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Pleurotus eryngii is a tasty and low-calorie mushroom containing abundant high-quality protein. This study aims to improve the digestibility of P. eryngii protein (PEP) and hence to facilitate its development as a healthy alternative protein. The extracted PEP was pretreated with 1000–5000 U of papain, neutral protease and alkaline protease. The Chyme collected from in vitro simulated gastrointestinal digestion was analyzed by fluorescence microscopy and protein particle analyzer, and the endpoint profiles of peptides and amino acids were determined by UHPLC-MS/MS and NanoLC-MS/MS. The particle size curve and fluorescence microscopy images jointly supported that protease hydrolysis improved decomposition and dispersion of PEP during digestion, particularly in the gastric phase. The impact on Zeta potential was minimal. Proteases effectively increased the abundance of amino acids after digestion, particularly L-isomer Lys and Arg Maximum release was achieved when pretreated with 5000 U of alkaline protease, reaching 7.54 times that of control. Pretreatments by proteases also notably increased digestive yields of 16,736–19,870 peptides, with the maximum reaching 1.70 times that of the control, which mainly consisted of small peptides composed of 7–15 amino acids with molecular weight below 800 Da. The findings indicated that protease hydrolysis, especially pretreatment with 5000 U of alkaline protease, effectively enhanced the digestibility of PEP, which shed light on providing enzymatic approaches for improving bioavailability and developing healthy fungal proteins. [ABSTRACT FROM AUTHOR]

Background and Objectives: Wheat gluten is a high‐quality, low‐priced, and nutritious plant protein. Applications of wheat gluten protein could be expanded by improving its solubility through enzymatic modifications. Herein, we investigated the effect of proteolytic processing on the physicochemical properties of wheat gluten protein. Findings: The degree of hydrolysis, trichloroacetic acid‐soluble nitrogen (TCA‐SN), and peptidyl nitrogen content of the enzymatic digestion products increased, and the TCA‐SN content exceeded 60% in all cases. The ζ‐potentials of all products after enzymatic digestion were significantly decreased and were negative. Three enzymatic solutions increased the stability of the solution system and the surface hydrophobicity of the enzymatic products to different degrees. The products after enzymatic digestion by alkaline protease had better antioxidant capacity. Conclusions: This study provides theoretical support for expanding the applications for wheat protein to achieve the added value of wheat products and enhance the utilization rate of these readily available raw materials. Significance and Novelty: This study investigates the dual enzyme stepwise enzymatic hydrolysis of wheat gluten protein by alkaline protease and neutral protease, which increases the water‐soluble protein content of wheat gluten protein. The resulting hydrolysate has a high peptide nitrogen content. [ABSTRACT FROM AUTHOR]

The effect of four enzyme preparations (EP), Bacillolysin, Agroprot, Protozyme, and Protozyme C (Russia), on the protein and peptide profiles of the protein isolate isolated from peas of the Focor variety, as well as on its smell and taste, was investigated in this work. It was shown that enzyme treatment can improve the odor characteristics of the isolate. Thus, it was possible to reduce significantly the severity of the bean and herbal smell. At the same time, enzyme treatment also improved the taste of the isolate: it was possible to reduce significantly the severity of disturbing flavors such as leguminous, astringent, bitter, and herbal. The results obtained allowed us to select EP (fungal acid aspartic protease) to improve the organoleptic parameters of pea protein isolates intended for the production of meat and dairy product analogs. [ABSTRACT FROM AUTHOR]

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Copyright of Journal of Chinese Institute of Food Science & Technology / Zhongguo Shipin Xuebao is the property of Journal of Chinese Institute of Food Science & Technology Periodical Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Food Research & Development is the property of Food Research & Development Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Journal of Food Safety & Quality is the property of Journal of Food Safety & Quality Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

The aim of this study was to investigate the effect of neutral protease treatment on the biochemical properties of various parts of Pacific oysters (Crassostrea gigas) under different storage conditions. The mechanism of quality degradation in the mantle, adductor muscle, gill, and trunk of treated oysters stored at −1.5 °C (superchilling) or 4 °C (refrigeration) for several days using different storage methods was studied. The results showed that the oyster treated with the enzyme exhibited higher glycogen content, flavor nucleotide content, and sensory scores compared to the control group. Superchilling at −1.5 °C was observed to slow the increase in total volatile basic nitrogen (TVB-N), total viable count (TVC), and pH, while maintaining sensory scores better than refrigeration at 4 °C. Both wet superchilling (WS) and dry exposed superchilling (DeS) methods effectively preserved freshness and quality at −1.5 °C. The freshness of the oysters' body trunk changed most significantly. K value, K′ value, and AEC value, as the evaluation indexes of oyster freshness, were affected by the storage medium. Therefore, neutral protease enhances the flavor of oysters in a short time, and oysters stored in wet superchilling or dry exposed superchilling conditions have an extended shelf life. [ABSTRACT FROM AUTHOR]

The effect of four enzyme preparations: bacillolysin, agroprot, protozyme and protozyme C (Russia) on solubility, emulsifying activity, emulsion stability, foaming and foam stability of isolates prepared from two varieties of peas was studied. It is shown that treatment with enzymes can increase the solubility of isolates at pH 5 by more than 7 times, the index of emulsifying activity at pH 5 by 1.5 to 2 times, and at pH 6 by almost 1.5 times; the stability index of the emulsion increased by about 20% at pH 5, and by 1.7 times (in one of the varieties) at pH 6; foaming increased by 2.4 to 3 times at pH 5, and at pH 6 by 1.8 to 3.7 times; foam stability increased by 25 to 33% at pH 5 and by more than 1.5 times (in one of the varieties) at pH 6. The results obtained made it possible to select an enzyme preparation (bacterial alkaline serine protease) to improve the parameters of pea protein isolates intended for the manufacture of analogs of fermented milk products. [ABSTRACT FROM AUTHOR]

Based on enzyme engineering and enzyme kinetics as the technical support and scientific basis, the combination of pig manure and corn stover was used as the substrate, 2% neutral protease was added, and the water content of the substrate was reduced to 56.01% at 80°C for 3 h in a biochemical reactor, a year-on-year decrease of 2.34%. The content of macromolecular organic matter was reduced to 63.79%, and after enzymatic hydrolysis, the organic content of substrates treated A and B reached 14.19% and 15.06%, respectively. The use of a large number of elements in plants had been increased to 6.87%, which met the NY 525-2012 organic fertilizer standard. Available-available phosphorus, avail- able potassium, total salt and total humic acid showed a downward trend. The mortality rate of roundworm eggs reached 92%, and the seed germination index reached 54%, which reached the standard of organic fertilizer rot. [ABSTRACT FROM AUTHOR]

Extruded corn flour can be easily agglomerated and poses poor flavor. Previous studies have found that phosphorylation heat treatment, enzyme treatment, edible gum dry heat treatment, and composite treatment could significantly reduce the agglomeration rate. The present study sought to explore the effects of sodium orthophosphate heat treatment (S1), neutral protease (S2), xanthan gum dry heat treatment (S3), and sodium orthophosphate-neutral protease treatment (S4) on the structural characteristics and flavor components of extruded corn flour (PCF) using scanning electron microscopy, Fourier transform infrared spectroscopy, X-ray diffraction, headspace solid phase microextraction, and gas chromatography mass spectrometry. The results showed that the extruded corn flour treated by these four methods exhibited an A-type crystal structure without the formation or introduction of any new chemical bonds or functional groups. The pores and cracks on the surface of starch microparticles significantly increased. Among them, the sodium orthophosphate+neutral protease treated powder had more surface pores and larger cracks than other samples. A total of 130 volatile components were identified by gas chromatography-mass spectrometry, including neutral protease treatment powder (57) > sodium orthophosphate+neutral protease treatment powder (46) > xanthan gum dry heat treatment powder (44) > PCF (41) > sodium orthophosphate heat treatment powder (40). ROAV analysis showed that the key flavor substances of extruded corn flour were mainly ketones. After different treatments, the overall flavor of the powder slightly differed, mainly aldehydes, with grass, orange, and fat flavor. The present study results could provide a theoretical basis to improve the flavor of blended grain powder.

Removing sericin from the periphery of silk without damage to silk fibroin (SF) to obtain high-molecular-weight SF is a major challenge in the field of SF-based biomaterials. In this study, four neutral proteases, subtilisin, trypsin, bromelain and papain, were used to degum silk, and the degumming efficiency of the proteases and their influence on the molecular weight (MW) of regenerated silk fibroin were studied. The results indicated that all four neutral proteases could remove sericin from silk almost completely, and they caused less damage to SF fibers than Na2CO3 degumming did. The degumming efficiency of trypsin and papain was strong, but they caused relatively high damage to SF, whereas bromelain caused the least damage. The results of sodium dodecyl sulfate–polyacrylamide gel electrophoresis, gel permeation chromatography and shear viscosity showed that the MWs of regenerated SF derived from neutral protease degumming were significantly higher than that of SF derived from Na2CO3 degumming. The MW of regenerated SF derived from bromelain degumming was the highest, while the MWs of regenerated SF derived from papain and trypsin degumming were relatively low. This study provides an efficient and environmentally friendly biological degumming method for obtaining high-molecular-weight silk fibroin. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate the effect of neutral protease treatment on the biochemical properties of various parts of Pacific oysters (Crassostrea gigas) under different storage conditions. The mechanism of quality degradation in the mantle, adductor muscle, gill, and trunk of treated oysters stored at −1.5 °C (superchilling) or 4 °C (refrigeration) for several days using different storage methods was studied. The results showed that the oyster treated with the enzyme exhibited higher glycogen content, flavor nucleotide content, and sensory scores compared to the control group. Superchilling at −1.5 °C was observed to slow the increase in total volatile basic nitrogen (TVB-N), total viable count (TVC), and pH, while maintaining sensory scores better than refrigeration at 4 °C. Both wet superchilling (WS) and dry exposed superchilling (DeS) methods effectively preserved freshness and quality at −1.5 °C. The freshness of the oysters’ body trunk changed most significantly. K value, K′ value, and AEC value, as the evaluation indexes of oyster freshness, were affected by the storage medium. Therefore, neutral protease enhances the flavor of oysters in a short time, and oysters stored in wet superchilling or dry exposed superchilling conditions have an extended shelf life.

For the advancement of porcine xenotransplantation for clinical use in type 1 diabetes mellitus, the concerns of a sustainable and safe digestion enzyme blend must be overcome. Incorporating good manufacturing practices (GMP) can facilitate this through utilizing GMP-grade enzymes. In conjunction, still taking into account the cost-effectiveness, a wide concern. We evaluated how GMP-grade enzyme blends impact our piglet islets and their long-term effects.  Preweaned porcine islets (PPIs) were isolated from 8- to 10-day-old pigs. Digestion enzyme blends, collagenase type V (Type V), collagenase AF-1 GMP-grade with collagenase NB 6 GMP-grade (AF-1 and NB 6), and collagenase AF-1 GMP-grade with collagenase neutral protease AF GMP-grade (AF-1 and NP AF) were compared. Islet quality control assessments, islet yield, viability, and function, were performed on days 3 and 7, and cell content was performed on day 7.  GMP-grade AF-1 and NB 6 (17,209 ± 2,730 islet equivalent per gram of pancreatic tissue [IE/g] on day 3, 9,001 ± 1,034 IE/g on day 7) and AF-1 and NP AF (17,214 ± 3,901 IE/g on day 3, 8,833 ± 2,398 IE/g on day 7) showed a significant increase in islet yield compared to Type V (4,618 ± 1,240 IE/g on day 3, 1,923 ± 704 IE/g on day 7). Islet size, viability, and function showed comparable results in all enzyme blends. There was no significant difference in islet cellular content between enzyme blends.  This study demonstrated a comparison of GMP-grade collagenase enzyme blends and a standard crude collagenase enzyme in preweaned-aged porcine, a novel topic in this age. GMP-grade enzyme blends of AF-1 and NB 6 and AF-1 and NP AF resulted in substantially higher yields and as effective PPIs compared to Type V. In the long run, considering costs, integrity, and sustainability, GMP-grade enzyme blends are more favorable for clinical application due to high reproducibility in comparison to undefined manufacturing processes of standard enzymes.

Trichophyton rubrum and T. mentagrophytes are the most common agents of dermatomycosis, a disease affecting millions worldwide. It has been widely recognized that secreted proteases are a key factor for host colonization. Dermatophytes have an unusually high amount of secreted protease, differentially expressed, and influenced by various conditions. This study analyzed the rule and expression of secreted deuterolysin protease of the M35 protein family in these two representative dermatophyte species. All strains secreted protease and could grow on keratin as the sole carbon and nitrogen source. Adding glucose to the keratin medium reduced the growth rate. Deuterolysin genes were most strongly expressed at acid conditions. NPIIc and NPIId expression was significantly higher than the other three deuterolysins. NPIIc had a high expression level in the two T. rubrum strains but a low expression in T. mentagrophytes strains. Both T. mentagrophytes strains had a high NPIId expression at low pH. NPIIc and NPIId deletion in T. rubrum caused a minor reduction in total protease activity, indicating the redundancy of protease in dermatophytes. It was postulated that protease gene enrichment in dermatophytes allows a sophisticated regulation of protease secretion to cope with changing conditions. Lay Summary: Nail infections and ringworm are caused by fungi called dermatophytes. About 20% of the world's population suffers from it at least once. Dermatophytes secrete skin protein-digesting enzymes. This study demonstrates the changing enzyme profile in response to different pH levels. [ABSTRACT FROM AUTHOR]

Using Ziziphi Spinosae semen as raw material, Ziziphi Spinosae semen protein was extracted by alkali extraction and acid precipitation method. The Ziziphi Spinosae semen protein was hydrolyzed by three different proteases (alkaline protease, neutral protease and papain). The functional properties and antioxidant activity of enzymatic hydrolysates from Ziziphi Spinosae semen protein were studied. The results showed that the solubility, oil holding capacity, foaming capacity and stability, emulsifying capacity and stability of protein hydrolysates from different Ziziphi Spinosae semen were improved in different degrees compared with protein hydrolysates from Ziziphi Spinosae semen, among them, papain hydrolysate had the strongest solubility, oil holding, foaming and emulsifying properties, and alkaline protease hydrolysate had the strongest foaming and emulsifying stability. In vitro antioxidant studies showed that compared with of Ziziphi Spinosae semen protein, the antioxidant activities of enzymatic hydrolysates obtained by different proteases were significantly improved, the alkaline protease DPPH scavenging ability, hydroxyl radical scavenging ability, super oxygen anion clearance and ABTS+ radical scavenging ability were higher than those of other enzymatic hydrolysates, when the mass concentration was 2.5 mg/mL, the highest DPPH radical scavenging rate was 95.83%, the superoxide anion radical scavenging rate was 44.77%, the ABTS radical scavenging rate was 90.84%, and the hydroxyl radical scavenging rate was 44.77%. Therefore, this study shows that the enzymatic hydrolysate of Ziziphi Spinosae semen protein has good food processing performance, and its antioxidant activity is significantly increased, protein sources can be used as a potential protein resource in food, and provides a theoretical basis for the application of Ziziphi Spinosae semen protease hydrolysate in food industry.

In this study, the effects of the thermal ultrasonic enzyme inactivation process on flavor enhancement in sea cucumber hydrolysates (SCHs) and its impact on the inactivation of neutral proteases (NPs) were investigated. The body wall of the sea cucumber was enzymatically hydrolyzed with NPs. On the one hand, the structure of NPs subjected to different enzyme inactivation methods was analyzed using ζ-potential, particle size, and Fourier transform infrared (FT-IR) spectroscopy. On the other hand, the microstructure and flavor changes of SCHs were examined through scanning electron microscopy, E -nose, and gas chromatography-ion mobility spectrometry (GC-IMS). The results indicated that thermal ultrasound treatment at 60 °C could greatly affect the structure of NPs, thereby achieving enzyme inactivation. Furthermore, this treatment generated more pleasant flavor compounds, such as pentanal and (E)-2-nonenal. Hence, thermal ultrasound treatment could serve as an alternative process to traditional heat inactivation of enzymes for improving the flavor of SCHs. • Thermal ultrasonic inactivation was comparable to traditional inactivation. • Ultrasound requires heat treatment assistance to inactivate enzyme. • Thermal ultrasound caused structural changes in neutral protease. • Thermal ultrasonic inactivation improved flavor of sea cucumber hydrolysates. [ABSTRACT FROM AUTHOR]

Sodium alginate and chitosan were cross-linked to form composite gel spheres, which were entrapped to immobilize the free neutral protease. The matrix of the immobilized neutral protease was detected and characterized by using Fourier transform infrared spectroscopy and energy-dispersive X-ray. The optimum immobilization conditions were determined by orthogonal test, in which the concentration of sodium alginate was 3.5%, CaCl2 2.5%, chitosan 2.5%, and immobilizing time 1.5 h. Meanwhile, the activities of immobilized neutral protease and free enzyme were compared. The results showed that the pH value of immobilized enzyme was 5–8, the relative activity was above 90%, the free enzyme was above 80%, the relative activity of immobilized enzyme was above 80% in 30–80 °C, and the free enzyme was above 64% in 40–80 °C. The immobilized enzyme is better than the free enzyme in the constant of pH and temperature. The relative activity of immobilized enzyme was 50% after six hydrolysis cycles, and 80% after 11 days of storage. [ABSTRACT FROM AUTHOR]

Neutral protease, pectinase and amylase were used to treat the tobacco flake concentrate from Guizhou raw materials and the effects of different enzyme treatments on the conventional chemical components and fragrance components in the concentrate of the treated samples were analyzed. The concentrated solution after enzyme treatment was coated on the tobacco sheet substrate and the sensory quality of tobacco sheet was evaluated. The results showed that the mass fractions of protein, reducing sugars, total sugars and amino acids in the concentrated solution treated with neutral protease were greatly reduced; 10 new fragrance ingredients including 4-cyclopentene-1,3-dione and β-ionone were found in the concentrated treated solution. The total content of aroma components in the concentrate had been increased from 0.907 mg/mL to 2.844 mg/mL, with an increase of 213.56%. There was no significant change in the conventional chemical components and flavor components in the concentrates treated with pectinase and amylase. Tobacco flakes coated with the concentrated solution treated with neutral protease were better than amylase group, pectinase group and blank group in terms of aroma, harmony, irritation and aftertaste. [ABSTRACT FROM AUTHOR]

High‐temperature pretreatment that is currently used in camellia oil extraction can have negative effects on the quality of camellia oil. In this study, the enzymatic pretreatment of camellia seeds is explored as an alternative to high‐temperature pretreatment. The main conditions for enzymatic pretreatment of camellia seeds including enzyme, pH, temperature, time, and buffer solution are optimized using the response surface methodology. Under the optimal conditions of enzymatic pretreatment, the oil recovery is close to 75%. Moreover, residual oil recovery from camellia seeds subjected to 1398 neutral protease pretreatment (4 g per kg seeds) and high‐temperature pretreatment are 5.62 ± 0.08% and 9.97 ± 0.18%, respectively. The enzymatic pretreatment is further applied to pre‐pressing solvent extraction of camellia oil, the cake oil recovery from camellia seeds subjected to enzymatic pretreatment is higher than that from high‐temperature pretreatment. These results show that enzymatic pretreatment of camellia seeds has potential for application in the oil industry. Practical Applications : This study suggests that enzymatic pretreatment can replace high‐temperature pretreatment and improve oil recovery and oil quality. Ultimately, this method can be used to extract camellia oil. [ABSTRACT FROM AUTHOR]

Copyright of China Brewing is the property of China Brewing Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Food Research & Development is the property of Food Research & Development Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Food & Machinery is the property of Food & Machinery Editorial Office and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of Food Research & Development is the property of Food Research & Development Editorial Department and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Abstract The strain Y1, with a notably high production of neutral protease, was isolated from naturally fermented broad beans and subsequently identified as Aspergillus oryzae, through the analysis of its morphology characteristics and 18S rDNA sequence. Naturally fermented broad beans are the main raw material in Sichuan broad-bean sauce. The neutral protease from Aspergillus oryzae Y1 was purified using ammonium sulphate precipitation and DEAE-Sepharose Fast Flow chromatography, which resulted in a 10.0-fold increase in the specific activity (2264.3 U/mg) and a recovery rate of 21%. The estimated molecular mass of the purified protease was approximately 45 kDa. The optimal pH and temperature of the purified protease were 7.0 and 55 °C, respectively. The heat resistance of the purified protease was significantly higher than the commercial protease. The effect of metal ions on the activity of the purified protease approximated that of commercial neutral protease. Furthermore, the maximum hydrolysis rate (V max) and apparent Michaelis–Menten constant (K m) values of the purified protease were 256.4103 μg/mL min and 20.0769 mg/mL, respectively. The purified protease had a higher affinity for the substrate than the commercial neutral protease. All the results suggest that this neutral protease exhibits the potential for application in industry due to its good resistance to high temperatures and wide range of acids and bases.

A fibroin-degrading bacterium was isolated in a medium containing fibroin as a sole nitrogen source, and then identified and characterized. The strain was designated Bacillus subtilis strain Bs5C, and the culture medium containing the extracellular proteolytic enzymes was found to hydrolyze cocoon fiber. A decomposition rate of 35% was achieved with the culture medium in optimized cocoon-degrading condition. In a second round of decomposition performed on the same fibers with additional culture medium, the decomposition rate reached 45.9%. To investigate the proteolytic enzymes degrading cocoon, the enzymes was purified from culture medium using ion exchange column and identified using LC-MS/MS analysis system. As a result, it was uncovered that the cocoon degradation was due to the neutral protease NprE of the strain Bs5C. Moreover, the cocoon fibers treated with semi-purified NprE enzyme solution were clearly split and degraded, as assessed by SEM, and solubilized peptides from fibroin and sericin were detected by LC-MS/MS. In conclusion, this study is the first report that peptides could be produced from cocoon by cultured medium of B. subtilis, and the newly isolated strain Bs5C and the NprE protease from the strain Bs5C could clearly be valuable for the production of silk peptides, which have various pharmacological and industrial applications. [ABSTRACT FROM AUTHOR]

Purpose: Human islet isolation requires a defined collagenase-protease enzyme combination for obtaining a successful islet yield. While different islet laboratories use different enzyme combinations, a systematic methodology to identify optimal enzyme combinations and their concentrations within a single donor pancreas has not been tested. In this study, we designed a trisected pancreas model to test efficacy of three clinical grade enzyme blends (VitaCyte, Roche, SERVA) within a single pancreas. Methods: Islet isolations were performed using brain-dead donor pancreases (n = 15) applying the enzyme-related design of experiments (DOEs) and the trisected model approach. After trimming, split each pancreas into three individual lobes (head, body, tail). As per the DOEs, the lobes were altered between different experiments, to minimize anatomical bias. Islets isolated from each lobe (27 lobes totally) were subjected to functional assessments. Insulin staining and islet area fraction were determined for tissue sections obtained from each lobe. Results: Utilizing the trisected model, we identified that the collagenase dose from three different vendors did not affect the pancreas digestion and islet yield, but islet morphology after isolation with the neutral protease and BP-protease was better than thermolysin. In addition, the head lobe yielded a lower islet mass and higher tissue volume compared to other two lobes, irrespective of enzyme combination used. Conclusions: This study demonstrates that the trisected model is a promising methodology in assessing donor and isolation associated parameters. Based on this study, we conclude that the donor characteristics and an optimal enzyme dose play a critical role in achieving higher islet yields. [ABSTRACT FROM AUTHOR]

Successful clinical human allo or auto-islet transplantation requires the recovery of a sufficient number of functional islets from either brain-dead or chronic pancreatitis pancreases respectively. In the last two decades (2000–2019), significant progress has been made in improving the human islet isolation procedures and in standardizing the use of different tissue dissociation enzyme (TDE; a mixture of collagenase and protease enzymes) blends to recover higher islet yields. This review presents information focusing on properties and role of TDE blends during the islet isolation process, particularly emphasizing on the current developments, associated challenges and future perspectives within the field. • Successful human islet isolation and transplantation indicate importance of intact class 1 collagenase. • Recombinant rC1rC2 collagenases have beneficial effects on human islet yield and function. • Increasing protease to collagenase dose improves recovery of human islets from young donor pancreases. • Low-cost enriched collagenase-purified protease enzyme mixtures for successful human islet isolation. [ABSTRACT FROM AUTHOR]

Introduction: Cost-effective production of proteases is one of the challenges facing the enzyme industry. In this regard, there are no reports concerning the influence of lowintensity electric current and nanoparticles on microbial neutral proteases production. This study aimed at investigating the effect of these treatments on the neutral protease production of Aeromonas hydrophila MSB16. Materials and methods: The protease production of A. hydrophila MSB16 in the presence of ionic and non-ionic surfactants (SDS, Tween 80, Triton X-100, and Span 80), glycerol as well as various nanoparticles (Ag, Fe III, Fe (0), Al, Zn, and Cu), was investigated. Furthermore, the effect of an electric current (50 μA) exposure to the bacterial cells during the logarithmic and stationary phase for 10 min and 20 min was studied. Results: According to the results, electric current exposure to the bacterial cells entered the stationary phase for 10 min and 20 min increased the protease production of A. hydrophila MSB16 by 48.2% and 59.1%, respectively. However, this positive effect was not observed for log phase-bacterial cells. Besides, glycerol, Tween 80, Span 80 and Ag nanoparticles enhanced the protease production of A. hydrophila MSB16 by 56.4%, 40.4%, 24.8%, and 12.5%, respectively. Discussion and conclusion: Non-ionic surfactants, glycerol, and electric current increased the protease production of A. hydrophila MSB16. Interestingly, the influence of electric current on the protease production of A. hydrophila MSB16 was dependent on the growth phase of the bacterium. [ABSTRACT FROM AUTHOR]

Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) is a widely used technique to analyze the purity of a protein. However, it is necessary to denature (via boiling) the samples before subjecting them to electrophoresis. In the case of protease-containing samples, autolysis of the protease can occur, affecting the accuracy of results. In this study, we investigated the methods for analyzing the purity of Dispase I, a thermolysin-like neutral protease. When we analyzed D protease, a neutral metalloprotease component of Dispase I and highly purified Dispase I using the conventional SDS-PAGE method, a large number of bands were detected in both cases. These bands (putative D protease fragments) were assumed to result from autolysis. To inactivate D protease (optimal pH 7–8), 0.05 M sulfuric acid was utilized (pH 0.7–2.5). Using a conventional sample preparation solution, acid-treated Dispase I samples (without boiling) were made, and SDS-PAGE (15% w/v gel) was carried out. Our findings show that autolysis was inhibited under strong acidic conditions, and protein denaturation was achieved by treatment with sulfuric acid and SDS without boiling. Using this modified SDS-PAGE method, the purities of Dispase I and the purified enzyme were determined to be approximately 80% and 98%, respectively. Furthermore, we demonstrated that this method can be applied for the analysis of other samples including non-acidic proteases (e.g., thermolysin, subtilisin, and trypsin) and protease-contaminated samples (a mixed solution of albumin and D protease). • Protease autolysis was inhibited in SDS-PAGE with sulfuric acid without boiling. • This method could measure the purity of the Dispase preparation for the first time. • This method is applicable to non-acidic proteases and protease-contaminated samples. [ABSTRACT FROM AUTHOR]

• The oil transfer was studied in the soybean emulsion hydrolyzed by protease. • The quantitative transfer of soybean oil extracted by petroleum ether was studied. • The organic extraction promoted the oil release and eliminated emulsification. • The process of enzyme-assisted aqueous extraction of soy oil was explored. • The process is lower economic cost and is easy to be industrialized. Enzyme-assisted aqueous Extraction of soybean oil is a promising green and safe alternative to traditional process. However, it is quite difficult to realize industrial application as the process needs large dosage of special enzyme and high-speed separation. To overcome these problems, the quantitative transfer of oil release during the hydrolysis of soybean protein by protease was studied. The results showed that the 1398 neutral protease with the total amount of only 720 IU/g soybean could be used to hydrolyze the soy protein, and the oil yield reached up to 90% after separation by the centrifugal force of 1960 × g. The enzymatic hydrolysis of soybean protein was beneficial to release and separate oil from soybean, and the separated protein could enrich almost 99.0% of oil from hydrolyzed emulsion. At the moment, the emulsification could be eliminated during the process of aqueous extraction with petroleum ether. [ABSTRACT FROM AUTHOR]

A propanol-tolerant neutral protease was purified and characterized from Bacillus sp. ZG20 in this study. This protease was purified to homogeneity with a specific activity of 26,655 U/mg. The recovery rate and purification fold of the protease were 13.7% and 31.5, respectively. The SDS-PAGE results showed that the molecular weight of the protease was about 29 kDa. The optimal temperature and pH of the protease were 45 °C and 7.0, respectively. The protease exhibited a good thermal- and pH stability, and was tolerant to 50% propanol. Mg2+, Zn2+, K+, Na+ and Tween-80 could improve its activity. The calculated Km and Vmax values of the protease towards α-casein were 12.74 mg/mL and 28.57 µg/(min mL), respectively. This study lays a good foundation for the future use of the neutral protease from Bacillus sp. ZG20. [ABSTRACT FROM AUTHOR]

Removal of protein from plant polysaccharide extract is often regarded as essential step for antioxidant activity evaluation. The aim of this study was to compare two different methods (neutral protease and Sevag) for efficient removal of protein from Lentinus edodes stipe polysaccharide extract. Orthogonal test was used to optimize the parameters of neutral protease deproteinization effects on polysaccharide extracted from Lentinus edodes stipe. Furthermore, antioxidant activity was also investigated by the two deproteinization methods. Results showed that the optimum parameters of deproteinization by neutral protease were pH of 6, temperature of 60 °C, time of 1 h, and enzyme amount 0.2%. Under these conditions, the polysaccharide retention rate was 92.29%, and the deproteinization rate was 84.30%. This result was higher than that of Sevag, which deproteinization rate and polysaccharide retention rate were 55.43% and 82.29%, respectively. The antioxidant activity of polysaccharide declined after the polysaccharide deproteinized for both techniques, however, the polysaccharide deproteinized by Sevag showed the lowest antioxidant activity. Therefore, neutral protease could be used for deproteinization of polysaccharide extracted from Lentinus edodes stipe. [ABSTRACT FROM AUTHOR]

Extracellular Polymeric Substances (EPS) are complex high-molecular-weight secretions from microorganisms produced by cell lysis and adsorbed organic matter in wastewater that has crucial effects on sludge dewaterability. In this study, the effects of neutral protease and sodium dodecyl sulfate (SDS) on the amount and characteristics of EPS and on the moisture distribution in mechanical dewatered sludge were observed. The treatment was effective, and the content of EPS increased by 241% after treatment with 0.15 g/g total suspended solid (TSS) SDS and 0.03 g/g TSS neutral protease. Moreover, the ratio of PN/PS increased from 5.76 (blank) to 7.81 (treated with 0.10 g/g TSS SDS and 0.03 g/g TSS neutral protease). The results also showed that the content of bound water decreased from 7.40 to 6.29 g/g TSS in response to the addition of 0.15 g/g TSS SDS and 0.03 g/g TSS neutral protease, and the percentage of moisture with a lower binding force increased from 96.07 to 97.60% after treatment, which indicated that the dewaterability of sludge improved after treatment. Furthermore, 3D-EEM analysis indicated that the aromatic proteins and soluble microbial by-product-like materials were dominant fluorescence materials in EPS, and the content of simple aromatic proteins such as tyrosine and tryptophan in LB-EPS decreased after the addition of neutral protease and SDS, while the fluorescence materials in S-EPS and TB-EPS induced no obvious change between the original and treated sludge. [ABSTRACT FROM AUTHOR]

In this work, we have reported an enzymatic activity and biochemical properties of extracellular proteases from Chromohalobacter salexigens BKL5 and Micrococcus luteus 11A. C. salexigens BKL5 and M. luteus 11A were previously isolated from Bledug Kuwu mud volcano and dietary industry wastewater treatment, respectively. Both bacterial strains were able to produce extracellular proteases, when grown on minimal agar medium supplemented with 1% of skim milk. Proteolytic indexes of C. salexigens BKL5 and M. luteus 11A were 2.5±0.14 and 2.9±0.42, respectively. Both extracellular proteases exhibited optimum enzymatic activity at pH 7, with specific activity of C. salexigens BKL5 was 13.3% higher than that of M. luteus 11A. Optimum temperature for enzymatic activity of both proteases was 45°C. Metal cofactor preferences assay showed that extracellular protease from C. salexigens BKL5 preferred Zn2+, meanwhile extracellular protease from M. luteus 11A mainly preferred Ca2+ ion. Metal cofactor preferences assay also suggested that crude extracellular protease from C. salexigens BKL5 was categorized as metalloprotease, meanwhile crude extracellular protease of M. luteus 11A was common neutral protease. The enzymatic stability assay against various salt concentrations showed that crude extracellular protease from C. salexigens BKL5 was more stable than that of M. luteus 11A.

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Hydrolysis can affect the foaming capacity and stability of egg white protein (EWP), while the relevance of structure and foaming properties were not elucidated. Herein, the changes of structure, physico-chemical and foaming properties of EWP hydrolysates with different hydrolysis times were investigated. The foaming ability was increased from 69.67 to 108.33% after 15 min of neutral protease-mediated hydrolysis. The results also indicated hydrolysis process increased the ratio of the polypeptides with small molecular weight (<10 kDa), this condition promotes EWP's adsorption to the air-water interface. Besides, intrinsic fluorescence spectroscopy indicated that the EWP's structure occurred to unfoldment and exposed to environment with strong polarity in hydrolysis, while the free sulfhydryl contents were increased. Meanwhile, the result of Fourier transforms infrared spectrometry confirmed these structural changes. This research demonstrated that enzymatic hydrolysis might become a beneficial method to obtain dominant foaming agents in the food process and industry. [Display omitted] • Moderate hydrolysis can improve the foaming property of egg white protein. • The 15 min hydrolysate had the most compact and uniform foam. • Molecular weight reduction of egg white protein benefits to foaming properties. • The interchange reactions of SH-SS conduct a contrary effect of foaming property. [ABSTRACT FROM AUTHOR]