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Dry-cured meat products are considerably popular around the world due to unique flavor. Proteolysis is one of the enzymatic reactions from which flavor substances are derived, which is affected by endogenous proteases. The purpose aimed to reveal the potential relationship between endogenous proteases and key flavor substances in dry-cured pork coppa in this paper. The dynamic changes of endogenous proteases activity, free amino acids, and volatiles during dry-cured pork coppa processing were characterized. The results showed that 5 kinds of free amino acids, Glu, Lys, Val, Ala, and Leu, were identified as significant contributors to taste. Meanwhile, key volatiles, such as hexanal, nonanal, octanal, benzaldehyde, 3-methyl butanoic acid, 2-methyl propanoic acid, and ethyl octanoate, greatly contributed to the flavor characteristics of dry-cured pork coppa. Further partial correlation analysis was performed to better elucidate the relationship among parameters. The results revealed that close relationship between endogenous proteases and key substances. RAP not only significantly affected the accumulation of key active-amino acids, but also affected the accumulation of ethyl octanoate, 2,3-pentanedione, and 2,3-octanedione by regulating the accumulation of octanoic acid and Leu. In addition, cathepsin B and D, DPP Ⅱ, DPP Ⅳ and RAP notably affected accumulation of hexanal.

Water-free transportation (WFT) causes shrimp (Penaeus vannamei) flesh quality deterioration. However, the roles of endogenous protease-induced protein hydrolysis have been neglected in the research. In the present study, calpain zymography, gelatinase zymography, the hematoxylin–eosin staining method, and other methods were applied to investigate the response of various endogenous proteases (cathepsin, calpain, and gelatinase), the myofibril fragmentation index (MFI), and the microscopic morphology of shrimp muscle during WFT in comparison with the shrimp under the conventional water transportation strategy (WT). The results showed that the total activity of proteases in shrimp muscle increased significantly (p ≤ 0.05) after simulated transportation. Cathepsins and gelatinases were activated during WFT. No significant (p > 0.05) changes of the activity of caspase-3 and the muscle cell apoptosis rate were detected in shrimp muscle cells after WFT. In addition, the MFI increased and the gap among muscle fiber bundles enlarged after WFT. Compared with WFT, no significant (p > 0.05) effect on the activities of calpain, gelatinase, and caspase-3 in the muscle of shrimp was found after WT, and only the activity of cathepsin L significantly increased (p ≤ 0.05). Based on the findings, we concluded that the activation of various endogenous proteases was induced during WFT.

NaCl is the main curing agent in dry-cured meat products, and a large amount of NaCl addition leads to high salt content of final products. Salt content and composition are important factors affecting the activity of endogenous proteases, which in turn could affect proteolysis as well as the quality of dry-cured meat products. With the increasing emphasis on the relationship between diet and health, reducing sodium content without sacrificing quality and safety of products is a great challenge for dry-cured meat industry. In this review, the change of endogenous proteases activity during processing, the potential relationship between sodium reduction strategy, endogenous proteases activity, and quality were summarized and discussed. The results showed that sodium replacement strategy and mediated-curing had a complementary advantage in influencing endogenous proteases activity. In addition, mediated-curing had the potential to salvage the negative effects of sodium substitution by affecting endogenous proteases. Based on the results, a sodium reduction strategy that sodium replacement in conjunction with mediated-curing based on endogenous proteases was proposed for the future perspective. [ABSTRACT FROM AUTHOR]

为了对薄壳山核桃仁中的主要蛋白质，特别是相对丰度较低的功能活性蛋白进行系统研究，以新鲜的薄壳山核桃仁为原料，利用电泳、液相串联质谱和氨基酸分析等方法，考察了薄壳山核桃仁的主要蛋白质和功能活性蛋白（抗氧化酶、内源性蛋白酶），并对内源性蛋白酶的最适pH和温度进行了研究。结果显示：薄壳山核桃仁的主要蛋白质包括11S球蛋白（58%）、7S球蛋白（17%）和9 kDa蛋白（15%），还含有0.239%超氧化物歧化酶、0.190%过氧化氢酶和0.282%内源性蛋白酶；只检测到1种蛋白酶抑制剂，并且其相对丰度极低；内源性蛋白酶在pH 3.5~5.5和40~60 ℃时可发挥水解活性，并在pH 4.5和50 ℃时活性最强；在氨基酸组成方面，薄壳山核桃仁蛋白的赖氨酸含量低（3.74%），但谷氨酸+谷氨酰胺含量（19.51%）和精氨酸含量（14.12%）高。说明国产薄壳山核桃适宜鲜食，对促进人体健康有益。 In order to systematically examine the major proteins in pecan kernels, especially the bioactive proteins with low relative abundance, the major proteins and bioactive proteins(antioxidant enzymes, endogenous proteases) in fresh pecan kernels were investigated by tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis, liquid chromatography tandem mass spectrometry and amino acid analysis.The optimum pH and temperature of the endogenous proteases were also studied. The results showed that the major proteins in pecan kernels included 11S globulins (58%), 7S globulins (17%) and 9 kDa proteins (15%), as well as 0.239% superoxide dismutases, 0.190% catalases and 0.282% endogenous proteases. Only one protease inhibitor was detected, and its relative abundance was extremely low. These endogenous proteases showed hydrolytic activity at pH 3.5-5.5 and 40-60 ℃, and optimal at pH 4.5 and 50 ℃. As for the amino acid composition, the proteins in pecan kernels contained a low content of lysine (3.74%), but a high content of glutamic acid + glutamine (19.51%) and arginine (14.12%). It is suggested that domestic pecans are suitable for fresh consumption, which is benifical for promoting human health.

Mung bean protein possesses several health benefits, and aqueous processing methods are used for its production. However, mung bean protein yields are different with different methods, which are actually different in conditions (e.g., pH, temperature, and time). Herein, liquid chromatography tandem mass spectrometry identified 28 endopeptidases and exopeptidases in mung bean protein extract, and the positions of 8S and 11S globulins on sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) gel were confirmed in our experimental conditions. The SDS-PAGE, trichloroacetic acid-nitrogen solubility index, and free amino acid analysis revealed that (1) 8S globulins showed strong resistance to the endopeptidases (optimal at pH 5 and 50 °C) at pH 3-9, and 11S globulin exhibit strong resistance expect at pH 3-3.5; (2) the exopeptidases (optimal at pH 6 and 50 °C) preferred to liberate methionine and tryptophan. These proteases negatively affected protein yield, and short production time and low temperature were recommended., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

The sea cucumber is an essential nutrient source and a significant economic marine resource associated with successful aquaculture. However, sea cucumbers are highly susceptible to autolysis induced by endogenous protease after postmortem, and the phenomenon of body wall “melting” occurs, which seriously affects the food quality of products and the degree of acceptance by consumers. To satisfy the growing demand for fresh or processed sea cucumbers, we must clarify the autolysis mechanism of sea cucumbers and the methods to achieve autolysis regulation. In this paper, the factors leading to the quality deterioration and texture softening of sea cucumbers are reviewed, with emphasis on enzymatic characteristics, the autolysis mechanism, the effects of autolysis on the physicochemical properties of the body wall of the sea cucumber, and the development of potential natural protease inhibitors. We aim to provide some reference in future preservation and processing processes for sea cucumbers, promote new processing and preservation technologies, and advance the sea cucumber industry’s development.

The aim of this study was to isolate the e ndogenous proteases in the shrimp head of Lito penaeus vannamei and investigate their enzymol ogy characterization. The results show that ther e are10 types of endogenous proteases existing in the shrimp head, which are successively iden tified as chymotrypsin, cathepsin L, collagenase, calpain, pepsin, metalloprotease, trypsin, serine protease, cathepsin B and MBSP, respectively. In addition, pH and temperature have a significan t impact on the activities of proteases P1, P4-P8, and their optimal pHs and temperature are in the range of 6.0-9.0 and 30-40 °C, respectively; proteases P5-P7 own better stability than that of other proteases; the optimal temperature for P1, P4-P8 stability is 30 °C, and that of other prot eases is 40 °C. In conclusion, proteases P1 to P10 are isolated and characterised, which is an i mportant support to develop high-quality condi ments and efficiently recover proteins from shri mp head waste. [ABSTRACT FROM AUTHOR]

This study evaluated the food safety and proximate composition of shrimp head (SH). Potentially toxic elements in SH were below European Union legislation limits. SH had a high content of tasting amino acids (sweet and umami amino acids was 57%) and a high content of functional amino acids (essential amino acids was 37%). Moreover, the changes of flavor and key umami molecules in SH were studied by sensory evaluation, electronic tongue, electronic nose, automated amino acid analyzer, and high performance liquid chromatography (HPLC). The results showed that the significant difference of flavor in SH happened during autolysis. SH with autolysis had the best umami taste at 6 h, which may result from the synergistic work of free amino acids and nucleotide related compounds. Additionally, the performance of endogenous proteases in SH was investigated to efficiently analyze autolysis. The optimum pH and temperature of endogenous proteases in SH were 7.5 and 50 °C, respectively. The autolysis of SH depends on two endogenous proteases (~50 kDa and ~75 kDa). These results suggest that the formation of flavor in SH during autolysis can be controlled, which could provide guidance for SH recycle. SH could consider as one of the food materials for producing condiments.

Water-free transportation (WFT) causes shrimp (Penaeus vannamei) flesh quality deterioration. However, the roles of endogenous protease-induced protein hydrolysis have been neglected in the research. In the present study, calpain zymography, gelatinase zymography, the hematoxylin–eosin staining method, and other methods were applied to investigate the response of various endogenous proteases (cathepsin, calpain, and gelatinase), the myofibril fragmentation index (MFI), and the microscopic morphology of shrimp muscle during WFT in comparison with the shrimp under the conventional water transportation strategy (WT). The results showed that the total activity of proteases in shrimp muscle increased significantly (p ≤ 0.05) after simulated transportation. Cathepsins and gelatinases were activated during WFT. No significant (p > 0.05) changes of the activity of caspase-3 and the muscle cell apoptosis rate were detected in shrimp muscle cells after WFT. In addition, the MFI increased and the gap among muscle fiber bundles enlarged after WFT. Compared with WFT, no significant (p > 0.05) effect on the activities of calpain, gelatinase, and caspase-3 in the muscle of shrimp was found after WT, and only the activity of cathepsin L significantly increased (p ≤ 0.05). Based on the findings, we concluded that the activation of various endogenous proteases was induced during WFT. [ABSTRACT FROM AUTHOR]

Sesame seeds is recognized as a good source of lipids and proteins, and its processing is mainly focused on oil production. Sesame is rich in a variety of essential amino acids for human body, which has great development and utilization value. However, the processing of sesame proteins is limited by its poor solubility. Due to the sustainability, economical costs and health benefits of plant proteins, researchers have greatly increased their interest in them. At present, aqueous extraction processing has the capability to realize the comprehensive utilizations of lipids and proteins in sesame. However, the cost of commercial proteases is considerable for large scale applications. Previous studies in our laboratory have confirmed that endogenous proteases in sesame seeds have high activity, and it has great potential for aqueous extraction processing of sesame. While preparing lipid products, endogenous proteases could be used to biologically convert sesame proteins into hydrolysate with high solubility and nutritional function. In addition, in the previous study, the effects of pH value, temperature, and time on proteolytic activity of endogenous proteases in sesame milk were studied, and the proteins were optimally hydrolyzed at pH 4.5 value and 50 ℃ for 6 h catalyzed by the endogenous proteases. In this paper, wet-peeled sesame was dried and pressed in order to obtain the sesame oil and pressed meal. Then, the pressed meal was then hydrolyzed into hydrolysate by endogenous proteases. And the effects of drying temperature on pressed oil yield and proteolytic activity of endogenous proteases in wet-peeled sesame were investigated, which provides reference for the practical applications of endogenous proteases. The results showed that under hydraulic pressing (80 MPa, 1 h), the oil yield of sesame and peeled sesame (50 ℃, 4 h) (the total lipids in sesame as 100%) were 81.76% and 83.57% respectively, indicating that peeling is conducive to the improvement of oil yield. The oil yield of wet-peeled sesame increased to 88.55%-90.98% after drying at 80-180 ℃ for 1 h. For the endogenous proteases, its activity decreased slowly at 80-130 ℃ and remained about 57% at 130 ℃, indicating that endogenous proteases had good thermal stability, then the activity decreased rapidly after 130 ℃ and inactivated at 180 ℃. In order to modify proteins in the pressed meal with endogenous proteases, sesame seeds were dried at 50 ℃ for 4 h to obtain sesame oil and low-denatured pressed meal. After grinding, the sesame milk prepared from pressed meal was incubated at pH 4.5 value and 50 ℃ for 6 h. And hydrolyzed sesame milk was separated into two fractions (skim and precipitation) by centrifugation (3 000 g, 5 min). The results showed that the proteins in the skim still accounted for 66% of total proteins in sesame milk after centrifugation, and the proteins in skim were composed of 38% small peptides (< 1 500 Da) and 37% free amino acids. In addition, there are many bioactive substances in the skim, such as ornithine, serotonin, DL-dopa and so on. The purposes of this study were to evaluate the thermal stability of endogenous proteases in sesame seeds and to develop a new method for production of sesame protein hydrolysate, which enhanced the solubility and bioactive properties of sesame proteins. [ABSTRACT FROM AUTHOR]

The study evaluated changes in total enzymatic activity and degradation of demineralized dentin following the application of universal or self-etch adhesives. The universal adhesives --Scotchbond Universal (SU) and All-Bond Universal (ABU) and self-etch adhesives --Adper Easy Bond (EB) and G-aenial Bond (GB) were used for 2 min pretreatment of the dentin beams. Phosphoric acid (PA) treatment as well as no treatment served as controls. Total enzymatic activity was analyzed before and after treatment, collagen degradation was assessed using mass loss, C-terminal telopeptide (CTX) and C-terminal-telopeptide of type I collagen (ICTP) release (24 h, 3-day, 3-week). Over three weeks of incubation, ICTP release of ABU treated beams was significantly higher than other groups (p<0.05), except for SU treated beams (p>0.05) and CTX release of GB treated beams was the highest among the groups with statistically significant difference (p<0.05). The results confirm that the universal adhesives tested have also potential to increase the enzymatic activity in dentin. [ABSTRACT FROM AUTHOR]

Identification and separation of endogenous proteases are important to find out their role in changes of biochemical process in post‐mortem muscle. Diverse studies were conducted to find out the activity of these enzymes but only a very few studies were carried out on avian species. In this study, a simple casein zymography method was explained for detection of calpain‐1 and calpain‐2 with the standardization of extraction buffer and that were quantified biochemically after one‐step separation and purification on anion exchange chromatography. DEAE Sephacel matrix was used to purify calpain‐1, calpain‐2, and calpastatin. SDS‐PAGE analysis showed native and autolyzed forms of calpains. Biochemical test indicated concentrations of calpain‐1, calpain‐2, and calpastatin being higher in breast than thigh muscles across all four breeds of chicken. A simple, effective, and low‐cost extraction, purification, and separation technique was developed for accurate identification of calpain‐1and calpain‐2 on casein zymography method and quantification by biochemical test. Novelty Impact Statement: Identification and isolation of endogenous proteases are necessary to determine their significance in the PM aging of chicken meat from various breeds. In this study, a straightforward casein zymography approach for in‐vitro detection of calpain‐1 and calpain‐2 activity is devised. This will aid in the optimization of PM aging duration for meat tenderization. [ABSTRACT FROM AUTHOR]

Endogenous proteases with high activity have been identified in sesame seeds. However, the hydrolyzing behaviors of endogenous proteases on proteins in sesame milk are not well understood. In this study, the endogenous proteases optimally hydrolyzed proteins at pH 4.5 and 50 °C for 6 h. Tricine-sodium dodecyl sulphate-polyacrylamide gel electrophoresis and liquid chromatography tandem mass spectrometry analyses revealed that endogenous proteases randomly cleaved the cleavable peptide bonds on 11S globulins, and amino acid analysis indicated that serine carboxypeptidases preferentially cleaved tryptophan, phenylalanine, methionine, tyrosine, and leucine. The hydrolyzed sesame milk was separated into cream, transparent skim, and precipitate fractions by centrifugation (3000g, 5 min). The major protein components in skim were 42% peptides (<1500 Da) and 35% free amino acids. The phytate in skim was greatly reduced by adjusting to neutral and alkaline pH. This study is meaningful at supplying a strategy for producing low-phyate sesame protein hydrolysate., (Copyright © 2022 Elsevier Ltd. All rights reserved.)

Summary: Effects of endogenous enzymes on texture deterioration of Macrobrachium rosenbergii during frozen storage (−18 °C) were investigated. The prawn was pretreated with iodoacetic acid (IAA) solution to inhibit cysteine proteases and/or phenylmethylsulfonyl fluoride (PMSF) solution to inhibit trypsin before frozen storage, and sterile distilled water was used as the control. After 16 weeks of cryopreservation, the shear force of the groups treated with enzyme inhibitors was significantly higher than that of sterile distilled water‐treated group (control; P < 0.05), while significantly fewer TCA‐dissolved peptides, α‐amino nitrogen and Myofibril fragmentation index (MFI) value were observed in all inhibitor‐treated groups (P < 0.05). Compared with PMSF treatment, IAA or IAA + PMSF treatment effectively alleviated softening and remarkably reduced the accumulation of TCA‐soluble peptides and AAN (α‐amino nitrogen) during frozen storage, with a lower MFI value and a higher sodium dodecyl sulphate‐polyacrylamide gel electrophoresis band intensity. Therefore, endogenous cysteine proteases in muscle play a crucial part in texture deterioration of giant freshwater prawn during cryopreservation. [ABSTRACT FROM AUTHOR]

Texture deterioration occurs in adductor muscle of scallop (Argopecten irradians) (AMS) after 5 d of cold storage. Principal component analysis indicated the texture deterioration resulted in significant decrease of hardness, springiness, adhesiveness and chewiness, but significantly increased cohesiveness. Endogenous proteases degraded structural proteins, among which cysteine proteases were mainly responsible for myofibrillar proteins (MPs) degradation, while serine proteases degraded both MPs and connective tissue proteins. Pearson coefficient analysis showed that texture indicators significantly correlated with structural protein indicators in AMS. To be more specific, the hardness, springiness, adhesiveness and chewiness negatively correlated with myofibrillar fragmentation index, soluble hydroxyproline (Hyp) and soluble glycosaminoglycans, but positively correlated with solubility of MPs and water holding capacity. Meanwhile, the cohesiveness positively correlated with soluble Hyp. The Taylor diagram and Hierarchical cluster analysis confirmed that the inhibitors of cysteine and serine proteases could effectively retard textural deterioration of AMS during 5 d of cold storage., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier Ltd. All rights reserved.)