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Screenin of Phospholipase D Producing Actinomycetes (방선균으로부터 Phospholipase D 생산균주의 탐색)

  • 손동화;심재용;윤석후
    • Microbiology and Biotechnology Letters
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    • v.22 no.4
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    • pp.333-339
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    • 1994
  • In order to screen microorganisms producing phospholipase D (PLD) [EC 3.1.4.4], culture broths of about 900 strains of soil bacteria were subjected to examine for the PLD activity. When the hydrolytic activity of PLD (H-activity) in the supernatant was determined, 64 strains produced PLD more than 0.3 unit/ml and all of them were actinomycetes. Among 26 culture broths tested, 6 ones had transphosphatidylation activity (T-activity) of 30~68%. When the strains except one were cultivated on 3 different media at 30$\circ$C for 3 days under aerobic condition, strain # 1090 on medium B (yeast extract 1%, peptone 1%, glucose 1.5%, glycerol 1%, CaCO$_{3}$ 0.4%, and pH 7.2) produced PLD with much higher H- and T-activity, which were 8.3 units/ml and 76.3%, respectively. Subsequently, time course of PLD production of the strain # 1090 during cultivation with aeration of 1 v/v/m and agitation of 400 rpm at 30$\circ$C for 5 days on medium B in jar fermentor was investigated. H-activty of PLD reached almost maximum (about 9 units/ml) after 32 hours and maximal T-activity was found to be about 80%.

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Germination Dependency of Antioxidative Activities in Brown Rice (현미의 발아정도에 의한 항산화활성의 변화)

  • Kang, Bo-Ra;Park, Mi-Jung;Lee, Heum-Sook
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.35 no.4
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    • pp.389-394
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    • 2006
  • The change of antioxidative character by germination of brown rice was evaluated. From the total methanolic extract of brown rice, 2.5 mm-germinated brown rice, and 5 mm-germinated brown rice, SOD-like activity and nitrite scavenging ability were identified as antioxidative character. SOD-like activities and nitrite scavenging abilities of all samples were changed dose-dependently and germination-dependently. After successive partitioning with hexane, ethyl acetate (EtOAc) and water, each fraction was tested for these activities. SOD-like activities of all fractions were increased by germination, and especially hexane fraction and EtOAc fraction of 5 mm-germinated brown rice had more strong activities than 50 ppm vitamin C. The $EC_{50}$ values of SOD-like activity showed a gradual decrease by germination and that of EtOAc fraction of 5 mm-germinated brown rice was 17 ppm, which was lower concentration than that of 50 ppm vitamin C. The $IC_{50}$ values of nitrite scavenging ability at PH 1.5 also underwent a great decrease by germination and germinated brown rice had the nitrite scavenging ability at lower concentration than brown rice. The results suggest that SOD-like activity and nitrite scavenging ability are thought to be enhanced by the germination effect.

Biochemical Characterizations of Phenylalanine Ammonia-Lyase and its Mutants to Develop an Enzymatic Therapy for Phenylketonuria (페닐케톤뇨증의 효소치료 개발을 위한 phenylalanine ammonia-lyase 및 유전자 변이형의 생화학적 특성)

  • Kim, Woo-Mi
    • Journal of Life Science
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    • v.19 no.9
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    • pp.1226-1231
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    • 2009
  • Enzyme substitution with recombinant phenylalanine ammonia-lyase (PAL, EC 4.3.1.5) is currently being explored for treatment of phenylketonuria (PKU), an autosomal recessive genetic disorder with mutations of the gene encoding phenylalanine-4-hydroxylase (EC 1.14.16.1). However, oral administration of PAL is limited because of proteolytic digestion in the gastrointestinal tract. The aim of this study was to determine the biochemical properties of PAL and delinate the susceptibility of wild-type PAL to pancreatic proteolysis by exploring several mutants, and to develop therapeutic drugs with PAL for PKU. The specific activity of PAL was assayed and its optimal pH, temperature stability, and intestinal protease susceptibility were investigated. Its $V_{max}$ values for phenylalanine and tyrosine were 1.77 and $0.47{\mu}mol$/ min/mg protein, respectively, and its $K_m$ values were $4.77{\times}10^{-4}$ and $4.37{\times}10^{-4}\;M$, respectively. PAL showed an optimal pH at 8.5, corresponding to the average pH range of the small intestine. It showed no loss of activity at $-80^{\circ}C$ for 5 months and possessed 93.4% of its activity under $4^{\circ}C$ for 4 wks. PAL was susceptible to chymotrypsin digestion and, to a lesser extent, to trypsin, elastase, carboxypeptidase A, and B. The trypsin and chymotrypsin cleaving sites were mutated to investigate protection from pancreatic digestion and the specific activities of these mutants were evaluated. The six mutants displayed low specific activities compared to the wild-type, suggesting that the primary trypsin and chymotrypsin cleaving sites may be essential for catalytic reaction. The PAL mutants could therefore be applied as a pretreatment modality without susceptibility to proteolytic attack, however, additional modification for enhancing enzymatic activity is needed to reduce the Phe levels effectively.

Soil Microbial Flora and Chemical Properties as Influenced by the Application of Pig Manure Compost (돈분퇴비의 시용이 배추재배지 토양의 미생물상 및 화학성에 미치는 영향)

  • Weon, Hang-Yeon;Kwon, Jang-Sik;Suh, Jang-Sun;Choi, Woo-Young
    • Korean Journal of Soil Science and Fertilizer
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    • v.32 no.1
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    • pp.76-83
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    • 1999
  • Studies were conducted during 2 months from May of 1997 to evaluate the effects of pig manure compost(PMC) on soil microbial flora. To do so, a field experiment of Chinese cabbage(Brassica campestris L.) was conducted in a randomized block design on a sandy loam soil and microbial floral characteristics in soils were analyzed. Treatments to control included the application of PMC at (A) $8Mg\;ha^{-1}$CM-8), (B) $29Mg\;ha^{-1}$(CM-2,9), and (C) $57Mg\;ha^{-1}$(CM-57), and of chemical fertilizer(D) at $320N-80P_2O_5-200K_2O\;kg\;ha^{-1}$(NPK). In each treatment, the rhizosphere and non-rhizosphere soils were tested for the analysis of microbial populations. The populations of bacteria, actinomycetes, and fungi increased in soils with the applications of PMC and chemical fertilizer, but that of Bacillus sp. decreased. However, the population of fluorescent Pseudomonas sp. was reduced in NPK plots only. With increasing application rates of PMC, the number of colony forming units(cfu) of bacteria (Pseudomonas sp. and actinomycetes) and fungi increased. in all PMC-treated plots, the population density peaked at early growth stage for bacteria(including Bacillus sp.), at late growth for fluorscent Pseudomonas sp., and at harvest for fungi and actinomycetes. The rhizosphere effect was greatest for fluorscent Pseudomonas sp. As the application rates of PMC increased, Total N, organic matter, available phosphate, and exchangeable -K, -Ca, and -Mg increased compared to control, but soil pH was lowered. In NPK plots, EC was 3.4-fold and exchangeable K was 5-fold higher than control.

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Characteristics of Manure and Estimation of Nutrient and Pollutant of Holstein Dairy Cattle (홀스타인 젖소 분뇨의 특성과 비료성분 및 오염물질 부하량 추정)

  • Choi, D.Y.;Choi, H.L.;Kwag, J.H.;Kim, J.H.;Choi, H.C.;Kwon, D.J.;Kang, H.S.;Yang, C.B.;Ahn, H.K.
    • Journal of Animal Science and Technology
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    • v.49 no.1
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    • pp.137-146
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    • 2007
  • This study was conducted to determine fertilizer nutrient and pollutant production of Holstein dairy cattle by estimating manure characteristics. The moisture content of feces was 83.9% and 95.1% for urine. The pH of feces and urine were in the ranges of 7.0~7.4 and 7.5~7.8, respectively. The average BOD5, COD, SS, T-N, T-P concentrations of the dairy feces were 18,294, 52,765, 102,889, 2,575, and 457mg/ℓ, respectively. Dairy urine showed lower levels of BOD5(5,455mg/ℓ), COD(8,089mg/ℓ), SS(593mg/ℓ), T-N(3,401mg/l), and T-P(13mg/ℓ) than feces. The total daily produced pollutant amounts of a dairy cow were 924.1g(Milking cow), 538.8g(Dry cow), 284.4g(Heifer) of BOD5, 2,336.5g (Milking cow), 1,651.8g(Dry cow), 734.1g(Heifer) of COD and 4,210.1g(Milking cow), 2,417.1g(Dry cow), 1,629.1g(Heifer) of SS and 194.8g(Milking cow), 96.4g(Dry cow), 58.3g(Heifer) of T-N and 24.0g(Milking cow), 10.2g(Dry cow), 6.1g(Heifer) of T-P. The calculated amount of pollutants produced by a 450kg dairy cow for one year were 181.3kg of BOD5, 492.5kg of COD, 899.9kg of SS, 36.0kg of T-N and 4.1kg of T-P. The total yearly estimated pollutant production from all head(497,261) of dairy cattle in Korea is 90,149 tons of BOD5, 244,890 tons of COD, 447,491 tons of SS, 17,898 tons of T-N and 2,008 tons of T-P. The fertilizer nutrient concentrations of dairy feces was 0.26% N, 0.1% P2O5 and 0.14% K2O. Urine was found to contain 0.34% N, 0.003% of P2O5 and 0.31% K2O. The total daily fertilizer nutrients produced by dairy cattle were 197.4g (Milking cow), 97.4g(Dry cow), and 57.9g(Heifer) of Nitrogen, 54.2g(Milking cow), 22.2g(Dry cow), and 14.2g(Heifer) of P2O5 and 110.8g(Milking cow), 80.4g (Dry cow), and 39.5g(Heifer) of K2O. The total yearly estimated fertilizer nutrient produced by a 450kg dairy animal is 36.2kg of N, 8.8kg of P2O5, 24.6kg of K2O. The estimated yearly fertilizer nutrient production from all dairy cattle in Korea is 18,000 tons of N, 4,397 tons of P2O5, 12,206 tons of K2O. Dairy manure contains useful trace minerals for crops, such as CaO and MgO, which are contained in similar levels to commercial compost being sold in the domestic market. Concentrations of harmful trace minerals, such as As, Cd, Hg, Pb, Cr, Cu, Ni, Zn, met the Korea compost standard regulations, with some of these minerals being in undetected amounts.