• Asian-Australasian Journal of Animal Sciences
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• v.30 no.11
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• pp.1612-1619
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• 2017

Objective: The objective of this study was to optimize ultrasonic-assisted enzymatic hydrolysis conditions, including enzyme-to-substrate (E/S) ratio, pH, and temperature, for producing porcine liver hydrolysates (PLHs) with the highest 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging activity by using response surface methodology (RSM). Methods: The study used RSM to determine the combination of hydrolysis parameters that maximized the antioxidant activity of our PLHs. Temperature ($40^{\circ}C$, $54^{\circ}C$, and $68^{\circ}C$), pH (8.5, 9.5, and 10.5), and E/S ratio (0.1%, 2.1%, and 4.1%) were selected as the independent variables and analyzed according to the preliminary experiment results, whereas DPPH free radical scavenging activity was selected as the dependent variable. Results: Analysis of variance showed that E/S ratio, pH, and temperature significantly affected the hydrolysis process (p<0.01). The optimal conditions for producing PLHs with the highest scavenging activity were as follows: E/S ratio, 1.4% (v/w); temperature, $55.5^{\circ}C$; and initial pH, 10.15. Under these conditions, the degree of hydrolysis, DPPH free radical scavenging activity, ferrous ion chelating ability, and reducing power of PLHs were 24.12%, 79%, 98.18%, and 0.601 absorbance unit, respectively. The molecular weight of most PLHs produced under these optimal conditions was less than 5,400 Da and contained 45.7% hydrophobic amino acids. Conclusion: Ultrasonic-assisted enzymatic hydrolysis can be applied to obtain favorable antioxidant hydrolysates from porcine liver with potential applications in food products for preventing lipid oxidation.

• Asian-Australasian Journal of Animal Sciences
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• v.14 no.5
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• pp.720-732
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• 2001

Skeletal muscle is of major economic importance since it is finally converted to meat for consumers. The increase in meat production with low costs of production may be achieved by optimizing muscle growth, whereas a high meat quality requires, among other factors, the optimization of intramuscular glycogen and fat stores. Thus, research in energy metabolism aims at controling muscle metabolism, but also liver and adipose tissue metabolism in order to optimize energy partitioning in favour of muscles. Liver is characterized by high anabolic and catabolic rates. Metabolic enzymes are regulated by nutrients through short-term regulation of their activities and long-term regulation of expression of their genes. Consequences of liver metabolic regulation on energy supply to muscles may affect protein deposition (and hence growth) as well as intramuscular energy stores. Adipose tissues are important body reserves of triglycerides, which result from the balance between lipogenesis and lipolysis. Both processes depend on the feeding level and on the nature of nutrients, which indirectly affect energy delivery to muscles. In muscles, the regulation of rate-limiting nutrient transporters, of metabolic enzyme activities and of ATP production, as well as the interactions between nutrients affect free energy availability for muscle growth and modify muscle metabolic characteristics which determine meat quality. The growth of tissues and organs, the number and the characteristics of muscle fibers depend, for a great part, on early events during the fetal life. They include variations in quantitative and qualitative nutrient supply to the fetus, and hence in maternal nutrition. During the postnatal life, muscle growth and characteristics are affected by the age and the genetic type of the animals, the feeding level and the diet composition. The latter determines the nature of available nutrients and the rate of nutrient delivery to tissues, thereby regulating metabolism. Physical activity at pasture also favours the orientation of muscle metabolism, towards the oxidative type. Consequently, breeding systems may be of a great importance during the postnatal life. Research is now directed towards the determination of individual tissue and organ energy requirements, a better knowledge of nutrient partitioning between and within organs and tissues. The discovery of new molecules (e. g. leptin), of new molecular mechanisms and of more powerful techniques (DNA chips) will help to achieve these objectives. The integration of the different levels of knowledge will finally allow scientists to formulate new types of diets adapted to sustain a production of high quality meat with lower costs of production.

• Journal of the Korean Wood Science and Technology
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• v.32 no.5
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• pp.12-19
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• 2004

This study was performed to investigate the optimum condition of protease production from submerged culture of oak mushroom (Lentinula edodes, Sanlim No. 5) and its enzymatic features. Among several combinations of media, the combination of wheat bran, corn flour, water and corn oil (WB+CF+W+ CO) yielded 84.8 U/g of maximum protease activity. This combination of ingredients, in spite of not being particularly protein-rich in comparison to the other media, allowed for good growth of the fungus and maximal protease production. Comparison of different growth medium liquids indicated that demineralized water afforded the best growth of the fungus and the highest protease activity. Acetate buffer and acidified water negatively affected The protease production peaked around 72 hr of incubation, and decreased thereafter. The molecular weights of produced protease were about 45,000 by Sephadex G-75 chromatography. The pH optimum for protease activity was 4, while maximal activity incubated at 37℃ for 1 hr was observed between pH 4~6. The optimum temperature of this protease was 55℃, and the enzyme was active over a broad temperature range (30~60℃), indicating that this protease would be suitable for a wide range of applications where. different pH and temperature are necessary, such as digestive aids, food industry, beer and tannery industries.

• Journal of Life Science
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• v.18 no.1
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• pp.52-57
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• 2008

Xylan is a major hemicellulose component of the cell walls of monocots and hardwood, representing up to 30% of the dry weight of these plants. To efficiently hydrolyze xylan, the endoxylanase gene from Bacillus sp. was expressed in B. subtilis DB431 by introducing the plasmid pJHKJ4. The total activity of the recombinant endoxylanase reached about 857 unit/ml by batch fermentation of B. subtilis DB431/pJHKJ4 in LB maltose medium. The majority (>92%) of endoxylanase was efficiently secreted into the culture medium. The recombinant endoxylanase hydrolyzed more the birchwood xylan efficiently than the other xylans. When 4 % concentration of xylan was used, the highest production of xylooligosaccharide was observed, and xylobiose and xylotriose were the major products. Optimal amount of enzyme and reaction time for producing xylooligosaccharide were found to be 10 unit and 1 hr, respectively. In addition, the temperature of $40^{\circ}C{\sim}50^{\circ}C$ gave the highest production of xylooligosaccharide. Consequently, the optimized conditions for the production of xylooligosaccharide through the hydrolysis of xylan were determined as follows: 10 unit endoxylanase, $50^{\circ}C$, 4% birchwood xylan, 1 hr reaction.

• Microbiology and Biotechnology Letters
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• v.47 no.3
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• pp.390-400
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• 2019

In this study, strains isolated from soil samples collected from Busan, Changwon, and Jeju Island were examined to verify their abilities of phosphate solubilization and nitrogen fixation, production of indole-3-acetic acid (IAA), siderophore, and 1-aminocylopropane-1-carboxylyic acid (ACC) deaminase in order to select strains that promote plant growth and play a role in biocontrol of pests or pathogens. According to the results of this study, most of the isolated strains were found to have ability of phosphate solubilization, nitrogen fixation, IAA production, siderophore production, and production of ACC deaminase. These isolated strains might help plant growth by directly improving absorption of nutrients essential for phosphate solubilization and nitrogen fixation. In addition, they can promote plant growth and control resistance to plant diseases through extracellular enzyme activity and antifungal activity. In addition, most of the selected strains were found to survive in various environmental conditions such as temperature, salinity, and pH. Therefore, Pseudomonas plecoglossicida ANG14, Pseudarthrobacter equi ANG28, Beijerinckia fluminensis ANG34, and Acinetobacter calcoaceticus ANG35 were finally selected through a comparative advantage analysis to suggest their potential as novel biological agents. Further studies are necessary in order to prove their efficacy as novel biological agents through formulation and optimization of effective microorganisms, their preservation period, and crop cultivation tests.

• Molecules and Cells
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• v.38 no.4
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• pp.318-326
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• 2015

We previously reported that the SbROMT3syn recombinant protein catalyzes the production of the methylated resveratrol derivatives pinostilbene and pterostilbene by methylating substrate resveratrol in recombinant E. coli. To further study the production of stilbene compounds in E. coli by the expression of enzymes involved in stilbene biosynthesis, we isolated three stilbene synthase (STS) genes from rhubarb, peanut, and grape as well as two resveratrol O-methyltransferase (ROMT) genes from grape and sorghum. The ability of RpSTS to produce resveratrol in recombinant E. coli was compared with other AhSTS and VrSTS genes. Out of three STS, only AhSTS was able to produce resveratrol from p-coumaric acid. Thus, to improve the solubility of RpSTS, VrROMT, and SbROMT3 in E. coli, we synthesized the RpSTS, VrROMT and SbROMT3 genes following codon-optimization and expressed one or both genes together with the cinnamate/4-coumarate:coenzyme A ligase (CCL) gene from Streptomyces coelicolor. Our HPLC and LC-MS analyses showed that recombinant E. coli expressing both ScCCL and RpSTSsyn led to the production of resveratrol when p-coumaric acid was used as the precursor. In addition, incorporation of SbROMT3syn in recombinant E. coli cells produced resveratrol and its mono-methylated derivative, pinostilbene, as the major products from p-coumaric acid. However, very small amounts of pterostilbene were only detectable in the recombinant E. coli cells expressing the ScCCL, RpSTSsyn and SbROMT3syn genes. These results suggest that RpSTSsyn exhibits an enhanced enzyme activity to produce resveratrol and SbROMT3syn catalyzes the methylation of resveratrol to produce pinostilbene in E. coli cells.

• Asian-Australasian Journal of Animal Sciences
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• v.24 no.11
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• pp.1529-1540
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• 2011

Silencing of a specific gene using RNAi (RNA interference) is a valuable tool for functional analysis of a target gene. However, information on RNAi for analysis of gene function in farm animals is relatively nil. In the present study, we have validated the interfering effects of siRNA (small interfering RNA) using both quantitative and qualitative gene silencing in buffalo granulosa cells. Qualitative gene knockdown was validated using a fluorescent vector, enhanced green fluorescence protein (EGFP) and fluorescently labeled siRNA (Cy3) duplex. While quantitatively, siRNA targeted against the luciferase and CYP19 mRNA was used to validate the technique. CYP19 gene, a candidate fertility gene, was selected as a model to demonstrate the technique optimization. However, to sustain the expression of CYP19 gene in culture conditions using serum is difficult because granulosa cells have the tendency to luteinize in presence of serum. Therefore, serum free culture conditions were optimized for transfection and were found to be more suitable for the maintenance of CYP19 gene transcripts in comparison to culture conditions with serum. Decline in fluorescence intensity of green fluorescent protein (EGFP) was observed following co-transfection with plasmid generating siRNA targeted against EGFP gene. Quantitative decrease in luminescence was seen when co-transfected with siRNA against the luciferase gene. A significant suppressive effect on the mRNA levels of CYP19 gene at 100 nM siRNA concentration was observed. Also, measurement of estradiol levels using ELISA (enzyme-linked immunosorbent assay) showed a significant decline in comparison to control. In conclusion, the present study validated gene silencing using RNAi in cultured buffalo granulosa cells which can be used as an effective tool for functional analysis of target genes.

• KSBB Journal
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• v.24 no.1
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• pp.101-105
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• 2009

Alcohol oxidation activities and optimization of extraction conditions of Rrhus verniciflua Stokes (RVS) extract were evaluated for the development of a functional biomaterial for improving liver function. When alcohol oxidation activities of RVS was analyzed, the Rrhus verniciflua Stokes bark (RVSB) were higher than the Rrhus verniciflua Stokes heartwood (RVSH). Alcohol oxidation activity value of RVSB increased in a concentration-dependent manner. In the comparative analysis between Hovenia dulcis Thunb (HOT) and Alnus japonica Steud (AJS) which was reported as a alcohol oxidation material, alcohol oxidation activity is much higher than the others. The experimental conditions were optimized for alcohol oxidation-active components production from RVSB. The extraction conditions such as temperature, time, pH and particle size were performed. It was recommended to extract the alcohol oxidation-active components from RVSB by hot water (pH 7.0) at $85^{\circ}C$ for 8 hours.

• Journal of the Korean Society of Food Science and Nutrition
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• v.40 no.10
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• pp.1430-1437
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• 2011

Synthesis conditions of cocoa butter equivalents were optimized using the response surface method (RSM) by interesterification of canola oil (Ca), palmitic ethyl ester (PEE), and stearic ethyl ester (StEE). The reaction was catalyzed by immobilized lipase (Lipozyme TLIM) from Thermomyces lanuginosa to produce structured lipids containing a composition of triacylglycerols similar to cocoa butter. Reaction conditions were optimized using D-optimal design with the three reaction factors of the substrate molar ratio of canola oil to palmitic ethyl ester and stearic ethyl ester (Ca : PEE : StEE=1:1:3, 1:1.66:5, 1:2:6, 1:2.33:7, 1:3:9, $X_1$), enzyme ratio (2~6%, $X_2$), and reaction time (30~270 min, $X_3$). The optimal conditions that minimized acyl-migration while maximizing 1-palmitoyl-2-oleoyl-3-stearoyl glycerol (POS), 1,3-distearoyl-2-oleoyl glycerol (SOS), and 1,3-dipalmitoyl-2-oleoyl glycerol (POP) were predicted, resulting in Ca : PEE : StEE=1:3:9, 6% of enzyme ratio, and 40 min of reaction time. The reaction product of structured lipids was synthesized again under the same conditions, showing 10.43 area% of acyl-migration, 25.31 area% of POS/PSO, 19.79 area% of SOS, and 11.22 area% of POP.

• Food Science and Preservation
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• v.18 no.5
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• pp.721-728
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• 2011

1(3)-palmitoyl-2-oleoyl-3(1)-stearoyl-(POS)-glycerol-enriched reaction products were synthesized from camellia oil, palmitic ethyl ester, and stearic ethyl ester via lipase-catalyzed interesterification. Response surface methodology (RSM) was employed to optimize the production of the POS-enriched reaction product (Y1, %) and the stearicand palmitic-acid contents at the sn-2 position due to acyl migration (Y2, %). The reaction factors were the enzyme amount (X1, 2-6%), reaction time (X2, 60-360 min), and substrate molar ratio of camellia oil to palmitic ethyl ester and stearic ethyl ester (X3, 1-3 mol). The predictive models for Y1 and Y2 were adequate and reproducible as no lack of fit was signified (0.128 and 0.237) and as there were satisfactory levels of R2 (0.968 and 0.990, respectively). The optimal conditions for the reaction product for maximizing Y1 while minimizing Y2 were predicted at the reaction combination of 5.86% enzyme amount, 60 min reaction time, and 1:3 substrate molar ratio (3 moles of palmitic ethyl ester and 3 moles of stearic ethyl ester). Actual reaction was performed under the same conditions as above, and the resulting product contained 20.19% TAG-P/O/S and 12.71% saturated fatty acids at the sn-2 position.