This study was carried out to identify the ACE (Angiotensin converting enzyme) inhibitory activity of casein hydrolysates for development of anti-hypertensive hydrolysates. Sodium caseinate was treated with six kinds of commercial proteases such as Flavourzyme, Protamex, Neutrase 1.5, Alcalase, Protease M, and Protease S for 8 h individually, and was then treated with the enzyme combination for 4 h at $45^{\circ}C$. The hydrolysate which had the highest ACE inhibitory effect was then hydrolysed successively with three digestive enzymes: pepsin, trypsin, and ${\alpha}$-chymotrypsin, at $37^{\circ}C$ for 4 h under conditions mimicking those of the gastrointestinal tract. UF (ultra filtration) treatment was applied to one of the secondary hydrolysates to determine ACE inhibitory activity. When sodium caseinate was hydrolysed by commercial proteases, the degree of hydrolysis (DH) showed 2.54 to 4.25% and after secondary hydrolysis, DH showed 4.30 to 5.22%. ACE inhibitory activity and $IC_{50}$ values decreased, and inhibition rates increased during hydrolysis. Protamex treatment showed the lowest $IC_{50}$ value ($516{\mu}g/mL$) and Flavourzyme hydrolysate showed the highest $IC_{50}$value ($866{\mu}g/mL$). As the first hydrolysate was treated with Flavourzyme, the ACE inhibitory activity increased. Neutrase hydrolysate had the highest activity with an $IC_{50}$ value ($282{\mu}g/mL$). When Neutrase plus Flavourzyme treatment was hydrolyzed by digestive enzymes, the $IC_{50}$ value ($597{\mu}g/mL$) was decreased statistically (p<0.05). As Neutrase plus Flavourzyme hydrolysate is treated by UF with MW cut-off 10,000, permeate showed $273{\mu}g/mL$ of $IC_{50}$ value, showed no difference, but retentate which has over MW 10,000 showed statistically different $IC_{50}$ value, $635{\mu}g/mL$ (p<0.05).
Cellulase is the key enzyme in the use of cellulose-based biomaterials. Because of its structure, cellulose is difficult to be degraded by enzymes. In order to utilize cellulose-based biomaterials efficiently, evolutionary wisdom of how to use enzymes accurately and harmoniously in a biological system is needed, such as the cellulose digestive system in animals. In this study, the symbiotic bacteria from snails, Achatina fulica, were identified and their cellulase activity was evaluated. The 16S rRNA sequence analysis of 100 aerobic bacteria showed that they belonged to 9 genus and almost half of the bacteria were Lactococcus spp. Among 100 identified strains, only two Aeromonas sp. strains showed cellulase activity. Aeromonas sp. KMBS020 had both endo-${\beta}$-glucanase and ${\beta}$-glucosidase activities but Aeromonas sp. KMBS018 had ${\beta}$-glucosidase activity only. None of the 100 bacterial colonies had any cellobiohydrolase activity.
Geun-Hui Nam;Young-Bin Lee;Sea-Hwan Sohn;In-Surk Jang
Korean Journal of Poultry Science
/
v.51
no.2
/
pp.107-116
/
2024
The study was conducted to examine age-related development of digestive organs, intestinal microvilli hydrolase, and hepatic antioxidant enzyme in Korean native chicks (KNC) aged from 0-d to 28-d of post-hatching. Body weight did not significantly increase from 0-d to 3-d-old, but after that remarkably increased from 3-d to 28-d-old (P<0.05). The relative weight (g/100 g of BW) of the proventriculus, gizzard, and liver was significantly higher at 3- and 7-d-old chicks than that of the other ages. The relative weight of the intestine, mucosal tissues, and pancreas was markedly developed at the ages of 3-, 14-, and (or) 21-d-old chicks (P<0.05). In the small intestine, the specific activities of maltase and sucrose were significantly higher at 14-d-old compared with the other ages (P<0.05). Leucine aminopeptidase activity showed a constant level from 0- to 28-d-old without significance. The specific activity of alkaline phosphatase was significantly higher at 0-d-old compared with the other ages (P<0.05). In the liver, the specific activities of superoxide dismutase, glutathione peroxidase, and glutathione S-transferase were shown to be lowest at 0-d-old, but they continued to increase as the age increased. The lipid peroxidation was significantly high at the age of 21-d (P<0.05), after that its level decreased at 28-d old. In conclusion, the KNC rapidly developed digestive organs and intestinal microvilli hydrolase activity from 3- to 14-d-old after hatching. Hepatic antioxidant enzyme activity continued to increase as the age increased after hatching, resulting in 28-d-old chicks showing the highest antioxidant enzyme activity in the KNC.
Black rockfish larvae and juveniles were reared for 95 days after parturition (DAP) in order to determine four enzyme activities (trypsin, pepsine-like enzyme, lipase, amylase) during ontogeny. Larvae were fed rotifers Brachionus plicatilis from 1 to 25 DAP, Artemia nauplii from 10 to 78 DAP and then gradually changed to pelleted feed from 30 DAP. Temperature was kept between $13.5{\sim}14.9^{\circ}C$. Trypsin and lipase activities were found in 2 DAP larvae ($7.0{\pm}1.5$ unit and $4.5{\pm}1.4$ unit, $mean{\pm}SD$, respectively). The evolution of both enzymes activities showed a profile marked by drastic increases between postflexion and juvenile stage. There is an increment on specific trypsin activity at 10 DAP, corresponding with the beginning of Artemia feeding. Pepsin-like enzyme activity was found at 11 DAP and increased drastically from 56 DAP, cor-responding with the initiation of juvenile stage. Amylase activity was also found at 11 DAP and maintained at a low level up to 38 DAP followed by a drastic increase from 39 DAP to 50 DAP. Considering our results of both trypsin and pepsin-like enzyme activities, it might be concluded that higher somatic growth of Sebastes inermis could be possible with the initiation juvenile of stage and the early juvenile stage is a suit-able period for feeding an artificial diet for fish.
A high protease-producing strain was isolated and identified from the digestive tract of octopus vulgaris by detecting a hydrolysis circle of protease and its activity. The strain was identified by morphology observation, biochemical experiments, and 16S rRNA sequence analysis. The protease obtained from the strain was purified by a three-step process involving ammonium sulfate precipitation, carboxy methyl-cellulose (CM-52) cation-exchange chromatography, and DEAE-Sephadex A50 anion-exchange chromatography. The properties of protease were characterized as well. The strain Bacillus sp. QDV-3, which produced the highest activity of protease, was isolated. On the basis of the phenotypic and biochemical characterization and 16S rRNA gene-sequencing studies, the isolate was identified as follows: domain: Bacteria; phylum: Firmicutes; class: Bacilli; order: Bacillales; family: Bacillaceae; and genus: Bacillus. The isolate was shown to have a 99.2% similarity with Bacillus flexus. A high active protease designated as QDV-E, with a molecular weight of 61.6 kDa, was obtained. The enzyme was found to be active in the pH range of 9.0-9.5 and its optimum temperature was $40^{\circ}C$. The protease activity retained more than 96% at the temperature of $50^{\circ}C$ for 60 min. Phenylmethylsulfonyl fluoride (PMSF) inhibited the enzyme activity, thus confirming that this protease isolated from Bacillus sp. QDV-3 is an alkaline serine protease. Metal ions, $Mn^{2+}$ and $Mg^{2+}$, were determined to enhance the protease activity, whereas $Ba^{2+}$, $Zn^{2+}$, and $Cu^{2+}$ were found to inactivate the enzyme.
Among different types of spoilage, microbial contamination can cause feed decomposition, which results in decreases in feed intake and productivity, infection, and breeding disorder. During the storage time, various microbes have a chance to inoculate with depreciation of feed and to infect the animals. We investigated bacteria that inhabit diverse feed ingredients and complete feed which have been stored for a few months. We isolated and identified 30 genera and 62 species of bacteria. Among these 62 species, 21 species were of non-pathogenic bacteria, 18 species were of pathogenic bacteria, 9 species were of opportunistic pathogens, and 14 species were of unknown bacteria. Pantoea allii and 24 species showed proteolytic enzyme activity. We also confirmed that 6 species including Pseudomonas psychrotolerans showed ${\alpha}$-amylase activity, and 29 species including Burkholderia vietnamiensis showed cellulase activity. Microbacterium testaceum and 3 species showed resistance to Ampicillin, Kanamycin, Streptomycin, Gentamicin, Carbenicillin, and Erythromycin ($50{\mu}g/mL$). Using mealworm larvae (Tenebrio molitor L.) as a model for pathogenicity, we confirmed that 8 species including Staphylococcus xylosus had pathogenicity for mealworm larvae. Especially, Enterobacter hormaechei, Staphylococcus xylosus, and Staphylococcus hominis were reported as being pathogenic for humans. This research suggests that hygienic management of animal feed is essential because beneficial and harmful bacteria can inhabit animal feed differently during storage and distribution.
Currently many studies aimed at enhancing efficacy of medicinal food on biological activity using bioconversion technology including fermentation process. In this study, the quality characteristics and antioxidative activity of fermented Crataegi fructus was investigated. The antioxidant activity of fermented Crataegi Fructus was assessed by various radical scavenging assays using DPPH (2,2-Diphenyl-1-picrylhydrazyl), FRAP (Ferric ion reducing antioxidant power), Reducing power and ABTS (2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid)). Moisture content of fermented Crataegi Fructus was $39.3{\pm}0.06%$. Contents of crude ash, crude protein, and crude fat were $0.20{\pm}0.01$, $1.77{\pm}0.04$, and $1.40{\pm}0.59%$, respectively. Moreover, the hunter's color values of fermented Crataegi Fructus were 79.24 (lightnees), 1.58 (redness), and 31.25 (yellowness), respectively. Total phenolic contents of fermented Crataegi Fructus were $3,015{\pm}250$ GAE ${\mu}g/g$. The antioxidative activities of fermented Crataegi Fructus significantly increased in a dose dependent manner. In addition, fermented Crataegi Fructus slightly (10.4%) inhibited ${\alpha}$-glucosidase activity; however, there was no inhibitory activity against ${\alpha}$-amylase. In terms of proteolytic activity, fermented Crataegi Fructus showed a strong activity than pancreatin (used as a positive control). These results indicate that fermented Crataegi Fructus can be used as a natural resource for material aiding digestion.
To investigate the efficacy of alternatives to antibiotics, the present study was conducted to compare the effects of antibiotic, lactic acid, a blend of commercial essential oils (EOs) and EOs in combination with lactic acid on growth performance and the functional activity of the gut in broiler chickens. A total of 168 broiler chickens were given the basal diet supplemented with 10 ppm colistin (T1), 0.1% lactic acid (T2), 25 ppm EOs (T3), 25 ppm EOs+0.1% lactic acid (T4), 50 ppm EOs (T5) or 50 ppm EOs+0.1% lactic acid (T6) in the period 3 to 35 days of age. As a result, the broiler chickens assigned to T4 group throughout the experimental period had apparently (p<0.05) greater body weight and total gain than these assigned to T1, T2, T3 and T5 groups. However, there was no difference in growth performance among the birds fed the diets supplemented with antibiotic (T1), lactic acid (T2) and EOs (T3 and T5) alone. The weights of digestive organs and the number of lactobacilli and E. coli in the lower ileum were not affected by dietary treatments. Total trypsin activity was significantly (p<0.05) greater in T4 than T1, T2, T3 and T5 groups. Total and specific pancreatic $\alpha$-amylase activities were significantly (p<0.05) enhanced in the broiler chickens fed T4 diet compared with these fed T1, T2 and T3 diets. However, there were no differences in growth performance and digestive enzyme activities including pancreatic trypsin and $\alpha$-amylase between T4 and T6 groups fed the diets supplemented with either low or high EOs levels in combination of lactic acid. In conclusion, a blend of commercial EOs combined with lactic acid showed significant increases in digestive enzyme activities of the pancreas and intestinal mucosa, leading to increase in growth performance.
This study was devised to elucidate whether soused shrimp exhibits a digestive action on boiled pork meats. and the mechanism by which sousing with a high concentration of sodium chloride preserves nutrients in foods for a prolonged pe\ulcornerriod. Protease was isolated from soused shrimp using a combination of ammonium sulfate fractionation. DEAE - cellulose ion exchange chromatography and gel filtra\ulcornertion. The isolated protease had specific activity of 1.560 units. 210 purification fo\ulcornerld with an yield of 38%. Its optimum pH and temperature were 8.0 and $43^{\circ}C$ respectively. The molecular weight of the enzyme was 35.000. The Km value of the enzyme for casein was 1.6 x $10^{-6}$ M The e=yme required the presence of cu\ulcornerpric ion to exhibit its full activity. Eighty eight percent of the enzyme activity was in\ulcornerhibited by 3.5M NaCI showing a reversibly linear decrease of the enzyme activity as NaCI concentration increased. The nature of the inhibition by NaCl was rever\ulcornersible and noncompetitive. The protease activity in soused shrimp was well preser\ulcornerved with the elapse of time at least in part due to NaCI induced suppression of autodigestion. The enzyme was denatured by acid easily. i.e. 1% of the original activity remained after staying at pH 2 for 10 minutes. which is within the norm\ulcorneral range of pH of the human stomach. Soused shrimp was observed to be one of those containing the highest protease activity compared with the other soused foo\ulcornerds such as soused oyster. squid. clam. and Pollack intestine with respect to spec\ulcornerific activities of dialized 1:4 whole homogenates(w/v) in 5 mM sodium phospha\ulcornerte - 2.4 mM j3 - mercaptoethanol buffer. pH 8.0. Casein and boiled meats including pork, beef, and chicken appeared to be the good substrates for the protease. Casein was the best. Therefore. the ingestion of boiled meats including pork together with soused sh\ulcornerrimp would help digestion of boiled pork in human not only by increasing appe\ulcornertite also by the direct proteolytic digestion of boiled meats by soused shrimp to\ulcorner some extent. And a high concentration of sodium chloride inhibited the protease activity reversibly in a remarkable degree, which ensued in a significant retardat\ulcornerion of autodigestion of protein in foods by proteases, and hereby contributed to the preservation of foods for an extended period.
Objective: This study was conducted to evaluate the effect of dietary energy and lipase supplementation on growth performance, nutrient digestibility, serum profiles, intestinal morphology, small intestinal digestive enzyme activities, biochemical index of intestinal development and noxious gas emission in weaning pigs. Methods: A total of 240 weaning pigs ([Yorkshire${\times}$Landrace]${\times}$Duroc) with an average body weight (BW) of $7.3{\pm}0.12kg$ were used in this 28-d experiment. Weaning pigs were randomly allocated to 4 dietary treatments in a $2{\times}2$ factorial arrangement with 2 levels of energy (net energy = 2,470 kcal/kg for low energy diet and 2,545 kcal/kg for basal diet) and 2 levels of lipase (0 and 1.5 U/g of lipase) according to BW and sex. There were 6 replications (pens) per treatment and 10 pigs per pen (5 barrows and 5 gilts). Results: Weaning pigs fed the low energy diet had lower (p<0.05) gain-to-feed ratio (G:F) throughout the experiment, apparent digestibility of dry matter, nitrogen, ether extract, and gross energy during d 0 to 14, average daily gain during d 15 to 28, lipase activity in duodenum and ileum and protein/DNA in jejunum (p<0.05), respectively. Lipase supplementation had no effect on growth performance but affected apparent nutrient digestibility (p<0.05) on d 14 and enhanced lipase activity in the duodenum and ileum and protease activity in duodenum and jejunum of pigs (p<0.05) fed the low energy diet. Lipase reduced serum low-density lipoprotein cholesterol (LDL-C) and triglyceride (TG), $NH_3$ production (p<0.05) from the feces. Conclusion: The low energy diet decreased G:F throughout the experiment and nutrient digestibility during d 0 to 14 as well as lipase activity in duodenum and ileum. Lipase supplementation increased nutrient digestibility during d 0 to 14 and exerted beneficial effects on lipase activity in duodenum and ileum as well as protease activity in duodenum and jejunum, while reduced serum LDL-C, TG and fecal $NH_3$.
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