• Korean Journal of Food Science and Technology
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• v.29 no.6
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• pp.1125-1130
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• 1997

The effects of enzyme combination, pH, acid washing and enzyme treatment sequence were investigated in the hydrolysis of soy protein. Comparing Alcalase vs. Neutrase/Alcalase, it appeared that Neutrase/Alcalase was more efficient than Alcalase alone, as the highest degree of hydrolysis (DH) was seen in Neutrase/Alcalase. A surprisingly high DH (more than 60%) was observed with Flavourzyme in the second hydrolysis. The separation of insolubles from the first hydrolysis had little effect on the second hydrolysis. When the washing water from the first hydrolysis was reused in the next hydrolysis, the DH and protein recovery were increased. The addition of calcium ion showed not so much positive effects by the stabilization of Neutrase on the Protein hydrolysis. The use of carbohydrase and repeated acid washing gave positive effects on DH. The simultaneous treatment using endoprotease and exoprotease with pH adjustment improved DH significantly.

• Parasites, Hosts and Diseases
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• v.49 no.3
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• pp.245-254
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• 2011

Many immune down-regulatory molecules have been isolated from parasites, including cystatin (cystain protease inhibitor). In a previous study, we isolated and characterized Type I cystatin (CsStefin-1) of the liver fluke, Clonorchis sinensis. To investigate whether the CsStefin-1 might be a new host immune modulator, we induced intestinal inflammation in mice by dextran sodium sulfate (DSS) and treated them with recombinant CsStefin-1 (rCsStefin-1). The disease activity index (DAI) increased in DSS only-treated mice. In contrast, the DAI value was significantly reduced in rCsStefin-1-treated mice than DSS only-treated mice. In addition, the colon length of DSS only-treated mice was shorter than that of rCsStefin-1 treated mice. The secretion levels of IFN-${\gamma}$ and TNF-${\alpha}$ in the spleen and mesenteric lymph nodes (MLNs) were significantly increased by DSS treatment, but the level of TNF-${\alpha}$ in MLNs was significantly decreased by rCsStefin-1 treatment. IL-10 production in both spleen and MLNs was significantly increased, and IL-$10^+F4/80^+$ macrophage cells were significantly increased in the spleen and MLNs of rCsStefin-1 treated mice after DSS treatment. In conclusion, rCsStefin-1 could reduce the intestinal inflammation occurring after DSS treatment, these effects might be related with recruitment of IL-10 secreting macrophages.

• Korean Journal of Food Science and Technology
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• v.30 no.4
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• pp.934-940
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• 1998

Kochujang, fermented hot pepper soybean paste, was prepared by traditional method, and irradiated with gamma ray of $^{60}Co{\;}up{\;}to{\;}15{\pm}1.5{\;}kGy$ for confirming main factor of kochujang fermentation. Nonirradiated (control) and irradiated samples kochujang were fermented at $25^{\circ}C$ and analyzed biochemical properties including enzyme activities and viable cell count during fermentation for 60 days. The total viable count in irradiated kochujang decreased to $10^4{\;}CFU/g$ which was $10^8{\;}CFU/g$ in the control. Because of a little changing enzyme activities of ${\alpha}{\cdot}{\beta}{\;}amylase$ and acid neutral protease by irradiation at the above level, amino type nitrogen which is the main quality reference of kochujang was comparable to the control. By irradiation, gas production was completely stopped which is one of biggest problems during distribution of kochujang. Total volume of gas produced at $25^{\circ}C$ from the control kochujang was 453 mL/100 g which was composed of over 90% of $CO_2$. The odor of irradiated kochujang was inferior to the control which seemed to be related to reduced microbial populations.

• Archives of Pharmacal Research
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• v.25 no.4
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• pp.505-510
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• 2002

An improved kinetic assay for prekallikrein activator (PKA), a potential vasodilator, has been developed to be used as an indicator for quality control during production of human albumin preparations. It consists of two reaction stages. In the first stage, PKA and prekallikrein are incubated at $37^{\circ}C$ for 45 min to allow the transformation into kallikrein. Kallikrein, a serine protease, catalyzes the splitting of p-nitroaniline (pNA) from its substrate H-D-Pro-Phe-Arg-pNA(S-2302). The rate at which pNA is released was measured spectrophotometrically at 405 nm. Prekallikrein, a substrate of PKA was purified by DEAE ion-exchange chromatography and the major potential variations in the assay were optimized; pH 8.0 and 150 mM sodium chloride were chosen to give a proper ionic strength. Reaction times in the range of 10 to 360 min provided linear dose-response curves. The concentration of prekallikrein was adjusted to fall between 1:1 and 1:3 dilutions to generate a linear standard calibration curve. Under the optimized conditions, reproducibility was checked. In a precision test, the coefficient of variation (CV) stayed within ${\pm}4%$ and the dose-response curve showed a good correlation (${r^2}=0.999$). An accuracy test with an international standard of PKA afforded a mean recovery of 97.5%.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.12
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• pp.1844-1853
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• 2019

Objective: We investigated how pituitary adenylate cyclase-activating polypeptide (PACAP) affects embryonic development during pre-in vitro maturation (pre-IVM) using porcine oocytes isolated from small follicles. Methods: We divided the follicles into the experimental groups by size (SF, small follicles; MF, medium follicles) and treated with and without PACAP and cultured for 18 hours (PreSF[-]PACAP; without PACAP, Pre-SF[+]PACAP; with PACAP) before undergoing IVM. The gene expression related to extracellular matrix formation (amphiregulin, epiregulin, and hyaluronan synthase 2 [HAS2]) and apoptosis (Bcl-2-associated X [BAX], B-cell lymphoma 2, and cysteine-aspartic acid protease 3) was investigated after maturation. The impact on developmental competence was assessed by the cleavage and blastocyst rate and total cell number of blastocysts in embryos generated from parthenogenesis (PA) and in vitro fertilization (IVF). Results: Cleavage rates in the Pre-SF(+)PACAP after PA were significantly higher than SF and Pre-SF(-)PACAP (p<0.05). The cleavage rates between MF and Pre- SF(+)PACAP groups yielded no notable differences after IVF. Pre-SF(+)PACAP displayed the higher rate of blastocyst formation and greater total cell number than SF and Pre-SF(-)PACAP (p<0.05). Cumulus cells showed significant upregulation of HAS2 mRNA in the Pre-SF(+)PACAP compared to the SF (p<0.05). In comparison to other groups, the Pre-SF(+)PACAP group displayed a downregulation in mRNA expression of BAX in matured oocytes (p<0.05). Conclusion: The PACAP treatment during pre-IVM improved the developmental potential of porcine oocytes derived from SF by regulating cumulus expansion and apoptosis of oocytes.

• The Plant Pathology Journal
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• v.35 no.5
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• pp.425-436
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• 2019

Botrytis cinerea, a major phytopathogenic fungus, has been reported to infect more than 200 crop species worldwide, and it causes massive losses in yield. The aim of this study was to evaluate the inhibitory abilities and effects of Bacillus amyloliquefaciens RS-25, Bacillus licheniformis MG-4, Bacillus subtilis Z-14, and Bacillus subtilis Pnf-4 and their culture filtrates and extracts against the gray mold caused by B. cinerea on postharvest tomato, strawberry, and grapefruit. The results revealed that the cells of Z-14, culture filtrate of RS-25, and cells of Z-14 showed the strongest biocontrol activity against the gray mold on the strawberry, grape, and tomato fruit, respectively. All the strains produced volatile organic compounds (VOCs), and the VOCs of Pnf-4 displayed the highest inhibition values. Based on headspace solid-phase microextraction in combination with gas chromatography-mass spectrometry, esters accounted for the largest percentage of the VOCs produced by RS-25, MG-4, Z-14, and Pnf-4 (36.80%, 29.58%, 30.78%, and 36.26%, respectively). All the strains showed potent cellulase and protease activities, but no chitinase activity. RS-25, Z-14, and MG-4, but not Pnf-4, grew on chrome azurol S agar, and an orange halo was formed around the colonies. All the strains showed biofilm formation, fruit colonization, and lipopeptide production, which may be the main modes of action of the antagonists against B. cinerea on the fruit. This study provides the basis for developing natural biocontrol agents against the gray mold caused by B. cinerea on postharvest fruit.

• Animal Bioscience
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• v.34 no.7
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• pp.1157-1168
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• 2021

Objective: The aim of this study was to investigate the effects of different amounts of wheat bran (WB) inclusion and postbiotics form by Saccharomyces cerevisiae and phytase co-fermented wheat bran (FWB) on the growth performance and health status of broilers. Methods: Study randomly allocated a total of 300 male broilers to a control and 4 treatment groups (5% WB, 5% FWB, 10% WB, and 10% FWB inclusion, respectively) with each pen having 20 broilers and 3 pens per treatment. Results: The WB does not contain enzymes, but there are 152.8, 549.2, 289.5, and 147.1 U/g dry matter xylanase, protease, cellulase and β-glucanase in FWB, respectively. Furthermore, FWB can decrease nitric oxide release of lipopolysaccharide stimulated chicken peripheral blood mononuclear cells by about two times. Results show that 10% FWB inclusion had significantly the highest weight gain (WG) at 1 to 21 d; 5% FWB had the lowest feed conversion rate at 22 to 35 d; 10% WB and 10% FWB inclusion have the highest villus height and Lactobacillus spp. number in caecum; and both 5% and 10% FWB can increase ash content in femurs. Compared to control group, all treatments increase mucin 2, and tight junction (TJ), such as occludin, claudin-1, zonula occludens-1, and mRNA expression in ileum by at least 5 folds. In chicken peripheral blood mononuclear cells, nicotinamide adenine dinucleotide phosphate-oxidase-1 mRNA expression decreases from 2 to 5 times, and glutamate-cysteine ligase catalytic subunit mRNA expression also increases in all treatment groups compared to control group. The mRNA expression of pro-inflammatory cytokines, including interleukin-6 (IL-6), nuclear factor-κB, and IL-1β, decreases in 5% and 10% FWB groups compared to control group. Conclusion: To summarize, both WB and FWB inclusion in broilers diets increase TJ mRNA expression and anti-oxidation and anti-inflammation, but up to 10% FWB groups have better WG in different stages of broiler development.

• Journal of Microbiology and Biotechnology
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• v.31 no.11
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• pp.1591-1600
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• 2021

Streptomyces coelicolor is a filamentous soil bacterium producing several kinds of antibiotics. S. coelicolor abs8752 is an abs (antibiotic synthesis deficient)-type mutation at the absR locus; it is characterized by an incapacity to produce any of the four antibiotics synthesized by its parental strain J1501. A chromosomal DNA fragment from S. coelicolor J1501, capable of complementing the abs- phenotype of the abs8752 mutant, was cloned and analyzed. DNA sequencing revealed that two complete ORFs (SCO6992 and SCO6993) were present in opposite directions in the clone. Introduction of SCO6992 in the mutant strain resulted in a remarkable increase in the production of two pigmented antibiotics, actinorhodin and undecylprodigiosin, in S. coelicolor J1501 and abs8752. However, introduction of SCO6993 did not show any significant difference compared to the control, suggesting that SCO6992 is primarily involved in stimulating the biosynthesis of antibiotics in S. coelicolor. In silico analysis of SCO6992 (359 aa, 39.5 kDa) revealed that sequences homologous to SCO6992 were all annotated as hypothetical proteins. Although a metalloprotease domain with a conserved metal-binding motif was found in SCO6992, the recombinant rSCO6992 did not show any protease activity. Instead, it showed very strong β-glucuronidase activity in an API ZYM assay and toward two artificial substrates, p-nitrophenyl-β-D-glucuronide and AS-BI-β-D-glucuronide. The binding between rSCO6992 and Zn²⁺ was confirmed by circular dichroism spectroscopy. We report for the first time that SCO6992 is a novel protein with β-glucuronidase activity, that has a distinct primary structure and physiological role from those of previously reported β-glucuronidases.

• Animal Bioscience
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• v.35 no.7
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• pp.1030-1038
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• 2022

Objective: Two experiments were conducted to evaluate the effect of enzyme complex (EC) on the standardized ileal digestibility (SID) of amino acids (AA) in corn gluten meal (60%) (CGM), soy protein concentrate (SPC), dried bovine plasma (DBP), and poultry offal meal (POM). Experiments I and II were conducted with broilers in the pre-starter (1 to 7 days of age) and starter (1 to 21 days of age) phases, respectively. Methods: The treatments consisted of a protein-free diet (PFD) containing feedstuffs either supplemented with EC (xylanase, amylase, and protease) or not. In Experiment I, a total of 360 one-day-old male Cobb-500 broiler chicks were randomly housed in 45 pens, resulting in five replicates with eight birds each, totalizing eight treatments and one PFD group. In Experiment II a total of 270 one-day-old male Cobb-500 broiler chicks were randomly housed in 45 pens, resulting in five replicates with six birds each, totalizing eight treatments and one PFD group. The PFD groups were used to assess the endogenous AA losses. The birds were slaughtered to collect the ileal content. Results: In the pre-starter phase, the SID of arginine, branched chain-aminoacids, glycine, serine, aspartate, and glutamic acid increased with EC addition. The EC improved the SID of arginine and glutamic acid of CGM; the SID of valine and cystine of SPC; the SID of leucine, glycine, and aspartate of POM and the SID of isoleucine of DBP. In the starter phase, the SID of isoleucine, phenylalanine and glycine increased in EC-supplemented diets. The EC improved the SID of isoleucine of DBP; the SID of phenylalanine of CGM and POM. The SID of AA of SPC was not influenced by the EC. Conclusion: The addition of an EC to broiler pre-starter and starter diets is efficient in increasing the SID of AA on SPC, POM, and DBP.

• Parasites, Hosts and Diseases
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• v.60 no.2
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• pp.117-126
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• 2022

Cystatin, a cysteine protease inhibitor found in many parasites, plays important roles in immune evasion. This study analyzed the molecular characteristics of a cystatin from Fasciola hepatica (FhCystatin) and expressed recombinant FhCystatin (rFhcystatin) to investigate the immune modulatory effects on lipopolysaccharide-induced proliferation, migration, cytokine secretion, nitric oxide (NO) production, and apoptosis in mouse macrophages. The FhCystatin gene encoded 116 amino acids and contained a conserved cystatin-like domain. rFhCystatin significantly inhibited the activity of cathepsin B. rFhCystatin bound to the surface of mouse RAW264.7 cells, significantly inhibited cell proliferation and promoted apoptosis. Moreover, rFhCystatin inhibited the expression of cellular nitric oxide, interleukin-6, and tumor necrosis factor-α, and promoted the expression of transforming growth factor-β and interleukin-10. These results showed that FhCystatin played an important role in regulating the activity of mouse macrophages. Our findings provide new insights into mechanisms underlying the immune evasion and contribute to the exploration of potential targets for the development of new drug to control F. hepatica infection.