• Asian-Australasian Journal of Animal Sciences
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• 제30권11호
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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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• 제14권5호
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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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• 제32권5호
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• pp.12-19
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• 2004

본 연구는 표고버섯균의 액체배양을 통한 protease효소의 최적생산 조건을 찾고, 생산된 protease 의 효소적 특성을 구명하고자 실시하였다. Protease 대량생산을 위한 공사균으로서 표고버섯균(Lentinula edodes)을 사용하였으며, 최적생산 조건을 구명하기 위하여 생산기질의 조성, 배양조건, 배양기간 등을 검토하였다. 배지의 조성별 protease 생산성에 있어서는 밀기울+옥수수가루+물+옥수수기름(WB+CF+W+CO)조합이 84.8 U/g으로서 가장 높은 protease 생산성을 보여주었고, 콩가루와 소맥분조합의 protease 생산성이 매우 낮은 것을 알 수 있었다. Protease 생산을 위한 배양매체로서 acetate buffer에서는 전혀 protease의 생산이 되지 않았으며, pH 6.2의 이온교환수가 80.5 U/g 의 가장 높은 protease 생산성을 나타냈다 이온교환수를 pH 4.0으로 조절하였을 때 생산성은 62.3 U/g 으로 낮아졌다. 배양기간에 따른 protease 생산성은 배양시간이 경과됨에 따라 증가하였으며, 72시간 배양으로 90.6 U/g 의 최대값에 달하였고, 그 이후 점차 감소하는 경향으로 나타났다 Sephadex G-75 chromatography 로 측정한 생산된 protease 의 분자량은 약 45,000 이었다. Lentinula edodes를 이용하여 생산된 protease의 효소특성을 상이한 pH 및 온도의 조건에서 조사한 결과, 최적 pH는 4에서 나타났으며, 37℃(1 시간)에서 측정한 활성은 pH 4~6 범위에 있었다. Protease의 온도안정성을 조사한 결과 최적온도는 55℃에서 나타났으며, 30~60℃의 넓은 온도범위에서 안정함을 알 수 있어서 이 효소는 소화용 제제, 식품공업, 맥주제조업 및 가죽무두질 등 널리 이용될 수 있을 것으로 생각된다.

• 생명과학회지
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• 제18권1호
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• pp.52-57
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• 2008

식물체 구성성분의 하나인 hemicellulose의 대부분을 차지하는 xylan은 농산 폐기물 또는 목재 성분의 약 30%를 차지하는 풍부한 자원물질이다. 이러한 xylan을 효과적으로 분해하기 위해 Bacillus에서 발현시킨 재조합 endoxylanase를 이용하여 기능성 식품소채인 xylooligosaccharide의 최적 생산 조건을 조사하였다. B. subtilis 재조합 균주 DB431/pJHKJ4를 이용한 발효조 회분배양 결과, endoxylanse의 총 활성은 857 unit/ml이며, 대부분이 세포밖으로 분비됨을 알 수 있었고, 분비효율은 92%로 나타났다. 재조합 endoxylanase를 이용하여 xylan으로부터 xylooligosaccharide 생성을 위한 최적 반응조건을 검토한 결과, xylooligosaccharide 생산을 위한 기질로는 birchwood xylan이 적합함을 알 수 있었고, 4%의 xylan 농도에서 가장 많은 xylooligosaccharide 을 생산하며, xylobiose와 xylotriose가 주생성물임을 알았다. 효소의 농토와 반응시간의 영향은 10 unit의 endoxylanase를 첨가하여 1시간 반응시켰을 때 가장 많은 양의 xylooligosaccharide을 생산할 수 있었고, 반응온도는 $40^{\circ}C{\sim}50^{\circ}C$가 적합함을 알았다. 결론적으로 재조합 endoxylanase을 이용한 xylooligosaccharide생성에는 10 unit endoxylanase과 4% birchwood xylan을 기질로 이용하여 $50^{\circ}C$에서 1시간 반응시키는 것이 최적반응 조건임을 알았다.

• 한국미생물·생명공학회지
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• 제47권3호
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• pp.390-400
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• 2019

본 연구에서 부산, 창원, 제주도 일대에서 채취한 토양으로부터 분리한 미생물을 이용하여 식물 생장 촉진활성 및 식물병원성 곰팡이에 대한 길항능을 확인하고자 하였으며, 분리주 간에 비교우위를 통해 Pseudomonas plecoglossicida ANG14, Pseudarthrobacter equi ANG28, Beijerinckia fluminensis ANG34, Acinetobacter calcoaceticus ANG35를 최종 선정하였다. 이들은 ACC deaminase 생성능, IAA 생성능, 질소 고정능, 인산 가용화능 및 siderophore 생성능을 모두 가지며, 일부 식물병원성 곰팡이에 대해 길항능을 가지는 것을 확인하였다. 특히 ANG35의 경우에는 7가지 식물병원성 곰팡이 중 6가지에 대해 비교적 높은 억제율을 나타내어 식물 생장 촉진뿐만 아니라 방제활성을 가지므로 식물이 생장하는 데 도움을 줄 것으로 기대된다. 또한, 4균주 간의 세포외 효소활성 synergy effect를 확인한 결과 단독배양을 했을 때보다 혼합배양 시 세포외 효소활성이 증가하는 것을 확인하였다. 따라서, 식물 생장촉진 활성 및 식물병원성 곰팡이에 대한 결과를 통해 새로운 생물학적 제제로서 이용 가능성을 제시한다.

• Molecules and Cells
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• 제38권4호
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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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• 제24권11호
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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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• 제24권1호
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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.

• 한국식품영양과학회지
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• 제40권10호
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• pp.1430-1437
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• 2011

본 연구에서는 카놀라유(canola oil)를 이용하여 코코아버터 유사 재구성지질 합성의 최적조건을 연구하였다. 반응기질로는 카놀라유와 palmitic ethyl ester와 stearic ethyl ester를 이용하였으며, 효소는 Lipozyme TLIM을 사용하여 반응기질의 몰비율과 반응효소의 양, 반응시간을 독립변수로 하고, D-optimal 계획에 의한 반응표면분석에 따라 합성을 실시하였다. 반응을 통해 acyl-migration은 최소로 일어나며, POS 및 SOS, POP의 함량은 최대로 합성되는 조건을 최적화 조건으로 결정하였다. D-optimal 계획에 의한 반응표면분석을 실행한 결과, sn-2 위치 지방산 조성의 변화를 일으키는 acyl-migration($Y_1$)은 최소로 일어나고 POS($Y_2$)와 SOS($Y_3$), POP($Y_4$)는 최대로 일어나는 조건은 기질의 몰비율은 canola oil : palmitic ethyl ester : stearic ethyl ester=1:3:9이며, 반응효소의 양은 6%이고, 반응시간은 40 min이었다. 이와 같은 반응조건으로 합성된 재구성지질의 acylmigration(% of palmitic acid+stearic acid, $Y_1$)은 10.43%, POS($Y_2$)의 함량은 25.31%, SOS($Y_3$)는 19.79%, 그리고 POP($Y_4$)는 11.22%로 측정되었다.

• 한국식품저장유통학회지
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• 제18권5호
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• pp.721-728
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• 2011

Camellia oil과 palmitic, 그리고 stearic ethyl ester를 기질로 하고 Lipozyme TLIM을 촉매로 이용하여 interesterification을 통한 POS를 많이 함유한 반응물을 합성하고자 하였다. 반응 조건(반응효소비율, 반응시간, 기질의 비율)을 독립변수로 하여 중심 합성 계획에 의한 반응 표면 분석을 통하여 POS의 함량은 높이고, sn-2 position의 acyl migration을 낮추어 합성하는 최적화 조건을 얻었다. POS를 많이 함유한 반응물의 합성 최적화 조건은 반응 효소양($X_1$)= 5.9%, 반응시간($X_2$)= 60 min, 그리고 기질비율($X_3$)= 1:3(1 mole의 camellia oil: 3 mole의 palmitic ethyl ester + 3 mole의 stearic ethyl ester)이었다. 이와 같은 조건으로 획득된 실재 효소반응 합성물의 TAG-P/O/S(palmitic, stearic과 oleic 지방산을 각각 1 분자씩 어느 위치이던지 함유한 TAG, 즉 POS 및 PSO/OSP/OPS/SPO)의 합성율($Y_1$)은 20.19%이었고, 이때 acyl migration 등에 의하여 sn-2 위치에 결합(이동)한 palmitic과 stearic acid의 양(P/S-sn-2)인 $Y_2$는 12.71% 이었다.