Acid Protease Research Articles

Characterizing the gastrointestinal development and digestive enzyme ontogeny of larval Amphiprion ocellaris

Clownfish (Amphiprion ocellaris) are the most recognizable and popular marine aquarium fish and are a key commodity for marine ornamental fish producers. Although these fish have been raised in captivity for decades, producers are still experiencing issues regarding larval survival, reliance on live feeds, and post-larval deformities. To improve the culture process, the ontogeny of larval development and digestive capacity should be elucidated. The morphological development, digestive enzyme ontogeny, and digestive tract development of A. ocellaris was examined in a 16-day trial. Standard microplate assays were conducted to quantify the activity of trypsin, lipase, and acid protease enzymes from 24 h before hatching to 15 days post hatch (DPH). Histology was used to visualize morphological changes in the digestive tract throughout the larval period. At hatching, A. ocellaris has functional eyes, an open mouth, and a coiled, differentiated digestive tract. Trypsin and lipase activities were detectable before hatching and increased significantly later in the larval period. Gastric gland formation was observed at 6 DPH, with stomach functionality occurring between 7 and 9 DPH, indicated by gastric gland proliferation and an increase in acid protease activity. The results from this study may be used to inform weaning protocols to maximize larval growth and survival while minimizing the use of costly live feeds.

Immunohistochemical analysis of cathepsin-D in benign and malignant salivary gland neoplasms. Evaluation of its role as a prognostic indicator.

Salivary gland tumors are histologically the most heterogeneous group of tumors. Traditional diagnostics and grading of these tumors often fail to provide any insight into patient's clinical course. Cathepsin-D is a lysosomal acid protease secreted in increased levels in several malignancies. However, its role in salivary gland tumors has not been studied extensively. The present study aims to assess the expression of cathepsin-D in malignant and benign salivary gland tumors and to compare its expression in these tumors. A total of 40 salivary gland tumors which included 10 cases each of adenoid cystic carcinoma, mucoepidermoid carcinoma, pleomorphic adenoma and Warthin's tumor were evaluated immunohistochemically for cathepsin-D expression. Intergroup comparison of cathepsin-D expression was done between the benign and malignant groups using the Mann-Whitney U-test. Intragroup comparison was also done using Kruskal-Wallis test. A statistically significant difference in the expression of cathepsin-D was observed between the benign and malignant groups. The malignant group showed a considerably higher cathepsin-D expression (mean value 6.284 ± 4.377) when compared to the benign group (mean value 2.281 ± 2.459). The differences in the immunopositivity between the malignant and benign groups were found to be highly significant (P = 0.004). Increased expression of cathepsin-D is observed in the malignant salivary gland neoplasms. This may prove to be a useful marker for the aggressive biologic behavior as well as invasive potential of salivary gland neoplasms.

The emerging role of Deubiquitinases (DUBs) in parasites: A foresight review.

Before the discovery of the proteasome complex, the lysosomes with acidic proteases and caspases in apoptotic pathways were thought to be the only pathways for the degradation of damaged, unfolded, and aged proteins. However, the discovery of 26S and 20S proteasome complexes in eukaryotes and microbes, respectively, established that the degradation of most proteins is a highly regulated ATP-dependent pathway that is significantly conserved across each domain of life. The proteasome is part of the ubiquitin-proteasome system (UPS), where the covalent tagging of a small molecule called ubiquitin (Ub) on the proteins marks its proteasomal degradation. The type and chain length of ubiquitination further determine whether a protein is designated for further roles in multi-cellular processes like DNA repair, trafficking, signal transduction, etc., or whether it will be degraded by the proteasome to recycle the peptides and amino acids. Deubiquitination, on the contrary, is the removal of ubiquitin from its substrate molecule or the conversion of polyubiquitin chains into monoubiquitin as a precursor to ubiquitin. Therefore, deubiquitylating enzymes (DUBs) can maintain the dynamic state of cellular ubiquitination by releasing conjugated ubiquitin from proteins and controlling many cellular pathways that are essential for their survival. Many DUBs are well characterized in the human system with potential drug targets in different cancers. Although, proteasome complex and UPS of parasites, like plasmodium and leishmania, were recently coined as multi-stage drug targets the role of DUBs is completely unexplored even though structural domains and functions of many of these parasite DUBs are conserved having high similarity even with its eukaryotic counterpart. This review summarizes the identification & characterization of different parasite DUBs based on in silico and a few functional studies among different phylogenetic classes of parasites including Metazoan (Schistosoma, Trichinella), Apicomplexan protozoans (Plasmodium, Toxoplasma, Eimeria, Cryptosporidium), Kinetoplastidie (Leishmania, Trypanosoma) and Microsporidia (Nosema). The identification of different homologs of parasite DUBs with structurally similar domains with eukaryotes, and the role of these DUBs alone or in combination with the 20S proteosome complex in regulating the parasite survival/death is further elaborated. We propose that small molecules/inhibitors of human DUBs can be potential antiparasitic agents due to their significant structural conservation.

Poly‐β‐hydroxybutyrate concentration, microbial enzymes activity, and nutritional value in biofloc system using different carbon sources and C/N ratios in common carp, Cyprinus carpio culture

AbstractThis experiment was designed with three carbon sources (Dextrin, wheat bran, and corn starch), each with two carbon to nitrogen (C/N) ratios (19 and 30) for 4 weeks. The data obtained from two variables (carbon sources, C/N ratio) were analyzed with a two‐way analysis of variance. At the end of the experiment, the results indicated that the use of varying carbon sources and C/N ratios significantly changed the nutritional value of microbial flocs (p < 0.05). Also, the results showed that both variables significantly affected the main groups of saturated, mono‐unsaturated, and polyunsaturated fatty acids in the microbial flocs (p < 0.05). Dextrin flowing corn starch at a low C/N ratio significantly increased poly‐β‐hydroxybutyrate accumulation in microbial flocs (p > 0.05). The microbial enzyme profile changed in all carbon sources but C/N ratios did not affect microbial enzymes in the treatments except in a significantly higher value for amylase and acidic protease activity at a C/N ratio of 30 (p < 0.05). Also, at the end of the experiment, total bacteria and Lactobacillus count were affected by carbon sources and the highest density was observed in wheat bran and Dextrin respectively. These findings may encourage farmers to consider specific carbon sources and C/N ratios based on culture conditions and species requirements.

An integrated pickling-bating technology for reducing ammonia-nitrogen and chloride pollution in leather manufacturing

Currently, ammonia-nitrogen (NH3–N), chloride (Cl−) and chromium (Cr) pollution from leather manufacturing processes attracts much attention and limited the sustainable development of the leather industry. In this paper, conventional trypsin bating and high-salt pickling procedures were substituted by a salt-free pickling-bating system, using acidic protease L80A and salt-free pickling agent naphthalene sulfonic acid (NSA); a novel integrated technology comprising non-ammonia bating, salt-free pickling and high-exhausting chrome-tanning methods was established to reduce NH3–N, Cl− and Cr pollution. The results indicated that the properties of the leather produced by the established method were better than the conventional method. Furthermore, the established integrated salt-free pickling-bating technology resulted in the almost elimination of NH3–N and chloride pollution from waste effluent. The total Cr utilization ratio was increased to 95% from 80%, the total Cr discharge was reduced by 74%. Scale-up experiments further verified the acceptability of this novel technology in terms of physical and organoleptic properties of the resultant leather. It can be easy to predict that the effluent liquors with a little amount of NH3–N, chloride and chromium can be disposed of conveniently. We suppose that the established novel technology is an environment-friendly technology and a way forward in the development of the modern sustainable leather industry.

CBD Supplementation Has a Positive Effect on the Activity of the Proteolytic System and Biochemical Markers of Honey Bees (Apis mellifera) in the Apiary.

Simple SummaryThe purpose of our study was to determine how CBD extract influences resistance in the hemolymph (insect blood) of honey bees in the hive test. The bees were divided into 3 groups: (CSy) bees fed with CBD in sugar syrup; (CSt) cotton strip with CBD placed in hive, (C) control bees fed sugar syrup. To determine the state of immunity, we used the analysis of the activity of the proteolytic system and biochemical markers, such as “liver tests”, and the concentration of selected ions and key compounds for the functioning of the organism. CBD extract increased the total protein concentration, proteases and their inhibitor activities in each age (except for acidic protease activities in the 21st and 28th day and alkaline protease inhibitor activities in the 28th day in the CSt group), increased concentrations of markers: ALP, AST, ALT; and glucose; triglycerides; cholesterol and creatinine. A decrease in concentration in experimental groups was noticed for urea acid and albumin compared to group C. Higher activities/concentrations of most of parameters were obtained in the CSy compared to the CSt and C. The CBD supplementation can positively influence bees’ resistance.We examined how CBD extract influences the activity of the immune system in the hemolymph of honey bees in the hive test. The bees were divided into 3 groups: (CSy) bees fed with CBD in sugar syrup with glycerin; (CSt) cotton strip with CBD placed in hive bees fed pure sugar syrup, (C) control bees fed sugar syrup with glycerin. CBD extract increased the total protein concentrations, proteases and their inhibitor activities in each age (the except for acidic protease activities in the 21st and 28th day and alkaline protease inhibitor activities in the 28th day in CSt group) in comparison with group C. In the groups with the extract there was also an increase in the enzymatic marker activities: ALP, AST (decrease on day 28 for CSt), ALT; and non-enzymatic marker concentrations: glucose; triglycerides; cholesterol and creatinine. The urea acid and albumin concentrations were lower in CSy and CSt groups compared to the C group (higher concentration of albumin was displayed by control bees). Higher activities/concentrations of most of biochemical parameters were obtained in the CSy compared to the CSt and C. CBD supplementation can positively influence workers’ immune system.

Asprosin promotes steroidogenesis and spermatogenesis with improved glucose metabolism in adult mice testis

Asprosin is an orexigenic adipokine that regulates appetite and glucose homeostasis in mammals. To date, only fragmentary findings are reported regarding its role in testicular activities. In the current investigation, immunolocalization and direct action of asprosin in adult mice testis was evaluated. Immunohistochemical and immunoblot studies were performed to analyse the testicular expression of asprosin. Intratesticular treatment of asprosin (0.1μg and 1.0μg per testis) was given to evaluate its direct action on testicular functions. Sertoli and Leydig cells were found to be immuno-positive for asprosin. Intratesticular administration of asprosin resulted into a significant increase in glucose and lactate levels along with enhanced expression of asprosin receptor OLFR734, insulin receptor (IR), glucose transporter 8 (GLUT 8), lactate dehydrogenase (LDH) activity and monocorboxylate transporters (MCT2 and 4). In addition, asprosin administration increased the testicular expression of cell proliferation (proliferating cell nuclear antigen: PCNA), cell survival (B cell lymphoma 2: Bcl2) and decreased germ cell apoptosis (Cysteine aspartic acid protease 3: Caspase 3) leading to increased sperm counts. Further, asprosin treatment resulted into increased level of total cholesterol, testosterone and steroidogenic markers (steroidogenic acute regulatory protein: StAR; 3beta-hydroxysteroid dehydrogenases: 3β HSD and 17beta-hydroxysteroid dehydrogenases: 17β HSD). Asprosin treatment promotes testicular glucose uptake and lactate synthesis to provide energy for steroidogenesis and spermatogenesis. The significant correlation between the asprosin-induced increased IR expression and increased testosterone, glucose and lactate levels suggests its role in increased survival and proliferation but decrease in germ cell apoptosis. This study proposed asprosin's role as an autocrine/paracrine regulator of testicular functions in adult mice.

Influence of the biofloc bacterial community on the digestive activity of Nile tilapia (Oreochromis niloticus)

High-throughput sequencing was performed to study the diversity of bacterial communities in the water of an aquaculture production system using Biofloc technology (BFT) and its effect on the activity of digestive enzymes in Nile tilapia, Oreochromis niloticus. Sex-reversed juveniles were reared in six circular indoor tanks (1 m3) for 12 weeks. The following treatments were performed: biofloc with 0% protein restriction in the diet (BFTH = high protein diet); biofloc with 11% dietary protein restriction (BFTL = low protein diet), and clear water control with 0% dietary protein restriction (CW). Biofloc groups showed a higher bacterial diversity and a completely different composition compared to CW, with significant increase of Actinobacteria and Planctomycetes and a decrease in Verrumicrobia and Fusobacteria. At the genus level, Flavobacterium, Thermomonas, Comamonas, Microbacterium, and Intestinibacter present in BFT significantly correlated with the activity of trypsin, aminopeptidase, and alkaline proteases. Twenty percent of the difference between the biofloc groups was due to the higher activity of acid proteases and aminopeptidases. The metagenome prediction revealed metabolic pathways differentially expressed in BFTH and BFTL. Most were similarly expressed in both treatments; except for the CoA degradation pathway I, which was expressed in BFTL, and the biosynthesis of isoprene II and mevalonate II in BFTH. The biofloc microbiota provided enzymes that, in synergy with the endogenous digestive enzymes of the fish, improved digestion and nutrient assimilation. The study provides knowledge on the exogenous enzymatic contribution and diversity of the bacterial groups present in the biofloc; the contribution of these microorganisms in digestive metabolism and fish performance is also highlighted.

Studies on the feeding habit and digestive enzyme activities in three small indigenous fish species from Assam, India

Knowledge of the feeding habit and the digestive physiology of a fish is important in making appropriate strategies for feed development and successful culture. Nutrient-rich small indigenous fish species (SIFs) are abundant in Assam, India. Puntius sophore, Mystus tengara, and Trichogaster fasciata of Gossaigaon, Assam are important SIFs for the local rural population, and also potential candidates for ornamental fish culture. The present study aims to evaluate the feeding habit and digestive enzyme activities of these species. Data obtained from the relative gut length and gut content analysis suggested that M. tengara is a carnivorous fish and the rest two fishes are omnivorous in habit. Further, the relative gut length was highest in T. fasciata (4.20±0.45) and lowest in M. tengara (0.55±0.11). Digestive enzyme activity indicates a correlation with the dietary habit of the fish. Further, total protease, trypsin, and amylase activity was reported highest in P. sophore. Acid protease pepsin was found to be significantly higher in M. tengara complementing its carnivorous habit and gut anatomy. The present study has established some important information on the digestive enzyme characteristics and feeding habits of the three fish species. This information might be useful in the development of suitable feed for the fish species for their culture.

Flavor differences of soybean and defatted soybean fermented soy sauce and its correlation with the enzyme profiles of the kojis.

Soybeans and defatted soybeans, commonly used as protein ingredients, have different flavors of their fermented soy sauce. Clarifying the differences between the two soy sauces, as well as the formation mechanism, is an important prerequisite for improving the flavor of defatted soybean soy sauce. To this goal, the aroma characteristics of two soy sauces and their volatile profiles were compared by sensory evaluation and gas chromatography-mass spectrometry, and eight enzyme activities and volatile profiles of matured koji were determined. Sensory results showed that the acids, fruity and cooked potato-like attributes were higher in whole soybean fermented soy sauce, whereas defatted soybean soy sauce exhibited higher smoky and malty attributes, closely related to the contents of aroma-active compounds in soy sauce, such as isobutyl acetate, 2/3-methylbutanal, acetic acid and 2/3-methylbutanoic acid. The content of most volatiles in the matured kojis showed a consistent trend with that of soy sauce: alcohols, acids, furan(one)s and ketones. Interestingly, acid protease and cellulase activities were 3.3 and 1.6 times higher in the whole soybean koji than in defatted soybean koji, respectively, whereas neutral protease, aminopeptidase, glucoamylase and β-glucosidase were approximately 2.0 times higher in defatted soybean koji. In summary, the flavor differences between soybean and defatted soybean fermented soy sauce were not only caused by the differences in the content of flavor precursors in the materials, but also closely related to the differences in the enzymatic profiles accumulated during the koji-making process. © 2022 Society of Chemical Industry.

Soil quality and r – K fungal communities in plantations after conversion from subtropical forest

Intensive clear-cutting of natural forests and conversion to monoculture plantations are ongoing worldwide, leading to the degradation of soil quality and microbial functions. Here, we compared soil quality index (SQI) and fungal communities in a natural forest (Forest) and four 5-year-old monoculture plantations, including Camellia oleifera (Oil), Amygdalus persica (Peach), Myrica rubra (Berry) and Cunninghamia lanceolate (Fir) in a subtropical region of China. After conversion, soil pH in the plantations rose up to 0.31, but organic carbon and total nitrogen contents, sucrase, acid protease, glutaminase, acid and alkaline phosphatase activities decreased by 83%, 59%, 40%, 64%, 66%, 94% and 59%, respectively. Correspondingly, the SQI dropped by 65%. High-throughput sequencing of the ITS1 region demonstrated an increase in α-diversity and a striking difference in β-diversity of fungi following conversion. Changes in the dominant fungal taxa following forest conversion to plantations were interpreted by r- and K-selection of life strategies. Conversion increased the fungal groups with r-strategies, such as Ascomycota and Zygomycota, but decreased the fungal groups with K-strategies, such as Basidiomycota. Genera affiliated to those phyla including Pseudophialophora, Rhytisma increased, but Russula decreased. Redundancy analysis and structural equation modeling indicated that the diversity and composition of fungal communities changed with soil degradation, which was mainly driven by increased pH and total phosphorus content, but decreased C/N ratio and C and N related enzymes activities. Overall, the conversion of forest to monoculture plantations decreased soil quality and the abundance of K-strategists, retarded the decomposition of persistent organic matter, but boosted the prevalence of r-strategists in a more diverse fungal community.

Enzyme activity of Aspergillus section Nigri strains isolated from the Korean fermentation starter, nuruk.

Aspergillus section Nigri is a fungus used industrially because of its ability to produce enzymes such as cellulolytic, amylolytic and proteolytic enzymes. In this study, we obtained twenty-eight strains of Aspergillus section Nigri from the traditional Korean fermentation starter, nuruk, which is known as a mixed culture of enzymatic filamentous fungi and yeasts. All strains were identified as Aspergillus section Nigri through combined phylogenetic analysis using partial β-tubulin and calmodulin gene sequences. The cellulase, amylase and protease activities of Korean strains were measured and compared with ten reference strains of Aspergillus niger. Most Korean strains showed higher cellulolytic activity than reference strains, and Aspergillus neoniger KCN5 showed the highest β-glucosidase activity. Two-thirds of the Korean strains showed similar levels of α- and glucoamylase activity as the reference strains. The protease activity of Aspergillus section Nigri strains was the highest at pH 3.0, and A. niger KSJ2 showed the highest acidic protease activity. By comparing ten reference strains and twenty-eight Korean strains, our results suggested useful Aspergillus section Nigri strains from nuruk with high enzyme activity, such as KCN5 and KSJ2, and their potential for industrial applications as enzyme producers.

Effects of intrinsic tannins on proteolysis dynamics, protease activity, and metabolome during sainfoin ensiling.

Condensed tannins (CT) from sainfoin have a high capacity to inhibit proteolysis. A previous study reported that CT from sainfoin can inhibit lactic acid bacteria activity and decrease ammonium-nitrogen (N) content during sainfoin ensiling; however, no study has focused on the metabolome of ensiled sainfoin. The objective of the present study was to investigate the effects of CT [following supplementation of deactivated CT with polyethylene glycol (PEG)] on protease activity, keystone bacteria, and metabolome during sainfoin ensiling. According to the results, PEG amendment increased non-protein N, amino acid, and soluble protein contents significantly (in the 49.08–59.41, 116.01–64.22, and 23.5–41.94% ranges, respectively, p < 0.05) during ensiling, whereas neutral detergent-insoluble protein and acid detergent-insoluble protein were decreased significantly (in the 55.98–64.71 and 36.58–57.55% ranges, respectively, p < 0.05). PEG supplementation increased aminopeptidase and acid protease activity after 3 days of ensiling (p < 0.05) and increased carboxypeptidase activity during the entire ensiling process (p < 0.05). The keystone bacteria changed following PEG addition (Stenotrophomonas, Pantoea, and Cellulosimicrobium in the control vs. Microbacterium, Enterococcus, and Brevundimonas in the PEG-treated group). In total, 510 metabolites were identified after 60 days of sainfoin ensiling, with 33 metabolites annotated in the Kyoto Encyclopedia of Genes and Genomes database. Among the metabolites, phospholipids were the most abundant (72.7% of 33 metabolites). In addition, 10 upregulated and 23 downregulated metabolites were identified in the PEG-treated group when compared with the control group, after 60 days of ensiling (p < 0.05). Pediococcus (correlated with 20 metabolites, R2 > 0.88, p < 0.05) and Lactobacillus (correlated with 16 metabolites, R2 > 0.88, p < 0.05) were the bacteria most correlated with metabolites. The results suggested antagonistic effects between Lactobacillus and Pediococcus during ensiling. The decreased proteolysis during sainfoin ensiling was mainly attributed to the inhibition of protease activity by CT, particularly carboxypeptidase activity. In addition, proteolysis decreased partly due to CT inhibiting Pediococcus activity during ensiling, with Pediococcus being significantly and positively correlated with dopamine after 60 days of ensiling (R2 = 0.8857, p < 0.05).

Isolation, identification and characterization of a novel antimicrobial peptide from Moringa oleifera seeds based on affinity adsorption

This study aimed to characterize a novel antimicrobial peptide (AMP) obtained from Moringa oleifera seed protein hydrolysates. Cell membrane chromatography and live bacteria adsorption were combined into a single step to efficiently isolate the active fraction of the AMP. Five peptides were identified by LC-MS/MS, among which the MCNDCGA peptide (termed MOp3) showed the greatest inhibitory effect against Staphylococcus aureus [minimum inhibitory concentration (MIC): 2 mg/mL]. MOp3 was identified as a hydrophobic anionic AMP rich in β-sheet structures with negligible hemolytic activity at 2.0 × MIC. MOp3 had good tolerance to salt solutions at 5 % and pH range 6.0–8.0, but was sensitive to high temperatures (>100 °C) and acid protease. Microscopic observation further revealed that MOp3 induced irreversible damage onto the cell membrane of S. aureus and interacted with dihydrofolate reductase and DNA gyrase by hydrogen bonding and hydrophobic interaction. These findings highlight the potential application of a new antimicrobial agent against S. aureus in the food industry.

Comparison of physicochemical characteristics and microbiome profiles of low-temperature Daqu with and without adding tartary buckwheat

There is increasing interest in the utilization of tartary buckwheat (TB) for making various fermented foods due to its unique starch structure and functional features. However, studies on Daqu starter produced with TB as a raw material are lacking. This study aimed to compare the differences between conventional low-temperature Daqu (NDQ; 65% barley and 35% pea) and Daqu made with TB (BDQ; 55% barley, 30% pea, and 15% TB) in physicochemical properties, microbial communities, and volatile and nonvolatile metabolites. Compared with NDQ, BDQ exhibited higher saccharifying power, liquefying power, acid protease activity, and lower pH value. High-throughput sequencing revealed that the microbial community structure of NDQ and BDQ had significant differences. NDQ were enriched in Kroppenstedtia, Bacillus, Saccharomycopsis, Issatchenkia, Cladosporium, and Aspergillus, while BDQ were dominated by Kroppenstedtia, Comamonas, Pseudomonas, Acinetobacter, Rhizopus, and Saccharomycopsis. There were notable distinctions in volatile and nonvolatile metabolomic profiles between NDQ and BDQ. The physicochemical differences between NDQ and BDQ were driven by their different microbiota, which also led to the inconsistency of their metabolite profiles. The bacterial community was significantly correlated with pH and acid protease activity, whereas the fungal community was only strongly correlated with saccharifying power. In addition, the correlation network demonstrated that the resident dominant microbes in Daqu had strong connections to numerous important volatile and nonvolatile compounds. This study provided a detailed insight into the microbiome characteristics that differ between the NDQ and BDQ, and it can be helpful to develop a TB-based starter applied in making traditional fermented foods.

Morphology and Physiology Response to Stratification during Seeds Epicotyl Dormancy Breaking of Paeonia emodi Wall. ex Royle

Paeonia emodi is a type of wild herbaceous peony with high ornamental and breeding value. Cold stratification is the only method to break its seed epicotyl dormancy to date, however, the key physiological factors during this process are not clear. In this study, rooted seeds of P. emodi were treated with 4 °C stratification, and the changes of the embryo, four nutrients, and two endogenous hormones in the seeds were investigated. The results showed that the plumule elongated at S6 (i.e., ten weeks of cold stratification), and grew to nearly the same length as the cotyledon at S9. Cold stratification increased starch consumption, significantly decreased soluble sugar content in the later stages, and decreased soluble protein content at S9, but it did not influence crude fat content. The activities of α-amylase and β-amylase increased significantly at S4 and S4 to S6, respectively. At S8 and S9, acid protease activity increased, and the increase in lipase activity continued throughout the whole process. At the same time, the ABA content decreased significantly after S6; from this stage, the ABA/GA3 ratio gradually decreased compared with that of the control, and the difference was significant at S9. Correlation analysis showed that the ABA/GA3 ratio was significantly correlated with starch content and α-amylase activity. It can be concluded that both carbohydrates and proteins were the energy supply for the epicotyl dormancy breaking of P. emodi seeds, rather than crude fat. Cold stratification promoted substance transformation by increasing the corresponding enzyme activities. The balance of ABA and GA3 suggested the key stage for the release of dormancy.

Effect of the surface charge of the acid protease on leather bating performance

Green and efficient acid protease has received increasing attention in bating wet blue (chrome tanned leather). However, acid proteases penetrate slowly in the wet blue, which reduces the dispersion efficiency and uniformity of hide collagen fibers. We explored the factors that affect the penetration of acid proteases in wet blue and the bating performance, including surface charge, molecular weight, particle size, and proteolytic activity of protease. The results show that the most important factor is the surface charge of the acid protease. The acid protease AP-2 derived from Trichoderma reesei had isoelectric point (pI) of 3.6 and penetrated the wet blue more deeply, thereby dispersing collagen fibers and softening the wet blue better than the pepsin (pI 1.9) and the acid protease AP-1 derived from Aspergillus usamii (pI 2.4). This finding is because the negatively charged pepsin and AP-1 are combined easily with the positively charged wet blue at the bating pH of 3.5 and stay on the surface layer rather than transfer into the inner layer of the wet blue. Therefore, choosing acid protease with the pI higher than the bating pH is important for deep penetration and good bating performance.

Marine sources for biotechnology: Preliminary digestive enzymes assessment of seven decapod species from the southwest Atlantic

Marine natural compounds are required by industries to improve and reduce costs of many biotechnological processes. In this work, specific activity of midgut gland digestive enzymes was measured in seven species of decapods. Alkaline and acid peptidase, lipase and amylase activities were registered in Pleoticus muelleri (Pm), Artemesia longinaris (Al), Paralomis granulosa (Pg), Lithodes santolla (Ls), Munida gregaria (Mg), Neohelice granulata (Ng), and Ovalipes trimaculatus (Ot). The highest activities for all the analyzed digestive enzymes were obtained in Pg, Ls, Mg and Ot, which were used for optimum temperature and pH measurements and to test their stability at different temperatures and pH. In general, optimal enzyme temperatures were between 30 and 50 °C in the four crustaceans. Peptidases and lipases from these species showed in general alkaline optimal pH except for those from Ot which presented high activity at the whole pH range (2–12). Amylase from Ot presented its optimal at pH 6–8 and 30–50 °C. Thermal stability of peptidases was elevated at 30 °C for the four species tested (Pg; Ls; Mg and Ot), lipases were stable mainly at 30–50 °C and amylase was stable at 10 °C, 30 °C and 50 °C during the first 30 min of the assay. In general, stability of the four enzymes from Pg was greater at alkaline pH than at acid. The other species showed stability at all pH for the four enzymes. The range of temperature and pH of the tested enzymes activity are coincident with those used in various biotechnological processes and therefore indicates the potential of these enzymes to be used for industrial procedures. Further, most of the species studied are converted into fishery residues, either by processing or by-catch, so their reuse would also contribute to the sustainability of the activity.

Comparison and application of the extraction method for the determination of enzymatic profiles in matured soybean koji

In this study, two methods of extracting crude enzymes from soy sauce koji were compared to investigate the effect of replacing the buffer extraction with distilled water extraction by measuring the enzyme activity. Results showed the enzyme activities of the two extracts in each koji sample were not significant differences. The stability of the different enzymes in the extracts varied. Acid protease, β-glucosidase and cellulase activities showed the best stability during the storage. Neutral protease, glucoamylase and aminopeptidase were moderately stable and their activities needed to be determined within 3 d of extract preparation, while xylanase activity was the least stable, decreasing by 40% on the second day of storage. Therefore, the recommended procedure for the analysis of koji enzyme profiles was as follows, xylanase was determined immediately after distilled water extraction, and then all other enzyme activity analyses need to be completed within 3 d to ensure the validity of enzyme analysis. This method was applied to the enzymatic analysis of 7 kojis made by different strains of Aspergillus oryzae, and to explore the association between the enzyme profile and the taste characteristics of seven fermented soy sauce. The results showed proteases were correlated with sour and richness, while carbohydrate-degrading enzymes were relatively more relevant to umami taste.

Interactions between biofiller-modified polymeric composites and wood-rotting fungi in terms of their biotechnological applications

The synthesis and characterization of polymeric cross-linked composites derived from bisphenol A glycerolate diacrylate (BPA.GDA) and N-vinyl-2–2pyrrolidone (NVP) are presented. Commercially available kraft lignin with different weight % (0, 5, and 20) was applied as an environmentally friendly filler. The composites were made by bulk polymerization using a photoinitiator (2-dimethoxy-2-phenylacetophenone) in special glass molds and cut into 10 mm × 10 mm fragments for testing. The chemical structures of the composites were confirmed by means of ATR-FTIR spectroscopy. Polymers modified with the addition of lignin were introduced into cultures of three species of white rot fungi: Cerrena unicolor, Pycnoporus sanquineus, and Abortiporus biennis. The biochemical and profilometric analyses showed clear changes both in the course of selected metabolic processes in the fungal cells and in the structure of the biomaterials. Clear changes in the activity of such enzymes as laccase, β-glucosidase, or acid proteases caused by the selected polymers were observed in the culture fluids.