, Luc J C; Kies, Arie K; Saris, Wim H M

    2007-08-01

    With the increasing knowledge about the role of nutrition in increasing exercise performance, it has become clear over the last 2 decades that amino acids, protein, and protein hydrolysates can play an important role. Most of the attention has been focused on their effects at a muscular level. As these nutrients are ingested, however, it also means that gastrointestinal digestibility and absorption can modulate their efficacy significantly. Therefore, discussing the role of amino acids, protein, and protein hydrolysates in sports nutrition entails holding a discussion on all levels of the metabolic route. On May 28-29, 2007, a small group of researchers active in the field of exercise science and protein metabolism presented an overview of the different aspects of the application of protein and protein hydrolysates in sports nutrition. In addition, they were asked to share their opinions on the future progress in their fields of research. In this overview, an introduction to the workshop and a short summary of its outcome is provided.

  1. Applications of Protein Hydrolysates in Biotechnology

    NASA Astrophysics Data System (ADS)

    Pasupuleti, Vijai K.; Holmes, Chris; Demain, Arnold L.

    By definition, protein hydrolysates are the products that are obtained after the hydrolysis of proteins and this can be achieved by enzymes, acid or alkali. This broad definition encompasses all the products of protein hydrolysis - peptides, amino acids and minerals present in the protein and acid/alkali used to adjust pH (Pasupuleti 2006). Protein hydrolysates contain variable side chains depending on the enzymes used. These side chains could be carboxyl, amino, imidazole, sulfhydryl, etc. and they may exert specific physiological roles in animal, microbial, insect and plant cells. This introductory chapter reviews the applications of protein hydrolysates in biotechnology. The word biotechnology is so broad and for the purpose of this book, we define it as a set of technologies such as cell culture technology, bioprocessing technology that includes fermentations, genetic engineering technology, microbiology, and so on. This chapter provides introduction and leads to other chapters on manufacturing and applications of protein hydrolysates in biotechnology.

  2. Immunogenicity evaluation of protein hydrolysates for hypoallergenic infant formulae.

    PubMed

    Cordle, C T; Mahmoud, M I; Moore, V

    1991-10-01

    Casein and soy protein were enzymatically hydrolyzed for potential use in a hypoallergenic infant formula. To assess the relative immunoreactivity of the hydrolysates, rabbits were immunized with either the intact proteins or the protein hydrolysates using a vigorous immunization protocol. Serum samples were tested using ELISA methods that quantitated IgG antibody specific for the immunizing protein hydrolysates and the corresponding intact proteins. The results showed that the protein hydrolysates had substantially lower immunogenicity than the parent proteins. Also, antibody specific for the parent protein showed very low cross-reactivity with the hydrolysates. Both of the protein hydrolysates seem to be promising candidates for use in hypoallergenic infant feeding systems.

  3. Characterisation of novel fungal and bacterial protease preparations and evaluation of their ability to hydrolyse meat myofibrillar and connective tissue proteins.

    PubMed

    Ryder, Kate; Ha, Minh; Bekhit, Alaa El-Din; Carne, Alan

    2015-04-01

    The catalytic capability of four commercially available food-grade fungal and bacterial protease preparations (AFP, FPII, F60K and HT) was evaluated over a range of pH, temperature and substrate conditions using esterase and caseinolytic activity assays and time course hydrolysis over 120 and 60 min of myofibrillar and connective tissue proteins, respectively. The protease preparations displayed similar casein hydrolysis kinetics and were active in hydrolysing BODIPY-FL casein to varying extents at postmortem aging meat pH (5.0-6.0). All of the four proteases exhibited selective hydrolytic activity towards meat myofibrillar proteins including myosin and actin. Significant hydrolysis of two meat tenderisation protein markers troponin T and desmin by the four proteases was detected by western blot. The results obtained indicate that the new fungal protease preparations AFP and FPII, bacterial protease preparation HT and the new source of fungal protease preparation F60K have potential for use in meat tenderising applications. PMID:25442543

  4. Production of enzymatic protein hydrolysates from freshwater catfish (Clarias batrachus)

    NASA Astrophysics Data System (ADS)

    Seniman, Maizatul Sarah Md; Yusop, Salma Mohamad; Babji, Abdul Salam

    2014-09-01

    Fish protein hydrolysate (FPH) was prepared from freshwater catfish (Clarias batrachus) by using Alcalase® 2.4L and Papain. The effect of hydrolysis time (30, 60, 120, 180 min) with enzyme concentration of 1% (v/w substrate); pH = 8.0, 7.0 was studied to determine the degree of hydrolysis (DH), peptide content, proximate composition and amino acid profile. Results showed that the highest DH of Alcalase and Papain FPH were 58.79% and 53.48% after 180 min at 55°C incubation respectively. The peptide content of both FPH increased as hydrolysis time increases. FPH showed higher crude protein content and lower fat, moisture and ash content compared to raw catfish. The major amino acids of both hydrolysates were Glu, Lys and Asp. Content of essential amino acids of Alcalase and Papain hydrolysates were 44.05% and 43.31% respectively.

  5. State of the Art Manufacturing of Protein Hydrolysates

    NASA Astrophysics Data System (ADS)

    Pasupuleti, Vijai K.; Braun, Steven

    The use of protein hydrolysates in microbiological media has been in existence for several decades and the basic manufacturing process of protein hydrolysates has remained the same. However, with increasing use of protein hydrolysates in specialized applications such as animal cell culture processes, the manufacturing of protein hydrolysates has dramatically improved and is still in its infancy to uncover the specific peptide, peptides and combination of individual amino acids that produce intended effects for that application. This will change as the protein hydrolysate manufacturers and end-users exchange information and work towards the common goal of developing the best protein hydrolysates for specific applications. This chapter will review the generic manufacturing of protein hydrolysates describing individual unit operations, problems faced by manufacturers and suggestions for obtaining consistent product and guidelines for the end-users in getting regulatory support and setting up reliable specifications. Finally the chapter concludes with future trends of protein hydrolysates.

  6. Stability of encapsulated beef-like flavourings prepared from enzymatically hydrolysed mushroom proteins with other precursors under conventional and microwave heating.

    PubMed

    Lotfy, Shereen N; Fadel, Hoda H M; El-Ghorab, Ahmed H; Shaheen, Mohamed S

    2015-11-15

    A comparative study was carried out between two beef-like flavourings prepared by conventional and microwave heating (CBF and MBF) of enzymatic hydrolysate of mushroom protein with other flavour precursors. GC-MS analysis of the isolated volatiles revealed that the thiol containing compounds were the predominate in both samples. However, MBF comprised higher concentration of these compounds (13.84 ± 0.06%) than CBF (10.74 ± 0.06%). The effect of microencapsulation with gum Arabic by using spray drying on the odour profile and volatile compounds of the two encapsulated samples (E-CBF and E-MBF) was investigated. The results revealed significant qualitative and quantitative variations in the volatiles of both samples. The highly volatile compounds decreased remarkably in concentration with encapsulation, while the pyrazines, thiazoles and disulphides showed opposite trend. The significant decrease in the thiol containing compounds in E-CBF and E-MBF were attributed to their oxidation to other compounds such as disulphide compounds which showed significant increase in the encapsulated samples.

  7. Stability of encapsulated beef-like flavourings prepared from enzymatically hydrolysed mushroom proteins with other precursors under conventional and microwave heating.

    PubMed

    Lotfy, Shereen N; Fadel, Hoda H M; El-Ghorab, Ahmed H; Shaheen, Mohamed S

    2015-11-15

    A comparative study was carried out between two beef-like flavourings prepared by conventional and microwave heating (CBF and MBF) of enzymatic hydrolysate of mushroom protein with other flavour precursors. GC-MS analysis of the isolated volatiles revealed that the thiol containing compounds were the predominate in both samples. However, MBF comprised higher concentration of these compounds (13.84 ± 0.06%) than CBF (10.74 ± 0.06%). The effect of microencapsulation with gum Arabic by using spray drying on the odour profile and volatile compounds of the two encapsulated samples (E-CBF and E-MBF) was investigated. The results revealed significant qualitative and quantitative variations in the volatiles of both samples. The highly volatile compounds decreased remarkably in concentration with encapsulation, while the pyrazines, thiazoles and disulphides showed opposite trend. The significant decrease in the thiol containing compounds in E-CBF and E-MBF were attributed to their oxidation to other compounds such as disulphide compounds which showed significant increase in the encapsulated samples. PMID:25976991

  8. Affinity purification of copper chelating peptides from chickpea protein hydrolysates.

    PubMed

    Megías, Cristina; Pedroche, Justo; Yust, Maria M; Girón-Calle, Julio; Alaiz, Manuel; Millan, Francisco; Vioque, Javier

    2007-05-16

    Chickpea protein hydrolysates obtained with alcalase and flavourzyme were used for purification of copper chelating peptides by affinity chromatography using copper immobilized on solid supports. The chelating activity of purified peptides was indirectly measured by the inhibition of beta-carotene oxidation in the presence of copper. Two protein hydrolysates, obtained after 10 and 100 min of hydrolysis, were the most inhibitory of beta-carotene oxidation. Purified copper chelating peptides from these protein hydrolysates contained 19.7 and 35.1% histidine, respectively, in comparison to 2.7 and 2.6% in the protein hydrolysates. Chelating peptides from hydrolysate obtained after 10 min of hydrolysis were the most antioxidative being 8.3 times more antioxidative than the hydrolysate, while chelating peptides purified from protein hydrolysate obtained after 100 min were 3.1 times more antioxidative than its hydrolysate. However, the histidine content was higher in peptides derived from the 100 min hydrolysate (19.7 against 35.1% in 10 min hydrolysate), indicating that this amino acid is not the only factor involved in the antioxidative activity, and other factors such as peptide size or amino acid sequence are also determinant. This manuscript shows that affinity chromatography is a useful procedure for purification of copper chelating peptides. This method can be extended to other metals of interest in nutrition, such as calcium, iron, or zinc. Purified chelating peptides, in addition to their antioxidative properties, may also be useful in food mineral fortification for increasing the bioavailability of these metals.

  9. 21 CFR 573.200 - Condensed animal protein hydrolysate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Condensed animal protein hydrolysate. 573.200... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.200 Condensed animal protein hydrolysate. (a) Identity. The...

  10. 21 CFR 573.200 - Condensed animal protein hydrolysate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Condensed animal protein hydrolysate. 573.200... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.200 Condensed animal protein hydrolysate. (a) Identity. The...

  11. 21 CFR 573.200 - Condensed animal protein hydrolysate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Condensed animal protein hydrolysate. 573.200... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.200 Condensed animal protein hydrolysate. (a) Identity. The...

  12. 21 CFR 573.200 - Condensed animal protein hydrolysate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Condensed animal protein hydrolysate. 573.200... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.200 Condensed animal protein hydrolysate. (a) Identity. The...

  13. 21 CFR 573.200 - Condensed animal protein hydrolysate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Condensed animal protein hydrolysate. 573.200... (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.200 Condensed animal protein hydrolysate. (a) Identity. The...

  14. Antioxidant activities of protein hydrolysates obtained from the housefly larvae.

    PubMed

    Zhang, Huan; Wang, Pan; Zhang, Ai-Jun; Li, Xuan; Zhang, Ji-Hong; Qin, Qi-Lian; Wu, Yi-Jun

    2016-09-01

    The housefly is an important resource insect and the housefly larvae are ideal source of food additives. The housefly larvae protein hydrolysates were obtained by enzymatic hydrolysis by alcalase and neutral proteinase. Their antioxidant activities were investigated, including the superoxide and hydroxyl radicalscavenging activity, 1,1-diphenyl-2-picrylhydrazyl (DPPH) scavenging activity, reducing power and metal chelating activity. The antioxidant activities of both hydrolysates increased with their increasing concentrations. The alcalase hydrolysate (AH) showed higher scavenging activities against hydroxyl radical and superoxide anion radical at low concentrations and higher metal-chelating activity than the neutral proteinase hydrolysate (NPH). The NPH exhibited higher scavenging activity against DPPH free radical and higher reducing power than the AH. Both hydrolysates showed more than 50% superoxide anion radical-scavenging activity at 10 μg/mL. These results indicate that both housefly larvae protein hydrolysates display high antioxidant activities and they could serve as potential natural antioxidant food additives. PMID:27630047

  15. Storage Stability of Food Protein Hydrolysates-A Review.

    PubMed

    Rao, Qinchun; Klaassen Kamdar, Andre; Labuza, Theodore P

    2016-05-18

    In recent years, mainly due to the specific health benefits associated with (1) the discovery of bioactive peptides in protein hydrolysates, (2) the reduction of protein allergenicity by protein hydrolysis, and (3) the improved protein digestibility and absorption of protein hydrolysates, the utilization of protein hydrolysates in functional foods and beverages has significantly increased. Although the specific health benefits from different hydrolysates are somewhat proven, the delivery and/or stability of these benefits is debatable during distribution, storage, and consumption. In this review, we discuss (1) the quality changes in different food protein hydrolysates during storage; (2) the resulting changes in the structure and texture of three food matrices, i.e., low moisture foods (LMF, aw < 0.6), intermediate moisture foods (IMF, 0.6 ≤ aw < 0.85), and high moisture foods (HMF, aw ≥ 0.85); and (3) the potential solutions to improve storage stability of food protein hydrolysates. In addition, we note there is a great need for evaluation of biofunction availability of bioactive peptides in food protein hydrolysates during storage.

  16. [Preparation and nutritional characteristics of a hydrolysate from pepitona (Arca zebra)].

    PubMed

    Arbej, J; Luna, G

    1985-12-01

    Two soluble products resulting from the hydrolysis of pepitona (Arca zebra) were prepared as flour. Papain at its optimum hydrolysis conditions, previously established, was the enzyme used (40 degrees C for two hours at a pH of 7 in the proportion of 0.3% weight/enzyme/100 g meat). The hydrolysate obtained was then subjected to two different dehydration techniques: drum drying at 121 degrees C and 18 seconds retention, and spray drying at 101 degrees C and 40 psi pressure. The products were then stored for a five-month period at a temperature of 25 degrees C +/- 2 degrees C, time during which chemical determinations were performed in both hydrolysates. Findings showed that the time of storage does exert a significant effect of deterioration on the products. The greater and more significant quality losses occur during the first two months. The dehydration techniques used also affect significantly the soluble nitrogen content, and non-protein nitrogen and soluble solids content, as well as color of pepitona hydrolysates. Spray-drying dehydration technique does not have a significant deteriorating effect. Biological studies undertaken demonstrated that the quality of both hydrolysates is satisfactory from the nutritional and amino acid composition points of view. A protein efficiency ratio (PER) of 2.27 and 2.29 was determined for the hydrolysate dehydrated by drum drier and for the dehydrated by spray drier, respectively. With regard to amino acid composition, both had satisfactory levels of essential amino acids, with a lysine content of 6.9 g/100 g protein for the hydrolysate dehydrated by drum drying, and 8.6 g/100 g protein for the other hydrolysate dehydrated by spray drying. PMID:3842922

  17. Production of bioactive peptide hydrolysates from deer, sheep and pig plasma using plant and fungal protease preparations.

    PubMed

    Bah, Clara S F; Bekhit, Alaa El-Din A; Carne, Alan; McConnell, Michelle A

    2015-06-01

    Plasma separated from deer, sheep and pig blood, obtained from abattoirs, was hydrolysed using protease preparations from plant (papain and bromelain) and fungal (FP400 and FPII) sources. Antioxidant and antimicrobial activities of the peptide hydrolysates obtained after 1, 2, 4 and 24h of hydrolysis, were investigated. The release of trichloroacetic acid-soluble peptides over the hydrolysis period was monitored using the o-phthaldialdehyde (OPA) assay, while the hydrolysis profiles were visualised using SDS-PAGE. The major plasma proteins in the animal plasmas were identified using MALDI-TOF-TOF MS. Hydrolysates of plasma generated with fungal proteases exhibited higher DPPH radical-scavenging, oxygen radical-scavenging capacity (ORAC) and ferric reducing antioxidant power (FRAP) than those generated with plant proteases for all three animal plasmas. No antimicrobial activity was detected in the hydrolysates. The results indicated that proteolytic hydrolysis of animal blood plasmas, using fungal protease preparations in particular, produces hydrolysates with high antioxidant properties.

  18. Production of bioactive peptide hydrolysates from deer, sheep and pig plasma using plant and fungal protease preparations.

    PubMed

    Bah, Clara S F; Bekhit, Alaa El-Din A; Carne, Alan; McConnell, Michelle A

    2015-06-01

    Plasma separated from deer, sheep and pig blood, obtained from abattoirs, was hydrolysed using protease preparations from plant (papain and bromelain) and fungal (FP400 and FPII) sources. Antioxidant and antimicrobial activities of the peptide hydrolysates obtained after 1, 2, 4 and 24h of hydrolysis, were investigated. The release of trichloroacetic acid-soluble peptides over the hydrolysis period was monitored using the o-phthaldialdehyde (OPA) assay, while the hydrolysis profiles were visualised using SDS-PAGE. The major plasma proteins in the animal plasmas were identified using MALDI-TOF-TOF MS. Hydrolysates of plasma generated with fungal proteases exhibited higher DPPH radical-scavenging, oxygen radical-scavenging capacity (ORAC) and ferric reducing antioxidant power (FRAP) than those generated with plant proteases for all three animal plasmas. No antimicrobial activity was detected in the hydrolysates. The results indicated that proteolytic hydrolysis of animal blood plasmas, using fungal protease preparations in particular, produces hydrolysates with high antioxidant properties. PMID:25624206

  19. Physiological Importance and Mechanisms of Protein Hydrolysate Absorption

    NASA Astrophysics Data System (ADS)

    Zhanghi, Brian M.; Matthews, James C.

    Understanding opportunities to maximize the efficient digestion and assimilation by production animals of plant- and animal-derived protein products is critical for farmers, nutritionists, and feed manufacturers to sustain and expand the affordable production of high quality animal products for human consumption. The challenge to nutritionists is to match gastrointestinal tract load to existing or ­inducible digestive and absorptive capacities. The challenge to feed manufacturers is to develop products that are efficient substrates for digestion, absorption, and/or both events. Ultimately, the efficient absorption of digesta proteins depends on the mediated passage (transport) of protein hydrosylate products as dipeptides and unbound amino acids across the lumen- and blood-facing membranes of intestinal absorptive cells. Data testing the relative efficiency of supplying protein as hydrolysates or specific dipeptides versus as free amino acids, and the response of animals in several physiological states to feeding of protein hydrolysates, are presented and reviewed in this chapter. Next, data describing the transport mechanisms responsible for absorbing protein hydrolysate digestion products, and the known and putative regulation of these mechanisms by their substrates (small peptides) and hormones are presented and reviewed. Several conclusions are drawn regarding the efficient use of protein hydrolysate-based diets for particular physiological states, the economically-practical application of which likely will depend on technological advances in the manufacture of protein hydrolysate products.

  20. Physiological properties of Scomber japonicus meat hydrolysate prepared by subcritical water hydrolysis.

    PubMed

    Choi, Jae-Suk; Moon, Hye Eun; Roh, Myong-Kyun; Ha, Yu-Mi; Lee, Bo-Bae; Cho, Kwang Keun; Choi, In Soon

    2016-01-01

    The health-beneficial biological activities, including antioxidant and tyrosinase inhibitory activities, of Scomber japonicus muscle protein hydrolysates prepared by subcritical water hydrolysis were investigated. After 5 min of subcritical hydrolysis at 140 degrees C, 59.76% of S. japonicus muscle protein was hydrolyzed, the highest degree of hydrolysis in all the groups were tested. According to the response surface methodology results, as the reaction temperature and reaction time became lower and shorter, the yield became higher. The highest 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging activity (90.63%) occurred in hydrolysates treated at 140 degrees C for 5 min, and the highest tyrosinase inhibitory activity (65.54%) was identified in hydrolysates treated at 200 degreesC for 15 min. Changes in the molecular weight distribution of S. japonicus muscle proteins after subcritical water hydrolysis were observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Subcritical water hydrolysis is a suitable technique for obtaining S.japonicus muscle protein hydrolysates with useful biological activities, within a short time (5-15 min).

  1. Preparation of gluten free bread enriched with green mussel (Perna canaliculus) protein hydrolysates and characterization of peptides responsible for mussel flavour.

    PubMed

    Vijaykrishnaraj, M; Roopa, B S; Prabhasankar, P

    2016-11-15

    Green mussel protein hydrolysates (GMPH) utilization for the enrichment of gluten-free bread followed by characterization of flavour peptides using chromatography and electronic nose techniques have been done. The degree of hydrolysis was carried out in each protease digest, and the higher degree of hydrolysis was observed in pepsin digestion. Gluten-free (GF) bread was formulated by using buckwheat flour (BWF), rice flour (RF) and chickpea flour (CPF) (70:20:10) and GMPH were added in the range of 0-20% in the GF bread for enrichment with GMPH. Radar plot of the electronic nose analysis showed that the sensors P30/2, T30/1 and T70/2 had a higher response to the GF bread and GMPH. Consequently, the peptide sequence was obtained manually by ESI-MS spectra of GMPH (KGYSSYICDK) and F-II (SSYCIVKICDK). Flavour quality was 97% discriminately comparable to the GMPH and F-II fractions. Mussel flavoured GF bread can be included in the celiac diet. PMID:27283688

  2. Effect of whey and casein protein hydrolysates on rheological, textural and sensory properties of cookies.

    PubMed

    Gani, Adil; Broadway, A A; Ahmad, Mudasir; Ashwar, Bilal Ahmad; Wani, Ali Abas; Wani, Sajad Mohd; Masoodi, F A; Khatkar, Bupinder Singh

    2015-09-01

    Milk proteins were hydrolyzed by papain and their effect on the rheological, textural and sensory properties of cookies were investigated. Water absorption (%) decreased significantly as the amount of milk protein concentrates and hydrolysates increased up to a level of 15 % in the wheat flour. Dough extensibility decreased with inrease in parental proteins and their hydrolysates in wheat flour, significantly. Similarly, the pasting properties also varied significantly in direct proportion to the quantity added in the wheat flour. The colour difference (ΔE) of cookies supplemented with milk protein concentrates and hydrolysates were significantly higher than cookies prepared from control. Physical and sensory characteristics of cookies at 5 % level of supplementation were found to be acceptable. Also the scores assigned by the judges for texture and colour were in good agreement with the measurements derived from the physical tests.

  3. [The use of chitosan solutions for defatting and clarification of protein hydrolysates].

    PubMed

    Novikov, Iu V; Mukhin, V A

    2001-01-01

    The possibility of the use of small amounts of chitosan for defatting and clarification of protein solutions prepared by enzymatic hydrolysis was tested. The following treatment conditions were shown to be optimal: chitosan concentration range, from 1.0 to 1.5 g per kg raw weight; pH of precipitation medium, from 8.0 to 8.5; and duration of incubation of protein hydrolysate solution with chitosan, less than 1 h. The hydrolysate defatting grade was found to depend on the degree of chitosan deacetylation. A possible mechanism of the chitosan-induced effects was suggested. The use of chitosan allows the mass fraction of enzyme protein hydrolysates to be reduced fourfold to fivefold.

  4. Chickpea protein hydrolysate as a substitute for serum in cell culture

    PubMed Central

    Vioque, Javier; Pedroche, Justo; Alaiz, Manuel; Yust, María M.; Megías, Cristina; Millán, Francisco

    2008-01-01

    The growth of mammalian cells in vitro requires the use of rich culture media that are prepared by combining serum with specific nutrient formulations. Serum, the most expensive component of culture media, provides a complex mixture of growth factors and nutrients. Protein hydrolysates that can support in vitro cell growth and eliminate or reduce the need to use serum have been obtained from different sources. Here we describe the use of two food grade proteases to produce a chickpea protein hydrolysate that has been added to cell culture medium in order to determine whether it can be used as a substitute for serum. Medium containing the hydrolysate has been tested using two human cells lines: the monocytic THP-1 cell line which grows in suspension, and the epithelial Caco-2 cell line which grows as a monolayer. The chickpea protein hydrolysate was a good substitute for serum in the first case, but did not allow growth of Caco-2 cells. Supplementation of culture media with this inexpensive and safe hydrolysate would greatly reduce the cost of cell culture. PMID:19003183

  5. Protein Hydrolysates as Hypoallergenic, Flavors and Palatants for Companion Animals

    NASA Astrophysics Data System (ADS)

    Nagodawithana, Tilak W.; Nelles, Lynn; Trivedi, Nayan B.

    Early civilizations have relied upon their good sense and experience to develop and improve their food quality. The discovery of soy sauce centuries ago can now be considered one of the earliest protein hydrolysates made by man to improve palatability of foods. Now, it is well known that such savory systems are not just sources for enjoyment but complex semiotic systems that direct the humans to satisfy the body's protein need for their sustenance. Recent developments have resulted in a wide range of cost effective savory flavorings, the best known of which are autolyzed yeast extracts and hydrolyzed vegetable proteins. New technologies have helped researchers to improve the savory characteristics of yeast extracts through the application of Maillard reaction and by generating specific flavor enhancers through the use of enzymes. An interesting parallel exists in the pet food industry, where a similar approach is taken in using animal protein hydrolysates to create palatability enhancers via Maillard reaction scheme. Protein hydrolysates are also utilized extensively as a source of nutrition to the elderly, young children and immuno-compromised patient population. These hydrolysates have an added advantage in having peptides small enough to avoid any chance of an allergenic reaction which sometimes occur with the consumption of larger sized peptides or proteins. Accordingly, protein hydrolysates are required to have an average molecular weight distribution in the range 800-1,500 Da to make them non-allergenic. The technical challenge for scientists involved in food and feed manufacture is to use an appropriate combination of enzymes within the existing economic constraints and other physical factors/limitations, such as heat, pH, and time, to create highly palatable, yet still nutritious and hypoallergenic food formulations.

  6. Rendered-protein hydrolysates for microbial synthesis of cyanophycin biopolymer

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cyanophycin is a poly(arginyl-aspartate) biopolymer produced and stored intracellularly by bacteria. Cyanophycin has been proposed as a renewable replacement for petrochemical-based industrial products. An abundant source of amino acids and nitrogen such as in the form of protein hydrolysates is n...

  7. Towards an Understanding of How Protein Hydrolysates Stimulate More Efficient Biosynthesis in Cultured Cells

    NASA Astrophysics Data System (ADS)

    Siemensma, André; Babcock, James; Wilcox, Chris; Huttinga, Hans

    In the light of the growing demand for high quality plant-derived hydrolysates (i.e., HyPep™ and UltraPep™ series), Sheffield Bio-Science has developed a new hydrolysate platform that addresses the need for animal-free cell culture medium supplements while also minimizing variability concerns. The platform is based upon a novel approach to enzymatic digestion and more refined processing. At the heart of the platform is a rationally designed animal component-free (ACF) enzyme cocktail that includes both proteases and non-proteolytic enzymes (hydrolases) whose activities can also liberate primary components of the polymerized non-protein portion of the raw material. This enzyme system is added during a highly optimized process step that targets specific enzyme-substrate reactions to expand the range of beneficial nutritional factors made available to cells in culture. Such factors are fundamental to improving the bio-performance of the culture system, as they provide not merely growth-promoting peptides and amino acids, but also key carbohydrates, lipids, minerals, and vitamins that improve both rate and quality of protein expression, and serve to improve culture life due to osmo-protectant and anti-apoptotic properties. Also of significant note is that, compared to typical hydrolysates, the production process is greatly reduced and requires fewer steps, intrinsically yielding a better-controlled and therefore more reproducible product. Finally, the more sophisticated approach to enzymatic digestion renders hydrolysates more amenable to sterile filtration, allowing hydrolysate end users to experience streamlined media preparation and bioreactor supplementation activities. Current and future development activities will evolve from a better understanding of the complex interactions within a handful of key biochemical pathways that impact the growth and productivity of industrially relevant organisms. Presented in this chapter are some examples of the efforts that

  8. Black bean (Phaseolus vulgaris L.) protein hydrolysates: Physicochemical and functional properties.

    PubMed

    Evangelho, Jarine Amaral do; Vanier, Nathan Levien; Pinto, Vânia Zanella; Berrios, Jose J De; Dias, Alvaro Renato Guerra; Zavareze, Elessandra da Rosa

    2017-01-01

    Black bean protein hydrolysates obtained from pepsin and alcalase digestions until 120min of hydrolysis were evaluated by gel electrophoresis, relative fluorescence intensity, emulsifying properties, light micrograph of emulsions and in vitro antioxidant activity. The emulsion stability of the bean protein hydrolysates were evaluated during 30days of storage. The pepsin-treated bean protein hydrolysates presented higher degree of hydrolysis than the alcalase-treated protein hydrolysates. The alcalase-treated bean protein hydrolysates showed higher surface hydrophobicity. Moreover, the protein hydrolysates obtained with alcalase digestion presented higher emulsion stability during 30-days than those obtained from pepsin digestion. The protein concentrate and especially the hydrolysates obtained from alcalase digestion had good emulsion stability and antioxidant activity. Thus, they could be exploited as protein supplements in the diet as nutritional and bioactive foods. PMID:27507499

  9. Physical and oxidative stability of fish oil-in-water emulsions stabilized with fish protein hydrolysates.

    PubMed

    García-Moreno, Pedro J; Guadix, Antonio; Guadix, Emilia M; Jacobsen, Charlotte

    2016-07-15

    The emulsifying and antioxidant properties of fish protein hydrolysates (FPH) for the physical and oxidative stabilization of 5% (by weight) fish oil-in-water emulsions were investigated. Muscle proteins from sardine (Sardina pilchardus) and small-spotted catshark (Scyliorhinus canicula) were hydrolyzed to degrees of hydrolysis (DH) of 3-4-5-6% with subtilisin. Sardine hydrolysates with low DH, 3% and 4%, presented the most effective peptides to physically stabilize emulsions with smaller droplet size. This implied more protein adsorbed at the interface to act as physical barrier against prooxidants. This fact might also be responsible for the higher oxidative stability of these emulsions, as shown by their lowest peroxide value and concentration of volatiles such as 1-penten-3-one and 1-penten-3-ol. Among the hydrolysates prepared from small-spotted catshark only the hydrolysate with DH 3% yielded a physically stable emulsion with low concentration of unsaturated aldehydes. These results show the potential of FPH as alternative protein emulsifiers for the production of oxidatively stable fish oil-in-water emulsions.

  10. Supplementation of Pork Patties with Bovine Plasma Protein Hydrolysates Augments Antioxidant Properties and Improves Quality

    PubMed Central

    Seo, Hyun-Woo

    2016-01-01

    This study investigated the effects of bovine plasma protein (PP) hydrolysates on the antioxidant and quality properties of pork patties during storage. Pork patties were divided into 4 groups: without butylated hydroxytoluene (BHT) and PP hydrolysates (control), 0.02% BHT (T1), 1% PP hydrolysates (T2), and 2% PP hydrolysates (T3). Pork patty supplemented with PP hydrolysates had higher pH values and lower weight loss during cooking than the control patties. Results showed that lightness and hardness both decreased upon the addition of PP hydrolysates. All samples containing BHT and PP hydrolysates had reduced TBARS and peroxide values during storage. In particular, 2% PP hydrolysates were more effective in delaying lipid oxidation than were the other treatments. It was concluded that treatment with 2% PP hydrolysates can enhance the acceptance of pork patty. PMID:27194928

  11. Protein hydrolysates and associated bacterial contaminants as oviposition attractants for the mosquito Culex quinquefasciatus.

    PubMed

    Beehler, J W; Millar, J G; Mulla, M S

    1994-10-01

    Six protein or protein hydrolysate solutions were tested for activity as attractants for ovipositing Culex quinquefasciatus mosquitoes in the laboratory. Four of these solutions (egg albumin, lactalbumin hydrolysate, casein hydrolysate and yeast hydrolysate) were attractive to ovipositing females at varying concentrations, when compared to distilled water controls. Soy hydrolysate was repellent at 1%, but not significantly attractive or repellent at lower concentrations. 'Nulure', a tephritid fly bait containing protein hydrolysate, also had no significant effect on oviposition behaviour. Gravid females mostly oviposited within the first 4 h of the scotophase, regardless of the presence or absence of an oviposition attractant. Lactalbumin hydrolysate 1% solution, with or without 0.1% neomycin antibiotic, was attractive to Cx quinquefasciatus. This effect was reduced by the presence of neomycin which, alone, had no effect on oviposition. Hence both lactalbumin hydrolysate and bacterial contaminants were shown to be attractive to gravid Cx quinquefasciatus.

  12. Use of Protein Hydrolysates in Industrial Starter Culture Fermentations

    NASA Astrophysics Data System (ADS)

    Ummadi, Madhavi (Soni); Curic-Bawden, Mirjana

    Lactic acid bacteria (LAB) have been used as starter cultures for fermenting foods long before the importance of microorganisms were recognized. The most important group of LAB are the lactococci, lactobacilli, streptococci, and pediococci. Additionally, bifidobacteria have been included as a probiotic, providing added value to the product. Since the genera involved are so diverse, the nutritional requirements (energy, carbon and nitrogen sources) differ significantly between and within species. Designing an optimum fermentation medium for production of active and vigorous LAB starter cultures and probiotics requires selecting the right raw ingredients, especially protein hydrolysates that can provide adequate nutrients for growth and viability. This chapter attempts to describe the application of various commercial protein hydrolysates used for production of dairy and meat starter cultures, with special emphasis on meeting the nitrogen requirements of industrially important LAB species.

  13. Generation, Fractionation, and Characterization of Iron-Chelating Protein Hydrolysate from Palm Kernel Cake Proteins.

    PubMed

    Zarei, Mohammad; Ghanbari, Rahele; Tajabadi, Naser; Abdul-Hamid, Azizah; Bakar, Fatimah Abu; Saari, Nazamid

    2016-02-01

    Palm kernel cake protein was hydrolyzed with different proteases namely papain, bromelain, subtilisin, flavourzyme, trypsin, chymotrypsin, and pepsin to generate different protein hydrolysates. Peptide content and iron-chelating activity of each hydrolysate were evaluated using O-phthaldialdehyde-based spectrophotometric method and ferrozine-based colorimetric assay, respectively. The results revealed a positive correlation between peptide contents and iron-chelating activities of the protein hydrolysates. Protein hydrolysate generated by papain exhibited the highest peptide content of 10.5 mM and highest iron-chelating activity of 64.8% compared with the other hydrolysates. Profiling of the papain-generated hydrolysate by reverse phase high performance liquid chromatography fractionation indicated a direct association between peptide content and iron-chelating activity in most of the fractions. Further fractionation using isoelectric focusing also revealed that protein hydrolysate with basic and neutral isoelectric point (pI) had the highest iron-chelating activity, although a few fractions in the acidic range also exhibited good metal chelating potential. After identification and synthesis of papain-generated peptides, GGIF and YLLLK showed among the highest iron-chelating activities of 56% and 53%, whereas their IC50 were 1.4 and 0.2 μM, respectively.

  14. The Use of Protein Hydrolysates for Weed Control

    NASA Astrophysics Data System (ADS)

    Christians, Nick; Liu, Dianna; Unruh, Jay Bryan

    Corn gluten meal, the protein fraction of corn (Zea mays L.) grain, is commercially used as a natural weed control agent and nitrogen source in horticultural crops and in the turf and ornamental markets. Corn gluten hydrolysate, a water soluble form of gluten meal, has also been proposed for the same purpose, although it could be sprayed on the soil rather than applied in the granular form. Five depeptides, glutaminyl-glutamine (Gln-Gln), glycinyl-alanine (Gly-Ala), alanyl-­glutamine (Ala-Glu), alanyl-asparagine (Ala-Asp), and alaninyl-alanine (Ala-Ala) and a pentapeptide leucine-serine-proline-alanine-glutamine (Leu-Ser-Pro-Ala-Gln) were identified as the active components of the hydrolysate. Microscopic analysis revealed that Ala-Ala acted on some metabolic process rather than directly on the mitotic apparatus. Similar to the chloracetamides and sulfonyl-urea hebicides, Ala-Ala inhibits cell division rather than disrupting of cell division processes. Cellular ultrastructure changes caused by exposure to Ala-Ala implicate Ala-Ala as having membrane-disrupting characteristics similar to several synthetic herbicides. The potential use of the hydrolysate and the peptides as weed controls is discussed.

  15. Antioxidant Effect and Functional Properties of Hydrolysates Derived from Egg-White Protein

    PubMed Central

    Cho, Dae-Yeon; Jo, Kyungae; Cho, So Young; Kim, Jin Man; Lim, Kwangsei; Suh, Hyung Joo

    2014-01-01

    This study utilized commercially available proteolytic enzymes to prepare egg-white protein hydrolysates (EPHs) with different degrees of hydrolysis. The antioxidant effect and functionalities of the resultant products were then investigated. Treatment with Neutrase yielded the most α-amino groups (6.52 mg/mL). Alcalase, Flavourzyme, Protamex, and Ficin showed similar degrees of α-amino group liberation (3.19-3.62 mg/mL). Neutrase treatment also resulted in the highest degree of hydrolysis (23.4%). Alcalase and Ficin treatment resulted in similar degrees of hydrolysis. All hydrolysates, except for the Flavourzyme hydrolysate, had greater radical scavenging activity than the control. The Neutrase hydrolysate showed the highest 2,2-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging activity (IC50=3.6mg/mL). Therefore, Neutrase was identified as the optimal enzyme for hydrolyzing egg-white protein to yield antioxidant peptides. During Neutrase hydrolysis, the reaction rate was rapid over the first 4 h, and then subsequently declined. The IC50 value was lowest after the first hour (2.99 mg/mL). The emulsifying activity index (EAI) of EPH treated with Neutrase decreased, as the pH decreased. The EPH foaming capacity was maximal at pH 3.6, and decreased at an alkaline pH. Digestion resulted in significantly higher 1,1-diphenyl-2-picrylhydrazyl (DPPH) and ABTS radical scavenging activity. The active peptides released from egg-white protein showed antioxidative activities on ABTS and DHHP radical. Thus, this approach may be useful for the preparation of potent antioxidant products. PMID:26761178

  16. Structural and Antihypertensive Properties of Enzymatic Hemp Seed Protein Hydrolysates.

    PubMed

    Malomo, Sunday A; Onuh, John O; Girgih, Abraham T; Aluko, Rotimi E

    2015-09-01

    The aim of this work was to produce antihypertensive protein hydrolysates through different forms of enzymatic hydrolysis (2% pepsin, 4% pepsin, 1% alcalase, 2% alcalase, 2% papain, and 2% pepsin + pancreatin) of hemp seed proteins (HSP). The hemp seed protein hydrolysates (HPHs) were tested for in vitro inhibitions of renin and angiotensin-converting enzyme (ACE), two of the enzymes that regulate human blood pressure. The HPHs were then administered orally (200 mg/kg body weight) to spontaneously hypertensive rats and systolic blood pressure (SBP)-lowering effects measured over a 24 h period. Size exclusion chromatography mainly showed a 300-9560 Da peptide size range for the HPHs, while amino acid composition data had the 2% pepsin HPH with the highest cysteine content. Fluorescence spectroscopy revealed higher fluorescence intensities for the peptides when compared to the unhydrolyzed hemp seed protein. Overall, the 1% alcalase HPH was the most effective (p < 0.05) SBP-reducing agent (-32.5 ± 0.7 mmHg after 4 h), while the pepsin HPHs produced longer-lasting effects (-23.0 ± 1.4 mmHg after 24 h). We conclude that an optimized combination of the fast-acting HPH (1% alcalase) with the longer-lasting HPHs (2% and 4% pepsin) could provide daily effective SBP reductions. PMID:26378569

  17. Structural and Antihypertensive Properties of Enzymatic Hemp Seed Protein Hydrolysates

    PubMed Central

    Malomo, Sunday A.; Onuh, John O.; Girgih, Abraham T.; Aluko, Rotimi E.

    2015-01-01

    The aim of this work was to produce antihypertensive protein hydrolysates through different forms of enzymatic hydrolysis (2% pepsin, 4% pepsin, 1% alcalase, 2% alcalase, 2% papain, and 2% pepsin + pancreatin) of hemp seed proteins (HSP). The hemp seed protein hydrolysates (HPHs) were tested for in vitro inhibitions of renin and angiotensin-converting enzyme (ACE), two of the enzymes that regulate human blood pressure. The HPHs were then administered orally (200 mg/kg body weight) to spontaneously hypertensive rats and systolic blood pressure (SBP)-lowering effects measured over a 24 h period. Size exclusion chromatography mainly showed a 300–9560 Da peptide size range for the HPHs, while amino acid composition data had the 2% pepsin HPH with the highest cysteine content. Fluorescence spectroscopy revealed higher fluorescence intensities for the peptides when compared to the unhydrolyzed hemp seed protein. Overall, the 1% alcalase HPH was the most effective (p < 0.05) SBP-reducing agent (−32.5 ± 0.7 mmHg after 4 h), while the pepsin HPHs produced longer-lasting effects (−23.0 ± 1.4 mmHg after 24 h). We conclude that an optimized combination of the fast-acting HPH (1% alcalase) with the longer-lasting HPHs (2% and 4% pepsin) could provide daily effective SBP reductions. PMID:26378569

  18. Structural and Antihypertensive Properties of Enzymatic Hemp Seed Protein Hydrolysates.

    PubMed

    Malomo, Sunday A; Onuh, John O; Girgih, Abraham T; Aluko, Rotimi E

    2015-09-10

    The aim of this work was to produce antihypertensive protein hydrolysates through different forms of enzymatic hydrolysis (2% pepsin, 4% pepsin, 1% alcalase, 2% alcalase, 2% papain, and 2% pepsin + pancreatin) of hemp seed proteins (HSP). The hemp seed protein hydrolysates (HPHs) were tested for in vitro inhibitions of renin and angiotensin-converting enzyme (ACE), two of the enzymes that regulate human blood pressure. The HPHs were then administered orally (200 mg/kg body weight) to spontaneously hypertensive rats and systolic blood pressure (SBP)-lowering effects measured over a 24 h period. Size exclusion chromatography mainly showed a 300-9560 Da peptide size range for the HPHs, while amino acid composition data had the 2% pepsin HPH with the highest cysteine content. Fluorescence spectroscopy revealed higher fluorescence intensities for the peptides when compared to the unhydrolyzed hemp seed protein. Overall, the 1% alcalase HPH was the most effective (p < 0.05) SBP-reducing agent (-32.5 ± 0.7 mmHg after 4 h), while the pepsin HPHs produced longer-lasting effects (-23.0 ± 1.4 mmHg after 24 h). We conclude that an optimized combination of the fast-acting HPH (1% alcalase) with the longer-lasting HPHs (2% and 4% pepsin) could provide daily effective SBP reductions.

  19. Mild protein hydrolysation of lactose-free milk further reduces milk-related gastrointestinal symptoms.

    PubMed

    Turpeinen, Anu; Kautiainen, Hanna; Tikkanen, Marja-Leena; Sibakov, Timo; Tossavainen, Olli; Myllyluoma, Eveliina

    2016-05-01

    Gastrointestinal symptoms associated with milk are common. Besides lactose, milk proteins may cause symptoms in sensitive individuals. We have developed a method for mild enzymatic hydrolysation of milk proteins and studied the effects of hydrolysed milk on gastrointestinal symptoms in adults with a self-diagnosed sensitive stomach. In a double blind, randomised placebo-controlled study, 97 subjects consumed protein-hydrolysed lactose-free milk or commercially available lactose-free milk for 10 d. Frequency of gastrointestinal symptoms during the study period was reported and a symptom score was calculated. Rumbling and flatulence decreased significantly in the hydrolysed milk group (P < 0·05). Also, the total symptom score was lower in subjects who consumed hydrolysed milk (P < 0·05). No difference between groups was seen in abdominal pain (P = 0·47) or bloating (P = 0·076). The results suggest that mild enzymatic protein hydrolysation may decrease gastrointestinal symptoms in adults with a sensitive stomach.

  20. Characterization and Potential Use of Cuttlefish Skin Gelatin Hydrolysates Prepared by Different Microbial Proteases

    PubMed Central

    Jridi, Mourad; Lassoued, Imen; Nasri, Rim; Ayadi, Mohamed Ali; Nasri, Moncef

    2014-01-01

    Composition, functional properties, and in vitro antioxidant activities of gelatin hydrolysates prepared from cuttlefish skin were investigated. Cuttlefish skin gelatin hydrolysates (CSGHs) were obtained by treatment with crude enzyme preparations from Bacillus licheniformis NH1, Bacillus mojavensis A21, Bacillus subtilis A26, and commercial alcalase. All CSGHs had high protein contents, 74.3–78.3%, and showed excellent solubility (over 90%). CSGH obtained by alcalase demonstrated high antioxidant activities monitored by β-carotene bleaching, DPPH radical scavenging, lipid peroxidation inhibition, and reducing power activity. Its antioxidant activity remained stable or increased in a wide range of pH (1–9), during heating treatment (100°C for 240 min) and after gastrointestinal digestion simulation. In addition, alcalase-CSGH was incorporated into turkey meat sausage to determine its effect on lipid oxidation during 35 days of storage period. At 0.5 mg/g, alcalase-CSGH delayed lipid oxidation monitored by TBARS and conjugated diene up to 10 days compared to vitamin C. The results reveal that CSGHs could be used as food additives possessing both antioxidant activity and functional properties. PMID:25025053

  1. Preparation of Egg White Liquid Hydrolysate (ELH) and Its Radical-Scavenging Activity

    PubMed Central

    Noh, Dong Ouk; Suh, Hyung Joo

    2015-01-01

    In the present study, an optimum protease was selected to hydrolyze the egg white liquid protein for the antioxidant peptides. Alcalase treatment yielded the highest amount of α-amino groups (15.27 mg/mL), while the control (no enzymatic hydrolysis) showed the lowest amount of α-amino groups (1.53 mg/mL). Alcalase also gave the highest degree of hydrolysis (DH) value (43.2%) and was more efficient for egg white liquid hydrolysis than the other enzymes. The Alcalase hydrolysate had the highest radical-scavenging activity (82.5%) at a concentration of 5.0 mg/mL. The conditions for enzymatic hydrolysis of egg white liquid with Alcalase were selected as substrate : water ratio of 2:1. Five percent Alacalse treatment did not show significant (P>0.05) increases of DH and α-amino nitrogen content after 24 h-hydrolysis. Thirty two hour-hydrolysis with 5% Alcalase is sufficient to make antioxidative egg white liquid hydrolysate from egg white liquid. DPPH and ABTS radical-scavenging activities were significantly (P<0.05) higher after enzymatic digestion. These results suggest that active peptides released from egg-white protein are effective radical-scavengers. Thus, this approach may be useful for the preparation of potent antioxidant products. PMID:26451355

  2. Purification and identification of antioxidant peptides from walnut (Juglans regia L.) protein hydrolysates.

    PubMed

    Chen, Ning; Yang, Hongmei; Sun, Yi; Niu, Jun; Liu, Shuying

    2012-12-01

    Walnut proteins were hydrolyzed separately using three different proteases to obtain antioxidant peptides. The antioxidant activities of the hydrolysates were measured using 1,1-diphenyl-2-picryl hydrazyl (DPPH) assay. Among hydrolysates, pepsin hydrolysate obtained by 3h exhibited the highest antioxidant activities, which could also quench the hydroxyl radical, chelate ferrous ion, exhibit reducing power and inhibit the lipid peroxidation. Then, 3-h pepsin hydrolysates were purified sequentially by ultrafiltration, gel filtration and RP-HPLC. The sequence of the peptide with the highest antioxidative activity was identified to be Ala-Asp-Ala-Phe (423.23 Da) using RP-HPLC-ESI-MS, which was identified for the first time from walnut protein hydrolysates. Last, the inhibition of the peptide on lipid peroxidation was similar with that of reduced glutathione (GSH). These results indicate that the protein hydrolysates and/or its isolated peptides may be effectively used as food additives.

  3. A novel multi-biofunctional protein from brown rice hydrolysed by endo/endo-exoproteases.

    PubMed

    Selamassakul, Orrapun; Laohakunjit, Natta; Kerdchoechuen, Orapin; Ratanakhanokchai, Khanok

    2016-06-15

    Brown rice, which is a less allergenic food grain and contains essential amino acids, was hydrolysed by bromelain and PROTEASE FP51® to improve its functionalities and taste for food applications. The hydrolysate prepared by bromelain (eb-RPH) had high protein solubility, surface hydrophobicity, low molecular weight peptides, hydrophobic amino acids (leucine, valine and glycine) and flavor amino acids (glutamic acid and aspartic acid). The eb-RPH exhibited higher 1,1-diphenyl-2-picrylhydrazyl (DPPH˙) and 2,2'-azino-bis 3-ethylbenzthiazoline-6-sulfonic (ABTS˙(+)) radical-scavenging activities of 76.62% and 52.96%, respectively, and possessed a better foaming capacity (221.76%) and emulsifying capacity (32.34%) than the hydrolysate prepared by PROTEASE FP51® (ep-RPH) did. The eb-RPH gave the desired taste, which is attributed to volatile flavor compounds (benzaldehyde, benzeneacetaldehyde and 2-acetyl-1-pyrroline) and non-volatile flavor compounds, such as monosodium glutamate, 5'-guanosine monophosphate and 5'-inosine monophosphate (0.07, 0.03 and 0.05 mg mL(-1), respectively). Brown rice peptides generated by bromelain were novel bioactive peptides with multifunctional properties. PMID:27186602

  4. Production of Defatted Palm Kernel Cake Protein Hydrolysate as a Valuable Source of Natural Antioxidants

    PubMed Central

    Zarei, Mohammad; Ebrahimpour, Afshin; Abdul-Hamid, Azizah; Anwar, Farooq; Saari, Nazamid

    2012-01-01

    The aim of this study was to produce a valuable protein hydrolysate from palm kernel cake (PKC) for the development of natural antioxidants. Extracted PKC protein was hydrolyzed using different proteases (alcalase, chymotrypsin, papain, pepsin, trypsin, flavourzyme, and bromelain). Subsequently, antioxidant activity and degree of hydrolysis (DH) of each hydrolysate were evaluated using DPPH• radical scavenging activity and O-phthaldialdehyde spectrophotometric assay, respectively. The results revealed a strong correlation between DH and radical scavenging activity of the hydrolysates, where among these, protein hydrolysates produced by papain after 38 h hydrolysis exhibited the highest DH (91 ± 0.1%) and DPPH• radical scavenging activity (73.5 ± 0.25%) compared to the other hydrolysates. In addition, fractionation of the most effective (potent) hydrolysate by reverse phase high performance liquid chromatography indicated a direct association between hydrophobicity and radical scavenging activity of the hydrolysates. Isoelectric focusing tests also revealed that protein hydrolysates with basic and neutral isoelectric point (pI) have the highest radical scavenging activity, although few fractions in the acidic range also exhibited good antioxidant potential. PMID:22942692

  5. Antioxidant and sensory properties of protein hydrolysate derived from Nile tilapia (Oreochromis niloticus) by one- and two-step hydrolysis.

    PubMed

    Yarnpakdee, Suthasinee; Benjakul, Soottawat; Kristinsson, Hordur G; Kishimura, Hideki

    2015-06-01

    Antioxidant and sensory properties of Nile tilapia protein hydrolysates prepared by one- and two-step hydrolysis using commercial proteases were investigated. Hydrolysates prepared using single protease including Alcalase (HA), Flavourzyme (HF), Protamex (HPr) and papain (HPa) had increases in antioxidant activities as the degree of hydrolysis (DH) increased up to 40 % (P < 0.05). Amongst all hydrolysates, HA having 40 % DH showed the highest antioxidant activities. When HA was further hydrolysed by papain, the resulting hydrolysate (HAPa) exhibited the highest antioxidant activities for all assays tested (P < 0.05). ABTS radical scavenging activity and metal chelating of HAPa generally remained constant in a wide pH range (1-11) and during heating at 30-100 °C. Both activities increased in the simulated gastrointestinal tract model system, especially in intestine condition. HAPa (100-1,000 ppm) could retard lipid oxidation in β-carotene-linoleate and lecithin-liposome model systems in a dose dependent manner. Peptides in both HA and HAPa with molecular weight of 513 Da and 1,484 Da possessed the strongest ABTS radical scavenging activity and metal chelating activity, respectively. The amino acid profile of both HA and HAPa contained a high amount of hydrophobic amino acids (38.26-38.85 %) and had glutamic acid/glutamine, lysine and aspartic acid/asparagine as the dominant amino acids. However, HAPa showed a higher acceptability than did HA, owing to the lower bitterness. Therefore, the use of Alcalase in combination with papain for hydrolysis of protein isolate rendered the hydrolysate with antioxidant properties and reduced bitterness, which could serve as the functional supplement. PMID:26028714

  6. Improved Functional Characteristics of Whey Protein Hydrolysates in Food Industry.

    PubMed

    Jeewanthi, Renda Kankanamge Chaturika; Lee, Na-Kyoung; Paik, Hyun-Dong

    2015-01-01

    This review focuses on the enhanced functional characteristics of enzymatic hydrolysates of whey proteins (WPHs) in food applications compared to intact whey proteins (WPs). WPs are applied in foods as whey protein concentrates (WPCs), whey protein isolates (WPIs), and WPHs. WPs are byproducts of cheese production, used in a wide range of food applications due to their nutritional validity, functional activities, and cost effectiveness. Enzymatic hydrolysis yields improved functional and nutritional benefits in contrast to heat denaturation or native applications. WPHs improve solubility over a wide range of pH, create viscosity through water binding, and promote cohesion, adhesion, and elasticity. WPHs form stronger but more flexible edible films than WPC or WPI. WPHs enhance emulsification, bind fat, and facilitate whipping, compared to intact WPs. Extensive hydrolyzed WPHs with proper heat applications are the best emulsifiers and addition of polysaccharides improves the emulsification ability of WPHs. Also, WPHs improve the sensorial properties like color, flavor, and texture but impart a bitter taste in case where extensive hydrolysis (degree of hydrolysis greater than 8%). It is important to consider the type of enzyme, hydrolysis conditions, and WPHs production method based on the nature of food application. PMID:26761849

  7. Improved Functional Characteristics of Whey Protein Hydrolysates in Food Industry.

    PubMed

    Jeewanthi, Renda Kankanamge Chaturika; Lee, Na-Kyoung; Paik, Hyun-Dong

    2015-01-01

    This review focuses on the enhanced functional characteristics of enzymatic hydrolysates of whey proteins (WPHs) in food applications compared to intact whey proteins (WPs). WPs are applied in foods as whey protein concentrates (WPCs), whey protein isolates (WPIs), and WPHs. WPs are byproducts of cheese production, used in a wide range of food applications due to their nutritional validity, functional activities, and cost effectiveness. Enzymatic hydrolysis yields improved functional and nutritional benefits in contrast to heat denaturation or native applications. WPHs improve solubility over a wide range of pH, create viscosity through water binding, and promote cohesion, adhesion, and elasticity. WPHs form stronger but more flexible edible films than WPC or WPI. WPHs enhance emulsification, bind fat, and facilitate whipping, compared to intact WPs. Extensive hydrolyzed WPHs with proper heat applications are the best emulsifiers and addition of polysaccharides improves the emulsification ability of WPHs. Also, WPHs improve the sensorial properties like color, flavor, and texture but impart a bitter taste in case where extensive hydrolysis (degree of hydrolysis greater than 8%). It is important to consider the type of enzyme, hydrolysis conditions, and WPHs production method based on the nature of food application.

  8. Improved Functional Characteristics of Whey Protein Hydrolysates in Food Industry

    PubMed Central

    Jeewanthi, Renda Kankanamge Chaturika; Lee, Na-Kyoung; Paik, Hyun-Dong

    2015-01-01

    This review focuses on the enhanced functional characteristics of enzymatic hydrolysates of whey proteins (WPHs) in food applications compared to intact whey proteins (WPs). WPs are applied in foods as whey protein concentrates (WPCs), whey protein isolates (WPIs), and WPHs. WPs are byproducts of cheese production, used in a wide range of food applications due to their nutritional validity, functional activities, and cost effectiveness. Enzymatic hydrolysis yields improved functional and nutritional benefits in contrast to heat denaturation or native applications. WPHs improve solubility over a wide range of pH, create viscosity through water binding, and promote cohesion, adhesion, and elasticity. WPHs form stronger but more flexible edible films than WPC or WPI. WPHs enhance emulsification, bind fat, and facilitate whipping, compared to intact WPs. Extensive hydrolyzed WPHs with proper heat applications are the best emulsifiers and addition of polysaccharides improves the emulsification ability of WPHs. Also, WPHs improve the sensorial properties like color, flavor, and texture but impart a bitter taste in case where extensive hydrolysis (degree of hydrolysis greater than 8%). It is important to consider the type of enzyme, hydrolysis conditions, and WPHs production method based on the nature of food application. PMID:26761849

  9. Chemical compositions and muddy flavour/odour of protein hydrolysate from Nile tilapia and broadhead catfish mince and protein isolate.

    PubMed

    Yarnpakdee, Suthasinee; Benjakul, Soottawat; Penjamras, Pimpimol; Kristinsson, Hordur G

    2014-01-01

    Chemical compositions and muddy compounds in dorsal and ventral muscles of Nile tilapia and broadhead catfish were comparatively studied. On a dry weight basis, Nile tilapia was rich in protein (93.1-93.8%), whilst broadhead catfish contained protein (55.2-59.5%) and lipid (36.6-42.4%) as the major constituents. Ventral portion had higher lipid or phospholipid contents with coincidentally higher geosmin and/or 2-methylisoborneol (2-MIB) contents. Geosmin was found in mince of Nile tilapia and broadhead catfish at levels of 1.5 and 3.2μg/kg, respectively. Broadhead catfish mince had 2-MIB at level of 0.8μg/kg, but no 2-MIB was detected in Nile tilapia counterpart. When pre-washing and alkaline solubilisation were applied for preparing protein isolate (PI), lipid and phospholipid contents were lowered with concomitant decrease in geosmin and 2-MIB contents. Protein hydrolysate produced from PI had a lighter colour and a lower amount of muddy compounds, compared with that prepared from mince. Therefore, PI from both Nile tilapia and broadhead catfish could serve as the promising proteinaceous material, yielding protein hydrolysate with the negligible muddy odour and flavour. PMID:24001833

  10. Use of Different Proteases to Obtain Flaxseed Protein Hydrolysates with Antioxidant Activity.

    PubMed

    Karamać, Magdalena; Kosińska-Cagnazzo, Agnieszka; Kulczyk, Anna

    2016-01-01

    The antioxidant activity of flaxseed protein hydrolysates obtained using five different enzymes was evaluated. Proteins were isolated from flaxseed cake and were separately treated with papain, trypsin, pancreatin, Alcalase and Flavourzyme. The degree of hydrolysis (DH) was determined as the percentage of cleaved peptide bonds using a spectrophotometric method with o-phthaldialdehyde. The distribution of the molecular weights (MW) of the hydrolysis products was profiled using Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Tricine-SDS-PAGE) and size exclusion-high performance liquid chromatography (SE-HPLC) separations. The antioxidant activities of the protein isolate and hydrolysates were probed for their radical scavenging activity using 2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonate) radical cation (ABTS(•+)) and photochemiluminescence (PCL-ACL) assays, and for their ferric reducing antioxidant power (FRAP) and ability to bind Fe(2+). The hydrolysates were more effective as antioxidants than the protein isolate in all systems. The PCL-ACL values of the hydrolysates ranged from 7.2 to 35.7 μmol Trolox/g. Both the FRAP and ABTS(•+) scavenging activity differed among the hydrolysates to a lower extent, with the ranges of 0.20-0.24 mmol Fe(2+)/g and 0.17-0.22 mmol Trolox/g, respectively. The highest chelating activity (71.5%) was noted for the pancreatin hydrolysate. In general, the hydrolysates obtained using Alcalase and pancreatin had the highest antioxidant activity, even though their DH (15.4% and 29.3%, respectively) and the MW profiles of the peptides varied substantially. The O₂(•-) scavenging activity and the ability to chelate Fe(2+) of the Flavourzyme hydrolysate were lower than those of the Alcalase and pancreatin hydrolysates. Papain was the least effective in releasing the peptides with antioxidant activity. The study showed that the type of enzyme used for flaxseed protein hydrolysis determines the antioxidant activity

  11. Use of Different Proteases to Obtain Flaxseed Protein Hydrolysates with Antioxidant Activity

    PubMed Central

    Karamać, Magdalena; Kosińska-Cagnazzo, Agnieszka; Kulczyk, Anna

    2016-01-01

    The antioxidant activity of flaxseed protein hydrolysates obtained using five different enzymes was evaluated. Proteins were isolated from flaxseed cake and were separately treated with papain, trypsin, pancreatin, Alcalase and Flavourzyme. The degree of hydrolysis (DH) was determined as the percentage of cleaved peptide bonds using a spectrophotometric method with o-phthaldialdehyde. The distribution of the molecular weights (MW) of the hydrolysis products was profiled using Tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Tricine-SDS-PAGE) and size exclusion-high performance liquid chromatography (SE-HPLC) separations. The antioxidant activities of the protein isolate and hydrolysates were probed for their radical scavenging activity using 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonate) radical cation (ABTS•+) and photochemiluminescence (PCL-ACL) assays, and for their ferric reducing antioxidant power (FRAP) and ability to bind Fe2+. The hydrolysates were more effective as antioxidants than the protein isolate in all systems. The PCL-ACL values of the hydrolysates ranged from 7.2 to 35.7 μmol Trolox/g. Both the FRAP and ABTS•+ scavenging activity differed among the hydrolysates to a lower extent, with the ranges of 0.20–0.24 mmol Fe2+/g and 0.17–0.22 mmol Trolox/g, respectively. The highest chelating activity (71.5%) was noted for the pancreatin hydrolysate. In general, the hydrolysates obtained using Alcalase and pancreatin had the highest antioxidant activity, even though their DH (15.4% and 29.3%, respectively) and the MW profiles of the peptides varied substantially. The O2•− scavenging activity and the ability to chelate Fe2+ of the Flavourzyme hydrolysate were lower than those of the Alcalase and pancreatin hydrolysates. Papain was the least effective in releasing the peptides with antioxidant activity. The study showed that the type of enzyme used for flaxseed protein hydrolysis determines the antioxidant activity of

  12. Whey Protein Concentrate Hydrolysate Prevents Bone Loss in Ovariectomized Rats.

    PubMed

    Kim, Jonggun; Kim, Hyung Kwan; Kim, Saehun; Imm, Ji-Young; Whang, Kwang-Youn

    2015-12-01

    Milk is known as a safe food and contains easily absorbable minerals and proteins, including whey protein, which has demonstrated antiosteoporotic effects on ovariectomized rats. This study evaluated the antiosteoporotic effect of whey protein concentrate hydrolysate (WPCH) digested with fungal protease and whey protein concentrate (WPC). Two experiments were conducted to determine (1) efficacy of WPCH and WPC and (2) dose-dependent impact of WPCH in ovariectomized rats (10 weeks old). In Experiment I, ovariectomized rats (n=45) were allotted into three dietary treatments of 10 g/kg diet of WPC, 10 g/kg diet of WPCH, and a control diet. In Experiment II, ovariectomized rats (n=60) were fed four different diets (0, 10, 20, and 40 g/kg of WPCH). In both experiments, sham-operated rats (n=15) were also fed a control diet containing the same amount of amino acids and minerals as dietary treatments. After 6 weeks, dietary WPCH prevented loss of bone, physical properties, mineral density, and mineral content, and improved breaking strength of femurs, with similar effect to WPC. The bone resorption enzyme activity (tartrate resistance acid phosphatase) in tibia epiphysis decreased in response to WPCH supplementation, while bone formation enzyme activity (alkaline phosphatase) was unaffected by ovariectomy and dietary treatment. Bone properties and strength increased as the dietary WPCH level increased (10 and 20 g/kg), but there was no difference between the 20 and 40 g/kg treatment. WPCH and WPC supplementation ameliorated bone loss induced by ovariectomy in rats. PMID:26367331

  13. Whey Protein Concentrate Hydrolysate Prevents Bone Loss in Ovariectomized Rats.

    PubMed

    Kim, Jonggun; Kim, Hyung Kwan; Kim, Saehun; Imm, Ji-Young; Whang, Kwang-Youn

    2015-12-01

    Milk is known as a safe food and contains easily absorbable minerals and proteins, including whey protein, which has demonstrated antiosteoporotic effects on ovariectomized rats. This study evaluated the antiosteoporotic effect of whey protein concentrate hydrolysate (WPCH) digested with fungal protease and whey protein concentrate (WPC). Two experiments were conducted to determine (1) efficacy of WPCH and WPC and (2) dose-dependent impact of WPCH in ovariectomized rats (10 weeks old). In Experiment I, ovariectomized rats (n=45) were allotted into three dietary treatments of 10 g/kg diet of WPC, 10 g/kg diet of WPCH, and a control diet. In Experiment II, ovariectomized rats (n=60) were fed four different diets (0, 10, 20, and 40 g/kg of WPCH). In both experiments, sham-operated rats (n=15) were also fed a control diet containing the same amount of amino acids and minerals as dietary treatments. After 6 weeks, dietary WPCH prevented loss of bone, physical properties, mineral density, and mineral content, and improved breaking strength of femurs, with similar effect to WPC. The bone resorption enzyme activity (tartrate resistance acid phosphatase) in tibia epiphysis decreased in response to WPCH supplementation, while bone formation enzyme activity (alkaline phosphatase) was unaffected by ovariectomy and dietary treatment. Bone properties and strength increased as the dietary WPCH level increased (10 and 20 g/kg), but there was no difference between the 20 and 40 g/kg treatment. WPCH and WPC supplementation ameliorated bone loss induced by ovariectomy in rats.

  14. Photosensitizing properties of protein hydrolysate-based fertilizers.

    PubMed

    Cavani, Luciano; Ter Halle, Alexandra; Richard, Claire; Ciavatta, Claudio

    2006-11-29

    The use of protein hydrolysate-based fertilizers (PHF) as adjuvant for pesticides or herbicides has been proposed; however, the behaviors of mixtures of PHFs and pesticides under solar light are not known, and various photochemical reactions may occur. The photosensitizing properties of PHFs were investigated in water solutions (0.8 g of total organic carbon L(-1)) within the wavelength range of 300-450 nm, using furfuryl alcohol (FA) as a probe to test the involvement of singlet oxygen and Irgarol 1051 as an example of organic pollutant. Two commercial PHFs and one standard PHF were studied, all of the products being of animal origin. PHFs photosensitize the transformation of FA (10(-4) M), and the kinetics of FA disappearance follows an apparent first-order rate law. Through the use of sodium azide (1 x 10(-3) M) as singlet oxygen scavenger and deuterium oxide (D2O) for increasing the singlet oxygen lifetime it was shown that singlet oxygen contributes largely to the phototransformation of FA. The replacement of water by D2O increases the apparent first-order rate constant 6 times, whereas the addition of sodium azide reduces it by approximately 90%. These results are confirmed using Irgarol 1051 (10(-5) M). The photosensitizing properties of PHFs might be due to pigments naturally present in tissues from which they are extracted or to compounds generated during the production processes. PMID:17117805

  15. Antioxidant activities of bambara groundnut (Vigna subterranea) protein hydrolysates and their membrane ultrafiltration fractions.

    PubMed

    Arise, Abimbola K; Alashi, Adeola M; Nwachukwu, Ifeanyi D; Ijabadeniyi, Oluwatosin A; Aluko, Rotimi E; Amonsou, Eric O

    2016-05-18

    In this study, the bambara protein isolate (BPI) was digested with three proteases (alcalase, trypsin and pepsin), to produce bambara protein hydrolysates (BPHs). These hydrolysates were passed through ultrafiltration membranes to obtain peptide fractions of different sizes (<1, 1-3, 3-5 and 5-10 kDa). The hydrolysates and their peptide fractions were investigated for antioxidant activities. The membrane fractions showed that peptides with sizes <3 kDa had significantly (p < 0.05) reduced surface hydrophobicity when compared with peptides >3 kDa. This is in agreement with the result obtained for the ferric reducing power, metal chelating and hydroxyl radical scavenging activities where higher molecular weight peptides exhibited better activity (p < 0.05) when compared to low molecular weight peptide fractions. However, for all the hydrolysates, the low molecular weight peptides were more effective diphenyl-1-picrylhydrazyl (DPPH) radical scavengers but not superoxide radicals when compared to the bigger peptides. In comparison with glutathione (GSH), BPHs and their membrane fractions had better (p < 0.05) reducing power and ability to chelate metal ions except for the pepsin hydrolysate and its membrane fractions that did not show any metal chelating activity. However, the 5-10 kDa pepsin hydrolysate peptide fractions had greater (88%) hydroxyl scavenging activity than GSH, alcalase and trypsin hydrolysates (82%). These findings show the potential use of BPHs and their peptide fraction as antioxidants in reducing food spoilage or management of oxidative stress-related metabolic disorders. PMID:27156453

  16. Comparison of the aggregation behavior of soy and bovine whey protein hydrolysates.

    PubMed

    Kuipers, Bas J H; Alting, Arno C; Gruppen, Harry

    2007-01-01

    Soy-derived proteins (soy protein isolate, glycinin, and beta-conglycinin) and bovine whey-derived proteins (whey protein isolate, alpha-lactalbumin, beta-lactoglobulin) were hydrolyzed using subtilisin Carlsberg, chymotrypsin, trypsin, bromelain, and papain. The (in)solubility of the hydrolysates obtained was studied as a function of pH. At neutral pH, all soy-derived protein hydrolysates, particularly those from glycinin, obtained by hydrolysis with subtilisin Carlsberg, chymotrypsin, bromelain, and papain showed a stronger aggregation compared to the non-hydrolyzed ones. This increase in aggregation was not observed upon hydrolysis by trypsin. None of the whey-derived protein hydrolysates exhibited an increase in aggregation at neutral pH. The high abundance of theoretical cleavage sites in the hydrophobic regions of glycinin probably explains the stronger exposure of hydrophobic groups than for the other proteins, which is suggested to be the driving force in the aggregate formation.

  17. Enzymatic protein hydrolysates from high pressure-pretreated isolated pea proteins have better antioxidant properties than similar hydrolysates produced from heat pretreatment.

    PubMed

    Girgih, Abraham T; Chao, Dongfang; Lin, Lin; He, Rong; Jung, Stephanie; Aluko, Rotimi E

    2015-12-01

    Isolated pea protein (IPP) dispersions (1%, w/v) were pretreated with high pressure (HP) of 200, 400, or 600 MPa for 5 min at 24 °C or high temperature (HT) for 30 min at 100 °C prior to hydrolysis with 1% (w/w) Alcalase. HP pretreatment of IPP at 400 and 600 MPa levels led to significantly (P<0.05) improved (>40%) oxygen radical absorption capacity (ORAC) of hydrolysates. 2,2-Diphenyl-1-picrylhydrazyl, superoxide radical and hydroxyl radical scavenging activities of pea protein hydrolysates were also significantly (P<0.05) improved (25%, 20%, and 40%, respectively) by HP pretreatment of IPP. Protein hydrolysates from HT IPP showed no ORAC, superoxide or hydroxyl scavenging activity but had significantly (P<0.05) improved (80%) ferric reducing antioxidant power. The protein hydrolysates had weaker antioxidant properties than glutathione but overall, the HP pretreatment was superior to HT pretreatment in facilitating enzymatic release of antioxidant peptides from IPP.

  18. Antioxidant activities of red tilapia (Oreochromis niloticus) protein hydrolysates as influenced by thermolysin and alcalase

    NASA Astrophysics Data System (ADS)

    Daud, Nur'Aliah; Babji, Abdul Salam; Yusop, Salma Mohamad

    2013-11-01

    The hydrolysis process was performed on fish meat from Red Tilapia (Oreochromis niloticus) by enzymes thermolysin and alcalase under optimum conditions. The hydrolysis was performed from 0 - 4 hours at 37°C. Hydrolysates after 2 hours incubation with thermolysin and alcalase had degree of hydrolysis of 76.29 % and 63.49 %, respectively. The freeze dried protein hydrolysate was tested for peptide content and characterized with respect to amino acid composition. The result of increased peptide content in Red Tilapia (O. Niloticus) hydrolysates obtained was directly proportional to the increase activities of different proteolytic enzymes. The result of amino acid composition showed that the sample used contained abundant Gly, Ala, Asp, Glu, Lys and Leu in residues or peptide sequences. Both enzymatic hydrolysates were tested for anti-oxidant activity with DPPH and ABTS assay. Alcalase yielded higher anti-oxidative activity than Thermolysin hydrolysates after 1 hour incubation, but both enzymes hydrolysates showed a significant decrease of anti-oxidant activity after 2 hours of incubation. Hydrolysates from Red Tilapia may contribute as a health promoting ingredient in functional foods to reduce oxidation stress caused by accumulated free radicals.

  19. Biostimulant action of a plant-derived protein hydrolysate produced through enzymatic hydrolysis.

    PubMed

    Colla, Giuseppe; Rouphael, Youssef; Canaguier, Renaud; Svecova, Eva; Cardarelli, Mariateresa

    2014-01-01

    The aim of this study was to evaluate the biostimulant action (hormone like activity, nitrogen uptake, and growth stimulation) of a plant-derived protein hydrolysate by means of two laboratory bioassays: a corn (Zea mays L.) coleoptile elongation rate test (Experiment 1), a rooting test on tomato cuttings (Experiment 2); and two greenhouse experiments: a dwarf pea (Pisum sativum L.) growth test (Experiment 3), and a tomato (Solanum lycopersicum L.) nitrogen uptake trial (Experiment 4). Protein hydrolysate treatments of corn caused an increase in coleoptile elongation rate when compared to the control, in a dose-dependent fashion, with no significant differences between the concentrations 0.75, 1.5, and 3.0 ml/L, and inodole-3-acetic acid treatment. The auxin-like effect of the protein hydrolysate on corn has been also observed in the rooting experiment of tomato cuttings. The shoot, root dry weight, root length, and root area were significantly higher by 21, 35, 24, and 26%, respectively, in tomato treated plants with the protein hydrolysate at 6 ml/L than untreated plants. In Experiment 3, the application of the protein hydrolysate at all doses (0.375, 0.75, 1.5, and 3.0 ml/L) significantly increased the shoot length of the gibberellin-deficient dwarf pea plants by an average value of 33% in comparison with the control treatment. Increasing the concentration of the protein hydrolysate from 0 to 10 ml/L increased the total dry biomass, SPAD index, and leaf nitrogen content by 20.5, 15, and 21.5%, respectively. Thus the application of plant-derived protein hydrolysate containing amino acids and small peptides elicited a hormone-like activity, enhanced nitrogen uptake and consequently crop performances. PMID:25250039

  20. Biostimulant action of a plant-derived protein hydrolysate produced through enzymatic hydrolysis

    PubMed Central

    Colla, Giuseppe; Rouphael, Youssef; Canaguier, Renaud; Svecova, Eva; Cardarelli, Mariateresa

    2014-01-01

    The aim of this study was to evaluate the biostimulant action (hormone like activity, nitrogen uptake, and growth stimulation) of a plant-derived protein hydrolysate by means of two laboratory bioassays: a corn (Zea mays L.) coleoptile elongation rate test (Experiment 1), a rooting test on tomato cuttings (Experiment 2); and two greenhouse experiments: a dwarf pea (Pisum sativum L.) growth test (Experiment 3), and a tomato (Solanum lycopersicum L.) nitrogen uptake trial (Experiment 4). Protein hydrolysate treatments of corn caused an increase in coleoptile elongation rate when compared to the control, in a dose-dependent fashion, with no significant differences between the concentrations 0.75, 1.5, and 3.0 ml/L, and inodole-3-acetic acid treatment. The auxin-like effect of the protein hydrolysate on corn has been also observed in the rooting experiment of tomato cuttings. The shoot, root dry weight, root length, and root area were significantly higher by 21, 35, 24, and 26%, respectively, in tomato treated plants with the protein hydrolysate at 6 ml/L than untreated plants. In Experiment 3, the application of the protein hydrolysate at all doses (0.375, 0.75, 1.5, and 3.0 ml/L) significantly increased the shoot length of the gibberellin-deficient dwarf pea plants by an average value of 33% in comparison with the control treatment. Increasing the concentration of the protein hydrolysate from 0 to 10 ml/L increased the total dry biomass, SPAD index, and leaf nitrogen content by 20.5, 15, and 21.5%, respectively. Thus the application of plant-derived protein hydrolysate containing amino acids and small peptides elicited a hormone-like activity, enhanced nitrogen uptake and consequently crop performances. PMID:25250039

  1. Antioxidant activity of protein hydrolysates derived from threadfin bream surimi byproducts.

    PubMed

    Wiriyaphan, Chompoonuch; Chitsomboon, Benjamart; Yongsawadigul, Jirawat

    2012-05-01

    Antioxidant activities of protein hydrolysates from threadfin bream surimi wastes, including frame, bone and skin (FBS) and refiner discharge (RD), were investigated. FBS and RD were rich in Lys, Glu, Gly, Pro, Asp, Leu, His, Tyr and Phe. FBS was hydrolysed to a greater extent than RD regardless of proteinases tested (Virgibacillus sp. SK33 proteinase, Alcalase, pepsin and trypsin). Pepsin-hydrolysed FBS, at a 5% degree of hydrolysis (DH), showed the highest antioxidant activity based on 2,2'-azinobis (3-ethyl-benzothiazoline-6-sulphonate) (ABTS) radical (0.455±0.054mg Trolox equivalents/mg leucine equivalents), ferric reducing antioxidant power (FRAP) (0.221±0.005mM Trolox equivalents) and inhibition of β-carotene bleaching assays. FBS hydrolysates showed higher antioxidant activity based on chemical assays than their RD counterparts. However, FBS and RD hydrolysates protected HepG2 cells against tert-butyl hydroperoxide-induced oxidative damage to a similar extent. Therefore, FBS and RD hydrolysates have a potential as antioxidative neutraceutical ingredients. PMID:26434269

  2. Immunomodulatory and hypoallergenic properties of milk protein hydrolysates in ICR mice.

    PubMed

    Pan, D D; Wu, Z; Liu, J; Cao, X Y; Zeng, X Q

    2013-08-01

    Approximately 2.5% of young children are allergic to cow milk. In this study, milk protein hydrolysates made from full-cream milk via enzymatic hydrolysis played a positive role in regulating the immune system of ICR mice. Milk protein hydrolysates enhanced immunity in mice by stimulating host immunity, probably by increasing the weight of certain immune system organs, improving the level of hemolysin in serum, and enhancing the phagocytosis of macrophages. Milk protein hydrolysates have the capability to reduce type I hypersensitivity by decreasing IgE levels, IL-4 in serum, and the release of histamine and bicarbonate in peritoneal mast cells, as well as enhancing transforming growth factor-β levels in the serum of ovalbumin-sensitized mice.

  3. Two novel antioxidant nonapeptides from protein hydrolysate of skate (Raja porosa) muscle.

    PubMed

    Hu, Fa-Yuan; Chi, Chang-Feng; Wang, Bin; Deng, Shang-Gui

    2015-04-01

    In the current study, the preparation conditions of neutrase hydrolysate (SMH) from skate (Raja porosa) muscle protein were optimized using orthogonal L9(3)4 tests, and R values indicated that pH was the most important factor affecting HO· scavenging activity of SMH. Under the optimum conditions of pH 7.0, enzymolysis temperature 60 °C, enzyme/substrate ratio (E/S) 2%, and enzymolysis time 5 h, EC50 of SMH on HO· was 2.14 ± 0.17 mg/mL. Using ultrafiltration, gel filtration chromatography, and RP-HPLC, two novel antioxidant nonapeptides (SP-A and SP-B) were isolated from SMH and their amino acid sequences were found to be APPTAYAQS (SP-A) and NWDMEKIWD (SP-B) with calculated molecular masses of 904.98 Da and 1236.38 Da, respectively. Both showed strong antioxidant activities. SP-A and SP-B exhibited good scavenging activities on HO· (EC50 0.390 and 0.176 mg/mL), DPPH· (EC50 0.614 and 0.289 mg/mL), and O2-· (EC50 0.215 and 0.132 mg/mL) in a dose-dependent manner. SP-B was also effective against lipid peroxidation in the model system. The aromatic (2Trp), acidic (2Asp and Glu), and basic (Lys) amino acid residues within the sequences of SP-B might account for its pronounced antioxidant activity. The results of this study suggested that protein hydrolysate and peptides from skate muscle might be effective as food additives for retarding lipid peroxidation occurring in foodstuffs. PMID:25854645

  4. Two Novel Antioxidant Nonapeptides from Protein Hydrolysate of Skate (Raja porosa) Muscle

    PubMed Central

    Hu, Fa-Yuan; Chi, Chang-Feng; Wang, Bin; Deng, Shang-Gui

    2015-01-01

    In the current study, the preparation conditions of neutrase hydrolysate (SMH) from skate (Raja porosa) muscle protein were optimized using orthogonal L9(3)4 tests, and R values indicated that pH was the most important factor affecting HO· scavenging activity of SMH. Under the optimum conditions of pH 7.0, enzymolysis temperature 60 °C, enzyme/substrate ratio (E/S) 2%, and enzymolysis time 5 h, EC50 of SMH on HO· was 2.14 ± 0.17 mg/mL. Using ultrafiltration, gel filtration chromatography, and RP-HPLC, two novel antioxidant nonapeptides (SP-A and SP-B) were isolated from SMH and their amino acid sequences were found to be APPTAYAQS (SP-A) and NWDMEKIWD (SP-B) with calculated molecular masses of 904.98 Da and 1236.38 Da, respectively. Both showed strong antioxidant activities. SP-A and SP-B exhibited good scavenging activities on HO· (EC50 0.390 and 0.176 mg/mL), DPPH· (EC50 0.614 and 0.289 mg/mL), and O2−· (EC50 0.215 and 0.132 mg/mL) in a dose-dependent manner. SP-B was also effective against lipid peroxidation in the model system. The aromatic (2Trp), acidic (2Asp and Glu), and basic (Lys) amino acid residues within the sequences of SP-B might account for its pronounced antioxidant activity. The results of this study suggested that protein hydrolysate and peptides from skate muscle might be effective as food additives for retarding lipid peroxidation occurring in foodstuffs. PMID:25854645

  5. Effect of foxtail millet protein hydrolysates on lowering blood pressure in spontaneously hypertensive rats

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to determine the effect of foxtail millet protein hydrolysates on lowering blood pressure in spontaneously hypertensive rats (SHRs). The protein of foxtail millet after extruding or fermenting and the raw foxtail millet was extracted and hydrolyzed by digestive protea...

  6. Maillard reaction products of rice protein hydrolysates with mono-, oligo- and polysaccharides

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Rice protein, a byproduct of rice syrup production, is abundant but, its lack of functionality prevents its wide use as a food ingredient. Maillard reaction products of (MRPs) hydrolysates from the limited hydrolysis of rice protein (LHRP) and various mono-, oligo- and polysaccharides were evaluat...

  7. Production and characterisation of hyaluronidase and elastase inhibitory protein hydrolysates from Venus clam.

    PubMed

    Sutthiwanjampa, Chanutchamon; Kim, Sang Moo

    2015-01-01

    The hydrolysates of fresh and boiled Venus clams with five different proteases for the production of low-molecular protein hydrolysates were optimised by response surface methodology. Alcalase hydrolysates exhibited the strongest hyaluronidase inhibitory activity. The optimum hydrolysis conditions of fresh and boiled clams were< enzyme-to-substrate ratio (E/S), 2.15%; time, 150 min; water-to-substrate ratio (W/S), 83.84 mL g(-1) for fresh clam, and E/S, 2.02%; time, 4.11 h; W/S, 69.74 mL g(-1) for boiled clam. The fresh and boiled clam protein hydrolysates were fractionated by S-200 HR size-exclusion chromatography, which resulted in one (FH1) and two (BH1 and BH2) fractions, respectively. BH1 exhibited the highest hyaluronidase and elastase inhibitory activities with specific activities of 141.15 and 81.36% mL mg(-1), respectively. Therefore, the boiled Venus clam hydrolysate might be developed as a cosmeceutical agent because of its strong hyaluronidase and elastase inhibitory activities.

  8. Kinetic characterization of Channa striatus muscle sarcoplasmic and myofibrillar protein hydrolysates.

    PubMed

    Ghassem, Masomeh; Fern, See Siau; Said, Mamot; Ali, Zainon Mohd; Ibrahim, Saadiah; Babji, Abdul Salam

    2014-03-01

    This study was conducted to evaluate the kinetic characteristics of proteolytic activity of proteases on Channa striatus protein fractions. Degree of hydrolysis (DH), amino acid composition and kinetic parameters of sarcoplasmic and myofibrillar proteins were investigated when incubated with proteinase K and thermolysin, separately. After 30 min incubation with proteases, a decrease in DH of sarcoplasmic protein was observed whereas, hydrolysis of myofibrillar protein with proteases took 2 h with an increase in DH. The major amino acids were glutamic acid (16.6%) in thermolysin- myofibrillar hydrolysate followed by aspartic acid (11.1%) in sarcoplasmic protein fraction with no enzyme treatment and lysine (10%) in thermolysin-myofibrillar hydrolysate. The apparent Michaelis constant of proteinase K was lower than thermolysin for both sarcoplasmic and myofibrillar proteins. However, rate of turnover and enzyme efficiency suggested that sarcoplasmic and myofibrillar proteins are suitable substrates for proteinase K and thermolysin hydrolytic reaction, respectively.

  9. Effect of cooking temperatures on protein hydrolysates and sensory quality in crucian carp (Carassius auratus) soup.

    PubMed

    Zhang, Jinjie; Yao, Yanjia; Ye, Xingqian; Fang, Zhongxiang; Chen, Jianchu; Wu, Dan; Liu, Donghong; Hu, Yaqin

    2013-06-01

    Cooking methods have a significant impact on flavour compounds in fish soup. The effects of cooking temperatures (55, 65, 75, 85, 95, and 100 °C) on sensory properties and protein hydrolysates were studied in crucian carp (Carassius auratus) soup. The results showed that the soup prepared at 85 °C had the best sensory quality in color, flavour, amour, and soup pattern. Cooking temperature had significant influence on the hydrolysis of proteins in the soup showed by SDS-PAGE result. The contents of water soluble nitrogen (WSN) and non-protein nitrogen (NPN) increased with the cooking temperature, but the highest contents of total peptides and total free amino acids (FAA) were obtained at the cooking temperature of 85 °C. The highest contents of umami-taste active amino acid and branched-chain amino acids were also observed in the 85 °C sample. In conclusion, a cooking temperature of 85 °C was preferred for more excellent flavor and higher nutritional value of crucian carp soup. PMID:24425950

  10. Chlorella Protein Hydrolysate Attenuates Glucose Metabolic Disorder and Fatty Liver in High-fat Diet-induced Obese Mice.

    PubMed

    Noguchi, Naoto; Yanagita, Teruyoshi; Rahman, Shaikh Mizanoor; Ando, Yotaro

    2016-07-01

    Chlorella (Parachlorella beijerinckii) powder is reported to show a preventive effect against metabolic syndromes such as arteriosclerosis, hyperlipidemia, and hypertension. Approximately 60% of the chlorella content is protein. In order to understand the role of chlorella protein, we prepared a chlorella protein hydrolysate (CPH) by protease treatment. Male C57BL/6 mice were divided into three groups: a normal diet group, high-fat diet (HFD) group, and high-fat diet supplemented with CPH (HFD+CPH) group. The CPH administration improved glucose intolerance, insulin sensitivity, and adipose tissue hypertrophy in the high-fat diet-fed mice. In addition, the HFD+CPH group had significantly decreased liver total cholesterol and triglyceride levels compared with those in the HFD group. Furthermore, the HFD+CPH group had a decreased level of monocyte chemotactic protein-1 (MCP-1) in serum and a lower MCP-1 mRNA expression level in adipose tissue compared with the HFD group. The present study suggests that chlorella protein hydrolysate can prevent a high-fat diet-induced glucose disorder and fatty liver by inhibiting adipocyte hypertrophy and reducing the MCP-1 protein and gene expression. PMID:27321121

  11. Regulation of Exacerbated Immune Responses in Human Peripheral Blood Cells by Hydrolysed Egg White Proteins

    PubMed Central

    Lozano-Ojalvo, Daniel; Molina, Elena; López-Fandiño, Rosina

    2016-01-01

    The anti-allergic potential of egg white protein hydrolysates (from ovalbumin, lysozyme and ovomucoid) was evaluated as their ability to hinder cytokine and IgE production by Th2-skewed human peripheral blood mononuclear cells (PBMCs), as well as the release of pro-inflammatory factors and generation of reactive oxygen species from Th1-stimulated peripheral blood leukocytes (PBLs). The binding to IgE of egg allergic patients was determined and the peptides present in the hydrolysates were identified. The hydrolysates with alcalase down-regulated the production of Th2-biased cytokines and the secretion of IgE to the culture media of Th2-skewed PBMCs, and they significantly neutralized oxidative stress in PBLs. The hydrolysates of ovalbumin and ovomucoid with pepsin helped to re-establish the Th1/Th2 balance in Th2-biased PBMCs, while they also inhibited the release of pro-inflammatory mediators and reduced oxidative stress in PBLs treated with inflammatory stimuli. The hydrolysates with alcalase, in addition to equilibrating Th2 differentiation, exhibited a low IgE-binding. Therefore, they would elicit mild allergic reactions while retaining T cell-stimulating abilities, which might correlate with an anti-allergic benefit. PMID:27007699

  12. Regulation of Exacerbated Immune Responses in Human Peripheral Blood Cells by Hydrolysed Egg White Proteins.

    PubMed

    Lozano-Ojalvo, Daniel; Molina, Elena; López-Fandiño, Rosina

    2016-01-01

    The anti-allergic potential of egg white protein hydrolysates (from ovalbumin, lysozyme and ovomucoid) was evaluated as their ability to hinder cytokine and IgE production by Th2-skewed human peripheral blood mononuclear cells (PBMCs), as well as the release of pro-inflammatory factors and generation of reactive oxygen species from Th1-stimulated peripheral blood leukocytes (PBLs). The binding to IgE of egg allergic patients was determined and the peptides present in the hydrolysates were identified. The hydrolysates with alcalase down-regulated the production of Th2-biased cytokines and the secretion of IgE to the culture media of Th2-skewed PBMCs, and they significantly neutralized oxidative stress in PBLs. The hydrolysates of ovalbumin and ovomucoid with pepsin helped to re-establish the Th1/Th2 balance in Th2-biased PBMCs, while they also inhibited the release of pro-inflammatory mediators and reduced oxidative stress in PBLs treated with inflammatory stimuli. The hydrolysates with alcalase, in addition to equilibrating Th2 differentiation, exhibited a low IgE-binding. Therefore, they would elicit mild allergic reactions while retaining T cell-stimulating abilities, which might correlate with an anti-allergic benefit. PMID:27007699

  13. The hypolipidemic effect and antithrombotic activity of Mucuna pruriens protein hydrolysates.

    PubMed

    Herrera Chalé, Francisco; Ruiz Ruiz, Jorge Carlos; Betancur Ancona, David; Acevedo Fernández, Juan José; Segura Campos, Maira Rubi

    2016-01-01

    Hydrolysates and peptide fractions (PF) obtained from M. pruriens protein concentrates with commercial and digestive enzymatic systems were studied for their hypolipidemic and antithrombotic activities. Hydrolysates obtained with Pepsin-Pancreatin (PP) and their peptide fractions inhibited cholesterol micellar solubility with a maximum value of 1.83% in PP. Wistar rats were used to evaluate the hypolipidemic effect of hydrolysates and PF. The higher reductions of cholesterol and triglyceride levels were exhibited by PP and both peptide fractions <1 kDa obtained from PP and Alcalase®-Flavourzyme® hydrolysate (AF) at a dose of 15 mg kg(-1) of animal weight. PF > 10 kDa from both hydrolysates showed the maximum antithrombotic activity with values of 33.33% for PF > 10 kDa from AF and 31.72% for PF > 10 kDa from PP. The results suggest that M. pruriens bioactive peptides with the hypolipidemic effect and antithrombotic activity might be utilized as nutraceuticals. PMID:26505152

  14. Enzymatic hydrolysis of rice protein with papain and antioxidation activity of hydrolysate

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The enzymatic hydrolysis technology of rice protein and the antioxidant activity of the hydrolysate were studied. Substrate concentration,enzyme dose,pH value and temperature were selected as factors to optimize the hydrolysis parameters with single—factor and orthogonal tests. Results show the opti...

  15. Protein Hydrolysates from Non-bovine and Plant Sources Replaces Tryptone in Microbiological Media

    NASA Astrophysics Data System (ADS)

    Ranganathan, Yamini; Patel, Shifa; Pasupuleti, Vijai K.; Meganathan, R.

    Tryptone (pancreatic digest of casein) is a common ingredient in laboratory and fermentation media for growing wild-type and genetically modified microorganisms. Many of the commercially manufactured products such as human growth hormone, antibiotics, insulin, etc. are produced by recombinant strains grown on materials derived from bovine sources. With the emergence of Bovine Spongiform Encephalopathy (BSE) and the consequent increase in Food and Drug Administration (FDA) regulations, elimination of materials of bovine origin from fermentation media is of paramount importance. To achieve this objective, a number of protein hydrolysates derived from non-bovine animal and plant sources were evaluated. Tryptone in Luria-Bertani (LB) broth was replaced with an equal quantity of alternate protein hydrolysates. Four of the six hydrolysates (one animal and three from plants) were found to efficiently replace the tryptone present in LB-medium as measured by growth rate and growth yield of a recombinant Escherichia coli strain. In addition, we have determined plasmid stability, inducibility and activity of the plasmid encoded β-galactosidase in the recombinant strain grown in the presence of various protein hydrolysates.

  16. Preparation of linear maltodextrins using a hyperthermophilic amylopullulanase with cyclodextrin- and starch-hydrolysing activities.

    PubMed

    Li, Xiaolei; Li, Dan

    2015-03-30

    A novel method for the preparation of linear maltodextrins from cyclodextrins and starch was proposed. To accomplish this process, an amylopullulanase from hyperthermophilic archaeon Caldivirga maquilingensis (CMApu) was characterized and used. CMApu with an estimated molecular mass of 62.7 kDa by SDS-PAGE had a maximal pullulan-hydrolysing activity at 100°C and pH 5.0. It could also hydrolyse amylopectin (AP), starch, β-CD and amylose (AM), in a decreasing order of relative activities from 88.96% to 57.17%. TLC and HPAEC analysis revealed that CMApu catalyzed the debranching and degrading reactions to produce linear malto-oligosaccharides (≤ G8-G1) from G8-β-CD and/or normal CDs, amylodextrins (DP6-96) from AM, and amylodextrins (DP1-76) from AP and potato starch. Our results showed that CMApu had a great potential for the industrial preparation of linear maltodextrins from normal starch instead of waxy starch, malto-oligosaccharides or sucrose. And the high optimal temperature of CMApu facilitated the simultaneous gelatinization and hydrolysis of cereal starch.

  17. Antioxidant potential of date (Phoenix dactylifera L.) seed protein hydrolysates and carnosine in food and biological systems.

    PubMed

    Ambigaipalan, Priyatharini; Shahidi, Fereidoon

    2015-01-28

    Date seed protein hydrolysates were evaluated for antioxidant activity as well as solubility and water-holding capacity in food and biological model systems. Date seed protein hydrolysates as well as carnosine exhibited >80% of solubility over a pH range of 2-12. The hydrolysates and carnosine at 0.5% (w/w) were also found to be effective in enhancing water-holding capacity and cooking yield in a fish model system, which was nearly similar to sodium tripolyphosphate (STPP; 0.3%, w/w). Incorporation of hydrolysates (200 ppm) in fish model systems resulted in the highest inhibition (30%) of oxidation in comparison to butylated hydroxytoluene (BHT; 9%). In addition, hydrolysates and carnosine inhibited β-carotene oxidation by 75%. The hydrolysates (0.1 mg/mL) inhibited LDL cholesterol oxidation by 60%, whereas carnosine inhibited oxidation by 80% after 12 h of incubation. Additionally, hydrolysates and carnosine effectively inhibited hydroxyl (6 mg/mL) and peroxyl (0.1 mg/mL) radical-induced DNA scission. Therefore, date seed protein hydrolysates could be used as a potential functional food ingredient for health promotion.

  18. Influence of peptides-phenolics interaction on the antioxidant profile of protein hydrolysates from Brassica napus.

    PubMed

    Hernández-Jabalera, Anaid; Cortés-Giraldo, Isabel; Dávila-Ortíz, Gloria; Vioque, Javier; Alaiz, Manuel; Girón-Calle, Julio; Megías, Cristina; Jiménez-Martínez, Cristian

    2015-07-01

    The role of the peptides-phenolic compounds (PC) interaction on the antioxidant capacity profile (ACP) of protein hydrolysates from rapeseed (Brassica napus) was studied in 36 hydrolysates obtained from a PC-rich and PC-reduced protein substrate. The latent profile analysis (LPA), with data of seven in vitro methods and one assay for cellular antioxidant activity (CAA), allowed identifying five distinctive groups of hydrolysates, each one with distinctive ACP. The interaction of peptides with naturally present PC diminished in vitro antioxidant activity in comparison with their PC-reduced counterparts. However, CAA increased when peptides-PC interaction occurred. The profile with the highest average CAA (62.41 ± 1.48%), shown by hydrolysates obtained by using alcalase, shared typical values of Cu(2+)-catalysed β-carotene oxidation (62.41 ± 0.43%), β-carotene bleaching inhibition (91.75 ± 0.22%) and Cu(2+)-chelating activity (74.53 ± 0.58%). The possibilities for a sample to exhibit ACP with higher CAA increased with each unit of positively charged amino acids, according to multinomial logistic regression analysis.

  19. In vitro antioxidant properties of chicken skin enzymatic protein hydrolysates and membrane fractions.

    PubMed

    Onuh, John O; Girgih, Abraham T; Aluko, Rotimi E; Aliani, Michel

    2014-05-01

    Chicken thigh and breast skin proteins were hydrolysed using alcalase or a combination of pepsin and pancreatin (PP), each at concentrations of 1-4%. The chicken skin protein hydrolysates (CSPHs) were then fractionated by membrane ultrafiltration into different molecular weight peptides (<1, 1-3, 3-5 and 5-10kDa) and analysed for antioxidant properties. Results showed that the CSPHs had a significantly (p<0.05) lower scavenging activity against DPPH radicals when compared to reduced glutathione. The chicken breast skin hydrolysates had significantly higher DPPH scavenging activity than the chicken thigh skin hydrolysates. DPPH scavenging and metal ion chelation increased significantly (p<0.05) from 29-40% to 86-89%, respectively with increasing proteolytic enzyme concentration. In contrast, the antioxidant properties decreased as peptide size increased. We conclude that CSPHs and their peptide fractions may be used as ingredients in the formulation of functional foods and nutraceuticals for the control and management of oxidative stress-related diseases.

  20. Angiotensin I-Converting Enzyme inhibitory and antioxidant activities and surfactant properties of protein hydrolysates as obtained of Amaranthus hypochondriacus L. grain.

    PubMed

    Soriano-Santos, J; Escalona-Buendía, H

    2015-04-01

    Even though some research has been carried out on surfactant properties of amaranth protein hydrolysates, their bio-functionality has not been studied yet. In this work amaranth grain Alb 1 and Glob were hydrolyzed (Alb 1H, Glob H) and foams and emulsions at optimal conditions (t, E/S, pH5) were prepared in order to assess techno-functional properties such as foaming (F) and emulsifying (E) (capacity (C) and stability (S)). FC and EC were much better for Glob H than for Alb H. Angiotensin I-converting enzyme-inhibitory activity was higher for Alb 1H (roughly 50 %) than that of Glob H (roughly 30 %). Scavenging of radicals activity (DPPH· or ABTS· (+) ) of Alb 1H and Glob H, at 2 mg/mL, was similar (approx. 40 %), but lower than Alb 1 (approx. 70 %), which was the best antioxidant. The low reducing power showed that hydrolysates barely donate an electron or hydrogen. Chelating activity on Cu(2+) was lower than that exhibited by Fe(2+,) which was remarkable, approx. 80 % as long as DH% > 10 %, where hydrolysates displayed high solubility (Alb 1H = 85 %, Glob H = 70 %) because of occurrence of 1-10 kDa peptides. Amaranth foams and emulsions prepared with protein hydrolysates have a potential as a nutraceutical food.

  1. Angiotensin I-Converting Enzyme inhibitory and antioxidant activities and surfactant properties of protein hydrolysates as obtained of Amaranthus hypochondriacus L. grain.

    PubMed

    Soriano-Santos, J; Escalona-Buendía, H

    2015-04-01

    Even though some research has been carried out on surfactant properties of amaranth protein hydrolysates, their bio-functionality has not been studied yet. In this work amaranth grain Alb 1 and Glob were hydrolyzed (Alb 1H, Glob H) and foams and emulsions at optimal conditions (t, E/S, pH5) were prepared in order to assess techno-functional properties such as foaming (F) and emulsifying (E) (capacity (C) and stability (S)). FC and EC were much better for Glob H than for Alb H. Angiotensin I-converting enzyme-inhibitory activity was higher for Alb 1H (roughly 50 %) than that of Glob H (roughly 30 %). Scavenging of radicals activity (DPPH· or ABTS· (+) ) of Alb 1H and Glob H, at 2 mg/mL, was similar (approx. 40 %), but lower than Alb 1 (approx. 70 %), which was the best antioxidant. The low reducing power showed that hydrolysates barely donate an electron or hydrogen. Chelating activity on Cu(2+) was lower than that exhibited by Fe(2+,) which was remarkable, approx. 80 % as long as DH% > 10 %, where hydrolysates displayed high solubility (Alb 1H = 85 %, Glob H = 70 %) because of occurrence of 1-10 kDa peptides. Amaranth foams and emulsions prepared with protein hydrolysates have a potential as a nutraceutical food. PMID:25829587

  2. Purification and characterization of high antioxidant peptides from duck egg white protein hydrolysates.

    PubMed

    Ren, Yao; Wu, Hui; Li, Xiaofeng; Lai, Furao; Xiao, Xinglong

    2014-10-01

    The hydrolysate from duck egg white protein (DEWP) prepared by "SEEP-Alcalase" at degree of hydrolysis (DH) value of 21% (namely HSA21) exhibited high antioxidant capacity in different oxidation systems. A consecutive chromatographic method was then developed for separation and purification of HSA21, including ion-exchange chromatography, macroporous adsorption resin (MAR) and gel filter chromatography. The final peptides "P21-3-75-B" were obtained with significantly enhanced antioxidant activity (p<0.05). It was further confirmed that the product mainly consisted of five oligopeptides (Mr: 202.1, 294.1, 382.1, 426.3, and 514.4Da). Furthermore, the antioxidant activity of P21-3-75-B kept stable after in vitro digestive simulation. Antioxidant capacity of the purified peptides was closely related to the molecular mass, hydrophobic amino acid residues, acidic amino acid and some antioxidant amino acids. This research provided a valuable route for producing new natural-source peptides with strong antioxidant capacity and high nutritious value for our daily intake.

  3. In vitro bioactive properties of intact and enzymatically hydrolysed whey protein: targeting the enteroinsular axis.

    PubMed

    Power-Grant, O; Bruen, C; Brennan, L; Giblin, L; Jakeman, P; FitzGerald, R J

    2015-03-01

    Enzymatically hydrolysed milk proteins have a variety of biofunctional effects some of which may be beneficial in the management of type 2 diabetes mellitus. The purpose of this study was to evaluate the effect of commercially available intact and hydrolysed whey protein ingredients (DH 32, DH 45) on markers of the enteroinsular axis (glucagon like peptide-1 secretion, dipeptidyl peptidase IV inhibition, insulin secretion and antioxidant activity) before and after simulated gastrointestinal digestion (SGID). A whey protein hydrolysate, DH32, significantly enhanced (P < 0.05) insulin secretion from BRIN BD11 β-cells compared to the positive control (16.7 mM glucose and 10 mM Ala). The whey protein hydrolysates inhibited dipeptidyl peptidase IV activity, yielding half maximal inhibitory concentration values (IC50) of 1.5 ± 0.1 and 1.1 ± 0.1 mg mL(-1) for the DH 32 and DH 45, samples respectively, and were significantly more potent than the intact whey (P < 0.05). Enzymatic hydrolysis of whey protein significantly enhanced (P < 0.05) its antioxidant activity compared to intact whey, as measured by the oxygen radical absorbance capacity assay (ORAC). This antioxidant activity was maintained (DH 32, P > 0.05) or enhanced (DH 45, P < 0.05) following SGID. Intact whey stimulated GLP-1 secretion from enteroendocrine cells compared to vehicle control (P < 0.05). This data confirm that whey proteins and peptides can act through multiple targets within the enteroinsular axis and as such may have glucoregulatory potential.

  4. Protein Hydrolysates Are Avoided by Herbivores but Not by Omnivores in Two-Choice Preference Tests

    PubMed Central

    Field, Kristin L.; Bachmanov, Alexander A.; Mennella, Julie A.; Beauchamp, Gary K.; Kimball, Bruce A.

    2009-01-01

    Background The negative sensory properties of casein hydrolysates (HC) often limit their usage in products intended for human consumption, despite HC being nutritious and having many functional benefits. Recent, but taxonomically limited, evidence suggests that other animals also avoid consuming HC when alternatives exist. Methodology/Principal Findings We evaluated ingestive responses of five herbivorous species (guinea pig, mountain beaver, gopher, vole, and rabbit) and five omnivorous species (rat, coyote, house mouse, white-footed mouse, and deer mouse; N = 16–18/species) using solid foods containing 20% HC in a series of two-choice preference tests that used a non-protein, cellulose-based alternative. Individuals were also tested with collagen hydrolysate (gelatin; GE) to determine whether it would induce similar ingestive responses to those induced by HC. Despite HC and GE having very different nutritional and sensory qualities, both hydrolysates produced similar preference score patterns. We found that the herbivores generally avoided the hydrolysates while the omnivores consumed them at similar levels to the cellulose diet or, more rarely, preferred them (HC by the white-footed mouse; GE by the rat). Follow-up preference tests pairing HC and the nutritionally equivalent intact casein (C) were performed on the three mouse species and the guinea pigs. For the mice, mean HC preference scores were lower in the HC v C compared to the HC v Cel tests, indicating that HC's sensory qualities negatively affected its consumption. However, responses were species-specific. For the guinea pigs, repeated exposure to HC or C (4.7-h sessions; N = 10) were found to increase subsequent HC preference scores in an HC v C preference test, which was interpreted in the light of conservative foraging strategies thought to typify herbivores. Conclusions/Significance This is the first empirical study of dietary niche-related taxonomic differences in ingestive responses to

  5. Selective separation and concentration of antihypertensive peptides from rapeseed protein hydrolysate by electrodialysis with ultrafiltration membranes.

    PubMed

    He, Rong; Girgih, Abraham T; Rozoy, Elodie; Bazinet, Laurent; Ju, Xing-Rong; Aluko, Rotimi E

    2016-04-15

    Rapeseed protein isolate was subjected to alcalase digestion to obtain a protein hydrolysate that was separated into peptide fractions using electrodialysis with ultrafiltration membrane (EDUF) technology. The EDUF process (6h duration) led to isolation of three peptide fractions: anionic (recovered in KCl-1 compartment), cationic (recovered in KCl-2 compartment), and those that remained in the feed compartment, which was labeled final rapeseed protein hydrolysate (FRPH). As expected the KCl-1 peptides were enriched in negatively-charged (43.57%) while KCl-2 contained high contents of positively-charged (28.35%) amino acids. All the samples inhibited angiotensin converting enzyme (ACE) and renin activities in dose-dependent manner with original rapeseed protein hydrolysate having the least ACE-inhibitory IC50 value of 0.0932±0.0037 mg/mL while FRPH and KCl-2 had least renin-inhibitory IC50 values of 0.47±0.05 and 0.55±0.06 mg/mL, respectively. Six hours after oral administration (100 mg/kg body weight) to spontaneously hypertensive rats, the FRPH produced the maximum systolic blood pressure reduction of -51 mmHg.

  6. Antioxidant activity of black bean (Phaseolus vulgaris L.) protein hydrolysates

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this work was to study the effect of enzymatic hydrolysis of black bean protein concentrate using different enzymes. Bean proteins were extracted and hydrolyzed over a period of 120 min using the enzymes pepsin or alcalase. The protein hydrolysates’ molecular weight was assayed by e...

  7. Study of Anti-Fatigue Effect in Rats of Ferrous Chelates Including Hairtail Protein Hydrolysates.

    PubMed

    Huang, Saibo; Lin, Huimin; Deng, Shang-Gui

    2015-12-01

    The ability of ferrous chelates including hairtail protein hydrolysates to prevent and reduce fatigue was studied in rats. After hydrolysis of hairtail surimi with papain, the hairtail protein hydrolysates (HPH) were separated into three groups by range of relative molecular weight using ultrafiltration membrane separation. Hairtail proteins were then chelated with ferrous ions, and the antioxidant activity, the amino acid composition and chelation rate of the three kinds of ferrous chelates including hairtail protein hydrolysates (Fe-HPH) were determined. Among the three groups, the Fe-HPH chelate showing the best conditions was selected for the anti-fatigue animal experiment. For it, experimental rats were randomly divided into seven groups. Group A was designated as the negative control group given distilled water. Group B, the positive control group, was given glutathione. Groups C, D and E were designated as the Fe-HPH chelate treatment groups and given low, medium, and high doses, respectively. Group F was designated as HPH hydrolysate treatment group, and Group G was designated as FeCl₂ treatment group. The different diets were orally administered to rats for 20 days. After that time, rats were subjected to forced swimming training after 1 h of gavage. Rats given Fe-FPH chelate had higher haemoglobin regeneration efficiency (HRE), longer exhaustive swimming time and higher SOD activity. Additionally, Fe-FPH chelate was found to significantly decrease the malondialdehyde content, visibly enhance the GSH-Px activity in liver and reduce blood lactic acid of rats. Fe-HPH chelate revealed an anti-fatigue effect, similar to or better than the positive control substance and superior to HPH or Fe when provided alone. PMID:26633476

  8. Study of Anti-Fatigue Effect in Rats of Ferrous Chelates Including Hairtail Protein Hydrolysates

    PubMed Central

    Huang, Saibo; Lin, Huimin; Deng, Shang-gui

    2015-01-01

    The ability of ferrous chelates including hairtail protein hydrolysates to prevent and reduce fatigue was studied in rats. After hydrolysis of hairtail surimi with papain, the hairtail protein hydrolysates (HPH) were separated into three groups by range of relative molecular weight using ultrafiltration membrane separation. Hairtail proteins were then chelated with ferrous ions, and the antioxidant activity, the amino acid composition and chelation rate of the three kinds of ferrous chelates including hairtail protein hydrolysates (Fe-HPH) were determined. Among the three groups, the Fe-HPH chelate showing the best conditions was selected for the anti-fatigue animal experiment. For it, experimental rats were randomly divided into seven groups. Group A was designated as the negative control group given distilled water. Group B, the positive control group, was given glutathione. Groups C, D and E were designated as the Fe-HPH chelate treatment groups and given low, medium, and high doses, respectively. Group F was designated as HPH hydrolysate treatment group, and Group G was designated as FeCl2 treatment group. The different diets were orally administered to rats for 20 days. After that time, rats were subjected to forced swimming training after 1 h of gavage. Rats given Fe-FPH chelate had higher haemoglobin regeneration efficiency (HRE), longer exhaustive swimming time and higher SOD activity. Additionally, Fe-FPH chelate was found to significantly decrease the malondialdehyde content, visibly enhance the GSH-Px activity in liver and reduce blood lactic acid of rats. Fe-HPH chelate revealed an anti-fatigue effect, similar to or better than the positive control substance and superior to HPH or Fe when provided alone. PMID:26633476

  9. Enhancement of ACE and prolyl oligopeptidase inhibitory potency of protein hydrolysates from sardine and tuna by-products by simulated gastrointestinal digestion.

    PubMed

    Martínez-Alvarez, Oscar; Batista, Irineu; Ramos, Cristina; Montero, Pilar

    2016-04-01

    This work was focused on the study of the bioactive potential of three fish protein hydrolysates, one of them prepared from industrial sardine by-products (head and viscera) and the others from tuna by-products (head, and muscle and viscera). These protein hydrolysates exhibited moderate ability to inhibit Angiotensin Converting Enzyme or ACE (IC50 between 0.24-1.16 mg dry weight per ml) and prolyl oligopeptidase or PO (IC50 between 3.30-9.57 mg ml(-1)), those obtained from tuna by-products being the most effective. Overall, ACE- and PO-inhibiting activities were enhanced by sequential nanofiltration through 3 and 1 kDa MWCO membranes (IC50 between 0.02-0.16 mg ml(-1) (ACE) and 1.10-4.21 mg ml(-1) (PO)). The inhibitory properties of the hydrolysates were greatly improved by in vitro gastric digestion, and were barely affected by further intestinal digestion. The digested tuna hydrolysates, mainly that from heads, proved to be the best source of PO- and ACE- inhibiting molecules (IC50 = 0.16 mg ml(-1) (ACE) and 1.04 mg ml(-1) (PO)) and could be potential new ingredients in food with interest in the prevention or treatment of cardiovascular and neurological diseases. PMID:27045751

  10. Identification of immunogenic maize proteins in a casein hydrolysate formula.

    PubMed

    Frisner, H; Rosendal, A; Barkholt, V

    2000-05-01

    Cow's milk-based formulas used for infants with cow's milk allergy are based on hydrolyzed proteins. The formulas that are successful in preventing allergic responses are extensively hydrolyzed. Nevertheless, reactions to such formulas are occasionally reported, and protein material of higher molecular weight than expected has been detected by binding immunoglobulin E (IgE) from patients' sera. This paper presents the identification of high-molecular-weight material in the extensively hydrolyzed casein formula, Nutramigen. The material was concentrated by simple centrifugation. The proteins in the pellet were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and protein-containing bands were analyzed by protein sequencing after electroblotting. The proteins were identified as maize zeins, which are water-insoluble proteins of apparent M(r) 20,000 and 23,000, presumably originating from the maize starch in Nutramigen. Rabbits immunized with this formula developed antibodies against zeins but not against milk proteins. The maize zeins are probably identical to the recently reported components in Nutramigen (1), detected by binding of IgE from milk allergic patients, but not correlated to clinical allergic reactivity. The clinical relevance of maize proteins in Nutramigen remains to be established. PMID:10893013

  11. Human insulinotropic response to oral ingestion of native and hydrolysed whey protein.

    PubMed

    Power, O; Hallihan, A; Jakeman, P

    2009-07-01

    The insulinotropic response to the ingestion of whey protein and whey protein hydrolysate, independent of carbohydrate, is not known. This study examined the effect of protein hydrolysis on the insulinotropic response to the ingestion of whey protein. Sixteen healthy males ingested a 500 mL solution containing either 45 g of whey protein (WPI) or whey protein hydrolysate (WPH). The estimated rate of gastric emptying was not altered by hydrolysis of the protein [18 (3) vs. 23 (3) min, n = 16; P = 0.15]. Maximum plasma insulin concentration (Cmax) occurred later (40 vs. 60 min) and was 28% [234 (26) vs. 299 (31) mM, P = 0.018] greater following ingestion of the WPH compared to the WPI leading to a 43% increase [7.6 (0.9) vs. 10.8 (2.6) nM, P = 0.21] in the AUC of insulin for the WPH. Of the amino acids with known insulinotropic properties only Phe demonstrated a significantly greater maximal concentration [C (max); 65 (2) vs. 72 (3) microM, n = 16; P = 0.01] and increase (+22%) in AUC following ingestion of the WPH. In conclusion, ingestion of whey protein is an effective insulin secretagogue. Hydrolysis of whey protein prior to ingestion augments the maximal insulin concentration by a mechanism that is unrelated to gastric emptying of the peptide solution.

  12. The Effects of Mechanically Deboned Chicken Hydrolysates on the Characteristics of Imitation Crab Stick

    PubMed Central

    Jin, Sang-Keun; Hwang, Jin-Won; Moon, Sungsil; Choi, Yeung-Joon; Kim, Gap-Don; Jung, Eun-Young; Yang, Han-Sul

    2014-01-01

    The effects of adding mechanically deboned chicken (MDC) hydrolysates on the quality characteristics of imitation crab stick (ICS) during storage were investigated. ICS was prepared from Alaska Pollack, chicken breast surimi, and protein hydrolysates enzymatically extracted from MDC. ICS samples were divided into 4 groups: without protein hydrolysate (control), added with 0.5% protein hydrolysate (T1), added with 1.0% protein hydrolysate (T2), and added with 1.5% protein hydrolysate (T3). Results showed that crude protein content did not differ significantly among the ICS samples (p>0.05). ICS sample added with MDC hydrolysates had higher crude fat and ash content but lower moisture content than the control (p<0.05). Lightness was significantly lower in T2 and T3 than in the other groups at 0 and 4 wk of storage. Also, whiteness decreased in the groups contained MDC hydrolysates. Breaking force and jelly strength were higher in samples containing MDC hydrolysates compared to control samples (p<0.05). Additionally, saturated fatty acid contents were lower in the groups containing MDC hydrolysates than in control sample groups (p<0.05). Polyunsaturated fatty acid (PUFA) and essential fatty acids (EFA) were significantly higher in T2 and T3 than the control samples. In particular, all samples containing MDC hydrolysates had reduced thiobarbituric acid-reactive substances (TBARS) values at 4 wk. Free radical scavenging activity also was increased with addition of MDC hydrolysates. PMID:26760938

  13. Isolation and structural elucidation of antioxidant peptides from oyster (Saccostrea cucullata) protein hydrolysate.

    PubMed

    Umayaparvathi, S; Meenakshi, S; Vimalraj, V; Arumugam, M; Balasubramanian, T

    2014-01-01

    Protein derived from the oyster (Saccostrea cucullata) was hydrolyzed using protease from Bacillus cereus SU12 for isolation of antioxidant peptides. The oyster hydrolysate exhibited a strong antioxidant potential in DPPH (85.7±0.37%) followed by Hydrogen peroxide radical scavenging activity (81.6±0.3%), Hydroxyl radical-scavenging activity (79.32±0.6%), Reducing power assay (2.63±0.2 OD at 700nm). Due to the high antioxidant potential, hydrolysate was fractionated in Sephadex G-25 gel filtration chromatography. The active peptide fraction was further purified by UPLC-MS. Totally 7 antioxidant peptides were collected. Among 7 peptides (SCAP 1-7), 3 peptides (SCAP 1, 3 and 7) had highest scavenging ability on DPPH radicals. The amino acid sequence and molecular mass of purified antioxidant peptides (SCAP1, SCAP3 and SCAP7) were determined by Q-TOF ESI mass spectroscopy and structures of the peptides were Leu-Ala-Asn-Ala-Lys (MW=515.29Da), Pro-Ser-Leu-Val-Gly-Arg-Pro-Pro-Val-Gly-Lys-Leu-Thr-Leu (MW=1432.89Da) and Val-Lys-Val-Leu-Leu-Glu-His-Pro-Val-Leu (MW=1145.75Da), respectively. The unique amino acid composition and sequence in the peptides might play an important role in expression of their antioxidant activity. The results of this study suggest that oyster protein hydrolysate is good source of natural antioxidants.

  14. Valorization of tomato waste proteins through production of antioxidant and antibacterial hydrolysates by proteolytic Bacillus subtilis: optimization of fermentation conditions.

    PubMed

    Moayedi, Ali; Hashemi, Maryam; Safari, Mohammad

    2016-01-01

    In this study, protein-rich waste of tomato processing industries was fermented by Bacillus subtilis A14h to produce hydrolysates with antioxidant and antibacterial activities. The effects of different levels of initial pH, incubation temperature, fermentation time, protein concentration and inoculum size on proteolytic activity, release of amino acids and peptides, antioxidant and antibacterial activities of hydrolysates were evaluated and optimized by using response surface methodology (RSM). Results showed that all the evaluated variables significantly influenced the hydrolysis and bioactivities of hydrolysates in polynomial models. Hydrolysates showed remarkable 2, 2'-diphenyl-1-picrylhydrazyl (DPPH) scavenging activity (up to 70 %), ferric ion reducing power, and inhibitory activity against B. cereus (up to 69.8 %) and E. coli (up to 29.8 %). Overall, good correlation between the concentration of amino acids and peptides, and antioxidant as well as antibacterial activities (in particular for B. cereus inhibition activity) was observed. Finally, optimum conditions for fermentative conversion of tomato waste proteins to antioxidant and antibacterial hydrolysates were established. Results of this study showed that tomato waste protein can be valorized to produce antioxidant and antibacterial hydrolysates in a fermentative system using B. subtilis A14h. PMID:26787958

  15. Fish protein hydrolysates affect cholesterol metabolism in rats fed non-cholesterol and high-cholesterol diets.

    PubMed

    Hosomi, Ryota; Fukunaga, Kenji; Arai, Hirofumi; Kanda, Seiji; Nishiyama, Toshimasa; Yoshida, Munehiro

    2012-03-01

    Fish consumption is well known to provide health benefits in both experimental animals and human subjects. Numerous studies have demonstrated the beneficial effects of various protein hydrolysates on lipid metabolism. In this context, this study examined the effect of fish protein hydrolysates (FPH) on cholesterol metabolism compared with the effect of casein. FPHs were prepared from Alaska pollock meat using papain as a protease. Male Wistar rats were divided into the following four dietary groups of seven rats each: either casein (20%) or FPH (10%) + casein (10%), with or without 0.5% cholesterol and 0.1% sodium cholate. Serum and liver lipid levels, fecal cholesterol and bile acid excretions, and the hepatic expression of genes encoding proteins involved in cholesterol homeostasis were examined. In rats fed the FPH diets compared with casein diets with or without cholesterol and sodium cholate, the indexes of cholesterol metabolism-namely, serum cholesterol, triglyceride, and low-density lipoprotein-cholesterol levels-were significantly lower, whereas fecal cholesterol and bile acid excretions were higher. Rats fed the FPH diets compared with casein with cholesterol exhibited a lower liver cholesterol level via an increased liver cholesterol 7α-hydroxylase (CYP7A1) expression level. This study demonstrates that the intake of FPH has hypocholesterolemic effects through the enhancement of fecal cholesterol and bile acid excretions and CYP7A1 expression levels. Therefore, fish peptides prepared by papain digestion might provide health benefits by decreasing the cholesterol content in the blood, which would contribute to the prevention of circulatory system diseases such as arteriosclerosis. PMID:22181072

  16. Changes in structural characteristics of antioxidative soy protein hydrolysates resulting from scavenging of hydroxyl radicals.

    PubMed

    Zhao, Jing; Xiong, Youling L; McNear, Dave H

    2013-02-01

    Antioxidant activity of soy protein (SP) and its hydrolyzed peptides has been widely reported. During scavenging of radicals, these antioxidative compounds would be oxidatively modified, but their fate is not understood. The objective of this study was to evaluate the structural characteristics of SP hydrolysates (SPHs), compared to intact SP, when used to neutralize hydroxyl radicals (•OH). SPHs with degree of hydrolysis (DH) 1 to 5 were prepared with Alcalase. Antioxidant activity of SPHs was confirmed by lipid oxidation inhibition measured with thiobarbituric acid-reactive substances, ability to scavenge 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) radicals, and ferrous ion chelation capability. Oxidation of SPHs was initiated by reaction with •OH generated from 0.1 mM FeCl(3) , 20 mM H(2) O(2) , and 1.0 mM ascorbate. After oxidative stress, carbonyl content of SPHs increased by 2- to 3-fold and sulfhydryl groups decreased by up to 42% compared to nonoxidized samples (P < 0.05). Methionine, histidine, and lysine residues were significantly reduced as a result of inactivating •OH (P < 0.05). Attenuated total reflectance-Fourier transform infrared and circular dichroism spectroscopy suggested the conversion of helical structure to strands and turns. Oxidatively modified SPHs had a lower intrinsic fluorescence intensity but similar solubility when compared to nonoxidized samples. These structural changes due to •OH stress may impact the ingredient interaction and functionality of SPHs in food products. PMID:23331209

  17. Isolation of peptides from an enzymatic hydrolysate of food proteins and characterization of their taste properties.

    PubMed

    Maehashi, K; Matsuzaki, M; Yamamoto, Y; Udaka, S

    1999-03-01

    Soybean protein, casein, bonito protein and chicken protein, each as foodstuff protein, were hydrolyzed with four proteinases; namely, pepsin, trypsin, alpha-chymotrypsin and bromelain. Since the chicken protein hydrolysate with bromelain possessed the most favorable umami taste, eleven peptides were isolated from the chicken protein hydrolysate by successive chromatography on ODS, Amberlite IR-120B, Amberlite IRA-410 and AG-50W; their structures were Asp-Ala, Asp-Val, Glu-Glu, Glu-Val, Ala-Asp-Glu, Ala-Glu-Asp, Asp-Glu-Glu, Asp-Glu-Ser, Glu-Glu-Asn, Ser-Pro-Glu, and Glu-Pro-Ala-Asp. Many of them did not show any umami taste by themselves, but Glu-Glu, Glu-Val, Ala-Asp-Glu, Ala-Glu-Asp, Asp-Glu-Glu, and Ser-Pro-Glu were recognized to enhance the umami taste of 0.02% 5'-inosine monophosphate (IMP). A combination of these peptides, especially 0.5% each of Glu-Glu, Glu-Val, Asp-Glu-Glu and Glu-Glu-Asn, with 0.02% IMP produced a delicious "full" umami taste. PMID:10227142

  18. Multifunctional cationic peptide fractions from flaxseed protein hydrolysates.

    PubMed

    Udenigwe, Chibuike C; Aluko, Rotimi E

    2012-03-01

    The aim of this work was to determine the multifunctional properties of flaxseed protein-derived cationic peptide fractions. Alcalase hydrolysis of flaxseed protein fractions liberated cationic peptides, which were separated into two major fractions (FI and FII) by chromatography using a cation-exchange column. Due to their cationic property, the peptide fractions bound and inactivated calmodulin (CaM, a negatively charged enzyme activator protein) with concomitant inhibition of CaM-dependent phosphodiesterase (CaMPDE); this activity was substantially reduced as CaM concentration increased. Enzyme kinetics studies showed competitive inhibition of CaMPDE by FI and FII with enzyme-inhibitor dissociation constants of 0.0202 and 0.0511 mg/ml, respectively. Only the FII peptides showed multifunctional activities by inhibiting CaMPDE, angiotensin converting enzyme (ACE) and renin. Separation of FII peptides by reverse phase HPLC resulted in eight fractions (FII-2 to FII-9) that inhibited the activities of CaMPDE, ACE, and renin but this multifunctional activity was more pronounced in FII-6. From LC-MS analysis, identified peptides present in FII fraction had molecular size range of 330-735 Da, which suggests potential for increased absorption. Potential peptide sequences were identified for each of the HPLC fractions and shown to contain either lysine or arginine as the positively charged amino acid residue. The multifunctional properties of the cationic peptide fractions can potentially enhance their use in targeting multiple symptoms of cardiovascular disease, considering that the excessive levels of CaM, CaMPDE, renin and ACE play important roles in enhancing progression and intensity of chronic human diseases. PMID:22327315

  19. Antioxidant and cryoprotective effects of Amur sturgeon skin gelatin hydrolysate in unwashed fish mince.

    PubMed

    Nikoo, Mehdi; Benjakul, Soottawat; Xu, Xueming

    2015-08-15

    Antioxidant and cryoprotective effects of Amur sturgeon skin gelatin hydrolysates prepared using different commercial proteases in unwashed fish mince were investigated. Gelatin hydrolysates prepared using either Alcalase or Flavourzyme, were effective in preventing lipid oxidation as evidenced by the lower thiobarbituric acid-reactive substances formation. Gelatin hydrolysates were able to retard protein oxidation as indicated by the retarded protein carbonyl formation and lower loss in sulfhydryl content. In the presence of gelatin hydrolysates, unwashed mince had higher transition temperature of myosin and higher enthalpy of myosin and actin as determined by differential scanning calorimetry. Based on low field proton nuclear magnetic resonance analysis, gelatin hydrolysates prevented the displacement of water molecules between the different compartments, thus stabilizing the water associated with myofibrils in unwashed mince induced by repeated freeze-thawing. Oligopeptides in gelatin hydrolysates more likely contributed to the cryoprotective effect. Thus, gelatin hydrolysate could act as both antioxidant and cryoprotectant in unwashed fish mince.

  20. Poultry feed based on protein hydrolysate derived from chrome-tanned leather solid waste: creating value from waste.

    PubMed

    Chaudhary, Rubina; Pati, Anupama

    2016-04-01

    Leather industry generates huge amount of chrome-containing leather solid waste which creates major environment problems to tanners worldwide. Chrome-tanned leather solid waste is primarily chromium complex of collagen protein. The presence of chromium limits its protein application in animal feed industry. The purified protein hydrolysate with zero chromium could be used in poultry feed. In this study, an attempt has been made to assess performance of poultry with purified protein hydrolysate as a feed derived from chrome-tanned leather waste as partial replacement of soyabean meal as a sole source of protein for growing broiler chickens. Growth study was conducted to evaluate the effect of feeding protein hydrolysate on performance and physiochemical characteristics of meat of broiler chickens. Two experimental diets containing various levels of protein hydrolysate (EI-20 % and EII-30 %) were evaluated. The comparative study was performed as control with soyabean meal. Daily feed intake, body weight gain and feed conversion ratio were measured from day 8 to day 35. At the end of the study, birds were randomly selected and slaughtered to evaluate for physiochemical characteristics of meat. Diet had significant effects on feed intake and body weight gain. Birds fed with 20 and 30 % protein hydrolysate consumed 9.5 and 17.5 % higher amount of feed and gained 6.5 and 16.6 % higher than soyabean meal-fed birds. The current study produced evidence that protein hydrolysate can replace up to 75 % of soyabean meal in broiler diets without affecting either growth performance or meat characteristics.

  1. Preparing Protein Samples

    NASA Technical Reports Server (NTRS)

    2000-01-01

    Cindy Barnes of University Space Research Association (USRA) at NASA's Marshall Space Flight Center pipettes a protein solution in preparation to grow crystals as part of NASA's structural biology program. Research on Earth helps scientists define conditions and specimens they will use in space experiments.

  2. Evaluation of the immunogenicity of protein hydrolysate formulas using laboratory animal hyperimmunization.

    PubMed

    Cordle, C T; Duska-McEwen, G; Janas, L M; Malone, W T; Hirsch, M A

    1994-02-01

    The rabbit hyperimmunization model has previously been used to evaluate candidate hypoallergenic protein ingredients. Use of the model has been expanded to include the evaluation of protein hydrolysate formulas. Each formula's immunological reactivity was determined by ELISA measurement of formula-specific rabbit antibody. Animals hyperimmunized with formulas containing extensively hydrolyzed proteins (Alimentum, Nutramigen, and Pregestimil) generated very weak immune responses (< 100 fold antibody increase). Products containing intact or partially hydrolyzed proteins (Similac, Enfamil, Good Start, Beba HA, and Nidina HA) elicited high level (> 10,000 fold increase) immune responses. Immunogenicity results were then compared to measurements of residual antigen content (by inhibition ELISA) and clinical performance. Correlation of formula immunogenicity, antigenicity and clinical performance indicates that the rabbit model is useful for screening "hypoallergenic" formulas to predict allergenic reactivity.

  3. Lower weight gain and hepatic lipid content in hamsters fed high fat diets supplemented with white rice protein, brown rice protein, and soy protein and their hydrolysates

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The physiological effects of the hydrolysates from white rice, brown rice, and soy isolate were compared to the original protein source. White rice, brown rice, and soy protein were hydrolyzed with the food grade enzyme, alcalase2.4 L®. Male Syrian hamsters were fed high-fat diets containing eithe...

  4. Production of bioactive protein hydrolysate using the yeasts isolated from soft chhurpi.

    PubMed

    Rai, Amit Kumar; Kumari, Reena; Sanjukta, Samurailatpam; Sahoo, Dinabandhu

    2016-11-01

    The aim of this work was to study the production of bioactive protein hydrolysates using yeasts isolated from chhurpi. For this, a total of 125 proteolytic yeasts were isolated and molecular identification was carried out by analysis of the restriction digestion pattern generated by digesting the PCR amplified internal transcribed spacer region and 5.8S rRNA gene (ITS1-5.8S-ITS2) using three endonucleases (HaeIII, CfoI and HinfI). The results obtained showed that different proteolytic yeasts were dominant in marketed products (Kluyveromyces marxianus and Issatchenkia orientalis) and samples from production centers (Trichosporon asahii, Saccharomyces cerevisiae and Exophiala dermatitidis). Proteolytic strains in individual groups showed their ability to hydrolyze milk protein and enhance antioxidant property. Among the isolates, fermentation using K. marxianus YMP45 and S. cerevisiae YAM14 resulted in higher antioxidant activity. This is the first report on application of yeast isolated from fermented food of North-East India for the production of bioactive protein hydrolysate. PMID:27494105

  5. Rice bran proteins and their hydrolysates modulate cholesterol metabolism in mice on hypercholesterolemic diets.

    PubMed

    Zhang, Huijuan; Wang, Jing; Liu, Yingli; Gong, Lingxiao; Sun, Baoguo

    2016-06-15

    The hypolipidemic properties of defatted rice bran protein (DRBP), fresh rice bran protein (FRBP), DRBP hydrolysates (DRBPH), and FRBP hydrolysates (FRBPH) were determined in mice on high fat diets for four weeks. Very low-density lipoprotein cholesterol (VLDL-C), low-density lipoprotein cholesterol (LDL-C) contents, and the hepatic total cholesterol content were reduced while fecal total cholesterol and total bile acid (TBA) contents were increased in the FRBPH diet group. The expression levels of hepatic genes for cholesterol biosynthesis HMG-CoAR and SREBP-2 were lowest in the FRBPH diet group. The mRNA level of HMG-CoAR was significantly positively correlated with the hepatic TG content (r = 0.82, P < 0.05). The mRNA levels of genes related to bile acid biosynthesis and cholesterol efflux, CYP7A1, ABCA1, and PPARγ were up-regulated in all test groups. The results suggest that FRBPH regulates cholesterol metabolism in mice fed the high fat and cholesterol diet by increasing fecal steroid excretion and expression levels of genes related to bile acid synthesis and cholesterol efflux, and the down-regulation of the expression levels of genes related to cholesterol biosynthesis. PMID:27216972

  6. Whey protein hydrolysates enhance water absorption in the perfused small intestine of anesthetized rats.

    PubMed

    Ito, Kentaro; Yamaguchi, Makoto; Noma, Teruyuki; Yamaji, Taketo; Itoh, Hiroyuki; Oda, Munehiro

    2016-08-01

    We evaluated the effect of whey protein hydrolysates (WPH) on the water absorption rate in the small intestine using a rat small intestine perfusion model. The rate was significantly higher with 5 g/L WPH than with 5 g/L soy protein hydrolysates or physiological saline (p < 0.05). WPH dose-dependently increased the water absorption rate in the range of 1.25-10.0 g/L. WPH showed a significantly higher rate than an amino acid mixture whose composition was equal to that of WPH (p < 0.05). The addition of 4-aminomethylbenzoic acid, an inhibitor of PepT1, significantly suppressed WPH's enhancement of water absorption (p < 0.05). The rate of water absorption was significantly correlated with that of peptides/amino acids absorption in WPH (r = 0.82, p < 0.01). These data suggest that WPH have a high water absorption-promoting effect, to which PepT1 contributes.

  7. Effect of whey protein hydrolysate on performance and recovery of top-class orienteering runners.

    PubMed

    Hansen, Mette; Bangsbo, Jens; Jensen, Jørgen; Bibby, Bo Martin; Madsen, Klavs

    2015-04-01

    This trial aimed to examine the effect of whey protein hydrolysate intake before and after exercise sessions on endurance performance and recovery in elite orienteers during a training camp. Eighteen elite orienteers participated in a randomized controlled intervention trial during a 1-week training camp (13 exercise sessions). Half of the runners (PRO-CHO) ingested a protein drink before (0.3 g kg(-1)) and a protein-carbohydrate drink after (0.3 g protein kg(-1) and 1 g carbohydrate kg(-1)) each exercise session. The others ingested energy and time-matched carbohydrate drinks (CHO). A 4-km run-test with 20 control points was performed before and on the last day of the intervention. Blood and saliva were obtained in the mornings, before and after run-tests, and after the last training session. During the intervention, questionnaires were fulfilled regarding psychological sense of performance capacity and motivation. PRO-CHO and not CHO improved performance in the 4-km run-test (interaction p < .05). An increase in serum creatine kinase was observed during the week, which was greater in CHO than PRO-CHO (interaction p < .01). Lactate dehydrogenase (p < .001) and cortisol (p = .057) increased during the week, but the change did not differ between groups. Reduction in sense of performance capacity during the intervention was greater in CHO (p < .05) than PRO-CHO. In conclusion, ingestion of whey protein hydrolysate before and after each exercise session improves performance and reduces markers of muscle damage during a strenuous 1-week training camp. The results indicate that protein supplementation in conjunction with each exercise session facilitates the recovery from strenuous training in elite orienteers.

  8. Synergistic inhibition of lipid oxidation by pea protein hydrolysate coupled with licorice extract in a liposomal model system.

    PubMed

    Zhang, Xin; Xiong, Youling L; Chen, Jie; Zhou, Liuming

    2013-09-01

    Fourteen pea protein hydrolysates (PPHs) were prepared using different proteases and tested for antioxidant activity in a liposomal model system under oxidative stress (100 μM FeCl3/2 mM ascorbate). Almost all PPHs inhibited lipid oxidation, and those prepared from heated protein with Flavourzyme (Fla-PPH) or Protamex (Pro-PPH) were the most effective. Remarkable synergistic effects were observed on both Fla-PPH and Pro-PPH with licorice extract (LE). Electron microscopy revealed a self-assembled network that appeared to provide crucial protection of liposome against oxidation. The presence of LE enhanced the antioxidant potential by producing a more compact network apparently via PPH-LE complexation. Zeta-potential measurements suggested electrostatic interactions are important driving forces for the accumulation of active peptides at the liposome interface. Peptides rich in leucine, lysine, glutamic acid, glutamine, valine, or proline with a hydrophobic N-terminus, as identified by mass spectrometry, were implicated in the antioxidative protection. PMID:23924409

  9. Synthesis and in vitro antioxidant functions of protein hydrolysate from backbones of Rastrelliger kanagurta by proteolytic enzymes

    PubMed Central

    Sheriff, Sheik Abdulazeez; Sundaram, Balasubramanian; Ramamoorthy, Baranitharan; Ponnusamy, Ponmurugan

    2013-01-01

    Every year, a huge quantity of fishery wastes and by-products are generated by fish processing industries. These wastes are either underutilized to produce low market value products or dumped leading to environmental issues. Complete utilization of fishery wastes for recovering value added products would be beneficial to the society and individual. The fish protein hydrolysates and derived peptides of fishery resources are widely used as nutritional supplements, functional ingredients, and flavor enhancers in food, beverage and pharmaceutical industries. Antioxidants from fishery resources have attracted the attention of researchers as they are cheaper in cost, easy to derive, and do not have side effects. Thus the present investigation was designed to produce protein hydrolysate by pepsin and papain digestion from the backbones of Rastrelliger kanagurta (Indian mackerel) and evaluate its antioxidant properties through various in vitro assays. The results reveal that both hydrolysates are potent antioxidants, capable of scavenging 46% and 36% of DPPH (1,1-diphenyl-2 picrylhydrazyl) and 58.5% and 37.54% of superoxide radicals respectively. The hydrolysates exhibit significant (p < 0.05) reducing power and lipid peroxidation inhibition. Among the two hydrolysates produced, pepsin derived fraction is superior than papain derived fraction in terms of yield, DH (Degree of hydrolysis), and antioxidant activity. PMID:24596496

  10. Hydrolysed proteins in preterm formula: influence on plasma aminoacids, blood fatty acids and insulinaemia.

    PubMed

    Agosti, M; Pugni, L; Ramenghi, L A; Mosca, F; Marini, A

    2003-09-01

    The data from the present investigation differ from those of the previous study. The new version of hydrolysed protein formula did not induce changes in insulinaemia or in the insulinaemia/glycaemia ratio in pre- and postprandial samples when compared with the intact protein formula. The investigation also confirmed that branch chain aminoacids regulate insulin secretion and that the length of chain is insulinotropic for fatty acids, while the degree of unsaturation is not able to reduce insulin secretion in the newborn baby. This could be caused by stimulation of insulin output activated by the C20:5 n 3; the effects of fatty acids are probably related more to the variable composition of cell membrane than to the fatty acids circulating levels.

  11. Influence of Amino Acid Compositions and Peptide Profiles on Antioxidant Capacities of Two Protein Hydrolysates from Skipjack Tuna (Katsuwonus pelamis) Dark Muscle.

    PubMed

    Chi, Chang-Feng; Hu, Fa-Yuan; Wang, Bin; Li, Zhong-Rui; Luo, Hong-Yu

    2015-04-27

    Influence of amino acid compositions and peptide profiles on antioxidant capacities of two protein hydrolysates from skipjack tuna (Katsuwonus pelamis) dark muscle was investigated. Dark muscles from skipjack tuna were hydrolyzed using five separate proteases, including pepsin, trypsin, Neutrase, papain and Alcalase. Two hydrolysates, ATH and NTH, prepared using Alcalase and Neutrase, respectively, showed the strongest antioxidant capacities and were further fractionated using ultrafiltration and gel filtration chromatography. Two fractions, Fr.A3 and Fr.B2, isolated from ATH and NTH, respectively, showed strong radical scavenging activities toward 2,2-diphenyl-1-picrylhydrazyl radicals (EC50 1.08% ± 0.08% and 0.98% ± 0.07%), hydroxyl radicals (EC50 0.22% ± 0.03% and 0.48% ± 0.05%), and superoxide anion radicals (EC50 1.31% ± 0.11% and 1.56% ± 1.03%) and effectively inhibited lipid peroxidation. Eighteen peptides from Fr.A3 and 13 peptides from Fr.B2 were isolated by reversed-phase high performance liquid chromatography, and their amino acid sequences were determined. The elevated antioxidant activity of Fr.A3 might be due to its high content of hydrophobic and aromatic amino acid residues (181.1 and 469.9 residues/1000 residues, respectively), small molecular sizes (3-6 peptides), low molecular weights (524.78 kDa), and amino acid sequences (antioxidant score 6.11). This study confirmed that a smaller molecular size, the presence of hydrophobic and aromatic amino acid residues, and the amino acid sequences were the key factors that determined the antioxidant activities of the proteins, hydrolysates and peptides. The results also demonstrated that the derived hydrolysates and fractions from skipjack tuna (K. pelamis) dark muscles could prevent oxidative reactions and might be useful for food preservation and medicinal purposes.

  12. Influence of Amino Acid Compositions and Peptide Profiles on Antioxidant Capacities of Two Protein Hydrolysates from Skipjack Tuna (Katsuwonus pelamis) Dark Muscle

    PubMed Central

    Chi, Chang-Feng; Hu, Fa-Yuan; Wang, Bin; Li, Zhong-Rui; Luo, Hong-Yu

    2015-01-01

    Influence of amino acid compositions and peptide profiles on antioxidant capacities of two protein hydrolysates from skipjack tuna (Katsuwonus pelamis) dark muscle was investigated. Dark muscles from skipjack tuna were hydrolyzed using five separate proteases, including pepsin, trypsin, Neutrase, papain and Alcalase. Two hydrolysates, ATH and NTH, prepared using Alcalase and Neutrase, respectively, showed the strongest antioxidant capacities and were further fractionated using ultrafiltration and gel filtration chromatography. Two fractions, Fr.A3 and Fr.B2, isolated from ATH and NTH, respectively, showed strong radical scavenging activities toward 2,2-diphenyl-1-picrylhydrazyl radicals (EC50 1.08% ± 0.08% and 0.98% ± 0.07%), hydroxyl radicals (EC50 0.22% ± 0.03% and 0.48% ± 0.05%), and superoxide anion radicals (EC50 1.31% ± 0.11% and 1.56% ± 1.03%) and effectively inhibited lipid peroxidation. Eighteen peptides from Fr.A3 and 13 peptides from Fr.B2 were isolated by reversed-phase high performance liquid chromatography, and their amino acid sequences were determined. The elevated antioxidant activity of Fr.A3 might be due to its high content of hydrophobic and aromatic amino acid residues (181.1 and 469.9 residues/1000 residues, respectively), small molecular sizes (3–6 peptides), low molecular weights (524.78 kDa), and amino acid sequences (antioxidant score 6.11). This study confirmed that a smaller molecular size, the presence of hydrophobic and aromatic amino acid residues, and the amino acid sequences were the key factors that determined the antioxidant activities of the proteins, hydrolysates and peptides. The results also demonstrated that the derived hydrolysates and fractions from skipjack tuna (K. pelamis) dark muscles could prevent oxidative reactions and might be useful for food preservation and medicinal purposes. PMID:25923316

  13. Purification and identification of adipogenesis inhibitory peptide from black soybean protein hydrolysate.

    PubMed

    Kim, Hyun Jeong; Bae, In Young; Ahn, Chang-Won; Lee, Suyong; Lee, Hyeon Gyu

    2007-11-01

    Adipogenesis inhibitory peptide was isolated and identified from black soybean (Rhynchosia volubilis Lour.) hydrolysate. An adipogenesis inhibitor was purified using consecutive methods including: ultrafiltration (MWCO; 3 and 10kDa), gel filtration chromatography (Superdex Peptide 10/300 GL column), and reverse-phase high-performance liquid chromatography (microBondapak C(18) column). Also, the adipogenesis inhibition effect of the purified peptide was measured by observation of droplet of 3T3-L1 adipocyte by Oil Red O staining in the highest active fraction in each step. The peptide was shown to inhibit the differentiation of the 3T3-L1 pre-adipocyte, which was confirmed by morphological study. The adipogenesis inhibitory peptide was purified 71.43-fold from black soybean hydrolysate throughout a five-step purification procedure. The adipogenesis inhibitor was identified to be a tripeptide, Ile-Gln-Asn, having an IC(50) value of 0.014 mg protein/ml. Furthermore, the synthetic tripeptide (Ile-Gln-Asn) exhibited the similar adipogenesis effects to the purified peptide. Thus, these results showed the potential anti-obesity effect of the purified peptide through control of adiposity.

  14. Sm-like protein enhanced tolerance of recombinant Saccharomyces cerevisiae to inhibitors in hemicellulosic hydrolysate.

    PubMed

    Gao, Lan; Xia, Liming

    2012-11-01

    A current challenge of the cellulosic ethanol industry is to improve the resistance of inhibitors present in biomass hydrolysates. RNA-binding protein gene lsm6 was cloned from industrial Saccharomyces cerevisiae ZU-E8, which is able to conferment glucose and xylose, and transformed into ZU-E8 via expression vector pRS426. The positive transformant ZU-910 with over-expressing lsm6 was identified on the culture plates using high concentration of acetate and re-screened by fermentation test. Fermentation by the recombinants was performed in a medium containing 80 g/L xylose and 2 g/L acetic acid or 20 g/L NH(4)Ac/NaAc. After 96 h shaking-flask fermentation, ZU-910 utilized 90.2% xylose with an ethanol yield of 26.9 g/L, which was 8.5- and 10-fold higher than ZU-E8. Further, in the corn stover hemicellulosic hydrolysate fermentation, both the xylose conversion and ethanol production by ZU-910 was larger by 50% and 40% than ZU-E8. ZU-910 has also enhanced tolerance against furfural and SO(4)(2-).

  15. Fabrication and characterization of the nano-composite of whey protein hydrolysate chelated with calcium.

    PubMed

    Xixi, Cai; Lina, Zhao; Shaoyun, Wang; Pingfan, Rao

    2015-03-01

    The nano-composites of whey protein hydrolysate (WPH) chelated with calcium were fabricated in aqueous solution at 30 °C for 20 min, with the ratio of hydrolysate to calcium 15 : 1 (w/w). UV scanning spectroscopy, fluorescent spectroscopy, Fourier transform infrared spectroscopy, dynamic light scattering and atomic force microscopy were applied to characterize the structure of the WPH-calcium chelate. The nano-composites showed the successful incorporation of calcium into the WPH, indicating the interaction between calcium and WPH. The chelation of calcium ions to WPH caused molecular folding and aggregation which led to the formation of a WPH-calcium chelate of nanoparticle size, and the principal sites of calcium-binding corresponded to the carboxyl groups and carbonyl groups of WPH. The WPH-calcium chelate demonstrated excellent stability and absorbability under both acidic and basic conditions, which was beneficial for calcium absorption in the gastrointestinal tract of the human body. Moreover, the calcium absorption of the WPH-calcium chelate on Caco-2 cells was significantly higher than those of calcium gluconate and CaCl₂ in vitro, suggesting the possible increase in calcium bioavailability. The findings suggest that the WPH-calcium chelate has the potential in making dietary supplements for improving bone health of the human body.

  16. The use of radiolabelling techniques to measure substantivity to, and penetration into, hair of protein hydrolysates.

    PubMed

    Jones, R T; Chahal, S P

    1997-10-01

    The use of 14C-labelled amino acids enables the measurement of both the total substantivity to hair and the degree of penetration into the hair shaft of amino acid mixtures derived from complete hydrolysis of proteins. The technique utilizes the fact that direct measurement of 14C radioactivity of the treated hair detects only the surface substantivity. Total substantivity can be determined following solubilization of the hair. Data obtained for wheat amino acids show significant penetration when used to treat hair from a shampoo or conditioner formulation. A similar technique has been investigated for a wheat protein partial hydrolysate using 14CNO for radiolabelling purposes and shows that significant penetration into hair can occur. L'utilisation d'amino-acides marques au 14C permet la mesure a la fois de l'absorption totale par les cheveux et du degre de penetration dans la fibre du cheveu de melanges d'amino-acides obtenus a partir d'une hydrolyse totale de proteines. La technique utilise le fait que la mesure directe de la radioactivite du 14C des cheveux traites ne detecte que l'absorption en surface. L'absorption totale peut etre determinee apres solubilisation des cheveux. Les donnees obtenues a partir d'amino-acides de ble montrent une penetration significative lors d'une utilisation pour traiter des cheveux a partir d'une formulation de shampoing ou d'apres-shampoing. Une technique similaire a ete exploree vis-a-vis d'un hydrolysat partiel d'une proteine de ble utilisant 14CNO a des fins de marquage, et montre qu'une penetration significative dans les cheveux peut avoir lieu. PMID:18507628

  17. Antioxidant Activities of Hydrolysates of Arca Subcrenata Prepared with Three Proteases

    PubMed Central

    Song, Liyan; Li, Tingfei; Yu, Rongmin; Yan, Chunyan; Ren, Shengfang; Zhao, Yu

    2008-01-01

    In order to get products with antioxidant activity from Arca subcrenata Lischke, the optimal hydrolase and hydrolysis conditions were investigated in the paper. Three proteases (neutrase, alcalase and papain) were applied to hydrolyze the homogenate of A. subcrenata. An orthogonal design was used to optimize hydrolysis conditions, and the pH-stat methods was used to determine the degree of hydrolysis. Viewed from the angle of reducing power, such as scavenging activities against α,α-diphenyl-β-picrylhydrazyl (DPPH) radical and hydrogen peroxide, the antioxidant activities of the alcalase hydrolysate (AH) were superior to neutrase hydrolysate (NH) and papain hydrolysate (PH), and its EC50 values in DPPH radical and hydrogen peroxide scavenging effect were 6.23 mg/ml and 19.09 mg/ml, respectively. Moreover, compared with products hydrolyzed by neutrase and papain, the molecular mass of AH was lower and its content of amino acid of peptides was higher. Therefore, alcalase was selected as the optimal enzyme to produce active ingredients since its hydrolysate exhibited the best antioxidant activity among them and possessed large amount of potential active peptides. PMID:19172198

  18. Preparation of squid skin collagen hydrolysate as an antihyaluronidase, antityrosinase, and antioxidant agent.

    PubMed

    Nakchum, Ladawan; Kim, Sang Moo

    2016-01-01

    A collagen was isolated from squid skin, a processing waste product. The biofunctional activities of enzymatic squid skin collagen hydrolysates were determined to produce a value-added material. Five low-molecular-mass hydrolysate fractions, F1 (>30 kD), F2 (10-30 kD), F3 (3-10 kD), F4 (1-3 kD), and F5 (<1 kD), were manufactured from its enzymatic hydrolysate by ultrafiltration. Fraction F3 had the strongest antihyaluronidase inhibitory activity. Gly, Val, and Pro were major amino acids in F3, while Met, Tyr, and His were minor ones. The molecular mass of F3 was in the range of 3.4 to 10 kD. F3 exhibited copper chelating ability in a concentration-dependent manner. The ferrous chelating ability of F3 was almost 50% at 200 µg/mL. F3 also inhibited tyrosinase activity by 39.65% at 1 mg/mL. Furthermore, F3 had stronger hydroxyl radical scavenging activity (IC50 = 149.94 µg/mL) than ascorbic acid (IC50 = 212.94 µg/mL). Therefore, the squid collagen hydrolysate can be utilized as a nutraceutical or cosmeceutical agent.

  19. Reduction of L-phenylalanine in protein hydrolysates using L-phenylalanine ammonia-lyase from Rhodosporidium toruloides.

    PubMed

    Castañeda, María Teresita; Adachi, Osao; Hours, Roque Alberto

    2015-10-01

    L-Phenylalanine ammonia-lyase (PAL, EC 4.3.1.25) from Rhodosporidium toruloides was utilized to remove L-phenylalanine (L-Phe) from different commercial protein hydrolysates. A casein acid hydrolysate (CAH, L-Phe ~2.28 %) was employed as a model substrate. t-Cinnamic acid resulting from deamination of L-Phe was extracted, analyzed at λ = 290 nm, and used for PAL activity determination. Optimum reaction conditions, optimized using successive Doehlert design, were 35 mg mL(-1) of CAH and 800 mU mL(-1) of PAL, while temperature and pH were 42 °C and 8.7, respectively. Reaction kinetics of PAL with CAH was determined under optimized conditions. Then, removal of L-Phe from CAH was tested. Results showed that more than 92 % of initial L-Phe was eliminated. Similar results were obtained with other protein hydrolysates. These findings demonstrate that PAL is a useful biocatalyst for L-Phe removal from protein hydrolysates, which can be evaluated as potential ingredients in foodstuffs for PKU patients.

  20. Soybean plant growth study conducted using purified protein hydrolysate-based fertilizer made from chrome-tanned leather waste.

    PubMed

    Pati, Anupama; Chaudhary, Rubina

    2015-12-01

    Leather processing discharges enormous amount of chrome containing leather solid waste which creates a major disposal problem. Chrome-tanned leather solid waste is a complex of collagen and chromium. The presence of chromium limits protein application in fertilizer industry. The purified protein hydrolysate with zero chromium could be used as a nitrogen source for fertilizer formulation. In this study, an attempt has been made to employ purified protein hydrolysate derived from chrome-tanned leather shavings (CTLS) in formulation of fertilizer. The formulated fertilizer (1–3 t ha(-1)) is employed as nitrogen source in production of soybean. Plant growth study demonstrates that formulated fertilizer dosage 3 t ha(-1) produced similar effects of commercial fertilizer-treated plants. Application of formulated fertilizer yielded higher seed in plant than commercial fertilizer.

  1. Effect of oven drying and freeze drying on the antioxidant and functional properties of protein hydrolysates derived from freshwater fish (Cirrhinus mrigala) using papain enzyme.

    PubMed

    Elavarasan, Krishnamoorthy; Shamasundar, Bangalore Aswathnarayan

    2016-02-01

    Fish protein hydrolysate (FPH) was prepared from fresh water fish Cirrhinus mrigala using papain and dried in oven (OD-FPH) and freeze dryer (FD-FPH). The electron micrographs of FD-FPH samples showed porous structure. The browning intensity of OD-FPH samples was higher than the FD-FPH samples. The DPPH (2, 2 Diphenyl-1-picrylhydrazyl) free radical scavenging activity and linoleic acid peroxidation inhibition activity of FPH were not affected by oven drying process. The sequential digestion of FPH with pepsin and pancreatin reduced the antioxidant properties in both OD-FPH and FD-FPH samples. The solubility of proteins in OD-FPH was lower at pH 5 while for that of FD-FPH it was at pH 7 with water as solvent. The surface active properties of FD-FPH samples were higher than OD-FPH samples. The oven drying of fish protein hydrolysates may be advocated considering the properties and cost of production. PMID:27162411

  2. Protein enrichment of an Opuntia ficus-indica cladode hydrolysate by cultivation of Candida utilis and Kluyveromyces marxianus

    PubMed Central

    Akanni, Gabriel B; du Preez, James C; Steyn, Laurinda; Kilian, Stephanus G

    2015-01-01

    BACKGROUND The cladodes of Opuntia ficus-indica (prickly pear cactus) have a low protein content; for use as a balanced feed, supplementation with other protein sources is therefore desirable. We investigated protein enrichment by cultivation of the yeasts Candida utilis and Kluyveromyces marxianus in an enzymatic hydrolysate of the cladode biomass. RESULTS Dilute acid pretreatment and enzymatic hydrolysis of sun-dried cladodes resulted in a hydrolysate containing (per litre) 45.5 g glucose, 6.3 g xylose, 9.1 g galactose, 10.8 g arabinose and 9.6 g fructose. Even though K. marxianus had a much higher growth rate and utilized l-arabinose and d-galactose more completely than C. utilis, its biomass yield coefficient was lower due to ethanol and ethyl acetate production despite aerobic cultivation. Yeast cultivation more than doubled the protein content of the hydrolysate, with an essential amino acid profile superior to sorghum and millet grains. CONCLUSIONS This K. marxianus strain was weakly Crabtree positive. Despite its low biomass yield, its performance compared well with C. utilis. This is the first report showing that the protein content and quality of O. ficus-indica cladode biomass could substantially be improved by yeast cultivation, including a comparative evaluation of C. utilis and K. marxianus. © 2014 The Authors. Journal of the Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry. PMID:25371280

  3. Protective effect of soluble eggshell membrane protein hydrolysate on cardiac ischemia/reperfusion injury

    PubMed Central

    Yang, Tao; Li, Yan; Ma, Meihu; Lin, Qinlu; Sun, Shuguo; Zhang, Bin; Feng, Xi; Liu, Junwen

    2015-01-01

    Background Soluble eggshell membrane protein (SEP) has been proved to hold the antioxidant activity. The functional role of SEP on cardioprotection was investigated in vivo and in vitro. Methods Rats and cardiomyocytes were pretreated with SP2, a hydrolysate attained from SEP, and then subjected to ischemia/reperfusion (I/R) or hypoxia/reoxygenation (H/R) and hydrogen peroxide, respectively. The measurement of myocardial infarct size, cell apoptosis assay, cell viability assay, and caspase activity assay were performed on rats and cardiomyocytes. Results The results showed that the treatment of SP2 induced the resistance to I/R or H/R injury on rats and cardiomyocytes as indicated by decreased infarct size and decreased cellular apoptosis. The cardioprotective roles of SP2 were partly resulted from the downregulated expression and activity of caspase-3 in which the effect was similar to the caspase inhibitor, z-VAD-fmk, and could be rescued by caspase activator, PAC-1. Conclusions This investigation has demonstrated that SP2 attenuated the damage of I/R and H/R on rats and cardiomyocytes by the caspase-dependent pathway. This cardioprotective effect of SP2 suggested a novel therapeutic agent of SEP for ischemic-related heart diseases. PMID:26699793

  4. Determination of free L- and D-alanine in hydrolysed protein fertilisers by capillary electrophoresis.

    PubMed

    Cavani, Luciano; Ciavatta, Claudio; Gessa, Carlo

    2003-01-24

    of racemisation of hydrolysed protein fertilisers (HPFs) using an The objective of this study was to determine the degree inexpensive and easy to handle analytical method for qualitative control of the products. Using a polyacrylamide coated capillary and a run buffer containing 0.1 M Tris-borate+2.5 mM EDTA-Na2+0.1% sodium dodecylsulfate+10 mM beta-cyclodextrin a quantitative separation of D- and L-alanine (Ala) was made from an not treated HPF sample derivatised with dansyl chlorine by capillary electrophoresis. The D-Ala:[D-Ala+L-Ala] ratio, called degree of racemisation (RD), was calculated. The analysis of ten commercial HPFs has shown that more than 60% of HPFs have an RD > or = 40%. while only one product has shown an RD <5%. These results showed that most of the HPFs on the market are obtained with strong hydrolytic processes and high contents of D-amino acids are probably less effective as plant nutrients or even potentially dangerous to plants.

  5. Regulatory effect of porcine plasma protein hydrolysates on pasting and gelatinization action of corn starch.

    PubMed

    Kong, Baohua; Niu, Haili; Sun, Fangda; Han, Jianchun; Liu, Qian

    2016-01-01

    The objective of this study was to investigate the regulatory effect of porcine plasma protein hydrolysates (PPPH) on the physicochemical, pasting, and gelatinization properties of corn starch (CS). The results showed that the solubility of CS markedly increased, whereas swelling power and gel penetration force decreased with increased PPPH concentration (P<0.05). Compared with native CS, PPPH significantly lowered peak viscosity, minimum viscosity, final viscosity, and total setback, whereas it increased breakdown and pasting temperature in rapid visco analyzer (RVA) measurement (P<0.05) and obviously enhanced the gelatinization temperature as determined in differential scanning calorimetry (DSC) (P<0.05). Confocal laser scanning microscopy (CLSM) showed that PPPH surrounded the starch granules at room temperature (25°C) and then formed a network with swollen starch granules during gelatinization. Atomic force microscopy (AFM) images indicated that the blocklet sizes of gelatinized CS-PPPH mixtures were smaller and more uniform than native CS. The results proved that pasting and gelatinization abilities of CS can be effectively influenced by adding PPPH.

  6. A novel hemp seed meal protein hydrolysate reduces oxidative stress factors in spontaneously hypertensive rats.

    PubMed

    Girgih, Abraham T; Alashi, Adeola M; He, Rong; Malomo, Sunday A; Raj, Pema; Netticadan, Thomas; Aluko, Rotimi E

    2014-12-01

    This report shows the antioxidant effects of a hemp seed meal protein hydrolysate (HMH) in spontaneously hypertensive rats (SHR). Defatted hemp seed meal was hydrolyzed consecutively with pepsin and pancreatin to yield HMH, which was incorporated into rat feed as a source of antioxidant peptides. Young (8-week old) SHRs were divided into three groups (8 rats/group) and fed diets that contained 0.0%, 0.5% or 1.0% (w/w) HMH for eight weeks; half of the rats were sacrificed for blood collection. After a 4-week washout period, the remaining 20-week old SHRs were fed for an additional four weeks and sacrificed for blood collection. Plasma total antioxidant capacity (TAC) and superoxide dismutase (SOD), catalase (CAT) and total peroxides (TPx) levels were determined. Results showed that plasma TAC, CAT and SOD levels decreased in the older 20-week old SHRs when compared to the young SHRs. The presence of HMH in the diets led to significant (p < 0.05) increases in plasma SOD and CAT levels in both young and adult SHR groups; these increases were accompanied by decreases in TPx levels. The results suggest that HMH contained antioxidant peptides that reduced the rate of lipid peroxidation in SHRs with enhanced antioxidant enzyme levels and total antioxidant capacity.

  7. A Novel Hemp Seed Meal Protein Hydrolysate Reduces Oxidative Stress Factors in Spontaneously Hypertensive Rats

    PubMed Central

    Girgih, Abraham T.; Alashi, Adeola M.; He, Rong; Malomo, Sunday A.; Raj, Pema; Netticadan, Thomas; Aluko, Rotimi E.

    2014-01-01

    This report shows the antioxidant effects of a hemp seed meal protein hydrolysate (HMH) in spontaneously hypertensive rats (SHR). Defatted hemp seed meal was hydrolyzed consecutively with pepsin and pancreatin to yield HMH, which was incorporated into rat feed as a source of antioxidant peptides. Young (8-week old) SHRs were divided into three groups (8 rats/group) and fed diets that contained 0.0%, 0.5% or 1.0% (w/w) HMH for eight weeks; half of the rats were sacrificed for blood collection. After a 4-week washout period, the remaining 20-week old SHRs were fed for an additional four weeks and sacrificed for blood collection. Plasma total antioxidant capacity (TAC) and superoxide dismutase (SOD), catalase (CAT) and total peroxides (TPx) levels were determined. Results showed that plasma TAC, CAT and SOD levels decreased in the older 20-week old SHRs when compared to the young SHRs. The presence of HMH in the diets led to significant (p < 0.05) increases in plasma SOD and CAT levels in both young and adult SHR groups; these increases were accompanied by decreases in TPx levels. The results suggest that HMH contained antioxidant peptides that reduced the rate of lipid peroxidation in SHRs with enhanced antioxidant enzyme levels and total antioxidant capacity. PMID:25493943

  8. Roe Protein Hydrolysates of Giant Grouper (Epinephelus lanceolatus) Inhibit Cell Proliferation of Oral Cancer Cells Involving Apoptosis and Oxidative Stress

    PubMed Central

    Yang, Jing-Iong; Tang, Jen-Yang; Liu, Ya-Sin; Wang, Hui-Ru; Lee, Sheng-Yang; Yen, Ching-Yu

    2016-01-01

    Roe protein hydrolysates were reported to have antioxidant property but the anticancer effects were less addressed, especially for oral cancer. In this study, we firstly used the ultrafiltrated roe hydrolysates (URH) derived from giant grouper (Epinephelus lanceolatus) to evaluate the impact of URH on proliferation against oral cancer cells. We found that URH dose-responsively reduced cell viability of two oral cancer cells (Ca9-22 and CAL 27) in terms of ATP assay. Using flow cytometry, URH-induced apoptosis of Ca9-22 cells was validated by morphological features of apoptosis, sub-G1 accumulation, and annexin V staining in dose-responsive manners. URH also induced oxidative stress in Ca9-22 cells in terms of reactive oxygen species (ROS)/superoxide generations and mitochondrial depolarization. Taken together, these data suggest that URH is a potential natural product for antioral cancer therapy. PMID:27195297

  9. Differential Growth Patterns Among Healthy Infants Fed Protein Hydrolysate or Cow-Milk Formulas

    PubMed Central

    Ventura, Alison K.; Beauchamp, Gary K.

    2011-01-01

    OBJECTIVE: Infant formulas differ considerably in composition and sensory profiles. In this randomized study, we examined whether healthy infants fed an extensively protein hydrolysate formula (PHF) would differ in feeding behavior and growth from those fed cow-milk formula (CMF). PATIENTS AND METHODS: Infants were randomly assigned to be fed CMF or PHF between 0.5 and 7.5 months of age. Each month for 7 months, infants were weighed and measured and then videotaped while being fed their assigned formula. Anthropometric z scores were calculated by using World Health Organization growth standards. Multilevel linear growth and piecewise mixed-effects models compared trajectories for growth measures and formula acceptance. RESULTS: When compared with infants fed CMF, infants fed PHF had significantly lower weight-for-length z scores across ages 2.5 to 7.5 months. There were no differences in length-for-age z scores, which indicate that group differences resulted from gains in weight, not length. Infants fed PHF also had significantly slower weight gain velocity compared with infants fed CMF. During the monthly assessments, PHF-fed infants consumed less formula to satiation than did CMF-fed infants across the study period. Maternal ratings of infants' acceptance of the formula did not differ at any age. CONCLUSIONS: z-score trajectories indicate that CMF-fed infants' weight gain was accelerated, whereas PHF-fed infants' weight gain was normative. Whether such differences in growth are because of differences in the protein content or amino acid profile of the formulas and, in turn, metabolism is unknown. Research on the long-term consequences of these early growth differences is needed. PMID:21187303

  10. Antihypertensive and free radical scavenging properties of enzymatic rapeseed protein hydrolysates.

    PubMed

    He, Rong; Alashi, Adeola; Malomo, Sunday A; Girgih, Abraham T; Chao, Dongfang; Ju, Xingrong; Aluko, Rotimi E

    2013-11-01

    In this study, rapeseed protein isolate (RPI) was digested with various proteases to produce rapeseed protein hydrolysates (RPHs), which were then separated into different peptide fractions (<1, 1-3, 3-5, and 5-10kDa) by membrane ultrafiltration. Membrane fractionation showed that peptides with sizes <3 kDa had significantly (p<0.05) reduced surface hydrophobicity when compared to the RPHs and peptide fractions with sizes >3 kDa. In contrast, the <3 kDa peptides showed significantly (p<0.05) higher oxygen radical scavenging ability when compared to the >3 kDa peptides and RPHs. In vitro inhibition of angiotensin I-converting enzyme (ACE) was significantly (p<0.05) higher for the Thermolysin, Proteinase K and Alcalase RPHs when compared to the pepsin+pancreatin (PP) and Flavourzyme RPHs. The Alcalase RPH had significantly (p<0.05) higher renin inhibition among the RPHs, while with the exception of Thermolysin, the 5-10 kDa peptide fraction had the least renin-inhibitory ability when compared to the <5 kDa peptide fractions. Oral administration (100mg/kg body weight) of the RPHs and RPI to spontaneously hypertensive rats (SHR) showed the Alcalase RPH to be the most effective in blood pressure (BP) reduction (∼24 mm Hg) while Proteinase K RPH was the least effective (∼5 mm Hg) after 8h. However, the PP RPH had the most prolonged effect with BP reduction of ∼20 mm Hg after 24h of oral administration. We conclude that the strong BP-lowering ability of Alcalase and PP RPHs could be due to high resistance of the peptides to structural degradation coupled with high absorption rate within the gastrointestinal tract.

  11. Antioxidant and ACE-inhibitory activities of hemp (Cannabis sativa L.) protein hydrolysates produced by the proteases AFP, HT, Pro-G, actinidin and zingibain.

    PubMed

    Teh, Sue-Siang; Bekhit, Alaa El-Din A; Carne, Alan; Birch, John

    2016-07-15

    Hemp protein isolates (HPIs) were hydrolysed by proteases (AFP, HT, ProG, actinidin and zingibain). The enzymatic hydrolysis of HPIs was evaluated through the degree of hydrolysis and SDS-PAGE profiles. The bioactive properties of the resultant hydrolysates (HPHs) were accessed through ORAC, DPPḢ scavenging and ACE-inhibitory activities. The physical properties of the resultant HPHs were evaluated for their particle sizes, zeta potential and surface hydrophobicity. HT had the highest rate of caseinolytic activity at the lowest concentration (0.1 mg mL(-1)) compared to other proteases that required concentration of 100 mg mL(-1) to achieve their maximum rate of caseinolytic activity. This led to the highest degree of hydrolysis of HPIs by HT in the SDS-PAGE profiles. Among all proteases and substrates, HT resulted in the highest bioactivities (ORAC, DPPḢ scavenging and ACE-inhibitory activities) generated from alkali extracted HPI in the shortest time (2 h) compared to the other protease preparations.

  12. Antioxidant and ACE-inhibitory activities of hemp (Cannabis sativa L.) protein hydrolysates produced by the proteases AFP, HT, Pro-G, actinidin and zingibain.

    PubMed

    Teh, Sue-Siang; Bekhit, Alaa El-Din A; Carne, Alan; Birch, John

    2016-07-15

    Hemp protein isolates (HPIs) were hydrolysed by proteases (AFP, HT, ProG, actinidin and zingibain). The enzymatic hydrolysis of HPIs was evaluated through the degree of hydrolysis and SDS-PAGE profiles. The bioactive properties of the resultant hydrolysates (HPHs) were accessed through ORAC, DPPḢ scavenging and ACE-inhibitory activities. The physical properties of the resultant HPHs were evaluated for their particle sizes, zeta potential and surface hydrophobicity. HT had the highest rate of caseinolytic activity at the lowest concentration (0.1 mg mL(-1)) compared to other proteases that required concentration of 100 mg mL(-1) to achieve their maximum rate of caseinolytic activity. This led to the highest degree of hydrolysis of HPIs by HT in the SDS-PAGE profiles. Among all proteases and substrates, HT resulted in the highest bioactivities (ORAC, DPPḢ scavenging and ACE-inhibitory activities) generated from alkali extracted HPI in the shortest time (2 h) compared to the other protease preparations. PMID:26948606

  13. Angiotensin I-Converting Enzyme (ACE) Inhibitory Activity and ACE Inhibitory Peptides of Salmon (Salmo salar) Protein Hydrolysates Obtained by Human and Porcine Gastrointestinal Enzymes

    PubMed Central

    Darewicz, Małgorzata; Borawska, Justyna; Vegarud, Gerd E.; Minkiewicz, Piotr; Iwaniak, Anna

    2014-01-01

    The objectives of the present study were two-fold: first, to detect whether salmon protein fractions possess angiotensin I-converting enzyme (ACE) inhibitory properties and whether salmon proteins can release ACE inhibitory peptides during a sequential in vitro hydrolysis (with commercial porcine enzymes) and ex vivo digestion (with human gastrointestinal enzymes). Secondly, to evaluate the ACE inhibitory activity of generated hydrolysates. A two-step ex vivo and in vitro model digestion was performed to simulate the human digestion process. Salmon proteins were degraded more efficiently by porcine enzymes than by human gastrointestinal juices and sarcoplasmic proteins were digested/hydrolyzed more easily than myofibrillar proteins. The ex vivo digested myofibrillar and sarcoplasmic duodenal samples showed IC50 values (concentration required to decrease the ACE activity by 50%) of 1.06 and 2.16 mg/mL, respectively. The in vitro hydrolyzed myofibrillar and sarcoplasmic samples showed IC50 values of 0.91 and 1.04 mg/mL, respectively. Based on the results of in silico studies, it was possible to identify 9 peptides of the ex vivo hydrolysates and 7 peptides of the in vitro hydrolysates of salmon proteins of 11 selected peptides. In both types of salmon hydrolysates, ACE-inhibitory peptides IW, IY, TVY and VW were identified. In the in vitro salmon protein hydrolysates an ACE-inhibitory peptides VPW and VY were also detected, while ACE-inhibitory peptides ALPHA, IVY and IWHHT were identified in the hydrolysates generated with ex vivo digestion. In our studies, we documented ACE inhibitory in vitro effects of salmon protein hydrolysates obtained by human and as well as porcine gastrointestinal enzymes. PMID:25123137

  14. Angiotensin I-converting enzyme (ACE) inhibitory activity and ACE inhibitory peptides of salmon (Salmo salar) protein hydrolysates obtained by human and porcine gastrointestinal enzymes.

    PubMed

    Darewicz, Małgorzata; Borawska, Justyna; Vegarud, Gerd E; Minkiewicz, Piotr; Iwaniak, Anna

    2014-08-13

    The objectives of the present study were two-fold: first, to detect whether salmon protein fractions possess angiotensin I-converting enzyme (ACE) inhibitory properties and whether salmon proteins can release ACE inhibitory peptides during a sequential in vitro hydrolysis (with commercial porcine enzymes) and ex vivo digestion (with human gastrointestinal enzymes). Secondly, to evaluate the ACE inhibitory activity of generated hydrolysates. A two-step ex vivo and in vitro model digestion was performed to simulate the human digestion process. Salmon proteins were degraded more efficiently by porcine enzymes than by human gastrointestinal juices and sarcoplasmic proteins were digested/hydrolyzed more easily than myofibrillar proteins. The ex vivo digested myofibrillar and sarcoplasmic duodenal samples showed IC50 values (concentration required to decrease the ACE activity by 50%) of 1.06 and 2.16 mg/mL, respectively. The in vitro hydrolyzed myofibrillar and sarcoplasmic samples showed IC50 values of 0.91 and 1.04 mg/mL, respectively. Based on the results of in silico studies, it was possible to identify 9 peptides of the ex vivo hydrolysates and 7 peptides of the in vitro hydrolysates of salmon proteins of 11 selected peptides. In both types of salmon hydrolysates, ACE-inhibitory peptides IW, IY, TVY and VW were identified. In the in vitro salmon protein hydrolysates an ACE-inhibitory peptides VPW and VY were also detected, while ACE-inhibitory peptides ALPHA, IVY and IWHHT were identified in the hydrolysates generated with ex vivo digestion. In our studies, we documented ACE inhibitory in vitro effects of salmon protein hydrolysates obtained by human and as well as porcine gastrointestinal enzymes.

  15. Fish protein hydrolysates: application in deep-fried food and food safety analysis.

    PubMed

    He, Shan; Franco, Christopher; Zhang, Wei

    2015-01-01

    Four different processes (enzymatic, microwave-intensified enzymatic, chemical, and microwave-intensified chemical) were used to produce fish protein hydrolysates (FPH) from Yellowtail Kingfish for food applications. In this study, the production yield and oil-binding capacity of FPH produced from different processes were evaluated. Microwave intensification significantly increased the production yields of enzymatic process from 42% to 63%. It also increased the production yields of chemical process from 87% to 98%. The chemical process and microwave-intensified chemical process produced the FPH with low oil-binding capacity (8.66 g oil/g FPH and 6.25 g oil/g FPH), whereas the microwave-intensified enzymatic process produced FPH with the highest oil-binding capacity (16.4 g oil/g FPH). The FPH from the 4 processes were applied in the formulation of deep-fried battered fish and deep-fried fish cakes. The fat uptake of deep-fried battered fish can be reduced significantly from about 7% to about 4.5% by replacing 1% (w/w) batter powder with FPH, and the fat uptake of deep-fried fish cakes can be significantly reduced from about 11% to about 1% by replacing 1% (w/w) fish mince with FPH. Food safety tests of the FPH produced by these processes demonstrated that the maximum proportion of FPH that can be safely used in food formulation is 10%, due to its high content of histamine. This study demonstrates the value of FPH to the food industry and bridges the theoretical studies with the commercial applications of FPH.

  16. Determination of Free-Form and Peptide Bound Pyrraline in the Commercial Drinks Enriched with Different Protein Hydrolysates

    PubMed Central

    Liang, Zhili; Li, Lin; Qi, Haiping; Zhang, Xia; Xu, Zhenbo; Li, Bing

    2016-01-01

    Pyrraline, a causative factor for the recent epidemics of diabetes and cardiovascular disease, is also employed as an indicator to evaluate heat damage and formation of advanced glycation end-products (AGEs) in foods. Peptide-enriched drinks (PEDs) are broadly consumed worldwide due to rapid rate of absorption and perceived health effects. It can be hypothesized that PED is an important source of pyrraline, especially peptide bound pyrraline (Pep-Pyr). In this study we determined free-form pyrraline (Free-Pyr) and Pep-Pyr in drinks enriched with whey protein hydrolysate (WPH), soy protein hydrolysate (SPH) and collagen protein hydrolysate (CPH). A detection method was developed using ultrahigh-performance liquid chromatography with UV-visible detector coupled with tandem mass spectrometry after solid-phase extraction (SPE). The SPE led to excellent recovery rates ranging between 93.2% and 98.5% and a high reproducibility with relative standard deviations (RSD) of <5%. The limits of detection and quantification obtained were 30.4 and 70.3 ng/mL, respectively. Pep-Pyr was identified as the most abundant form (above 96 percent) of total pyrraline, whereas Free-Pyr was present in a small proportion (less than four percent) of total pyrraline. The results indicate that PED is an important extrinsic source of pyrraline, especially Pep-Pyr. As compared with CPH- and SPH-enriched drinks, WPH-enriched drinks contained high content of Pep-Pyr. The Pep-Pyr content is associated with the distribution of peptide lengths and the amino acid compositions of protein in PEDs. PMID:27384561

  17. Antioxidant properties of enzymatic hydrolysates from royal jelly.

    PubMed

    Nagai, Takeshi; Inoue, Reiji; Suzuki, Nobutaka; Nagashima, Toshio

    2006-01-01

    Enzymatic hydrolysates were prepared from royal jelly using three enzymes (pepsin, trypsin, and papain), and their antioxidative properties were evaluated. The yield of these hydrolysates was very high, about 20-26% on a raw weight basis. In comparison with the antioxidative activities of water extract and alkaline extract of royal jelly, the antioxidative activities and scavenging activities against active oxygen species such as superoxide anion radical and hydroxyl radical of each hydrolysate were high in the sample with a low protein concentration. These results suggest that once royal jelly is hydrolyzed using enzyme, the hydrolysate possesses much higher antioxidative activity and scavenging activity against active oxygen species. Royal jelly will act as a medicinal food in the human body. PMID:17004899

  18. The effect of a partially hydrolysed formula based on rice protein in the treatment of infants with cow’s milk protein allergy

    PubMed Central

    Reche, M; Pascual, C; Fiandor, A; Polanco, I; Rivero-Urgell, M; Chifre, R; Johnston, S; Martín-Esteban, M

    2010-01-01

    Reche M, Pascual C, Fiandor A, Polanco I, Rivero-Urgell M, Chifre R, Johnston S, Martín-Esteban M. The effect of a partially hydrolysed formula based on rice protein in the treatment of infants with cow’s milk protein allergy. Pediatr Allergy Immunol 2010: 21: 577–585. © 2010 John Wiley & Sons A/S Infants diagnosed with allergy to cow’s milk protein (CMP) are fed extensively hydrolysed cow’s milk formulas, modified soy formulas or even amino acid-based formulas. Hydrolysed rice protein infant formulas have become available and have been shown to be well tolerated by these infants. A prospective open, randomized clinical study to compare the clinical tolerance of a new hydrolysed rice protein formula (HRPF) with an extensively hydrolysed CMP formula (EHF) in the feeding of infants with IgE-mediated cow’s milk allergy. Ninety-two infants (46 boys and 46 girls, mean age 4.3 months, range 1.1–10.1 months) diagnosed with IgE-mediated cow’s milk allergy were enrolled in the study. Clinical tolerance to the formula products was tested. Clinical evaluation included skin prick tests with whole cow’s milk, soya and rice as well as antigens of CMP (beta-lactoglobulin, alpha-lactalbumin, casein and bovine seroalbumin), HRPF and EHF and specific IgE determinations to CMP using CAP technology. Patients were randomized to receive either an EHF based on CMP or a new HRPF. Follow-up was at 3, 6, 12, 18 and 24 months. Growth parameters were measured at each visit. One infant showed immediate allergic reaction to EHF, but no reaction was shown by any infant in the HRPF group. The number of infants who did not become tolerant to CMP during the study was not statistically different between the two groups. Measurement of IgE levels of infants allergic to CMP during the study showed no significant differences between the two formula groups. Growth parameters were in the normal range and similar between groups. In this study, the HRPF was well tolerated by infants with

  19. Purification and characterisation of a glutamic acid-containing peptide with calcium-binding capacity from whey protein hydrolysate.

    PubMed

    Huang, Shun-Li; Zhao, Li-Na; Cai, Xixi; Wang, Shao-Yun; Huang, Yi-Fan; Hong, Jing; Rao, Ping-Fan

    2015-02-01

    The bioavailability of dietary ionised calcium is affected by intestinal basic environment. Calcium-binding peptides can form complexes with calcium to improve its absorption and bioavailability. The aim of this study was focused on isolation and characterisation of a calcium-binding peptide from whey protein hydrolysates. Whey protein was hydrolysed using Flavourzyme and Protamex with substrate to enzyme ratio of 25:1 (w/w) at 49 °C for 7 h. The calcium-binding peptide was isolated by DEAE anion-exchange chromatography, Sephadex G-25 gel filtration and reversed phase high-performance liquid chromatography (RP-HPLC). A purified peptide of molecular mass 204 Da with strong calcium binding ability was identified on chromatography/electrospray ionisation (LC/ESI) tandem mass spectrum to be Glu-Gly (EG) after analysis and alignment in database. The calcium binding capacity of EG reached 67·81 μg/mg, and the amount increased by 95% compared with whey protein hydrolysate complex. The UV and infrared spectrometer analysis demonstrated that the principal sites of calcium-binding corresponded to the carboxyl groups and carbonyl groups of glutamic acid. In addition, the amino group and peptide amino are also the related groups in the interaction between EG and calcium ion. Meanwhile, the sequestered calcium percentage experiment has proved that EG-Ca is significantly more stable than CaCl2 in human gastrointestinal tract in vitro. The findings suggest that the purified dipeptide has the potential to be used as ion-binding ingredient in dietary supplements.

  20. [Complex of zinc with enzymatic hydrolysate of pigspleen protein - in vivo investigation].

    PubMed

    Zorin, S N; Sidorova, Iu S; Zilova, I S; Mazo, V K

    2014-01-01

    Experimental sample of complex zinc with enzymatic hydrolysate of pigspleen protein (Zn-EHPS) was produced in laboratory with ultra- and nanofiltration technologies; its composition was analyzed; male rats Wister (initial body weight 132±2,4 g) were under observation in vivo: rats weregiven zinc-deficient ration (ZDR) "Zinc Deficient Diet, Egg White Base", containing not more than 1,3 mg zinc/kg, and water ad libitum during 18 days. Animals were divided into 3 groups (10 in each): group I was given ZDR and distilled water; group 2 - ZDR and water zinc sulphate solution (ZnSO4) with zinc concentration of 16,3 mcg/ml; group 3 - ZDR and water Zn-EHPS solution with zinc concentration of 16,3 mcg/ml. All rats were daily observed with registration of water consumption. Unconditioned reflex (photophobia) or crossing time from light to darkness on special system before the beginning of experiment was tested out. An increase of body weight (ΔW) during the experiment, unconditioned reflex stability and training to conditioned passive avoidance reflex (CPAR) after paw painful electric stimulation were studied. On 18 day in 16 hours after paw painful electric stimulation alkaline phosphatase activity (AP) and corticosterone concentration in rat blood serum were examined. Insignificant changes in distribution of peptide fractions according its molecular mass and practical complete remove of free metal ions from final product were observed when Zn-EHPS-technology were used. During experiment in all groups correspondently ΔW was 23,87 and 82% (p<0,05); AP activity - 1,70±0,06; 3,10±0,25; 4,19± 0,26 mcmol/l/s (p<0,01); corticosterone concentration - 20,7±7,2; 21,5±6,3; 22,1±16,5 ng/ml (p>0,1) were observed. Significant (p<0,05) worse change of unconditioned reflex (photophobia) in group 1 (ZDR) in comparison with group 2 and group 3 (latent period - LP, s: 50,9±12,0; 18,2±16,0; 16,0±4,2) as well as to initial test (LP, s - 16,8±8,0) was found on the 14th day of the

  1. Production of bioactive peptide hydrolysates from deer, sheep, pig and cattle red blood cell fractions using plant and fungal protease preparations.

    PubMed

    Bah, Clara S F; Carne, Alan; McConnell, Michelle A; Mros, Sonya; Bekhit, Alaa El-Din A

    2016-07-01

    Protease preparations from plant (papain and bromelain) and fungal (FP400 and FPII) sources were used to hydrolyze the red blood cell fractions (RBCFs) separated from deer, sheep, pig, and cattle abattoir-sourced blood. After 1, 2, 4 and 24h of hydrolysis, the antioxidant and antibacterial activities of the peptide hydrolysates obtained were investigated. The increase in trichloroacetic acid-soluble peptides over the hydrolysis period was examined using the o-phthaldialdehyde (OPA) assay and the hydrolysis profiles were illustrated using SDS-PAGE. Papain generated RBCF hydrolysates exhibited higher ferric reducing antioxidant power (FRAP) and oxygen radical absorbance capacity (ORAC) compared to those generated with bromelain, FP400 and FPII. At certain concentrations, 24h hydrolysates of RBCF using FP400 and FPII were able to inhibit the growth of Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa. The results indicated that the use of proteases from plant or fungal sources can produce animal blood hydrolysates with antioxidant and antimicrobial activities. PMID:26920319

  2. Production of bioactive peptide hydrolysates from deer, sheep, pig and cattle red blood cell fractions using plant and fungal protease preparations.

    PubMed

    Bah, Clara S F; Carne, Alan; McConnell, Michelle A; Mros, Sonya; Bekhit, Alaa El-Din A

    2016-07-01

    Protease preparations from plant (papain and bromelain) and fungal (FP400 and FPII) sources were used to hydrolyze the red blood cell fractions (RBCFs) separated from deer, sheep, pig, and cattle abattoir-sourced blood. After 1, 2, 4 and 24h of hydrolysis, the antioxidant and antibacterial activities of the peptide hydrolysates obtained were investigated. The increase in trichloroacetic acid-soluble peptides over the hydrolysis period was examined using the o-phthaldialdehyde (OPA) assay and the hydrolysis profiles were illustrated using SDS-PAGE. Papain generated RBCF hydrolysates exhibited higher ferric reducing antioxidant power (FRAP) and oxygen radical absorbance capacity (ORAC) compared to those generated with bromelain, FP400 and FPII. At certain concentrations, 24h hydrolysates of RBCF using FP400 and FPII were able to inhibit the growth of Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa. The results indicated that the use of proteases from plant or fungal sources can produce animal blood hydrolysates with antioxidant and antimicrobial activities.

  3. Anti-inflammatory and antioxidant activities, functional properties and mutagenicity studies of protein and protein hydrolysate obtained from Prosopis alba seed flour.

    PubMed

    Cattaneo, Florencia; Sayago, Jorge Esteban; Alberto, María Rosa; Zampini, Iris Catiana; Ordoñez, Roxana Mabel; Chamorro, Verónica; Pazos, Adriana; Isla, María Inés

    2014-10-15

    Prosopis species are considered multipurpose trees and shrubs by FAO and their fruit constitute a food source for humans and animals. According to the "Código Alimentario Argentino", "algarrobo flour" is produced by grinding the whole mature pod, but in the traditional process most of the seeds are discarded. In this paper, the flour from seed was obtained. Then, the proteins were extracted and enzymatic hydrolysis was carried out. According to their amino acid profile and chemical score (>100%), the Prosopis alba proteins, are not deficient in essential amino acids considering the amount of amino acid necessary by adults. The protein isolate showed a good solubility (pH 7.4-9), emulsificant capacity, oil binding capacity and water adsorption capacity. The antioxidant ability of proteins was significantly increased with hydrolysis (SC50 values: 50-5μg/mL, respectively). Inhibitory activity of pro-inflammatory enzymes (lipoxygenase and phospholipase) was described. The mutagenicity/antimutagenicity of proteins and protein hydrolysates from seed flour were also analysed. The results suggest that P. alba cotyledon flour could be a new alternative in the formulation of functional foods not only for its high protein content but also by the biological and functional properties of its proteins and protein hydrolysates.

  4. Supplementation with a whey protein hydrolysate enhances recovery of muscle force-generating capacity following eccentric exercise.

    PubMed

    Buckley, Jonathan D; Thomson, Rebecca L; Coates, Alison M; Howe, Peter R C; DeNichilo, Mark O; Rowney, Michelle K

    2010-01-01

    There is evidence that protein hydrolysates can speed tissue repair following damage and may therefore be useful for accelerating recovery from exercise induced muscle damage. The potential for a hydrolysate (WPI(HD)) of whey protein isolate (WPI) to speed recovery following eccentric exercise was evaluated by assessing effects on recovery of peak isometric torque (PIT). In a double-blind randomised parallel trial, 28 sedentary males had muscle soreness (MS), serum creatine kinase (CK) activity, plasma TNFalpha, and PIT assessed at baseline and after 100 maximal eccentric contractions (ECC) of their knee extensors. Participants then consumed 250 ml of flavoured water (FW; n=11), or FW containing 25 g WPI (n=11) or 25 g WPI(HD) (n=6) and the assessments were repeated 1, 2, 6 and 24h later. PIT decreased approximately 23% following ECC, remained suppressed in FW and WPI, but recovered fully in WPI(HD) by 6h (P=0.006, treatment x time interaction). MS increased following ECC (P<0.001 for time), and remained elevated with no difference between groups (P=0.61). TNFalpha and CK did not change (P>0.45). WPI(HD) may be a useful supplement for assisting athletes to recover from fatiguing eccentric exercise.

  5. Transglutaminase cross-linking effect on sensory characteristics and antioxidant activities of Maillard reaction products from soybean protein hydrolysates.

    PubMed

    Song, Na; Tan, Chen; Huang, Meigui; Liu, Ping; Eric, Karangwa; Zhang, Xiaoming; Xia, Shuqin; Jia, Chengsheng

    2013-01-01

    To improve the yield of Maillard peptides, a microbial transglutaminase (MTGase) was used to increase the content of 1000-5000Da peptides in soybean protein hydrolysates by using a cross-linking reaction. The sensory characteristics and antioxidant activities of corresponding Maillard Reaction Products (MSPC) was then evaluated. After cross-linking treatment the content of 1000-5000Da peptides in protein hydrolysates and the yield of Maillard peptides increased by 21.19% and 8.71%, respectively, which contributed to the improved mouthfulness of MSPC. The bitter amino acids were significantly decreased and the umami acids were markedly increased in MSPC. Volatile compounds identified by GC-MS analysis showed that the content of the important meaty flavour compounds (such as 2-methyl-3-furanthiol, bis(2-methyl-3-furyl)disulfide) of MSPC were dramatically higher than that of MRPs from uncross-linking peptides. Combined with sensory evaluation, it was confirmed that MTGase cross-linking improved the flavour Characteristics and did not affect the antioxidant activity of MSPC.

  6. Anticancer Activity of a Hexapeptide from Skate (Raja porosa) Cartilage Protein Hydrolysate in HeLa Cells.

    PubMed

    Pan, Xin; Zhao, Yu-Qin; Hu, Fa-Yuan; Chi, Chang-Feng; Wang, Bin

    2016-01-01

    In this study, the hexapeptide Phe-Ile-Met-Gly-Pro-Tyr (FIMGPY), which has a molecular weight of 726.9 Da, was separated from skate (Raja porosa) cartilage protein hydrolysate using ultrafiltration and chromatographic methods, and its anticancer activity was evaluated in HeLa cells. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay indicated that FIMGPY exhibited high, dose-dependent anti-proliferation activities in HeLa cells with an IC50 of 4.81 mg/mL. Acridine orange/ethidium bromide (AO/EB) fluorescence staining and flow cytometry methods confirmed that FIMGPY could inhibit HeLa cell proliferation by inducing apoptosis. Western blot assay revealed that the Bax/Bcl-2 ratio and relative intensity of caspase-3 in HeLa cells treated with 7-mg/mL FIMGPY were 2.63 and 1.83, respectively, significantly higher than those of the blank control (p < 0.01). Thus, FIMGPY could induce apoptosis by upregulating the Bax/Bcl-2 ratio and caspase-3 activation. Using a DNA ladder method further confirmed that the anti-proliferation activity of FIMGPY was attributable to its role in inducing apoptosis. These results suggest that FIMGPY from skate cartilage protein hydrolysate may have applications as functional foods and nutraceuticals for the treatment and prevention of cancer. PMID:27537897

  7. Anticancer Activity of a Hexapeptide from Skate (Raja porosa) Cartilage Protein Hydrolysate in HeLa Cells

    PubMed Central

    Pan, Xin; Zhao, Yu-Qin; Hu, Fa-Yuan; Chi, Chang-Feng; Wang, Bin

    2016-01-01

    In this study, the hexapeptide Phe-Ile-Met-Gly-Pro-Tyr (FIMGPY), which has a molecular weight of 726.9 Da, was separated from skate (Raja porosa) cartilage protein hydrolysate using ultrafiltration and chromatographic methods, and its anticancer activity was evaluated in HeLa cells. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay indicated that FIMGPY exhibited high, dose-dependent anti-proliferation activities in HeLa cells with an IC50 of 4.81 mg/mL. Acridine orange/ethidium bromide (AO/EB) fluorescence staining and flow cytometry methods confirmed that FIMGPY could inhibit HeLa cell proliferation by inducing apoptosis. Western blot assay revealed that the Bax/Bcl-2 ratio and relative intensity of caspase-3 in HeLa cells treated with 7-mg/mL FIMGPY were 2.63 and 1.83, respectively, significantly higher than those of the blank control (p < 0.01). Thus, FIMGPY could induce apoptosis by upregulating the Bax/Bcl-2 ratio and caspase-3 activation. Using a DNA ladder method further confirmed that the anti-proliferation activity of FIMGPY was attributable to its role in inducing apoptosis. These results suggest that FIMGPY from skate cartilage protein hydrolysate may have applications as functional foods and nutraceuticals for the treatment and prevention of cancer. PMID:27537897

  8. Anticancer Activity of a Hexapeptide from Skate (Raja porosa) Cartilage Protein Hydrolysate in HeLa Cells.

    PubMed

    Pan, Xin; Zhao, Yu-Qin; Hu, Fa-Yuan; Chi, Chang-Feng; Wang, Bin

    2016-01-01

    In this study, the hexapeptide Phe-Ile-Met-Gly-Pro-Tyr (FIMGPY), which has a molecular weight of 726.9 Da, was separated from skate (Raja porosa) cartilage protein hydrolysate using ultrafiltration and chromatographic methods, and its anticancer activity was evaluated in HeLa cells. Methylthiazolyldiphenyl-tetrazolium bromide (MTT) assay indicated that FIMGPY exhibited high, dose-dependent anti-proliferation activities in HeLa cells with an IC50 of 4.81 mg/mL. Acridine orange/ethidium bromide (AO/EB) fluorescence staining and flow cytometry methods confirmed that FIMGPY could inhibit HeLa cell proliferation by inducing apoptosis. Western blot assay revealed that the Bax/Bcl-2 ratio and relative intensity of caspase-3 in HeLa cells treated with 7-mg/mL FIMGPY were 2.63 and 1.83, respectively, significantly higher than those of the blank control (p < 0.01). Thus, FIMGPY could induce apoptosis by upregulating the Bax/Bcl-2 ratio and caspase-3 activation. Using a DNA ladder method further confirmed that the anti-proliferation activity of FIMGPY was attributable to its role in inducing apoptosis. These results suggest that FIMGPY from skate cartilage protein hydrolysate may have applications as functional foods and nutraceuticals for the treatment and prevention of cancer.

  9. Purification and characterization of novel antioxidant peptides of different molecular weights from mackerel Pneumatophorus japonicus protein hydrolysate

    NASA Astrophysics Data System (ADS)

    Wang, Xueqin; Xing, Ronge; Liu, Song; Yu, Huahua; Li, Kecheng; Chen, Zuoyuan; Li, Pengcheng

    2015-01-01

    Mackerel ( Pneumatophorus japonic u s) proteins were hydrolyzed by five proteases: trypsin, papain, neutrase, acid protease, and flavourzyme. The hydrolysate treated by neutrase exhibited the highest antioxidant activity. Response surface methodology (RSM) was employed to optimize the hydrolysis conditions in an effort to obtain a mackerel protein hydrolysate (MPH) with the highest DPPH radical scavenging activity. The MPH was fractioned using a series of ultrafiltration membranes and five fractions, namely, MPH-I (>10 kDa), MPH-II (10-2.5 kDa), MPH-III (1-2.5 kDa), MPH-IV (0.4-1 kDa), and MPH-V (below 0.4 kDa), were obtained. DPPH radical scavenging activity, reducing power, hydroxyl radical scavenging activity, and the lipid peroxidation inhibition capability of these fractions were evaluated. The fractions in molecular weights <2.5 kDa (MPH-III, MPH-IV, and MPH-V), which occupied 93.4% of the total fractions, showed the strongest antioxidant activity; and the antioxidant activities of the three fractions are similar to each other. Using SP Sephadex C-25 and Sephadex G-25 columns, eight fractions were obtained from the MPH (<2.5 kDa). The isolated peptide I (1 664 kDa) displayed the highest DPPH radical scavenging activity. Therefore, MPH is a potential source of antioxidant peptides.

  10. The effects of inulin supplementation of diets with or without hydrolysed protein sources on digestibility, faecal characteristics, haematology and immunoglobulins in dogs.

    PubMed

    Verlinden, A; Hesta, M; Hermans, J M; Janssens, G P J

    2006-11-01

    Dogs with food allergy are often treated by giving a diet with hydrolysed protein sources. Prebiotics might also be successful in prevention and treatment of allergic disease through their effect on the colonic microflora, analogous to studies on probiotics in allergic children. The present study was set up to investigate the effect of supplementing inulin (IN) to commercial hypoallergenic dog diets on apparent nutrient digestibility, faecal characteristics, haematology and Ig in dogs. Supplementation of 3 % IN did not affect faecal pH, food and water intake and urine production. Compared with the intact protein diet with a limited number of ingredients (L), the diet with a hydrolysed protein source (H) resulted in an increased water intake (P<0.001), which could be due to the osmotic effect of free amino acids. Faeces production was increased by IN due to increased faecal moisture content. Increased faeces production on the H diet was mainly due to a higher DM excretion. Subsequently, the apparent digestibility coefficient (ADC) of DM was lower in the H diet group. A similar result was noted for ADC of diethyl ether extract and crude ash. The ADC of crude protein was higher in the H diet group, whereas IN decreased the ADC of crude protein. Differences in the ADC of crude protein among the different diets disappeared after correction for a higher faecal biomass, except for the dogs fed the L+IN diet. Total faecal IgA concentrations were lower in the H group (P<0.05) because of lower antigenic stimulation of hydrolysed protein, which implies that hydrolysed protein is really hypoallergenic. The present study indicates that the use of hydrolysed protein diets for canine food allergy treatment can affect digestibility and that combination with IN affected apparent protein digestibility but not IgA response. PMID:17092385

  11. The effects of inulin supplementation of diets with or without hydrolysed protein sources on digestibility, faecal characteristics, haematology and immunoglobulins in dogs.

    PubMed

    Verlinden, A; Hesta, M; Hermans, J M; Janssens, G P J

    2006-11-01

    Dogs with food allergy are often treated by giving a diet with hydrolysed protein sources. Prebiotics might also be successful in prevention and treatment of allergic disease through their effect on the colonic microflora, analogous to studies on probiotics in allergic children. The present study was set up to investigate the effect of supplementing inulin (IN) to commercial hypoallergenic dog diets on apparent nutrient digestibility, faecal characteristics, haematology and Ig in dogs. Supplementation of 3 % IN did not affect faecal pH, food and water intake and urine production. Compared with the intact protein diet with a limited number of ingredients (L), the diet with a hydrolysed protein source (H) resulted in an increased water intake (P<0.001), which could be due to the osmotic effect of free amino acids. Faeces production was increased by IN due to increased faecal moisture content. Increased faeces production on the H diet was mainly due to a higher DM excretion. Subsequently, the apparent digestibility coefficient (ADC) of DM was lower in the H diet group. A similar result was noted for ADC of diethyl ether extract and crude ash. The ADC of crude protein was higher in the H diet group, whereas IN decreased the ADC of crude protein. Differences in the ADC of crude protein among the different diets disappeared after correction for a higher faecal biomass, except for the dogs fed the L+IN diet. Total faecal IgA concentrations were lower in the H group (P<0.05) because of lower antigenic stimulation of hydrolysed protein, which implies that hydrolysed protein is really hypoallergenic. The present study indicates that the use of hydrolysed protein diets for canine food allergy treatment can affect digestibility and that combination with IN affected apparent protein digestibility but not IgA response.

  12. Glucosamine-induced glycation of hydrolysed meat proteins in the presence or absence of transglutaminase: Chemical modifications and taste-enhancing activity.

    PubMed

    Hong, Pui Khoon; Ndagijimana, Maurice; Betti, Mirko

    2016-04-15

    Salt reduction in food is a challenging task. The food processing sector has adopted taste enhancers to replace salt partially. In this study, a flavour enhancer formulation (liquid seasoning) was produced using enzymatically hydrolysed poultry proteins isolate (PPI). The PPI obtained through the isoelectric solubilisation precipitation process (ISP) was hydrolysed with Alcalase and glycated with glucosamine (GlcN) at moderate temperatures (37/50°C) in the presence or absence of transglutaminase (TGase). The glycated hydrolysates showed reduced fluorescence advanced glycated end-products (AGE) and a reduced amount of alpha-dicarbonyl compounds (α-DC). An untrained consumer panel ranked the meat protein hydrolysate seasoning saltier than the salty standard seasoning solution (p<0.05) regardless of GlcN glycation (both tested at 0.3M Na(+)). GlcN treatments showed a tendency (p=0.0593) to increase savouriness. Free glutamic acid and free aspartic acid found in the PPI hydrolysate likely increased the salty perception. PMID:26675851

  13. Affinity purification and characterisation of zinc chelating peptides from rapeseed protein hydrolysates: possible contribution of characteristic amino acid residues.

    PubMed

    Xie, Ningning; Huang, Jingjing; Li, Bo; Cheng, Jianghua; Wang, Zhuochen; Yin, Junfeng; Yan, Xiaoming

    2015-04-15

    Zinc is an essential trace element for human growth and development. In this work, zinc-chelating peptides from rapeseed protein hydrolysates produced with alcalase were investigated by affinity chromatography with immobilized zinc and Sephadex G-25 gel filtration. Four small peptides, namely, Ala-Arg, Asn-Ser-Met (NSM), Gly-Lys-Arg, and Glu-Pro-Ser-His, were obtained and identified by reversed-phase high-performance liquid chromatography and electrospray ionization mass spectrometry. The zinc-chelating ability of the four peptides was further validated by inductively coupled plasma atomic emission spectrometry (ICP-AES). NSM was found to exhibit the highest zinc-chelating rate, which was better than that of reduced glutathione. We speculated that the Asn residue at the amino-terminus might facilitate this zinc-chelating ability. Therefore, utilizing small peptides from rapeseed protein as novel carriers for zinc supplement was feasible.

  14. Fluorescence spectroscopy and principal component analysis of soy protein hydrolysate fractions and the potential to assess their antioxidant capacity characteristics.

    PubMed

    Ranamukhaarachchi, Sahan A; Peiris, Ramila H; Moresoli, Christine

    2017-02-15

    The potential of intrinsic fluorescence and principal component analysis (PCA) to characterize the antioxidant capacity of soy protein hydrolysates (SPH) during sequential ultrafiltration (UF) and nanofiltration (NF) was evaluated. SPH was obtained by enzymatic hydrolysis of soy protein isolate. Antioxidant capacity was measured by Oxygen Radical Absorbance Capacity (ORAC) and Folin Ciocalteau Reagent (FCR) assays together with fluorescence excitation-emission matrices (EEM). PCA of the fluorescence EEMs revealed two principal components (PC1-tryptophan, PC2-tyrosine) that captured significant variance in the fluorescence spectra. Regression models between antioxidant capacity and PC1 and PC2 displayed strong linear correlations for NF fractions and a weak linear correlation for UF fractions. Clustering of UF and NF fractions according to ORACFPCA and FCRFPCA was observed. The ability of this method to extract information on contributions by tryptophan and tyrosine amino acid residues to the antioxidant capacity of SPH fractions was demonstrated. PMID:27664660

  15. Food protein hydrolysates as a source of dipeptidyl peptidase IV inhibitory peptides for the management of type 2 diabetes.

    PubMed

    Power, Orla; Nongonierma, A B; Jakeman, P; FitzGerald, R J

    2014-02-01

    The prevalence of type 2 diabetes mellitus (T2DM) is increasing and it is estimated that by 2030 approximately 366 million people will be diagnosed with this condition. The use of dipeptidyl peptidase IV (DPP-IV) inhibitors is an emerging strategy for the treatment of T2DM. DPP-IV is a ubiquitous aminodipeptidase that cleaves incretins such as glucagon like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), resulting in a loss in their insulinotropic activity. Synthetic DPP-IV drug inhibitors are being used to increase the half-life of the active GLP-1 and GIP. Dietary intervention is accepted as a key component in the prevention and management of T2DM. Therefore, identification of natural food protein-derived DPP-IV inhibitors is desirable. Peptides with DPP-IV inhibitory activity have been identified in a variety of food proteins. This review aims to provide an overview of food protein hydrolysates as a source of the DPP-IV inhibitory peptides with particular focus on milk proteins. In addition, the proposed modes of inhibition and structure-activity relationship of peptide inhibitors are discussed. Milk proteins and associated peptides also display insulinotropic activity and help regulate blood glucose in healthy and diabetic subjects. Therefore, milk protein derived peptide inhibitors may be a unique multifunctional peptide approach for the management of T2DM.

  16. Efficient hydrolysis of corncob residue through cellulolytic enzymes from Trichoderma strain G26 and L-lactic acid preparation with the hydrolysate.

    PubMed

    Xie, Lulu; Zhao, Jin; Wu, Jian; Gao, Mingfu; Zhao, Zhewei; Lei, Xiangyun; Zhao, Yi; Yang, Wei; Gao, Xiaoxue; Ma, Cuiyun; Liu, Huanfei; Wu, Fengjuan; Wang, Xingxing; Zhang, Fengwei; Guo, Pengyuan; Dai, Guifu

    2015-10-01

    To prepare fermentable hydrolysate from corncob residue (CCR), Trichoderma strain G26 was cultured on medium containing CCR for production of cellulolytic enzymes through solid-state fermentation (SSF), resulting in 71.3 IU/g (FPA), 136.2 IU/g (CMCase), 85.1 IU/g (β-glucosidase) and 11,344 IU/g (xylanase), respectively. Through a three-stage saccharification strategy, CCR was hydrolyzed by the enzymatic solution (6.5 FPU/ml) into fermentable hydrolysate containing 60.1g/l glucose (81.2% cellulose was converted at solid loading of 12.5%), 21.4% higher than that by the one-stage method. And then the hydrolysate was used to produce L-lactic acid by a previous screened strain Bacillus coagulans ZX25 in the submerged fermentation. 52.0 g/l L-lactic acid was obtained after fermentation for 44 h, with 86.5% glucose being converted to L-lactic acid. The results indicate that the strains and the hydrolysis strategy are promising for commercial production of L-lactic acid from CCR and other biomass. PMID:26143000

  17. Single-Cell Protein Production by the Acid-Tolerant Fungus Scytalidium acidophilum from Acid Hydrolysates of Waste Paper †

    PubMed Central

    Ivarson, K. C.; Morita, H.

    1982-01-01

    The bioconversion of waste paper to single-cell protein at pH <1 by Scytalidium acidophilum is described. Waste paper pretreated with 72% H2SO4 at 4°C was diluted with water to a pH of <0.1 and hydrolyzed. This yielded an adequate sugar-containing substrate for the growth of the fungus. A total of 97% of the sugars (glucose, galactose, mannose, xylose, arabinose) in the hydrolysates were converted to cell biomass. Microbial contamination was not observed. Based on the sugars consumed, S. acidophilum produced higher yields in shake cultures than many other Fungi Imperfecti. In aerated cultures, productivity increased, and yields of 43 to 46% containing 44 to 47% crude protein were obtained. This compares favorably with Candida utilis, a yeast used commercially to produce single-cell protein. The chemical constituents and the essential amino acids of the fungal cells were similar to those of other fungi. The nucleic acid content was characteristic of microbes containing low levels of nucleic acid. The advantages of using S. acidophilum for single-cell protein production are discussed. PMID:16345970

  18. An extensively hydrolysed rice protein-based formula in the management of infants with cow's milk protein allergy: preliminary results after 1 month

    PubMed Central

    Vandenplas, Yvan; De Greef, Elisabeth; Hauser, Bruno

    2014-01-01

    Background Guidelines recommend extensively hydrolysed cow's milk protein formulas (eHF) in the treatment of infants diagnosed with cow's milk protein allergy (CMPA). Extensively hydrolysed rice protein infant formulas (eRHFs) have recently become available, and could offer a valid alternative. Methods A prospective trial was performed to evaluate the clinical tolerance of a new eRHF in infants with a confirmed CMPA. Patients were followed for 1 month. Clinical tolerance of the eRHF was evaluated with a symptom-based score (SBS) and growth (weight and length) was monitored. Results Thirty-nine infants (mean age 3.4 months, range 0.5–6 months) diagnosed with CMPA were enrolled. All infants tolerated the eRHF and experienced a normal growth. Conclusions In accordance with current guidelines, this eRHF is tolerated by more than 90% of children with proven CMPA with a 95% CI, and is an adequate alternative to cow's milk-based eHF. Trial registration number ClinicalTrials.gov NCT01998074. PMID:24914098

  19. Selective separation of peptides contained in a rapeseed (Brassica campestris L.) protein hydrolysate using UF/NF membranes.

    PubMed

    Tessier, B; Harscoat-Schiavo, C; Marc, I

    2006-05-17

    The ability of a charged UF membrane to fractionate the small peptides found in a rapeseed protein enzymatic hydrolysate, according to their charge characteristics, was investigated. The complexity of such a hydrolysate has required the setting up of technological alternatives to isolate the small peptides, to obtain a more efficient separation among the numerous peptide species. A preliminary step consisted of precipitation followed by filtration with a 3000 g/mol molecular weight cutoff (MWCO) membrane to obtain a solution concentrated in small peptides. The possibility of fractionating these small peptides by a charged 1000 g/mol MWCO membrane was investigated. The study enabled us to assess the contribution of electrostatic interactions during fractionation. The effect of pH and ionic strength on the peptide transmission was studied. The ionic strength contribution was considered by studying the effect on the selectivity of a desalting step by nanofiltration on a 500 g/mol MWCO membrane. Peptide transmission was lower at pH 9 than pH 4, and it was the lowest at pH 9 and low ionic strength. Ionic strength had a significant influence at pH 9 but showed no influence at pH 4. The amino acid analysis and capillary electrophoresis revealed that negatively charged (acid) peptides were found in lower proportions in the permeate. The opposite trend was observed for basic peptides, whereas neutral peptides were found in the same proportion in the retentate and the permeate. These results can be explained, according to the Donnan theory, by the existence of attractive and repulsive forces at the membrane-solution interface. Selectivity between basic and acid peptides was as high as 1.90 at pH 9 and low ionic strength. A rough sketch of a membrane-based process is proposed to fractionate rapeseed peptide mixtures. Results obtained were reproducible within 10%.

  20. Contribution to aroma characteristics of mutton process flavor from the enzymatic hydrolysate of sheep bone protein assessed by descriptive sensory analysis and gas chromatography olfactometry.

    PubMed

    Zhan, Ping; Tian, Honglei; Zhang, Xiaoming; Wang, Liping

    2013-03-15

    Changes in the aroma characteristics of mutton process flavors (MPFs) prepared from sheep bone protein hydrolysates (SBPHs) with different degrees of hydrolysis (DH) were evaluated using gas chromatography-mass spectrometry (GC-MS), gas chromatography-olfactometry (GC-O), and descriptive sensory analysis (DSA). Five attributes (muttony, meaty, roasted, mouthful, and simulate) were selected to assess MPFs. The results of DSA showed a distinct difference among the control sample MPF0 and other MPF samples with added SBPHs for different DHs of almost all sensory attributes. MPF5 (DH 25.92%) was the strongest in the muttony, meaty, and roasted attributes, whereas MPF6 (DH 30.89%) was the strongest in the simulate and roasted attributes. Thirty-six compounds were identified as odor-active compounds for the evaluation of the sensory characteristics of MPFs via GC-MS-O analysis. The results of correlation analysis among odor-active compounds, molecular weight, and DSA further confirmed that the SBPH with a DH range of 25.92-30.89% may be a desirable precursor for the sensory characteristics of MPF. PMID:23416288

  1. Effect of Shrimp Chitin and Shrimp Chitin Hydrolysate on the Freeze-Induced Denaturation, and on the Amount of Unfreezable Water of Wanieso Lizardfish Myofibrillar Protein

    NASA Astrophysics Data System (ADS)

    Somjit, Kingduean; Kongpun, Orawan; Osatomi, Kiyoshi; Hara, Kenji; Nozaki, Yukinori

    In view of potential utilization of shrimp waste, shrimp chitin (SC) and shrimp chitin hydrolysate (SCH) were prepared from 3 kinds of shrimp species, namely: black tiger shrimp Penaeus monodon, endeavour shrimp Metapenaeus endeavouri and giant freshwater shrimp Macrobrachium rosenbergii. The effects of 5% SC and SCH (dry weight) on the state of water and on the denaturation of wanieso lizardfish Saurida wanieso myofibrillar protein (Mf) were evaluated based on changes in Mf Ca-ATPase activity and the amount of unfreezable water during frozen storage. Each effect was compared with those of Mf without additives (control) and Mf with glucose. The changes in Ca-ATPase activity of control and Mf with SC during frozen storage were exhibited biphasic pattern while those of SCH and glucose exhibited monophasic pattern. The amount of unfreezable water of Mf with SC was lower than that of control while those of Mf with SCH and glucose were higher than that of control. Present findings suggested that the preventive effect of SCH on freeze-induced denaturation of Mf is caused by the stabilizing the hydrated water molecule surrounding the Mf.

  2. A novel angiotensin-І converting enzyme (ACE) inhibitory peptide from gastrointestinal protease hydrolysate of silkworm pupa (Bombyx mori) protein: Biochemical characterization and molecular docking study.

    PubMed

    Wu, Qiongying; Jia, Junqiang; Yan, Hui; Du, Jinjuan; Gui, Zhongzheng

    2015-06-01

    Silkworm pupa (Bombyx mori) protein was hydrolyzed using gastrointestinal endopeptidases (pepsin, trypsin and α-chymotrypsin). Then, the hydrolysate was purified sequentially by ultrafiltration, gel filtration chromatography and RP-HPLC. A novel ACE inhibitory peptide, Ala-Ser-Leu, with the IC50 value of 102.15μM, was identified by IT-MS/MS. This is the first report of Ala-Ser-Leu from natural protein. Lineweaver-Burk plots suggest that the peptide is a competitive inhibitor against ACE. The molecular docking studies revealed that the ACE inhibition of Ala-Ser-Leu is mainly attributed to forming very strong hydrogen bonds with the S1 pocket (Ala354) and the S2 pocket (Gln281 and His353). The results indicate that silkworm pupa (B. mori) protein or its gastrointestinal protease hydrolysate could be used as a functional ingredient in auxiliary therapeutic foods against hypertension.

  3. Cholesterol regulatory effects and antioxidant activities of protein hydrolysates from zebra blenny (Salaria basilisca) in cholesterol-fed rats.

    PubMed

    Ktari, Naourez; Belguith-Hadriche, Olfa; Ben Amara, Ibtissem; Ben Hadj, Aïda; Turki, Mouna; Makni-Ayedi, Fatma; Boudaouara, Tahia; El Feki, Abdelfattah; Boualga, Ahmed; Ben Salah, Riadh; Nasri, Moncef

    2015-07-01

    This study aims to explore the hypocholesterolemic effects and antioxidative activities of zebra blenny protein hydrolysates (ZBPHs) in rats fed with a hypercholesterolemic diet. The rats were fed during eight weeks a standard laboratory diet (normal rats), a high-cholesterol diet (HCD) (1%) or a HCD and orally treated with ZBPHs or undigested zebra blenny proteins (UZBPs) (400 mg per kg per day). Results showed that a hypercholesterolemic diet induced the increase of total cholesterol (TC), triglycerides (TG), and low-density lipoprotein cholesterol (LDL-C). Treatment with ZBPHs increased the level of high-density lipoprotein cholesterol (HDL-C) and decreased significantly the levels of TC, TG, and LDL-C. In addition, ZBPH treatment showed significant normalization of thiobarbituric acid-reactive substance (TBARS) levels as well as catalase, superoxide dismutase (SOD), and glutathione peroxidase (GPx) activities in renal and hepatic tissues. Furthermore, ZBPHs may also exert significant protective effects on liver and kidney functions, evidenced by a marked decrease in the level of serum urea, uric acid, creatinine, alkaline phosphatase (ALP), and alanine aminotransferase (ALAT). Histological studies confirmed that ZBPHs effectively protected the livers and kidneys against hypercholesterolemia-mediated oxidative damage. Therefore, the study strengthens the hypothesis that ZBPHs can be used as novel antioxidants and hypocholesterolemic compounds against hyperlipidemia induced atherosclerosis. PMID:26065510

  4. Optimization of Maillard reaction with ribose for enhancing anti-allergy effect of fish protein hydrolysates using response surface methodology.

    PubMed

    Yang, Sung-Yong; Kim, Se-Wook; Kim, Yoonsook; Lee, Sang-Hoon; Jeon, Hyeonjin; Lee, Kwang-Won

    2015-06-01

    Halibut is served on sushi and as sliced raw fish fillets. We investigated the optimal conditions of the Maillard reaction (MR) with ribose using response surface methodology to reduce the allergenicity of its protein. A 3-factored and 5-leveled central composite design was used, where the independent variables were substrate (ribose) concentration (X1, %), reaction time (X2, min), and pH (X3), while the dependent variables were browning index (Y1, absorbance at 420nm), DPPH scavenging (Y2, EC50 mg/mL), FRAP (Y3, mM FeSO4/mg extract) and β-hexosaminidase release (Y4, %). The optimal conditions were obtained as follows: X1, 28.36%; X2, 38.09min; X3, 8.26. Maillard reaction products of fish protein hydrolysate (MFPH) reduced the amount of nitric oxide synthesis compared to the untreated FPH, and had a significant anti-allergy effect on β-hexosaminidase and histamine release, compared with that of the FPH control. We concluded that MFPH, which had better antioxidant and anti-allergy activities than untreated FPH, can be used as an improved dietary source. PMID:25624251

  5. Reverse-phase HPLC separation of hemp seed (Cannabis sativa L.) protein hydrolysate produced peptide fractions with enhanced antioxidant capacity.

    PubMed

    Girgih, Abraham T; Udenigwe, Chibuike C; Aluko, Rotimi E

    2013-03-01

    Hemp seed protein hydrolysate (HPH) was produced through simulated gastrointestinal tract (GIT) digestion of hemp seed protein isolate followed by partial purification and separation into eight peptide fractions by reverse-phase (RP)-HPLC. The peptide fractions exhibited higher oxygen radical absorbance capacity as well as scavenging of 2,2-diphenyl-1-picrylhydrazyl, superoxide and hydroxyl radicals when compared to HPH. Radical scavenging activities of the fractionated peptides increased as content of hydrophobic amino acids or elution time was increased, with the exception of hydroxyl radical scavenging that showed decreased trend. Glutathione (GSH), HPH and the RP-HPLC peptide fractions possessed low ferric ion reducing ability but all had strong (>60 %) metal chelating activities. Inhibition of linoleic acid oxidation by some of the HPH peptide fractions was higher at 1 mg/ml when compared to that observed at 0.1 mg/ml peptide concentration. Peptide separation resulted in higher concentration of some hydrophobic amino acids (especially proline, leucine and isoleucine) in the fractions (mainly F5 and F8) when compared to HPH. The elution time-dependent increased concentrations of the hydrophobic amino acids coupled with decreased levels of positively charged amino acids may have been responsible for the significantly higher (p < 0.05) antioxidant properties observed for some of the peptide fractions when compared to the unfractionated HPH. In conclusion, the antioxidant activity of HPH after simulated GIT digestion is mainly influenced by the amino acid composition of some of its peptides.

  6. Antioxidative activities of hydrolysates from edible birds nest using enzymatic hydrolysis

    NASA Astrophysics Data System (ADS)

    Muhammad, Nurul Nadia; Babji, Abdul Salam; Ayub, Mohd Khan

    2015-09-01

    Edible bird's nest protein hydrolysates (EBN) were prepared via enzymatic hydrolysis to investigate its antioxidant activity. Two types of enzyme (alcalase and papain) were used in this study and EBN had been hydrolysed with different hydrolysis time (30, 60, 90 and 120 min). Antioxidant activities in EBN protein hydrolysate were measured using DPPH, ABTS+ and Reducing Power Assay. From this study, increased hydrolysis time from 30 min to 120 min contributed to higher DH, as shown by alcalase (40.59%) and papain (24.94%). For antioxidant assay, EBN hydrolysed with papain showed higher scavenging activity and reducing power ability compared to alcalase. The highest antioxidant activity for papain was at 120 min hydrolysis time with ABTS (54.245%), DPPH (49.78%) and Reducing Power (0.0680). Meanwhile for alcalase, the highest antioxidant activity was at 30 min hydrolysis time. Even though scavenging activity for EBN protein hydrolysates were high, the reducing power ability was quite low as compared to BHT and ascorbic Acid. This study showed that EBN protein hydrolysate with alcalase and papain treatments potentially exhibit high antioxidant activity which have not been reported before.

  7. The Maillard reaction of a shrimp by-product protein hydrolysate: chemical changes and inhibiting effects of reactive oxygen species in human HepG2 cells.

    PubMed

    Zha, Fengchao; Wei, Binbin; Chen, Shengjun; Dong, Shiyuan; Zeng, Mingyong; Liu, Zunying

    2015-06-01

    Recently, much attention has been given to improving the antioxidant activity of protein hydrolysates via the Maillard reaction, but little is known about the cellular antioxidant activity of Maillard reaction products (MRPs) from protein hydrolysates. We first investigated chemical characterization and the cellular antioxidant activity of MRPs in a shrimp (Litopenaeus vannamei) by-product protein hydrolysate (SBH)-glucose system at 110 °C for up to 10 h of heating. Solutions of SBH and glucose were also heated alone as controls. The Maillard reaction greatly resulted in the increase of hydroxymethylfurfural (HMF) and browning intensity, high molecular weight fraction, and reduction of the total amino acid in SBH with the heating time, which correlated well with the free radical scavenging activity of MRPs. MRPs had stronger inhibiting effects on oxidative stress of human HepG2 cells than the original SBH, and its cellular antioxidant activity strongly correlated with free radical scavenging activity, but less affected by the browning intensity and HMF level. The caramelization of glucose partially affected the HMF level and free radical scavenging activity of MRPs, but it was not related to the cellular antioxidant activity. The cellular antioxidant activity of MRPs for 5 h of heating time appeared to reach a maximum level, which was mainly due to carbonyl ammonia condensation reaction. In conclusion, the Maillard reaction is a potential method to increase the cellular antioxidant activity of a shrimp by-product protein hydrolysate, but the higher HMF levels and the lower amino acid content in MRPs should also be considered.

  8. Antioxidant activity of Cod (Gadus morhua) protein hydrolysates: in vitro assays and evaluation in 5% fish oil-in-water emulsion.

    PubMed

    Farvin, K H Sabeena; Lystbæk Andersen, Lisa; Hauch Nielsen, Henrik; Jacobsen, Charlotte; Jakobsen, Greta; Johansson, Inez; Jessen, Flemming

    2014-04-15

    Cod protein hydrolysates were fractionated according to the molecular mass into three fractions of >5 kDa, 3-5 kDa and <3 kDa using an ultrafiltration membrane system. The antioxidative activity of the crude hydrolysates and the fractions were investigated, both in in vitro assays (DPPH radical scavenging activity, reducing power, Fe²⁺ chelating activity and inhibition of lipid oxidation in liposome model system), and in 5% fish oil-in-water emulsions. The <3 kDa fractions had very good radical scavenging activity, Fe²⁺ chelating activity and reducing power while the fraction 3-5 kDa resulted in higher protection against oxidation in the liposome model system. When tested in 5% oil-in-water emulsions, all the fractions, including the crude protein hydrolysate, were able to protect fish oil against oxidation in an iron induced oxidation system. However, none of the peptide fractions were effective in preventing tocopherol loss and showed no tocopherol sparing property. Volatile oxidation products showed an interaction between the aldehydes and the protein/peptides added in the emulsions, and this needs further investigation. PMID:24295714

  9. Use of viscera extract from hybrid catfish (Clarias macrocephalus × Clarias gariepinus) for the production of protein hydrolysate from toothed ponyfish (Gazza minuta) muscle.

    PubMed

    Klomklao, Sappasith; Kishimura, Hideki; Benjakul, Soottawat

    2013-01-15

    Proteolytic activity of viscera extract from hybrid catfish (Clarias macrocephalus × Clarias gariepinus) was studied. The optimal pH and temperature were 9.0 and 50°C, respectively, when toothed ponyfish (Gazza minuta) muscle was used as a substrate. When viscera extract from hybrid catfish was used for the production of protein hydrolysate from toothed ponyfish muscle, extract concentration, reaction time, and fish muscle/buffer ratio affected the hydrolysis and nitrogen recovery (NR) (p<0.05). Optimum conditions for toothed ponyfish muscle hydrolysis were 3.5% hybrid catfish viscera extract, 15 min reaction time and fish muscle/buffer ratio of 1:3 (w/v). High correlation between the degree of hydrolysis (DH) and NR (R(2)=0.974) was observed. Freeze-dried hydrolysate had a high protein content (89.02%, dry weight basis) and it was brownish yellow in colour (L(∗)=63.67, a(∗)=6.33, b(∗)=22.41). The protein hydrolysate contained a high amount of essential amino acids (48.22%) and had arginine and lysine as the dominant amino acids. PMID:23122156

  10. Post-exercise whey protein hydrolysate supplementation induces a greater increase in muscle protein synthesis than its constituent amino acid content.

    PubMed

    Kanda, Atsushi; Nakayama, Kyosuke; Fukasawa, Tomoyuki; Koga, Jinichiro; Kanegae, Minoru; Kawanaka, Kentaro; Higuchi, Mitsuru

    2013-09-28

    It is well known that ingestion of a protein source is effective in stimulating muscle protein synthesis after exercise. In addition, there are numerous reports on the impact of leucine and leucine-rich whey protein on muscle protein synthesis and mammalian target of rapamycin (mTOR) signalling. However, there is only limited information on the effects of whey protein hydrolysates (WPH) on muscle protein synthesis and mTOR signalling. The aim of the present study was to compare the effects of WPH and amino acids on muscle protein synthesis and the initiation of translation in skeletal muscle during the post-exercise phase. Male Sprague–Dawley rats swam for 2 h to depress muscle protein synthesis. Immediately after exercise, the animals were administered either carbohydrate (CHO), CHO plus an amino acid mixture (AA) or CHO plus WPH. At 1 h after exercise, the supplements containing whey-based protein (AA and WPH) caused a significant increase in the fractional rate of protein synthesis (FSR) compared with CHO. WPH also caused a significant increase in FSR compared with AA. Post-exercise ingestion of WPH caused a significant increase in the phosphorylation of mTOR levels compared with AA or CHO. In addition, WPH caused greater phosphorylation of ribosomal protein S6 kinase and eukaryotic initiation factor 4E-binding protein 1 than AA and CHO. In contrast, there was no difference in plasma amino acid levels following supplementation with either AA or WPH. These results indicate that WPH may include active components that are superior to amino acids for stimulating muscle protein synthesis and initiating translation.

  11. Zn(II) chelating with peptides found in sesame protein hydrolysates: identification of the binding sites of complexes.

    PubMed

    Wang, Chan; Wang, Chan; Li, Bo; Li, Haixia

    2014-12-15

    Two metal-chelating peptides identified from sesame protein hydrolysates, Ser-Met (SM) and Asn-Cys-Ser (NCS) were chemically synthesized in order to study zinc-peptide complexes. The stability constants of two peptides and their zinc complexations were measured by pH-potentiometric techniques, and the molecular masses of the Zn-peptide complexes were determined by electrospray ionization mass spectrometry. The structures of the complexes were studied by infrared spectroscopy and quantum chemistry and possible conformations of the Zn-peptide complexes were determined by molecular modeling to obtain direct insights into the molecular mechanism of Zn(2+) chelation with peptides. The results confirmed that the zinc ion-to-ligand ratio is 1:1 for the two peptides and that water is involved in the formation of the complexes. Among the functional groups of SM and NCS, the carboxyl, hydroxyl, and sulfhydryl groups showed the strongest bonding abilities with Zn(2+) and the carbonyl group of the peptide bond and water (O) regularly participate in coordination by weaker interactions with Zn(2+).

  12. Effects of fish protein hydrolysate on growth performance and humoral immune response in large yellow croaker (Pseudosciaena crocea R.)* §

    PubMed Central

    Tang, Hong-gang; Wu, Tian-xing; Zhao, Zhan-yu; Pan, Xiao-dong

    2008-01-01

    We investigated the effects of fish protein hydrolysate (FPH) on growth performance and humoral immune response of the large yellow croaker (Pseudosciaena crocea R.). One thousand and two hundred large yellow croakers [initial average weight: (162.75±23.85) g] were divided into four groups and reared in floating sea cages (3 m×3 m×3 m). The animals were fed with 4 diets: basal diet only (control) or diets supplemented with 5%, 10% and 15% (w/w) FPH. The results show that dietary FPH levels significantly influenced the growth and immunity of the large yellow croaker. Compared with the control group, total weight gain (TWG) in all treatment groups, relative weight gain (RWG) and specific growth rate (SGR) in fish fed with diets supplemented with 10% and 15% FPH were significantly increased (P<0.05). Similar results were observed in immune parameters [lysozyme activity, serum complements, immunoglobulin M (IgM)]. Lysozyme activity, complement C4 and IgM were also significantly increased (P<0.05) in fish fed with diets supplemented with 10% and 15% FPH, while complement C3 level was significantly increased (P<0.05) in all treatment groups. In general, with the supplementation of FPH, particularly at dose of 10%, the growth performance and immunity of the large yellow croaker can be improved effectively. PMID:18763300

  13. Evaluation of the cross‐reactivity of antigens in Glupearl 19S and other hydrolysed wheat proteins in cosmetics

    PubMed Central

    Nakamura, Masashi; Yagami, Akiko; Hara, Kazuhiro; Sano‐Nagai, Akiyo; Kobayashi, Tsukane

    2016-01-01

    Summary Background In Japan, over 2000 users of a facial soap containing Glupearl 19S (GP19S), a hydrolysed wheat protein (HWP), developed immediate‐type systemic wheat allergy (HWP‐IWA), and ∼70% of them developed associated contact urticaria. Objectives We investigated whether HWP‐IWA patients cross‐react with other HWPs, and analysed HWP antigenic characteristics. Methods We used 10 types of HWP that are commercially available as cosmetic ingredients, and 16 subjects with HWP‐IWA. We performed an enzyme‐linked immunosorbent assay (ELISA) to evaluate the reactivity to each HWP, and western blotting to evaluate the characteristics of the antigens by using HWP‐IWA patients' serum IgE antibodies. We also performed prick tests with the HWPs. Results The patients reacted to four other HWPs in addition to GP19S, according to ELISA, and this was confirmed by strong reactions in the prick tests to the same four types of HWP. Smears of antigens with molecular weights ranging from the high range to the low range were seen on western blotting with the four HWPs that showed strong reactions in the ELISA and prick tests. Conclusions HWP‐IWA patients cross‐react with other HWPs. The antigens that they cross‐reacted to had a molecular weight distribution similar to that of GP19S present in the HWPs. PMID:27027256

  14. Modification of soy protein hydrolysates by Maillard reaction: Effects of carbohydrate chain length on structural and interfacial properties.

    PubMed

    Li, Weiwei; Zhao, Haibo; He, Zhiyong; Zeng, Maomao; Qin, Fang; Chen, Jie

    2016-02-01

    This study investigated the effects of carbohydrate chain length on the structural and interfacial properties of the Maillard reaction conjugates of soy protein hydrolysates (Mw>30 kDa). The covalent attachment of sugars to soy peptides was confirmed by amino acid analysis and examination of the Fourier-transform infrared spectra. The results suggested that the emulsion stability of the conjugates increased as the length of the carbohydrate chains increased. The surface activity measurement revealed that the soy peptide-dextran conjugates were closely packed and that each molecule occupied a small area of the interface. It was further confirmed that the soy peptide-dextran conjugates formed a thick adsorbed layer at the oil-water interface, as observed in the confocal laser scanning micrographs. The interfacial layer of soy peptides was rheologically complex with broad linear viscoelastic region and strong elastic modulus, and the soy peptide-dextran conjugates might form multilayer adsorption at the interface. This study suggested that the improved surface properties of the soy peptide-dextran conjugates were a result of the strong membrane formed by the closely packed molecular and multilayer adsorption at the interface, which provided steric hindrance to flocculation. PMID:26655794

  15. Investigation of enzymatic hydrolysis conditions on the properties of protein hydrolysate from fish muscle (Collichthys niveatus) and evaluation of its functional properties.

    PubMed

    Shen, Qing; Guo, Rui; Dai, Zhiyuan; Zhang, Yanping

    2012-05-23

    This study was carried out to investigate the enzymatic hydrolysis conditions on the properties of protein hydrolysate from fish muscle of the marine fish species Collichthys niveatus. About 160 fish samples were tested, and the analyzed fish species was found to be a lean fish with low fat (1.77 ± 0.01%) and high protein (16.76 ± 1.21%). Fish muscle of C. niveatus was carefully collected and hydrolyzed with four commercial enzymes: Alcalase, Neutrase, Protamex, and Flavourzyme under the conditions recommended by the manufacturers. Among the tested proteases, Neutrase catalyzed the hydrolysis process most effectively since the hydrolysate generated by Neutrase has the highest content of sweet and umami taste amino acids (SUA). The effect of hydrolysis conditions was further optimized using response surface methodology (RSM), and the optimum values for temperature, pH, and enzyme/substrate ratio (E/S ratio) were found to be 40.7 °C, 7.68, and 0.84%, respectively. Finally, the amino acid composition of the hydrolysate was analyzed by AccQ·Tag derivatization and HPLC-PDA determination. Major amino acids of the muscle of C. niveatus were threonine, glutamic acid, phenyalanine, tryptophan, and lysine, accounting for respectively 10.92%, 10.85%, 10.79%, 9.86%, and 9.76% of total amino acid content. The total content of essential amino acids was 970.7 ng·mL(-1), while that of nonessential amino acids was 709.1 ng·mL(-1). The results suggest that the fish muscle and its protein hydrolysate from C. niveatus provide a versatile supply of the benefits and can be incorporated as supplements in health-care foods.

  16. Development of an enzymatic fish hydrolysate and its use in instant soup bases.

    PubMed

    Gálvez, A; Morales de Léon, J; Bourges Rodríguez, H

    1985-12-01

    The successful conservation of fish products, at low costs, is a subject of special interest in the developing countries. Conscious of this fact, our group has been studying several fish conservation methods, such as autolysis with high salt concentrations, and has obtained a sauce of high nutritive value and long shelf life. Nevertheless, the reaction process takes from four to six months. In the study herein reported, the hydrolysis was accelerated and controlled by using the following enzymes: papain, HT proteolytic, and Brew (N) zyme. The hydrolysate was then mixed with cereals to prepare instant soups. As results indicated, the best hydrolysate was obtained with Brew (N) zyme at 50 degrees C and 8.30 hours. This hydrolysate contains 93.0 g/100 g crude protein with a protein efficiency ratio (PER) and a net protein utilization (NPU) of 60% that of casein's NPU as well as a content of 0.8% ether extract. The lowest-cost mixtures with the highest nutritive value were: hydrolysate-wheat-soymeal, and hydrolysate-rice-soymeal, with 38.3 and 29.7 protein per 100 g of mixture, respectively, and a NPU of 79.0 and 79.8% in relation to casein, respectively. The soups prepared had a satisfactory acceptance rating. There were no significant differences in flavor and aroma at a confidence level of 95%. The cost per gram of protein is about US$ 0.22 per kg. PMID:3842931

  17. Angiotensin-I converting enzyme inhibitory activity of hydrolysates from oat (Avena sativa) proteins by in silico and in vitro analyses.

    PubMed

    Cheung, Imelda W Y; Nakayama, Satoko; Hsu, Monica N K; Samaranayaka, Anusha G P; Li-Chan, Eunice C Y

    2009-10-14

    The potential for producing antihypertensive peptides from oat proteins through enzymatic hydrolysis was assessed in silico and confirmed in vitro. Thermolysin (EC 3.4.24.27) was predicted using BIOPEP database as the enzyme that would theoretically produce the most angiotensin I converting enzyme (ACE) inhibitory peptides from oat. Experimental evidence confirmed that strong ACE-inhibitory activity was produced under various hydrolysis conditions. Hydrolysates produced under high enzyme-to-substrate ratio (3%) short time (20 min) (HEST) and low enzyme-to-substrate ratio (0.1%) long time (120 min) (LELT) conditions had IC(50) values of 30 and 50 microg/mL, respectively. After simulated gastrointestinal digestion, the IC(50) of the HEST hydrolysate was 35 microg/mL whereas the IC(50) of the LELT hydrolysate was higher at 85 microg/mL. Ultrafiltration revealed that potent ACE-inhibitory peptides had molecular weights below 3 kDa. This study demonstrates the usefulness of in silico analysis to select enzymes for hydrolysis of proteins not previously examined as sources of bioactive peptides.

  18. Antihypertensive Properties of a Pea Protein Hydrolysate during Short- and Long-Term Oral Administration to Spontaneously Hypertensive Rats.

    PubMed

    Girgih, Abraham T; Nwachukwu, Ifeanyi D; Onuh, John O; Malomo, Sunday A; Aluko, Rotimi E

    2016-05-01

    This study investigated short-term (24 h) and long-term (5 wk) systolic blood pressure (SBP)-lowering effects in spontaneously hypertensive rats (SHR) of a 5 kDa membrane pea protein hydrolysate permeate (PPH-5) produced through thermoase hydrolysis of pea protein isolate (PPI). Amino acid analysis showed that the PPH-5 had lower contents of sulfur-containing amino acids than the PPI. Size-exclusion chromatography indicated mainly low molecular weight (<10 kDa) peptides in PPH-5 but not in the PPI. The PPH-5 had renin and angiotensin converting enzyme inhibition IC50 values of 0.57 and 0.10 mg/mL (P < 0.05), respectively, and consisted mainly of peptides with 2 to 6 amino acids. Mass spectrometry analysis revealed mainly hydrophilic tetrapeptide sequences. After a single oral administration (100 mg/kg body weight) to SHR, the unheated PPI showed weakest (P < 0.05) SBP-lowering effect with a -4 mm Hg maximum when compared to -25 mm Hg for heat-treated PPI and -36 mm Hg for PPH-5. Incorporation of the PPH-5 as 0.5% or 1% (w/w) casein substitute in the SHR diet produced maximum SBP reductions of -22 or -26 mm Hg (P < 0.05), respectively after 3 wk. In comparison, the unhydrolyzed PPI produced a maximum SBP reduction of -17 mm Hg also after 3 wk. Potency of the pea products decreased in the 4th and 5th wk, though SBP values of the treated rats were still lower than the untreated control. We conclude that the antihypertensive potency of PPH-5 may have been due to the presence of easily absorbed hydrophilic peptides. PMID:27037677

  19. Fermentative recovery of lipids and proteins from freshwater fish head waste with reference to antimicrobial and antioxidant properties of protein hydrolysate.

    PubMed

    Ruthu; Murthy, Pushpa S; Rai, Amit Kumar; Bhaskar, N

    2014-09-01

    Effectiveness of fermentation using lactic acid bacteria (LAB) for recovering lipids and proteins simultaneously from freshwater fish head (FWH) was evaluated. Three different proteolytic LAB (Pediococcus acidilactici NCIM5368, Enterococcus faecium NCIM5335 and Pediococcus acidilactici FD3) isolated from fish processing wastes were employed in the fermentation process. The fermentation conditions involved 10 % (w/w) glucose, 2 % (w/w) NaCl and 10 % (v/w) LAB cultures at 37 °C. The process resulted in 38.4 % of degree of hydrolysis (in case proteins) and a recovery of 63.6 % of the oil present in the material. The fermentation process did not affect the fatty acid profile of lipids. The hydrolyzed protein rich fermentation liquor exhibited DPPH radical scavenging activity (EC50 - 5.1 mg protein) as well as antagonistic properties towards several bacterial and fungal pathogens. The minimum inhibitory concentrations (MIC) of fermentated liquor (with E. faecium NCIM5335 as starter) were 10 and 12 mg/ml for Listeria monocytogenes and Salmonella itridicus, respectively. A higher MIC (60 and 96 mg/ml for Aspergillus ochraceus and Penicillium chrysogenum, respectively) was observed in case of fungal pathogens. Both the oil and protein hydrolysate rich liquor from fish head can be used as biofunctional ingredients in both human food as well as livestock feed formulations.

  20. Concentration-dependent displacement of cholesterol in micelles by hydrophobic rice bran protein hydrolysates

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The recent production of rice bran oil in Asia and the U.S. has resulted in large quantities of defatted rice bran as a low-value byproduct. Peptides from soy, milk, and other foods have been shown to have the potential hypocholesterolemic property and rice bran protein (RBP) may also contain bioact...

  1. Enzymatic generation of whey protein hydrolysates under pH-controlled and non pH-controlled conditions: Impact on physicochemical and bioactive properties.

    PubMed

    Le Maux, Solène; Nongonierma, Alice B; Barre, Chloé; FitzGerald, Richard J

    2016-05-15

    Enzymatic hydrolysis of whey protein (WP) was carried out under pH-controlled and non pH-controlled conditions using papain and a microbial-derived alternative (papain-like activity). The impact of such conditions on physicochemical and bioactive properties was assessed. WP hydrolysates (WPH) generated with the same enzyme displayed similar degree of hydrolysis. However, their reverse-phase liquid chromatograph mass spectrometry peptide profiles differed. A significantly higher oxygen radical absorbance capacity (ORAC) value was obtained for WP hydrolysed with papain at constant pH of 7.0 compared to the associated WPH generated without pH regulation. In contrast, there was no significant effect of pH regulation on dipeptidyl peptidase IV (DPP-IV) properties. WP hydrolysed with papain-like activity under pH regulation at 7.0 displayed higher ORAC activity and DPP-IV inhibitory properties compared to the associated WPH generated without pH regulation. This study has demonstrated that pH conditions during WPH generation may impact on peptide release and therefore on WPH bioactive properties.

  2. Effect of Mechanically Deboned Chicken Meat Hydrolysates on the Physicochemical Properties of Imitation Fish Paste

    PubMed Central

    Jin, Sang-Keun; Go, Gwang-woong; Jung, Eun-Young; Lim, Hyun-Jung; Yang, Han-Sul; Park, Jae-Hong

    2014-01-01

    This study investigated on the effects of adding mechanically deboned chicken meat (MDCM) hydrolysates on the quality properties of imitation fish paste (IFP) during storage. IFP was prepared from Alaska Pollack, spent laying hens surimi and protein hydrolysates which were enzymatically extracted from MDCM. The study was designed as a 3×4 factorial design with three MDCM hydrolysate content groups (0%, 0.4%, and 0.8%) and four storage times (0, 2, 4, and 6 weeks). Addition of MDCM hydrolysates increased crude fat content but lowered water content (p<0.05). The breaking force of IFP, an indicator of gel formation, increased in treated groups compared to control (p<0.05). Angiotensin I-converting enzyme (ACE) activity was inhibited and free radical scavenging activity increased with increasing MDCM hydrolysate content (p<0.05). In conclusion, the addition of MDCM to IFP improves gel characteristics. Additionally, protein hydrolysates from MDCM serve as a potential source of ACE inhibiting peptides. PMID:25049933

  3. Scientist prepare Lysozyme Protein Crystal

    NASA Technical Reports Server (NTRS)

    1996-01-01

    Dan Carter and Charles Sisk center a Lysozyme Protein crystal grown aboard the USML-2 shuttle mission. Protein isolated from hen egg-white and functions as a bacteriostatic enzyme by degrading bacterial cell walls. First enzyme ever characterized by protein crystallography. It is used as an excellent model system for better understanding parameters involved in microgravity crystal growth experiments. The goal is to compare kinetic data from microgravity experiments with data from laboratory experiments to study the equilibrium.

  4. Feather keratin hydrolysates obtained from microbial keratinases: effect on hair fiber

    PubMed Central

    2013-01-01

    Background Hair is composed mainly of keratin protein and a small amount of lipid. Protein hydrolysates, in particular those with low molecular weight distribution have been known to protect hair against chemical and environmental damage. Many types of protein hydrolysates from plants and animals have been used in hair and personal care such as keratin hydrolysates obtained from nails, horns and wool. Most of these hydrolysates are obtained by chemical hydrolysis and hydrothermal methods, but recently hydrolyzed hair keratin, feather keratin peptides, and feather meal peptides have been obtained by enzymatic hydrolysis using Bacillus spp in submerged fermentation. Results Keratin peptides were obtained by enzymatic hydrolysis of keratinases using Bacillus subtilis AMR. The microorganism was grown on a feather medium, pH 8.0 (1% feathers) and supplemented with 0.01% of yeast extract, for 5 days, at 28°C with agitation. The supernatant containing the hydrolysates was colleted by centrifugation and ultra filtered in an AMICON system using nano–membranes (Millipore – YC05). The Proteins and peptides were analyzed using HPTLC and MALDI-TOF-MS. Commercial preparations of keratin hydrolysates were used as a comparative standard. After five days the feather had been degraded (90-95%) by the peptidases and keratinases of the microorganism. MALDI-TOF mass spectrometry showed multiple peaks that correspond to peptides in the range of 800 to 1079 Daltons and the commercial hydrolysate was in the range of 900 to 1400 Da. HPTLC showed lower molecular mass peptides and amino acids in the enzymatic hydrolysate when compared with the commercial hydrolysate . A mild shampoo and a rinse off conditioner were formulated with the enzymatic hydrolysate and applied to hair fibers to evaluate the hydration, with and without heat, using a Corneometer® CM 825. The hydration was more efficient with heat, suggesting a more complete incorporation of hydrolysates into the fibers

  5. Buffalo Cheese Whey Proteins, Identification of a 24 kDa Protein and Characterization of Their Hydrolysates: In Vitro Gastrointestinal Digestion.

    PubMed

    Bassan, Juliana C; Goulart, Antonio J; Nasser, Ana L M; Bezerra, Thaís M S; Garrido, Saulo S; Rustiguel, Cynthia B; Guimarães, Luis H S; Monti, Rubens

    2015-01-01

    Milk whey proteins are well known for their high biological value and versatile functional properties, characteristics that allow its wide use in the food and pharmaceutical industries. In this work, a 24 kDa protein from buffalo cheese whey was analyzed by mass spectrometry and presented homology with Bos taurus beta-lactoglobulin. In addition, the proteins present in buffalo cheese whey were hydrolyzed with pepsin and with different combinations of trypsin, chymotrypsin and carboxypeptidase-A. When the TNBS method was used the obtained hydrolysates presented DH of 55 and 62% for H1 and H2, respectively. Otherwise for the OPA method the DH was 27 and 43% for H1 and H2, respectively. The total antioxidant activities of the H1 and H2 samples with and without previous enzymatic hydrolysis, determined by DPPH using diphenyl-p-picrylhydrazyl radical, was 4.9 and 12 mM of Trolox equivalents (TE) for H2 and H2Dint, respectively. The increased concentrations for H1 and H2 samples were approximately 99% and 75%, respectively. The in vitro gastrointestinal digestion efficiency for the samples that were first hydrolyzed was higher compared with samples not submitted to previous hydrolysis. After in vitro gastrointestinal digestion, several amino acids were released in higher concentrations, and most of which were essential amino acids. These results suggest that buffalo cheese whey is a better source of bioavailable amino acids than bovine cheese whey.

  6. Buffalo Cheese Whey Proteins, Identification of a 24 kDa Protein and Characterization of Their Hydrolysates: In Vitro Gastrointestinal Digestion

    PubMed Central

    Bassan, Juliana C.; Goulart, Antonio J.; Nasser, Ana L. M.; Bezerra, Thaís M. S.; Garrido, Saulo S.; Rustiguel, Cynthia B.; Guimarães, Luis H. S.; Monti, Rubens

    2015-01-01

    Milk whey proteins are well known for their high biological value and versatile functional properties, characteristics that allow its wide use in the food and pharmaceutical industries. In this work, a 24 kDa protein from buffalo cheese whey was analyzed by mass spectrometry and presented homology with Bos taurus beta-lactoglobulin. In addition, the proteins present in buffalo cheese whey were hydrolyzed with pepsin and with different combinations of trypsin, chymotrypsin and carboxypeptidase-A. When the TNBS method was used the obtained hydrolysates presented DH of 55 and 62% for H1 and H2, respectively. Otherwise for the OPA method the DH was 27 and 43% for H1 and H2, respectively. The total antioxidant activities of the H1 and H2 samples with and without previous enzymatic hydrolysis, determined by DPPH using diphenyl-p-picrylhydrazyl radical, was 4.9 and 12 mM of Trolox equivalents (TE) for H2 and H2Dint, respectively. The increased concentrations for H1 and H2 samples were approximately 99% and 75%, respectively. The in vitro gastrointestinal digestion efficiency for the samples that were first hydrolyzed was higher compared with samples not submitted to previous hydrolysis. After in vitro gastrointestinal digestion, several amino acids were released in higher concentrations, and most of which were essential amino acids. These results suggest that buffalo cheese whey is a better source of bioavailable amino acids than bovine cheese whey. PMID:26465145

  7. Buffalo Cheese Whey Proteins, Identification of a 24 kDa Protein and Characterization of Their Hydrolysates: In Vitro Gastrointestinal Digestion.

    PubMed

    Bassan, Juliana C; Goulart, Antonio J; Nasser, Ana L M; Bezerra, Thaís M S; Garrido, Saulo S; Rustiguel, Cynthia B; Guimarães, Luis H S; Monti, Rubens

    2015-01-01

    Milk whey proteins are well known for their high biological value and versatile functional properties, characteristics that allow its wide use in the food and pharmaceutical industries. In this work, a 24 kDa protein from buffalo cheese whey was analyzed by mass spectrometry and presented homology with Bos taurus beta-lactoglobulin. In addition, the proteins present in buffalo cheese whey were hydrolyzed with pepsin and with different combinations of trypsin, chymotrypsin and carboxypeptidase-A. When the TNBS method was used the obtained hydrolysates presented DH of 55 and 62% for H1 and H2, respectively. Otherwise for the OPA method the DH was 27 and 43% for H1 and H2, respectively. The total antioxidant activities of the H1 and H2 samples with and without previous enzymatic hydrolysis, determined by DPPH using diphenyl-p-picrylhydrazyl radical, was 4.9 and 12 mM of Trolox equivalents (TE) for H2 and H2Dint, respectively. The increased concentrations for H1 and H2 samples were approximately 99% and 75%, respectively. The in vitro gastrointestinal digestion efficiency for the samples that were first hydrolyzed was higher compared with samples not submitted to previous hydrolysis. After in vitro gastrointestinal digestion, several amino acids were released in higher concentrations, and most of which were essential amino acids. These results suggest that buffalo cheese whey is a better source of bioavailable amino acids than bovine cheese whey. PMID:26465145

  8. Preparation of Yeast Hydrolysate Enriched in Cyclo-His-Pro (CHP) by Enzymatic Hydrolysis and Evaluation of Its Functionality.

    PubMed

    Lee, Hyun Jung; Son, Heung Soo; Park, Chung; Suh, Hyung Joo

    2015-12-01

    In this study, we attempted to enrich cyclo-His-Pro (CHP) using enzymatic hydrolysis of yeast and to evaluate the functionality of yeast hydrolysate (YH)-enriched CHP. Flavourzyme offered a better performance in enhancing CHP content than other proteases. The CHP enrichment conditions were optimized as follows: addition of 1% Flavourzyme, 48-h incubation at 60°C, and pH 6.0. The CHP content significantly increased by 20-fold after ultra-filtration (UF). Maximal CHP translation was obtained after heating for 8 h at 50°C and pH 7.0. YH showed poor foaming capacity between pH 3.0 to 9.0. The emulsifying activities of YHs were slightly higher at near acidic pH. Increase in heating temperature and time resulted in decreased CHP content. The results indicate that YH is more heat stable after UF. Therefore, the CHP in YH after UF can be used as a food additive with physiological CHP activity and high heat stability.

  9. Preparation of Yeast Hydrolysate Enriched in Cyclo-His-Pro (CHP) by Enzymatic Hydrolysis and Evaluation of Its Functionality

    PubMed Central

    Lee, Hyun Jung; Son, Heung Soo; Park, Chung; Suh, Hyung Joo

    2015-01-01

    In this study, we attempted to enrich cyclo-His-Pro (CHP) using enzymatic hydrolysis of yeast and to evaluate the functionality of yeast hydrolysate (YH)-enriched CHP. Flavourzyme offered a better performance in enhancing CHP content than other proteases. The CHP enrichment conditions were optimized as follows: addition of 1% Flavourzyme, 48-h incubation at 60°C, and pH 6.0. The CHP content significantly increased by 20-fold after ultra-filtration (UF). Maximal CHP translation was obtained after heating for 8 h at 50°C and pH 7.0. YH showed poor foaming capacity between pH 3.0 to 9.0. The emulsifying activities of YHs were slightly higher at near acidic pH. Increase in heating temperature and time resulted in decreased CHP content. The results indicate that YH is more heat stable after UF. Therefore, the CHP in YH after UF can be used as a food additive with physiological CHP activity and high heat stability. PMID:26770916

  10. In silico and in vitro analyses of the angiotensin-I converting enzyme inhibitory activity of hydrolysates generated from crude barley (Hordeum vulgare) protein concentrates.

    PubMed

    Gangopadhyay, Nirupama; Wynne, Kieran; O'Connor, Paula; Gallagher, Eimear; Brunton, Nigel P; Rai, Dilip K; Hayes, Maria

    2016-07-15

    Angiotensin-I-converting enzyme (ACE-I) plays a key role in control of hypertension, and type-2 diabetes mellitus, which frequently co-exist. Our current work utilised in silico methodologies and peptide databases as tools for predicting release of ACE-I inhibitory peptides from barley proteins. Papain was the enzyme of choice, based on in silico analysis, for experimental hydrolysis of barley protein concentrate, which was performed at the enzyme's optimum conditions (60 °C, pH 6.0) for 24 h. The generated hydrolysate was subjected to molecular weight cut-off (MWCO) filtration, following which the non-ultrafiltered hydrolysate (NUFH), and the generated 3 kDa and 10 kDa MWCO filtrates were assessed for their in vitro ACE-I inhibitory activities. The 3 kDa filtrate (1 mg/ml), that demonstrated highest ACE-I inhibitory activity of 70.37%, was characterised in terms of its peptidic composition using mass spectrometry and 1882 peptides derived from 61 barley proteins were identified, amongst which 15 peptides were selected for chemical synthesis based on their predicted ACE-I inhibitory properties. Of the synthesized peptides, FQLPKF and GFPTLKIF were most potent, demonstrating ACE-I IC50 values of 28.2 μM and 41.2 μM respectively. PMID:26948626

  11. Assessment of the angiotensin-I-converting enzyme (ACE-I) inhibitory and antioxidant activities of hydrolysates of bovine brisket sarcoplasmic proteins produced by papain and characterisation of associated bioactive peptidic fractions.

    PubMed

    Di Bernardini, Roberta; Mullen, Anne Maria; Bolton, Declan; Kerry, Joseph; O'Neill, Eileen; Hayes, Maria

    2012-01-01

    The main objective was to investigate the angiotensin-I-converting enzyme (ACE-I) inhibitory and antioxidant activities of sarcoplasmic proteins isolated from the brisket muscle (Pectoralis profundus) of 3 (Bos taurus) cattle and hydrolysed with papain for 24 h at 37°C. Sarcoplasmic protein hydrolysates were ultra-filtered using molecular weight cut off (MWCO) membranes and 10-kDa and 3-kDa filtrates were obtained. The total sarcoplasmic protein extracts and the 3-kDa filtrates were tested for angiotensin I-converting enzyme inhibitory (ACE-I) activities. The total hydrolysates, 10-kDa and 3-kDa filtrates were also tested for their associated antioxidant activities using the 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity assay, the ferric ion reducing antioxidant power (FRAP) assay and the Fe(2+) metal chelating ability assay. The peptidic content of the total hydrolysates, the 10-kDa and the 3-kDa filtrates were analysed using an ORBITRAP mass spectrometer, and mass spectral data obtained were analysed using TurboSEQUEST. Eleven peptides were characterised from the total hydrolysates, fifteen from the 10-kDa filtrate fractions, whilst nine peptides were characterised from the 3-kDa filtrate fractions. Similarities between the amino acid sequences of the peptides identified in this study and previously identified antioxidant and ACE-I inhibitory peptides detailed in the BIOPEP database were outlined. PMID:21880436

  12. Angiotensin I-converting enzyme (ACE) inhibitory activity and structural properties of oven- and freeze-dried protein hydrolysate from fresh water fish (Cirrhinus mrigala).

    PubMed

    Elavarasan, K; Shamasundar, B A; Badii, Faraha; Howell, Nazlin

    2016-09-01

    The angiotensin I-converting enzyme (ACE) inhibitory activity and structural properties of oven-dried (OD-FPH) and freeze-dried (FD-FPH) protein hydrolysates derived from fresh water fish (Cirrhinus mrigala) muscle, using papain, were investigated. Amino acid profiles indicated a higher proportion of hydrophobic residues in OD-FPH and hydrophilic residues in FD-FPH samples. Fourier transform infrared (FT-IR) spectra revealed random coil structure in OD-FPH and β-sheet in FD-FPH samples. The approximate molecular weight of peptides in OD-FPH and FD-FPH was in the range of 7030-339Da. The IC50 values for ACE inhibition by OD-FPH and FD-FPH samples were found to be 1.15 and 1.53mg of proteinml(-1), respectively. The ACE-inhibitory activity of OD-FPH was more stable (during sequential digestion, using pepsin and pancreatin) than that of FD-FPH sample. The study suggested that the ACE inhibitory activity of protein hydrolysate was not affected by oven-drying. PMID:27041318

  13. Angiotensin I-converting enzyme (ACE) inhibitory activity and structural properties of oven- and freeze-dried protein hydrolysate from fresh water fish (Cirrhinus mrigala).

    PubMed

    Elavarasan, K; Shamasundar, B A; Badii, Faraha; Howell, Nazlin

    2016-09-01

    The angiotensin I-converting enzyme (ACE) inhibitory activity and structural properties of oven-dried (OD-FPH) and freeze-dried (FD-FPH) protein hydrolysates derived from fresh water fish (Cirrhinus mrigala) muscle, using papain, were investigated. Amino acid profiles indicated a higher proportion of hydrophobic residues in OD-FPH and hydrophilic residues in FD-FPH samples. Fourier transform infrared (FT-IR) spectra revealed random coil structure in OD-FPH and β-sheet in FD-FPH samples. The approximate molecular weight of peptides in OD-FPH and FD-FPH was in the range of 7030-339Da. The IC50 values for ACE inhibition by OD-FPH and FD-FPH samples were found to be 1.15 and 1.53mg of proteinml(-1), respectively. The ACE-inhibitory activity of OD-FPH was more stable (during sequential digestion, using pepsin and pancreatin) than that of FD-FPH sample. The study suggested that the ACE inhibitory activity of protein hydrolysate was not affected by oven-drying.

  14. Housefly larvae hydrolysate: orthogonal optimization of hydrolysis, antioxidant activity, amino acid composition and functional properties

    PubMed Central

    2013-01-01

    Background Antioxidant, one of the most important food additives, is widely used in food industry. At present, antioxidant is mostly produced by chemical synthesis, which would accumulate to be pathogenic. Therefore, a great interest has been developed to identify and use natural antioxidants. It was showed that there are a lot of antioxidative peptides in protein hydrolysates, possessing strong capacity of inhibiting peroxidation of macro-biomolecular and scavenging free redicals in vivo. Enzymatic hydrolysis used for preparation of antioxidative peptides is a new hot-spot in the field of natural antioxidants. It reacts under mild conditions, with accurate site-specific degradation, good repeatability and few damages to biological activity of protein. Substrates for enzymatic hydrolysis are usually plants and aqua-animals. Insects are also gaining attention because of their rich protein and resource. Antioxidative peptides are potential to be exploited as new natural antioxidant and functional food. There is a huge potential market in medical and cosmetic field as well. Result Protein hydrolysate with antioxidant activity was prepared from housefly larvae, by a two-step hydrolysis. Through orthogonal optimization of the hydrolysis conditions, the degree of hydrolysis was determined to be approximately 60%. Fractionated hydrolysate at 25 mg/mL, 2.5 mg/mL and 1 mg/mL exhibited approximately 50%, 60% and 50% of scavenging capacity on superoxide radicals, 1, 1-Diphenyl-2-picrylhydrazyl radicals and hydroxyl radicals, respectively. Hydrolysate did not exhibit substantial ion chelation. Using a linoneic peroxidation system, the inhibition activity of hydrolysate at 20 mg/mL was close to that of 20 μg/mL tertiary butylhydroquinone, suggesting a potential application of hydrolysate in the oil industry as an efficient antioxidant. The lyophilized hydrolysate presented almost 100% solubility at pH 3-pH 9, and maintained nearly 100% activity at pH 5-pH 8 at 0

  15. Pepsin-pancreatin protein hydrolysates from extruded amaranth inhibit markers of atherosclerosis in LPS-induced THP-1 macrophages-like human cells by reducing expression of proteins in LOX-1 signaling pathway

    PubMed Central

    2014-01-01

    Background Atherosclerosis is considered a progressive disease that affects arteries that bring blood to the heart, to the brain and to the lower end. It derives from endothelial dysfunction and inflammation, which play an important role in the thrombotic complications of atherosclerosis. Cardiovascular disease is the leading cause of death around the world and one factor that can contribute to its progression and prevention is diet. Our previous study found that amaranth hydrolysates inhibited LPS-induced inflammation in human and mouse macrophages by preventing activation of NF-κB signaling. Furthermore, extrusion improved the anti-inflammatory effect of amaranth protein hydrolysates in both cell lines, probably attributed to the production of bioactive peptides during processing. Therefore, the objective of this study was to compare the anti-atherosclerotic potential of pepsin-pancreatin hydrolysates from unprocessed and extruded amaranth in THP-1 lipopolysaccharide-induced human macrophages and suggest the mechanism of action. Results Unprocessed amaranth hydrolysate (UAH) and extruded amaranth hydrolysate (EAH) showed a significant reduction in the expression of interleukin-4 (IL-4) (69% and 100%, respectively), interleukin-6 (IL-6) (64% and 52%, respectively), interleukin-22 (IL-22) (55% and 70%, respectively). Likewise, UAH and EAH showed a reduction in the expression of monocyte-chemo attractant protein-1 (MCP-1) (35% and 42%, respectively), transferrin receptor-1 (TfR-1) (48% and 61%, respectively), granulocyte-macrophage colony-stimulating factor (GM-CSF) (59% and 63%, respectively), and tumor necrosis factor-α (TNF-α) (60% and 63%, respectively). Also, EAH reduced the expression of lectin-like oxidized low-density lipoprotein receptor-1 (LOX-1) (27%), intracellular adhesion molecule-1 (ICAM-1) (28%) and matrix metalloproteinase-9 (MMP-9) (19%), important molecular markers in the atherosclerosis pathway. EAH, led to a reduction of 58, 52 and 79% for

  16. Preparation of Soluble Proteins from Escherichia coli

    PubMed Central

    Wingfield, Paul T.

    2014-01-01

    Purification of human IL-1β is used in this unit as an example of the preparation of soluble proteins from E. coli. Bacteria containing IL-1β are lysed, and IL-1 β in the resulting supernatant is purified by anion-exchange chromatography, salt precipitation and cation-exchange chromatography, and then concentrated. Finally, the IL-1 β protein is applied to a gel-filtration column to separate it from remaining higher- and lower-molecular-weight contaminants, the purified protein is stored frozen or is lyophilized. The purification protocol described is typical for a protein that is expressed in fairly high abundance (i.e., >5% total protein) and accumulates in a soluble state. Also, the purification procedure serves as an example of how use classical protein purifications methods which may also be used in conjunction with the affinity-based methods now more commonly used. PMID:25367009

  17. Preparation of Soluble Proteins from Escherichia coli.

    PubMed

    Wingfield, Paul T

    2014-01-01

    Purification of human IL-1β is used in this unit as an example of the preparation of a soluble protein from E. coli. Bacteria containing IL-1β are lysed, and IL-1 β in the resulting supernatant is purified by anion-exchange chromatography, salt precipitation, and cation-exchange chromatography, and then concentrated. Finally, the IL-1 β protein is applied to a gel-filtration column to separate it from remaining higher- and lower-molecular-weight contaminants, the purified protein is stored frozen or is lyophilized. The purification protocol described is typical for a protein that is expressed in fairly high abundance (i.e., >5% total protein) and accumulates in a soluble state. In addition, the purification procedure serves as an example of how to use classical protein purifications methods, which may also be used in conjunction with the affinity-based methods now more commonly used. PMID:25367009

  18. Actinopyga lecanora Hydrolysates as Natural Antibacterial Agents

    PubMed Central

    Ghanbari, Raheleh; Ebrahimpour, Afshin; Abdul-Hamid, Azizah; Ismail, Amin; Saari, Nazamid

    2012-01-01

    Actinopyga lecanora, a type of sea cucumber commonly known as stone fish with relatively high protein content, was explored as raw material for bioactive peptides production. Six proteolytic enzymes, namely alcalase, papain, pepsin, trypsin, bromelain and flavourzyme were used to hydrolyze A. lecanora at different times and their respective degrees of hydrolysis (DH) were calculated. Subsequently, antibacterial activity of the A. lecanora hydrolysates, against some common pathogenic Gram positive bacteria (Bacillus subtilis and Staphylococcus aureus) and Gram negative bacteria (Escherichia coli, Pseudomonas aeruginosa, and Pseudomonas sp.) were evaluated. Papain hydrolysis showed the highest DH value (89.44%), followed by alcalase hydrolysis (83.35%). Bromelain hydrolysate after one and seven hours of hydrolysis exhibited the highest antibacterial activities against Pseudomonas sp., P. aeruginosa and E. coli at 51.85%, 30.07% and 30.45%, respectively compared to the other hydrolysates. Protein hydrolysate generated by papain after 8 h hydrolysis showed maximum antibacterial activity against S. aureus at 20.19%. The potent hydrolysates were further fractionated using RP-HPLC and antibacterial activity of the collected fractions from each hydrolysate were evaluated, wherein among them only three fractions from the bromelain hydrolysates exhibited inhibitory activities against Pseudomonas sp., P. aeruginosa and E. coli at 24%, 25.5% and 27.1%, respectively and one fraction of papain hydrolysate showed antibacterial activity of 33.1% against S. aureus. The evaluation of the relationship between DH and antibacterial activities of papain and bromelain hydrolysates revealed a meaningful correlation of four and six order functions. PMID:23222684

  19. Angiotensin-I Converting Enzyme (ACE) Inhibitory and Anti-Oxidant Activities of Sea Cucumber (Actinopyga lecanora) Hydrolysates.

    PubMed

    Ghanbari, Raheleh; Zarei, Mohammad; Ebrahimpour, Afshin; Abdul-Hamid, Azizah; Ismail, Amin; Saari, Nazamid

    2015-01-01

    In recent years, food protein-derived hydrolysates have received considerable attention because of their numerous health benefits. Amongst the hydrolysates, those with anti-hypertensive and anti-oxidative activities are receiving special attention as both activities can play significant roles in preventing cardiovascular diseases. The present study investigated the angiotensin-I converting enzyme (ACE) inhibitory and anti-oxidative activities of Actinopyga lecanora (A. lecanora) hydrolysates, which had been prepared by alcalase, papain, bromelain, flavourzyme, pepsin, and trypsin under their optimum conditions. The alcalase hydrolysate showed the highest ACE inhibitory activity (69.8%) after 8 h of hydrolysis while the highest anti-oxidative activities measured by 2,2-diphenyl 1-1-picrylhydrazyl radical scavenging (DPPH) (56.00%) and ferrous ion-chelating (FIC) (59.00%) methods were exhibited after 24 h and 8 h of hydrolysis, respectively. The ACE-inhibitory and anti-oxidative activities displayed dose-dependent trends, and increased with increasing protein hydrolysate concentrations. Moreover, strong positive correlations between angiotensin-I converting enzyme (ACE) inhibitory and anti-oxidative activities were also observed. This study indicates that A. lecanora hydrolysate can be exploited as a source of functional food owing to its anti-oxidant as well as anti-hypertension functions. PMID:26690117

  20. Angiotensin-I Converting Enzyme (ACE) Inhibitory and Anti-Oxidant Activities of Sea Cucumber (Actinopyga lecanora) Hydrolysates

    PubMed Central

    Ghanbari, Raheleh; Zarei, Mohammad; Ebrahimpour, Afshin; Abdul-Hamid, Azizah; Ismail, Amin; Saari, Nazamid

    2015-01-01

    In recent years, food protein-derived hydrolysates have received considerable attention because of their numerous health benefits. Amongst the hydrolysates, those with anti-hypertensive and anti-oxidative activities are receiving special attention as both activities can play significant roles in preventing cardiovascular diseases. The present study investigated the angiotensin-I converting enzyme (ACE) inhibitory and anti-oxidative activities of Actinopyga lecanora (A. lecanora) hydrolysates, which had been prepared by alcalase, papain, bromelain, flavourzyme, pepsin, and trypsin under their optimum conditions. The alcalase hydrolysate showed the highest ACE inhibitory activity (69.8%) after 8 h of hydrolysis while the highest anti-oxidative activities measured by 2,2-diphenyl 1-1-picrylhydrazyl radical scavenging (DPPH) (56.00%) and ferrous ion-chelating (FIC) (59.00%) methods were exhibited after 24 h and 8 h of hydrolysis, respectively. The ACE-inhibitory and anti-oxidative activities displayed dose-dependent trends, and increased with increasing protein hydrolysate concentrations. Moreover, strong positive correlations between angiotensin-I converting enzyme (ACE) inhibitory and anti-oxidative activities were also observed. This study indicates that A. lecanora hydrolysate can be exploited as a source of functional food owing to its anti-oxidant as well as anti-hypertension functions. PMID:26690117

  1. Atlantic salmon (Salmo salar) protein hydrolysate in diets for weaning piglets ─ effect on growth performance, intestinal morphometry and microbiota composition.

    PubMed

    Opheim, Margareth; Strube, Mikael Lenz; Sterten, Hallgeir; Øverland, Margareth; Kjos, Nils Petter

    2016-01-01

    Salmon protein hydrolysates (SPH) from two different rest raw materials were evaluated in diets for weaning piglets. Four experimental diets were included in the study: a diet based on plant protein with soy protein as the main protein source (Diet PP), a diet based on fishmeal in exchange for soy protein (Diet FM) and two diets in which different SPH replaced fishmeal in the FM diet. The experimental diets were fed to piglets from the day of weaning until 32 d postweaning. In addition to the record of performance data, an intestinal sampling for mucosal morphometry and microbiota 16S rRNA gene sequencing were performed at day 11 on a subset of the animals. The duodenal villi absorption area was significantly larger in piglets receiving Diets SPH compared with Diet PP (p < 0.02). A significant positive correlation between duodenal villi height and average daily gain during the first 11 d postweaning was detected. Only small differences in intestinal microbiota community and no differences in growth performance were detected between the experimental diets. To conclude, SPH seem to be an interesting novel protein source in weanling piglets.

  2. Angiotensin-I Converting Enzyme (ACE) Inhibitory and Anti-Hypertensive Effect of Protein Hydrolysate from Actinopyga lecanora (Sea Cucumber) in Rats

    PubMed Central

    Sadegh Vishkaei, Mahdokht; Ebrahimpour, Afshin; Abdul-Hamid, Azizah; Ismail, Amin; Saari, Nazamid

    2016-01-01

    Food protein hydrolysates are known to exhibit angiotensin converting enzyme (ACE) inhibitory properties and can be used as a novel functional food for prevention of hypertension. This study evaluated the ACE inhibitory potentials of Actinopyga lecanora proteolysate (ALP) in vivo. The pre-fed rats with ALP at various doses (200, 400, 800 mg/kg body weight) exhibited a significant (p ≤ 0.05) suppression effect after inducing hypertension. To determine the optimum effective dose that will produce maximal reduction in blood pressure, ALP at three doses was fed to the rats after inducing hypertension. The results showed that the 800 mg/kg body weight dose significantly reduced blood pressure without noticeable negative physiological effect. In addition, there were no observable changes in the rats’ heart rate after oral administration of the ALP. It was concluded that Actinopyga lecanora proteolysate could potentially be used for the development of functional foods and nutraceuticals for prevention and treatment of hypertension. PMID:27706040

  3. A fish protein hydrolysate alters fatty acid composition in liver and adipose tissue and increases plasma carnitine levels in a mouse model of chronic inflammation

    PubMed Central

    2013-01-01

    Background There is growing evidence that fish protein hydrolysate (FPH) diets affect mitochondrial fatty acid metabolism in animals. The aim of the study was to determine if FPH could influence fatty acid metabolism and inflammation in transgene mice expressing human tumor necrosis factor alpha (hTNFα). Methods hTNFα mice (C57BL/6 hTNFα) were given a high-fat (23%, w/w) diet containing 20% casein (control group) or 15% FPH and 5% casein (FPH group) for two weeks. After an overnight fast, blood, adipose tissue, and liver samples were collected. Gene expression and enzyme activity was analysed in liver, fatty acid composition was analyzed in liver and ovarian white adipose tissue, and inflammatory parameters, carnitine, and acylcarnitines were analyzed in plasma. Results The n-3/n-6 fatty acid ratio was higher in mice fed the FPH diet than in mice fed the control diet in both adipose tissue and liver, and the FPH diet affected the gene expression of ∆6 and ∆9 desaturases. Mice fed this diet also demonstrated lower hepatic activity of fatty acid synthase. Concomitantly, a lower plasma INF-γ level was observed. Plasma carnitine and the carnitine precursor γ-butyrobetaine was higher in the FPH-group compared to control, as was plasma short-chained and medium-chained acylcarnitine esters. The higher level of plasma acetylcarnitine may reflect a stimulated mitochondrial and peroxisomal β-oxidation of fatty acids, as the hepatic activities of peroxisomal acyl-CoA oxidase 1 and mitochondrial carnitine palmitoyltransferase-II were higher in the FPH-fed mice. Conclusions The FPH diet was shown to influence hepatic fatty acid metabolism and fatty acid composition. This indicates that effects on fatty acid metabolism are important for the bioactivity of protein hydrolysates of marine origin. PMID:24098955

  4. Extensive protein hydrolysate formula effectively reduces regurgitation in infants with positive and negative challenge tests for cow’s milk allergy

    PubMed Central

    Vandenplas, Y; De Greef, E

    2014-01-01

    Aim Cow’s milk protein allergy (CMPA) is treated using an elimination diet with an extensive protein hydrolysate. We explored whether a thickened or nonthickened version was best for infants with suspected CMPA, which commonly causes regurgitation/vomiting. Methods Diagnosis of CMPA was based on a positive challenge test. We compared the efficacy of two casein extensive hydrolysates (eCH), a nonthickened version (NT-eCH) and a thickened version (T-eCH), using a symptom-based score covering regurgitation, crying, stool consistency, eczema, urticarial and respiratory symptoms. Results A challenge was performed in 52/72 infants with suspected CMPA and was positive in 65.4%. All confirmed CMPA cases tolerated eCH. The symptom-based score decreased significantly in all infants within a month, and the highest reduction was in those with confirmed CMPA. Regurgitation was reduced in all infants (6.4 ± 3.2–2.8 ± 2.9, p < 0.001), but fell more with the T-eCH (−4.2 ± 3.2 regurgitations/day vs. −3.0 ± 4.5, ns), especially in infants with a negative challenge (−3.9 ± 4.0 vs. −1.9 ± 3.4, ns). Conclusion eCH fulfilled the criteria for a hypoallergenic formula, and the NT-eCH and T-eCH formulas both reduced CMPA symptoms. The symptom-based score is useful for evaluating how effective dietary treatments are for CMPA. PMID:24575806

  5. Influence of the rapeseed protein hydrolysis process on CHO cell growth.

    PubMed

    Chabanon, G; Alves da Costa, L; Farges, B; Harscoat, C; Chenu, S; Goergen, J-L; Marc, A; Marc, I; Chevalot, I

    2008-10-01

    Different protein hydrolysates were prepared from enzymatic hydrolyses of a rapeseed isolate (>90% protein content) using different commercial enzymes of non-animal origin. The extent of hydrolysis was controlled to produce hydrolysates corresponding to various degrees of hydrolysis (DH) from 5 to 30. These hydrolysates were characterized according to their solubility and size peptide pattern. Different growth behaviours of Chinese Hamster Ovary cells were observed when these various hydrolysates were added in serum-free medium containing transferrin, albumin and insulin. Hydrolysates from low degree of hydrolysis generally did not exhibit significant positive effect on cell growth; conversely hydrolysates from extensive hydrolysis, corresponding to a major low molecular size peptides content, usually allowed an increase of the maximal cell density. However, depending on the enzyme used, the supplementation with hydrolysates corresponding to a high degree of hydrolysis and composed of at least 70% peptides with a molecular size under 1kDa, led to different maximal cell density values, indicating the importance of enzyme specificity and consequently the nature of the released peptides. This result showed that the positive influence of the rapeseed hydrolysates on cell growth was not only due to a nutritional support tied to the addition of small peptides but may be related to the presence of peptides exhibiting growth or survival factor effects. Furthermore, total substitution of proteins (transferrin, albumin and insulin) in the cell culture medium by some rapeseed hydrolysates appeared to be a promising alternative to improve the cell growth in protein-free media.

  6. Effects of various feed supplements containing fish protein hydrolysate or fish processing by-products on the innate immune functions of juvenile coho salmon (oncorhynchus kisutch)

    USGS Publications Warehouse

    Murray, A.L.; Pascho, R.J.; Alcorn, S.W.; Fairgrieve, W.T.; Shearer, K.D.; Roley, D.

    2003-01-01

    Immunomodulators administered to fish in the diet have been shown in some cases to enhance innate immune defense mechanisms. Recent studies have suggested that polypeptide fractions found in fish protein hydrolysates may stimulate factors in fish important for disease resistance. For the current study, groups of coho salmon were reared on practical feeds that contained either fish meal (Control diet), fish meal supplemented with cooked fish by-products, or fish meal supplemented with hydrolyzed fish protein alone, or with hydrolyzed fish protein and processed fish bones. For each diet group, three replicate tanks of fish were fed the experimental diets for 6 weeks. Morphometric measurements, and serologic and cellular assays were used to evaluate the general health and immunocompetence of fish in the various feed groups. Whereas the experimental diets had no effect on the morphometric and cellular measurements, fish fed cooked by-products had increased leucocrit levels and lower hematocrit levels than fish from the other feed groups. Innate cellular responses were increased in all feed groups after feeding the four experimental diets compared with pre-feed results. Subgroups of fish from each diet group were also challenged with Vibrio anguillarum (ca. 7.71 ?? 105 bacteria ml-1) at 15??C by immersion. No differences were found in survival among the various feed groups.

  7. Protein hydrolysate of salted duck egg white as a substitute of phosphate and its effect on quality of Pacific white shrimp (Litopenaeus vannamei).

    PubMed

    Kaewmanee, Thammarat; Benjakul, Soottawat; Visessanguan, Wonnop

    2009-10-01

    Protein hydrolysate from salted egg white (PHSEW) with different degrees of hydrolysis (DH) (3%, 6%, and 9%) was produced using pepsin. Disappearance of proteins with molecular weight (MW) of 108 and 85 kDa with the concomitant formation of proteins with MW of 23, 20, 13, and 5 kDa was observed in PHSEW. The use of PHSEW for quality improvement of Pacific white shrimp (Litopenaeus vannamei) was investigated. Shrimp soaked in 4% NaCl containing 7% PHSEW and 2.5% mixed phosphates (0.625% sodium acid pyrophosphate [SAPP] and 1.875% tetrasodium pyrophosphate [TSPP]) had the highest cooking yield with the lowest cooking loss (P < 0.05), compared with shrimps with other treatments. Nevertheless, no difference in weight gain was obtained in comparison with those treated with 4% NaCl containing 3.5% mixed phosphate (P > 0.05). Cooked shrimp treated with 4% NaCl containing 7% PHSEW and 2.5% mixed phosphate or those treated with 4% NaCl containing 3.5% mixed phosphate had the higher score of appearance, texture, and overall likeness but less shear force, in comparison with the control (no treatment) (P < 0.05). Microstructure study revealed that muscle fibers of cooked shrimp from both treatments had the swollen fibrils and gaps, while the control had the swollen compact structure. Therefore, use of PHSEW could reduce phosphate residue in shrimps without an adverse effect on sensory properties.

  8. Preparation and use of reverse protein microarrays.

    PubMed

    Pin, Elisa; Federici, Giulia; Petricoin, Emanuel F

    2014-01-01

    Reverse-phase protein array (RPPA) is a multiplex, high-throughput proteomic technique for profiling the activation status of signal transduction pathways involved in cancer survival and progression, potentially allowing for identification of new biomarkers and drug targets. On RPPA, the entire patient proteome is immobilized on a spot and single proteins can be quantified across a set of samples, spotted on the same array, with high specificity and sensitivity. Array immunostaining and signal amplification systems are used to generate a signal proportional to the concentration of the analyte. Dedicated scanners and software are used to detect spots, measure intensity, subtract background, normalize signal, and generate a numeric value as output. The generated output file is then analyzed using several different bioinformatic and biostatistical tools. In this unit, the RPPA procedure is described in depth, from sample handling and preparation to data analysis, with particular emphasis on tissue sample analysis. PMID:24510676

  9. Short communication: Identification of iron-binding peptides from whey protein hydrolysates using iron (III)-immobilized metal ion affinity chromatography and reversed phase-HPLC-tandem mass spectrometry.

    PubMed

    Cruz-Huerta, Elvia; Martínez Maqueda, Daniel; de la Hoz, Lucia; da Silva, Vera S Nunes; Pacheco, Maria Teresa Bertoldo; Amigo, Lourdes; Recio, Isidra

    2016-01-01

    Peptides with iron-binding capacity obtained by hydrolysis of whey protein with Alcalase (Novozymes, Araucaria, PR, Brazil), pancreatin, and Flavourzyme (Novozymes) were identified. Hydrolysates were subjected to iron (III)-immobilized metal ion affinity chromatography, and the bound peptides were sequenced by mass spectrometry. Regardless of the enzyme used, the domains f(42-59) and f(125-137) from β-lactoglobulin enclosed most of identified peptides. This trend was less pronounced in the case of peptides derived from α-lactalbumin, with sequences deriving from diverse regions. Iron-bound peptides exhibited common structural characteristics, such as an abundance of Asp, Glu, and Pro, as revealed by mass spectrometry and AA analysis. In conclusion, this characterization of iron-binding peptides helps clarify the relationship between peptide structure and iron-chelating activity and supports the promising role of whey protein hydrolysates as functional ingredients in iron supplementation treatments.

  10. Short communication: Identification of iron-binding peptides from whey protein hydrolysates using iron (III)-immobilized metal ion affinity chromatography and reversed phase-HPLC-tandem mass spectrometry.

    PubMed

    Cruz-Huerta, Elvia; Martínez Maqueda, Daniel; de la Hoz, Lucia; da Silva, Vera S Nunes; Pacheco, Maria Teresa Bertoldo; Amigo, Lourdes; Recio, Isidra

    2016-01-01

    Peptides with iron-binding capacity obtained by hydrolysis of whey protein with Alcalase (Novozymes, Araucaria, PR, Brazil), pancreatin, and Flavourzyme (Novozymes) were identified. Hydrolysates were subjected to iron (III)-immobilized metal ion affinity chromatography, and the bound peptides were sequenced by mass spectrometry. Regardless of the enzyme used, the domains f(42-59) and f(125-137) from β-lactoglobulin enclosed most of identified peptides. This trend was less pronounced in the case of peptides derived from α-lactalbumin, with sequences deriving from diverse regions. Iron-bound peptides exhibited common structural characteristics, such as an abundance of Asp, Glu, and Pro, as revealed by mass spectrometry and AA analysis. In conclusion, this characterization of iron-binding peptides helps clarify the relationship between peptide structure and iron-chelating activity and supports the promising role of whey protein hydrolysates as functional ingredients in iron supplementation treatments. PMID:26601589

  11. Porcine Splenic Hydrolysate has Antioxidant Activity in vivo and in vitro

    PubMed Central

    Yoon, Taek Joon

    2014-01-01

    The antioxidant capacity of porcine splenic hydrolysate (PSH) was studied in vitro and in vivo. Peptide hydrolysates were prepared, using the proteolytic enzyme Alcalase®. The molecular weights of PSH were 37,666, 10,673, 6,029, and 2,918 g/mol. Rats were fed a 5% (w/v) PSH diet, instead of a casein diet, for 4 wk. The food intake, body weight gain, and liver weight of rats in the PSH group were similar to those in the control (CONT) group. There were no differences in the serum total cholesterol, triglyceride, total protein, or albumin levels between PSH and CONT groups. However, the level of in vivo hepatic lipid peroxidation in PSH group was significantly lower than that in CONT. In vivo hepatic catalase and glutathione peroxidase activities in the PSH group were significantly higher than those in the control group. The in vitro protein digestibility of PSH was lower than that of casein. The in vitro trolox equivalent antioxidant capacity of PSH was significantly higher than that of the peptide hydrolysate from casein. The in vitro radical scavenging activities of PSH were significantly higher than those of the peptide hydrolysate from casein. The present findings suggest that porcine splenic peptides improve the antioxidant status in rats by enhancing hepatic catalase and GSH-Px activities, and indicate a potential mechanism of radical scavenging activity during gastrointestinal passage. PMID:26761173

  12. Generation of bioactive peptide hydrolysates from cattle plasma using plant and fungal proteases.

    PubMed

    Bah, Clara S F; Bekhit, Alaa El-Din A; McConnell, Michelle A; Carne, Alan

    2016-12-15

    Four protease preparations from plant and fungal sources (papain, bromelain, FP400 and FPII) were used to hydrolyse plasma which was separated from slaughterhouse cattle blood. The o-phthaldialdehyde assay was used to follow the release of TCA-soluble peptides over a 24h period. Hydrolysis profiles were displayed using SDS-PAGE. The in vitro antioxidant and antimicrobial activities of the hydrolysates were determined. The results showed that hydrolysates of cattle plasma generated with fungal protease FPII had higher antioxidant activities. Overall than hydrolysates generated with papain, bromelain and FP400. None of the hydrolysates demonstrated antimicrobial activity. The FPII peptide hydrolysate was fractionated using gel permeation chromatography, OFFGEL isoelectric focusing and RP-HPLC. The RP-HPLC fraction with highest antioxidant activity contained 15 novel peptide sequences. The use of protease FPII to hydrolyse cattle plasma resulted in a hydrolysate with high antioxidant properties and unique peptide sequences. PMID:27451160

  13. Generation of bioactive peptide hydrolysates from cattle plasma using plant and fungal proteases.

    PubMed

    Bah, Clara S F; Bekhit, Alaa El-Din A; McConnell, Michelle A; Carne, Alan

    2016-12-15

    Four protease preparations from plant and fungal sources (papain, bromelain, FP400 and FPII) were used to hydrolyse plasma which was separated from slaughterhouse cattle blood. The o-phthaldialdehyde assay was used to follow the release of TCA-soluble peptides over a 24h period. Hydrolysis profiles were displayed using SDS-PAGE. The in vitro antioxidant and antimicrobial activities of the hydrolysates were determined. The results showed that hydrolysates of cattle plasma generated with fungal protease FPII had higher antioxidant activities. Overall than hydrolysates generated with papain, bromelain and FP400. None of the hydrolysates demonstrated antimicrobial activity. The FPII peptide hydrolysate was fractionated using gel permeation chromatography, OFFGEL isoelectric focusing and RP-HPLC. The RP-HPLC fraction with highest antioxidant activity contained 15 novel peptide sequences. The use of protease FPII to hydrolyse cattle plasma resulted in a hydrolysate with high antioxidant properties and unique peptide sequences.

  14. In vivo antitumor and antioxidative effects of a rapeseed meal protein hydrolysate on an S180 tumor-bearing murine model.

    PubMed

    Xue, Zhaohui; Yu, Wancong; Wu, Moucheng; Wang, Jiehua

    2009-11-01

    The antitumor and antioxidative activities of a rapeseed protein hydrolysate (RSCH) obtained from rapeseed meal were evaluated by using an in vivo S180 tumor-bearing Kunming mice model. Tumor-bearing female mice were given RSCH for 10 at doses of 0, 50, 100, and 150 mg/kg/d by gastric perfusion. RSCH significantly decreased the tumor weight by 44% and 53% in the 100 and 150 mg/kg/d groups, respectively, without causing mortality or growth retardation. The thymus and spleen indices (organ weight relative to body weight) were increased significantly in the 150 mg/kg/d group. The phagocytic capability of coeliac macrophages and delayed-type hypersensitivity (DTH) were significantly increased in tumor-bearing mice treated with RSCH at 150 mg/kg/d. RSCH administration also enhanced the superoxide dismutase activity and reduced the serum level of thiobarbituric acid reactive substances. Our results show that an oral RSCH administration had an antitumor protective effect and may improve immune function by reducing free radical formation and oxidative stress in a murine model.

  15. Rice Bran Protein Hydrolysates Improve Insulin Resistance and Decrease Pro-inflammatory Cytokine Gene Expression in Rats Fed a High Carbohydrate-High Fat Diet.

    PubMed

    Boonloh, Kampeebhorn; Kukongviriyapan, Veerapol; Kongyingyoes, Bunkerd; Kukongviriyapan, Upa; Thawornchinsombut, Supawan; Pannangpetch, Patchareewan

    2015-08-01

    A high carbohydrate-high fat (HCHF) diet causes insulin resistance (IR) and metabolic syndrome (MS). Rice bran has been demonstrated to have anti-dyslipidemic and anti-atherogenic properties in an obese mouse model. In the present study, we investigated the beneficial effects of rice bran protein hydrolysates (RBP) in HCHF-induced MS rats. After 12 weeks on this diet, the HCHF-fed group was divided into four subgroups, which were orally administered RBP 100 or 500 mg/kg, pioglitazone 10 mg/kg, or tap water for a further 6 weeks. Compared with normal diet control group, the MS rats had elevated levels of blood glucose, lipid, insulin, and HOMA-IR. Treatment with RBP significantly alleviated all those changes and restored insulin sensitivity. Additionally, RBP treatment increased adiponectin and suppressed leptin levels. Expression of Ppar-γ mRNA in adipose tissues was significantly increased whereas expression of lipogenic genes Srebf1 and Fasn was significantly decreased. Levels of mRNA of proinflammatory cytokines, Il-6, Tnf-α, Nos-2 and Mcp-1 were significantly decreased. In conclusion, the present findings support the consumption of RBP as a functional food to improve insulin resistance and to prevent the development of metabolic syndrome. PMID:26247962

  16. Rice Bran Protein Hydrolysates Improve Insulin Resistance and Decrease Pro-inflammatory Cytokine Gene Expression in Rats Fed a High Carbohydrate-High Fat Diet.

    PubMed

    Boonloh, Kampeebhorn; Kukongviriyapan, Veerapol; Kongyingyoes, Bunkerd; Kukongviriyapan, Upa; Thawornchinsombut, Supawan; Pannangpetch, Patchareewan

    2015-08-01

    A high carbohydrate-high fat (HCHF) diet causes insulin resistance (IR) and metabolic syndrome (MS). Rice bran has been demonstrated to have anti-dyslipidemic and anti-atherogenic properties in an obese mouse model. In the present study, we investigated the beneficial effects of rice bran protein hydrolysates (RBP) in HCHF-induced MS rats. After 12 weeks on this diet, the HCHF-fed group was divided into four subgroups, which were orally administered RBP 100 or 500 mg/kg, pioglitazone 10 mg/kg, or tap water for a further 6 weeks. Compared with normal diet control group, the MS rats had elevated levels of blood glucose, lipid, insulin, and HOMA-IR. Treatment with RBP significantly alleviated all those changes and restored insulin sensitivity. Additionally, RBP treatment increased adiponectin and suppressed leptin levels. Expression of Ppar-γ mRNA in adipose tissues was significantly increased whereas expression of lipogenic genes Srebf1 and Fasn was significantly decreased. Levels of mRNA of proinflammatory cytokines, Il-6, Tnf-α, Nos-2 and Mcp-1 were significantly decreased. In conclusion, the present findings support the consumption of RBP as a functional food to improve insulin resistance and to prevent the development of metabolic syndrome.

  17. Seed vigour studies in corn, soybean and tomato in response to fish protein hydrolysates and consequences on phenolic-linked responses.

    PubMed

    Horii, Akiyo; McCue, Patrick; Shetty, Kalidas

    2007-08-01

    Seed priming with fish protein hydrolysates (FPH) has been studied for the enhancement of seed vigour of corn, soybean and tomato. The influence of FPH at 2.5 mL/L and 5.0 mL/L on traditional agronomic parameters for seed vigour (germination percentage, seedling weight, seedling height) and potential new vigour-associated parameters (phenolic content, antioxidant activity, guaiacol peroxidase (GuPX) activity, chlorophyll content) was investigated. FPH treatment preferentially stimulated seedling vigour in the following order: soybean>tomato>corn. For soybean, FPH at 2.5 mL/L and 5 mL/L improved the majority of the seed vigour parameters (seedling weight and height, phenolic content, antioxidant activity and chlorophyll content, and lignification-associated GuPX activity). Similarly, for tomato, FPH at 2.5 mL/L stimulated seedling weight and height, GuPX activity and chlorophyll content. However, FPH did not stimulate corn seed vigour. Our results suggest an ability of proline precursor-rich FPH to improve of plant growth and development (e.g., seed vigour) in phenolic-rich plant species through modulation of phenolic and chlorophyll metabolisms.

  18. [Antigenicity and allergenicity of hypoallergenic hydrolysate for infant nutrition].

    PubMed

    Görtler, I; Urbanek, R

    1990-09-01

    The antigenicity/allergenicity of protein components in hypoallergenic formulae is altered by hydrolysis. Two different hydrolysate formulae, hydrolysate 1 = cow milk based; hydrolysate 2 = soya/beef collagen based, were investigated with respect to their specific IgG/IgE binding capacities using the sera of 41 healthy controls, 40 atopic and 12 cow milk allergic subjects. Furthermore, histamine liberation from basophils on incubation with milk proteins and the hydrolysates was measured in 5 healthy and 3 cow milk allergic individuals. Nearly all probands demonstrated specific IgG binding with hypoallergenic formulae. Anti-hydrolysate 1 IgG titres were higher than titres against hydrolysate 2 in the cow milk allergic and healthy populations. Nonhydrolyzed cow milk elicited IgE binding in all cow milk allergic sera. IgE-antibody for hydrolysate 2 could only be demonstrated in one atopic subject. Hydrolysate 1 showed binding capacity for the IgE-antibody of one atopic and 3/12 cow milk allergic patients. Histamine liberation followed in-vitro incubation with both hydrolysates in one out of 3 cow milk allergic subjects and two out of these cases following incubation to unprocessed cow milk protein. A decreased antigenicity/allergenicity can be demonstrated for the two hydrolysates investigated, however antigenic/allergenic reactivity is still present to some degree. Therefore, the therapeutic application of hypoallergenic formulae in patients with specific sensitization to cow milk should be based on the results of the above-mentioned in-vitro parameters and if necessary skin tests and oral challenges.

  19. Biocatalytic conversion of poultry processing leftovers: Optimization of hydrolytic conditions and peptide hydrolysate characterization.

    PubMed

    Nikolaev, I V; Sforza, S; Lambertini, F; Ismailova, D Yu; Khotchenkov, V P; Volik, V G; Dossena, A; Popov, V O; Koroleva, O V

    2016-04-15

    Peptide hydrolysate (PH) was produced by deep controllable bioconversion of poultry processing leftovers (broiler necks), by means of a multienzyme composition, containing four commercially available enzyme preparations (Alcalase, Neutrase, Flavourzyme, Protamex). The design of multienzyme composition (MEC) was applied to yield a hydrolysate with adjusted properties, including minimized antigenicity and bitterness. The protein recovery was optimized using Box-Behnken response surface design. The individual and interactive effects of hydrolysis conditions (time, hydromodule and MEC dosage) were studied. The experimental data were analyzed by ANOVA method and a well-predictive, second order polynomial model was developed using multiple regression analysis. Optimal hydrolysis conditions were found to be: hydrolysis time 3 h, hydromodule 2.25 l/kg and dosage of MEC 0.25%. The corresponding predicted value for protein recovery was 75.34%, 2 times higher compared to traditional long-term heating hydrolysis. The PH obtained is a low allergenic product with high antioxidant capacity.

  20. Lipolysis stimulating peptides of potato protein hydrolysate effectively suppresses high-fat-diet-induced hepatocyte apoptosis and fibrosis in aging rats

    PubMed Central

    Chiang, Wen-Dee; Huang, Chih Yang; Paul, Catherine Reena; Lee, Zong-Yan; Lin, Wan-Teng

    2016-01-01

    Background Non-alcoholic fatty liver disease (NAFLD) is one of the most common outcomes of obesity and is characterized by the accumulation of triglycerides, increased tissue apoptosis, and fibrosis. NAFLD is more common among elderly than in younger age groups, and it causes serious hepatic complications. Objective In this study, alcalase treatment derived potato protein hydrolysate (APPH) with lipolysis-stimulating property has been evaluated for its efficiency to provide hepato-protection in a high-fat-diet (HFD)-fed aging rats. Design Twenty-four-month-old SD rats were randomly divided into six groups (n=8): aged rats fed with standard chow, HFD-induced aged obese rats, HFD with low-dose (15 mg/kg/day) APPH treatment, HFD with moderate (45 mg/kg/day) APPH treatment, HFD with high (75 mg/kg/day) APPH treatment, and HFD with probucol. Results APPH was found to reduce the NAFLD-related effects in rat livers induced by HFD and all of the HFD-fed rats exhibited heavier body weight than those with control chow diet. However, the HFD-induced hepatic fat accumulation was effectively attenuated in rats administered with low (15 mg/kg/day), moderate (45 mg/kg/day), and high (75 mg/kg/day) doses of APPH. APPH oral administration also suppressed the hepatic apoptosis- and fibrosis-related proteins induced by HFD. Conclusions Our results thus indicate that APPH potentially attenuates hepatic lipid accumulation and anti-apoptosis and fibrosis effects in HFD-induced rats. APPH may have therapeutic potential in the amelioration of NAFLD liver damage. PMID:27415158

  1. Determination of nutritional and bioactive properties of peptides in enzymatic pea, chickpea, and mung bean protein hydrolysates.

    PubMed

    Aluko, Rotimi E

    2008-01-01

    Within the primary structure of many pea and mung bean proteins are peptide sequences that can potentially be used in the formulation of therapeutic products for the treatment and prevention of human diseases. However, these peptide sequences need protease treatments before they can be released free of the parent proteins. Unlike chemical hydrolysis, enzymatic treatment enables more efficient tailoring of peptide products without formation of toxic by-products or destruction of amino acids. This review provides information on current methods that have been used to convert inactive pea and mung bean proteins into bioactive peptides. It focuses on 3 main bioactive properties, such as inhibitions of (1) angiotensin converting enzyme (ACE) activity; (2) calmodulin (CaM)-dependent enzymes; and (3) copper-chelating activity. ACE is an established marker for hypertension, high levels of some CaM-dependent enzymes are risk factors for various human diseases including cancer and Alzheimer's disease, and high vascular copper concentrations may potentiate atherosclerosis. Also reviewed are the production and evaluation of activity of hypoallergenic peptides that may offer protection against anaphylactic reactions. The 3 main proteins discussed are chickpea, mung bean, and field pea.

  2. Chronic treatment with a tryptophan-rich protein hydrolysate improves emotional processing, mental energy levels and reaction time in middle-aged women.

    PubMed

    Mohajeri, M H; Wittwer, J; Vargas, K; Hogan, E; Holmes, A; Rogers, P J; Goralczyk, R; Gibson, E L

    2015-01-28

    Common pharmacological treatments of mood disorders aim to modulate serotonergic neurotransmission and enhance serotonin levels in the brain. Brain serotonin levels are dependent on the availability of its food-derived precursor essential amino acid tryptophan (Trp). We tested the hypothesis that delivery of Trp via food may serve as an alternative treatment, and examined the effects of a Trp-rich, bioavailable dietary supplement from egg protein hydrolysate on cognitive and emotional functions, mood state, and sleep quality. In a randomised, placebo-controlled, parallel trial, fifty-nine mentally and physically healthy women aged 45-65 years received placebo (n 30) or the supplement (n 29) (both as 0.5 g twice per d) for 19 d. Emotional processing was significantly changed by supplementation, exhibiting a shift in bias away from negative stimuli. The results for the Affective Go/No-Go Task exhibited a slowing of responses to negative words, suggesting reduced attention to negative emotional stimuli. The results for the Facial Emotional Expression Rating Task also supported a shift away from attention to negative emotions and a bias towards happiness. An increase in arousal-like symptoms, labelled 'high energy', shorter reaction times and a slight benefit to sustained attention were observed in the treated subjects. Finally, when the supplement was taken 60-90 min before bedtime, a feeling of happiness before going to bed was consistently reported. In summary, daily consumption of a low-dose supplement containing bioavailable Trp may have beneficial effects on emotional and cognitive functions. PMID:25572038

  3. Chronic treatment with a tryptophan-rich protein hydrolysate improves emotional processing, mental energy levels and reaction time in middle-aged women.

    PubMed

    Mohajeri, M H; Wittwer, J; Vargas, K; Hogan, E; Holmes, A; Rogers, P J; Goralczyk, R; Gibson, E L

    2015-01-28

    Common pharmacological treatments of mood disorders aim to modulate serotonergic neurotransmission and enhance serotonin levels in the brain. Brain serotonin levels are dependent on the availability of its food-derived precursor essential amino acid tryptophan (Trp). We tested the hypothesis that delivery of Trp via food may serve as an alternative treatment, and examined the effects of a Trp-rich, bioavailable dietary supplement from egg protein hydrolysate on cognitive and emotional functions, mood state, and sleep quality. In a randomised, placebo-controlled, parallel trial, fifty-nine mentally and physically healthy women aged 45-65 years received placebo (n 30) or the supplement (n 29) (both as 0.5 g twice per d) for 19 d. Emotional processing was significantly changed by supplementation, exhibiting a shift in bias away from negative stimuli. The results for the Affective Go/No-Go Task exhibited a slowing of responses to negative words, suggesting reduced attention to negative emotional stimuli. The results for the Facial Emotional Expression Rating Task also supported a shift away from attention to negative emotions and a bias towards happiness. An increase in arousal-like symptoms, labelled 'high energy', shorter reaction times and a slight benefit to sustained attention were observed in the treated subjects. Finally, when the supplement was taken 60-90 min before bedtime, a feeling of happiness before going to bed was consistently reported. In summary, daily consumption of a low-dose supplement containing bioavailable Trp may have beneficial effects on emotional and cognitive functions.

  4. Light backscatter fiber optic sensor: a new tool for predicting the stability of pork emulsions containing antioxidative potato protein hydrolysate.

    PubMed

    Nieto, Gema; Xiong, Youling L; Payne, Fred; Castillo, Manuel

    2015-02-01

    The objective of this study was to determine whether light backscatter response from fresh pork meat emulsions is correlated to final product stability indices. A specially designed fiber optic measurement system was used in combination with a miniature fiber optic spectrometer to determine the intensity of light backscatter within the wavelength range 300-1100 nm (UV/VIS/NIR) at different radial distances (2, 2.5 and 3mm) with respect to the light source in pork meat emulsions with two fat levels (15%, 30%) and two levels (0, 2.5%) of the natural antioxidant hydrolyzed potato protein (HPP). Textural parameters (hardness, deformability, cohesiveness and breaking force), cooking loss, TBARS (1, 2, 3, and 7 days of storage at 4 °C) and CIELAB color coordinates of cooked emulsions were measured. The light backscatter was directly correlated with cooking losses, color, breaking force and TBARS. The optical configuration proposed would compensate for the emulsion heterogeneity, maximizing the existing correlation between the optical signal and the emulsion quality metrics.

  5. Light backscatter fiber optic sensor: a new tool for predicting the stability of pork emulsions containing antioxidative potato protein hydrolysate.

    PubMed

    Nieto, Gema; Xiong, Youling L; Payne, Fred; Castillo, Manuel

    2015-02-01

    The objective of this study was to determine whether light backscatter response from fresh pork meat emulsions is correlated to final product stability indices. A specially designed fiber optic measurement system was used in combination with a miniature fiber optic spectrometer to determine the intensity of light backscatter within the wavelength range 300-1100 nm (UV/VIS/NIR) at different radial distances (2, 2.5 and 3mm) with respect to the light source in pork meat emulsions with two fat levels (15%, 30%) and two levels (0, 2.5%) of the natural antioxidant hydrolyzed potato protein (HPP). Textural parameters (hardness, deformability, cohesiveness and breaking force), cooking loss, TBARS (1, 2, 3, and 7 days of storage at 4 °C) and CIELAB color coordinates of cooked emulsions were measured. The light backscatter was directly correlated with cooking losses, color, breaking force and TBARS. The optical configuration proposed would compensate for the emulsion heterogeneity, maximizing the existing correlation between the optical signal and the emulsion quality metrics. PMID:25460135

  6. A whey protein hydrolysate promotes insulinotropic activity in a clonal pancreatic β-cell line and enhances glycemic function in ob/ob mice.

    PubMed

    Gaudel, Celine; Nongonierma, Alice B; Maher, Samuel; Flynn, Sarah; Krause, Mauricio; Murray, Brian A; Kelly, Phillip M; Baird, Alan W; FitzGerald, Richard J; Newsholme, Philip

    2013-07-01

    Whey protein hydrolysates (WPHs) represent novel antidiabetic agents that affect glycemia in animals and humans, but little is known about their insulinotropic effects. The effects of a WPH were analyzed in vitro on acute glucose-induced insulin secretion in pancreatic BRIN-BD11 β cells. WPH permeability across Caco-2 cell monolayers was determined in a 2-tiered intestinal model. WPH effects on insulin resistance were studied in vivo following an 8-wk oral ingestion (100 mg/kg body weight) by ob/ob (OB-WPH) and wild-type mice (WT-WPH) compared with vehicle control (OB and WT groups) using a 2 × 2 factorial design, genotype × treatment. BRIN-BD11 cells showed a robust and reproducible dose-dependent insulinotropic effect of WPH (from 0.01 to 5.00 g/L). WPH bioactive constituents were permeable across Caco-2 cell monolayers. In the OB-WPH and WT-WPH groups, WPH administration improved glucose clearance after a glucose challenge (2 g/kg body weight), as indicated by differences in the area under curves (AUCs) (P ≤ 0.05). The basal plasma glucose concentration was not affected by WPH treatment in either genotype. The plasma insulin concentration was lower in the OB-WPH than in the OB group (P ≤ 0.005) but was similar between the WT and WT-WPH groups; the interaction genotype × treatment was significant (P ≤ 0.005). Insulin release from pancreatic islets isolated from the OB-WPH group was greater (P ≤ 0.005) than that from the OB group but did not differ between the WT-WPH and WT groups; the interaction genotype × treatment was not significant. In conclusion, an 8-wk oral administration of WPH improved blood glucose clearance, reduced hyperinsulinemia, and restored the pancreatic islet capacity to secrete insulin in response to glucose in ob/ob mice. Hence, it may be useful in diabetes management.

  7. In vitro digestion of short-dough biscuits enriched in proteins and/or fibres using a multi-compartmental and dynamic system (2): Protein and starch hydrolyses.

    PubMed

    Villemejane, C; Denis, S; Marsset-Baglieri, A; Alric, M; Aymard, P; Michon, C

    2016-01-01

    The influence of protein and/or fibre enrichment on the nutritional properties of biscuits was studied in terms of proteolysis and amylolysis. Biscuits were digested using a multi-compartmental and dynamic system that simulates the main physiological digestive functions of the upper tract of healthy adult humans: the TIM-1. A control biscuit and three biscuits enriched in proteins and/or fibres were digested under the same conditions. Samples were collected in each compartment of the TIM-1 (stomach, duodenum, jejunum and ileum) at different times of digestion and analysed in terms of proteolysis and amylolysis. Results indicate that both formulation and processing impacted the digestive fate of the biscuits. Incorporating proteins or fibres in biscuits lowered or delayed proteolysis. Moreover a protein-plus-fibre additional or synergic effect was observed. Biscuits enriched in proteins and/or fibres displayed a higher amylolysis degree than the control biscuit, probably due to lower starch amounts and higher gelatinization degrees.

  8. Ethanolic fermentation of lignocellulose hydrolysates

    SciTech Connect

    Hahn-Haegerdal, B.

    1996-12-31

    This minireview discusses various factors which require consideration for the ethanolic fermentation of lignocellulose hydrolysates. The production of an alternative transportation fuel requires pretreatment of the biomass and detoxification to enhance the fermentability. Recombinant DNA technology makes it possible to engineer new microorganisms for efficient ethanol production from all sugars present in the hydrolysates. 60 refs.

  9. Electrodeposited gels prepared from protein alloys

    PubMed Central

    Lin, Yinan; Wang, Siran; Chen, Ying; Wang, Qianrui; Burke, Kelly A; Spedden, Elise M; Staii, Cristian; Weiss, Anthony S; Kaplan, David L

    2015-01-01

    Aim Silk-tropoelastin alloys, composed of recombinant human tropoelastin and regenerated Bombyx mori silk fibroin, are an emerging, versatile class of biomaterials endowed with tunable combinations of physical and biological properties. Electrodeposition of these alloys provides a programmable means to assemble functional gels with both spatial and temporal controllability. Materials & methods Tropoelastin-modified silk was prepared by enzymatic coupling between tyrosine residues. Hydrogel coatings were electrodeposited using two wire electrodes. Results & discussion Mechanical characterization and in vitro cell culture revealed enhanced adhesive capability and cellular response of these alloy gels as compared with electrogelled silk alone. Conclusion These electro-depositable silk-tropoelastin alloys constitute a suitable coating material for nanoparticle-based drug carriers and offer a novel opportunity for on-demand encapsulation/release of nanomedicine. PMID:25816881

  10. Ethanolic fermentation of pentoses in lignocellulose hydrolysates

    SciTech Connect

    Hahn-Haegerdal, B.; Linden, T.; Senac, T.; Skoog, K.

    1991-12-31

    In the fermentation of lignocellulose hydrolysates to ethanol, two major problems are encountered: the fermentation of the pentose sugar xylose, and the presence of microbial inhibitors. Xylose can be directly fermented with yeasts; such as Pachysolen tannophilus, Candida shehatae, and Pichia stipis, or by isomerization of xylose to xylulose with the enzyme glucose (xylose) isomerase, and subsequent fermentation with bakers yeast, Saccharomyces cerevisiae. The direct fermentation requires low, carefully controlled oxygenation, as well as the removal of inhibitors. Also, the xylose-fermenting yeasts have a limited ethanol tolerance. The combined isomerization and fermentation with XI and S. cerevisiae gives yields and productivities comparable to those obtained in hexose fermentations without oxygenation and removal of inhibitors. However, the enzyme is not very stable in a lignocellulose hydrolysate, and S. cerevisiae has a poorly developed pentose phosphate shunt. Different strategies involving strain adaptation, and protein and genetic engineering adopted to overcome these different obstacles, are discussed.

  11. Long-Term Fungal Inhibition by Pisum sativum Flour Hydrolysate during Storage of Wheat Flour Bread

    PubMed Central

    Lavecchia, Anna; Gramaglia, Valerio; Gobbetti, Marco

    2015-01-01

    In order to identify antifungal compounds from natural sources to be used as ingredients in the bakery industry, water/salt-soluble extracts (WSE) from different legume flour hydrolysates obtained by the use of a fungal protease were assayed against Penicillium roqueforti DPPMAF1. The agar diffusion assays allowed the selection of the pea (Pisum sativum) hydrolysate as the most active. As shown by the hyphal radial growth rate, the WSE had inhibitory activity towards several fungi isolated from bakeries. The MIC of the WSE was 9.0 mg/ml. Fungal inhibition was slightly affected by heating and variations in pH. The antifungal activity was attributed to three native proteins (pea defensins 1 and 2 and a nonspecific lipid transfer protein [nsLTP]) and a mixture of peptides released during hydrolysis. The three proteins have been reported previously as components of the defense system of the plant. Five peptides were purified from WSE and were identified as sequences encrypted in leginsulin A, vicilin, provicilin, and the nsLTP. To confirm antifungal activity, the peptides were chemically synthesized and tested. Freeze-dried WSE were used as ingredients in leavened baked goods. In particular, breads made by the addition of 1.6% (wt/wt) of the extract and fermented by baker's yeast or sourdough were characterized for their main chemical, structural, and sensory features, packed in polyethylene bags, stored at room temperature, and compared to controls prepared without pea hydrolysate. Artificially inoculated slices of a bread containing the WSE did not show contamination by fungi until at least 21 days of storage and behaved like the bread prepared with calcium propionate (0.3%, wt/wt). PMID:25862230

  12. Long-Term Fungal Inhibition by Pisum sativum Flour Hydrolysate during Storage of Wheat Flour Bread.

    PubMed

    Rizzello, Carlo Giuseppe; Lavecchia, Anna; Gramaglia, Valerio; Gobbetti, Marco

    2015-06-15

    In order to identify antifungal compounds from natural sources to be used as ingredients in the bakery industry, water/salt-soluble extracts (WSE) from different legume flour hydrolysates obtained by the use of a fungal protease were assayed against Penicillium roqueforti DPPMAF1. The agar diffusion assays allowed the selection of the pea (Pisum sativum) hydrolysate as the most active. As shown by the hyphal radial growth rate, the WSE had inhibitory activity towards several fungi isolated from bakeries. The MIC of the WSE was 9.0 mg/ml. Fungal inhibition was slightly affected by heating and variations in pH. The antifungal activity was attributed to three native proteins (pea defensins 1 and 2 and a nonspecific lipid transfer protein [nsLTP]) and a mixture of peptides released during hydrolysis. The three proteins have been reported previously as components of the defense system of the plant. Five peptides were purified from WSE and were identified as sequences encrypted in leginsulin A, vicilin, provicilin, and the nsLTP. To confirm antifungal activity, the peptides were chemically synthesized and tested. Freeze-dried WSE were used as ingredients in leavened baked goods. In particular, breads made by the addition of 1.6% (wt/wt) of the extract and fermented by baker's yeast or sourdough were characterized for their main chemical, structural, and sensory features, packed in polyethylene bags, stored at room temperature, and compared to controls prepared without pea hydrolysate. Artificially inoculated slices of a bread containing the WSE did not show contamination by fungi until at least 21 days of storage and behaved like the bread prepared with calcium propionate (0.3%, wt/wt). PMID:25862230

  13. Long-Term Fungal Inhibition by Pisum sativum Flour Hydrolysate during Storage of Wheat Flour Bread.

    PubMed

    Rizzello, Carlo Giuseppe; Lavecchia, Anna; Gramaglia, Valerio; Gobbetti, Marco

    2015-06-15

    In order to identify antifungal compounds from natural sources to be used as ingredients in the bakery industry, water/salt-soluble extracts (WSE) from different legume flour hydrolysates obtained by the use of a fungal protease were assayed against Penicillium roqueforti DPPMAF1. The agar diffusion assays allowed the selection of the pea (Pisum sativum) hydrolysate as the most active. As shown by the hyphal radial growth rate, the WSE had inhibitory activity towards several fungi isolated from bakeries. The MIC of the WSE was 9.0 mg/ml. Fungal inhibition was slightly affected by heating and variations in pH. The antifungal activity was attributed to three native proteins (pea defensins 1 and 2 and a nonspecific lipid transfer protein [nsLTP]) and a mixture of peptides released during hydrolysis. The three proteins have been reported previously as components of the defense system of the plant. Five peptides were purified from WSE and were identified as sequences encrypted in leginsulin A, vicilin, provicilin, and the nsLTP. To confirm antifungal activity, the peptides were chemically synthesized and tested. Freeze-dried WSE were used as ingredients in leavened baked goods. In particular, breads made by the addition of 1.6% (wt/wt) of the extract and fermented by baker's yeast or sourdough were characterized for their main chemical, structural, and sensory features, packed in polyethylene bags, stored at room temperature, and compared to controls prepared without pea hydrolysate. Artificially inoculated slices of a bread containing the WSE did not show contamination by fungi until at least 21 days of storage and behaved like the bread prepared with calcium propionate (0.3%, wt/wt).

  14. Mechanism of the discrepancy in the enzymatic hydrolysis efficiency between defatted peanut flour and peanut protein isolate by Flavorzyme.

    PubMed

    Zheng, Lin; Zhao, Yijun; Xiao, Chuqiao; Sun-Waterhouse, Dongxiao; Zhao, Mouming; Su, Guowan

    2015-02-01

    Both defatted peanut flour (DPF) and peanut protein isolate (PPI) are widely used to prepare peanut protein hydrolysates. To compare their enzymatic hydrolysis efficiencies, DPF and PPI were hydrolysed by Alcalase, Neutrase, Papain, Protamex and Flavorzyme. Alcalase and Flavorzyme were found to be the most efficient proteases to hydrolyse both DPF and PPI. The efficiency was comparable to each other when using Alcalase, while PPI was hydrolysed less efficiently than DPF when using Flavorzyme. Analysis of changes in the protein solubility, subunit and conformation, and amino acid composition of DPF, PPI and their Flavorzyme hydrolysis residues indicated that the PPI preparation process had minimal effect on it, but peptide aggregation via non-covalent bonding (including hydrophobic interactions and hydrogen bonds) during hydrolysis and/or thermal treatment after hydrolysis were likely responsible for the reduced hydrolysis efficiency of PPI by Flavorzyme.

  15. Evaluation of Catfish Skin Hydrolysates as a Glazing Material for Air-Blast Frozen Shrimp

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Catfish is one of the most widely consumed seafood in the United States. A by-product of this consumption is a large quantity of catfish skin (CS), approximately 8,200 metric tons in 2014. Enzymatic hydrolysis is used to produce protein hydrolysates from the skin. These hydrolysates have considerabl...

  16. Preparation of Modified Films with Protein from Grouper Fish

    PubMed Central

    Tecante, A.; Granados-Navarrete, S.; Martínez-García, C.

    2016-01-01

    A protein concentrate (PC) was obtained from Grouper fish skin and it was used to prepare films with different amounts of sorbitol and glycerol as plasticizers. The best performing films regarding resistance were then modified with various concentrations of CaCl2, CaSO4 (calcium salts), and glucono-δ-lactone (GDL) with the purpose of improving their mechanical and barrier properties. These films were characterized by determining their mechanical properties and permeability to water vapor and oxygen. Formulations with 5% (w/v) protein and 75% sorbitol and 4% (w/v) protein with a mixture of 15% glycerol and 15% sorbitol produced adequate films. Calcium salts and GDL increased the tensile fracture stress but reduced the fracture strain and decreased water vapor permeability compared with control films. The films prepared represent an attractive alternative for being used as food packaging materials. PMID:27597950

  17. Preparation of Modified Films with Protein from Grouper Fish

    PubMed Central

    Tecante, A.; Granados-Navarrete, S.; Martínez-García, C.

    2016-01-01

    A protein concentrate (PC) was obtained from Grouper fish skin and it was used to prepare films with different amounts of sorbitol and glycerol as plasticizers. The best performing films regarding resistance were then modified with various concentrations of CaCl2, CaSO4 (calcium salts), and glucono-δ-lactone (GDL) with the purpose of improving their mechanical and barrier properties. These films were characterized by determining their mechanical properties and permeability to water vapor and oxygen. Formulations with 5% (w/v) protein and 75% sorbitol and 4% (w/v) protein with a mixture of 15% glycerol and 15% sorbitol produced adequate films. Calcium salts and GDL increased the tensile fracture stress but reduced the fracture strain and decreased water vapor permeability compared with control films. The films prepared represent an attractive alternative for being used as food packaging materials.

  18. Preparation of Modified Films with Protein from Grouper Fish.

    PubMed

    Valdivia-López, M A; Tecante, A; Granados-Navarrete, S; Martínez-García, C

    2016-01-01

    A protein concentrate (PC) was obtained from Grouper fish skin and it was used to prepare films with different amounts of sorbitol and glycerol as plasticizers. The best performing films regarding resistance were then modified with various concentrations of CaCl2, CaSO4 (calcium salts), and glucono-δ-lactone (GDL) with the purpose of improving their mechanical and barrier properties. These films were characterized by determining their mechanical properties and permeability to water vapor and oxygen. Formulations with 5% (w/v) protein and 75% sorbitol and 4% (w/v) protein with a mixture of 15% glycerol and 15% sorbitol produced adequate films. Calcium salts and GDL increased the tensile fracture stress but reduced the fracture strain and decreased water vapor permeability compared with control films. The films prepared represent an attractive alternative for being used as food packaging materials.

  19. Preparation of Modified Films with Protein from Grouper Fish.

    PubMed

    Valdivia-López, M A; Tecante, A; Granados-Navarrete, S; Martínez-García, C

    2016-01-01

    A protein concentrate (PC) was obtained from Grouper fish skin and it was used to prepare films with different amounts of sorbitol and glycerol as plasticizers. The best performing films regarding resistance were then modified with various concentrations of CaCl2, CaSO4 (calcium salts), and glucono-δ-lactone (GDL) with the purpose of improving their mechanical and barrier properties. These films were characterized by determining their mechanical properties and permeability to water vapor and oxygen. Formulations with 5% (w/v) protein and 75% sorbitol and 4% (w/v) protein with a mixture of 15% glycerol and 15% sorbitol produced adequate films. Calcium salts and GDL increased the tensile fracture stress but reduced the fracture strain and decreased water vapor permeability compared with control films. The films prepared represent an attractive alternative for being used as food packaging materials. PMID:27597950

  20. An ignored variable: solution preparation temperature in protein crystallization.

    PubMed

    Chen, Rui-Qing; Lu, Qin-Qin; Cheng, Qing-Di; Ao, Liang-Bo; Zhang, Chen-Yan; Hou, Hai; Liu, Yong-Ming; Li, Da-Wei; Yin, Da-Chuan

    2015-01-19

    Protein crystallization is affected by many parameters, among which certain parameters have not been well controlled. The temperature at which the protein and precipitant solutions are mixed (i.e., the ambient temperature during mixing) is such a parameter that is typically not well controlled and is often ignored. In this paper, we show that this temperature can influence protein crystallization. The experimental results showed that both higher and lower mixing temperatures can enhance the success of crystallization, which follows a parabolic curve with an increasing ambient temperature. This work illustrates that the crystallization solution preparation temperature is also an important parameter for protein crystallization. Uncontrolled or poorly controlled room temperature may yield poor reproducibility in protein crystallization.

  1. An Automatic Refolding Apparatus for Preparative-Scale Protein Production

    PubMed Central

    Feng, Yanye; Zhang, Ming; Zhang, Linlin; Zhang, Ting; Ding, Jianfeng; Zhuang, Yingping; Wang, Xiaoning; Yang, Zhong

    2012-01-01

    may provide a powerful tool for preparative scale protein production. PMID:23029296

  2. Partial Molecular Characterization of Arctium minus Aspartylendopeptidase and Preparation of Bioactive Peptides by Whey Protein Hydrolysis.

    PubMed

    Cimino, Cecilia V; Colombo, María Laura; Liggieri, Constanza; Bruno, Mariela; Vairo-Cavalli, Sandra

    2015-08-01

    In this article, we report the cloning of an aspartic protease (AP) from flowers of Arctium minus (Hill) Bernh. (Asteraceae) along with the use of depigmented aqueous flower extracts, as a source of APs, for the hydrolysis of whey proteins. The isolated cDNA encoded a protein product with 509 amino acids called arctiumisin, with the characteristic primary structure organization of typical plant APs. Bovine whey protein hydrolysates, obtained employing the enzyme extracts of A. minus flowers, displayed inhibitory angiotensin-converting enzyme (ACE) and antioxidant activities. Hydrolysates after 3 and 5 h of reaction (degree of hydrolysis 2.4 and 5.6, respectively) and the associated peptide fraction with molecular weight below 3 kDa were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, matrix-assisted laser desorption ionization/time of flight mass spectrometry, and reverse phase-high-performance liquid chromatography. The results obtained in this study demonstrate the viability of using proteases from A. minus to increase the antioxidant and inhibitory ACE capacity of whey proteins.

  3. Partial Molecular Characterization of Arctium minus Aspartylendopeptidase and Preparation of Bioactive Peptides by Whey Protein Hydrolysis.

    PubMed

    Cimino, Cecilia V; Colombo, María Laura; Liggieri, Constanza; Bruno, Mariela; Vairo-Cavalli, Sandra

    2015-08-01

    In this article, we report the cloning of an aspartic protease (AP) from flowers of Arctium minus (Hill) Bernh. (Asteraceae) along with the use of depigmented aqueous flower extracts, as a source of APs, for the hydrolysis of whey proteins. The isolated cDNA encoded a protein product with 509 amino acids called arctiumisin, with the characteristic primary structure organization of typical plant APs. Bovine whey protein hydrolysates, obtained employing the enzyme extracts of A. minus flowers, displayed inhibitory angiotensin-converting enzyme (ACE) and antioxidant activities. Hydrolysates after 3 and 5 h of reaction (degree of hydrolysis 2.4 and 5.6, respectively) and the associated peptide fraction with molecular weight below 3 kDa were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, matrix-assisted laser desorption ionization/time of flight mass spectrometry, and reverse phase-high-performance liquid chromatography. The results obtained in this study demonstrate the viability of using proteases from A. minus to increase the antioxidant and inhibitory ACE capacity of whey proteins. PMID:25575270

  4. Towards generation of bioactive peptides from meat industry waste proteins: Generation of peptides using commercial microbial proteases.

    PubMed

    Ryder, Kate; Bekhit, Alaa El-Din; McConnell, Michelle; Carne, Alan

    2016-10-01

    Five commercially available food-grade microbial protease preparations were evaluated for their ability to hydrolyse meat myofibrillar and connective tissue protein extracts to produce bioactive peptides. A bacterial-derived protease (HT) extensively hydrolysed both meat protein extracts, producing peptide hydrolysates with significant in vitro antioxidant and ACE inhibitor activities. The hydrolysates retained bioactivity after simulated gastrointestinal hydrolysis challenge. Gel permeation chromatography sub-fractionation of the crude protein hydrolysates showed that the smaller peptide fractions exhibited the highest antioxidant and ACE inhibitor activities. OFFGEL electrophoresis of the small peptides of both hydrolysates showed that low isoelectric point peptides had antioxidant activity; however, no consistent relationship was observed between isoelectric point and ACE inhibition. Cell-based assays indicated that the hydrolysates present no significant cytotoxicity towards Vero cells. The results indicate that HT protease hydrolysis of meat myofibrillar and connective tissue protein extracts produces bioactive peptides that are non-cytotoxic, should be stable in the gastrointestinal tract and may contain novel bioactive peptide sequences.

  5. Towards generation of bioactive peptides from meat industry waste proteins: Generation of peptides using commercial microbial proteases.

    PubMed

    Ryder, Kate; Bekhit, Alaa El-Din; McConnell, Michelle; Carne, Alan

    2016-10-01

    Five commercially available food-grade microbial protease preparations were evaluated for their ability to hydrolyse meat myofibrillar and connective tissue protein extracts to produce bioactive peptides. A bacterial-derived protease (HT) extensively hydrolysed both meat protein extracts, producing peptide hydrolysates with significant in vitro antioxidant and ACE inhibitor activities. The hydrolysates retained bioactivity after simulated gastrointestinal hydrolysis challenge. Gel permeation chromatography sub-fractionation of the crude protein hydrolysates showed that the smaller peptide fractions exhibited the highest antioxidant and ACE inhibitor activities. OFFGEL electrophoresis of the small peptides of both hydrolysates showed that low isoelectric point peptides had antioxidant activity; however, no consistent relationship was observed between isoelectric point and ACE inhibition. Cell-based assays indicated that the hydrolysates present no significant cytotoxicity towards Vero cells. The results indicate that HT protease hydrolysis of meat myofibrillar and connective tissue protein extracts produces bioactive peptides that are non-cytotoxic, should be stable in the gastrointestinal tract and may contain novel bioactive peptide sequences. PMID:27132822

  6. Ultrasonic-Assisted Enzymolysis to Improve the Antioxidant Activities of Peanut (Arachin conarachin L.) Antioxidant Hydrolysate

    PubMed Central

    Yu, Lina; Sun, Jie; Liu, Shaofang; Bi, Jie; Zhang, Chushu; Yang, Qingli

    2012-01-01

    The objective of this work is to provide a theoretical basis for preparing peanut antioxidant hydrolysate in order to improve its antioxidant activities. Therefore, response surface methodology (RSM) based on the Box-Behnken design was used to optimize ultrasonic-assisted enzymolysis for the purpose of preparing peanut antioxidant hydrolysate. Results indicated that the DPPH free radical scavenging activity of peanut hydrolysate could reach 90.06% under the following optimum conditions: ultrasonic power of 150.0 w, reaction temperature of 62.0 °C, incubation time of 25.0 min, and initial pH value of 8.5. The DPPH free radical scavenging rate of peanut hydrolysate from ultrasonic-assisted enzymolysis improved comparing with that of peanut hydrolysate from protease hydrolysis alone. The peanut antioxidant hydrolysate was found to display eight improved kinds of antioxidant activities. In conclusion, the optimal ultrasonic-assisted enzymolysis technology conditions described in this paper, appear to be beneficial for preparing peanut antioxidant hydrolysate. PMID:22942751

  7. Temperature-dependent FTIR spectra of collagen and protective effect of partially hydrolysed fucoidan

    NASA Astrophysics Data System (ADS)

    Pielesz, Anna

    2014-01-01

    FTIR spectra of collagen (PC) and partially hydrolysed fucoidan (PHF) incorporated into collagen films were investigated at different temperatures between 20 °C and 100 °C. Changes within the bands of amide I, amide II and amide III may indicate stabilization of collagen by hydrogen bonds during its interaction with partially hydrolysed fucoidan. Spectroscopic studies revealed that partially hydrolysed fucoidan was bound to the collagen without affecting its triple helicity. Interactions of fucoidan with H2SO4 (mild acid hydrolysis), leading to changes of the sulphated band positions in the 800-590 cm-1 region of IR spectra were observed. The effect of partially hydrolysed fucoidan on glucose-mediated collagen glycation and cross-linking of proteins in vitro was evaluated. It was observed that partially hydrolysed fucoidan incorporated into collagen films can be used as therapeutically active biomaterials that speed up the process of wound healing and may increase the anticancer activity of fucoidan.

  8. Preparation of 2D crystals of membrane proteins for high-resolution electron crystallography data collection.

    PubMed

    Abeyrathne, Priyanka D; Chami, Mohamed; Pantelic, Radosav S; Goldie, Kenneth N; Stahlberg, Henning

    2010-01-01

    Electron crystallography is a powerful technique for the structure determination of membrane proteins as well as soluble proteins. Sample preparation for 2D membrane protein crystals is a crucial step, as proteins have to be prepared for electron microscopy at close to native conditions. In this review, we discuss the factors of sample preparation that are key to elucidating the atomic structure of membrane proteins using electron crystallography.

  9. Approaches to High-Performance Preparative Chromatography of Proteins

    NASA Astrophysics Data System (ADS)

    Sun, Yan; Liu, Fu-Feng; Shi, Qing-Hong

    Preparative liquid chromatography is widely used for the purification of chemical and biological substances. Different from high-performance liquid chromatography for the analysis of many different components at minimized sample loading, high-performance preparative chromatography is of much larger scale and should be of high resolution and high capacity at high operation speed and low to moderate pressure drop. There are various approaches to this end. For biochemical engineers, the traditional way is to model and optimize a purification process to make it exert its maximum capability. For high-performance separations, however, we need to improve chromatographic technology itself. We herein discuss four approaches in this review, mainly based on the recent studies in our group. The first is the development of high-performance matrices, because packing material is the central component of chromatography. Progress in the fabrication of superporous materials in both beaded and monolithic forms are reviewed. The second topic is the discovery and design of affinity ligands for proteins. In most chromatographic methods, proteins are separated based on their interactions with the ligands attached to the surface of porous media. A target-specific ligand can offer selective purification of desired proteins. Third, electrochromatography is discussed. An electric field applied to a chromatographic column can induce additional separation mechanisms besides chromatography, and result in electrokinetic transport of protein molecules and/or the fluid inside pores, thus leading to high-performance separations. Finally, expanded-bed adsorption is described for process integration to reduce separation steps and process time.

  10. Preparation and functional evaluation of antihypertensive polypeptides from rice based on ultrasonic pretreatment

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Enzymolysis was used for preparation of antihypertensive peptide from rice. Following studies were conducted:ultrasonic pretreatment of substrate protein, ultrafilter of hydrolysate and test of anti-digestive enzyme degradation and one-time feeding of spontaneously hypertensive rats (SHR) of antihyp...

  11. The Heart Protection Effect of Alcalase Potato Protein Hydrolysate Is through IGF1R-PI3K-Akt Compensatory Reactivation in Aging Rats on High Fat Diets

    PubMed Central

    Hu, Wei-Syun; Ting, Wei-Jen; Chiang, Wen-Dee; Pai, Peiying; Yeh, Yu-Lan; Chang, Chung-Ho; Lin, Wan-Teng; Huang, Chih-Yang

    2015-01-01

    The prevalence of obesity is high in older adults. Alcalase potato protein hydrolysate (APPH), a nutraceutical food, might have greater benefits and be more economical than hypolipidemic drugs. In this study, serum lipid profiles and heart protective effects were evaluated in high fat diet (HFD) induced hyperlipidemia in aging rats treated with APPH (15, 45 and 75 mg/kg/day) and probucol (500 mg/kg/day). APPH treatments reduced serum triacylglycerol (TG), total cholesterol (TC), and low density lipoprotein (LDL) levels to the normal levels expressed in the control group. Additionally, the IGF1R-PI3K-Akt survival pathway was reactivated, and Fas-FADD (Fas-associated death domain) induced apoptosis was inhibited by APPH treatments (15 and 45 mg/kg/day) in HFD aging rat hearts. APPH (75 mg/kg/day) rather than probucol (500 mg/kg/day) treatment could reduce serum lipids without affecting HDL expression. The heart protective effect of APPH in aging rats with hyperlipidemia was through lowering serum lipids and enhancing the activation of the compensatory IGF1R-PI3K-Akt survival pathway. PMID:25950762

  12. Nine novel angiotensin I-converting enzyme (ACE) inhibitory peptides from cuttlefish (Sepia officinalis) muscle protein hydrolysates and antihypertensive effect of the potent active peptide in spontaneously hypertensive rats.

    PubMed

    Balti, Rafik; Bougatef, Ali; Sila, Assaâd; Guillochon, Didier; Dhulster, Pascal; Nedjar-Arroume, Naima

    2015-03-01

    This study aimed to identify novel ACE inhibitory peptides from the muscle of cuttlefish. Proteins were hydrolyzed and the hydrolysates were then subjected to various types of chromatography to isolate the active peptides. Nine ACE inhibitory peptides were isolated and their molecular masses and amino acid sequences were determined using ESI-MS and ESI-MS/MS, respectively. The structures of the most potent peptides were identified as Val-Glu-Leu-Tyr-Pro, Ala-Phe-Val-Gly-Tyr-Val-Leu-Pro and Glu-Lys-Ser-Tyr-Glu-Leu-Pro. The first peptide displayed the highest ACE inhibitory activity with an IC50 of 5.22μM. Lineweaver-Burk plots suggest that Val-Glu-Leu-Tyr-Pro acts as a non-competitive inhibitor against ACE. Furthermore, antihypertensive effects in spontaneously hypertensive rats (SHR) also revealed that oral administration of Val-Glu-Leu-Tyr-Pro can decrease systolic blood pressure significantly (p<0.01). These results suggest that the Val-Glu-Leu-Tyr-Pro would be a beneficial ingredient for nutraceuticals and pharmaceuticals acting against hypertension and its related diseases.

  13. The Heart Protection Effect of Alcalase Potato Protein Hydrolysate Is through IGF1R-PI3K-Akt Compensatory Reactivation in Aging Rats on High Fat Diets.

    PubMed

    Hu, Wei-Syun; Ting, Wei-Jen; Chiang, Wen-Dee; Pai, Peiying; Yeh, Yu-Lan; Chang, Chung-Ho; Lin, Wan-Teng; Huang, Chih-Yang

    2015-01-01

    The prevalence of obesity is high in older adults. Alcalase potato protein hydrolysate (APPH), a nutraceutical food, might have greater benefits and be more economical than hypolipidemic drugs. In this study, serum lipid profiles and heart protective effects were evaluated in high fat diet (HFD) induced hyperlipidemia in aging rats treated with APPH (15, 45 and 75 mg/kg/day) and probucol (500 mg/kg/day). APPH treatments reduced serum triacylglycerol (TG), total cholesterol (TC), and low density lipoprotein (LDL) levels to the normal levels expressed in the control group. Additionally, the IGF1R-PI3K-Akt survival pathway was reactivated, and Fas-FADD (Fas-associated death domain) induced apoptosis was inhibited by APPH treatments (15 and 45 mg/kg/day) in HFD aging rat hearts. APPH (75 mg/kg/day) rather than probucol (500 mg/kg/day) treatment could reduce serum lipids without affecting HDL expression. The heart protective effect of APPH in aging rats with hyperlipidemia was through lowering serum lipids and enhancing the activation of the compensatory IGF1R-PI3K-Akt survival pathway. PMID:25950762

  14. Nine novel angiotensin I-converting enzyme (ACE) inhibitory peptides from cuttlefish (Sepia officinalis) muscle protein hydrolysates and antihypertensive effect of the potent active peptide in spontaneously hypertensive rats.

    PubMed

    Balti, Rafik; Bougatef, Ali; Sila, Assaâd; Guillochon, Didier; Dhulster, Pascal; Nedjar-Arroume, Naima

    2015-03-01

    This study aimed to identify novel ACE inhibitory peptides from the muscle of cuttlefish. Proteins were hydrolyzed and the hydrolysates were then subjected to various types of chromatography to isolate the active peptides. Nine ACE inhibitory peptides were isolated and their molecular masses and amino acid sequences were determined using ESI-MS and ESI-MS/MS, respectively. The structures of the most potent peptides were identified as Val-Glu-Leu-Tyr-Pro, Ala-Phe-Val-Gly-Tyr-Val-Leu-Pro and Glu-Lys-Ser-Tyr-Glu-Leu-Pro. The first peptide displayed the highest ACE inhibitory activity with an IC50 of 5.22μM. Lineweaver-Burk plots suggest that Val-Glu-Leu-Tyr-Pro acts as a non-competitive inhibitor against ACE. Furthermore, antihypertensive effects in spontaneously hypertensive rats (SHR) also revealed that oral administration of Val-Glu-Leu-Tyr-Pro can decrease systolic blood pressure significantly (p<0.01). These results suggest that the Val-Glu-Leu-Tyr-Pro would be a beneficial ingredient for nutraceuticals and pharmaceuticals acting against hypertension and its related diseases. PMID:25306378

  15. Characterization of the sporulation-related gamma-D-glutamyl-(L)meso-diaminopimelic-acid-hydrolysing peptidase I of Bacillus sphaericus NCTC 9602 as a member of the metallo(zinc) carboxypeptidase A family. Modular design of the protein.

    PubMed Central

    Hourdou, M L; Guinand, M; Vacheron, M J; Michel, G; Denoroy, L; Duez, C; Englebert, S; Joris, B; Weber, G; Ghuysen, J M

    1993-01-01

    The sporulation-related gamma-D-glutamyl-(L)meso-diaminopimelic-acid-hydrolysing peptidase I of Bacillus sphaericus NCTC 9602 has been analysed by proton-induced X-ray emission. It contains 1 equivalent Zn2+ per mol of protein. As derived from gene cloning and sequencing, the B. sphaericus Zn peptidase I is a two-module protein. A 100-amino-acid-residue N-terminal domain consisting of two tandem segments of similar sequences, is fused to a 296-amino-acid-residue C-terminal catalytic domain. The catalytic domain belongs to the Zn carboxypeptidase A family, the closest match being observed with the Streptomyces griseus carboxypeptidase [Narahashi (1990) J. Biochem. 107, 879-886] and with the family prototype, bovine carboxypeptidase A. The catalytic domain of the B. sphaericus peptidase I possesses, distributed along the amino-acid sequence, peptide segments, a triad His162-Glu165-His307 and a dyad Tyr347-Glu366 that are equivalent to secondary structures, the zinc-binding triad His69-Glu72-His196 and the catalytic dyad Tyr248-Glu270 of bovine carboxypeptidase A respectively. The N-terminal repeats of the B. sphaericus peptidase I have similarity with the C-terminal repeats of the Enterococcus hirae muramidase 2, the Streptococcus (now Enterococcus) faecalis autolysin and the Bacillus phi PZA and phi 29 lysozymes, to which a role in the recognition of a particular moiety of the bacterial cell envelope has been tentatively assigned. Detergents enhance considerably the specific activity of the B. sphaericus peptidase I. Images Figure 1 PMID:8503890

  16. Development toward rapid and efficient screening for high performance hydrolysate lots in a recombinant monoclonal antibody manufacturing process.

    PubMed

    Luo, Ying; Pierce, Karisa M

    2012-07-01

    Plant-derived hydrolysates are widely used in mammalian cell culture media to increase yields of recombinant proteins and monoclonal antibodies (mAbs). However, these chemically varied and undefined raw materials can have negative impact on yield and/or product quality in large-scale cell culture processes. Traditional methods that rely on fractionation of hydrolysates yielded little success in improving hydrolysate quality. We took a holistic approach to develop an efficient and reliable method to screen intact soy hydrolysate lots for commercial recombinant mAb manufacturing. Combined high-resolution (1) H nuclear magnetic resonance (NMR) spectroscopy and partial least squares (PLS) analysis led to a prediction model between product titer and NMR fingerprinting of soy hydrolysate with cross-validated correlation coefficient R(2) of 0.87 and root-mean-squared-error of cross-validation RMSECV% of 11.2%. This approach screens for high performance hydrolysate lots, therefore ensuring process consistency and product quality in the mAb manufacturing process. Furthermore, PLS analysis was successful in discerning multiple markers (DL-lactate, soy saccharides, citrate and succinate) among hydrolysate components that positively and negatively correlate with titer. Interestingly, these markers correlate to the metabolic characteristics of some strains of taxonomically diverse lactic acid bacteria (LAB). Thus our findings indicate that LAB strains may exist during hydrolysate manufacturing steps and their biochemical activities may attribute to the titer enhancement effect of soy hydrolysates. PMID:22641483

  17. Protein-peptide interaction: study of heat-induced aggregation and gelation of β-lactoglobulin in the presence of two peptides from its own hydrolysate.

    PubMed

    Kosters, Hans A; Wierenga, Peter A; de Vries, Renko; Gruppen, Harry

    2013-05-01

    Two peptides, [f135-158] and [f135-162]-SH, were used to study the binding of the peptides to native β-lactolobulin, as well as the subsequent effects on aggregation and gelation of β-lactoglobulin. The binding of the peptide [f135-158] to β-lactoglobulin at room temperature was confirmed by SELDI-TOF-MS. It was further illustrated by increased turbidity of mixed solutions of peptide and protein (at pH 7), indicating association of proteins and peptides in larger complexes. At pH below the isoelectric point of the protein, the presence of peptides did not lead to an increased turbidity, showing the absence of complexation. The protein-peptide complexes formed at pH 7 were found to dissociate directly upon heating. After prolonged heating, extensive aggregation was observed, whereas no aggregation was seen for the pure protein or pure peptide solutions. The presence of the free sulfhydryl group in [f135-162]-SH resulted in a 10 times increase in the amount of aggregation of β-lactoglobulin upon heating, illustrating the additional effect of the free sulfhydryl group. Subsequent studies on the gel strength of heat-induced gels also showed a clear difference between these two peptides. The replacement of additional β-lactoglobulin by [f135-158] resulted in a decrease in gel strength, whereas replacement by peptide [f135-162]-SH increased gel strength.

  18. Radiation hydrolysate of tuna cooking juice with enhanced antioxidant properties

    NASA Astrophysics Data System (ADS)

    Choi, Jong-il; Sung, Nak-Yun; Lee, Ju-Woon

    2012-08-01

    Tuna protein hydrolysates are of increasing interest because of their potential application as a source of bioactive peptides. Large amounts of tuna cooking juice with proteins and extracts are produced during the process of tuna canning, and these cooking juice wastes cause environmental problems. Therefore, in this study, cooking juice proteins were hydrolyzed by irradiation for their utilization as functional additives. The degree of hydrolysis of tuna cooking juice protein increased from 0% to 15.1% at the absorbed doses of 50 kGy. To investigate the antioxidant activity of the hydrolysate, it was performed the ferric reducing antioxidant power (FRAP) assay, and the lipid peroxidation inhibitory and superoxide radical scavenging activities were measured. The FRAP values increased from 1470 μM to 1930 μM and IC50 on superoxide anion was decreased from 3.91 μg/mL to 1.29 μg/mL at 50 kGy. All of the antioxidant activities were increased in the hydrolysate, suggesting that radiation hydrolysis, which is a simple process that does not require an additive catalysts or an inactivation step, is a promising method for food and environmental industries.

  19. SynProt: A Database for Proteins of Detergent-Resistant Synaptic Protein Preparations

    PubMed Central

    Pielot, Rainer; Smalla, Karl-Heinz; Müller, Anke; Landgraf, Peter; Lehmann, Anne-Christin; Eisenschmidt, Elke; Haus, Utz-Uwe; Weismantel, Robert; Gundelfinger, Eckart D.; Dieterich, Daniela C.

    2012-01-01

    Chemical synapses are highly specialized cell–cell contacts for communication between neurons in the CNS characterized by complex and dynamic protein networks at both synaptic membranes. The cytomatrix at the active zone (CAZ) organizes the apparatus for the regulated release of transmitters from the presynapse. At the postsynaptic side, the postsynaptic density constitutes the machinery for detection, integration, and transduction of the transmitter signal. Both pre- and postsynaptic protein networks represent the molecular substrates for synaptic plasticity. Their function can be altered both by regulating their composition and by post-translational modification of their components. For a comprehensive understanding of synaptic networks the entire ensemble of synaptic proteins has to be considered. To support this, we established a comprehensive database for synaptic junction proteins (SynProt database) primarily based on proteomics data obtained from biochemical preparations of detergent-resistant synaptic junctions. The database currently contains 2,788 non-redundant entries of rat, mouse, and some human proteins, which mainly have been manually extracted from 12 proteomic studies and annotated for synaptic subcellular localization. Each dataset is completed with manually added information including protein classifiers as well as automatically retrieved and updated information from public databases (UniProt and PubMed). We intend that the database will be used to support modeling of synaptic protein networks and rational experimental design. PMID:22737123

  20. Whey or Casein Hydrolysate with Carbohydrate for Metabolism and Performance in Cycling.

    PubMed

    Oosthuyse, T; Carstens, M; Millen, A M E

    2015-07-01

    The protein type most suitable for ingestion during endurance exercise is undefined. This study compared co-ingestion of either 15 g/h whey or casein hydrolysate with 63 g/h fructose: maltodextrin (0.8:1) on exogenous carbohydrate oxidation, exercise metabolism and performance. 2 h postprandial, 8 male cyclists ingested either: carbohydrate-only, carbohydrate-whey hydrolysate, carbohydrate-casein hydrolysate or placebo-water in a crossover, double-blind design during 2 h of exercise at 60%W max followed by a 16-km time trial. Data were evaluated by magnitude-based inferential statistics. Exogenous carbohydrate oxidation, measured from (13)CO2 breath enrichment, was not substantially influenced by co-ingestion of either protein hydrolysate. However, only co-ingestion of carbohydrate-casein hydrolysate substantially decreased (98% very likely decrease) total carbohydrate oxidation (mean±SD, 242±44; 258±47; 277±33 g for carbohydrate-casein, carbohydrate-whey and carbohydrate-only, respectively) and substantially increased (93% likely increase) total fat oxidation (92±14; 83±27; 73±19 g) compared with carbohydrate-only. Furthermore, only carbohydrate-casein hydrolysate ingestion resulted in a faster time trial (-3.6%; 90% CI: ±3.2%) compared with placebo-water (95% likely benefit). However, neither protein hydrolysate enhanced time trial performance when compared with carbohydrate-only. Under the conditions of this study, ingesting carbohydrate-casein, but not carbohydrate-whey hydrolysate, favourably alters metabolism during prolonged moderate-strenuous cycling without substantially altering cycling performance compared with carbohydrate-only.

  1. A defence-related Olea europaea β-glucosidase hydrolyses and activates oleuropein into a potent protein cross-linking agent

    PubMed Central

    Koudounas, Konstantinos; Banilas, Georgios; Michaelidis, Christos; Demoliou, Catherine; Rigas, Stamatis; Hatzopoulos, Polydefkis

    2015-01-01

    Oleuropein, the major secoiridoid compound in olive, is involved in a sophisticated two-component defence system comprising a β-glucosidase enzyme that activates oleuropein into a toxic glutaraldehyde-like structure. Although oleuropein deglycosylation studies have been monitored extensively, an oleuropein β-glucosidase gene has not been characterized as yet. Here, we report the isolation of OeGLU cDNA from olive encoding a β-glucosidase belonging to the defence-related group of terpenoid-specific glucosidases. In planta recombinant protein expression assays showed that OeGLU deglycosylated and activated oleuropein into a strong protein cross-linker. Homology and docking modelling predicted that OeGLU has a characteristic (β/α)8 TIM barrel conformation and a typical construction of a pocket-shaped substrate recognition domain composed of conserved amino acids supporting the β-glucosidase activity and non-conserved residues associated with aglycon specificity. Transcriptional analysis in various olive organs revealed that the gene was developmentally regulated, with its transcript levels coinciding well with the spatiotemporal patterns of oleuropein degradation and aglycon accumulation in drupes. OeGLU upregulation in young organs reflects its prominent role in oleuropein-mediated defence system. High gene expression during drupe maturation implies an additional role in olive secondary metabolism, through the degradation of oleuropein and reutilization of hydrolysis products. PMID:25697790

  2. A defence-related Olea europaea β-glucosidase hydrolyses and activates oleuropein into a potent protein cross-linking agent.

    PubMed

    Koudounas, Konstantinos; Banilas, Georgios; Michaelidis, Christos; Demoliou, Catherine; Rigas, Stamatis; Hatzopoulos, Polydefkis

    2015-04-01

    Oleuropein, the major secoiridoid compound in olive, is involved in a sophisticated two-component defence system comprising a β-glucosidase enzyme that activates oleuropein into a toxic glutaraldehyde-like structure. Although oleuropein deglycosylation studies have been monitored extensively, an oleuropein β-glucosidase gene has not been characterized as yet. Here, we report the isolation of OeGLU cDNA from olive encoding a β-glucosidase belonging to the defence-related group of terpenoid-specific glucosidases. In planta recombinant protein expression assays showed that OeGLU deglycosylated and activated oleuropein into a strong protein cross-linker. Homology and docking modelling predicted that OeGLU has a characteristic (β/α)8 TIM barrel conformation and a typical construction of a pocket-shaped substrate recognition domain composed of conserved amino acids supporting the β-glucosidase activity and non-conserved residues associated with aglycon specificity. Transcriptional analysis in various olive organs revealed that the gene was developmentally regulated, with its transcript levels coinciding well with the spatiotemporal patterns of oleuropein degradation and aglycon accumulation in drupes. OeGLU upregulation in young organs reflects its prominent role in oleuropein-mediated defence system. High gene expression during drupe maturation implies an additional role in olive secondary metabolism, through the degradation of oleuropein and reutilization of hydrolysis products.

  3. Alteration of Gene Expression Profile in Kidney of Spontaneously Hypertensive Rats Treated with Protein Hydrolysate of Blue Mussel (Mytilus edulis) by DNA Microarray Analysis

    PubMed Central

    Feng, Junli; Dai, Zhiyuan; Zhang, Yanping; Meng, Lu; Ye, Jian; Ma, Xuting

    2015-01-01

    Marine organisms are rich sources of bioactive components, which are often reported to have antihypertensive effects. However, the underlying mechanisms have yet to be fully identified. The aim of this study was to investigate the antihypertensive effect of enzymatic hydrolysis of blue mussel protein (HBMP) in rats. Peptides with in vitro ACE inhibitory activity were purified from HBMP by ultrafiltration, gel filtration chromatography and reversed-phase high performance liquid chromatography. And the amino acid sequences of isolated peptides were estimated to be Val-Trp, Leu-Gly-Trp, and Met-Val-Trp-Thr. To study its in vivo action, spontaneously hypertensive rats (SHRs) were orally administration with high- or low-dose of HBMP for 28 days. Major components of the renin-angiotensin (RAS) system in serum of SHRs from different groups were analyzed, and gene expression profiling were performed in the kidney of SHRs, using the Whole Rat Genome Oligonucleotide Microarray. Results indicated although genes involved in RAS system were not significantly altered, those related to blood coagulation system, cytokine and growth factor, and fatty acids metabolism were remarkablely changed. Several genes which were seldom reported to be implicated in pathogenesis of hypertension also showed significant expression alterations after oral administration of HBMP. These data provided valuable information for our understanding of the molecular mechanisms that underlie the potential antihypertensive activities of HBMP, and will contribute towards increased value-added utilization of blue mussel protein. PMID:26517713

  4. Preparation of Extracellular Matrix Protein Fibers for Brillouin Spectroscopy.

    PubMed

    Edginton, Ryan S; Mattana, Sara; Caponi, Silvia; Fioretto, Daniele; Green, Ellen; Winlove, C Peter; Palombo, Francesca

    2016-01-01

    Brillouin spectroscopy is an emerging technique in the biomedical field. It probes the mechanical properties of a sample through the interaction of visible light with thermally induced acoustic waves or phonons propagating at a speed of a few km/sec. Information on the elasticity and structure of the material is obtained in a nondestructive contactless manner, hence opening the way to in vivo applications and potential diagnosis of pathology. This work describes the application of Brillouin spectroscopy to the study of biomechanics in elastin and trypsin-digested type I collagen fibers of the extracellular matrix. Fibrous proteins of the extracellular matrix are the building blocks of biological tissues and investigating their mechanical and physical behavior is key to establishing structure-function relationships in normal tissues and the changes which occur in disease. The procedures of sample preparation followed by measurement of Brillouin spectra using a reflective substrate are presented together with details of the optical system and methods of spectral data analysis. PMID:27684584

  5. Identifying inhibitory compounds in lignocellulosic biomass hydrolysates using an exometabolomics approach

    PubMed Central

    2014-01-01

    Background Inhibitors are formed that reduce the fermentation performance of fermenting yeast during the pretreatment process of lignocellulosic biomass. An exometabolomics approach was applied to systematically identify inhibitors in lignocellulosic biomass hydrolysates. Results We studied the composition and fermentability of 24 different biomass hydrolysates. To create diversity, the 24 hydrolysates were prepared from six different biomass types, namely sugar cane bagasse, corn stover, wheat straw, barley straw, willow wood chips and oak sawdust, and with four different pretreatment methods, i.e. dilute acid, mild alkaline, alkaline/peracetic acid and concentrated acid. Their composition and that of fermentation samples generated with these hydrolysates were analyzed with two GC-MS methods. Either ethyl acetate extraction or ethyl chloroformate derivatization was used before conducting GC-MS to prevent sugars are overloaded in the chromatograms, which obscure the detection of less abundant compounds. Using multivariate PLS-2CV and nPLS-2CV data analysis models, potential inhibitors were identified through establishing relationship between fermentability and composition of the hydrolysates. These identified compounds were tested for their effects on the growth of the model yeast, Saccharomyces. cerevisiae CEN.PK 113-7D, confirming that the majority of the identified compounds were indeed inhibitors. Conclusion Inhibitory compounds in lignocellulosic biomass hydrolysates were successfully identified using a non-targeted systematic approach: metabolomics. The identified inhibitors include both known ones, such as furfural, HMF and vanillin, and novel inhibitors, namely sorbic acid and phenylacetaldehyde. PMID:24655423

  6. Gelatin hydrolysates from farmed Giant catfish skin using alkaline proteases and its antioxidative function of simulated gastro-intestinal digestion.

    PubMed

    Ketnawa, Sunantha; Martínez-Alvarez, Oscar; Benjakul, Soottawat; Rawdkuen, Saroat

    2016-02-01

    This work aims to evaluate the ability of different alkaline proteases to prepare active gelatin hydrolysates. Fish skin gelatin was hydrolysed by visceral alkaline-proteases from Giant catfish, commercial trypsin, and Izyme AL®. All antioxidant activity indices of the hydrolysates increased with increasing degree of hydrolysis (P<0.05). The hydrolysates obtained with Izyme AL® and visceral alkaline-proteases showed the highest and lowest radical scavenging capacity, while prepared with commercial trypsin was the most effective in reducing ferric ions and showed the best metal chelating properties. The hydrolysate obtained with Izyme AL® showed the lowest iron reducing ability, but provided the highest average molecular weight (⩾ 7 kDa), followed by commercial trypsin (2.2 kDa) and visceral alkaline-proteases (1.75 kDa). After in vitro gastrointestinal digestion, the hydrolysates showed significant higher radical scavenging, reducing ferric ions and chelating activities. Gelatin hydrolysates, from fish skin, could serve as a potential source of functional food ingredients for health promotion.

  7. Gelatin hydrolysates from farmed Giant catfish skin using alkaline proteases and its antioxidative function of simulated gastro-intestinal digestion.

    PubMed

    Ketnawa, Sunantha; Martínez-Alvarez, Oscar; Benjakul, Soottawat; Rawdkuen, Saroat

    2016-02-01

    This work aims to evaluate the ability of different alkaline proteases to prepare active gelatin hydrolysates. Fish skin gelatin was hydrolysed by visceral alkaline-proteases from Giant catfish, commercial trypsin, and Izyme AL®. All antioxidant activity indices of the hydrolysates increased with increasing degree of hydrolysis (P<0.05). The hydrolysates obtained with Izyme AL® and visceral alkaline-proteases showed the highest and lowest radical scavenging capacity, while prepared with commercial trypsin was the most effective in reducing ferric ions and showed the best metal chelating properties. The hydrolysate obtained with Izyme AL® showed the lowest iron reducing ability, but provided the highest average molecular weight (⩾ 7 kDa), followed by commercial trypsin (2.2 kDa) and visceral alkaline-proteases (1.75 kDa). After in vitro gastrointestinal digestion, the hydrolysates showed significant higher radical scavenging, reducing ferric ions and chelating activities. Gelatin hydrolysates, from fish skin, could serve as a potential source of functional food ingredients for health promotion. PMID:26304317

  8. [The complexes of copper, manganese and chromium with enzymatic hydrolysate of pig spleen: research in vitro].

    PubMed

    Zorin, S N; Sidorova, Yu S; Pleten, A P; Mazo, V K

    2016-01-01

    This report describes the preparation and the results of physical and chemical analysis of complexes of enzymatic hydrolysate of pig spleen (EHPS) with manganese, copper and chromium. The complexes were prepared using schemes including the reaction of complexation of inorganic cations with EHPS-peptides structures and application of membrane technology. The process of microfiltration of the resulting mixtures was carried out in tangential flow and low molecular weight fractions were collected. Solutions of copper and manganese complexes with EHPS were subjected to nanofiltration to remove inorganic ions from the reaction mixture. The obtained preparations were lyophilic dried and the molecular weight distribution of the protein fractions in Cu-EHPS, Mn-EHPS and Cr-EHPS complexes was analyzed by exclusion medium pressure liquid chromatography. The percentage relation of fractions with specific molecular weight range was calculated by applying the weighted integration of chromatograms. The determination of copper, manganese and chromium levels in the complexes was performed by atomic absorption method. The content of microelements in the preparations is for copper 16.5 ± 0.3 mg/g, for manganese--24.9 ± 0.5 mg/g and for chromium--2.5 ± 0.2 mg/g.

  9. [The complexes of copper, manganese and chromium with enzymatic hydrolysate of pig spleen: research in vitro].

    PubMed

    Zorin, S N; Sidorova, Yu S; Pleten, A P; Mazo, V K

    2016-01-01

    This report describes the preparation and the results of physical and chemical analysis of complexes of enzymatic hydrolysate of pig spleen (EHPS) with manganese, copper and chromium. The complexes were prepared using schemes including the reaction of complexation of inorganic cations with EHPS-peptides structures and application of membrane technology. The process of microfiltration of the resulting mixtures was carried out in tangential flow and low molecular weight fractions were collected. Solutions of copper and manganese complexes with EHPS were subjected to nanofiltration to remove inorganic ions from the reaction mixture. The obtained preparations were lyophilic dried and the molecular weight distribution of the protein fractions in Cu-EHPS, Mn-EHPS and Cr-EHPS complexes was analyzed by exclusion medium pressure liquid chromatography. The percentage relation of fractions with specific molecular weight range was calculated by applying the weighted integration of chromatograms. The determination of copper, manganese and chromium levels in the complexes was performed by atomic absorption method. The content of microelements in the preparations is for copper 16.5 ± 0.3 mg/g, for manganese--24.9 ± 0.5 mg/g and for chromium--2.5 ± 0.2 mg/g. PMID:27228705

  10. Comparison of Antioxidant Activities of Hydrolysates of Domestic and Imported Skim Milk Powders Treated with Papain

    PubMed Central

    Ha, Go Eun; Chang, Oun Ki; Han, Gi Sung; Ham, Jun Sang; Park, Beom-Young; Jeong, Seok-Geun

    2015-01-01

    Milk proteins have many potential sequences within their primary structure, each with a specific biological activity. In this study, we compared and investigated the bioactivities of hydrolysates of the domestic (A, B) and imported (C, D) skim milk powders generated using papain digestion. MALDI-TOF analysis revealed that all milk powder proteins were intact, indicating no autolysis. Electrophoretic analysis of hydrolysates showed papain treatment caused degradation of milk proteins into peptides of various size. The antioxidant activity of the hydrolysates, determined using 2,2-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) and total phenolic contents (TPC) assays, increased with incubation times. In all skim milk powders, the antioxidant activities of hydrolysates were highest following 24 h papain treatment (TPC: A, 196.48 μM GE/L; B, 194.52 μM GE/L; C, 194.76 μM GE/L; D, 163.75 μM GE/L; ABTS: A, 75%; B, 72%; C, 72%; D, 57%). The number of peptide derived from skim milk powders, as determined by LC-MS/MS, was 308 for A, 283 for B, 208 for C, and 135 for D. Hydrolysate A had the highest antioxidant activity and the most potential antioxidant peptides amongst the four skim milk powder hydrolysates. A total of 4 β-lactoglobulin, 4 αs1-casein, and 56 β-casein peptide fragments were identified as potential antioxidant peptides in hydrolysate A by LC-MS/MS. These results suggest that domestic skim milk could have applications in various industries, i.e., in the development of functional foods. PMID:26761850

  11. Effects of a non-starch polysaccharidase enzyme preparation from Thermomyces lanuginosus on energy and protein metabolism and milk yield of dairy cattle.

    PubMed

    Jurkovich, V; Brydl, E; Rafai, P; Könyves, L; Tegzes, L; Kutasi, J; Bata, A; Nagy, G; Bartyik, J; Fülöp, A

    2002-01-01

    Non-starch polysaccharides (NSPs) form an integral part of the cell walls in plants and represent considerable available energy when degraded into absorbable mono-, di-, tri- and oligosaccharides. The ruminal microflora hydrolyses a good part of NSPs, however, recently there have been attempts to enhance the rate of utilisation by using external polysaccharidase enzymes. In the present study the effects of an enzyme preparation (Rumino-Zyme) high in xylanase activity were studied on ruminal volatile fatty acid (VFA) concentration, parameters of energy and protein metabolism, milk yield, feed conversion ratio (FCR) and body condition score of high-yielding dairy cows. A lignolytic enzyme preparation produced by the thermophilic fungus Thermomyces lanuginosus was applied in the present experiment and fed to dairy cows at 34 g/day dosage in the period between calving and the 110th day of lactation. This preparation increased VFA concentration in the rumen from about 32 days after calving and onward. Increased VFA concentration was followed by an about 5 to 10% increase in milk production and an almost 0.1% increase in butterfat production. Increased VFA concentration produced more balanced energy metabolism in the experimental cows as indicated by the lower incidence rate of hyperketonaemia, and lower acetoacetic acid and non-esterified fatty acid (NEFA) concentration in the blood of the experimental cows. Aspartate aminotransferase (AST) activity was tendentiously higher in the control group and the proportion of cows that had AST activity higher than 100 U/l was also higher in the control group. Both control and experimental cows showed balanced protein and acid-base metabolism throughout the experiment. Enhanced VFA concentration contributed to an improvement in energy balance in the experimental cows with a resultant improvement of feed intake and feed utilisation. Due to the more balanced energy metabolism postparturient body condition loss of the treated cows

  12. A novel preparation of milk protein/polyethylene terephthalate fabric

    NASA Astrophysics Data System (ADS)

    Zhou, J. F.; Zheng, D. D.; Zhong, L.; Zhang, F. X.; Zhang, G. X.

    2016-07-01

    In this work, -NH2 groups were introduced to polyethylene terephthalate (PET) fibers by nitration and reduction method, and then milk protein was grafted on the nitrated and reduced PET (NR PET) fibers by sucrose glycidyl ether crosslinking agent. FTIR suggested the milk protein was successfully grafted on PET fiber surface. SEM images showed a layer of substance covered on the PET fiber surface. DSC demonstrated an excellent thermal stability of milk protein/PET fiber. The moisture regain was improved by milk protein/PET fiber. Moreover, the crease recovery angle and stiffness were retained by the milk protein/PET fabric.

  13. Protein profile of mature soybean seeds and prepared soybean milk.

    PubMed

    Capriotti, Anna Laura; Caruso, Giuseppe; Cavaliere, Chiara; Samperi, Roberto; Stampachiacchiere, Serena; Zenezini Chiozzi, Riccardo; Laganà, Aldo

    2014-10-01

    The soybean (Glycine max (L.) Merrill) is economically the most important bean in the world, providing a wide range of vegetable proteins. Soybean milk is a colloidal solution obtained as water extract from swelled and ground soybean seeds. Soybean proteins represent about 35-40% on a dry weight basis and they are receiving increasing attention with respect to their health effects. However, the soybean is a well-recognized allergenic food, and therefore, it is urgent to define its protein components responsible for the allergenicity in order to develop hypoallergenic soybean products for sensitive people. The main aim of this work was the characterization of seed and milk soybean proteome and their comparison in terms of protein content and specific proteins. Using a shotgun proteomics approach, 243 nonredundant proteins were identified in mature soybean seeds.

  14. Biofunctional Properties of Enzymatic Squid Meat Hydrolysate

    PubMed Central

    Choi, Joon Hyuk; Kim, Kyung-Tae; Kim, Sang Moo

    2015-01-01

    Squid is one of the most important commercial fishes in the world and is mainly utilized or consumed as sliced raw fish or as processed products. The biofunctional activities of enzymatic squid meat hydrolysate were determined to develop value-added products. Enzymatic squid hydrolysate manufactured by Alcalase effectively quenched 1,1-diphenyl-2-picrylhydrazyl radical, hydroxyl radical, and hydrogen peroxide radical with IC50 values of 311, 3,410, and 111.5 μg/mL, respectively. Angiotensin I-converting enzyme inhibitory activity of squid hydrolysate was strong with an IC50 value of 145.1 μg/mL, while tyrosinase inhibitory activity with an IC50 value of 4.72 mg/mL was moderately low. Overall, squid meat hydrolysate can be used in food or cosmetic industries as a bioactive ingredient and possibly be used in the manufacture of seasoning, bread, noodle, or cosmetics. PMID:25866752

  15. Membrane Extraction for Detoxification of Biomass Hydrolysates

    SciTech Connect

    Grzenia, D. L.; Schell, D. J.; Wickramasinghe, S. R.

    2012-05-01

    Membrane extraction was used for the removal of sulfuric acid, acetic acid, 5-hydroxymethyl furfural and furfural from corn stover hydrolyzed with dilute sulfuric acid. Microporous polypropylene hollow fiber membranes were used. The organic extractant consisted of 15% Alamine 336 in: octanol, a 50:50 mixture of oleyl alcohol:octanol or oleyl alcohol. Rapid removal of sulfuric acid, 5-hydroxymethyl and furfural was observed. The rate of acetic acid removal decreased as the pH of the hydrolysate increased. Regeneration of the organic extractant was achieved by back extraction into an aqueous phase containing NaOH and ethanol. A cleaning protocol consisting of flushing the hydrolysate compartment with NaOH and the organic phase compartment with pure organic phase enabled regeneration and reuse of the module. Ethanol yields from hydrolysates detoxified by membrane extraction using 15% Alamine 336 in oleyl alcohol were about 10% higher than those from hydrolysates detoxified using ammonium hydroxide treatment.

  16. Hydrolysates of lignocellulosic materials for biohydrogen production.

    PubMed

    Chen, Rong; Wang, Yong-Zhong; Liao, Qiang; Zhu, Xun; Xu, Teng-Fei

    2013-05-01

    Lignocellulosic materials are commonly used in bio-H2 production for the sustainable energy resource development as they are abundant, cheap, renewable and highly biodegradable. In the process of the bio-H2 production, the pretreated lignocellulosic materials are firstly converted to monosaccharides by enzymolysis and then to H2 by fermentation. Since the structures of lignocellulosic materials are rather complex, the hydrolysates vary with the used materials. Even using the same lignocellulosic materials, the hydrolysates also change with different pretreatment methods. It has been shown that the appropriate hydrolysate compositions can dramatically improve the biological activities and bio-H2 production performances. Over the past decades, hydrolysis with respect to different lignocellulosic materials and pretreatments has been widely investigated. Besides, effects of the hydrolysates on the biohydrogen yields have also been examined. In this review, recent studies on hydrolysis as well as their effects on the biohydrogen production performance are summarized.

  17. Effects of Ca(OH)(2) treatments ("overliming") on the composition and toxicity of bagasse hemicellulose hydrolysates.

    PubMed

    Martinez, A; Rodriguez, M E; York, S W; Preston, J F; Ingram, L O

    2000-09-01

    Hemicellulose syrups from dilute sulfuric acid hydrolysates of hemicellulose contain inhibitors that prevent efficient fermentation by yeast or bacteria. It is well known that the toxicity of these hydrolysate syrups can be ameliorated by optimized "overliming" with Ca(OH)(2). We have investigated the optimization of overliming treatments for sugar cane bagasse hydrolysates (primarily pentose sugars) using recombinant Escherichia coli LY01 as the biocatalyst. A comparison of composition before and after optimal overliming revealed a substantial reduction in furfural, hydroxymethylfurfural, and three unidentified high-performance liquid chromatography (HPLC) peaks. Organic acids (acetic, formic, levulinic) were not affected. Similar changes have been reported after overliming of spruce hemicellulose hydrolysates (Larsson et al., 1999). Our studies further demonstrated that the extent of furan reduction correlated with increasing fermentability. However, furan reduction was not the sole cause for reduced toxicity. After optimal overliming, bagasse hydrolysate was rapidly and efficiently fermented (>90% yield) by LY01. During these studies, titration, and conductivity were found to be in excellent agreement as methods to estimate sulfuric acid content. Titration was also found to provide an estimate of total organic acids in hydrolysate, which agreed well with the sum of acetic, levulinic, and formic acids obtained by HPLC. Titration of acids, measurement of pH before and after treatment, and furan analyses are proposed as relatively simple methods to monitor the reproducibility of hydrolysate preparations and the effectiveness of overliming treatments.

  18. Complex Physiology and Compound Stress Responses during Fermentation of Alkali-Pretreated Corn Stover Hydrolysate by an Escherichia coli Ethanologen

    PubMed Central

    Schwalbach, Michael S.; Tremaine, Mary; Marner, Wesley D.; Zhang, Yaoping; Bothfeld, William; Higbee, Alan; Grass, Jeffrey A.; Cotten, Cameron; Reed, Jennifer L.; da Costa Sousa, Leonardo; Jin, Mingjie; Balan, Venkatesh; Ellinger, James; Dale, Bruce; Kiley, Patricia J.

    2012-01-01

    The physiology of ethanologenic Escherichia coli grown anaerobically in alkali-pretreated plant hydrolysates is complex and not well studied. To gain insight into how E. coli responds to such hydrolysates, we studied an E. coli K-12 ethanologen fermenting a hydrolysate prepared from corn stover pretreated by ammonia fiber expansion. Despite the high sugar content (∼6% glucose, 3% xylose) and relatively low toxicity of this hydrolysate, E. coli ceased growth long before glucose was depleted. Nevertheless, the cells remained metabolically active and continued conversion of glucose to ethanol until all glucose was consumed. Gene expression profiling revealed complex and changing patterns of metabolic physiology and cellular stress responses during an exponential growth phase, a transition phase, and the glycolytically active stationary phase. During the exponential and transition phases, high cell maintenance and stress response costs were mitigated, in part, by free amino acids available in the hydrolysate. However, after the majority of amino acids were depleted, the cells entered stationary phase, and ATP derived from glucose fermentation was consumed entirely by the demands of cell maintenance in the hydrolysate. Comparative gene expression profiling and metabolic modeling of the ethanologen suggested that the high energetic cost of mitigating osmotic, lignotoxin, and ethanol stress collectively limits growth, sugar utilization rates, and ethanol yields in alkali-pretreated lignocellulosic hydrolysates. PMID:22389370

  19. Sample Preparation for Mass Spectrometry Analysis of Protein-Protein Interactions in Cancer Cell Lines and Tissues.

    PubMed

    Beigbeder, Alice; Vélot, Lauriane; James, D Andrew; Bisson, Nicolas

    2016-01-01

    A precisely controlled network of protein-protein interactions constitutes the basis for functional signaling pathways. This equilibrium is more often than not disrupted in cancer cells, by the aberrant expression or activation of oncogenic proteins. Therefore, the analysis of protein interaction networks in cancer cells has become crucial to expand our comprehension of the molecular underpinnings of tumor formation and progression. This protocol describes a sample preparation method for the analysis of signaling complexes by mass spectrometry (MS), following the affinity purification of a protein of interest from a cancer cell line or a solid tumor. In particular, we provide a spin tip-based protease digestion procedure that offers a more rapid and controlled alternative to other gel-based and gel-free methods. This sample preparation protocol represents a useful strategy to identify protein interactions and to gain insight into the molecular mechanisms that contribute to a given cancer phenotype. PMID:27581032

  20. Molten salt destruction of base hydrolysate

    SciTech Connect

    Watkins, B.E.; Kanna, R.L.; Chambers, R.D.; Upadhye, R.S.; Promeda, C.O.

    1996-10-01

    There is a great need for alternatives to open burn/open detonation of explosives and propellants from dismantled munitions. LANL has investigated the use of base hydrolysis for the demilitarization of explosives. Hydrolysates of Comp B, Octol, Tritonal, and PBXN-109 were processed in the pilot molten salt unit (in building 191). NOx and CO emissions were found to be low, except for CO from PBXN-109 processing. This report describes experimental results of the destruction of the base hydrolysates.

  1. Preparation of Polypyrrole-Protein Composite Films and the Electrochemically Controlled Release of Proteins.

    PubMed

    Jin, Juan; Huang, Zhongbing; Yin, Guangfu; Lin, Jiangli; Li, Qiuping; Han, Donghui

    2016-03-01

    It is fabricated that an electrically controlled release system based on the (poly-L-lactic acid)-mixed polypyrrole (PLLA-PPy) films through casting film of PPy and PLLA mixed solution on the glass plate, in which polyglutamic acid (PGlu)-doped PPy nanoparticles (NPs) with -50 nm diameter are synthesized via chemical oxidation. Surface conductivity of the composite film is (3.33 ± 2.01) x 10(-3) S/cm. Bovine serum albumin (BSA), as a model protein drug, is chemically linked onto the composite film via carbodiimide chemistry due to the good surface nano-structure of PLLA-PPy film. The releases of BSA from PLLA-PPy film under constant current and constant voltage can be achieved using the two-electrode electrochemical system. 6 h accumulative releases of BSA are 276 μg/cm2 and 176 μg/cm2 under 3 mA and 1 V electrical stimulation, respectively, accompanied with de-doping of PGlu and separation of a part of PPy NPs from the composite film. The results of cell experiment indicate that PGlu-doped PPy NPs in the prepared composite film have good cyto-compatibility. These results suggest that PPy-PLLA composite film would be able to be applied in the construction of degradable protein-drug-loaded scaffold for nerve tissue repair. PMID:27455630

  2. Biological and functional properties of proteolytic enzyme-modified egg protein by-products

    PubMed Central

    Pokora, Marta; Eckert, Ewelina; Zambrowicz, Aleksandra; Bobak, Łukasz; Szołtysik, Marek; Dąbrowska, Anna; Chrzanowska, Józefa; Polanowski, Antoni; Trziszka, Tadeusz

    2013-01-01

    Enzymatic hydrolysis led to improve functional properties and biological activity of protein by-products, which can be further used as protein ingredients for food and feed applications. The effects of proteolytic enzyme modification of egg-yolk protein preparation (YP) and white protein preparation (WP), obtained as the by-products left during the course of lecithin, lysozyme, and cystatin isolation on their biological and functional properties, were evaluated by treating a commercial Neutrase. The antihypertensive and antioxidative properties of YP and WP hydrolysates were evaluated based on their angiotensin-converting enzyme (ACE)-inhibitory activity and radical scavenging (DPPH) capacity, ferric reducing power, and chelating of iron activity. The functionality of obtained hydrolysates was also determined. Neutrase caused a degree of hydrolysis (DH) of YP and WP by-products: 27.6% and 20.9%, respectively. In each of them, mixture of peptides with different molecular masses were also observed. YP hydrolysate showed high levels of antioxidant activity. The scavenging capacity, ferric reducing power, and chelating capacity were observed at the level: 0.44 μmol/L Trolox mg−1, 177.35 μg Fe2+ mg−1, and 549.87 μg Fe2+ mg−1, respectively. YP hydrolysate also exhibited significant ACE-inhibitory activity, in which the level was 59.2 μg. Protein solubility was significantly improved as the DH increased. WP hydrolysate showed high water-holding capacity of 43.2. This study indicated that YP and WP hydrolysates could be used in foods as natural antioxidants and functionality enhancers. PMID:24804027

  3. Preparation of protein concentrates from whey and seed products

    SciTech Connect

    Saunders, R.M.; Kohler, G.O.

    1980-01-01

    Whey is mixed with a seed product (e.g., cereal, legumes, oil seeds, flour, etc.) and the pH of the mixture adjusted to 9-10. The resultant mixture is treated to separate a juice from the fibrous residue; in a preferred embodiment of the subsequent process, a protein concentrate is recovered from the juice by adding an acid to it to adjust the pH to 3-4 and subsequently adding sodium hexametaphosphate in an amount sufficient to precipitate the protein product. After adjustment of the pH to 7, a protein concentrate may be obtained by drying the alkaline extract.

  4. Protein quality of several multi-component, commercially prepared foods.

    PubMed

    Miller, J; Wang, Y Y; Beuchat, L R

    1979-09-01

    Eleven commercial food products that may be considered by consumers to be major sources of protein were evaluated. Most of these items contained significant amounts of carbohydrate in the form of cereal products or purified sugars. Seven of the products were at least equal to casein in protein efficiency ratio (PER). Canned spaghetti and meat balls, the TV dinner, canned chicken-and-vegetable baby food, and food bars had PER values between 91 and 73 per cent of that of casein, while PER values for ground beef and high-protein cereal were significantly higher than casein. PMID:469137

  5. Complex coacervation of collagen hydrolysate extracted from leather solid wastes and chitosan for controlled release of lavender oil.

    PubMed

    Ocak, Buğra

    2012-06-15

    In the world, approximately 600,000 metric tonnes of chromium-containing solid wastes are generated by the leather industry each year. Environmental concerns and escalating landfill costs are becoming increasingly serious problems to the leather industry and seeking solutions to these problems is a prime concern in much research today. In this study, solid collagen-based protein hydrolysate was isolated from chromium-tanned leather wastes and its chemical properties were determined. Microcapsules of collagen hydrolysate (CH) - chitosan (C) crosslinked with glutaraldehyde (GA) containing Lavender oil (LO) were prepared by complex coacervation method. The effects of various processing parameters, including the CH to C ratio, LO content, and GA, on the oil load (%), oil content (%), encapsulation efficiency (%) and release rate of LO from microcapsules were investigated. As the ratio of C present in the CH/C mixture and crosslinking density increased, the release rate of LO from microcapsules slowed down. Optical and scanning electron microscopy images illustrated that the LO microcapsules were spherical in shape. Fourier transform infrared spectroscopy (FTIR) studies confirmed that there was no significant interaction between CH/C complex and LO.

  6. Preparation, characterization and functional properties of flax seed protein isolate.

    PubMed

    Kaushik, Pratibha; Dowling, Kim; McKnight, Stafford; Barrow, Colin J; Wang, Bo; Adhikari, Benu

    2016-04-15

    Flaxseed protein isolate (FPI) was extracted from flaxseeds, and its amino acid composition and functional properties (solubility, thermal stability, emulsifying properties and electrostatic charge density, water holding and fat absorption capacities) were determined. The highest purity of FPI (90.6%) was achieved by extraction at 60°C. FPI had a low lysine to arginine ratio of 0.25, which is desired in heart-healthy foods and infant formulas. The denaturation temperature of FPI was 105°C. FPI had the highest emulsion activity index (375.51 m(2)/g), highest emulsion stability index (179.5 h) and zeta potential (-67.4 mV) when compared to those of other commonly used proteins, such as sodium caseinate (SC), whey protein isolate (WPI), gelatin (Gel) and soy protein isolate (SPI). The average emulsion droplet size of emulsions stabilized by these proteins was in the order SCproteins. PMID:26616943

  7. Preparation, characterization and functional properties of flax seed protein isolate.

    PubMed

    Kaushik, Pratibha; Dowling, Kim; McKnight, Stafford; Barrow, Colin J; Wang, Bo; Adhikari, Benu

    2016-04-15

    Flaxseed protein isolate (FPI) was extracted from flaxseeds, and its amino acid composition and functional properties (solubility, thermal stability, emulsifying properties and electrostatic charge density, water holding and fat absorption capacities) were determined. The highest purity of FPI (90.6%) was achieved by extraction at 60°C. FPI had a low lysine to arginine ratio of 0.25, which is desired in heart-healthy foods and infant formulas. The denaturation temperature of FPI was 105°C. FPI had the highest emulsion activity index (375.51 m(2)/g), highest emulsion stability index (179.5 h) and zeta potential (-67.4 mV) when compared to those of other commonly used proteins, such as sodium caseinate (SC), whey protein isolate (WPI), gelatin (Gel) and soy protein isolate (SPI). The average emulsion droplet size of emulsions stabilized by these proteins was in the order SCproteins.

  8. Isolation of oleaginous yeast (Rhodosporidium toruloides) mutants tolerant of sugarcane bagasse hydrolysate.

    PubMed

    Kitahara, Yuki; Yin, Tie; Zhao, Xuebing; Wachi, Masaaki; Du, Wei; Liu, Dehua

    2014-01-01

    Rhodosporidium toruloides is a lipid-producing yeast, the growth of which is severely suppressed when hydrolysates of lignocellulosic biomass are used as carbon source. This is probably due to the toxic substances, such as organic acids, furans, and phenolic compounds produced during the preparation of the hydrolysates. In order to solve this problem, R. toruloides cultures were subjected to atmospheric room-temperature plasma mutagenesis, resulting in the isolation of mutants showing tolerance to sugarcane bagasse hydrolysate (SBH). Three mutant strains, M11, M13, and M18, were found to grow with producing lipids with SBH as carbon source. M11 in particular appeared to accumulate higher levels (up to 60% of dry cell weight) of intracellular lipids. Further, all three mutant strains showed tolerance of vanillin, furfural, and acetic acid, with different spectra, suggesting that different genetic determinants are involved in SBH tolerance.

  9. Purification of glycosylphosphatidylinositol-anchored proteins by modified triton X-114 partitioning and preparative gel electrophoresis.

    PubMed

    Ko, Y G; Thompson, G A

    1995-01-01

    Triton X-114 phase partitioning has frequently been used to obtain preparations enriched in glycosylphosphatidylinositol (GPI)-anchored proteins and other hydrophobic proteins from crude cellular homogenates. We have developed a new modification of this phase-partitioning technique which allows two distinct GPI-anchored proteins of Tetrahymena mimbres to be separated from other hydrophobic as well as hydrophilic proteins and recovered in approximately 90% yield. The unique feature of the new method is a 24-h incubation of the first Triton X-114 extract at -20 degrees C. This improves the partitioning of GPI-anchored proteins into the detergent phase while promoting the aggregation of other hydrophobic proteins. Individual GPI-anchored proteins in the detergent phase are then purified to near homogeneity by one-step preparative sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. By substituting Triton X-114/water mixtures for the conventional SDS buffer used to collect proteins eluting from the preparative gel, the purified GPI-anchored protein can be rapidly concentrated from relatively large volumes of eluate by phase partitioning at 32 degrees C. The method is also effective in separating mammalian GPI-anchored alkaline phosphatase from other proteins. It is likely to be of general utility in characterizing the GPI anchor structures associated with nonabundant and abundant GPI-anchored proteins coexisting within the same cell type.

  10. DSCG binding protein and process for preparing same

    SciTech Connect

    Pecht, I.; Mazurek, N.

    1987-07-28

    An essentially pure protein is described consisting essentially of the protein, (CBP), present in nature in membranes of basophile cells and in mast cells, having a molecular weight of about 60,000 +- 2,000 determined by SDS polyacrylamide electrophoresis an isoelectric point of about 3.9 and an amino acid composition of about 4 units of asparagine, 3 units of threonine and serine, 3 units glycine, 2 units alanine, 2 units proline, 1 unit cysteine, 2 units valine, 1 unit methionine, 1 unit isoleucine, 2 units leucine, 1 unit tyrosine, 1 unit phenylalanine, 2 units histamine, 2 units lysine and 1 unit arginine. The protein is able to build calcium and having a calcium dependent affinity to the disodium salt of 1,2 bis(-2 carboxychromon-5-yloxy)-2-hydroxy propane (DSCG).

  11. Preparation and evaluation of tara-modified proteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Quebracho, a vegetable tannin, can be used to modify gelatin to produce a product that has been applied effectively as a filler in leather processing, as described in our previous report. In this ongoing study, another vegetable tannin tara is examined for its possible application in protein modifi...

  12. Determination of protein carbonyls in plasma, cell extracts, tissue homogenates, isolated proteins: Focus on sample preparation and derivatization conditions

    PubMed Central

    Weber, Daniela; Davies, Michael J.; Grune, Tilman

    2015-01-01

    Protein oxidation is involved in regulatory physiological events as well as in damage to tissues and is thought to play a key role in the pathophysiology of diseases and in the aging process. Protein-bound carbonyls represent a marker of global protein oxidation, as they are generated by multiple different reactive oxygen species in blood, tissues and cells. Sample preparation and stabilization are key steps in the accurate quantification of oxidation-related products and examination of physiological/pathological processes. This review therefore focuses on the sample preparation processes used in the most relevant methods to detect protein carbonyls after derivatization with 2,4-dinitrophenylhydrazine with an emphasis on measurement in plasma, cells, organ homogenates, isolated proteins and organelles. Sample preparation, derivatization conditions and protein handling are presented for the spectrophotometric and HPLC method as well as for immunoblotting and ELISA. An extensive overview covering these methods in previously published articles is given for researchers who plan to measure protein carbonyls in different samples. PMID:26141921

  13. Functional properties of autolysate and enzymatic hydrolysates from yam tsukuneimo (Dioscorea opposita Thunb.) tuber mucilage tororo: antioxidative activity and antihypertensive activity.

    PubMed

    Nagai, Takeshi; Suzuki, Nobutaka; Kai, Norihisa; Tanoue, Yasuhiro

    2014-12-01

    Yam tsukuneimo tuber mucilage tororo hydrolysates were prepared by autolysis and three different peptic enzymes. Except for pepsin hydrolysate, tororo was perfectly digested. Each hydrolysate for 100 mg/ml significantly prolonged the induction period of auto-oxidation of linoleic acid, which was similar to 5 mM ascorbic acid. These hydrolysates also possessed high scavenging activities such as superoxide anion radicals, hydroxyl radicals, and DPPH radicals. Moreover, high antihypertensive activities were detected in these hydrolysates except for autolysate, which were similar to various fermented foods such as miso, natto, sake, cheese, and so on. Present findings suggest that yam tsukuneimo tuber mucilage tororo may be useful for preventing diseases associated with reactive oxygen species and blood pressure in the body system and it can fully absorb the useful components from it to digest using the gastrointestinal enzymes.

  14. Preparative Protein Production from Inclusion Bodies and Crystallization: A Seven-Week Biochemistry Sequence

    ERIC Educational Resources Information Center

    Peterson, Megan J.; Snyder, W. Kalani; Westerman, Shelley; McFarland, Benjamin J.

    2011-01-01

    We describe how to produce and purify proteins from "Escherichia coli" inclusion bodies by adapting versatile, preparative-scale techniques to the undergraduate laboratory schedule. This 7-week sequence of experiments fits into an annual cycle of research activity in biochemistry courses. Recombinant proteins are expressed as inclusion bodies,…

  15. Properties of proteins in cancer procoagulant preparations that are detected by anti-tissue factor antibodies.

    PubMed

    Raasi, Shari; Mielicki, Wojciech P; Gordon, Stuart G; Korte, Wolfgang

    2004-08-15

    Cancer procoagulant (CP) and tissue factor (TF; only in complex with Factor VIIa (FVIIa)) can activate FX to FXa. Controversy still exists whether or not CP is an entity different from TF, or whether CP activity is due to contamination of CP preparations with TF/FVIIa complex. We therefore looked for proteins in CP preparations that were detected by anti-TF antibodies and then sequenced these proteins. One- and two-dimensional gels of CP and TF were used to identify proteins immunoreactive to monoclonal anti-CP and anti-TF antibodies (Mabs). Those proteins in the CP preparation recognized by anti-TF antibodies were sequenced. Angiotensinogen precursor, alpha-1-antitrypsin precursor, and vitamin D-binding protein were identified along with one so far unidentified sequence; however, no TF-sequences were identified. Also, no proteins with the correct molecular weight for TF were identified using anti-TF antibodies. It seems possible that CP preparations contain proteins that have some epitopes similar to the epitopes recognized in TF by anti-TF Mab. However, these proteins do neither have the molecular weight nor the amino acid sequence of TF.

  16. Preparative Procedures Markedly Influence the Appearance and Structural Integrity of Protein Storage Vacuoles in Soybean Seeds

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In legumes, vacuoles serve as the final depository for storage proteins. The protein storage vacuoles (PSVs) of soybean contain electron-transparent globoid regions in which phytic acid (myo-inositol-1,2,3,4,5,6-hexakisphosphate) is sequestered. Here, I report the effect of preparative procedures o...

  17. Preparative Purification of Recombinant Proteins: Current Status and Future Trends

    PubMed Central

    Saraswat, Mayank; Ravidá, Alessandra; Holthofer, Harry

    2013-01-01

    Advances in fermentation technologies have resulted in the production of increased yields of proteins of economic, biopharmaceutical, and medicinal importance. Consequently, there is an absolute requirement for the development of rapid, cost-effective methodologies which facilitate the purification of such products in the absence of contaminants, such as superfluous proteins and endotoxins. Here, we provide a comprehensive overview of a selection of key purification methodologies currently being applied in both academic and industrial settings and discuss how innovative and effective protocols such as aqueous two-phase partitioning, membrane chromatography, and high-performance tangential flow filtration may be applied independently of or in conjunction with more traditional protocols for downstream processing applications. PMID:24455685

  18. Visualizing Proteins and Macromolecular Complexes by Negative Stain EM: from Grid Preparation to Image Acquisition

    PubMed Central

    Booth, David S.; Avila-Sakar, Agustin; Cheng, Yifan

    2011-01-01

    Single particle electron microscopy (EM), of both negative stained or frozen hydrated biological samples, has become a versatile tool in structural biology 1. In recent years, this method has achieved great success in studying structures of proteins and macromolecular complexes 2, 3. Compared with electron cryomicroscopy (cryoEM), in which frozen hydrated protein samples are embedded in a thin layer of vitreous ice 4, negative staining is a simpler sample preparation method in which protein samples are embedded in a thin layer of dried heavy metal salt to increase specimen contrast 5. The enhanced contrast of negative stain EM allows examination of relatively small biological samples. In addition to determining three-dimensional (3D) structure of purified proteins or protein complexes 6, this method can be used for much broader purposes. For example, negative stain EM can be easily used to visualize purified protein samples, obtaining information such as homogeneity/heterogeneity of the sample, formation of protein complexes or large assemblies, or simply to evaluate the quality of a protein preparation. In this video article, we present a complete protocol for using an EM to observe negatively stained protein sample, from preparing carbon coated grids for negative stain EM to acquiring images of negatively stained sample in an electron microscope operated at 120kV accelerating voltage. These protocols have been used in our laboratory routinely and can be easily followed by novice users. PMID:22215030

  19. [Comparative studies on the enzymatic absorption of protein hydrolysates in the small intestine of the rat. 3. The absorption trypsins thermatatic and trypsin-thermatatic hydrolysates of a fava bean protein isolate compared to an equimolar mixture of free amino acids].

    PubMed

    Proll, J; Friedrich, M; Noack, J; Noack, R

    1985-01-01

    Tryptic, thermitatic, and tryptic-thermitatic Faba bean protein hydrolyzates as well as their equimolar mixture of amino acids were perfused through proximal and distal parts of the intestine (10 cm length) of non-narcotized rats. The total amino-acid concentration of the perfused solution was 50 mM. The absorption of nitrogen and total amino acids from the tryptic and tryptic-thermitatic hydrolyzates was lower than that from the amino-acid mixture, the absorption from the thermitatic hydrolyzate was in accordance with that from the amino-acid mixture. The absorption pattern of the amino acids which preferably undergo a peptidic absorption is similar with the three hydrolyzates: in the proximal intestinal part this concerns glutamic acid and serine, in the distal intestinal part--methionine, alanine, glycin, and serine. The absorption pattern of the amino acids is different between the three hydrolyzates and the amino-acid mixture. Between the absorption pattern of the amino acids from the three hydrolyzates little differences were evident only in the proximal intestinal part. The coefficients of variation of the tryptic-thermitatic hydrolyzates are in accordance with those of the amino-acid mixture, whereas that of the thermitatic hydrolyzates is significantly lower. In the distal intestinal part all supplied forms are more rapidly absorbed than in the proximal part of the intestine.

  20. Efficient preparation and analysis of membrane and membrane protein systems.

    PubMed

    Javanainen, Matti; Martinez-Seara, Hector

    2016-10-01

    Molecular dynamics (MD) simulations have become a highly important technique to consider lipid membrane systems, and quite often they provide considerable added value to laboratory experiments. Rapid development of both software and hardware has enabled the increase of time and size scales reachable by MD simulations to match those attainable by several accurate experimental techniques. However, until recently, the quality and maturity of software tools available for building membrane models for simulations as well as analyzing the results of these simulations have seriously lagged behind. Here, we discuss the recent developments of such tools from the end-users' point of view. In particular, we review the software that can be employed to build lipid bilayers and other related structures with or without embedded membrane proteins to be employed in MD simulations. Additionally, we provide a brief critical insight into force fields and MD packages commonly used for membrane and membrane protein simulations. Finally, we list analysis tools that can be used to study the properties of membrane and membrane protein systems. In all these points we comment on the respective compatibility of the covered tools. We also share our opinion on the current state of the available software. We briefly discuss the most commonly employed tools and platforms on which new software can be built. We conclude the review by providing a few ideas and guidelines on how the development of tools can be further boosted to catch up with the rapid pace at which the field of membrane simulation progresses. This includes improving the compatibility between software tools and promoting the openness of the codes on which these applications rely. This article is part of a Special Issue entitled: Biosimulations edited by Ilpo Vattulainen and Tomasz Róg. PMID:26947184

  1. Influenza vaccines: from whole virus preparations to recombinant protein technology.

    PubMed

    Huber, Victor C

    2014-01-01

    Vaccination against influenza represents our most effective form of prevention. Historical approaches toward vaccine creation and production have yielded highly effective vaccines that are safe and immunogenic. Despite their effectiveness, these historical approaches do not allow for the incorporation of changes into the vaccine in a timely manner. In 2013, a recombinant protein-based vaccine that induces immunity toward the influenza virus hemagglutinin was approved for use in the USA. This vaccine represents the first approved vaccine formulation that does not require an influenza virus intermediate for production. This review presents a brief history of influenza vaccines, with insight into the potential future application of vaccines generated using recombinant technology.

  2. Phosphate acceptor amino acid residues in structural proteins of rhabdoviruses.

    PubMed

    Sokol, F; Tan, K B; McFalls, M L; Madore, P

    1974-07-01

    Partial acid hydrolysates of the [(32)P]phosphate- or [(3)H]serine-labeled proteins of purified vesicular stomatitis, rabies, Lagos bat, Mokola, or spring viremia of carp virions and of purified intracellular nucleocapsids of these viruses have been analyzed by paper electrophoresis for the presence of phosphorylated amino acids. Both phosphoserine and phosphothreonine, with the former predominant, were present in virion and nucleocapsid preparations that contained phosphoproteins. An exception was the fish rhabdovirus, which contained only phosphoserine. When vesicular stomatitis or rabies virus proteins were phosphorylated in a cell-free system by the virion-associated protein kinase and analyzed for the presence of phosphorylated amino acid residues, phosphoserine was again found to be more abundant than phosphothreonine. After in vitro protein phosphorylation, another phospho-compound, possibly a third phosphoamino acid, was detected in the partial acid hydrolysates of these viruses. PMID:4365328

  3. Preparation of iron bound succinylated milk protein concentrate and evaluation of its stability.

    PubMed

    Shilpashree, B G; Arora, Sumit; Sharma, Vivek; Bajaj, Rajesh Kumar; Tomar, S K

    2016-04-01

    Major problems associated with the fortification of soluble iron salts include chemical reactivity and incompatibility with other components. Milk protein concentrate (MPC) are able to bind significant amount of iron due to the presence of both casein and whey protein. MPC in its native state possess very poor solubility, therefore, succinylated derivatives of MPC (succ. MPC) were also used for the preparation of protein-iron complex. Preparation of the complex involved centrifugation (to remove insoluble iron), ultrafiltration (to remove unbound iron) and lyophilisation (to attain in dry form). Iron binding ability of MPC enhanced significantly (P<0.05) upon succinylation. Stability of bound iron from both varieties of complexes was monitored under different conditions encountered during processing. Higher stability (P<0.05) of bound iron was observed in succ. MPC-iron complex than native protein complex. This method could be adopted for the production of stable iron enriched protein, an organic iron source. PMID:26593557

  4. Preparation of iron bound succinylated milk protein concentrate and evaluation of its stability.

    PubMed

    Shilpashree, B G; Arora, Sumit; Sharma, Vivek; Bajaj, Rajesh Kumar; Tomar, S K

    2016-04-01

    Major problems associated with the fortification of soluble iron salts include chemical reactivity and incompatibility with other components. Milk protein concentrate (MPC) are able to bind significant amount of iron due to the presence of both casein and whey protein. MPC in its native state possess very poor solubility, therefore, succinylated derivatives of MPC (succ. MPC) were also used for the preparation of protein-iron complex. Preparation of the complex involved centrifugation (to remove insoluble iron), ultrafiltration (to remove unbound iron) and lyophilisation (to attain in dry form). Iron binding ability of MPC enhanced significantly (P<0.05) upon succinylation. Stability of bound iron from both varieties of complexes was monitored under different conditions encountered during processing. Higher stability (P<0.05) of bound iron was observed in succ. MPC-iron complex than native protein complex. This method could be adopted for the production of stable iron enriched protein, an organic iron source.

  5. Iron-Binding Capacity of Defatted Rice Bran Hydrolysate and Bioavailability of Iron in Caco-2 Cells.

    PubMed

    Foong, Lian-Chee; Imam, Mustapha Umar; Ismail, Maznah

    2015-10-21

    The present study was aimed at utilizing defatted rice bran (DRB) protein as an iron-binding peptide to enhance iron uptake in humans. DRB samples were treated with Alcalase and Flavourzyme, and the total extractable peptides were determined. Furthermore, the iron-binding capacities of the DRB protein hydrolysates were determined, whereas iron bioavailability studies were conducted using an in vitro digestion and absorption model (Caco-2 cells). The results showed that the DRB protein hydrolysates produced by combined Alcalase and Flavourzyme hydrolysis had the best iron-binding capacity (83%) after 90 min of hydrolysis. The optimal hydrolysis time to produce the best iron-uptake in Caco-2 cells was found to be 180 min. The results suggested that DRB protein hydrolysates have potent iron-binding capacities and may enhance the bioavailability of iron, hence their suitability for use as iron-fortified supplements. PMID:26435326

  6. [Fractional and amino acid composition of krill proteins and the potential for obtaining protein preparations].

    PubMed

    Orlova, T A; Churina, E E; Kuranova, L K

    1985-01-01

    Studies of the fractional composition of krill proteins demonstrated that the content of protein fractions changes depending on the time of krill catch. The highest amount of water-soluble proteins is contained by krill caught in December (64%), of salt-soluble by krill caught in June (12%), base-soluble by krill caught in May, September and February (34%). Krill protein contains from 50 to 60% of water- and salt-soluble fractions. Analysis of the amino acid composition of krill proteins showed that it does not differ essentially from that of adequate food proteins.

  7. Use of a purified outer membrane protein F (porin) preparation of Pseudomonas aeruginosa as a protective vaccine in mice.

    PubMed Central

    Gilleland, H E; Parker, M G; Matthews, J M; Berg, R D

    1984-01-01

    The outer membrane protein F (porin) from the PAO1 strain of Pseudomonas aeruginosa was purified by two different methods. One procedure involved separation by column chromatography of proteins extracted from isolated outer membranes, whereas the other involved extraction from gels after slab polyacrylamide gel electrophoresis of proteins extracted from cell envelopes. Both procedures yielded protein F preparations which successfully immunized mice from subsequent challenge with the PAO1 strain. The protein F preparations contained small quantities of lipopolysaccharide (LPS). This level of LPS contamination protected immunized mice from challenge with the homologous LPS serotype strain. However, immunization of mice with protein F preparations from the PAO1 strain also afforded protection against challenge with two different LPS serotype strains. This protective ability was lost when the protein F preparation was treated with papain before use as a vaccine. These observations support the conclusion that protein F has protective ability, which is not due to LPS contamination, when given as a vaccine. After immunization with the protein F preparation, mice showed an increase in antibody titer to the purified protein F preparation by enzyme-linked immunosorbent assay. Mice were protected passively by administration of rabbit antisera raised to the protein F preparation. These results indicate that the protein F preparation elicits a specific humoral antibody response in immunized animals. Our results suggest that purified protein F has potential as an effective vaccine for P. aeruginosa. Images PMID:6323316

  8. Protein distribution in lupin protein isolates from Lupinus angustifolius L. prepared by various isolation techniques.

    PubMed

    Muranyi, Isabel S; Volke, Daniela; Hoffmann, Ralf; Eisner, Peter; Herfellner, Thomas; Brunnbauer, Markus; Koehler, Peter; Schweiggert-Weisz, Ute

    2016-09-15

    Differences in the protein distribution of various protein isolates from Lupinus angustifolius L. Vitabor were identified as affected by the isolation procedure (alkaline and/or salt-induced extraction followed by isoelectric and/or dilutive precipitation). Protein isolates extracted in alkaline solution showed higher protein yields (26.4-31.7%) compared to salt-induced extraction (19.8-30.0%) or combined alkaline and salt-induced extraction (23.3-25.6%). Chemical variations among the protein isolates especially occurred within the albumins. Protein isolates precipitated isoelectrically showed the highest contents, whereas protein isolates precipitated by dilutive showed the lowest contents of conglutin δ. Furthermore, the alkaline subunits of conglutin α and conglutin γ decreased during alkaline extraction compared to salt-induced extraction. A decrease in protein-bound polar and basic amino acids was shown after protein isolation. In contrast, the amounts of nonpolar, aliphatic, aromatic, hydroxylated and sulfur-rich amino acids were higher in the lupin protein isolates compared to the lupin flakes. However, the functional side chains could not be related to the specific molecular arrangements of the protein isolates, as a similar amino acid composition was found among the protein isolates. PMID:27080873

  9. Protein distribution in lupin protein isolates from Lupinus angustifolius L. prepared by various isolation techniques.

    PubMed

    Muranyi, Isabel S; Volke, Daniela; Hoffmann, Ralf; Eisner, Peter; Herfellner, Thomas; Brunnbauer, Markus; Koehler, Peter; Schweiggert-Weisz, Ute

    2016-09-15

    Differences in the protein distribution of various protein isolates from Lupinus angustifolius L. Vitabor were identified as affected by the isolation procedure (alkaline and/or salt-induced extraction followed by isoelectric and/or dilutive precipitation). Protein isolates extracted in alkaline solution showed higher protein yields (26.4-31.7%) compared to salt-induced extraction (19.8-30.0%) or combined alkaline and salt-induced extraction (23.3-25.6%). Chemical variations among the protein isolates especially occurred within the albumins. Protein isolates precipitated isoelectrically showed the highest contents, whereas protein isolates precipitated by dilutive showed the lowest contents of conglutin δ. Furthermore, the alkaline subunits of conglutin α and conglutin γ decreased during alkaline extraction compared to salt-induced extraction. A decrease in protein-bound polar and basic amino acids was shown after protein isolation. In contrast, the amounts of nonpolar, aliphatic, aromatic, hydroxylated and sulfur-rich amino acids were higher in the lupin protein isolates compared to the lupin flakes. However, the functional side chains could not be related to the specific molecular arrangements of the protein isolates, as a similar amino acid composition was found among the protein isolates.

  10. Protein Chips Compatible with MALDI Mass Spectrometry Prepared by Ambient Ion Landing.

    PubMed

    Pompach, Petr; Benada, Oldřich; Rosůlek, Michal; Darebná, Petra; Hausner, Jiří; Růžička, Viktor; Volný, Michael; Novák, Petr

    2016-09-01

    We present a technology that allows the preparation of matrix-assisted laser desorption/ionization (MALDI)-compatible protein chips by ambient ion landing of proteins and successive utilization of the resulting protein chips for the development of bioanalytical assays. These assays are based on the interaction between the immobilized protein and the sampled analyte directly on the protein chip and subsequent in situ analysis by MALDI mass spectrometry. The electrosprayed proteins are immobilized on dry metal and metal oxide surfaces, which are nonreactive under normal conditions. The ion landing of electrosprayed protein molecules is performed under atmospheric pressure by an automated ion landing apparatus that can manufacture protein chips with a predefined array of sample positions or any other geometry of choice. The protein chips prepared by this technique are fully compatible with MALDI ionization because the metal-based substrates are conductive and durable enough to be used directly as MALDI plates. Compared to other materials, the nonreactive surfaces show minimal nonspecific interactions with chemical species in the investigated sample and are thus an ideal substrate for selective protein chips. Three types of protein chips were used in this report to demonstrate the bioanalytical applications of ambient ion landing. The protein chips with immobilized proteolytic enzymes showed the usefulness for fast in situ peptide MALDI sequencing; the lectin-based protein chips showed the ability to enrich glycopeptides from complex mixtures with subsequent MALDI analysis, and the protein chips with immobilized antibodies were used for a novel immunoMALDI workflow that allowed the enrichment of antigens from the serum followed by highly specific MALDI detection. PMID:27478994

  11. Production of feather hydrolysates with antioxidant, angiotensin-I converting enzyme- and dipeptidyl peptidase-IV-inhibitory activities.

    PubMed

    Fontoura, Roberta; Daroit, Daniel J; Correa, Ana P F; Meira, Stela M M; Mosquera, Mauricio; Brandelli, Adriano

    2014-09-25

    The antioxidant and antihypertensive activities of feather hydrolysates obtained with the bacterium Chryseobacterium sp. kr6 were investigated. Keratin hydrolysates were produced with different concentrations of thermally denatured feathers (10-75 g l(-1)) and initial pH values (6.0-9.0). Soluble proteins accumulated in high amounts in media with 50 and 75 g l(-1) of feathers, reaching values of 18.5 and 22 mg ml(-1), respectively, after 48 hours of cultivation. In media with 50 g l(-1) of feathers, initial pH had minimal effect after 48 hours. Maximal protease production was observed after 24 hours of cultivation, and feather concentration and initial pH values showed no significant effect on enzyme yields at this time. Feather hydrolysates displayed in vitro antioxidant properties, and optimal antioxidant activities were observed in cultures with 50 g l(-1) feathers, at initial pH 8.0, after 48 hours growth at 30°C. Also, feather hydrolysates were demonstrated to inhibit the angiotesin I-converting enzyme by 65% and dipeptidyl peptidase-IV by 44%. The bioconversion of an abundant agroindustrial waste such as chicken feathers can be utilized as a strategy to obtain hydrolysates with antioxidant and antihypertensive activities. Feather hydrolysates might be employed as supplements in animal feed, and also as a potential source of bioactive molecules for feed, food and drug development.

  12. Peat hydrolysate medium optimization for pullulan production.

    PubMed

    Boa, J M; Leduy, A

    1984-07-01

    Peat hydrolysate, a diluted acid-autoclaved extract of peat, was used as a substrate for the production of the extracellular polysaccharide pullulan by three strains of Aureobasidium pullulans, 140B, 142, and 2552. It was found that the addition of (NH(4))(2)SO(4) and K(2)HPO(4) as sources of nitrogen and phosphate, respectively, is not necessary for the polysaccharide production. The economically optimized culture medium for large-scale production of pullulan contains peat hydrolysate, 0.05% NaCl, 0.02% MgSO(4), and 0.01% antifoam FG-10. The initial pH of peat hydrolysate medium is adjusted to its optimum value of 6.0 with Ca(OH)(2). The total ingredient cost for the production of each kilogram of pullulan with optimized medium is only 1/10 of that with the nonoptimized medium. In this study, a zero cost for peat hydrolysate was assumed, since it is an effluent of the peat and peat processing industries.

  13. Hydrolysates of lignocellulosic materials for biohydrogen production

    PubMed Central

    Chen, Rong; Wang, Yong-Zhong; Liao, Qiang; Zhu, Xun; Xu, Teng-Fei

    2013-01-01

    Lignocellulosic materials are commonly used in bio-H2 production for the sustainable energy resource development as they are abundant, cheap, renewable and highly biodegradable. In the process of the bio-H2 production, the pretreated lignocellulosic materials are firstly converted to monosaccharides by enzymolysis and then to H2 by fermentation. Since the structures of lignocellulosic materials are rather complex, the hydrolysates vary with the used materials. Even using the same lignocellulosic materials, the hydrolysates also change with different pretreatment methods. It has been shown that the appropriate hydrolysate compositions can dramatically improve the biological activities and bio-H2 production performances. Over the past decades, hydrolysis with respect to different lignocellulosic materials and pretreatments has been widely investigated. Besides, effects of the hydrolysates on the biohydrogen yields have also been examined. In this review, recent studies on hydrolysis as well as their effects on the biohydrogen production performance are summarized. [BMB Reports 2013; 46(5): 244-251] PMID:23710634

  14. A novel family VIII carboxylesterase hydrolysing third- and fourth-generation cephalosporins.

    PubMed

    Jeon, Jeong Ho; Lee, Hyun Sook; Lee, Jung Hun; Koo, Bon-Sung; Lee, Chang-Muk; Lee, Sang Hee; Kang, Sung Gyun; Lee, Jung-Hyun

    2016-01-01

    A metagenomic library was constructed from a soil sample of spindle tree-rhizosphere. From this library, one clone with esterase activity was selected. The sequence analysis revealed an open reading frame (EstSTR1) encoded protein of 390 amino acids. EstSTR1 is a family VIII carboxylesterase and retains the S-X-X-K motif conserved in both family VIII carboxylesterases and class C β-lactamases. The estSTR1 gene was overexpressed in E. coli and the recombinant protein was purified by purified by metal chelating affinity chromatography and size-exclusion chromatography. EstSTR1 hydrolysed p-nitrophenyl esters, exhibited the highest activity toward p-nitrophenyl butyrate. Furthermore, EstSTR1 could hydrolyse third- and fourth-generation cephalosporins (cefotaxime and cefepime) as well as first-generation cephalosporin (cephalothin). Site-directed mutagenesis studies revealed that a catalytic residue, Ser71, of EstSTR1 plays an essential role in hydrolysing both antibiotics and p-nitrophenyl esters. We demonstrate that a metagenome-derived carboxylesterase displays β-lactam-hydrolysing activities toward third- and fourth-generation cephalosporins. PMID:27186489

  15. Soybean and casein hydrolysates induce grapevine immune responses and resistance against Plasmopara viticola

    PubMed Central

    Lachhab, Nihed; Sanzani, Simona M.; Adrian, Marielle; Chiltz, Annick; Balacey, Suzanne; Boselli, Maurizio; Ippolito, Antonio; Poinssot, Benoit

    2014-01-01

    Plasmopara viticola, the causal agent of grapevine downy mildew, is one of the most devastating grape pathogen in Europe and North America. Although phytochemicals are used to control pathogen infections, the appearance of resistant strains and the concern for possible adverse effects on environment and human health are increasing the search for alternative strategies. In the present investigation, we successfully tested two protein hydrolysates from soybean (soy) and casein (cas) to trigger grapevine resistance against P. viticola. On Vitis vinifera cv. Marselan plants, the application of soy and cas reduced the infected leaf surface by 76 and 63%, as compared to the control, respectively. Since both hydrolysates might trigger the plant immunity, we investigated their ability to elicit grapevine defense responses. On grapevine cell suspensions, a different free cytosolic calcium signature was recorded for each hydrolysate, whereas a similar transient phosphorylation of two MAP kinases of 45 and 49 kDa was observed. These signaling events were followed by transcriptome reprogramming, including the up-regulation of defense genes encoding pathogenesis-related (PR) proteins and the stilbene synthase enzyme responsible for the biosynthesis of resveratrol, the main grapevine phytoalexin. Liquid chromatography analyses confirmed the production of resveratrol and its dimer metabolites, δ- and ε-viniferins. Overall, soy effects were more pronounced as compared to the cas ones. Both hydrolysates proved to act as elicitors to enhance grapevine immunity against pathogen attack. PMID:25566290

  16. A Simple and Rapid Method for Preparing a Cell-Free Bacterial Lysate for Protein Synthesis

    PubMed Central

    Kaduri, Maya; Shainsky-Roitman, Janna; Goldfeder, Mor; Ivanir, Eran; Benhar, Itai; Shoham, Yuval; Schroeder, Avi

    2016-01-01

    Cell-free protein synthesis (CFPS) systems are important laboratory tools that are used for various synthetic biology applications. Here, we present a simple and inexpensive laboratory-scale method for preparing a CFPS system from E. coli. The procedure uses basic lab equipment, a minimal set of reagents, and requires less than one hour to process the bacterial cell mass into a functional S30-T7 extract. BL21(DE3) and MRE600 E. coli strains were used to prepare the S30-T7 extract. The CFPS system was used to produce a set of fluorescent and therapeutic proteins of different molecular weights (up to 66 kDa). This system was able to produce 40–150 μg-protein/ml, with variations depending on the plasmid type, expressed protein and E. coli strain. Interestingly, the BL21-based CFPS exhibited stability and increased activity at 40 and 45°C. To the best of our knowledge, this is the most rapid and affordable lab-scale protocol for preparing a cell-free protein synthesis system, with high thermal stability and efficacy in producing therapeutic proteins. PMID:27768741

  17. Electron Cryomicroscopy of Membrane Proteins: Specimen Preparation for Two-Dimensional Crystals and Single Particles

    PubMed Central

    Schmidt-Krey, Ingeborg; Rubinstein, John L.

    2010-01-01

    Membrane protein structure and function can be studied by two powerful and highly complementary electron cryomicroscopy (cryo-EM) methods: electron crystallography of two-dimensional (2D) crystals and single particle analysis of detergent-solubilized protein complexes. To obtain the highest-possible resolution data from membrane proteins, whether prepared as 2D crystals or single particles, cryo-EM samples must be vitrified with great care. Grid preparation for cryo-EM of 2D crystals is possible by back-injection, the carbon sandwich technique, drying in sugars before cooling in the electron microscope, or plunge-freezing. Specimen grids for single particle cryo-EM studies of membrane proteins are usually produced by plunge-freezing protein solutions, supported either by perforated or a continuous carbon film substrate. This review outlines the different techniques available and the suitability of each method for particular samples and studies. Experimental considerations in sample preparation and preservation include the protein itself and the presence of lipid or detergent. The appearance of cryo-EM samples in different conditions is also discussed. PMID:20678942

  18. Radioactive labeling of proteins in cultured postimplantation mouse embryos. I. Influence of the embryo preparation method

    SciTech Connect

    Nowak, J.; Klose, J. )

    1989-07-01

    Conditions for optimum incorporation of radioactive amino acids into proteins of cultured postimplantation mouse embryos were investigated under the aspect of using these proteins for two-dimensional electrophoretic separations followed by fluorography. The aim was to obtain highly radioactive proteins under conditions as physiological as possible. Embryos at Days 10, 11, and 12 of gestation were prepared in different ways and incubated for 4 h in Tyrode's solution containing ({sup 3}H)amino acids (mixture) at a concentration of 27 microCi/ml medium. The preparations were: (a) yolk sac opened, placenta and blood circulation intact; (b) yolk sac and amnion opened, placenta and blood circulation intact (Day 10 embryos only); (c) placenta, yolk sac, and amnion removed (embryo naked); (d) naked embryos cut randomly into pieces (Day 10 embryos only). After incubation whole embryos or certain parts (tail, liver, rest body) were investigated by determining the radioactivity taken up by the protein. The results are given in dpm per mg protein per embryo. Radioactivity of proteins was about 3 times higher in naked embryos than in embryos left in their yolk sacs. This was true for all three stages investigated. However, the degree of radioactivity in the various parts of naked embryos differed by a factor of 15, whereas radioactivity was evenly distributed in embryos incubated in their yolk sacs. Therefore, embryos prepared according to the first method (see above) fulfilled the conditions required at the best.

  19. An effective approach to prepare uniform protein Zn2+ nanoparticles under mild conditions

    NASA Astrophysics Data System (ADS)

    Yuan, Weien; Wu, Fei; Geng, Yan; Xu, Songlin; Jin, Tuo

    2007-04-01

    We report a novel yet simple method to prepare nanometre-sized dense and spherical protein-Zn2+ particles, a system useful for delivery of structurally delicate protein therapeutics. To demonstrate the preparation method, a model protein, bovine serum albumin (BSA) and zinc acetate were added to a solution of polyethylene glycol, followed by freeze-drying. Protein-Zn2+ particles were harvested by washing the lyophilized powder using dichloromethane or acetonitrile to remove PEG. By varying formulation parameters, such as molecular weight and concentration of PEG, particles of desired diameters can easily be obtained in the range of 30-350 nm. This particulate system was characterized using electron microscopy, laser scattering, x-ray diffraction (XRD) and differential scanning calorimeter (DSC) for morphology, particle sizes, crystalline structure and thermal properties, respectively. The results proved that the said protein-Zn2+ particles are spherical, glassy and uniform in size. For the mechanism of how the PEG continuous phase facilitates the formation of spherical protein-Zn2+ particles (which are irregular in shape when formed by spray-drying), we hypothesize that freezing-induced phase separation occurred during the freezing process, during which the BSA-Zn2+ complex was 'squeezed' into each dispersed particle. For structural integrity, the protein reconstituted from the particles showed the same SEC-HPLC charts as the original samples, suggesting the particle forming process did not induce protein aggregation.

  20. Fibrous scaffolds loaded with protein prepared by blend or coaxial electrospinning.

    PubMed

    Ji, Wei; Yang, Fang; van den Beucken, Jeroen J J P; Bian, Zhuan; Fan, Mingwen; Chen, Zhi; Jansen, John A

    2010-11-01

    The aim of the present study was to fabricate polycaprolactone-based nanofibrous scaffolds with incorporated protein via either the blend or coaxial electrospinning technique. Both techniques were compared with respect to processing set-up and scaffold characteristics as well as the release kinetics and biological activity of the loaded protein. Bovine serum albumin was used as a model protein to determine release profiles, while alkaline phosphatase was used to determine protein activity after the electrospinning process. Coaxial electrospinning resulted in a uniform fiber morphology with a core-shell structure, and a homogeneous protein distribution throughout the core of the fibers. In contrast, blend electrospinning formed bead-like fibers with a heterogeneous protein distribution in the fibers. The coaxial scaffold exhibited more sustained release profiles than the comparative blend scaffold, and the additive poly(ethylene glycol) (PEG) in the coaxial scaffold accelerated protein release. Both electrospinning processes decreased the biological activity of the incorporated protein, but coaxial electrospinning with PEG as an additive showed up to 75% preservation of the initial biological activity. Thus, coaxial electrospinning was demonstrated to be superior to blend electrospinning for the preparation of nanofibrous scaffolds with a uniform fibrous structure and protein distribution and sustained protein release kinetics as well as high preservation of the protein activity.

  1. Preparation and application of vegetable proteins, especially proteins from sunflower seed, for human consumption. An approach.

    PubMed

    Gassmann, B

    1983-01-01

    About 80% of the world protein production are of vegetable origin. More than half the vegetable protein is fed to animals, whereas merely 10 kg protein per capita are obtained from meat, milk and eggs per year. Therefore, and because of rising prices for raw materials and energy the production and the firsthand utilisation of proteinacous plants for foodstuffs are a worldwide problem. As future source of protein for human nutrition sunflower seed and oil extraction residues from sunflower seed, respectively, are of great significance. Sunflower seed does not contain anti-nutritive and toxical compounds. After crossing of species having a high oil content, the today cultivated sunflower hybrids bring seeds containing 17-22% crude protein and 30-52% oil. The cultivation also has led to a considerable reduction of the hull content. In processing of sunflower proteins colour problems occur resulting from finely ground particles of dark hulls and from polyphenolic acids which are easily oxidized and converted into brown polymerics. Essential components of the sunflower protein production are, therefore, the at least 98% dehulling before processing as well as the separation of polyphenolic acids and/or the prevention of their oxidation. In principle, the variation and combination of technological steps in pre-treating and defatting of sunflower seed, in extracting, precipitating, washing and drying of proteins, the chemical modification of proteins obtained, the interaction with neutral salts or complexing agents, and the admixture of lysine or proteins of high lysine content allow to adapt sunflower proteins to each type of application.

  2. [Construction of cTnC-linker-TnI (P) Genes, Expression of Fusion Protein and Preparation of Lyophilized Protein].

    PubMed

    Song, Xiaoli; Liu, Xiaoyun; Cai, Lei; Wu, Jianwei; Wang, Jihua

    2015-12-01

    In order to construct and express human cardiac troponin C-linker-troponin I(P) [ cTnC-linker-TnI(P)] fusion protein, detect its activity and prepare lyophilized protein, we searched the CDs of human cTnC and cTnI from GenBank, synthesized cTnC and cTnI(30-110aa) into cloning vector by a short DNA sequence coding for 15 neutral amino acid residues. pCold I-cTnC-linker-TnI(P) was constructed and transformed into E. coli BL21(DE3). Then, cTnC-linker-TnI(P) fusion protein was induced by isopropyl-β-D-thiogalactopyranoside (IPTG). Soluable expression of cTnC-linker-TnI(P) in prokaryotic system was successfully obtained. The fusion protein was purified by Ni²⁺ Sepharose 6 Fast Flow affinity chromatography with over 95% purity and prepared into lyophilized protein. The activity of purified cTnC-linker-TnI(P) and its lyophilized protein were detected by Wondfo Finecare™ cTnI Test. Lyophilized protein of cTnC-linker-TnI(P) was stable for 10 or more days at 37 °C and 4 or more months at 25 °C and 4 °C. The expression system established in this research is feasible and efficient. Lyophilized protein is stable enough to be provided as biological raw materials for further research. PMID:27079099

  3. Preparative purification of antibodies with protein A—an alternative to conventional chromatography

    NASA Astrophysics Data System (ADS)

    Holschuh, Karl; Schwämmle, Achim

    2005-05-01

    Protein A coated magnetic particles are used for the preparative purification of antibodies from up to 100 L cell culture supernatant. The comparison with conventional column and expanded bed chromatography results in similar yield and purity of the product but much faster separation times.

  4. Wheat gluten hydrolysate affects race performance in the triathlon.

    PubMed

    Koikawa, Natsue; Aoki, Emi; Suzuki, Yoshio; Sakuraba, Keishoku; Nagaoka, Isao; Aoki, Kazuhiro; Shimmura, Yuki; Sawaki, Keisuke

    2013-07-01

    Wheat gluten hydrolysate (WGH) is a food ingredient, prepared by partial enzymatic digestion of wheat gluten, which has been reported to suppress exercise-induced elevation of serum creatinine kinase (CK) activity. However, its effects on athletic performance have not yet been elucidated. This is the presentation of an experiment performed on five female college triathletes who completed an Olympic distance triathlon with or without ingestion of 21 g of WGH during the cycling leg. The experiment was performed in a crossover double-blind manner. The race time of the running leg and thus the total race time was significantly shorter when WGH was ingested. However, serum CK levels exhibited no apparent differences between the two WGH or placebo groups.

  5. Effects and mechanism of dual-frequency power ultrasound on the molecular weight distribution of corn gluten meal hydrolysates.

    PubMed

    Jin, Jian; Ma, Haile; Wang, Bei; Yagoub, Abu El-Gasim A; Wang, Kai; He, Ronghai; Zhou, Cunshan

    2016-05-01

    The impact of dual-frequency power ultrasound (DPU) on the molecular weight distribution (MWD) of corn gluten meal (CGM) hydrolysates and its mechanism were investigated in the present study. The mechanism was studied from aspects of structural and nano-mechanical characteristics of the major protein fractions of CGM, viz. zein and glutelin. The results of molecular weight distribution indicated that DPU pretreatment of CGM was beneficial to the preparation of peptides with molecular weights of 200-1000Da. Moreover, FTIR spectral analysis and atomic force microscopy characterization showed that the DPU pretreatment changed the contents of secondary structure of proteins, decreased the particle height and surface roughness of glutelin, reduced the Young's modulus and stiffness of zein while increased its adhesion force. In conclusion, DPU pretreatment of proteins before proteolysis is an efficient alternative method to produce short-chain peptides because of its positive effects originating from acoustic cavitation on the molecular conformation, nano-structures and nano-mechanical properties of proteins as well.

  6. Effects and mechanism of dual-frequency power ultrasound on the molecular weight distribution of corn gluten meal hydrolysates.

    PubMed

    Jin, Jian; Ma, Haile; Wang, Bei; Yagoub, Abu El-Gasim A; Wang, Kai; He, Ronghai; Zhou, Cunshan

    2016-05-01

    The impact of dual-frequency power ultrasound (DPU) on the molecular weight distribution (MWD) of corn gluten meal (CGM) hydrolysates and its mechanism were investigated in the present study. The mechanism was studied from aspects of structural and nano-mechanical characteristics of the major protein fractions of CGM, viz. zein and glutelin. The results of molecular weight distribution indicated that DPU pretreatment of CGM was beneficial to the preparation of peptides with molecular weights of 200-1000Da. Moreover, FTIR spectral analysis and atomic force microscopy characterization showed that the DPU pretreatment changed the contents of secondary structure of proteins, decreased the particle height and surface roughness of glutelin, reduced the Young's modulus and stiffness of zein while increased its adhesion force. In conclusion, DPU pretreatment of proteins before proteolysis is an efficient alternative method to produce short-chain peptides because of its positive effects originating from acoustic cavitation on the molecular conformation, nano-structures and nano-mechanical properties of proteins as well. PMID:26703201

  7. Endotoxin depletion of recombinant protein preparations through their preferential binding to histidine tags.

    PubMed

    Mack, Laura; Brill, Boris; Delis, Natalia; Groner, Bernd

    2014-12-01

    The presence of endotoxins in preparations of recombinantly produced therapeutic proteins poses serious problems for patients. Endotoxins can cause fever, respiratory distress syndromes, intravascular coagulation, or endotoxic shock. A number of methods have been devised to remove endotoxins from protein preparations using separation procedures based on molecular mass or charge properties. Most of the methods are limited in their endotoxin removal capacities and lack general applicability. We are describing a biotechnological approach for endotoxin removal. This strategy exploits the observation that endotoxins form micelles that expose negative charges on their surface, leading to preferential binding of endotoxins to cationic surfaces, allowing the separation from their resident protein. Endotoxins exhibit high affinity to stretches of histidines, which are widely used tools to facilitate the purification of recombinant proteins. They bind to nickel ions and are the basis for protein purification from cellular extracts by immobilized metal affinity chromatography. We show that the thrombin-mediated cleavage of two histidine tags from the purified recombinant protein and the adsorption of these histidine tags and their associated endotoxins to a nickel affinity column result in an appreciable depletion of the endotoxins in the purified protein fraction.

  8. Chemical and cellular antioxidative properties of threadfin bream (Nemipterus spp.) surimi byproduct hydrolysates fractionated by ultrafiltration.

    PubMed

    Wiriyaphan, Chompoonuch; Xiao, Hang; Decker, Eric A; Yongsawatdigul, Jirawat

    2015-01-15

    Protein hydrolysate from frame, bone and skin (FBSH) of threadfin bream was prepared using Virgibacillus sp. SK33 proteinase and fractionated using sequential ultrafiltration membranes with molecular weight cut-offs (MWCO) of 30, 5 and 1 kDa, respectively. Four fractions, namely FBSH-I (>30 kDa), FBSH-II (5-30 kDa), FBSH-III (1-5 kDa), and FBSH-IV (<1 kDa), were obtained. All fractions were rich in Lys, Glu/Gln, Gly, Pro, Ala, Asp/Asn, and Arg. FBSH-III and FBSH-IV showed the highest surface hydrophobicity measured by 8-anilino-1-naphthalenesulfonic acid (ANS) probe (p<0.05). FBSH-III showed the highest antioxidant activity and cytoprotective effects against tert-butyl hydroperoxide (TBHP)-induced cytotoxicity of Caco-2 cells. In addition, FBSH-III inhibited lactate dehydrogenase (LDH) leakage and intracellular reactive species (ROS) production in a dose-dependent manner. FBSH-III retained antioxidant activity and cytoprotective capacity after in vitro simulated gastrointestinal digestion. These results suggested that FBSH-III might potentially be nutraceutical peptides with antioxidative properties. PMID:25148952

  9. Human protein C: new preparations. Effective replacement therapy for some clotting disorders.

    PubMed

    2003-02-01

    (1) Depending on its severity, congenital protein C deficiency can cause a variety of problems, such as increasing the frequency of venous thrombosis in high risk situations; recurrent venous thrombosis; skin necrosis at the start of treatment with a vitamin K antagonist; and severe thrombotic events in neonates. For many years the only available replacement treatment consisted of fresh frozen plasma which, among other adverse effects, carries a risk of hypervolemia. (2) Two human protein C concentrates prepared from donated blood have been given marketing authorisation in Europe for intravenous replacement therapy (Ceprotin from Baxter, and Protexel from LFB). (3) Their clinical files contain only retrospective case series (22 children with severe deficiency treated with Ceprotin; and 10 patients of various ages and with different degrees of severity treated with Protexel). The two preparations have not been compared with each other. (4) In patients with severe protein C deficiency, including neonates, replacement therapy with human protein C is effective, especially for treating cutaneous thrombosis and preventing thrombosis in high risk situations. (5) In patients with moderate deficiency, a short-course of human protein C prophylaxis reduces the frequency of thrombosis in high risk situations. (6) In long-term prophylaxis, human protein C replacement therapy, added to ongoing (but inadequately effective) vitamin K antagonist therapy, seems to reduce the risk of recurrent venous thrombosis even though it has some constraints. (7) The adverse effects of the two preparations are poorly documented. Allergic reactions and bleeding have been reported. Human protein C is a blood product, and therefore carries a risk of infection. (8) Ceprotin offers a small advantage, being available in two dose strengths: for a given dose the volume injected is halved. (9) In practice, Ceprotin and Protexel are the reference drugs for replacement therapy of constitutional protein C

  10. Hydrogen production from paper sludge hydrolysate.

    PubMed

    Kádár, Zsófia; De Vrije, Truus; Budde, Miriam A W; Szengyel, Zsolt; Réczey, Kati; Claassen, Pieternel A M

    2003-01-01

    The main objective of this study was to develop a system for the production of "renewable" hydrogen. Paper sludge is a solid industrial waste yielding mainly cellulose, which can be used, after hydrolysis, as a feedstock in anaerobic fermentation by (hyper)thermophilic organisms, such as Thermotoga elfii and Caldicellulosiruptor saccharolyticus. Tests on different medium compositions showed that both bacteria were able to produce hydrogen from paper sludge hydrolysate, but the amount of produced hydrogen and the requirement for other components differed. Hydrogen production by T. elfii strongly depended on the presence of yeast extract and salts. By contrast, C. saccharolyticus was less dependent on medium components but seemed to be inhibited by a component present in the sludge hydrolysate. Utilization of xylose was preferred over glucose by C. saccharolyticus.

  11. Calcium pectinate gel bead intended for oral protein delivery: preparation improvement and formulation development.

    PubMed

    Si, Luqin; Zhao, Ying; Huang, Jiangeng; Li, Sha; Zhai, Xuezhen; Li, Gao

    2009-07-01

    Calcium pectinate gel (CPG) micrometer-sized beads (microbeads) containing insulin, as a model amphoteric protein, were prepared by ionotropic gelation technique together with an air compressor. The influences of phosphate buffer, pH as well as calcium and pectin concentrations of cross-linking solution on the characteristics and release profiles of microbeads were investigated. With the aid of compressed air flow, the mean diameters of beads were successfully decreased to micron-sized. The results showed that all the factors investigated greatly affected the entrapment efficiencies and release profiles of the microbeads. Suitable formulation concentrations should be considered and great care should be taken to maintain the pH of working solutions at or close to isoelectric point of protein loaded during the whole preparation process. Hence, CPG microbeads of perfect spherical shape, uniform sizes, enhanced mechanical strength, good entrapment efficiencies and delayed release profiles were prepared for a load of amphoteric protein and peptide drugs, without any use of organic solvents or harsh ingredients. Therefore, CPG microbeads could be a promising carrier for oral controlled-release systems of amphoteric protein and peptide drugs.

  12. Preparation and Extraction of Insoluble (Inclusion-Body) Proteins from Escherichia coli

    PubMed Central

    Palmer, Ira; Wingfield, Paul T.

    2013-01-01

    High-level expression of many recombinant proteins in Escherichia coli leads to the formation of highly aggregated protein commonly referred to as inclusion bodies. Inclusion bodies are normally formed in the cytoplasm; however, if a secretion vector is used, they can form in the periplasmic space. Inclusion bodies can be recovered from cell lysates by low speed centrifugation. Following preextaction (or washing) protein is extracted from washed pellets using guanidine·HCl. The solubilized and unfolded protein is either directly folded as described in UNIT 6.1 or further purified by gel filtration in the presence of guanidine·HCl as described here. A support protocol describes the removal of guanidine·HCl from column fractions so they can be monitored by SDS-PAGE. High-level expression of many recombinant proteins in Escherichia coli leads to the formation of highly aggregated protein commonly referred to as inclusion bodies (UNITS 5.1 & 6.1). Inclusion bodies are normally formed in the cytoplasm; alternatively, if a secretion vector is used, they can form in the periplasmic space. Inclusion bodies are not restricted to E. coli; they can also form in yeast, mammalian, and insect cells. Inclusion bodies recovered from cell lysates by low-speed centrifugation are heavily contaminated with E. coli cell wall and outer membrane components. The latter are largely removed by selective extraction with detergents and low concentrations of either urea or guanidine·HCl to produce so-called washed pellets. These basic steps result in a significant purification of the recombinant protein, which usually makes up ~60% of the washed pellet protein. The challenge, therefore, is not to purify the recombinant-derived protein, but to solubilize it and then fold it into native and biologically active protein. Basic Protocol 1 describes preparation of washed pellets and solubilization of the protein using guanidine·HCl. The extracted protein, which is unfolded, is either directly

  13. Improved characterization of EV preparations based on protein to lipid ratio and lipid properties.

    PubMed

    Osteikoetxea, Xabier; Balogh, Andrea; Szabó-Taylor, Katalin; Németh, Andrea; Szabó, Tamás Géza; Pálóczi, Krisztina; Sódar, Barbara; Kittel, Ágnes; György, Bence; Pállinger, Éva; Matkó, János; Buzás, Edit Irén

    2015-01-01

    In recent years the study of extracellular vesicles has gathered much scientific and clinical interest. As the field is expanding, it is becoming clear that better methods for characterization and quantification of extracellular vesicles as well as better standards to compare studies are warranted. The goal of the present work was to find improved parameters to characterize extracellular vesicle preparations. Here we introduce a simple 96 well plate-based total lipid assay for determination of lipid content and protein to lipid ratios of extracellular vesicle preparations from various myeloid and lymphoid cell lines as well as blood plasma. These preparations included apoptotic bodies, microvesicles/microparticles, and exosomes isolated by size-based fractionation. We also investigated lipid bilayer order of extracellular vesicle subpopulations using Di-4-ANEPPDHQ lipid probe, and lipid composition using affinity reagents to clustered cholesterol (monoclonal anti-cholesterol antibody) and ganglioside GM1 (cholera toxin subunit B). We have consistently found different protein to lipid ratios characteristic for the investigated extracellular vesicle subpopulations which were substantially altered in the case of vesicular damage or protein contamination. Spectral ratiometric imaging and flow cytometric analysis also revealed marked differences between the various vesicle populations in their lipid order and their clustered membrane cholesterol and GM1 content. Our study introduces for the first time a simple and readily available lipid assay to complement the widely used protein assays in order to better characterize extracellular vesicle preparations. Besides differentiating extracellular vesicle subpopulations, the novel parameters introduced in this work (protein to lipid ratio, lipid bilayer order, and lipid composition), may prove useful for quality control of extracellular vesicle related basic and clinical studies.

  14. [Protein assay by the modified Dumas method applied to preparations of plasma proteins].

    PubMed

    Blondel, P; Vian, L

    1993-01-01

    Quantify protein according Pharmacopoeia method, based on Kjeldahl method, needs a long time to do. The development of an automaton which used the modified Dumas method divide the analysis time by 15 (6 minutes versus over 90 minutes). The results show no statistical differences between official method and this one. PMID:8154798

  15. Preparation of phytantriol cubosomes by solvent precursor dilution for the delivery of protein vaccines.

    PubMed

    Rizwan, S B; Assmus, D; Boehnke, A; Hanley, T; Boyd, B J; Rades, T; Hook, S

    2011-09-01

    Different delivery strategies to improve the immunogenicity of peptide/protein-based vaccines are currently under investigation. In this study, the preparation and physicochemical characterisation of cubosomes, a novel lipid-based particulate system currently being explored for vaccine delivery, was investigated. Cubosomes were prepared from a liquid precursor mixture containing phytantriol or glycerylmonooleate (GMO), F127 for particle stabilisation, and a hydrotrope (ethanol or polyethylene glycol (PEG(200)) or propylene glycol (PG)). Several liquid precursors were prepared, and the effect of varying the concentrations of F127 and the hydrotrope on cubosome formation was investigated. Formulations were prepared by fragmentation for comparison. The model protein ovalbumin (Ova) was also entrapped within selected formulations. Submicron-sized particles (180-300 nm) were formed spontaneously upon dilution of the liquid precursors, circumventing the need for the preformed cubic phase used in traditional fragmentation-based methods. The nanostructure of the phytantriol dispersions was determined to be cubic phase using SAXS whilst GMO dispersions had a reverse hexagonal nanostructure coexisting with cubic phase. The greatest entrapment of Ova was within phytantriol cubosomes prepared from liquid precursors. Release of Ova from the various formulations was sustained; however, release was significantly faster and the extent of release was greater from fragmented dispersions compared to liquid precursor formulations. Taken together, these results suggest that phytantriol cubosomes can be prepared using liquid precursors and that it is a suitable alternative to GMO. Furthermore, the high entrapment and the slow release of Ova in vitro highlight the potential of phytantriol cubosomes prepared using liquid precursors as a novel vaccine delivery system.

  16. COMPARATIVE PROFILING OF HIGHLY ENRICHED 22L AND CHANDLER MOUSE SCRAPIE PRION PROTEIN PREPARATIONS

    PubMed Central

    Moore, Roger A.; Timmes, Andrew; Wilmarth, Phillip A.; Priola, Suzette A.

    2013-01-01

    Transmissible spongiform encephalopathies (TSEs) or prion diseases are characterized by the accumulation of an aggregated isoform of the prion protein (PrP). This pathological isoform, termed PrPSc, appears to be the primary component of the TSE infectious agent or prion. However, it is not clear to what extent other protein co-factors may be involved in TSE pathogenesis or whether there are PrPSc-associated proteins which help to determine TSE strain-specific disease phenotypes. We enriched PrPSc from the brains of mice infected with either 22L or Chandler TSE strains and examined the protein content of these samples using nanospray liquid chromatography-tandem mass spectrometry. These samples were compared to “mock” PrPSc preparations from uninfected brains. Prion protein was the major component of the infected samples and ferritin was the most abundant impurity. By contrast, mock enrichments contained no detectable prion protein but did contain a significant amount of ferritin. Of the total proteins identified, 32% were found in both mock and infected samples. The similarities between PrPSc samples from 22L and Chandler TSE strains suggest that the non-PrPSc protein components found in standard enrichment protocols are not strain-specific. PMID:20518029

  17. Expression, purification and antibody preparation of PCV2 Rep and ORF3 proteins.

    PubMed

    Peng, Zhiyuan; Ma, Teng; Pang, Daxin; Su, Dan; Chen, Fuwang; Chen, Xinrong; Guo, Ning; Ouyang, Ting; Ouyang, Hongsheng; Ren, Linzhu

    2016-05-01

    Rep and ORF3 proteins are important functional proteins of porcine circovirus 2 (PCV2). Here, Rep and ORF3 genes were cloned, expressed and used to raise polyclonal antibodies. The result showed the recombinant plasmids of Rep and ORF3 genes constructed in this study were expressed efficiently in the prokaryotic system, and the recombinant proteins had antigenicity and immunogenicity. Furthermore, reactivity and specificity of the antiserums were characterized by western blot and indirect immunofluorescent assays. The results elucidated that polyclonal antiserum prepared with Rep or ORF3 had good reactivity and specificity against PCV2, or the Rep and ORF3 expressed in PK-15 cells, respectively. The Rep protein is promising for PCV2 antibody and vaccine development. These results will be helpful for further studies focusing on pathogenesis of PCV2 and serology diagnostic test or vaccine development against PCV2. PMID:26812108

  18. Step-by-step preparation of proteins for mass spectrometric analysis.

    PubMed

    Franz, Thomas; Li, Xinping

    2015-01-01

    Nowadays, identification of proteins from biological samples by mass spectrometry is widely used. In principle there are two scenarios. Proteins are pre-fractionated in some way, e.g. by gel electrophoresis or are analyzed as complex mixture (shot gun). Shot gun proteomics became recently more popular, because of technological developments on the mass spectrometer side which allows now the identification of several thousand proteins from complex biological matrix. However, in many cases it is still useful to separate proteins first in a gel. But not only mass spectrometer technology made progress. This is also true for the sample preparation. Recently, protocols and techniques were developed which make the analysis of starting material in the low microgram range possible and also simplify the whole procedure. Detailed protocols will be described allowing also inexperienced beginners to get good results.

  19. Biodegradable polymersomes with an ionizable membrane: facile preparation, superior protein loading, and endosomal pH-responsive protein release.

    PubMed

    Li, Shaoke; Meng, Fenghua; Wang, Zhongjuan; Zhong, Yinan; Zheng, Meng; Liu, Haiyan; Zhong, Zhiyuan

    2012-09-01

    Novel biodegradable polymersomes containing an ionizable membrane were developed for efficient loading and rapid intracellular release of proteins. The polymersomes were prepared from poly(ethylene glycol)-b-poly(trimethylene carbonate) (PEG-PTMC) block copolymer derivatives containing acrylate, carboxylic acid, and amine groups along PTMC block, which are denoted as PEG-PTMC(AC), PEG-PTMC(COOH), and PEG-PTMC(NH(2)), respectively. Notably, nano-sized polymersomes (95.1-111.6nm) were formed by directly dispersing these copolymers in phosphate buffer at room temperature. Both FITC-labeled bovine serum albumin (FITC-BSA) and cytochrome C (FITC-CC) were readily loaded into PEG-PTMC(COOH) and PEG-PTMC(NH(2)) polymersomes with remarkably high loading levels. Interestingly, in vitro release studies showed that PEG-PTMC(COOH) and PEG-PTMC(NH(2)) polymersomes had pH-responsive protein release behaviors in which significantly faster protein release was observed at endosomal pH than at physiological pH. MTT assays indicated that these polymersomes had low cytotoxicity. Furthermore, confocal laser scanning microscope (CLSM) observations revealed that FITC-CC loaded polymersomes efficiently delivered proteins into MCF-7 cells following 24h incubation. Importantly, flow cytometry showed that CC-loaded polymersomes induced markedly enhanced apoptosis in MCF-7 cells as compared to free CC. These novel membrane ionizable biodegradable polymersomes have appeared as highly promising nanocarriers for efficient intracellular protein delivery.

  20. [Preparation and detection of anti-influenza A virus polymerase basic protein 1 polyclonal antibody].

    PubMed

    Qin, Yujie; Zhang, Tinghong; Ye, Xin

    2016-01-01

    Influenza A virus is an enveloped virus that belongs to the Orthomyxoviridae family. It has 8 negative RNA segments that encode 16 viral proteins. The viral polymerase consists of 3 proteins (PB 1, PB2 and PA) which plays an important role in the transcription and replication of the influenza A virus. Polymerase basic protein 1 (PB 1) is a critical member of viral polymerase complex. In order to further study the function of PB1, we need to prepare the PB1 antibody with good quality. Therefore, we amplified PB1 conserved region (nt1648-2265) by PCR and cloned it into pET-30a vector, and transformed into Escherichia coli BL2 1. The expression of His tagged PB 1 protein was induced by IPTG, and His-PB 1 proteins were purified by Ni-NTA resin. For preparation of PB 1 protein antiserum, rabbits were immunized with His-PB 1 fusion protein 3 times. Then the titer of PB 1 polyclonal antibody was measured by indirect ELISA. The antibody was purified by membrane affinity purification and subjected to immunoblotting analysis. Data showed that PB1 antibody can recognize PB 1 protein from WSN virus infected or pCMV FLAG-PB 1 transfected cells. Meanwhile, PB 1 antibody can also recognize specifically other subtype strains of influenza A virus such as H9N2 and H3N2. PB 1 polyclonal antibody we generated will be a useful tool to study the biological function of PB1. PMID:27363203

  1. Infant food from quality protein maize and chickpea: optimization for preparing and nutritional properties.

    PubMed

    Alarcón-Valdez, C; Milán-Carrillo, J; Cárdenas-Valenzuela, O G; Mora-Escobedo, R; Bello-Pérez, L A; Reyes-Moreno, C

    2005-06-01

    The present study had two objectives: to determine the best combination of nixtamalized maize flour (NMF) from quality protein maize and extruded chickpea flour (ECF) for producing an infant food, and to evaluate the nutritional properties of the optimized NMF/ECF mixture and the infant food. Response surface methodology (RSM) was applied to determine the best combination of NMF/ECF; the experimental design (Lattice simplex) generated 11 assays. Mixtures from each assay were evaluated for true protein and available lysine. Each one of 11 mixtures was used for preparing 11 infant foods that were sensory evaluated for acceptability. A common optimum value for the three response variables was obtained utilizing the desirability method. The best combination of NMF/ECF for producing an infant food was NMF = 26.7%/ECF = 73.3%; this optimized mixture had a global desirability of 0.87; it contained 19.72% dry matter (DM) proteins, 6.10% (DM) lipids, 71.45% (DM) carbohydrates, and 2.83% (DM) minerals; its essential amino acids profile covered the amino acids requirements for children 10-12 years old. The infant food prepared from optimized mixture had an in vitro protein digestibility of 87.9%, and a calculated protein efficiency ratio of 1.86. Infant food could be used to support the growth of infants in developing countries.

  2. Dissecting a complex chemical stress: chemogenomic profiling of plant hydrolysates

    PubMed Central

    Skerker, Jeffrey M; Leon, Dacia; Price, Morgan N; Mar, Jordan S; Tarjan, Daniel R; Wetmore, Kelly M; Deutschbauer, Adam M; Baumohl, Jason K; Bauer, Stefan; Ibáñez, Ana B; Mitchell, Valerie D; Wu, Cindy H; Hu, Ping; Hazen, Terry; Arkin, Adam P

    2013-01-01

    The efficient production of biofuels from cellulosic feedstocks will require the efficient fermentation of the sugars in hydrolyzed plant material. Unfortunately, plant hydrolysates also contain many compounds that inhibit microbial growth and fermentation. We used DNA-barcoded mutant libraries to identify genes that are important for hydrolysate tolerance in both Zymomonas mobilis (44 genes) and Saccharomyces cerevisiae (99 genes). Overexpression of a Z. mobilis tolerance gene of unknown function (ZMO1875) improved its specific ethanol productivity 2.4-fold in the presence of miscanthus hydrolysate. However, a mixture of 37 hydrolysate-derived inhibitors was not sufficient to explain the fitness profile of plant hydrolysate. To deconstruct the fitness profile of hydrolysate, we profiled the 37 inhibitors against a library of Z. mobilis mutants and we modeled fitness in hydrolysate as a mixture of fitness in its components. By examining outliers in this model, we identified methylglyoxal as a previously unknown component of hydrolysate. Our work provides a general strategy to dissect how microbes respond to a complex chemical stress and should enable further engineering of hydrolysate tolerance. PMID:23774757

  3. Collagen hydrolysate based collagen/hydroxyapatite composite materials

    NASA Astrophysics Data System (ADS)

    Ficai, Anton; Albu, Madalina Georgiana; Birsan, Mihaela; Sonmez, Maria; Ficai, Denisa; Trandafir, Viorica; Andronescu, Ecaterina

    2013-04-01

    The aim of this study was to study the influence of collagen hydrolysate (HAS) on the formation of ternary collagen-hydrolysate/hydroxyapatite composite materials (COLL-HAS/HA). During the precipitation process of HA, a large amount of brushite is resulted at pH = 7 but, practically pure HA is obtained at pH ⩾ 8. The FTIR data reveal the duplication of the most important collagen absorption bands due to the presence of the collagen hydrolysate. The presence of collagen hydrolysate is beneficial for the management of bone and joint disorders such as osteoarthritis and osteoporosis.

  4. Virion proteins and the perspectives of gene manipulations in vaccine preparation.

    PubMed

    Tickhonenko, T I

    1985-05-01

    The achievements and perspectives of genetic manipulations are described aiming at preparation of first generation subunit vaccines based on the synthesis in bacterial and eukaryotic cells of full-sized virion proteins expressing the main antigenic determinants. The preparation of such vaccines in bacterial cells seems out of perspective in the case of influenza, human hepatitis B, foot- and - mouth disease and some other viruses due to the peculiarities of structure and synthesis as well as low immunogenicity of the monomeric form of virion polypeptides. However, biotechnological procedures using eukaryotic cells and higher eukaryotic vectors and in part also yeast cells allowed to obtain full-sized virion proteins in a highly immunogenic state with good effects.

  5. Optimization of proteomic sample preparation procedures for comprehensive protein characterization of pathogenic systems

    SciTech Connect

    Brewer, Heather M.; Norbeck, Angela D.; Adkins, Joshua N.; Manes, Nathan P.; Ansong, Charles; Shi, Liang; Rikihisa, Yasuko; Kikuchi, Takane; Wong, Scott; Estep, Ryan D.; Heffron, Fred; Pasa-Tolic, Ljiljana; Smith, Richard D.

    2008-12-19

    The elucidation of critical functional pathways employed by pathogens and hosts during an infectious cycle is both challenging and central to our understanding of infectious diseases. In recent years, mass spectrometry-based proteomics has been used as a powerful tool to identify key pathogenesis-related proteins and pathways. Despite the analytical power of mass spectrometry-based technologies, samples must be appropriately prepared to characterize the functions of interest (e.g. host-response to a pathogen or a pathogen-response to a host). The preparation of these protein samples requires multiple decisions about what aspect of infection is being studied, and it may require the isolation of either host and/or pathogen cellular material.

  6. [Preparation of procion brilliant blue-doped silica nanorods and their recognition properties for proteins].

    PubMed

    Deng, Qiliang; Chen, Yang; Wu, Jianhua; Li, Yanli; Liu, Chunjing; Song, Weijing

    2011-09-01

    Protein enrichment and separation is one of the pivotal preliminary steps of proteomics studies, which is important to medical diagnosis and treatment. In this study, procion brilliant blue-doped silica nanorod was prepared via self-assembly sol-gel technology without any additional template. Procion brilliant blue was covalently linked to 3-aminopropyltriethyloxy silane in ethanol. Tetraethylorthosilane (TEOS) was then added into the mixture, subsequently hydrolyzed and co-condensed for 3 h under stirring. The resulted nanorods were isolated by centrifugation, re-dispersed in deionized water, and centrifuged again. This wash process was repeated three times. Finally, the nanorods were dried under vacuum. Procion brilliant blue acted simultaneously as a self-assembly template during the preparation process, and subsequently as recognition probe for proteins. Scanning electron microscopy (SEM) image showed that the nanotubes were 2-16 microm in length and 200-500 nm in diameter. The obtained nanorods were further characterized by Fourier transform infrared spectroscopy (FTIR), thermogravimetric analysis (TGA) and differential scanning calorimetry (DSA), separately. All these results indicated that procion brilliant blue were successfully doped into silica nanorods. The recognition property of nanorods for bovine serum albumin (BSA) was investigated under static condition. The resulted nanorods showed high binding capacity (57.6 mg/g) for BSA and fast adsorption equilibrium (within 60 min). The nanorods were also evaluated with four typical proteins, hemoglobin, trypsin, lysozyme and pepsin, with different relative molecular masses and isoelectric points. The results indicated that the prepared nanorods exhibited the highest binding capacity for trypsin (87.5 mg/g) and the least binding for hemoglobin (Hb, 3.0 mg/g). This easy preparation protocol and excellent recognition property make the prepared materials a bright future in proteomics research. PMID:22233075

  7. The preparation and properties of folate-binding protein from cow's milk.

    PubMed Central

    Salter, D N; Scott, K J; Slade, H; Andrews, P

    1981-01-01

    An improved affinity-chromatographic method for the preparation of folate-binding protein from cow's milk is described. Under dissociating conditions the protein appeared homogeneous in the ultracentrifuge, with a molecular weight of 35 000 +/- 1500, but it was heterogeneous on electrophoresis and ion-exchange chromatography and evidently consisted of several glycoproteins with similar molecular weights that all bound folic acid. Overall, the protein contained a high proportion of half-cystine (18 residues/molecule) and 10.3% of carbohydrate. At saturation it bound approx. 1 mol of folate/mol of protein at pH 7.2. Equilibrium-dialysis measurements of the binding of folic acid and 5-methyltetrahydrofolate to the purified protein gave non-linear Scatchard plots, the shapes of which depended on pH. The results were interpreted in terms of ligand binding to a polymerizing system in which the affinity of ligand for monomer was greater than its affinity for polymer. When the protein concentration was similar to that in cow's milk, dissociation constants (Kd) for folate and 5-methyltetrahydrofolate were 3 nM and 5 nM respectively at pH 7.2 and 37 degrees C, whereas Kd for the binding of folate to monomer was about 50 pM. The properties of the binding protein are discussed in relation to its possible role in folate absorption in the gut. PMID:7305943

  8. Preparation of mesoporous silica thin films by photocalcination method and their adsorption abilities for various proteins.

    PubMed

    Kato, Katsuya; Nakamura, Hitomi; Yamauchi, Yoshihiro; Nakanishi, Kazuma; Tomita, Masahiro

    2014-07-01

    Mesoporous silica (MPS) thin film biosensor platforms were established. MPS thin films were prepared from tetraethoxysilane (TEOS) via using sol-gel and spin-coating methods using a poly-(ethylene oxide)-block-poly-(propylene oxide)-block-poly-(ethylene oxide) triblock polymer, such as P123 ((EO)20(PO)70(EO)20) or F127 ((EO)106(PO)70(EO)106), as the structure-directing agent. The MPS thin film prepared using P123 as the mesoporous template and treated via vacuum ultraviolet (VUV) irradiation to remove the triblock copolymer had a more uniform pore array than that of the corresponding film prepared via thermal treatment. Protein adsorption and enzyme-linked immunosorbent assay (ELISA) on the synthesized MPS thin films were also investigated. VUV-irradiated MPS thin films adsorbed a smaller quantity of protein A than the thermally treated films; however, the human immunoglobulin G (IgG) binding efficiency was higher on the former. In addition, protein A-IgG specific binding on MPS thin films was achieved without using a blocking reagent; i.e., nonspecific adsorption was inhibited by the uniform pore arrays of the films. Furthermore, VUV-irradiated MPS thin films exhibited high sensitivity for ELISA testing, and cytochrome c adsorbed on the MPS thin films exhibited high catalytic activity and recyclability. These results suggest that MPS thin films are attractive platforms for the development of novel biosensors. PMID:24857463

  9. Preparation, characterization and in-vitro evaluation of sustained release protein-loaded nanoparticles based on biodegradable polymers

    PubMed Central

    Mukherjee, Biswajit; Santra, Kousik; Pattnaik, Gurudutta; Ghosh, Soma

    2008-01-01

    Controlled drug delivery technology of proteins/peptides from biodegradable nanoparticles has emerged as one of the eminent areas to overcome formulation associated problems of the macromolecules. The purpose of the present investigation was to develop protein-loaded nanoparticles using biodegradable polymer poly l-lactide-co-glycolidic acid (PLGA) with bovine serum albumin (BSA) as a model protein. Despite many studies available with PLGA-based protein-loaded nanoparticles, production know-how, process parameters, protein loading, duration of protein release, narrowing polydispersity of particles have not been investigated enough to scale up manufacturing of protein-loaded nanoparticles in formulations. Different process parameters such as protein/polymer ratio, homogenizing speed during emulsifications, particle surface morphology and surface charges, particle size analysis and in-vitro protein release were investigated. The in-vitro protein release study suggests that release profile of BSA from nanoparticles could be modulated by changing protein-polymer ratios and/or by varying homogenizing speed during multiple-emulsion preparation technique. The formulation prepared with protein-polymer ratio of 1:60 at 17,500 rpm gave maximum protein-loading, minimum polydispersion with maximally sustained protein release pattern, among the prepared formulations. Decreased (10,000 rpm) or enhanced (24,000 rpm) homogenizing speeds resulted in increased polydispersion with larger particles having no better protein-loading and -release profiles in the present study. PMID:19337417

  10. Ex vivo digestion of carp muscle tissue--ACE inhibitory and antioxidant activities of the obtained hydrolysates.

    PubMed

    Borawska, J; Darewicz, M; Vegarud, G E; Iwaniak, A; Minkiewicz, P

    2015-01-01

    In the digestive tract of humans, bioactive peptides, i.e. protein fragments impacting the physiological activity of the body, may be released during the digestion of food proteins, including those of fish. The aim of the study was to establish the method of human ex vivo digestion of carp muscle tissue and evaluate the angiotensin I-converting enzyme inhibitory and antioxidant activities of hydrolysates obtained after digestion. It was found that the hydrolysates of carp muscle tissue obtained with the three-stage method of simulated ex vivo digestion showed ACE inhibitory as well as antioxidative activities. It was demonstrated that the degree of hydrolysis depended on the duration of individual stages and the degree of comminution of the examined material. Although the applied gastric juices initiated the process of hydrolysis of carp muscle tissue, the duodenal juices caused a rapid increase in the amount of hydrolysed polypeptide bonds. The antihypertensive and antioxidative activities of the hydrolysates of carp muscle tissue increased together with progressive protein degradation. However, the high degree of protein hydrolysis does not favour an increase in the activity of free radical scavenging. The presented results are an example of the first preliminary screening of the potential health-promoting biological activity of carp muscle tissue in an ex vivo study.

  11. Semiautomated Sample Preparation for Protein Stability and Formulation Screening via Buffer Exchange.

    PubMed

    Ying, William; Levons, Jaquan K; Carney, Andrea; Gandhi, Rajesh; Vydra, Vicky; Rubin, A Erik

    2016-06-01

    A novel semiautomated buffer exchange process workflow was developed to enable efficient early protein formulation screening. An antibody fragment protein, BMSdab, was used to demonstrate the workflow. The process afforded 60% to 80% cycle time and scientist time savings and significant material efficiencies. These efficiencies ultimately facilitated execution of this stability work earlier in the drug development process, allowing this tool to inform the developability of potential candidates for development from a formulation perspective. To overcome the key technical challenges, the protein solution was buffer-exchanged by centrifuge filtration into formulations for stability screening in a 96-well plate with an ultrafiltration membrane, leveraging automated liquid handling and acoustic volume measurements to allow several cycles of exchanges. The formulations were transferred into a vacuum manifold and sterile filtered into a rack holding 96 glass vials. The vials were sealed with a capmat of individual caps and placed in stability stations. Stability of the samples prepared by this process and by the standard process was demonstrated to be comparable. This process enabled screening a number of formulations of a protein at an early pharmaceutical development stage with a short sample preparation time.

  12. Preparation of λN-GST fusion protein for affinity immobilization of RNA.

    PubMed

    Di Tomasso, Geneviève; Lampron, Philipe; Omichinski, James G; Legault, Pascale

    2012-01-01

    Affinity purification of in vitro transcribed RNA is becoming an attractive alternative to purification using standard denaturing gel electrophoresis. Affinity purification is particularly advantageous because it can be performed in a few hours under non-denaturing conditions. However, the performance of affinity purification methods can vary tremendously depending on the RNA immobilization matrix. It was previously shown that RNA immobilization via an optimized λN-GST fusion protein bound to glutathione-Sepharose resin allows affinity purification of RNA with very high purity and yield. This Chapter outlines the experimental procedure employed to prepare the λN-GST fusion protein used for RNA immobilization in successful affinity purifications of RNA. It describes the details of protein expression and purification as well as routine quality control analyses. PMID:23065558

  13. The comparison of protein-entrapped liposomes and lipoparticles: preparation, characterization, and efficacy of cellular uptake

    PubMed Central

    Chang, Wei-Kuo; Tai, Yu-Ju; Chiang, Chiao-Hsi; Hu, Chieh-Shen; Hong, Po-Da; Yeh, Ming-Kung

    2011-01-01

    Fluorescein isothiocyanate-conjugated bovine serum albumin (FITC-BSA)-loaded polyethylene glycol (PEG)-modified liposomes and lipoparticles with high protein entrapment were developed. The lipid formula of the liposomes contained PEGylated lipids and unsaturated fatty acids for enhancing membrane fluidity and effective delivery into cells. The preparation techniques, lipid content, and PEG-modified lipoparticle ratios were evaluated. The PEG-modified lipoparticles prepared by ethanol injection extrusion (100 nm pore size) achieve a population of blank liposomes with a mean size of 125 ± 2.3 nm and a zeta potential of −12.4 ± 1.5 mV. The average particle size of the PEG-modified lipoparticles was 133.7 ± 8.6 nm with a zeta potential of +13.3 mV. Lipoparticle conformation was determined using transmission electron microscopy and field-emission scanning electron microscopy. The FITC-BSA encapsulation efficiency was dramatically increased from 19.0% for liposomes to 59.7% for lipoparticles. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) results confirmed the preparation process, and an 8-hour leaching test did not harm the protein structure. Once prepared, the physical and chemical stability of the PEG-modified lipoparticle formulations was satisfactory over 90 days. In vitro retention tests indicated that the 50% retention time for the protein-containing lipoparticles was 7.9 hours, substantially longer than the liposomes at 3.3 hours. A Caco-2 cell model was used for evaluating the cytotoxicity and cell uptake efficiency of the PEG-modified lipoparticles. At a lipid content below 0.25 mM, neither the liposomes nor the lipoparticles caused significant cellular cytotoxicity (P < 0.01) and FITC-BSA was significantly taken up into cells within 60 minutes (P < 0.01). PMID:22072876

  14. Feather hydrolysate from Streptomyces sampsonii GS 1322: A potential low cost soil amendment.

    PubMed

    Jain, Richa; Jain, Aakanchha; Rawat, Neha; Nair, Meera; Gumashta, Raghvendra

    2016-06-01

    Process parameters for obtaining hydrolysate from hen feathers, i.e., initial pH (5.0-9.0) and incubation period (1-6 day), were set and studied, using Streptomyces sampsonii GS 1322 in submerged and solid state conditions. Under submerged conditions, complete hydrolysis of feathers was observed on fifth day [initial pH 8.0, 28 ± 2°C, shaking (150 rpm)] with release of soluble protein (2985 μg ml(-1)) and amino acids (2407 μg ml(-1)). Cell free hydrolysate showed hydrolysis of casein (18 mm), gelatin (26 mm), collagen (31 mm), hemoglobin (23 mm) and Tween 80 (35 mm) while 445 U keratinase activity. Total soluble protein reached 5 mg ml(-1) in solid state conditions. During Pot experimentation using barren agriculture soil the effect of feather hydrolysate on wheat crop were recorded. Significant increase (p<0.01) in wheat seed germination was observed in treated soils as compared to untreated. Treatment significantly increased plant height from 30 to 60 days and 30-90 days (p<0.001). Treated soil showed an increase in total microbial count, proteolytic activity, phosphate solubilizing bacteria and ammonifying bacteria, whereas pathogenic fungi load was reduced. S. sampsonii GS 1322 can be used for bio-processing of poultry wastes yielding feather hydrolysate rich in proteins and amino acids for development of low-cost organic amendment to accelerate wheat crop growth in barren agricultural land.

  15. Feather hydrolysate from Streptomyces sampsonii GS 1322: A potential low cost soil amendment.

    PubMed

    Jain, Richa; Jain, Aakanchha; Rawat, Neha; Nair, Meera; Gumashta, Raghvendra

    2016-06-01

    Process parameters for obtaining hydrolysate from hen feathers, i.e., initial pH (5.0-9.0) and incubation period (1-6 day), were set and studied, using Streptomyces sampsonii GS 1322 in submerged and solid state conditions. Under submerged conditions, complete hydrolysis of feathers was observed on fifth day [initial pH 8.0, 28 ± 2°C, shaking (150 rpm)] with release of soluble protein (2985 μg ml(-1)) and amino acids (2407 μg ml(-1)). Cell free hydrolysate showed hydrolysis of casein (18 mm), gelatin (26 mm), collagen (31 mm), hemoglobin (23 mm) and Tween 80 (35 mm) while 445 U keratinase activity. Total soluble protein reached 5 mg ml(-1) in solid state conditions. During Pot experimentation using barren agriculture soil the effect of feather hydrolysate on wheat crop were recorded. Significant increase (p<0.01) in wheat seed germination was observed in treated soils as compared to untreated. Treatment significantly increased plant height from 30 to 60 days and 30-90 days (p<0.001). Treated soil showed an increase in total microbial count, proteolytic activity, phosphate solubilizing bacteria and ammonifying bacteria, whereas pathogenic fungi load was reduced. S. sampsonii GS 1322 can be used for bio-processing of poultry wastes yielding feather hydrolysate rich in proteins and amino acids for development of low-cost organic amendment to accelerate wheat crop growth in barren agricultural land. PMID:26906933

  16. Early diagnosis of typhoid fever by an enzyme immunoassay using Salmonella typhi outer membrane protein preparations.

    PubMed

    Verdugo-Rodríguez, A; López-Vidal, Y; Puente, J L; Ruíz-Placios, G M; Calva, E

    1993-04-01

    An enzyme immunoassay (EIA) for detection of serum antibodies in patients with typhoid fever was developed using Salmonella typhi outer membrane protein (OMP) preparations as antigen. Acute phase (first week) sera from adult typhoid fever patients were tested as well as sera from the following control groups: adult travellers with diarrhea caused by enterotoxigenic Escherichia coli, children infected with Campylobacter jejuni, healthy Mexican adult blood donors, and adults with septicemia caused by other organisms. At a 1:3,125 serum dilution, the mean absorbance values were 1.41 in the typhoid fever patients, and 0.57, 0.55, 0.51 and 0.52 in the respective control groups. Inhibition EIA studies using OMP preparations or lipopolysaccharide (LPS) as free antigen indicated that proteins can play an important role in the detection of antibodies in early typhoid fever. This EIA may be useful for the diagnosis of typhoid fever since results were obtained within about five hours and in an endemic area antibodies against Salmonella typhi OMP preparations appear early in the course of the disease.

  17. Pickering emulsions stabilized by whey protein nanoparticles prepared by thermal cross-linking.

    PubMed

    Wu, Jiande; Shi, Mengxuan; Li, Wei; Zhao, Luhai; Wang, Ze; Yan, Xinzhong; Norde, Willem; Li, Yuan

    2015-03-01

    A Pickering (o/w) emulsion was formed and stabilized by whey protein isolate nanoparticles (WPI NPs). Those WPI NPs were prepared by thermal cross-linking of denatured WPI proteins within w/o emulsion droplets at 80°C for 15 min. During heating of w/o emulsions containing 10% (w/v) WPI proteins in the water phase, the emulsions displayed turbid-transparent-turbid phase transitions, which is ascribed to the change in the size of the protein-containing water droplets caused by thermal cross-linking between denatured protein molecules. The transparent stage indicated the formation of WPI NPs. WPI NPs of different sizes were obtained by varying the mixing speed. WPI NPs of 200-500 nm were selected to prepare o/w Pickering emulsions because of their good stability against coalescence. By Confocal Laser Scanning Microscopy, it was observed that WPI NPs were closely packed and distributed at the surface of the emulsion droplets. By measuring water contact angles of WPI NPs films, it was found that under most conditions WPI NPs present good partial wetting properties, but that at the isoelectric point (pI) and high ionic strength the particles become more hydrophobic, resulting in less stable Pickering emulsion. Thus, at pH above and below the pI of WPI NPs and low to moderate ionic strengths (1-10 mM), and with a WPI NPs concentration of 2% (w/v), a stable Pickering emulsion can be obtained. The results may provide useful information for applications of WPI NPs in environmentally friendly and food grade applications, notably in food, pharmaceutical and cosmetic products.

  18. Coagulant properties of Moringa oleifera protein preparations: application to humic acid removal.

    PubMed

    Santos, Andréa F S; Paiva, Patrícia M G; Teixeira, José A C; Brito, António G; Coelho, Luana C B B; Nogueira, Regina

    2012-01-01

    This work aimed to characterize the coagulant properties of protein preparations from Moringa oleifera seeds in the removal of humic acids from water. Three distinct preparations were assayed, namely extract (seeds homogenized with 0.15 M NaCl), fraction (extract precipitated with 60% w/v ammonium sulphate) and cMoL (protein purified with guar gel column chromatography). The extract showed the highest coagulant activity in a protein concentration between 1 mg/L and 180 mg/L at pH 7.0. The zeta potential of the extract (-10 mV to -15 mV) was less negative than that of the humic acid (-41 mV to -42 mV) in a pH range between 5.0 and 8.0; thus, the mechanism that might be involved in this coagulation activity is adsorption and neutralization of charges. Reduction of total organic carbon (TOC) and dissolved organic carbon (DOC) was observed in water samples containing 9 mg/L carbon as humic acid when treated with 1 mg/L of the extract. A decrease in colour and in the aromatic content of the treated water was also observed. These results suggested that the extract from M. oleifera seeds in a low concentration (1 mg/L) can be an interesting natural alternative for removing humic acid from water in developing countries. The extract dose determined in the present study does not impart odour or colour to the treated water.

  19. DNA substrate preparation for atomic force microscopy studies of protein-DNA interactions.

    PubMed

    Buechner, Claudia N; Tessmer, Ingrid

    2013-12-01

    Protein-DNA interactions provide fundamental control mechanisms over biologically essential processes such as DNA replication, transcription, and repair. However, many details of these mechanisms still remain unclear. Atomic force microscopy (AFM) analyses provide unique and important structural and functional information on such protein-DNA interactions at the level of the individual molecules. The high sensitivity of the method with topographical visualization of all sample components also demands for extremely clean and pure materials. Here, we provide an overview of molecular biology-based approaches to produce DNA substrates for AFM imaging as well as other types of experiments, such as optical or magnetic tweezers, that profit from controllable substrate properties in long DNA fragments. We present detailed strategies to produce different types of motifs in DNA that are frequently employed targets of protein interactions. Importantly, the presented preparation techniques imply exact knowledge of the location of the introduced specific target sites within the DNA fragments, allowing for a distinction between specific and non-specific protein-DNA interactions in the AFM images and for separate conformational analyses of the different types of protein-DNA complexes.

  20. Preparation of photoreactive phospholipid polymer nanoparticles to immobilize and release protein by photoirradiation.

    PubMed

    Chen, Weixin; Inoue, Yuuki; Ishihara, Kazuhiko

    2015-11-01

    Photoreactive and cytocompatible polymer nanoparticles for immobilizing and releasing proteins were prepared. A water-soluble and amphiphilic phospholipid polymer, poly(2-methacryloyloxyethyl phosphorylcholine (MPC)-co-n-butyl methacrylate (BMA)-co-4-(4-(1-methacryloyloxyethyl)-2-methoxy-5-nitrophenoxy) butyric acid (PL)) (PMB-PL) was synthesized. The PMB-PL underwent a cleavage reaction at the PL unit with photoirradiation at a wavelength of 365 nm. Additionally, the PMB-PL took polymer aggregate in aqueous medium and was used to modify the surface of biodegradable poly(L-lactic acid) (PLA) nanoparticle as an emulsifier. The morphology of the PMB-PL/PLA nanoparticle was spherical and approximately 130 nm in diameter. The carboxylic acid group in the PL unit could immobilize proteins by covalent bonding. The bound proteins were released by a photoinduced cleavage reaction. Within 60s, up to 90% of the immobilized proteins was released by photoirradiation. From these results and with an understanding of the fundamental properties of MPC polymers, we concluded that PMB-PL/PLA nanoparticles have the potential to be used as smart carriers to deliver proteins to biological systems, such as the inside of living cells.

  1. Optimized extract preparation methods and reaction conditions for improved yeast cell-free protein synthesis.

    PubMed

    Hodgman, C Eric; Jewett, Michael C

    2013-10-01

    Cell-free protein synthesis (CFPS) has emerged as a powerful platform technology to help satisfy the growing demand for simple, affordable, and efficient protein production. In this article, we describe a novel CFPS platform derived from the popular bio-manufacturing organism Saccharomyces cerevisiae. By developing a streamlined crude extract preparation protocol and optimizing the CFPS reaction conditions we were able to achieve active firefly luciferase synthesis yields of 7.7 ± 0.5 µg mL(-1) with batch reactions lasting up to 2 h. This duration of synthesis is the longest ever reported for a yeast CFPS batch reaction. Furthermore, by removing extraneous processing steps and eliminating expensive reagents from the cell-free reaction, we have increased relative product yield (µg protein synthesized per $ reagent cost) over an alternative commonly used method up to 2000-fold from ∼2 × 10(-4) to ∼4 × 10(-1)  µg $(-1) , which now puts the yeast CPFS platform on par with other eukaryotic CFPS platforms commercially available. Our results set the stage for developing a yeast CFPS platform that provides for high-yielding and cost-effective expression of a variety of protein therapeutics and protein libraries.

  2. Incorporation of strawberries preparation in yoghurt: impact on phytochemicals and milk proteins.

    PubMed

    Oliveira, Ana; Alexandre, Elisabete M C; Coelho, Marta; Lopes, Cláudia; Almeida, Domingos P F; Pintado, Manuela

    2015-03-15

    An immediate decrease in the total antioxidant activity (23%) and total phenolic content (14%) was observed after addition of strawberry preparations to yoghurt. The total anthocyanin content did not change immediately, but decreased 24% throughout the yoghurt shelf-life. The individual compounds, (+)-catechin (60%), (-)-epicatechin (60%), kaempferol (33%) and quercetin-3-rutinoside (29%) decreased after 24h in the yoghurt made with the strawberry preparation. During the remaining period of storage these compounds increased by 47%, 6%, 4% and 18%, respectively. Pelargonidin-3-glucoside decreased 49% after 28 d. Immediately after the addition of the strawberry preparation to yoghurt, β-lactoglobulin decreased to values lower than the limit of detection and α-lactalbumin by approximately 34%, and was reduced further slowly throughout yoghurt self-life. An immediate interaction between the carrageenan present in the strawberry preparation and β-LG was observed. The variations of both polyphenols and protein in the presence of carrageenan and the potential interactions were discussed. PMID:25308682

  3. Hydrolysed egg displays strong decrease in allergenicity and is well tolerated by egg-allergic patients.

    PubMed

    Ballmer-Weber, B K; Brockow, K; Fiocchi, A; Theler, B; Vogel, L; Ring, J; Szépfalusi, Z; Mazzina, O; Schaller, R; Fritsché, R; Vissers, Y M; Nutten, S

    2016-05-01

    Food allergies are believed to be on the rise, and currently, management relies on the avoidance of the food. Hen's egg allergy is after cow's milk allergy the most common food allergy; eggs are used in many food products and thus difficult to avoid. A technological process using a combination of enzymatic hydrolysis and heat treatment was designed to produce modified hen's egg with reduced allergenic potential. Biochemical (SDS-PAGE, Size exclusion chromatography and LC-MS/MS) and immunological (ELISA, immunoblot, RBL-assays, animal model) analysis showed a clear decrease in intact proteins as well as a strong decrease of allergenicity. In a clinical study, 22 of the 24 patients with a confirmed egg allergy who underwent a double-blind food challenge with the hydrolysed egg remained completely free of symptoms. Hydrolysed egg products may be beneficial as low-allergenic foods for egg-allergic patients to extent their diet.

  4. Improved conjugation and purification strategies for the preparation of protein-polysaccharide conjugates.

    PubMed

    Suárez, N; Massaldi, H; Franco Fraguas, L; Ferreira, F

    2008-12-12

    A glycoconjugate constituted by the Streptococcus pneumoniae serotype 14 capsular polysaccharide (CPS14) and bovine serum albumin (BSA) was prepared, and the unique properties of Sephadex LH-20 were used to separate the conjugate from the unconjugated material. The strength of this approach consists in its capacity to produce pure polysaccharide-protein conjugate in good yield and free from unconjugated material, a common residual contaminant of this type of immunobiologicals. The CPS14-BSA conjugate prepared via an improved 1-cyano-4-dimethylaminopyridinium tetrafluoroborate (CDAP)-activation technique was characterized chemically and its immunogenicity was evaluated in mice. The purified conjugate, unlike the corresponding polysaccharide, produced a T-cell-dependent response in this species. PMID:18992885

  5. Preparation of inulin-type fructooligosaccharides using fast protein liquid chromatography coupled with refractive index detection.

    PubMed

    Li, J; Cheong, K L; Zhao, J; Hu, D J; Chen, X Q; Qiao, C F; Zhang, Q W; Chen, Y W; Li, S P

    2013-09-20

    A fast protein liquid chromatography coupled with refractive index detection (FPLC-RID) method was firstly developed for preparation and purification of fructooligosaccharides with different degree of polymerization from burdock, Arctium lappa. After extraction with 60% ethanol and decolorization with MCI gel CHP20P, total fructooligosaccharides were purified on Bio-Gel P-2 column eluted with water at the flow rate of 0.3 ml/min, which was the optimized conditions. The obtained fructooligosaccharides with degree of polymerization of 3-9 were identified based on their methylation analysis, MS and NMR data. This method has the advantages of high automation, good recovery and easy performance, which could be used for preparation of FOS from other sources, as well as other targeted compounds without UV absorbance.

  6. Preparation of inulin-type fructooligosaccharides using fast protein liquid chromatography coupled with refractive index detection.

    PubMed

    Li, J; Cheong, K L; Zhao, J; Hu, D J; Chen, X Q; Qiao, C F; Zhang, Q W; Chen, Y W; Li, S P

    2013-09-20

    A fast protein liquid chromatography coupled with refractive index detection (FPLC-RID) method was firstly developed for preparation and purification of fructooligosaccharides with different degree of polymerization from burdock, Arctium lappa. After extraction with 60% ethanol and decolorization with MCI gel CHP20P, total fructooligosaccharides were purified on Bio-Gel P-2 column eluted with water at the flow rate of 0.3 ml/min, which was the optimized conditions. The obtained fructooligosaccharides with degree of polymerization of 3-9 were identified based on their methylation analysis, MS and NMR data. This method has the advantages of high automation, good recovery and easy performance, which could be used for preparation of FOS from other sources, as well as other targeted compounds without UV absorbance. PMID:23962565

  7. Improved conjugation and purification strategies for the preparation of protein-polysaccharide conjugates.

    PubMed

    Suárez, N; Massaldi, H; Franco Fraguas, L; Ferreira, F

    2008-12-12

    A glycoconjugate constituted by the Streptococcus pneumoniae serotype 14 capsular polysaccharide (CPS14) and bovine serum albumin (BSA) was prepared, and the unique properties of Sephadex LH-20 were used to separate the conjugate from the unconjugated material. The strength of this approach consists in its capacity to produce pure polysaccharide-protein conjugate in good yield and free from unconjugated material, a common residual contaminant of this type of immunobiologicals. The CPS14-BSA conjugate prepared via an improved 1-cyano-4-dimethylaminopyridinium tetrafluoroborate (CDAP)-activation technique was characterized chemically and its immunogenicity was evaluated in mice. The purified conjugate, unlike the corresponding polysaccharide, produced a T-cell-dependent response in this species.

  8. Amino acid incorporation into the protein of mitochondrial preparations from cerebral cortex and spinal cord.

    PubMed

    Bachelard, H S

    1966-07-01

    1. Washed guinea-pig cerebral-cortex mitochondria incorporate [(14)C]leucine into their protein at a rate comparable with the rates reported for liver or heart mitochondria only if the mitochondria are separated from myelin and nerve endings by density-gradient centrifugation. 2. The non-mitochondrial components (myelin and nerve endings) of brain mitochondrial preparations incorporated [(14)C]leucine at a negligible rate. 3. The mitochondria do not require an exogenous supply of energy or a full supply of amino acids to support the process. 4. The incorporation rate was linear up to 2hr. aerobic incubation at 30 degrees and was inhibited by chloramphenicol, only slightly by actinomycin D and not by penicillin or pretreatment with ribonuclease. The observed incorporation is considered to be unlikely to be due to contaminating cytoplasmic ribosomes or bacteria. 5. The process was also studied in mitochondrial preparations from rabbit cerebral cortex and spinal cord.

  9. A novel method for preparation of HAMLET-like protein complexes.

    PubMed

    Permyakov, Sergei E; Knyazeva, Ekaterina L; Leonteva, Marina V; Fadeev, Roman S; Chekanov, Aleksei V; Zhadan, Andrei P; Håkansson, Anders P; Akatov, Vladimir S; Permyakov, Eugene A

    2011-09-01

    Some natural proteins induce tumor-selective apoptosis. α-Lactalbumin (α-LA), a milk calcium-binding protein, is converted into an antitumor form, called HAMLET/BAMLET, via partial unfolding and association with oleic acid (OA). Besides triggering multiple cell death mechanisms in tumor cells, HAMLET exhibits bactericidal activity against Streptococcus pneumoniae. The existing methods for preparation of active complexes of α-LA with OA employ neutral pH solutions, which greatly limit water solubility of OA. Therefore these methods suffer from low scalability and/or heterogeneity of the resulting α-LA - OA samples. In this study we present a novel method for preparation of α-LA - OA complexes using alkaline conditions that favor aqueous solubility of OA. The unbound OA is removed by precipitation under acidic conditions. The resulting sample, bLA-OA-45, bears 11 OA molecules and exhibits physico-chemical properties similar to those of BAMLET. Cytotoxic activities of bLA-OA-45 against human epidermoid larynx carcinoma and S. pneumoniae D39 cells are close to those of HAMLET. Treatment of S. pneumoniae with bLA-OA-45 or HAMLET induces depolarization and rupture of the membrane. The cells are markedly rescued from death upon pretreatment with an inhibitor of Ca(2+) transport. Hence, the activation mechanisms of S. pneumoniae death are analogous for these two complexes. The developed express method for preparation of active α-LA - OA complex is high-throughput and suited for development of other protein complexes with low-molecular-weight amphiphilic substances possessing valuable cytotoxic properties.

  10. Determinants of protein elution rates from preparative ion-exchange adsorbents.

    PubMed

    Angelo, James M; Lenhoff, Abraham M

    2016-04-01

    The rate processes involved in elution in preparative chromatography can affect both peak resolution and hence selectivity as well as practical factors such as facility fit. These processes depend on the physical structure of the adsorbent particles, the amount of bound solute, the solution conditions for operation or some combination of these factors. Ion-exchange adsorbents modified with covalently attached or grafted polymer layers have become widely used in preparative chromatography. Their often easily accessible microstructures offer substantial binding capacities for biomolecules, but elution has sometimes been observed to be undesirably slow. In order to determine which physicochemical phenomena control elution behavior, commercially available cellulosic, dextran-grafted and unmodified agarose materials were characterized here by their elution profiles at various conditions, including different degrees of loading. Elution data were analyzed under the assumption of purely diffusion-limited control, including the role of pore structure properties such as porosity and tortuosity. In general, effective elution rates decreased with the reduction of accessible pore volume, but differences among different proteins indicated the roles of additional factors. Additional measurements and analysis, including the use of confocal laser scanning microscopy to observe elution within single chromatographic particles, indicated the importance of protein association within the particle during elution. The use of protein stabilizing agents was explored in systems presenting atypical elution behavior, and l-arginine and disaccharide excipients were shown to alleviate the effects for one protein, lysozyme, in the presence of sodium chloride. Incorporation of these excipients into eluent buffer gave rise to faster elution and significantly lower pool volumes in elution from polymer-modified adsorbents.

  11. Proteomic tools for environmental microbiology--a roadmap from sample preparation to protein identification and quantification.

    PubMed

    Wöhlbrand, Lars; Trautwein, Kathleen; Rabus, Ralf

    2013-10-01

    The steadily increasing amount of (meta-)genomic sequence information of diverse organisms and habitats has a strong impact on research in microbial physiology and ecology. In-depth functional understanding of metabolic processes and overall physiological adaptation to environmental changes, however, requires application of proteomics, as the context specific proteome constitutes the true functional output of a cell. Considering the enormous structural and functional diversity of proteins, only rational combinations of various analytical approaches allow a holistic view on the overall state of the cell. Within the past decade, proteomic methods became increasingly accessible to microbiologists mainly due to the robustness of analytical methods (e.g. 2DE), and affordability of mass spectrometers and their relative ease of use. This review provides an overview on the complex portfolio of state-of-the-art proteomics and highlights the basic principles of key methods, ranging from sample preparation of laboratory or environmental samples, via protein/peptide separation (gel-based or gel-free) and different types of mass spectrometric protein/peptide analyses, to protein identification and abundance determination. PMID:23894077

  12. Preparation and recognition of surface molecularly imprinted core-shell microbeads for protein in aqueous solutions

    NASA Astrophysics Data System (ADS)

    Lu, Yan; Yan, Chang-Ling; Gao, Shu-Yan

    2009-04-01

    In this paper, a surface molecular imprinting technique was reported for preparing core-shell microbeads of protein imprinting, and bovine hemoglobin or bovine serum albumin were used as model proteins for studying the imprinted core-shell microbeads. 3-Aminophenylboronic acid (APBA) was polymerized onto the surface of polystyrene microbead in the presence of the protein templates to create protein-imprinted core-shell microbeads. The various samples were characterized using scanning electron microscopy (SEM), transmission electron microscopy (TEM), Raman spectroscopy, X-ray photoelectron spectroscopy (XPS) and Brunauer-Emmett-Teller (BET) methods. The effect of pH on rebinding of the template hemoglobin, the specific binding and selective recognition were studied for the imprinted microbeads. The results show that the bovine hemoglobin-imprinted core-shell microbeads were successfully created. The shell was a sort of imprinted thin films with porous structure and larger surface areas. The imprinted microbeads have good selectivity for templates and high stability. Due to the recognition sites locating at or closing to the surface, these imprinted microbeads have good property of mass-transport. Unfortunately, the imprint technology was not successfully applied to imprinting bovine serum albumin (BSA).

  13. Fermentation and recovery of glutamic acid from palm waste hydrolysate by Ion-exchange resin column.

    PubMed

    Das, K; Anis, M; Azemi, B M; Ismail, N

    1995-12-01

    Glutamic acid produced from palm waste hydrolysate by fermentation with Brevibacterium lactofermentum ATCC 13869 is produced with a remarkably high yield compared with that produced from pure glucose as a carbon source. The produce yield is 70 g/L with glucose, wherease, when palm waste hydrolysate is the fermentation medium in the same bioreactor under same conditions, it is 88 g/L. The higher yield may be attributed to the fact that this organism has the ability to convert sugars other than only glucose present in the hydrolysate. Bioreactor conditions most conducive for maximum production are pH 7.5, temperature of 30 degrees rmentation period of 48 h, inoculum size 6%, substrate concentration of 10 g per 100 mL, yeast extract 0.5 g per 100 mL as a suitable N source, and biotin at a concentration of 10 pg/L. Palm waste hydrolysate used in this study was prepared by enzymic saccharification of treated palm press fiber under conditions that yielded a maximum of 30 g/L total reducing sugars. Glutamic acid from fermentation broth was recovered by using a chromatographic column (5cm x 60 cm) packed with a strong ion-exchange resin. The filtered broth containing glutamic acid and other inorganic ions was fed to the fully charged column. The broth was continuously recycled at a flow rate of 50 mL/min (retention time of 55 min) until glutamic acid was fully adsorbed on the column leaving other ions in the effluent. Recovery was done by eluting with urea and sodium hydroxide for total displacement of glutamic acid from the resin. The eluent containing 88 g/L of glutamic acid was concentrated by evaporation to obtain solid crystals of the product. (c) 1995 John Wiley & Sons, Inc.

  14. Preparation and use of Xenopus egg extracts to study DNA replication and chromatin associated proteins

    PubMed Central

    Gillespie, Peter J.; Gambus, Agnieszka; Blow, J. Julian

    2012-01-01

    The use of cell-free extracts prepared from eggs of the South African clawed toad, Xenopus laevis, has led to many important discoveries in cell cycle research. These egg extracts recapitulate the key nuclear transitions of the eukaryotic cell cycle in vitro under apparently the same controls that exist in vivo. DNA added to the extract is first assembled into a nucleus and is then efficiently replicated. Progression of the extract into mitosis then allows the separation of paired sister chromatids. The Xenopus cell-free system is therefore uniquely suited to the study of the mechanisms, dynamics and integration of cell cycle regulated processes at a biochemical level. In this article we describe methods currently in use in our laboratory for the preparation of Xenopus egg extracts and demembranated sperm nuclei for the study of DNA replication in vitro. We also detail how DNA replication can be quantified in this system. In addition, we describe methods for isolating chromatin and chromatin-bound protein complexes from egg extracts. These recently developed and revised techniques provide a practical starting point for investigating the function of proteins involved in DNA replication. PMID:22521908

  15. Cellular uptake of beta-carotene from protein stabilized solid lipid nano-particles prepared by homogenization-evaporation method

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Using a homogenization-evaporation method, beta-carotene (BC) loaded nano-particles were prepared with different ratios of food-grade sodium caseinate (SC), whey protein isolate (WPI), or soy protein isolate (SPI) to BC and evaluated for their physiochemical stability, in vitro cytotoxicity, and cel...

  16. Method to produce succinic acid from raw hydrolysates

    DOEpatents

    Donnelly, Mark I.; Sanville-Millard, Cynthia Y.; Nghiem, Nhuan Phu

    2004-06-01

    A method for producing succinic acid from industrial-grade hydrolysates is provided, comprising supplying an organism that contains mutations for the genes ptsG, pflB, and ldhA, allowing said organism to accumulate biomass, and allowing said organism to metabolize the hydrolysate. Also provided is a bacteria mutant characterized in that it produces succinic acid from substrate contained in industrial-grade hydrolysate in a ratio of between 0.6:1 and 1.3:1 succinic acid to substrate.

  17. Bio-mimetic mineralization potential of collagen hydrolysate obtained from chromium tanned leather waste.

    PubMed

    Banerjee, Pradipta; Madhu, S; Chandra Babu, N K; Shanthi, C

    2015-04-01

    Hydroxyapatite (HA) ceramics serve as an alternative to autogenous-free bone grafting by virtue of their excellent biocompatibility. However, chemically synthesized HA lacks the strong load-bearing capacity as required by bone. The bio-mimetic growth of HA crystals on collagen surface provides a feasible solution for synthesizing bone substitutes with the desired properties. This study deals with the utilization of the collagen hydrolysate recovered from leather waste as a substrate for promoting HA crystal growth. Bio-mimetic growth of HA was induced by subjecting the hydrolysate to various mineralization conditions. Parameters that would have a direct effect on crystal growth were varied to determine the optimal conditions necessary. Maximum mineralization was achieved with a combination of 10mM of CaCl2, 5mM of Na2HPO4, 100mM of NaCl and 0.575% glutaraldehyde at a pH of 7.4. The metal-protein interactions leading to formation of HA were identified through Fourier-transform infrared (FTIR) spectroscopy and x-ray diffraction (XRD) studies. The crystal dimensions were determined to be in the nanoscale range by atomic force microscopy (AFM) and scanning electron microscopy (SEM). The size and crystallinity of bio-mimetically grown HA indicate that hydrolysate from leather waste can be used as an ideal alternative substrate for bone growth.

  18. Fish skin gelatin hydrolysates produced by visceral peptidase and bovine trypsin: Bioactivity and stability.

    PubMed

    Ketnawa, Sunantha; Benjakul, Soottawat; Martínez-Alvarez, Oscar; Rawdkuen, Saroat

    2017-01-15

    The peptidase from the viscera of farmed giant catfish was used for producing gelatin hydrolysates (HG) and compared with those produced from commercial bovine trypsin (HB). The degree of hydrolysis (DH) observed suggests that proteolytic cleavage rapidly occurred within the first 120min of incubation, and there was higher DH in HG than in HB. HG demonstrated the highest ACE-inhibitory activity, DPPH, ABTS radical scavenging activity, and FRAP. HB showed the highest FRAP activity. The DPPH radical scavenging activity of HG was quite stable over the pH range of 1-11, but it increased slightly when the heating duration time reached 240min at 100°C. The ACE-inhibitory activity of HG showed the highest stability at a pH of 7, and it remained very stable at 100°C for over 15-240min. The visceral peptidase from farmed giant catfish could be an alternative protease for generating protein hydrolysates with desirable bioactivities. The resulting hydrolysates showed good stability, making them potential functional ingredients for food formulations. PMID:27542490

  19. Effect of spray drying on the sensory and physical properties of hydrolysed casein using gum arabic as the carrier.

    PubMed

    Subtil, S F; Rocha-Selmi, G A; Thomazini, M; Trindade, M A; Netto, F M; Favaro-Trindade, C S

    2014-09-01

    This study was aimed at spray drying hydrolysed casein using gum Arabic as the carrier agent, in order to decrease the bitter taste. Three formulations with differing proportions of hydrolysed casein: gum Arabic (10:90, 20:80 and 30:70) were prepared and characterized. They were evaluated for their moisture content, water activity, hygroscopicity, dispersibility in water and in oil, particle size and distribution, particle morphology, thermal behaviour (DSC) and bitter taste by a trained sensory panel using a paired-comparison test (free samples vs. spray dried samples). The proportion of hydrolysed casein did not affect the morphology of the microspheres. The spray drying process increased product stability and modified the dissolution time, but had no effect on the ability of the material to dissolve in either water or oil. The sensory tests showed that the spray drying process using gum Arabic as the carrier was efficient in attenuating or masking the bitter taste of the hydrolysed casein. PMID:25190858

  20. Preparation of resveratrol-enriched and poor allergic protein peanut sprout from ultrasound treated peanut seeds.

    PubMed

    Yu, Miao; Liu, Hongzhi; Shi, Aimin; Liu, Li; Wang, Qiang

    2016-01-01

    Peanut sprout is a kind of high quality natural food which has important effect on health-care. It contains abundant bioactive substances such as resveratrol and lower fat. Naturally, resveratrol occurs in stilbene phytoalexin phenolic compound produced in response to a variety of biotic and abiotic stresses. In this study, the influence of ultrasonic stimulation on the resveratrol accumulate in germinant peanut prepared from three varieties (FH12, FH18, and BS1016) in the dry state before steeping were investigated. All experiments were performed using an ultrasonic cleaner bath operating at three frequencies (28, 45 and 100 kHz) for 20 min at constant temperature 30°C. The resulted amounts of resveratrol in peanut sprout were increasing by 2.25, 3.34, and 1.71 times compared with the control group of peanut germinated from FH12, FH18, and BS1016, respectively, after 3d with decreasing the amounts of allergic protein. After ultrasound, the germination rate and total sugar content increased slightly while the crude fat decreased and protein remained unchanged. Overall, the study results indicated that ultrasound treatment combined with germination can be an effective method for producing enriched-resveratrol and poor allergic protein peanut sprout as a functional vegetable. PMID:26384916

  1. Preparation of resveratrol-enriched and poor allergic protein peanut sprout from ultrasound treated peanut seeds.

    PubMed

    Yu, Miao; Liu, Hongzhi; Shi, Aimin; Liu, Li; Wang, Qiang

    2016-01-01

    Peanut sprout is a kind of high quality natural food which has important effect on health-care. It contains abundant bioactive substances such as resveratrol and lower fat. Naturally, resveratrol occurs in stilbene phytoalexin phenolic compound produced in response to a variety of biotic and abiotic stresses. In this study, the influence of ultrasonic stimulation on the resveratrol accumulate in germinant peanut prepared from three varieties (FH12, FH18, and BS1016) in the dry state before steeping were investigated. All experiments were performed using an ultrasonic cleaner bath operating at three frequencies (28, 45 and 100 kHz) for 20 min at constant temperature 30°C. The resulted amounts of resveratrol in peanut sprout were increasing by 2.25, 3.34, and 1.71 times compared with the control group of peanut germinated from FH12, FH18, and BS1016, respectively, after 3d with decreasing the amounts of allergic protein. After ultrasound, the germination rate and total sugar content increased slightly while the crude fat decreased and protein remained unchanged. Overall, the study results indicated that ultrasound treatment combined with germination can be an effective method for producing enriched-resveratrol and poor allergic protein peanut sprout as a functional vegetable.

  2. Effect of tannic acid-fish scale gelatin hydrolysate hybrid nanoparticles on intestinal barrier function and α-amylase activity.

    PubMed

    Wu, Shao-Jung; Ho, Yi-Cheng; Jiang, Shun-Zhou; Mi, Fwu-Long

    2015-07-01

    Practical application of tannic acid is limited because it readily binds proteins to form insoluble aggregates. In this study, tannic acid was self-assembled with fish scale gelatin hydrolysates (FSGH) to form stable colloidal complex nanoparticles. The nanoparticles prepared from 4 mg ml(-1) tannic acid and 4 mg ml(-1) FSGH had a mean particle size of 260.8 ± 3.6 nm, and showed a positive zeta potential (20.4 ± 0.4 mV). The nanoparticles acted as effective nano-biochelators and free radical scavengers because they provided a large number of adsorption sites for interaction with heavy metal ions and scavenging free radicals. The maximum adsorption capacity for Cu(2+) ions was 123.5 mg g(-1) and EC50 of DPPH radical scavenging activity was 21.6 ± 1.2 μg ml(-1). Hydroxyl radical scavenging effects of the nanoparticles were investigated by electron spin resonance spectroscopy. The copper-chelating capacity and free radical scavenging activity of the nanoparticles were associated with their capacity to inhibit Cu(2+) ion-induced barrier impairment and hyperpermeability of Caco-2 intestinal epithelial tight junction (TJ). However, α-amylase inhibitory activity of the nanoparticles was significantly lower than that of free tannic acid. The results suggest that the nanoparticles can ameliorate Cu(2+) ion induced intestinal epithelial TJ dysfunction without severely inhibiting the activity of the digestive enzymes. PMID:26069899

  3. Acute allergic skin response as a new tool to evaluate the allergenicity of whey hydrolysates in a mouse model of orally induced cow's milk allergy.

    PubMed

    van Esch, Betty C A M; Schouten, Bastiaan; Hofman, Gerard A; van Baalen, Ton; Nijkamp, Frans P; Knippels, Léon M J; Willemsen, Linette E M; Garssen, Johan

    2010-06-01

    Hypoallergenic milk formulae are used for cow's milk allergic infants and may be a good option for infants at risk. Clinical studies have shown that the protein source or the hydrolysis methodology used may influence the effectiveness in infants stressing the importance of adequate pre-clinical testing of hypoallergenic formulae in an in vivo model of orally induced cow's milk allergy. This study was undertaken to introduce a new read-out system to measure the residual allergenicity of whey hydrolysates on both the sensitization and challenge phase of orally induced cow's milk allergy in mice. Mice were sensitized orally to whey or a partial whey hydrolysate (pWH) to measure the residual sensitizing capacity. To predict the residual allergenicity of hydrolysates, whey allergic mice were challenged in the ear with pWH, extensive whey hydrolysate or an amino acid-based formula. An acute allergic skin response (ear swelling at 1 h), whey-specific serum antibodies, and local MCP-1 concentrations were measured. In contrast to whey, oral sensitization with pWH did not result in the induction of whey-specific antibodies, although a minor residual skin response to whey was observed after challenge. Skin exposure to whey hydrolysates showed a hydrolysation dependent reduction of the acute allergic skin response in whey allergic mice. In contrast to whey, skin exposure to pWH did not enhance tissue MCP-1 levels. The acute allergic skin response in mice orally sensitized to cow's milk proteins reveals a new pre-clinical tool which might provide information about the residual sensitizing capacity of hydrolysates supporting the discussion on the use of hypoallergenic formulae in high risk children. This mouse model might be a relevant model for the screening of new hypoallergenic formulae aimed to prevent or treat cow's milk allergy.

  4. Modelling of the production of ACE inhibitory hydrolysates of horse mackerel using proteases mixtures.

    PubMed

    Pérez-Gálvez, R; Morales-Medina, R; Espejo-Carpio, F; Guadix, A; Guadix, E M

    2016-09-14

    Fish protein hydrolysates from Mediterranean horse mackerel were produced by using a mixture of two commercial endoproteases (i.e. subtilisin and trypsin) at different levels of substrate concentration (2.5 g L(-1), 5 g L(-1), and 7.5 g L(-1) of protein), temperature (40 °C, 47.5 °C, and 55 °C) and percentage of subtilisin in the enzyme mixture (0%, 25%, 50%, 75% and 100%). A crossed mixture process model was employed to predict the degree of hydrolysis (DH) and the ACE inhibitory activity of the final hydrolysates as a function of the experimental factors. Both models were optimized for a maximum DH and ACE inhibition. A maximum DH (17.1%) was predicted at 2.54 g L(-1) of substrate concentration, 40 °C and an enzyme mixture comprising 38.3% of subtilisin and 61.7% of trypsin. Although its proteolytic activity is limited, the presence of trypsin in the enzyme mixture allowed obtaining higher degrees of hydrolysis at low temperatures, which is desirable to minimize thermal deactivation of the proteins. Similarly, a percentage of ACE inhibition above 48% was attained at 2.5 g L(-1) of protein, 40 °C and a 1 : 1 mixture of both proteases. Higher values of ACE inhibition could be attained by increasing both the temperature and the amount of trypsin in the enzyme mixture (e.g. 50% ACE inhibition at 55 °C and 81.5% of trypsin). Finally, those hydrolysates exhibiting the highest levels of ACE inhibition were subjected to simulated gastrointestinal digestion. These assays confirmed the resistance of active fractions against their degradation by digestive enzymes. PMID:27526864

  5. Accelerated shelf-life testing of quality loss for a commercial hydrolysed hen egg white powder.

    PubMed

    Rao, Qinchun; Rocca-Smith, Jeancarlo R; Schoenfuss, Tonya C; Labuza, Theodore P

    2012-11-15

    In recent years, due to the specific health benefits associated with bioactive peptides and the reduction of protein allergenicity by enzymatic hydrolysis, the utilisation of protein hydrolysates in functional foods and beverages for both protein supplementation and clinical use has significantly increased. However, few studies have explored the moisture-induced effects on food protein hydrolysates, and the resulting changes in the structure and texture of the food matrix as well as the loss in functional properties of bioactive peptides during storage. The main purpose of this study is to determine the influence of water activity (a(w)) on the storage quality of a commercial spray-dried hydrolysed hen egg white powder (HEW). During storage at 45 °C for two months at different a(w)s (0.05-0.79), the selected physicochemical properties of the HEW samples were analysed. Overall, the effect of a(w) on the colour change of HEW at 45 °C for one month was similar to that of HEW after four months at 23 °C due to the presence of a small amount of glucose in HEW. Several structural changes occurred at a(w)s from 0.43 to 0.79 including agglomeration, stickiness and collapse. Kinetic analysis showed a first-order hyperbolic model fit for the change in the L(∗) value, the total colour difference (ΔE(∗)) and the fluorescence intensity (FI). There was a high correlation between colour change and fluorescence, as expected for the Maillard reaction. The reduction in the remaining free amino groups was about 5% at a(w) 0.50 and 6% at a(w) 0.79 after one month storage. In summary, during storage, the Maillard reaction and/or its resulting products could decrease the nutritional value and the quality of HEW. PMID:22868115

  6. Preparation and characterization of milk protein films and their application for packaging of Cheddar cheese.

    PubMed

    Wagh, Y R; Pushpadass, Heartwin A; Emerald, F Magdaline Eljeeva; Nath, B Surendra

    2014-12-01

    Casein and whey protein concentrate (WPC) films, plasticized with glycerol and sorbitol independently, were prepared by casting. The film thickness, water vapour and oxygen permeation and tensile and moisture sorption properties of the films were determined. The tensile strength (TS), tensile strain (TE) and elastic modulus (EM) of the films ranged from 0.71 to 4.58 MPa, 19.22 to 66.63 % and 2.05 to 6.93 MPa, respectively. The film properties were influenced by the type of biopolymer (casein and whey protein concentrate), plasticizer and its concentration. Increasing the plasticizer concentration, increased the film thickness, TE and water vapour permeability (WVP), but decreased the TS and EM. As the concentration of plasticizer increased to the highest level, the film thickness increased from 0.168 to 0.305 mm for glycerol-plasticized films and from 0.251 to 0.326 mm for sorbitol-plasticized films. The film thickness increased because the amount of plasticizer in the film network increased and the amount of biopolymer remained same. Casein films showed superior tensile properties as compared to WPC films. The WVP of both casein and WPC films lied between 3.87 and 13.97 g.mm./(m(2).h.kPa). The moisture sorption isotherms of both films were typical of high-protein material, and were adequately described by the GAB model. The oxygen permeability of casein films was relatively lower than that of WPC films, regardless of the plasticizer used. The sensory data revealed that the organoleptic quality of Cheddar cheese was unaffected by milk-protein film packaging. PMID:25477643

  7. Identification of the major T-cell antigens present in the Brucella melitensis B115 protein preparation, Brucellergene OCB.

    PubMed

    Denoel, P A; Vo, T K; Weynants, V E; Tibor, A; Gilson, D; Zygmunt, M S; Limet, J N; Letesson, J J

    1997-09-01

    Brucellergene is a commercial allergen prepared from Brucella melitensis strain B115 and containing at least 20 cytoplasmic proteins. These proteins were separated by SDS-PAGE. The unstained gel was divided into 18 fractions and proteins were eluted from the gel fractions. The capacity of the separated proteins to elicit delayed-type hypersensitivity (DTH) in infected guinea-pigs or to induce the production of interferon-gamma (IFN-gamma) by blood cells from infected cattle was evaluated. The biological activity of the corresponding protein fractions blotted on to nitrocellulose was measured in a lymphocyte blastogenesis assay. Among the 18 fractions tested, two-spanning the mol. wt ranges 17-22 (fraction 8) and 35-42-kDa (fraction 17)-showed the maximum biological activity in the three tests. These fractions contain two antigens, the Brucella bacterioferritin (BFR) and P39 proteins. Both proteins are good candidates for the detection of cellular immunity to Brucella. PMID:9291893

  8. A pull-down method with a biotinylated bait protein prepared by cell-free translation using a puromycin linker.

    PubMed

    Mochizuki, Yuki; Kohno, Fumiaki; Nishigaki, Koichi; Nemoto, Naoto

    2013-03-01

    In this paper, we demonstrate a novel pull-down method that dramatically reduces the cost and preparation time of a bait protein by cell-free translation with a puromycin linker. With the C-terminus of the bait protein linked to biotin through a puromycin molecule after the translation reaction and subsequent mRNA degradation by RNase, the prey protein was easily pulled down by streptavidin-coated magnetic beads in a test tube. Three fluorescent prey protein types were tested and confirmed by gel electrophoresis to be pulled down easily and rapidly, depending on their affinity.

  9. Microspheres for protein delivery prepared from amphiphilic multiblock copolymers. 2. Modulation of release rate.

    PubMed

    Bezemer, J M; Radersma, R; Grijpma, D W; Dijkstra, P J; van Blitterswijk, C A; Feijen, J

    2000-07-01

    Amphiphilic multiblock copolymers, based on hydrophilic poly(ethylene glycol) (PEG) blocks and hydrophobic poly(butylene terephthalate) (PBT) blocks were used as matrix material for protein-loaded microspheres. The efficiency of lysozyme entrapment by a double emulsion method was found to depend on the swelling behavior of the polymers in water and decreased from 100% for polymers with a degree of swelling of less than 1.8 to 11% for PEG-PBT copolymers with a degree of swelling of 3.6. The particle size could be controlled by varying the concentration of the polymer solution used in the microsphere preparation. An increase in the polymer concentration resulted in a proportional increase in the particle size. The in vitro release profiles of the encapsulated model protein lysozyme could be precisely tailored by variation of the copolymer composition and the size of the microspheres. Both a slow continuous release of lysozyme, and a fast release which was completed within a few days could be obtained. The release behavior, attributed to a combination of diffusion and polymer degradation, could be described by a previously developed model. PMID:10825558

  10. Preparation of silica coated cobalt ferrite magnetic nanoparticles for the purification of histidine-tagged proteins

    NASA Astrophysics Data System (ADS)

    Aygar, Gülfem; Kaya, Murat; Özkan, Necati; Kocabıyık, Semra; Volkan, Mürvet

    2015-12-01

    Surface modified cobalt ferrite (CoFe2O4) nanoparticles containing Ni-NTA affinity group were synthesized and used for the separation of histidine tag proteins from the complex matrices through the use of imidazole side chains of histidine molecules. Firstly, CoFe2O4 nanoparticles with a narrow size distribution were prepared in an aqueous solution using the controlled co-precipitation method. In order to obtain small CoFe2O4 agglomerates, oleic acid and sodium chloride were used as dispersants. The CoFe2O4 particles were coated with silica and subsequently the surface of these silica coated particles (SiO2-CoFe2O4) was modified by amine (NH2) groups in order to add further functional groups on the silica shell. Then, carboxyl (-COOH) functional groups were added to the SiO2-CoFe2O4 magnetic nanoparticles through the NH2 groups. After that Nα,Nα-Bis(carboxymethyl)-L-lysine hydrate (NTA) was attached to carboxyl ends of the structure. Finally, the surface modified nanoparticles were labeled with nickel (Ni) (II) ions. Furthermore, the modified SiO2-CoFe2O4 magnetic nanoparticles were utilized as a new system that allows purification of the N-terminal His-tagged recombinant small heat shock protein, Tpv-sHSP 14.3.

  11. A FRET based method to study protein thiol oxidation in histological preparations

    PubMed Central

    Mastroberardino, Pier G.; Orr, Adam L.; Hu, Xiaoping; Na, Hye Mee; Greenamyre, J. Timothy

    2008-01-01

    Cysteine residues in proteins have important biological roles. For example, disulfides bonds are important structural elements; additionally, reversible oxidation of thiols to disulfides functions as a molecular switch and constitutes an early response to oxidative damage Because organs are heterogeneous structures composed of diverse cell types, there is a compelling need for a histological approach to investigate thiol oxidation in situ in order to address the role of specific cell types in oxidative imbalance. Here we describe a fluorescence technique – which can be used in association with standard immunological staining procedures – to detect variations in disulfides in histological preparations. Moreover, by monitoring the fluorescence resonance energy transfer (FRET) between a labeled specific primary antibody and the thiol probe described here, this method can detect thiol oxidation in candidate proteins of interest. When applied to an animal model of Parkinson’s disease, our technique demonstrated that thiol oxidation occurs selectively in the dopaminergic neurons of the substantia nigra, the same neurons that are lost selectively in the disease. In summary, this technique provides a new, powerful tool to provide further understanding of oxidative imbalance, a phenomenon common to many diseases. PMID:18620047

  12. Dry powders of stable protein formulations from aqueous solutions prepared using supercritical CO(2)-assisted aerosolization.

    PubMed

    Sellers, S P; Clark, G S; Sievers, R E; Carpenter, J F

    2001-06-01

    We report on the use of a new supercritical carbon dioxide-assisted aerosolization coupled with bubble drying technology to prepare stabilized, dry, finely divided powders from aqueous protein formulations. In this study, the feasibility of this new technology was tested using two model proteins, lysozyme and lactate dehydrogenase (LDH). In the absence of excipients, lysozyme was observed to undergo perturbations of secondary structure observed by solid-state infrared spectroscopy. In the presence of sucrose, this unfolding was minimized. Lysozyme did not, however, undergo irreversible loss of activity, as all lysozyme powders generated by supercritical CO(2)-assisted aerosolization (with or without excipients) regained almost complete activity on reconstitution. The more labile LDH suffered irrecoverable loss of activity on reconstituting after supercritical CO(2)-assisted aerosolization and bubble drying in the absence of carbohydrate stabilizers. LDH could be stabilized throughout the nebulization, drying, and rehydration processes with the addition of sucrose, and almost complete preservation of activity was achieved with the further addition of a surface active agent, such as Tween 20, to the aqueous formulation prior to processing.

  13. Preparation of protein imprinted materials by hierarchical imprinting techniques and application in selective depletion of albumin from human serum

    NASA Astrophysics Data System (ADS)

    Liu, Jinxiang; Deng, Qiliang; Tao, Dingyin; Yang, Kaiguang; Zhang, Lihua; Liang, Zhen; Zhang, Yukui

    2014-06-01

    Hierarchical imprinting was developed to prepare the protein imprinted materials, as the artificial antibody, for the selective depletion of HSA from the human serum proteome. Porcine serum albumin (PSA) was employed as the dummy template for the fabrication of the recognition sites. To demonstrate the advantages of the hierarchical imprinting, molecularly imprinted polymers prepared by hierarchical imprinting technique (h-MIPs) were compared with those obtained by bulk imprinting (b-MIPs), in terms of the binding capacity, adsorption kinetics, selectivity and synthesis reproducibility. The binding capacity of h-MIPs could reach 12 mg g-1. And saturation binding could be reached in less than 20 min for the h-MIPs. In the protein mixture, h-MIPs exhibit excellent selectivity for PSA, with imprinting factors as about 3.6, much higher than those for non-template proteins. For the proteomic application, the identified protein group number in serum treated by h-MIPs was increased to 422, which is 21% higher than that obtained from the original serum, meanwhile the identified protein group number for the Albumin Removal kit was only 376. The results demonstrate that protein imprinted polymers prepared by hierarchical imprinting technique, might become the artificial antibodies for the selective depletion of high abundance proteins in proteome study.

  14. Efficient DNP NMR of membrane proteins: sample preparation protocols, sensitivity, and radical location.

    PubMed

    Liao, Shu Y; Lee, Myungwoon; Wang, Tuo; Sergeyev, Ivan V; Hong, Mei

    2016-03-01

    Although dynamic nuclear polarization (DNP) has dramatically enhanced solid-state NMR spectral sensitivities of many synthetic materials and some biological macromolecules, recent studies of membrane-protein DNP using exogenously doped paramagnetic radicals as polarizing agents have reported varied and sometimes surprisingly limited enhancement factors. This motivated us to carry out a systematic evaluation of sample preparation protocols for optimizing the sensitivity of DNP NMR spectra of membrane-bound peptides and proteins at cryogenic temperatures of ~110 K. We show that mixing the radical with the membrane by direct titration instead of centrifugation gives a significant boost to DNP enhancement. We quantify the relative sensitivity enhancement between AMUPol and TOTAPOL, two commonly used radicals, and between deuterated and protonated lipid membranes. AMUPol shows ~fourfold higher sensitivity enhancement than TOTAPOL, while deuterated lipid membrane does not give net higher sensitivity for the membrane peptides than protonated membrane. Overall, a ~100 fold enhancement between the microwave-on and microwave-off spectra can be achieved on lipid-rich membranes containing conformationally disordered peptides, and absolute sensitivity gains of 105-160 can be obtained between low-temperature DNP spectra and high-temperature non-DNP spectra. We also measured the paramagnetic relaxation enhancement of lipid signals by TOTAPOL and AMUPol, to determine the depths of these two radicals in the lipid bilayer. Our data indicate a bimodal distribution of both radicals, a surface-bound fraction and a membrane-bound fraction where the nitroxides lie at ~10 Å from the membrane surface. TOTAPOL appears to have a higher membrane-embedded fraction than AMUPol. These results should be useful for membrane-protein solid-state NMR studies under DNP conditions and provide insights into how biradicals interact with phospholipid membranes. PMID:26873390

  15. Purification of Antioxidant Peptides by High Resolution Mass Spectrometry from Simulated Gastrointestinal Digestion Hydrolysates of Alaska Pollock (Theragra chalcogramma) Skin Collagen

    PubMed Central

    Sun, Liping; Chang, Weidan; Ma, Qingyu; Zhuang, Yongliang

    2016-01-01

    In this study, the stable collagen hydrolysate was prepared by alcalase hydrolysis and twice simulated gastrointestinal digestion from Alaska pollock skin. The characteristics of hydrolysates and antioxidant activities in vitro, including 2,2′-azino-bis (3-ethylbenzothiazoline-6-sulfonic acid) radical (ABTS•+) scavenging activity, ferric-reducing antioxidant power (FRAP) and hydroxyl radical (OH·) scavenging activity, were determined. After twice simulated gastrointestinal digestion of skin collagen (SGI-2), the degree of hydrolysis (DH) reached 26.17%. The main molecular weight fractions of SGI-2 were 1026.26 and 640.53 Da, accounting for 59.49% and 18.34%, respectively. Amino acid composition analysis showed that SGI-2 had high content of total hydrophobic amino acid (307.98/1000). With the simulated gastrointestinal digestion progressing, the antioxidant activities increased significantly (p < 0.05). SGI-2 was further purified by gel filtration chromatography, ion exchange chromatography and high performance liquid chromatography, and the A1a3c–p fraction with high hydroxyl radical scavenging activity (IC50 = 7.63 μg/mL) was obtained. The molecular weights and amino acid sequences of key peptides of A1a3c–p were analyzed using high resolution mass spectrometry (LC-ESI-LTQ-Orbitrap-MS) combined with de novo software and UniProt of MaxQuant software. Four peptides were identified from A1a3c–p, including YGCC (444.1137 Da) and DSSCSG (554.1642 Da) identified by de novo software and NNAEYYK (900.3978 Da) and PAGNVR (612.3344 Da) identified by UniProt of MaxQuant software. The molecular weights and amino acid sequences of four peptides were in accordance with the features of antioxidant peptides. The results indicated that different peptides were identified by different data analysis software according to spectrometry mass data. Considering the complexity of LC-ESI-LTQ-Orbitrap-MS, it was necessary to use the different methods to identify the key peptides

  16. Development of a yeast strain for xylitol production without hydrolysate detoxification as part of the integration of co-product generation within the lignocellulosic ethanol process.

    PubMed

    Huang, Chiung-Fang; Jiang, Yi-Feng; Guo, Gia-Luen; Hwang, Wen-Song

    2011-02-01

    The present study verified an applicable technology of xylitol bioconversion as part of the integration of co-product generation within second-generation bioethanol processes. A newly isolated yeast strain, Candida tropicalis JH030, was shown to have a capacity for xylitol production from hemicellulosic hydrolysate without detoxification. The yeast gives a promising xylitol yield of 0.71 g(p) g(s)(-1) from non-detoxified rice straw hydrolysate that had been prepared by the dilute acid pretreatment under severe conditions. The yeast's capacity was also found to be practicable with various other raw materials, such as sugarcane bagasse, silvergrass, napiergrass and pineapple peel. The lack of a need to hydrolysate detoxification enhances the potential of this newly isolated yeast for xylitol production and this, in turn, has the capacity to improve economics of lignocellulosic ethanol production. PMID:21095119

  17. A Standard Operating Procedure (SOP) for the preparation of intra- and extracellular proteins of Clostridium acetobutylicum for proteome analysis.

    PubMed

    Schwarz, Katrin; Fiedler, Tomas; Fischer, Ralf-Jörg; Bahl, Hubert

    2007-02-01

    We report on the development of a Standard Operating Procedure (SOP) for extraction and handling of intra- and extracellular protein fractions of Clostridium acetobutylicum ATCC 824 for reproducible high quality two-dimensional gel electrophoresis (2-DE) analyses. Standardized cells from a phosphate-limited chemostat were used to evaluate different protein preparation methods. For the preparation of the secretome, a dialysis/ultrafiltration procedure resulted in higher protein yields and proved to be more reliable compared to different precipitation methods using TCA, DOC-TCA, acetone, and PEG 6000. Sonication was found to be the most efficient method among different tested techniques of cell disruption for the analysis of the intracellular proteome. Furthermore, the effect of protease inhibitors and sample storage conditions were tested for both intra- and extracellular protein samples. Significant changes in the protein pattern were observed depending on the addition of protease inhibitors. 2-DE gels with a pH gradient from 4 to 7 prepared according to the developed SOP contained at least 736 intracellular and 324 extracellular protein spots.

  18. Amylase binding to starch granules under hydrolysing and non-hydrolysing conditions.

    PubMed

    Dhital, Sushil; Warren, Frederick J; Zhang, Bin; Gidley, Michael J

    2014-11-26

    Although considerable information is available about amylolysis rate, extent and pattern of granular starches, the underlying mechanisms of enzyme action and interactions are not fully understood, partly due to the lack of direct visualisation of enzyme binding and subsequent hydrolysis of starch granules. In the present study, α-amylase (AA) from porcine pancreas was labelled with either fluorescein isothiocyanate (FITC) or tetramethylrhodamine isothiocyanate (TRITC) fluorescent dye with maintenance of significant enzyme activity. The binding of FITC/TRITC-AA conjugate to the surface and interior of granules was studied under both non-hydrolysing (0 °C) and hydrolysing (37 °C) conditions with confocal microscopy. It was observed that enzyme binding to maize starch granules under both conditions was more homogenous compared with potato starch. Enzyme molecules appear to preferentially bind to the granules or part of granules that are more susceptible to enzymic degradation. The specificity is such that fresh enzyme added after a certain time of incubation binds at the same location as previously bound enzyme. By visualising the enzyme location during binding and hydrolysis, detailed information is provided regarding the heterogeneity of granular starch digestion. PMID:25256464

  19. Amylase binding to starch granules under hydrolysing and non-hydrolysing conditions.

    PubMed

    Dhital, Sushil; Warren, Frederick J; Zhang, Bin; Gidley, Michael J

    2014-11-26

    Although considerable information is available about amylolysis rate, extent and pattern of granular starches, the underlying mechanisms of enzyme action and interactions are not fully understood, partly due to the lack of direct visualisation of enzyme binding and subsequent hydrolysis of starch granules. In the present study, α-amylase (AA) from porcine pancreas was labelled with either fluorescein isothiocyanate (FITC) or tetramethylrhodamine isothiocyanate (TRITC) fluorescent dye with maintenance of significant enzyme activity. The binding of FITC/TRITC-AA conjugate to the surface and interior of granules was studied under both non-hydrolysing (0 °C) and hydrolysing (37 °C) conditions with confocal microscopy. It was observed that enzyme binding to maize starch granules under both conditions was more homogenous compared with potato starch. Enzyme molecules appear to preferentially bind to the granules or part of granules that are more susceptible to enzymic degradation. The specificity is such that fresh enzyme added after a certain time of incubation binds at the same location as previously bound enzyme. By visualising the enzyme location during binding and hydrolysis, detailed information is provided regarding the heterogeneity of granular starch digestion.

  20. Glycyl endopeptidase from papaya latex: partial purification and use for production of fish gelatin hydrolysate.

    PubMed

    Karnjanapratum, Supatra; Benjakul, Soottawat

    2014-12-15

    An aqueous two-phase system (ATPS) in combination with ammonium sulphate ((NH4)2SO4) precipitation was applied to fractionate glycyl endopeptidase from the papaya latex of Red Lady and Khack Dum cultivars. ATPS containing polyethylene glycol (PEG 2000 and 6000) and salts ((NH4)2SO4 and MgSO4) at different concentrations were used. Glycyl endopeptidase with high purification fold (PF) and yield was found in the salt-rich bottom phase of ATPS with 10%PEG 6000-10% (NH4)2SO4. When ATPS fraction from Red Lady cultivar was further precipitated with 40-60% saturation of (NH4)2SO4, PF of 2.1-fold with 80.23% yield was obtained. Almost all offensive odorous compounds, particularly benzyl isothiocyanate, were removed from partially purified glycyl endopeptidase (PPGE). The fish gelatin hydrolysates prepared using PPGE showed higher ABTS radical scavenging activity and less odour, compared with those of crude extract (CE). Thus antioxidative gelatin hydrolysate with negligible undesirable odour could be prepared with the aid of PPGE. PMID:25038693

  1. Preparation of recombinant protein spotted arrays for proteome-wide identification of kinase targets.

    PubMed

    Im, Hogune; Snyder, Michael

    2013-04-01

    Protein microarrays allow unique approaches for interrogating global protein interaction networks. Protein arrays can be divided into two categories: antibody arrays and functional protein arrays. Antibody arrays consist of various antibodies and are appropriate for profiling protein abundance and modifications. Functional full-length protein arrays employ full-length proteins with various post-translational modifications. A key advantage of the latter is rapid parallel processing of large number of proteins for studying highly controlled biochemical activities, protein-protein interactions, protein-nucleic acid interactions, and protein-small molecule interactions. This unit presents a protocol for constructing functional yeast protein microarrays for global kinase substrate identification. This approach enables the rapid determination of protein interaction networks in yeast on a proteome-wide level. The same methodology can be readily applied to higher eukaryotic systems with careful consideration of overexpression strategy.

  2. Preparation and Delivery of Protein Microcrystals in Lipidic Cubic Phase for Serial Femtosecond Crystallography.

    PubMed

    Ishchenko, Andrii; Cherezov, Vadim; Liu, Wei

    2016-09-20

    Membrane proteins (MPs) are essential components of cellular membranes and primary drug targets. Rational drug design relies on precise structural information, typically obtained by crystallography; however MPs are difficult to crystallize. Recent progress in MP structural determination has benefited greatly from the development of lipidic cubic phase (LCP) crystallization methods, which typically yield well-diffracting, but often small crystals that suffer from radiation damage during traditional crystallographic data collection at synchrotron sources. The development of new-generation X-ray free-electron laser (XFEL) sources that produce extremely bright femtosecond pulses has enabled room temperature data collection from microcrystals with no or negligible radiation damage. Our recent efforts in combining LCP technology with serial femtosecond crystallography (LCP-SFX) have resulted in high-resolution structures of several human G protein-coupled receptors, which represent a notoriously difficult target for structure determination. In the LCP-SFX technique, LCP is recruited as a matrix for both growth and delivery of MP microcrystals to the intersection of the injector stream with an XFEL beam for crystallographic data collection. It has been demonstrated that LCP-SFX can substantially improve the diffraction resolution when only sub-10 µm crystals are available, or when the use of smaller crystals at room temperature can overcome various problems associated with larger cryocooled crystals, such as accumulation of defects, high mosaicity and cryocooling artifacts. Future advancements in X-ray sources and detector technologies should make serial crystallography highly attractive and practicable for implementation not only at XFELs, but also at more accessible synchrotron beamlines. Here we present detailed visual protocols for the preparation, characterization and delivery of microcrystals in LCP for serial crystallography experiments. These protocols include

  3. Changes of protein solutions during storage: a study of albumin pharmaceutical preparations.

    PubMed

    Christiansen, Cathrine; Skotland, Tore

    2010-03-05

    During the production of air-filled albumin microspheres, to be used as an ultrasound contrast agent, it was observed that some albumin lots could not be used owing to albumin precipitation. In order to understand the reason for these lot-to-lot variations, 24 lots of 5% (w/v) human albumin pharmaceutical preparations were analysed. The results revealed that the good albumin lots all contained <0.03 mol of free SH groups per mol of albumin. The precipitation observed with other lots was most probably due to higher amounts of free SH groups. The lower amount of free SH groups in the good lots correlated with: (i) a yellow colour of the solutions and a UV-visible spectrum similar to that observed for non-enzymatic glycosylation; (ii) a decreased fructosamine content; (iii) an increased mobility against the anode in isoelectric focusing; and (iv) an increased truncation of the two N-terminal amino acids. No, or only small, differences were observed for the amounts of albumin dimer, albumin aggregates and protein impurities, and these could not account for the albumin precipitation. The differences observed between the albumin lots were most probably due to varying storage times and/or storage conditions, and incubation experiments revealed changes in all parameters that differed between the good and bad lots. Increasing the storage temperature or exposing the solutions to light resulted in a faster decrease of free SH groups and increase of the yellow colouration. It is likely that at least some of the changes observed were due to reactive degradation products formed from the stabilizer N-acetyl-L-tryptophan. The results presented should also be of interest regarding the storage of monoclonal antibodies and other proteins used in pharmaceuticals.

  4. Preparation and Delivery of Protein Microcrystals in Lipidic Cubic Phase for Serial Femtosecond Crystallography.

    PubMed

    Ishchenko, Andrii; Cherezov, Vadim; Liu, Wei

    2016-01-01

    Membrane proteins (MPs) are essential components of cellular membranes and primary drug targets. Rational drug design relies on precise structural information, typically obtained by crystallography; however MPs are difficult to crystallize. Recent progress in MP structural determination has benefited greatly from the development of lipidic cubic phase (LCP) crystallization methods, which typically yield well-diffracting, but often small crystals that suffer from radiation damage during traditional crystallographic data collection at synchrotron sources. The development of new-generation X-ray free-electron laser (XFEL) sources that produce extremely bright femtosecond pulses has enabled room temperature data collection from microcrystals with no or negligible radiation damage. Our recent efforts in combining LCP technology with serial femtosecond crystallography (LCP-SFX) have resulted in high-resolution structures of several human G protein-coupled receptors, which represent a notoriously difficult target for structure determination. In the LCP-SFX technique, LCP is recruited as a matrix for both growth and delivery of MP microcrystals to the intersection of the injector stream with an XFEL beam for crystallographic data collection. It has been demonstrated that LCP-SFX can substantially improve the diffraction resolution when only sub-10 µm crystals are available, or when the use of smaller crystals at room temperature can overcome various problems associated with larger cryocooled crystals, such as accumulation of defects, high mosaicity and cryocooling artifacts. Future advancements in X-ray sources and detector technologies should make serial crystallography highly attractive and practicable for implementation not only at XFELs, but also at more accessible synchrotron beamlines. Here we present detailed visual protocols for the preparation, characterization and delivery of microcrystals in LCP for serial crystallography experiments. These protocols include

  5. Detection of antibacterial activity of an enzymatic hydrolysate generated by processing rainbow trout by-products with trout pepsin.

    PubMed

    Wald, Maleen; Schwarz, Karin; Rehbein, Hartmut; Bußmann, Bettina; Beermann, Christopher

    2016-08-15

    Trout by-product hydrolysates, generated using trout pepsin, were characterized and studied in terms of their antibacterial effects against food contaminants and fish farming pathogens. After a hydrolysis time of 25 min, the hydrolysates demonstrated inhibitory activity against several gram-positive and gram-negative bacteria. The degree of hydrolysis (DH) was found to exert a considerable influence on antibacterial activity, with a significant increase in the observed inhibitory effect at the beginning of hydrolysis. The highest antibacterial activity was obtained at a DH of 30% (enzyme/protein ratio 0.04 U/mg of protein, enzyme activity 6.5 U/mg protein, hydrolysis conditions 37°C, pH 3.0). The highest antibacterial activity detected was against the fish farming bacteria Flavobacterium psychrophilum and Renibacterium salmoninarum, with minimal inhibition concentrations of 2mg/ml and 5mg/ml, respectively. The amino acid determination of the hydrolysate (DH 30%) revealed that lysine, leucine, alanine, arginine, glycine, aspartic acid and glutamic acid residues represented the major amino acids. PMID:27006234

  6. Microbial utilization and biopolyester synthesis of bagasse hydrolysates.

    PubMed

    Yu, Jian; Stahl, Heiko

    2008-11-01

    Cellulosic biomass is a potentially inexpensive renewable feedstock for the biorefineries of fuels, chemicals and materials. Sugarcane bagasse was pretreated in dilute acid solution under moderately severe conditions, releasing sugars and other hydrolysates including volatile organic acids, furfurals and acid soluble lignin. Utilization of the hydrolysates by an aerobic bacterium, Ralstonia eutropha, was investigated to determine if the organic inhibitors can be removed for potential recycling and reuse of the process water. Simultaneous biosynthesis of polyhydroxyalkanoates (PHAs) for the production of value-added bioplastics was also investigated. An inhibitory effect of hydrolysates on microbial activity was observed, but it could be effectively relieved by using (a) a large inoculum, (b) a diluted hydrolysate solution, and (c) a tolerant strain, or a combination of the three. The major organic inhibitors including formic acid, acetic acid, furfural and acid soluble lignin were effectively utilized and removed to low concentration levels (less than 100ppm) while at the same time, PHA biopolyesters were synthesized and accumulated to 57wt% of cell mass under appropriate C/N ratios. Poly(3-hydroxybutyrate) was the predominant biopolyester formed on the hydrolysates, but the cells could also synthesize co-polyesters that exhibit high ductility. PMID:18474421

  7. Use of fish hydrolysates and fishmeal by-products of the Alaskan fishing industry in diets for Pacific white shrimp (Litopenaeus vannamei)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The suitability of four fish hydrolysates and two fishmeals derived from by-products of the Alaskan fishing industry, as menhaden fishmeal replacements in shrimp diets was determined. A control diet (30% crude protein and 8.5% crude lipid) was produced with menhaden meal (13% of diet). Experimental ...

  8. Preparation of a porcine plasma protein composite film and its application.

    PubMed

    Lee, Ji-Hyun; Song, Kyung Bin

    2015-01-01

    To use blood released from slaughtering houses, a porcine plasma protein (PPP)/nanoclay composite film was prepared. The tensile strength and elongation at break values of the PPP composite film with 5% nanoclay were 10.01 MPa and 6.55%, respectively. The PPP composite film containing 1% grapefruit seed extract (GSE) was applied to pork meat, and the populations of inoculated Escherichia coli O157:H7 and Listeria monocytogenes in the pork meat packaged with the PPP composite film decreased by 0.8 and 1.0 log CFU/g, respectively, after 7 days of storage compared to the populations of the control. In addition, thiobarbituric acid values in the pork meat packaged with the PPP composite film were less than those of the control sample during storage. These results suggest that the PPP nanocomposite film containing 1% GSE can be used as a packaging material to maintain the quality of pork meat. PMID:25248798

  9. Preparation of calcium hydroxyapatite nanoparticles using microreactor and their characteristics of protein adsorption.

    PubMed

    Kandori, Kazuhiko; Kuroda, Tomohiko; Togashi, Shigenori; Katayama, Erika

    2011-02-01

    The calcium hydroxyapatite Ca(10)(PO(4))(6)(OH)(2) (Hap) nanoparticles were prepared by using microreactor and employed these Hap nanoparticles to clarify the adsorption behavior of proteins. The size of Hap particles produced by the microreactor reduced in the order of a hardness of the reaction conditions for mixing Ca(OH)(2) and H(3)PO(4) aqueous solutions, such as flow rates of both solutions and temperature. Finally, the size of the smallest Hap nanoparticle became 2 × 15 nm(2), similar to that of BSA molecule (4 × 14 nm(2)). It is noteworthy that the smallest Hap nanoparticles still possesses rodlike shape, suggesting that particles are grown along c-axis even though the reactants mixed very rapidly in narrow channels of the microreactors. The X-ray diffraction patterns of the Hap nanoparticles revealed that the crystallinity of the materials produced by the microreactor is low. The FTIR measurement indicated that the Hap nanoparticles produced by microreactor were carbonate-substituted type B Hap, where the carbonate ions replace the phosphate ions in the crystal lattice. All the adsorption isotherms of acidic bovine serum albumin (BSA), neutral myoglobin (MGB), and basic lysozyme (LSZ) onto Hap nanoparticles from 1 × 10(-4) mol/dm(3) KCl solution were the Langmuirian type. The saturated amounts of adsorbed BSA (n(S)(BSA)) for the Hap nanoparticles produced by microreactor were decreased with decrease in the mean particle length, and finally it reduced to zero for the smallest Hap nanoparticles. Similar results were observed for the adsorption of LSZ; the saturated amounts of adsorbed LSZ (n(S)(LSZ)) also reduced to zero for the smallest Hap nanoparticles. However, in the case of MGB, the saturated mounts of adsorbed MGB (n(S)(MGB)) are also depressed with decreased in their particle size, but about half of MGB molecules still adsorbed onto the smallest Hap nanoparticles. This difference in the protein adsorption behavior was explained by the difference

  10. Global regulator engineering significantly improved Escherichia coli tolerances toward inhibitors of lignocellulosic hydrolysates.

    PubMed

    Wang, Jianqing; Zhang, Yan; Chen, Yilu; Lin, Min; Lin, Zhanglin

    2012-12-01

    Lignocellulosic biomass is regarded as the most viable source of feedstock for industrial biorefinery, but the harmful inhibitors generated from the indispensable pretreatments prior to fermentation remain a daunting technical hurdle. Using an exogenous regulator, irrE, from the radiation-resistant Deinococcus radiodurans, we previously showed that a novel global regulator engineering (GRE) approach significantly enhanced tolerances of Escherichia coli to alcohol and acetate stresses. In this work, an irrE library was subjected to selection under various stresses of furfural, a typical hydrolysate inhibitor. Three furfural tolerant irrE mutants including F1-37 and F2-1 were successfully obtained. The cells containing these mutants reached OD(600) levels of 4- to 16-fold of that for the pMD18T cells in growth assay under 0.2% (v/v) furfural stress. The cells containing irrE F1-37 and F2-1 also showed considerably reduced intracellular oxygen species (ROS) levels under furfural stress. Moreover, these two irrE mutants were subsequently found to confer significant cross tolerances to two other most common inhibitors, 5-hydroxymethyl-2-furaldehyde (HMF), vanillin, as well as real lignocellulosic hydrolysates. When evaluated in Luria-Bertani (LB) medium supplemented with corn stover cellulosic hydrolysate (prepared with a solid loading of 30%), the cells containing the mutants exhibited lag phases markedly shortened by 24-44 h in comparison with the control cells. This work thus presents a promising step forward to resolve the inhibitor problem for E. coli. From the view of synthetic biology, irrE can be considered as an evolvable "part" for various stresses. Furthermore, this GRE approach can be extended to exploit other exogenous global regulators from extremophiles, and the native counterparts in E. coli, for eliciting industrially useful phenotypes.

  11. Characterisation and foaming properties of hydrolysates derived from rapeseed isolate.

    PubMed

    Larré, C; Mulder, W; Sánchez-Vioque, R; Lazko, J; Bérot, S; Guéguen, J; Popineau, Y

    2006-04-15

    Two hydrolysis methods used to obtain rapeseed isolate derivates were compared: chemical hydrolysis performed under alkaline conditions and pepsic proteolysis performed under acidic conditions. The mean molecular weights obtained for the hydrolysates varied from 26 to 2.5 kDa, depending on the level of hydrolysis. Further characterisation showed that, at the same level of hydrolysis, the chemical hydrolysates differed by their charges and hydrophobicity from those derived from enzymatic digestion. Analysis of the foaming properties showed, for both cases, that a limited degree of hydrolysis, around 3%, was sufficient to optimise the foaming properties of the isolate despite the different physicochemical properties of the peptides generated. The study of foaming properties at basic, neutral and acidic pHs showed that the hydrolysate solutions yielded dense foams which drained slowly and which maintained a very stable volume under the three pH conditions tested.

  12. Succinic acid production from lignocellulosic hydrolysate by Basfia succiniciproducens.

    PubMed

    Salvachúa, Davinia; Smith, Holly; St John, Peter C; Mohagheghi, Ali; Peterson, Darren J; Black, Brenna A; Dowe, Nancy; Beckham, Gregg T

    2016-08-01

    The production of chemicals alongside fuels will be essential to enhance the feasibility of lignocellulosic biorefineries. Succinic acid (SA), a naturally occurring C4-diacid, is a primary intermediate of the tricarboxylic acid cycle and a promising building block chemical that has received significant industrial attention. Basfia succiniciproducens is a relatively unexplored SA-producing bacterium with advantageous features such as broad substrate utilization, genetic tractability, and facultative anaerobic metabolism. Here B. succiniciproducens is evaluated in high xylose-content hydrolysates from corn stover and different synthetic media in batch fermentation. SA titers in hydrolysate at an initial sugar concentration of 60g/L reached up to 30g/L, with metabolic yields of 0.69g/g, and an overall productivity of 0.43g/L/h. These results demonstrate that B. succiniciproducens may be an attractive platform organism for bio-SA production from biomass hydrolysates. PMID:27179951

  13. Biodegradable nanoparticles for protein delivery: analysis of preparation conditions on particle morphology and protein loading, activity and sustained release properties.

    PubMed

    Coleman, Jason; Lowman, Anthony

    2012-01-01

    PLGA particles have been extensively used as a sustained drug-delivery system, but there are multiple drawbacks when delivering proteins. The focus of this work is to address the most significant disadvantages to the W/O/W double emulsion procedure and demonstrate that simple changes to this procedure can have significant changes to particle size and dispersity and considerable improvements to protein loading, activity and sustained active protein release. A systematic approach was taken to analyze the effects of the following variables: solvent miscibility (dichloromethane (DCM), ethyl acetate, acetone), homogenization speed (10 000-25 000 rpm), PLGA concentration (10-30 mg/ml) and additives in both the organic (sucrose acetate isobutyrate (SAIB)) and aqueous (bovine serum albumin (BSA)) phases. Increasing solvent miscibility decreased particle size, dispersity and protein denaturation, while maintaining adequate protein loading. Increasing solvent miscibility also lowered the impact of homogenization on particle size and dispersity and protein activity. Changes to PLGA concentration demonstrated a minimum impact on particle size and dispersity, but showed an inverse relationship between protein encapsulation efficiency and particle protein weight percent. Most particles tested provided sustained release of active protein over 60 days. Increasing solvent miscibility resulted in increases in the percent of active protein released. When subjected to synthesis conditions with DCM as the solvent, BSA as a stabilizer resulted in the maximum stabilization of protein at a concentration of 100 mg/ml. At this concentration, BSA allowed for increases in the total amount of active protein delivered for all three solvents. The benefit of SAIB was primarily increased protein loading.

  14. A comparative in vitro study of the digestibility of heat- and high pressure-induced gels prepared from industrial milk whey proteins

    NASA Astrophysics Data System (ADS)

    He, Jin-Song; Mu, Tai-Hua; Wang, Juan

    2013-06-01

    We undertook this study to compare the digestibility of heat- and high pressure-induced gels produced from whey protein isolate (WPI). To simulate in vivo gastrointestinal digestion of WPI gels, a pepsin-trypsin digestion system was used. The in vitro protein digestibility of WPI gels induced by high pressure (400 MPa and 30 min; P-gel) and those induced by heat (80°C and 30 min; H-gel) was compared using a protein concentration of 0.14 g mL-1. The in vitro protein digestibility of P-gels was significantly greater than that of H-gels (p<0.05). The size-exclusion chromatography profiles of the hydrolysates showed that the P-gel generated more and smaller peptides than natural WPI and H-gels. Furthermore, Sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis showed some soluble disulfide-mediated aggregation in the P-gel, while there was more insoluble aggregation in the H-gel than the P-gel. The P-gel was more sensitive to proteinase than the H-gel, which was related to the content of S-S bonds, and this in turn could be attributed to the differences in the gelation mechanism between the H-gel and P-gel.

  15. Influence of protein composition and hydrolysis method on intestinal absorption of protein in man.

    PubMed

    Keohane, P P; Grimble, G K; Brown, B; Spiller, R C; Silk, D B

    1985-09-01

    An intestinal perfusion technique has been used in normal human subjects to investigate the influence that starter protein composition and hydrolysis procedure have on absorption of amino acid residues from partial enzymic hydrolysates of whole protein. Five starter proteins were studied. Three (egg albumin, casein/soy/lactalbumin, and lactalbumin) were hydrolysed by papain, a second lactalbumin starter protein, and a meat/soy/lactalbumin blend were hydrolysed by a porcine pancreatic enzyme system. Irrespective of starter protein composition or hydrolysis method used, four amino acid residues (threonine, glutamic acid, phenylalanine, and histidine) were absorbed significantly faster from all hydrolysates compared with absorption from their equivalent free amino acid mixtures. In contrast, both starter protein composition and hydrolysis method influenced absorption characteristics of up to nine other amino acid residues. PMID:4029718

  16. Preparation and in vivo evaluation of a novel stabilized linker for 211At labeling of protein.

    PubMed

    Talanov, Vladimir S; Garmestani, Kayhan; Regino, Celeste A S; Milenic, Diane E; Plascjak, Paul S; Waldmann, Thomas A; Brechbiel, Martin W

    2006-05-01

    Significant improvement of in vivo stability of 211At-labeled radioimmunoconjugates achieved upon employment of a recently reported new linker, succinimidyl N-2-(4-[211At]astatophenethyl)succinamate (SAPS), prompted additional studies of its chemistry. The 211At radiolabeling of succinimidyl N-2-(4-tributylstannylphenethyl)succinamate (1) was noted to decline after storage at -15 degrees C for greater than 6 months. Compound 1 was found to degrade via a ring closure reaction with the formation of N-2-(4-tributylstannylphenethyl)succinimide (3), and a modified procedure for the preparation of 1 was developed. The N-methyl structural analog of 1, succinimidyl N-2-(4-tributylstannylphenethyl)-N-methyl succinamate (SPEMS), was synthesized to investigate the possibility of improving the stability of reagent-protein linkage chemistry. Radiolabeling of SPEMS with 211At generates succinimidyl N-2-(4-[211At]astatophenethyl)-N-methyl succinamate (Methyl-SAPS), with yields being consistent for greater than 1 year. Radiolabelings of 1 and SPEMS with 125I generated succinimidyl N-2-(4-[125I]iodophenethyl)succinamate (SIPS) and succinimidyl N-2-(4-[125I]iodophenethyl)-N-methyl succinamate (Methyl-SIPS), respectively, and showed no decline in yields. Methyl-SAPS, SAPS, Methyl-SIPS and SIPS were conjugated to Herceptin for a comparative assessment in LS-174T xenograft-bearing mice. The conjugates of Herceptin with Methyl-SAPS or Methyl-SIPS demonstrated immunoreactivity equivalent to if not superior to the SAPS and SIPS paired analogs. The in vivo studies also revealed that the N-methyl modification resulted in a superior statinated product. PMID:16720238

  17. Protein-loaded microspheres prepared by sequential adsorption of dextran sulphate and protamine on melamine formaldehyde core.

    PubMed

    Balabushevich, Nadezda G; Larionova, Natalia I

    2009-11-01

    Polyelectrolyte multilayer microspheres were fabricated by layer-by-layer self-assembly of a dextran sulphate and a protamine on melamine formaldehyde cores, followed by the partial decomposition of the core. Effects of pH on the encapsulation of proteins and enzymes with different physico-chemical properties (insulin, aprotinin, peroxidase, glucose oxidase (GOD), catalase (Cat)) in the prepared microspheres were then studied. This method of protein encapsulation demonstrated a high loading capacity and efficiency. The protein incorporation and release was regulated by the pH of the solution. Encapsulated enzymes retained a high specific activity depending on the amount of protein incorporated. Bienzyme system GOD/Cat immobilized in the microspheres was suitable for the glucose content assay.

  18. Absorbent alginate fibres modified with hydrolysed chitosan for wound care dressings--II. Pilot scale development.

    PubMed

    Sweeney, I R; Miraftab, M; Collyer, G

    2014-02-15

    Fibres have been used extensively in wound dressing applications as they provide a high surface area for absorption, ease of fabrication and softness. It is common practice for commercial wound dressings to be produced from natural materials, such a marine polysaccharides, as they are predominantly biocompatible, non-toxic, and often display bioactive properties, such as inherent antimicrobial activity. In this study hydrolysed chitosans were utilised as a sole coagulant for the production of alginate-chitosan fibres via a one-step, direct wet-spinning extrusion process. The levels of chitosan incorporated into the fibres were analysed quantitatively via elemental analysis and qualitatively by staining using Amido Black 10B. It was estimated that the fibres contained between 4.50 and 5.10% (wt.%) chitosan. The presence of chitosan improved tensile properties such as elongation and tenacity of the base alginate fibres. The increased incorporation of chitosan into the fibres also improved the absorption of the fibres in both saline and distilled water; reaching maximum of >30 g/g and >50 g/g, respectively. This work suggests that the observed hydrolysed chitosan content within the fibre may be optimal for the preparation of a novel fibre for wound care application.

  19. Development of silane-hydrolysate binder for UV-resistant thermal control coatings

    NASA Technical Reports Server (NTRS)

    Patterson, W. J.

    1981-01-01

    Detailed characterizaton and formulation studies were performed on a methyltriakoxysilane hydrolysate as a binder for thermal control coatings. The binder was optimized by varying hydrolysis temperature, time, catalyst type, and water concentration. The candidate coating formulations, based on this binder with TiO2 pigment, were optimized via a detailed series of sprayed test panels that included the parameters of binder/pigment ratio, ethanol content, pigment particle size, coating thickness and cure conditions. A typical optimized coating was prepared by acetic acid catalyzed hydrolysis of methyltriethoxysilane with 3.25 mol-equivalents of water over a 24 hour period at room temperature. The resulting hydrolysate was directly mixed with pre-milled TiO2 (12 grams pigment/26 grams binder) to yield a sprayable consistency. Panels were sprayed to result in a nominal cure coating thickness of 2 mils. Cure was affected by air drying for 24 hr at room temperature plus 72 hr at 150 F. These coatings are typically extremely tough and abrasion-resistant, with an absorptance (alpha) of 0.20 and emittance (e) of 0.89. No significant coating damage was observed in the mandrel bend test, even after exposure to thermal cycling from -160 to 160 F. Vacuum exposure of the coatings for 930 hours at 1 equivalent UV sun resulted in no visible degradation and no significant increase in absorptance.

  20. Polymersomes prepared from thermoresponsive fluorescent protein-polymer bioconjugates: capture of and report on drug and protein payloads.

    PubMed

    Wong, Chin Ken; Laos, Alistair J; Soeriyadi, Alexander H; Wiedenmann, Jörg; Curmi, Paul M G; Gooding, J Justin; Marquis, Christopher P; Stenzel, Martina H; Thordarson, Pall

    2015-04-27

    Polymersomes provide a good platform for targeted drug delivery and the creation of complex (bio)catalytically active systems for research in synthetic biology. To realize these applications requires both spatial control over the encapsulation components in these polymersomes and a means to report where the components are in the polymersomes. To address these twin challenges, we synthesized the protein-polymer bioconjugate PNIPAM-b-amilFP497 composed of thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) and a green-fluorescent protein variant (amilFP497). Above 37 °C, this bioconjugate forms polymersomes that can (co-)encapsulate the fluorescent drug doxorubicin and the fluorescent light-harvesting protein phycoerythrin 545 (PE545). Using fluorescence lifetime imaging microscopy and Förster resonance energy transfer (FLIM-FRET), we can distinguish the co-encapsulated PE545 protein inside the polymersome membrane while doxorubicin is found both in the polymersome core and membrane.

  1. Polymersomes prepared from thermoresponsive fluorescent protein-polymer bioconjugates: capture of and report on drug and protein payloads.

    PubMed

    Wong, Chin Ken; Laos, Alistair J; Soeriyadi, Alexander H; Wiedenmann, Jörg; Curmi, Paul M G; Gooding, J Justin; Marquis, Christopher P; Stenzel, Martina H; Thordarson, Pall

    2015-04-27

    Polymersomes provide a good platform for targeted drug delivery and the creation of complex (bio)catalytically active systems for research in synthetic biology. To realize these applications requires both spatial control over the encapsulation components in these polymersomes and a means to report where the components are in the polymersomes. To address these twin challenges, we synthesized the protein-polymer bioconjugate PNIPAM-b-amilFP497 composed of thermoresponsive poly(N-isopropylacrylamide) (PNIPAM) and a green-fluorescent protein variant (amilFP497). Above 37 °C, this bioconjugate forms polymersomes that can (co-)encapsulate the fluorescent drug doxorubicin and the fluorescent light-harvesting protein phycoerythrin 545 (PE545). Using fluorescence lifetime imaging microscopy and Förster resonance energy transfer (FLIM-FRET), we can distinguish the co-encapsulated PE545 protein inside the polymersome membrane while doxorubicin is found both in the polymersome core and membrane. PMID:25736460

  2. Complementary analysis of the hard and soft protein corona: sample preparation critically effects corona composition

    NASA Astrophysics Data System (ADS)

    Winzen, S.; Schoettler, S.; Baier, G.; Rosenauer, C.; Mailaender, V.; Landfester, K.; Mohr, K.

    2015-02-01

    Here we demonstrate how a complementary analysis of nanocapsule-protein interactions with and without application media allows gaining insights into the so called hard and soft protein corona. We have investigated how both human plasma and individual proteins (human serum albumin (HSA), apolipoprotein A-I (ApoA-I)) adsorb and interact with hydroxyethyl starch (HES) nanocapsules possessing different functionalities. To analyse the hard protein corona we used sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and a protein quantitation assay. No significant differences were observed with regards to the hard protein corona. For analysis of the soft protein corona we characterized the nanocapsule-protein interaction with isothermal titration calorimetry (ITC) and dynamic light scattering (DLS). DLS and ITC measurements revealed that a high amount of plasma proteins were adsorbed onto the capsules' surface. Although HSA was not detected in the hard protein corona, ITC measurements indicated the adsorption of an HSA amount similar to plasma with a low binding affinity and reaction heat. In contrast, only small amounts of ApoA-I protein adsorb to the capsules with high binding affinities. Through a comparison of these methods we have identified ApoA-I to be a component of the hard protein corona and HSA as a component of the soft corona. We demonstrate a pronounced difference in the protein corona observed depending on the type of characterization technique applied. As the biological identity of a particle is given by the protein corona it is crucial to use complementary characterization techniques to analyse different aspects of the protein corona.Here we demonstrate how a complementary analysis of nanocapsule-protein interactions with and without application media allows gaining insights into the so called hard and soft protein corona. We have investigated how both human plasma and individual proteins (human serum albumin (HSA), apolipoprotein A

  3. Xylitol bioproduction in hemicellulosic hydrolysate obtained from sorghum forage biomass.

    PubMed

    Camargo, Danielle; Sene, Luciane; Variz, Daniela Inês Loreto Saraiva; Felipe, Maria das Graças de Almeida

    2015-04-01

    This study evaluated the biotechnological production of xylitol from sorghum forage biomass. The yeast Candida guilliermondii was cultivated in hemicellulosic hydrolysates obtained from biomass of three sorghum varieties (A, B, and C). First, the biomass was chemically characterized and subjected to dilute acid hydrolysis to obtain the hemicellulosic hydrolysates which were vacuum-concentrated and detoxified with activated charcoal. The hemicellulosic hydrolysates (initial pH 5.5) were supplemented with nutrients, and fermentations were conducted in 125-mL Erlenmeyer flasks containing 50 mL medium, under 200 rpm, at 30 °C for 96 h. Fermentations were evaluated by determining the parameters xylitol yield (Y P/S ) and productivity (QP), as well as the activities of the enzymes xylose reductase (XR) and xylitol dehydrogenase (XDH). There was no significant difference among the three varieties with respect to the contents of cellulose, hemicellulose, and lignin, although differences were found in the hydrolysate fermentability. Maximum xylitol yield and productivity values for variety A were 0.35 g/g and 0.16 g/L.h(-1), respectively. It was coincident with XR (0.25 U/mg prot) and XDH (0.17 U/mg prot) maximum activities. Lower values were obtained for varieties B and C, which were 0.25 and 0.17 g/g for yield and 0.12 and 0.063 g/L.h(-1) for productivity.

  4. Development of Silane Hydrolysate Binder for Thermal-Control Coatings

    NASA Technical Reports Server (NTRS)

    Patterson, W. J.

    1983-01-01

    Technical report describes theoretical and experimental development of methyltriethoxysilane (MTES) hydrolysate binder for white, titanium dioxidepigmented thermal-control coatings often needed on satellites. New coating is tougher and more abrasion-resistant than conventional coating, S-13G, which comprises zinc oxide in hydroxyl-therminated dimethylsiloxane binder.

  5. A quantitative assessment of glycolipid and protein associated with paired helical filament preparations from Alzheimer's diseased brain.

    PubMed

    Goux, Warren J.; Liu, Bingcam; Shumburo, Abdurahman M.; Parikh, Samir; Sparkman, Dennis R.

    2001-10-01

    Protease resistant paired helical filaments (prcPHF) can be isolated from the brains of Alzheimer's diseased patients. A second type of PHF, A68 PHF, may be extracted in soluble form from brain homogenate and induced to form filaments in vitro. Here we use a variety of analytical techniques to assess the protein, carbohydrate and fatty acid composition of prcPHF and A68 PHF. High-field ^1H NMR of both PHF preparations display similar fatty acid and carbohydrate proton resonances, consistent with the presence of a structurally similar glycolipid. Carbohydrate analysis showed that both preparations contained greater than 82% less than 12% C16:1 was significantly lower in A68 PHF than in prcPHF, both preparations contained otherwise similar fatty acid profiles with the most abundant lipid component being oleic acid (C18:1, 29.3 +/- 9.0%) followed by palmitic (C16:0, 28.5 +/- 5.6%) 17.6 +/- 7.5%) preparations revealed a profile reasonably consistent with that previously determined for PHF-tau but significantly higher in glycine and lower in lysine than would be predicted from the cDNA sequence. On a weight per cent basis, protein accounted for about 51% A68 PHF samples but only about 10% Carbohydrate and fatty acid accounted for about 39% A68 PHF samples but 74% preparations showed strong correlations between the total amount of tau protein and fatty acid. These results suggest that a glycolipid component forms an integral part of the PHF structure.

  6. Protein immobilization on Ni(II) ion patterns prepared by microcontact printing and dip-pen nanolithography.

    PubMed

    Wu, Chien-Ching; Reinhoudt, David N; Otto, Cees; Velders, Aldrik H; Subramaniam, Vinod

    2010-02-23

    An indirect method of protein patterning by using Ni(II) ion templates for immobilization via a specific metal-protein interaction is described. A nitrilotriacetic acid (NTA)-terminated self-assembled monolayer (SAM) allows oriented binding of histidine-tagged proteins via complexation with late first-row transition metal ions, such as Ni(II). Patterns of nickel(II) ions were prepared on NTA SAM-functionalized glass slides by microcontact printing (microCP) and dip-pen nanolithography (DPN) to obtain micrometer and submicrometer scale patterns. Consecutive dipping of the slides in 6His-protein solutions resulted in the formation of protein patterns, as was subsequently proven by AFM and confocal fluorescence microscopy. This indirect method prevents denaturation of fragile biomolecules caused by direct printing or writing of proteins. Moreover, it yields well-defined patterned monolayers of proteins and, in principle, is indifferent for biomolecules with a high molecular weight. This approach also enabled us to characterize the transfer of Ni(II) ions on fundamental parameters of DPN, such as writing speeds and tip-surface contact times, while writing with the smallest possible ink "molecules" (i.e., metal ions). PMID:20104881

  7. Enzyme Hydrolysates from Stichopus horrens as a New Source for Angiotensin-Converting Enzyme Inhibitory Peptides

    PubMed Central

    Forghani, Bita; Ebrahimpour, Afshin; Bakar, Jamilah; Abdul Hamid, Azizah; Hassan, Zaiton; Saari, Nazamid

    2012-01-01

    Stichopus horrens flesh was explored as a potential source for generating peptides with angiotensin-converting enzyme (ACE) inhibitory capacity using 6 proteases, namely alcalase, flavourzyme, trypsin, papain, bromelain, and protamex. Degree of hydrolysis (DH) and peptide profiling (SDS-PAGE) of Stichopus horrens hydrolysates (SHHs) was also assessed. Alcalase hydrolysate showed the highest DH value (39.8%) followed by flavourzyme hydrolysate (32.7%). Overall, alcalase hydrolysate exhibited the highest ACE inhibitory activity (IC50 value of 0.41 mg/mL) followed by flavourzyme hydrolysate (IC50 value of 2.24 mg/mL), trypsin hydrolysate (IC50 value of 2.28 mg/mL), papain hydrolysate (IC50 value of 2.48 mg/mL), bromelain hydrolysate (IC50 value of 4.21 mg/mL), and protamex hydrolysate (IC50 value of 6.38 mg/mL). The SDS-PAGE results showed that alcalase hydrolysate represented a unique pattern compared to others, which yielded potent ACE inhibitory peptides with molecular weight distribution lower than 20 kDa. The evaluation of the relationship between DH and IC50 values of alcalase and flavourzyme hydrolysates revealed that the trend between those parameters was related to the type of the protease used. We concluded that the tested SHHs would be used as a potential source of functional ACE inhibitory peptides for physiological benefits. PMID:22927875

  8. Complementary analysis of the hard and soft protein corona: sample preparation critically effects corona composition.

    PubMed

    Winzen, S; Schoettler, S; Baier, G; Rosenauer, C; Mailaender, V; Landfester, K; Mohr, K

    2015-02-21

    Here we demonstrate how a complementary analysis of nanocapsule-protein interactions with and without application media allows gaining insights into the so called hard and soft protein corona. We have investigated how both human plasma and individual proteins (human serum albumin (HSA), apolipoprotein A-I (ApoA-I)) adsorb and interact with hydroxyethyl starch (HES) nanocapsules possessing different functionalities. To analyse the hard protein corona we used sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and a protein quantitation assay. No significant differences were observed with regards to the hard protein corona. For analysis of the soft protein corona we characterized the nanocapsule-protein interaction with isothermal titration calorimetry (ITC) and dynamic light scattering (DLS). DLS and ITC measurements revealed that a high amount of plasma proteins were adsorbed onto the capsules' surface. Although HSA was not detected in the hard protein corona, ITC measurements indicated the adsorption of an HSA amount similar to plasma with a low binding affinity and reaction heat. In contrast, only small amounts of ApoA-I protein adsorb to the capsules with high binding affinities. Through a comparison of these methods we have identified ApoA-I to be a component of the hard protein corona and HSA as a component of the soft corona. We demonstrate a pronounced difference in the protein corona observed depending on the type of characterization technique applied. As the biological identity of a particle is given by the protein corona it is crucial to use complementary characterization techniques to analyse different aspects of the protein corona.

  9. Origin of D-amino acids detected in the acid hydrolysates of purified Escherichia coli β-galactosidase.

    PubMed

    Miyamoto, Tetsuya; Sekine, Masae; Ogawa, Tetsuhiro; Hidaka, Makoto; Homma, Hiroshi; Masaki, Haruhiko

    2015-12-10

    In previous report, we detected D-amino acids in the acid hydrolysates of purified recombinant β-galactosidase. Here, we employed a deuterium-hydrogen exchange method to discriminate innate D-amino acids from those generated during hydrolytic incubation. After hydrolysis of β-galactosidase in DCl/D2O, amino acids were derivatized with NBD-F and separated on a reverse-phase column, followed by liquid chromatography-tandem mass spectrometry equipped with a chiral column. Our results show an absence of innate D-amino acid residues in the protein and suggest that the protein undergoes isomerization during a very early stage of hydrolytic incubation.

  10. Identification of a NFκB inhibitory peptide from tryptic β-casein hydrolysate.

    PubMed

    Malinowski, J; Klempt, M; Clawin-Rädecker, I; Lorenzen, P Chr; Meisel, H

    2014-12-15

    Several bioactive peptides are encrypted within the sequence of major milk proteins, requiring enzymatic proteolysis for release and activation. The present study aimed at the identification of potential anti-inflammatory activities in tryptic hydrolysates of bovine β-casein. Inflammatory processes involve in most cases an activation of Nuclear factor Kappa-light-chain enhancer of activated B cells (NFκB), which is a pro-inflammatory transcription factor of several genes. Hence, a NFκB reporter cell line was established, and TNF-α mediated activation of NFκB was used as a measurement. Bovine β-casein (β-CN) was hydrolysed by trypsin and fractionated by ultrafiltration. Total proteolysate as well as the fraction containing peptides between 1 and 5 kDa showed an inhibitory effect in the cell-based assay, while the fraction containing molecules smaller than 1 kDa did not. This anti-inflammatory effect was ascribed to a group of large, hydrophobic peptides, which were identified using LC-MS. The main peptide was synthesised and showed a significant anti-inflammatory effect in HEK(nfkb-RE)-cells. Thus, for the first time, a casein-derived peptide having an anti-inflammatory effect in vitro has been identified.

  11. Heavy Metal Complexation of Thiol-Containing Peptides from Soy Glycinin Hydrolysates

    PubMed Central

    Ding, Xiuzhen; Hua, Yufei; Chen, Yeming; Zhang, Caimeng; Kong, Xiangzhen

    2015-01-01

    Many thiol-containing molecules show heavy metal complexation ability and are used as antidotes. In this study, the potential function associated with thiol-containing peptides (TCPs) from soy protein hydrolysates as natural detoxicants for heavy metals is reported. TCPs enriched by Thiopropyl-Sepharose 6B covalent chromatography had different molecular weight distributions as well as different numbers of proton dissociable groups, depending on the proteases and degree of hydrolysis. The major contribution of sulfhydryl groups was confirmed by the largest pH decrease between 8.0 and 8.5 of the pH titration curves. The complexation of TCPs with heavy metalswas evaluated by stability constants (βn) of TCP-metal complexes whose stoichiometry was found to be 1:1 (ML) and 1:2 (ML2). TCPs from degree of hydrolysis of 25% hydrolysates gave high affinities towards Hg2+, Cd2+, and Pb2+ (giving similar or even bigger lgβ values than that of glutathione). A significantly positive correlation was found between the logarithm of stability constants for ML2 (lgβ2) and the sulfhydryl group content. Molecular weight distribution of TCPs affected the complexation with Pb2+ notably more than Hg2+ and Cd2+. These results suggest that soy TCPs have the potential to be used in the formulation of functional foods to counteract heavy metal accumulation in humans. PMID:25867477

  12. Effect of fermentation conditions on L-lactic acid production from soybean straw hydrolysate.

    PubMed

    Wang, Juan; Wang, Qunhui; Xu, Zhong; Zhang, Wenyu; Xiang, Juan

    2015-01-01

    Four types of straw, namely, soybean, wheat, corn, and rice, were investigated for use in lactic acid production. These straws were mainly composed of cellulose, hemicellulose, and lignin. After pretreatment with ammonia, the cellulose content increased, whereas the hemicellulose and lignin contents decreased. Analytical results also showed that the liquid enzymatic hydrolysates were primarily composed of glucose, xylose, and cellobiose. Preliminary experiments showed that a higher lactic acid concentration could be obtained from the wheat and soybean straw. However, soybean straw was chosen as the substrate for lactic acid production owing to its high protein content. The maximum lactic acid yield (0.8 g/g) and lactic acid productivity (0.61 g/(l/h)) were obtained with an initial reducing sugar concentration of 35 g/l at 30°C when using Lactobacillus casei (10% inoculum) for a 42 h fermentation period. Thus, the experimental results demonstrated the feasibility of using a soybean straw enzymatic hydrolysate as a substrate for lactic acid production. PMID:25152056

  13. Effect of fermentation conditions on L-lactic acid production from soybean straw hydrolysate.

    PubMed

    Wang, Juan; Wang, Qunhui; Xu, Zhong; Zhang, Wenyu; Xiang, Juan

    2015-01-01

    Four types of straw, namely, soybean, wheat, corn, and rice, were investigated for use in lactic acid production. These straws were mainly composed of cellulose, hemicellulose, and lignin. After pretreatment with ammonia, the cellulose content increased, whereas the hemicellulose and lignin contents decreased. Analytical results also showed that the liquid enzymatic hydrolysates were primarily composed of glucose, xylose, and cellobiose. Preliminary experiments showed that a higher lactic acid concentration could be obtained from the wheat and soybean straw. However, soybean straw was chosen as the substrate for lactic acid production owing to its high protein content. The maximum lactic acid yield (0.8 g/g) and lactic acid productivity (0.61 g/(l/h)) were obtained with an initial reducing sugar concentration of 35 g/l at 30°C when using Lactobacillus casei (10% inoculum) for a 42 h fermentation period. Thus, the experimental results demonstrated the feasibility of using a soybean straw enzymatic hydrolysate as a substrate for lactic acid production.

  14. Enhanced xylose fermentation and hydrolysate inhibitor tolerance of Scheffersomyces shehatae for efficient ethanol production from non-detoxified lignocellulosic hydrolysate.

    PubMed

    Senatham, Srisuda; Chamduang, Thada; Kaewchingduang, Yotin; Thammasittirong, Anon; Srisodsuk, Malee; Elliston, Adam; Roberts, Ian N; Waldron, Keith W; Thammasittirong, Sutticha Na-Ranong

    2016-01-01

    Effective conversion of xylose into ethanol is important for lignocellulosic ethanol production. In the present study, UV-C mutagenesis was used to improve the efficiency of xylose fermentation. The mutated Scheffersomyces shehatae strain TTC79 fermented glucose as efficiently and xylose more efficiently, producing a higher ethanol concentration than the wild-type. A maximum ethanol concentration of 29.04 g/L was produced from 71.31 g/L xylose, which was 58.95 % higher than that of the wild-type. This mutant also displayed significantly improved hydrolysate inhibitors tolerance and increased ethanol production from non-detoxified lignocellulosic hydrolysates. The ethanol yield, productivity and theoretical yield by TTC79 from sugarcane bagasse hydrolysate were 0.46 g/g, 0.20 g/L/h and 90.61 %, respectively, while the corresponding values for the wild-type were 0.20 g/g, 0.04 g/L/h and 39.20 %, respectively. These results demonstrate that S. shehatae TTC79 is a useful non-recombinant strain, combining efficient xylose consumption and high inhibitor tolerance, with potential for application in ethanol production from lignocellulose hydrolysates. PMID:27462488

  15. Pressurized liquid extraction-assisted mussel cytosol preparation for the determination of metals bound to metallothionein-like proteins.

    PubMed

    Santiago-Rivas, Sandra; Moreda-Piñeiro, Antonio; Bermejo-Barrera, Pilar; Moreda-Piñeiro, Jorge; Alonso-Rodríguez, Elia; Muniategui-Lorenzo, Soledad; López-Mahía, Purificación; Prada-Rodríguez, Darío

    2007-11-01

    The possibilities of pressurized liquid extraction (PLE) have been novelty tested to assist the cytosol preparation from wet mussel soft tissue before the determination of metals bound to metallothionein-like proteins (MLPs). Results obtained after PLE were compared with those obtained after a classical blending procedure for mussel cytosolic preparation. Isoforms MLP-1 (retention time of 4.1 min) and MLP-2 (retention time of 7.4 min) were separated by anion exchange high-performance liquid chromatography (HPLC) and the concentrations of Ba, Cu, Mn, Sr and Zn bound to MLP isoforms were directly measured by inductively coupled plasma-atomic emission spectrometry (ICP-OES) as a multi-element detector. The optimized PLE-assisted mussel cytosol preparation has consisted of one extraction cycle at room temperature and 1500 psi for 2 min. Since separation between the solid mussel residue and the extract (cytosol) is performed by the PLE system, the cytosol preparation method is faster than conventional cytosol preparation methods by cutting/blending using Ultraturrax or Stomacher devices. PMID:17950055

  16. Lipid Production of Heterotrophic Chlorella sp. from Hydrolysate Mixtures of Lipid-Extracted Microalgal Biomass Residues and Molasses.

    PubMed

    Zheng, Hongli; Ma, Xiaochen; Gao, Zhen; Wan, Yiqin; Min, Min; Zhou, Wenguang; Li, Yun; Liu, Yuhuan; Huang, He; Chen, Paul; Ruan, Roger

    2015-10-01

    This study investigated the feasibility of lipid production of Chlorella sp. from waste materials. Lipid-extracted microalgal biomass residues (LMBRs) and molasses were hydrolyzed, and their hydrolysates were analyzed. Five different hydrolysate mixture ratios (w/w) of LMBRs/molasses (1/0, 1/1, 1/4, 1/9, and 0/1) were used to cultivate Chlorella sp. The results showed that carbohydrate and protein were the two main compounds in the LMBRs, and carbohydrate was the main compound in the molasses. The highest biomass concentration of 5.58 g/L, Y biomass/sugars of 0.59 g/g, lipid productivity of 335 mg/L/day, and Y lipids/sugars of 0.25 g/g were obtained at the hydrolysate mixture ratio of LMBRs/molasses of 1/4. High C/N ratio promoted the conversion of sugars into lipids. The lipids extracted from Chlorella sp. shared similar lipid profile of soybean oil and is therefore a potential viable biodiesel feedstock. These results showed that Chlorella sp. can utilize mixed sugars and amino acids from LMBRs and molasses to accumulate lipids efficiently, thus reducing the cost of microalgal biodiesel production and improving its economic viability.

  17. Levanase from Bacillus subtilis hydrolyses β-2,6 fructosyl bonds in bacterial levans and in grass fructans.

    PubMed

    Jensen, Susanne L; Diemer, Mikkel B; Lundmark, Maria; Larsen, Flemming H; Blennow, Andreas; Mogensen, Helle K; Nielsen, Tom H

    2016-04-01

    A Levanase, LevB, from Bacillus subtilis 168, was expressed as a His6-tagged protein in Escherichia coli. The enzyme was purified and characterised for its activity and substrate specificity. LevB has a pH optimum of 6.0-6.5 and a maximum observed specific activity of 3 U mg(-1) using levan from Erwinia herbicola as substrate. Hydrolysis products were analysed by HPAEC, TLC, and NMR using chicory root inulin, mixed linkage fructans purified from ryegrass (Lolium perenne) and levan from E. herbicola as substrates. This revealed that LevB is an endolevanase that selectively cleaves the (β-2,6) fructosyl bonds and does not hydrolyse inulin. Ryegrass fructans and bacterial levan was hydrolysed partially releasing oligosaccharides, but together with exoinulinase, LevB hydrolysed both ryegrass fructans and bacterial levan to near completion. We suggest that LevB can be used as a tool to achieve more structural information on complex fructans and to achieve complete degradation and quantification of mixed linkage fructans. PMID:26773563

  18. Hydrolysable tannin fed to entire male pigs affects intestinal production, tissue deposition and hepatic clearance of skatole.

    PubMed

    Čandek-Potokar, M; Škrlep, M; Batorek Lukač, N; Zamaratskaia, G; Prevolnik Povše, M; Velikonja Bolta, Š; Kubale, V; Bee, G

    2015-05-01

    The effect of adding hydrolysable tannins to the diet of fattening boars was studied. Performance, reproductive organ weights, salivary gland morphology, boar taint compounds and skatole metabolism were evaluated. At 123 days of age and 52 ± 6 kg liveweight, 24 Landrace × Large White boars were assigned within a litter to four treatment groups: control (T0 fed mixture with 13.2 MJ/kg, 17.5% crude proteins) and three experimental diets for which the T0 diet was supplemented with 1%, 2% and 3% of hydrolysable tannin-rich extract (T1, T2 and T3, respectively). Pigs were kept individually with ad libitum access to feed and water and slaughtered at 193 days of age and 122 ± 10 kg liveweight. Adding hydrolysable tannins to the diet had no negative effect on growth performance at 1% and 2%, whereas the 3% inclusion reduced feed intake and resulted in an adaptive response of the salivary glands (particularly parotid gland hypertrophy). Relative to T0, fat tissue skatole concentration was increased in the T1 group, but was similar in T2 and T3. Across treatments tissue skatole concentrations were proportional to the activity of hepatic CYP450. The results indicate the potential of tannin supplementation to reduce boar taint although further investigations are needed in order to establishing optimal dosage. PMID:25890671

  19. Protein adsorption resistance of PVP-modified polyurethane film prepared by surface-initiated atom transfer radical polymerization

    NASA Astrophysics Data System (ADS)

    Yuan, Huihui; Qian, Bin; Zhang, Wei; Lan, Minbo

    2016-02-01

    An anti-fouling surface of polyurethane (PU) film grafted with Poly(N-vinylpyrrolidone) (PVP) was prepared through surface-initiated atom transfer radical polymerization (SI-ATRP). And the polymerization time was investigated to obtain PU films with PVP brushes of different lengths. The surface properties and protein adsorption of modified PU films were evaluated. The results showed that the hydrophilicity of PU-PVP films were improved with the increase of polymerization time, which was not positive correlation with the surface roughness due to the brush structure. Additionally, the protein resistance performance was promoted when prolonging the polymerization time. The best antifouling PU-PVP (6.0 h) film reduced the adsoption level of bovine serum albumin (BSA), lysozyme (LYS), and brovin serum fibrinogen (BFG) by 93.4%, 68.3%, 85.6%, respectively, compared to the unmodified PU film. The competitive adsorption of three proteins indicated that LYS preferentially adsorbed on the modified PU film, while BFG had the lowest adsorption selectivity. And the amount of BFG on PU-PVP (6.0 h) film reduced greatly to 0.08 μg/cm2, which was almost one-tenth of its adsorption from the single-protein system. Presented results suggested that both hydrophilicity and surface roughness might be the important factors in all cases of protein adsorption, and the competitive or selective adsorption might be related to the size of the proteins, especially on the non-charged films.

  20. [Preparation and characterization of the recombinant protein containing immunomimetic peptide of benzo[a]pyrene].

    PubMed

    Apal'ko, S V; Lunin, V G; Filipenko, M L; Matveeva, V A; Liashchuk, A M; Lavrova, N V; Sherina, E A; Aver'ianov, A V; Kostianko, M V; Glushkov, A N

    2011-01-01

    Two recombinant plasmids were constructed. The first plasmid contained the hybrid gene composed of immunomimetic peptide of benzo[a]pyrene, of the protein pIII of bacteriophage M13 and of cellulose binding domain encoding sequences. The second plasmid contained the hybrid gene composed of the signal peptide of the protein pIII of bacteriophage M13, of immunomimetic peptide of benzo[a]pyrene, of the protein pill of bacteriophage M13 and of cellulose binding domain sequences. The obtained recombinant plasmids were used in expression of chimeric protein containing immunomimetic peptide ofbenzo[a]pyrene based on strain E. coli M15. The lack of the recombinant protein expression using first plasmid was demonstrated. In the same time, it was shown that accumulation of recombinant protein contained immunomimetic peptide with signal peptide of the protein pIIIl of bacteriophage was present. This chimeric protein was produced in "mature" (without signal peptide) and "unprocessing" (with signal peptide) forms. Using the Western-blot analysis, it was shown that the "mature" form only specifically bound to the B2 monoclonal antibody against benzo[a]pyrene. Thus, we expressed, purified, and characterized the recombinant protein containing immunomimetic peptide of benzo[a]pyrene.

  1. Hydrolysates of citrus plants stimulate melanogenesis protecting against UV-induced dermal damage.

    PubMed

    Chiang, Hsiu-Mei; Lin, Jen-Wen; Hsiao, Pei-Ling; Tsai, Shang-Yuan; Wen, Kuo-Ching

    2011-04-01

    The sun-tanning process occurs as a spontaneous response to ultraviolet (UV) irradiation. UV will induce tanning and DNA damage, processes that can lead to photoaging and skin disorders such as hyperpigmentation and cancer. The pigment melanin protects skin from UV damage; therefore, an efficient melanin-promoting suntan lotion could be highly beneficial. In this study, a process was developed to increase the content of naringenin in citrus extracts and to determine whether a higher naringenin content of citrus would induce melanogenesis. Melanin content and tyrosinase expression in mouse B16 melanoma cells were assayed after treatment with citrus plant extracts and their hydrolysates. The results indicate that hydrolysis increased the naringenin content in citrus extracts and that citrus preparations stimulated cellular melanogenesis and tyrosinase expression. It is suggested that this method is applicable to the industrial production of melanin-promoting suntan lotions with antiphotocarcinogenic properties derived from citrus rind and citrus products.

  2. Protein preparation, crystallization and preliminary X-ray crystallographic analysis of SMU.961 protein from the caries pathogen Streptococcus mutans.

    PubMed

    Gao, Xiong-Zhuo; Li, Lan-Fen; Su, Xiao-Dong; Zhao, XiaoJun; Liang, Yu-He

    2007-10-01

    The smu.961 gene encodes a putative protein of 183 residues in Streptococcus mutans, a major pathogen in human dental caries. The gene was cloned into expression vector pET28a and expressed in a substantial quantity in Escherichia coli strain BL21 (DE3) with a His tag at its N-terminus. The recombinant protein SMU.961 was purified to homogeneity in a two-step procedure consisting of Ni2+-chelating and size-exclusion chromatography. Crystals suitable for X-ray diffraction were obtained by the hanging-drop vapour-diffusion method and diffracted to 2.9 A resolution at beamline I911-3, MAX-II-lab, Sweden. The crystal belonged to space group C2, with unit-cell parameters a = 98.62, b = 73.73, c = 184.73 A, beta = 98.82 degrees.

  3. Functional and antioxidant properties of hydrolysates of sardine (S. pilchardus) and horse mackerel (T. mediterraneus) for the microencapsulation of fish oil by spray-drying.

    PubMed

    Morales-Medina, R; Tamm, F; Guadix, A M; Guadix, E M; Drusch, S

    2016-03-01

    The functionality of fish protein hydrolysates (FPH) for the microencapsulation of fish oil was investigated. Muscle protein from sardine (Sardina pilchardus) and horse mackerel (Trachurus mediterraneus) was hydrolysed using Alcalase or trypsin. Physically stable emulsions suitable for spray-drying were obtained when using FPH with a degree of hydrolysis of 5%. Microencapsulation efficiency amounted to 98±0.1% and oxidative stability of the encapsulated oil over a period of twelve weeks was in a similar range as it is reported for other matrix systems. Therefore, the suitability of FPH for use in spray-dried emulsions has been shown for the first time. Since no clear correlation between the antioxidative activity of the FPH and the course of lipid oxidation could be established future research is required to more specifically characterise the molecular structure of the peptides and its impact on protein alteration and role in lipid oxidation.

  4. Remodeling of the plasma membrane in preparation for sperm-egg recognition: roles of acrosomal proteins.

    PubMed

    Tanphaichitr, Nongnuj; Kongmanas, Kessiri; Kruevaisayawan, Hathairat; Saewu, Arpornrad; Sugeng, Clarissa; Fernandes, Jason; Souda, Puneet; Angel, Jonathan B; Faull, Kym F; Aitken, R John; Whitelegge, Julian; Hardy, Daniel; Berger, Trish; Baker, Mark

    2015-01-01

    The interaction of sperm with the egg's extracellular matrix, the zona pellucida (ZP) is the first step of the union between male and female gametes. The molecular mechanisms of this process have been studied for the past six decades with the results obtained being both interesting and confusing. In this article, we describe our recent work, which attempts to address two lines of questions from previous studies. First, because there are numerous ZP binding proteins reported by various researchers, how do these proteins act together in sperm-ZP interaction? Second, why do a number of acrosomal proteins have ZP affinity? Are they involved mainly in the initial sperm-ZP binding or rather in anchoring acrosome reacting/reacted spermatozoa to the ZP? Our studies reveal that a number of ZP binding proteins and chaperones, extracted from the anterior sperm head plasma membrane, coexist as high molecular weight (HMW) complexes, and that these complexes in capacitated spermatozoa have preferential ability to bind to the ZP. Zonadhesin (ZAN), known as an acrosomal protein with ZP affinity, is one of these proteins in the HMW complexes. Immunoprecipitation indicates that ZAN interacts with other acrosomal proteins, proacrosin/acrosin and sp32 (ACRBP), also present in the HMW complexes. Immunodetection of ZAN and proacrosin/acrosin on spermatozoa further indicates that both proteins traffic to the sperm head surface during capacitation where the sperm acrosomal matrix is still intact, and therefore they are likely involved in the initial sperm-ZP binding step.

  5. Preparation, composition and functional properties of pennycress (Thlaspi arvense L.) seed protein isolates

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study evaluated two methods, saline extraction (SE) and conventional acid precipitation (AP), to recover proteins from pennycress (Thlaspi arvense L.) seed meal. SE was done using 0.1 M NaCl at 50ºC while AP involved alkaline extraction (pH 10) first followed by protein precipitation at pH 4. C...

  6. Protein preparation, crystallization and preliminary X-ray crystallographic analysis of SMU.961 protein from the caries pathogen Streptococcus mutans

    SciTech Connect

    Gao, Xiong-Zhuo; Li, Lan-Fen; Su, Xiao-Dong; Zhao, XiaoJun; Liang, Yu-He

    2007-10-01

    The SMU.961 protein from S. mutans was crystallized and preliminary characterization of the crystals, which diffracted to 2.9 Å resolution, shows them to belong to space group C2. The smu.961 gene encodes a putative protein of 183 residues in Streptococcus mutans, a major pathogen in human dental caries. The gene was cloned into expression vector pET28a and expressed in a substantial quantity in Escherichia coli strain BL21 (DE3) with a His tag at its N-terminus. The recombinant protein SMU.961 was purified to homogeneity in a two-step procedure consisting of Ni{sup 2+}-chelating and size-exclusion chromatography. Crystals suitable for X-ray diffraction were obtained by the hanging-drop vapour-diffusion method and diffracted to 2.9 Å resolution at beamline I911-3, MAX-II-lab, Sweden. The crystal belonged to space group C2, with unit-cell parameters a = 98.62, b = 73.73, c = 184.73 Å, β = 98.82°.

  7. A Biocompatible and Biodegradable Protein Hydrogel with Green and Red Autofluorescence: Preparation, Characterization and In Vivo Biodegradation Tracking and Modeling

    NASA Astrophysics Data System (ADS)

    Ma, Xiaoyu; Sun, Xiangcheng; Hargrove, Derek; Chen, Jun; Song, Donghui; Dong, Qiuchen; Lu, Xiuling; Fan, Tai-Hsi; Fu, Youjun; Lei, Yu

    2016-01-01

    Because of its good biocompatibility and biodegradability, albumins such as bovine serum albumin (BSA) and human serum albumin (HSA) have found a wide range of biomedical applications. Herein, we report that glutaraldehyde cross-linked BSA (or HSA) forms a novel fluorescent biological hydrogel, exhibiting new green and red autofluorescence in vitro and in vivo without the use of any additional fluorescent labels. UV-vis spectra studies, in conjunction with the fluorescence spectra studies including emission, excitation and synchronous scans, indicated that three classes of fluorescent compounds are presumably formed during the gelation process. SEM, FTIR and mechanical tests were further employed to investigate the morphology, the specific chemical structures and the mechanical strength of the as-prepared autofluorescent hydrogel, respectively. Its biocompatibility and biodegradability were also demonstrated through extensive in vitro and in vivo studies. More interestingly, the strong red autofluorescence of the as-prepared hydrogel allows for conveniently and non-invasively tracking and modeling its in vivo degradation based on the time-dependent fluorescent images of mice. A mathematical model was proposed and was in good agreement with the experimental results. The developed facile strategy to prepare novel biocompatible and biodegradable autofluorescent protein hydrogels could significantly expand the scope of protein hydrogels in biomedical applications.

  8. Effect of Different Tumbling Marination Methods and Time on the Water Status and Protein Properties of Prepared Pork Chops

    PubMed Central

    Gao, Tian; Li, Jiaolong; Zhang, Lin; Jiang, Yun; Yin, Maowen; Liu, Yang; Gao, Feng; Zhou, Guanghong

    2015-01-01

    The combined effect of tumbling marination methods (vacuum continuous tumbling marination, CT; vacuum intermittent tumbling marination, IT) and effective tumbling time (4, 6, 8, and 10 h) on the water status and protein properties of prepared pork chops was investigated. Results showed that regardless of tumbling time, CT method significantly decreased the muscle fiber diameter (MD) and significantly increased the total moisture content, product yield, salt soluble proteins (SSP) solubility, immobilized water component (p<0.05) compared with IT method. With the effective tumbling time increased from 4 h to 10 h, the fat content and the MD were significantly decreased (p<0.05), whereas the SSP solubility of prepared pork chops increased firstly and then decreased. Besides, an interactive effect between CT method and effective tumbling time was also observed for the chemical composition and proportion of immobilized water (p<0.05). These results demonstrated that CT method of 8 h was the most beneficial for improving the muscle structure and water distribution status, increasing the water-binding capacity and accelerating the marinade efficiency of pork chops; and thus, it should be chosen as the most optimal treatment method for the processing production of prepared pork chops. PMID:26104408

  9. A Biocompatible and Biodegradable Protein Hydrogel with Green and Red Autofluorescence: Preparation, Characterization and In Vivo Biodegradation Tracking and Modeling

    PubMed Central

    Ma, Xiaoyu; Sun, Xiangcheng; Hargrove, Derek; Chen, Jun; Song, Donghui; Dong, Qiuchen; Lu, Xiuling; Fan, Tai-Hsi; Fu, Youjun; Lei, Yu

    2016-01-01

    Because of its good biocompatibility and biodegradability, albumins such as bovine serum albumin (BSA) and human serum albumin (HSA) have found a wide range of biomedical applications. Herein, we report that glutaraldehyde cross-linked BSA (or HSA) forms a novel fluorescent biological hydrogel, exhibiting new green and red autofluorescence in vitro and in vivo without the use of any additional fluorescent labels. UV-vis spectra studies, in conjunction with the fluorescence spectra studies including emission, excitation and synchronous scans, indicated that three classes of fluorescent compounds are presumably formed during the gelation process. SEM, FTIR and mechanical tests were further employed to investigate the morphology, the specific chemical structures and the mechanical strength of the as-prepared autofluorescent hydrogel, respectively. Its biocompatibility and biodegradability were also demonstrated through extensive in vitro and in vivo studies. More interestingly, the strong red autofluorescence of the as-prepared hydrogel allows for conveniently and non-invasively tracking and modeling its in vivo degradation based on the time-dependent fluorescent images of mice. A mathematical model was proposed and was in good agreement with the experimental results. The developed facile strategy to prepare novel biocompatible and biodegradable autofluorescent protein hydrogels could significantly expand the scope of protein hydrogels in biomedical applications. PMID:26813916

  10. imFASP: An integrated approach combining in-situ filter-aided sample pretreatment with microwave-assisted protein digestion for fast and efficient proteome sample preparation.

    PubMed

    Zhao, Qun; Fang, Fei; Wu, Ci; Wu, Qi; Liang, Yu; Liang, Zhen; Zhang, Lihua; Zhang, Yukui

    2016-03-17

    An integrated sample preparation method, termed "imFASP", which combined in-situ filter-aided sample pretreatment and microwave-assisted trypsin digestion, was developed for preparation of microgram and even nanogram amounts of complex protein samples with high efficiency in 1 h. For imFASP method, proteins dissolved in 8 M urea were loaded onto a filter device with molecular weight cut off (MWCO) as 10 kDa, followed by in-situ protein preconcentration, denaturation, reduction, alkylation, and microwave-assisted tryptic digestion. Compared with traditional in-solution sample preparation method, imFASP method generated more protein and peptide identifications (IDs) from preparation of 45 μg Escherichia coli protein sample due to the higher efficiency, and the sample preparation throughput was significantly improved by 14 times (1 h vs. 15 h). More importantly, when the starting amounts of E. coli cell lysate decreased to nanogram level (50-500 ng), the protein and peptide identified by imFASP method were improved at least 30% and 44%, compared with traditional in-solution preparation method, suggesting dramatically higher peptide recovery of imFASP method for trace amounts of complex proteome samples. All these results demonstrate that the imFASP method developed here is of high potential for high efficient and high throughput preparation of trace amounts of complex proteome samples. PMID:26920773

  11. Using mass spectrometry to detect hydrolysed gluten in beer that is responsible for false negatives by ELISA.

    PubMed

    Colgrave, Michelle L; Goswami, Hareshwar; Blundell, Malcolm; Howitt, Crispin A; Tanner, Gregory J

    2014-11-28

    Gluten is the collective name for a class of proteins found in wheat, rye, barley and oats. Eating gluten triggers an inappropriate auto-immune reaction in ∼70 million people globally affected by coeliac disease, where the gut reacts to gluten proteins and this triggers an immune response, resulting in intestinal inflammation and damage. Gluten-free foods are now commonplace, however, it is difficult to accurately determine the gluten content of products claiming to be gluten-free using current methodologies as the antibodies are non-specific, show cross-reactivity and have different affinities for the different classes of gluten. The measurement of gluten in processed products is further confounded by modifications to the proteins that occur during processing and in some case hydrolysis of the proteins. In this study, LC-MS/MS was used to profile whole beer, and two beer fractions representing hydrolysed hordeins (<30 kDa) and hordein peptide fragments (<10 kDa). Subsequently, multiple reaction monitoring (MRM) MS enabled the relative quantification of selected peptide fragments in beer and revealed that certain classes of hordein were prone to hydrolysis (B- and D-hordein). Furthermore, select beers contained very high levels of gluten-derived fragments. Strikingly, those beers that contained high levels of B-hordein fragments gave near zero values by ELISA. The hydrolysed fragments that persist in beer show a dose-dependent suppression of ELISA measurement of gluten despite using a hordein standard for calibration of the assay. The development of MS-based methodology for absolute quantification of gluten is required for the accurate assessment of gluten, including hydrolysed forms, in food and beverages to support the industry, legislation and to protect consumers suffering from CD.

  12. Preparation of Core-Shell Hybrid Materials by Producing a Protein Corona Around Magnetic Nanoparticles

    NASA Astrophysics Data System (ADS)

    Weidner, A.; Gräfe, C.; von der Lühe, M.; Remmer, H.; Clement, J. H.; Eberbeck, D.; Ludwig, F.; Müller, R.; Schacher, F. H.; Dutz, S.

    2015-07-01

    Nanoparticles experience increasing interest for a variety of medical and pharmaceutical applications. When exposing nanomaterials, e.g., magnetic iron oxide nanoparticles (MNP), to human blood, a protein corona consisting of various components is formed immediately. The composition of the corona as well as its amount bound to the particle surface is dependent on different factors, e.g., particle size and surface charge. The actual composition of the formed protein corona might be of major importance for cellular uptake of magnetic nanoparticles. The aim of the present study was to analyze the formation of the protein corona during in vitro serum incubation in dependency of incubation time and temperature. For this, MNP with different shells were incubated in fetal calf serum (FCS, serving as protein source) within a water bath for a defined time and at a defined temperature. Before and after incubation the particles were characterized by a variety of methods. It was found that immediately (seconds) after contact of MNP and FCS, a protein corona is formed on the surface of MNP. This formation led to an increase of particle size and a slight agglomeration of the particles, which was relatively constant during the first minutes of incubation. A longer incubation (from hours to days) resulted in a stronger agglomeration of the FCS incubated MNP. Quantitative analysis (gel electrophoresis) of serum-incubated particles revealed a relatively constant amount of bound proteins during the first minutes of serum incubation. After a longer incubation (>20 min), a considerably higher amount of surface proteins was determined for incubation temperatures below 40 °C. For incubation temperatures above 50 °C, the influence of time was less significant which might be attributed to denaturation of proteins during incubation. Overall, analysis of the molecular weight distribution of proteins found in the corona revealed a clear influence of incubation time and temperature on corona

  13. Preparation of Core-Shell Hybrid Materials by Producing a Protein Corona Around Magnetic Nanoparticles.

    PubMed

    Weidner, A; Gräfe, C; von der Lühe, M; Remmer, H; Clement, J H; Eberbeck, D; Ludwig, F; Müller, R; Schacher, F H; Dutz, S

    2015-12-01

    Nanoparticles experience increasing interest for a variety of medical and pharmaceutical applications. When exposing nanomaterials, e.g., magnetic iron oxide nanoparticles (MNP), to human blood, a protein corona consisting of various components is formed immediately. The composition of the corona as well as its amount bound to the particle surface is dependent on different factors, e.g., particle size and surface charge. The actual composition of the formed protein corona might be of major importance for cellular uptake of magnetic nanoparticles. The aim of the present study was to analyze the formation of the protein corona during in vitro serum incubation in dependency of incubation time and temperature. For this, MNP with different shells were incubated in fetal calf serum (FCS, serving as protein source) within a water bath for a defined time and at a defined temperature. Before and after incubation the particles were characterized by a variety of methods. It was found that immediately (seconds) after contact of MNP and FCS, a protein corona is formed on the surface of MNP. This formation led to an increase of particle size and a slight agglomeration of the particles, which was relatively constant during the first minutes of incubation. A longer incubation (from hours to days) resulted in a stronger agglomeration of the FCS incubated MNP. Quantitative analysis (gel electrophoresis) of serum-incubated particles revealed a relatively constant amount of bound proteins during the first minutes of serum incubation. After a longer incubation (>20 min), a considerably higher amount of surface proteins was determined for incubation temperatures below 40 °C. For incubation temperatures above 50 °C, the influence of time was less significant which might be attributed to denaturation of proteins during incubation. Overall, analysis of the molecular weight distribution of proteins found in the corona revealed a clear influence of incubation time and temperature on

  14. Preparation and photolithography of self-assembled monolayers of 10-mercaptodecanylphosphonic acid on glass mediated by zirconium for protein patterning.

    PubMed

    Han, Xuemingyue; Sun, Shuqing; He, Tao

    2013-08-01

    Self-assembled monolayers (SAMs) formed by adsorption of octadecylphosphonic acid (ODPA) on zirconium mediated glass substrates were prepared. In this sandwich structure, Zr(4+) was used as a bi-linker to bind phosphonic acid head group in ODPA to glass substrates. The contact angle of the as-prepared SAMs was measured to be around 104°. X-ray photoelectron spectroscopy (XPS) characterization indicated the modification of Zr(4+) on glass substrates was critical for the formation of reasonably dense, well-ordered SAMs similar in quality to those typically formed on other metal oxide surfaces. Bifunctional molecule, 10-mercaptodecanylphosphonic acid (MDPA), bearing thiol terminal groups for various chemical reactions, was synthesized and formed SAMs on glass using the same approach, which allowed us to control the surface chemistry and functionality through photooxidation of the thiol terminal group. Photopatterning of proteins was performed first by exposing the SAMs to UV light through a mask, followed by protein immobilization to the masked regions through a heterobifunctional linker, while the exposed areas prohibit nonspecific protein absorption. The present strategy, which combined the SAMs assembly and photolithography, offered a facile approach for the fabrication of biomolecule patterning and could be applied to construction of biochips and other applications.

  15. Preparation and photolithography of self-assembled monolayers of 10-mercaptodecanylphosphonic acid on glass mediated by zirconium for protein patterning.

    PubMed

    Han, Xuemingyue; Sun, Shuqing; He, Tao

    2013-08-01

    Self-assembled monolayers (SAMs) formed by adsorption of octadecylphosphonic acid (ODPA) on zirconium mediated glass substrates were prepared. In this sandwich structure, Zr(4+) was used as a bi-linker to bind phosphonic acid head group in ODPA to glass substrates. The contact angle of the as-prepared SAMs was measured to be around 104°. X-ray photoelectron spectroscopy (XPS) characterization indicated the modification of Zr(4+) on glass substrates was critical for the formation of reasonably dense, well-ordered SAMs similar in quality to those typically formed on other metal oxide surfaces. Bifunctional molecule, 10-mercaptodecanylphosphonic acid (MDPA), bearing thiol terminal groups for various chemical reactions, was synthesized and formed SAMs on glass using the same approach, which allowed us to control the surface chemistry and functionality through photooxidation of the thiol terminal group. Photopatterning of proteins was performed first by exposing the SAMs to UV light through a mask, followed by protein immobilization to the masked regions through a heterobifunctional linker, while the exposed areas prohibit nonspecific protein absorption. The present strategy, which combined the SAMs assembly and photolithography, offered a facile approach for the fabrication of biomolecule patterning and could be applied to construction of biochips and other applications. PMID:23524079

  16. Dried blood spot proteomics: surface extraction of endogenous proteins coupled with automated sample preparation and mass spectrometry analysis.

    PubMed

    Martin, Nicholas J; Bunch, Josephine; Cooper, Helen J

    2013-08-01

    Dried blood spots offer many advantages as a sample format including ease and safety of transport and handling. To date, the majority of mass spectrometry analyses of dried blood spots have focused on small molecules or hemoglobin. However, dried blood spots are a potentially rich source of protein biomarkers, an area that has been overlooked. To address this issue, we have applied an untargeted bottom-up proteomics approach to the analysis of dried blood spots. We present an automated and integrated method for extraction of endogenous proteins from the surface of dried blood spots and sample preparation via trypsin digestion by use of the Advion Biosciences Triversa Nanomate robotic platform. Liquid chromatography tandem mass spectrometry of the resulting digests enabled identification of 120 proteins from a single dried blood spot. The proteins identified cross a concentration range of four orders of magnitude. The method is evaluated and the results discussed in terms of the proteins identified and their potential use as biomarkers in screening programs.

  17. Dried Blood Spot Proteomics: Surface Extraction of Endogenous Proteins Coupled with Automated Sample Preparation and Mass Spectrometry Analysis

    NASA Astrophysics Data System (ADS)

    Martin, Nicholas J.; Bunch, Josephine; Cooper, Helen J.

    2013-08-01

    Dried blood spots offer many advantages as a sample format including ease and safety of transport and handling. To date, the majority of mass spectrometry analyses of dried blood spots have focused on small molecules or hemoglobin. However, dried blood spots are a potentially rich source of protein biomarkers, an area that has been overlooked. To address this issue, we have applied an untargeted bottom-up proteomics approach to the analysis of dried blood spots. We present an automated and integrated method for extraction of endogenous proteins from the surface of dried blood spots and sample preparation via trypsin digestion by use of the Advion Biosciences Triversa Nanomate robotic platform. Liquid chromatography tandem mass spectrometry of the resulting digests enabled identification of 120 proteins from a single dried blood spot. The proteins identified cross a concentration range of four orders of magnitude. The method is evaluated and the results discussed in terms of the proteins identified and their potential use as biomarkers in screening programs.

  18. Lipid bilayer preparations of membrane proteins for oriented and magic-angle spinning solid-state NMR samples

    PubMed Central

    Das, Nabanita; Murray, Dylan T; Cross, Timothy A

    2014-01-01

    Solid-state NMR spectroscopy has been used successfully for characterizing the structure and dynamics of membrane proteins as well as their interactions with other proteins in lipid bilayers. such an environment is often necessary for achieving native-like structures. sample preparation is the key to this success. Here we present a detailed description of a robust protocol that results in high-quality membrane protein samples for both magic-angle spinning and oriented-sample solid-state NMR. the procedure is demonstrated using two proteins: CrgA (two transmembrane helices) and rv1861 (three transmembrane helices), both from Mycobacterium tuberculosis. the success of this procedure relies on two points. First, for samples for both types of NMR experiment, the reconstitution of the protein from a detergent environment to an environment in which it is incorporated into liposomes results in ‘complete’ removal of detergent. second, for the oriented samples, proper dehydration followed by rehydration of the proteoliposomes is essential. By using this protocol, proteoliposome samples for magic-angle spinning NMR and uniformly aligned samples (orientational mosaicity of <1°) for oriented-sample NMR can be obtained within 10 d. PMID:24157546

  19. Synthetic virus-like particles prepared via protein corona formation enable effective vaccination in an avian model of coronavirus infection.

    PubMed

    Chen, Hui-Wen; Huang, Chen-Yu; Lin, Shu-Yi; Fang, Zih-Syun; Hsu, Chen-Hsuan; Lin, Jung-Chen; Chen, Yuan-I; Yao, Bing-Yu; Hu, Che-Ming J

    2016-11-01

    The ongoing battle against current and rising viral infectious threats has prompted increasing effort in the development of vaccine technology. A major thrust in vaccine research focuses on developing formulations with virus-like features towards enhancing antigen presentation and immune processing. Herein, a facile approach to formulate synthetic virus-like particles (sVLPs) is demonstrated by exploiting the phenomenon of protein corona formation induced by the high-energy surfaces of synthetic nanoparticles. Using an avian coronavirus spike protein as a model antigen, sVLPs were prepared by incubating 100 nm gold nanoparticles in a solution containing an optimized concentration of viral proteins. Following removal of free proteins, antigen-laden particles were recovered and showed morphological semblance to natural viral particles under nanoparticle tracking analysis and transmission electron microscopy. As compared to inoculation with free proteins, vaccination with the sVLPs showed enhanced lymphatic antigen delivery, stronger antibody titers, increased splenic T-cell response, and reduced infection-associated symptoms in an avian model of coronavirus infection. Comparison to a commercial whole inactivated virus vaccine also showed evidence of superior antiviral protection by the sVLPs. The study demonstrates a simple yet robust method in bridging viral antigens with synthetic nanoparticles for improved vaccine application; it has practical implications in the management of human viral infections as well as in animal agriculture.

  20. Synthetic virus-like particles prepared via protein corona formation enable effective vaccination in an avian model of coronavirus infection.

    PubMed

    Chen, Hui-Wen; Huang, Chen-Yu; Lin, Shu-Yi; Fang, Zih-Syun; Hsu, Chen-Hsuan; Lin, Jung-Chen; Chen, Yuan-I; Yao, Bing-Yu; Hu, Che-Ming J

    2016-11-01

    The ongoing battle against current and rising viral infectious threats has prompted increasing effort in the development of vaccine technology. A major thrust in vaccine research focuses on developing formulations with virus-like features towards enhancing antigen presentation and immune processing. Herein, a facile approach to formulate synthetic virus-like particles (sVLPs) is demonstrated by exploiting the phenomenon of protein corona formation induced by the high-energy surfaces of synthetic nanoparticles. Using an avian coronavirus spike protein as a model antigen, sVLPs were prepared by incubating 100 nm gold nanoparticles in a solution containing an optimized concentration of viral proteins. Following removal of free proteins, antigen-laden particles were recovered and showed morphological semblance to natural viral particles under nanoparticle tracking analysis and transmission electron microscopy. As compared to inoculation with free proteins, vaccination with the sVLPs showed enhanced lymphatic antigen delivery, stronger antibody titers, increased splenic T-cell response, and reduced infection-associated symptoms in an avian model of coronavirus infection. Comparison to a commercial whole inactivated virus vaccine also showed evidence of superior antiviral protection by the sVLPs. The study demonstrates a simple yet robust method in bridging viral antigens with synthetic nanoparticles for improved vaccine application; it has practical implications in the management of human viral infections as well as in animal agriculture. PMID:27552321

  1. Preparation of amyloid-like fibrils containing magnetic iron oxide nanoparticles: Effect of protein aggregation on proton relaxivity

    SciTech Connect

    Viktor Andersson, B.; Skoglund, Caroline; Uvdal, Kajsa; Solin, Niclas

    2012-03-23

    Highlights: Black-Right-Pointing-Pointer Preparation of amyloid materials labeled with magnetic iron oxide nanoparticles. Black-Right-Pointing-Pointer Characterization of amyloid materials by electron tomography. Black-Right-Pointing-Pointer Influence of protein aggregation on the magnetic nanoparticle properties. -- Abstract: A method to prepare amyloid-like fibrils functionalized with magnetic nanoparticles has been developed. The amyloid-like fibrils are prepared in a two step procedure, where insulin and magnetic nanoparticles are mixed simply by grinding in the solid state, resulting in a water soluble hybrid material. When the hybrid material is heated in aqueous acid, the insulin/nanoparticle hybrid material self assembles to form amyloid-like fibrils incorporating the magnetic nanoparticles. This results in magnetically labeled amyloid-like fibrils which has been characterized by Transmission Electron Microscopy (TEM) and electron tomography. The influence of the aggregation process on proton relaxivity is investigated. The prepared materials have potential uses in a range of bio-imaging applications.

  2. Purification and characterization of a novel angiotensin I-converting enzyme inhibitory peptide derived from an enzymatic hydrolysate of duck skin byproducts.

    PubMed

    Lee, Seung-Jae; Kim, Yon-Suk; Kim, Seong-Eun; Kim, Eun-Kyung; Hwang, Jin-Woo; Park, Tae-Kyu; Kim, Bo Kyung; Moon, Sang-Ho; Jeon, Byong-Tae; Jeon, You-Jin; Ahn, Chang-Bum; Je, Jae-Young; Park, Pyo-Jam

    2012-10-10

    An angiotensin I-converting enzyme (ACE) inhibitory peptide was isolated and identified from hydrolysates of duck skin byproducts. Duck skin byproducts were hydrolyzed using nine proteases (Alcalase, Collagenase, Flavourzyme, Neutrase, papain, pepsin, Protamex, trypsin, and α-chymotrypsin) to produce an antihypertensive peptide. Of the various hydrolysates produced, the α-chymotrypsin hydrolysate exhibited the highest ACE inhibitory activity. The hydrolysate was purified using fast protein liquid chromatography (FPLC) and high-performance liquid chromatography (HPLC). The amino acid sequence of the ACE inhibitory peptide was identified as a hexapeptide Trp-Tyr-Pro-Ala-Ala-Pro, with a molecular weight of 693.90 Da. The peptide had an IC50 value of 137 μM, and the inhibitory pattern of the purified ACE inhibitor from duck skin byproducts was determined to be competitive by Lineweaver-Burk plots. In addition, the peptide was synthesized and the ACE inhibitory activity was verified in vivo. Spontaneously hypertensive rats (SHR) exhibited significantly decreased blood pressure and heart rate after peptide injection. Taken together, the results suggest that Trp-Tyr-Pro-Ala-Ala-Pro may be useful as a new antihypertensive agent.

  3. Bitter peptide from hemoglobin hydrolysate: isolation and characterization.

    PubMed

    Aubes-Dufau, I; Capdevielle, J; Seris, J L; Combes, D

    1995-05-01

    Two separation methods, ultrafiltration and 2-butanol extraction, have shown that a peptide is the major agent responsible for bitterness in peptic hemoglobin hydrolysates. It was easily purified from these complex mixtures by specific hydrophobic adsorption on Superose 12, a gel-filtration column, which could constitute an original and interesting method for bitterness detection. The bitter peptide which corresponded to VV-hemorphin 7, the fragment 32-40 of the beta chain of bovine hemoglobin, is first generated during proteolysis, then hydrolysed by pepsin. It exhibited a strong bitterness at 0.25 mM equivalent to 0.073 mM quinine sulfate or 21 mM caffeine.

  4. Preparation and antioxidant activities of oligosaccharides from Crassostrea gigas.

    PubMed

    Wu, Shengjun; Huang, Xiaolian

    2017-02-01

    Oligosaccharides were prepared from Crassostrea gigas by hydrolysis of polysaccharide in C. gigas with peroxide oxygen (H2O2). The hydrolysates were cleared of protein, filtered, ultrafiltered and precipitated with absolute ethanol to give C. gigas oligosaccharides (CGOs). Factors affecting CGO yields, i.e., reaction time, temperature, and H2O2 concentration, were optimised as follows: 2.96h reaction time, 84.71°C reaction temperature, and 2.46% H2O2 concentration. Under these conditions, the maximum yield of CGOs reached 10.61%. The CGOs were then partially characterised by Fourier transform infrared spectroscopy, UV spectroscopy, monosaccharide composition, and antioxidant activities. Results indicate that CGOs possessed strong hydroxyl radical activity, 2,2-diphenyl-β-picrylhydrazyl-radical-scavenging activity and reducing capacity at a concentration of 100μg/mL. PMID:27596415

  5. Proteomic Challenges: Sample Preparation Techniques for Microgram-Quantity Protein Analysis from Biological Samples

    PubMed Central

    Feist, Peter; Hummon, Amanda B.

    2015-01-01

    Proteins regulate many cellular functions and analyzing the presence and abundance of proteins in biological samples are central focuses in proteomics. The discovery and validation of biomarkers, pathways, and drug targets for various diseases can be accomplished using mass spectrometry-based proteomics. However, with mass-limited samples like tumor biopsies, it can be challenging to obtain sufficient amounts of proteins to generate high-quality mass spectrometric data. Techniques developed for macroscale quantities recover sufficient amounts of protein from milligram quantities of starting material, but sample losses become crippling with these techniques when only microgram amounts of material are available. To combat this challenge, proteomicists have developed micro-scale techniques that are compatible with decreased sample size (100 μg or lower) and still enable excellent proteome coverage. Extraction, contaminant removal, protein quantitation, and sample handling techniques for the microgram protein range are reviewed here, with an emphasis on liquid chromatography and bottom-up mass spectrometry-compatible techniques. Also, a range of biological specimens, including mammalian tissues and model cell culture systems, are discussed. PMID:25664860

  6. Recombinant Preparation, Biochemical Analysis, and Structure Determination of Sirtuin Family Histone/Protein Deacylases.

    PubMed

    Suenkel, B; Steegborn, C

    2016-01-01

    Lysine acetylation is long known as a regulatory posttranslational modification of histone proteins and is emerging as a ubiquitous intracellular protein modification. Additional lysine acylations such as succinylation and glutarylation have also been found on histones and other proteins. Acylations are reversibly attached through nonenzymatic acylation mechanisms and the action of protein acyl transferases and protein deacylases (PDACs). Sirtuins are an evolutionary defined class of PDACs and act as metabolic sensors by catalyzing a unique deacylation reaction that requires the cosubstrate NAD(+). Sirtuins are found in all domains of life, and the mammalian sirtuin family comprises seven isoforms in different cellular compartments. They regulate a wide range of cellular targets and functions, such as energy metabolism and stress responses, and they have been implicated in aging processes and aging-related diseases. A large body of functional, biochemical, biophysical, and structural work on isolated sirtuins has provided many important insights that complement the many physiological studies on this enzyme family. They enabled the comprehensive structural and biochemical analysis of sirtuin catalysis, substrate selectivity, and regulation. Here, we describe the recombinant production of sirtuin proteins, with an emphasis on the mammalian isoforms. We then describe their application in activity and binding assays and for crystal structure analysis. We provide protocols for these procedures, and we discuss typical pitfalls in studying this enzyme family and how to avoid them. This information will support further molecular studies on sirtuin mechanisms and functions. PMID:27372754

  7. Preparation of hollow nickel silicate nanospheres for separation of His-tagged proteins.

    PubMed

    Wu, Yonghui; Chang, Guanxiao; Zhao, Yanbao; Zhang, Yu

    2014-01-14

    Hollow nickel silicate nanospheres (NiSiO3 NSs) with hierarchical shells were hydrothermally synthesized by using silica spheres as a template. The NiSiO3 NSs have an average diameter of 250 nm with a shell thickness of 50 nm, and the hierarchical shell consists of a large number of sheets. By taking advantage of the high affinity of Ni(2+) toward histidine-tagged (His-tagged) proteins, hollow NiSiO3 NSs can be used to enrich and separate His-tagged proteins directly from a mixture of lysed cells. Results indicated that the hollow NiSiO3 NSs presented negligible nonspecific protein adsorption and a high protein binding ability with a high binding capacity of 13.2 mmol g(-1). Their specificity and affinity toward His-tagged proteins remained after recycling 5 times. The hollow NiSiO3 NSs are especially suitable for rapid purification of His-tagged proteins. PMID:24149676

  8. Simulation of the continuous fermentation of manioc hydrolysate

    SciTech Connect

    Bonomi, A.; Aboutboul, H.; Schmidell, W.

    1981-01-01

    The simulation of the continuous fermentation of manioc hydrolysate utilizing a yeast strain of Saccharomyces cerevisiae isolated from the commercial pressed yeast largely employed in Brazilian distilleries is described. The model used in the simulation is derived from batch experimental runs. In order to assess the economical competitiveness of the continuous fermentation, some additional concepts, such as cell recycle, and two fermentors connected in series with and without feed division of fresh substrate, are analyzed and compared.

  9. Kinetic studies of cellodextrins hydrolyses by exocellulase from trichoderma reesei

    SciTech Connect

    Teh-An Hsu, Cheng-Shung Gong; Tsao, G.T.

    1980-11-01

    The kinetics of the hydrolyses of cellotriose and of cellotetraose by cellobiohydrolase were studied using a convenient integral technique. Reaction mechanisms and mathematical models were postulated to describe the reactions. The end-products of the reaction were found to be inhibitory toward hydrolysis in a competitive mode. Hydrolysis of cellotraose produces cellobiose and hydrolysis of cellotriose produces cellobiose and glucose. Both sugars inhibit the enzyme with cellobiose being a stronger inhibitor.

  10. Scope of Hydrolysable Tannins as Possible Antimicrobial Agent.

    PubMed

    Ekambaram, Sanmuga Priya; Perumal, Senthamil Selvan; Balakrishnan, Ajay

    2016-07-01

    Hydrolysable tannins (HTs) are secondary metabolites from plants, which are roughly classified into gallotannins and ellagitannins having gallic acid and ellagic acid residues respectively attached to the hydroxyl group of glucose by ester linkage. The presence of hexahydroxydiphenoyl and nonahydroxyterphenoyl moieties is considered to render antimicrobial property to HTs. HTs also show considerable synergy with antibiotics. Nevertheless, they have low pharmacokinetic property. The present review presents the scope of HTs as future antimicrobial agent. Copyright © 2016 John Wiley & Sons, Ltd. PMID:27062587

  11. Alternative preparation of inclusion bodies excludes interfering non-protein contaminants and improves the yield of recombinant proinsulin.

    PubMed

    Mackin, Robert B

    2014-01-01

    The goal of simple, high-yield expression and purification of recombinant human proinsulin has proven to be a considerable challenge. First, proinsulin forms inclusion bodies during bacterial expression. While this phenomenon can be exploited as a capture step, conventionally prepared inclusion bodies contain significant amounts of non-protein contaminants that interfere with subsequent chromatographic purification. Second, the proinsulin molecules within the inclusion bodies are incorrectly folded, and likely cross-linked to one another, making it difficult to quantify the amount of expressed proinsulin. Third, proinsulin is an intermediate between the initial product of ribosomal translation (preproinsulin) and the final product secreted by pancreatic beta cells (insulin). Therefore, to be efficiently produced in bacteria, it must be produced as an N-terminally extended fusion protein, which has to be converted to authentic proinsulin during the purification scheme. To address all three of these problems, while simultaneously streamlining the procedure and increasing the yield of recombinant proinsulin, we have made three substantive modifications to our previous method for producing proinsulin:.•Conditions for the preparation of inclusion bodies have been altered so contaminants that interfere with semi-preparative reversed-phase chromatography are excluded while the proinsulin fusion protein is retained at high yield.•Aliquots are taken following important steps in the procedure and the quantity of proinsulin-related polypeptide in the sample is compared to the amount present prior to that step.•Final purification is performed using a silica-based reversed-phase matrix in place of a polystyrene-divinylbenzene-based matrix.

  12. Gel-aided sample preparation (GASP)--a simplified method for gel-assisted proteomic sample generation from protein extracts and intact cells.

    PubMed

    Fischer, Roman; Kessler, Benedikt M

    2015-04-01

    We describe a "gel-assisted" proteomic sample preparation method for MS analysis. Solubilized protein extracts or intact cells are copolymerized with acrylamide, facilitating denaturation, reduction, quantitative cysteine alkylation, and matrix formation. Gel-aided sample preparation has been optimized to be highly flexible, scalable, and to allow reproducible sample generation from 50 cells to milligrams of protein extracts. This methodology is fast, sensitive, easy-to-use on a wide range of sample types, and accessible to nonspecialists. PMID:25515006

  13. Extensively and partially hydrolysed infant formulas for allergy prophylaxis

    PubMed Central

    Oldaeus, G; Anjou, K; Bjorksten, B; Moran, J; Kjellman, N

    1997-01-01

    Accepted 17 March 1997
 The allergy preventive effect of extensively (N) and partially (PH) hydrolysed cows' milk formulas compared with a regular formula (RM) was assessed in 155 infants with a family history of allergy. No cows' milk was given during the first nine months of life and no egg and fish up to 12 months of age. Breast feeding mothers avoided the same foods. At weaning the infants were randomised to one of the formula groups. The cumulative incidence of atopic symptoms at 18 months was 51, 64, and 84% in the N, PH, and RM groups, respectively. From 6 to 18 months there were significantly less cumulative atopic symptoms in the N group compared with the RM group, and significantly less than the PH group up to 6 (N= 25%; PH = 46%) and 9 months (N = 34%, PH = 58%). At 9 months significantly fewer infants in the N group (10%) than in the PH group (33%) had a positive skin prick test to eggs. The findings support an allergy preventive effect of an extensively hydrolysed formula, but not of a partially hydrolysed formula, during the first 18 months of life of high risk infants.

 PMID:9279143

  14. Bioflavour production from orange peel hydrolysate using immobilized Saccharomyces cerevisiae.

    PubMed

    Lalou, Sofia; Mantzouridou, Fani; Paraskevopoulou, Adamantini; Bugarski, Branko; Levic, Steva; Nedovic, Victor

    2013-11-01

    The rising trend of bioflavour synthesis by microorganisms is hindered by the high manufacturing costs, partially attributed to the cost of the starting material. To overcome this limitation, in the present study, dilute-acid hydrolysate of orange peel was employed as a low-cost, rich in fermentable sugars substrate for the production of flavour-active compounds by Saccharomyces cerevisiae. With this purpose, the use of immobilized cell technology to protect cells against the various inhibitory compounds present in the hydrolysate was evaluated with regard to yeast viability, carbon and nitrogen consumption and cell ability to produce flavour active compounds. For cell immobilization the encapsulation in Ca alginate beads was used. The results were compared with those obtained using free-cell system. Based on the data obtained immobilized cells showed better growth performance and increased ability for de novo synthesis of volatile esters of "fruity" aroma (phenylethyl acetate, ethyl hexanoate, octanoate, decanoate and dodecanoate) than those of free cells. The potential for in situ production of new formulations containing flavour-active compounds derive from yeast cells and also from essential oil of orange peel (limonene, α-terpineol) was demonstrated by the fact that bioflavour mixture was found to accumulate within the beads. Furthermore, the ability of the immobilized yeast to perform efficiently repeated batch fermentations of orange peel hydrolysate for bioflavour production was successfully maintained after six consecutive cycles of a total period of 240 h. PMID:23995224

  15. Detoxification of sugarcane bagasse hydrolysate improves ethanol production by Candida shehatae NCIM 3501.

    PubMed

    Chandel, Anuj Kumar; Kapoor, Rajeev Kumar; Singh, Ajay; Kuhad, Ramesh Chander

    2007-07-01

    Sugarcane bagasse hydrolysis with 2.5% (v/v) HCl yielded 30.29g/L total reducing sugars along with various fermentation inhibitors such as furans, phenolics and acetic acid. The acid hydrolysate when treated with anion exchange resin brought about maximum reduction in furans (63.4%) and total phenolics (75.8%). Treatment of hydrolysate with activated charcoal caused 38.7% and 57.5% reduction in furans and total phenolics, respectively. Laccase reduced total phenolics (77.5%) without affecting furans and acetic acid content in the hydrolysate. Fermentation of these hydrolysates with Candida shehatae NCIM 3501 showed maximum ethanol yield (0.48g/g) from ion exchange treated hydrolysate, followed by activated charcoal (0.42g/g), laccase (0.37g/g), overliming (0.30g/g) and neutralized hydrolysate (0.22g/g). PMID:17011776

  16. In vitro protein phosphorylation in head preparations from normal and mutant Drosophila melanogaster.

    PubMed

    Buxbaum, J D; Dudai, Y

    1987-10-01

    We have characterized protein phosphorylation in vitro in subcellular fractions from Drosophila melanogaster heads. Optimal conditions for the incorporation of 32P into proteins, and its dependence on ATP, divalent cations, and cyclic nucleotides have been determined, as well as the effect of inhibitors of ATPase, protein phosphatase, and protein kinase on protein phosphorylation. Among these inhibitors, Zn2+ was found to affect the incorporation of 32P into specific bands and p-hydroxymercuribenzoate was found to be most suited for freezing the activity of both kinases and phosphatases. Cyclic AMP-dependent protein kinase (cAMP-dPK) activity was present in both supernatant (S2) and particulate (P2) fractions, with the majority (60-85%, depending on the homogenization medium) being associated with S2, as determined by phosphorylation of exogenous synapsin I. cAMP-dPK catalyzed the phosphorylation of at least 18 endogenous polypeptides in S2 and at least 10 endogenous polypeptides in P2. These proteins could be classified on the basis of the extent of stimulation of phosphorylation by cyclic nucleotides, dependence on c