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Factors Affecting the Survival Rates of Frozen-Thawed Spermatozoa in Equine by Glycerol Concentration and Freezing Speed (말 정액 동결시 Glycerol 농도와 동결 속도가 생존율에 미치는 영향)

  • Choi, Sun-Ho;Kim, Sung-Jae;Cho, Sang-Rae;Choe, Chang-Yong;Son, Jun-Kyu;Yoo, Yong-Hee;Cho, Young-Jae;Choi, Gui-Cheol;Moon, Yun-Young
    • Reproductive and Developmental Biology
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    • v.34 no.3
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    • pp.271-274
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    • 2010
  • This study was conducted to investigate the survival rate of frozen-thawed spermatozoa in equine by glycerol concentration and freezing speed Two stallions (1 Thoroughbred-13 year old and 1 Arab-7 year old) bred in Korea Racing authority was examined for 1 times in a couple of weeks. Semen was collected by condom method standing heated mare and were centrifuged 650 g for 15 min. and isolated the seminal plasma. Thick fraction of semen was diluted EDTA-Lactose-egg yolk diluents to 1:1 and contained in 0.5 ml straw as $6{\sim}14{\times}10^7\;cells/ml$. Final concentrations of glycerol were 3, 5 and 7% in cryopreseved diluents and added 4 times for 2 hours equilibration. For the freezing, equilibrated straws were located 3 or 5 em above $LN_2$ gas for 5 or 10 min. Survival rates of pre-frozen sperm were $65.0{\pm}13.2%$, $68.3{\pm}10.4%$, $66.7{\pm}11.5%$ and post-frozen were $53.3{\pm}23.1%$, $45.0{\pm}15.0%$, $50.0{\pm}18.0%$ in 3, 5, 7% glycerol concentration, respectively. There was no difference between glycerol concentrations. Survival rates of frozen-thawed sperm on freezing speed were $36.7{\pm}10.4%$, $40.0{\pm}7.1%$, $30.0{\pm}13.2%$ at 3 cm-5 min and $33.3{\pm}11.5%$, $31.7{\pm}2.9%$, $21.7{\pm}10.4%$ at 3 cm-10 min in 3, 5, 7% glycerol concentration, respectively. Survival rates of frozen-thawed sperm on freezing speed were $43.3{\pm}15.3%$, $32.0{\pm}17.9%$, $22.3{\pm}15.7%$ at 5cm-5 min and were $47.5{\pm}15.0%$, $43.3{\pm}12.6%$, $48.3{\pm}15.3%$ at 5cm-10 min in 3, 5, 7% glycerol concentration, respectively. There were significantly different between groups (p<0.05). These results suggest that glycerol concentration did not affect cryopreservation of stallion semen within 3~7% but freezing speed affects. In our experiment, the best cryopreservation condition was at 5 cm above $LN_2$ gas for 10 min for pre-freezing and 7% of glycerol concentration. These results lead to commercial AI with frozen-thawed stallion semen.

Changes in the Myocardial Antioxidant Enzyme System by Post-Ischemic Reperfusion During Corontory Artery Bypass Operations (관상동맥우회술시 심근허혈후 재관류에 의한 활성산소 방어효소계의 변화)

  • 김응중;김기봉
    • Journal of Chest Surgery
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    • v.29 no.8
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    • pp.850-860
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    • 1996
  • Oxygen free radicals and their metabolites have been implicated as possible causes of reperrusion injury In animal models. Their role in the clinical setting is still controversial. The aim of this study was to evaluate the degree of tissue damage, oxidative stress. and changes in the antioxidant enzyme system in patients undergoing cor nary artery bypass graft operations(CABG) with myocardial protection by cold blood cardioplegia. In patients undergoing CABG(n:10). the levels of lactate dehydrogenate(LDH), creatine phosphokinase MB fraction(CK-MB), and malondialdehyde(M DA) were measured In the coronary sinus effluent before aortic cross clamping and 20 minutes after reperfusion. At the same time, the myocardial tissue activities of superoxide dismutase(SOD). catalase(CAT), glutathione peroxiddse(GSHPX), glutathione reductase (GSSGRd), and glucose 6-phosphate dehydrogenate(GfPDH ) were determined in the right atrial auricle excised before aortic cross clamping and in the left atrial auricle excised 20 minutes after reperfuslon. The levels of increased significantly after reperrusion(p< U.05). There were no significant changes in CAT and CfPDH levels. Western blot analysis was performed to study the induction of antioxidant enzyme and demonstrated increased amount of Cu,Zn-SOD.

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The Compost Inoculation Effect on the Cellulose Degradation in Bench-scale Composting of Food Waste and Paper Mixture (음식쓰레기의 실험실 규모 퇴비화에서 셀룰로스 분해에 대한 퇴비 식종효과)

  • Shin, Hang-Sik;Jeong, Yeon-Koo;Hwang, Eung-Ju
    • Journal of the Korea Organic Resources Recycling Association
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    • v.6 no.1
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    • pp.21-30
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    • 1998
  • The effects of compost inoculation on the degradation of cellulosic fraction in composting of food waste and paper mixture were investigated by bench-scale composting. With the increase of seed inoculation, the time to reach the peaks of temperature, $CO_2$ evolution rate, and ammonia evolution rate was reduced, indicating that seed compost had beneficial effects on the enhanced degradation of organic materials at the early stage of composting. However, the final conversion of organic matters and the loss of ammonia were not affected by the amount of seed compost inoculated. The increasing of seed inoculum also resulted in the higher level of cellulase activity at initial stages and rapid rise to the maximums, suggesting that initial supply of sufficient cellulolytic microorganisms might facilitate the evolution of cellulase activity. The cellulose was degraded substantially during the increasing phase of cellulase activity, while they showed similar values at the end of 20 days composting. As a result, the seed inoculation seemed to be effective to the enhanced evolution of cellulase activity and cellulose degradation at initial stage of composting. But it did not contribute to increase the final degradation of cellulose after the entire composting reaction of 20 days.

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The Effects of Polygala Tenuifolia DM Fraction on CT105-injuried Neuronal Cells (원지 디클로로메탄분획이 CT105에 의한 신경세포 상해에 미치는 영향)

  • Lee Sang Won;Kim Sang Ho;Kim Tae Heon;Kang Hyung Won;Lyu Yeoung Su
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.18 no.2
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    • pp.507-516
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    • 2004
  • Alzheimer's disease(AD) is a geriatric dementia that is widespread in old age. In the near future AD will be the commom disease in public health service. Although a variety of oriental presciptions in study POD(Polygala tenuifolia extracted from dichlorometan) have been traditionally utilized for the treatment of AD, their pharmacological effects and action mechanisms have not yet fully elucidated. It has been widely believed that AP peptide divided from APP causes apoptotic neurotoxicity in AD brain. However, recent evidence suggests that CT105, carboxy terminal 105 aminoacids peptide fragment of APP, may be an important factor causing neurotoxicity in AD. SK-N-SH cells expressed with CT105 exhibited remarkable apoptotic cell damage. Based on morphological observations by phase contrast microscope and NO formation in the culture media, the CT105-induced cell death was significantly inhibited by POD. In addition, AD is one of brain degeneration disease. So We studied on herbal medicine that have a relation of brain degeneration. From old times, In Oriental Medicine, PO water extract has been used for disease in relation to brain degeneration. We were examined by ROS formation, neurite outgrowth assay and DPPH scravage assay. Additionally, we investigated the association between the CT105 and neurite degeneration caused by CT105-induced apoptotic response in neurone cells. We studied on the regeneratory and inhibitory effects of anti-Alzheimer disease in pCT105-induced neuroblastoma cell lines by POD. Findings from our experiments have shown that POD inhibits the synthesis or activities of CT105, which has neurotoxityies and apoptotic activities in cell line. In addition, treatment of POD(>50 ㎍/㎖ for 12 hours) partially prevented CT(105)-induced cytotoxicity in SK-N-SH cell lines, and were inhibited by the treatment with its. POD(>50 ㎍/㎖ for 12 hours) repaired CT105-induced neurite outgrowth when SK-N-SH cell lines was transfected with CT105. As the result of this study, In POD group, the apoptosis in the nervous system is inhibited, the repair against the degerneration of Neuroblastoma cells by CT105 expression is promoted. Decrease of memory induced by injection of scopolamin into rat was also attenuted by POD, based on passive avoidance test. Taken together, POD exhibited inhibition of CT105-induced apoptotic cell death. POD was found to reduce the activity of AchE and induced about the CA1 in rat hippocampus. Base on these findings, POD may be beneficial for the treatment of AD.

Analysis of Mobile Cadmium from Soils with Functionalized Magnetic Beads (기능성 자성입자를 이용할 토양의 유동성 카드뮴 분석)

  • So Hyungsuk;Nah In-Wook;Hwang Kyung-Yub;Shin Hyun Chul;Kim Beom-Suk;Yoo Yeong-Seok
    • Journal of Soil and Groundwater Environment
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    • v.10 no.1
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    • pp.13-17
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    • 2005
  • Potential for measuring mobile cadmium concentration in sandy soil using polymer magnetic beads with carboxyl groups was investigated. Experiments for extracting cadmium were performed with contaminated soils, de-ionized water and magnetic beads. In this neutral experimental condition, reacting cadmium with magnetic beads indicate total amount of cadmium that can be moved in soil. The results showed that the mobile fraction of cadmium in soil could be combined with magnetic beads in short time. After binding between cadmium and magnetic beads, the beads were separated from soil suspension by outer magnetic force. The bound cadmium was dissolved from magnetic beads by acid solutions, which were then analyzed by atomic absorption spectroscopy (AAS). This method can determine mobile heavy metals in sandy soil effectively than existing method which use pollutant chemicals to environments such as EDTA.

Growth Temperature-Dependent Conversion of De novo-Synthesized Unsaturated Fatty Acids into Polyhydroxyalkanoic Acid and Membrane Cyclopropane Fatty Acids in the Psychrotrophic Bacterium Pseudomonas fluorescens BM07

  • LEE , HO-JOO;RHO, JONG-KOOK;YOON, SUNG-CHUL
    • Journal of Microbiology and Biotechnology
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    • v.14 no.6
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    • pp.1217-1226
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    • 2004
  • A psychrotrophic bacterial strain, Pseudomonas fluorescens BM07, synthesized unsaturated fatty acids (UFA) from fructose in response to lowering of growth temperature, and incorporated them into both polyhydroxyalkanoic acid (PHA) and membrane lipid. The blocking of PHA synthesis by adding 5 mM 2-bromooctanoic acid to the growth medium, containing 70 mM fructose, was found to be a useful means to profile the composition of membrane lipid by gas chromatography. As the growth temperature changed from 35 to $50^{\circ}C$, the total content of two UFA, 3-hydroxy-cis-5­dodecenoic acid ($C_{12:1}$) and 3-hydroxy-cis-7-tetradecenoic acid ($C_{14:1}$), in PHA increased from 31 to 44 $mol\%$. The growth at lower temperatures also led to an increase in the level of two major UFA, palmitoleic acid (C16:1 cis9) and cis-vaccenic acid (C18:1 cis11), in membrane lipid. A fraction of these membrane-lipid UFA was converted to their corresponding cyclopropane fatty acids (CFA). The CFA conversion was a function of culture time, exhibiting biphasic increase before and after entering the stationary phase. However, pH changes in growth media had no effect on the CFA conversion, which is contrary to the case of E. coli reported. The cells grown at $30^{\circ}C$ responded to a cold shock (lowering the medium temperature down to $10^{\circ}C$) by increasing the level of C16:1 cis9 and C 18: I cis II up to that of $10^{\circ}C$-grown control cells and concomitantly decreasing the relative level of cis-9,10­methylenehexadecanoic acid (the CFA converted from C16:1 cis9) from 14 to 8 $mol\%$, whereas the 10-grown cells exhibited little change in the lipid composition when exposed to a warmer environment of $30^{\circ}C$ for 12 h. Based on this one- way response, we suggest that this psychrotrophic strain responds more efficiently and sensitively to a cold shock than to a hot shock. It is also suggested that BM07 strain is a good producer of two unsaturated 3-hydroxyacids, $C_{12:1}\;and\;C_{141:1}$.

Effects of Microbacterium laevaniformans Levans Molecular Weight on Cytotoxicity

  • Oh, Im-Kyung;Yoo, Sang-Ho;Bae, In-Young;Cha, Jae-Ho;Lee, Hyeon-Gyu
    • Journal of Microbiology and Biotechnology
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    • v.14 no.5
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    • pp.985-990
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    • 2004
  • Levans produced from Microbacterium laevaniformans were isolated, characterized, and fractionated by molecular weight. TLC, HPLC, and GC-MS analyses of the exopolysaccharide showed that it was a fructan-type polymer and was composed of (2,6)- and (2,1)-glycosidic linkages. $^{13}C$-NMR analysis proved that the polysaccharide was mainly a $\beta$-(2,6)-linked levan-type polysaccharide. To investigate the cytotoxicity of the acetone-precipitated levan fractions such as M1, M2, and M3, HepG2, P388D1, U937, SNU-1, and SNUC2A cell lines were screened. Among the cell lines tested, the cytotoxicity of M1- M3 fractions were detected from only SNU-1 and HepG2 cells at the dosage level of $100-800\mu\textrm{g}ml$. The M2 fraction M_r$, 80,000) at 400 $mu{g/ml}$ had the greatest cell growth inhibition (84.6%) on SNU-1, while the M1 $(M_r$, 50,000) at $800\mu\textrm{g}ml$ showed the greatest (46.32%) on HepG2. To obtain more uniform M_r$ fractions of levan, the levan was further fractionated from S1 $(M_r$ 1,000,000) to S5 $(M_r$ 10,000) using gel permeation chromatography. Again, the S1-S5 fractions had strong cytotoxicity on SNU-1 and HepG2 cell lines. The greatest inhibition effects of S4 $(M_r$ 80,000) on SNU-1 and S5 $(M_r$ 10,000) on HepG2 were shown to be 49.5% and 73.0%, respectively. The cytotoxicity of the levan fractions was more effective on SNU-1 than on HepG2. Although the relationship between the Mw and the cytotoxicity was not clear, smaller $M_r$, fractions of levan showed greater growth inhibition effect on the cancer cell lines in general. Therefore, it was indicated that a specific Mw class of levan is responsible for the effective cytotoxicity.

Evaluation of Novel Constitutive Expression Vectors Equipped with Mined Promoters from Metagenome (메타게놈에서 발굴한 프로모터를 장착한 새로운 항시발현 벡터의 가치평가)

  • Han, Sang-Soo;Kim, Geun-Joong
    • Microbiology and Biotechnology Letters
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    • v.36 no.4
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    • pp.260-267
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    • 2008
  • The choice of expression vector is very important for industrial production of proteins. Therefore, the systematic mining of promoters over a wider range of genetic resource and/or host is required. We previously reported a novel bidirectional reporting system (pBGR) for the isolation of promoters from metagenome and screened useful promoters that functioned constitutively in E. coli under general culture conditions. Among them, three promoter sequences including each upstream region were amplified by PCR and used to construct new expression vectors. To facilitate subcloning, a multi-cloning site was incorporated into the downstream region of the revere primer sequence. At these sites, GFP, esterase and $\beta$-glucosidase were subcloned and analyzed the constitutive expression ability of new promoter in terms of protein solubility and expression level. As a result, these vectors expressed the proteins constitutively to a level of $2{\sim}3%$ of the total cell protein in soluble fraction (>80 %). This study suggested that excavation of metagenomic promoters for construction of expression vector in a certain strain could provide a way for the development of the expression systems.

Hydrogenation and Electrochemical Characteristics of Amorphous-nanostructured Mg-based Alloys

  • Gebert, A.;Khorkounov, B.;Schultz, L.
    • Journal of Powder Materials
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    • v.13 no.5 s.58
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    • pp.327-335
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    • 2006
  • In the development of new hydrogen absorbing materials for a next generation of metal hydride electrodes for rechargeable batteries, metastable Mg-Ni-based compounds find currently special attention. Amor phous-nanocrystalline $Mg_{63}Ni_{30}Y_7$ and $Mg_{50}Ni_{30}Y_{20}$ alloys were produced by mechanical alloying and melt-spinning and characterized by means of XRD, TEM and DSC. On basis of mechanically alloyed Mg-Ni-Y powders, complex hydride electrodes were fabricated and their electrochemical behaviour in 6M KOH (pH=14,8) was investigated. The electrodes made from $Mg_{63}Ni_{30}Y_7$ powders, which were prepared under use of a SPEX shaker mill, with a major fraction of nanocrystalline phase reveal a higher electrochemical activity far hydrogen reduction and a higher maximum discharge capacity (247 mAh/g) than the electrodes from alloy powder with predominantly amorphous microstructure (216 mAh/g) obtained when using a Retsch planetary ball mill at low temperatures. Those discharge capacities are higher that those fur nanocrystalline $Mg_2Ni$ electrodes. However, the cyclic stability of those alloy powder electrodes was low. Therefore, fundamental stability studies were performed on $Mg_{63}Ni_{30}Y_7$ and $Mg_{50}Ni_{30}Y_{20}$ ribbon samples in the as-quenched state and after cathodic hydrogen charging by means of anodic and cathodic polarisation measurements. Gradual oxidation and dissolution of nickel governs the anodic behaviour before a passive state is attained. A stabilizing effect of higher fractions of yttrium in the alloy on the passivation was detected. During the cathodic hydrogen charging process the alloys exhibit a change in the surface state chemistry, i.e. an enrichment of nickel-species, causing preferential oxidation and dissolution during subsequent anodization. The effect of chemical pre-treatments in 1% HF and in $10\;mg/l\;YCl_3/1%\;H_2O_2$ solution on the surface degradation processes was investigated. A HF treatment can improve their anodic passivation behavior by inhibiting a preferential nickel oxidation-dissolution at low polarisation, whereas a $YCl_3/H_2O_2$ treatment has the opposite effect. Both pre-treatment methods lead to an enhancement of cathodically induced surface degradation processes.

Effect of Benzoyl Peroxide on the Activity of Drug-metabolizing Enzyme System and Lipid Peroxidation in Rats (Benzoyl peroxide가 흰쥐의 지질과산화현상에 미치는 영향)

  • Lee, H.W.;Rhee, K.S.;Hong, S.U.
    • The Korean Journal of Pharmacology
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    • v.18 no.1
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    • pp.55-63
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    • 1982
  • Lipid peroxidation is the reaction of oxidative deterioration of polyunsaturated lipids and this peroxidation involves the direct reaction of oxygen and lipid to form free radical intermediates, which can lead to autocatalysis. As results of the extensive studies on the lipid peroxidation by many authors, the relationship between lipid peroxidation and the drug metabolizing system as well as the actions of free radicals on the peroxidation was reasonably well known. For a long time, the mechanism of hepatotoxicity of $CCl_4$ was not clearly understood. However, it is now quite well established that $CCl_4$ is activated in vivo to a free radical which is a highly reactive molecule. Therefore, lipid peroxidation which induces the reduction of cytochrome P-450 and aminopyrine demethylase activity is known as decisive event of $CCl_4$ hepatotoxicity. On the other hand, it was also reported that singlet molecular oxygen produces lipid peroxidation in liver microsomes. In this study the effects of benzoyl peroxide on the lipid peroxidation and drug-metabolizing enzyme were examined. Benzoyl peroxide mixed with starch and phosphates etc. is usually used as a food additive for flour bleaching and maturing purpose because of its oxidative property. Albino rats were used for the experimental animals. Benzoyl peroxide was suspended in soybean oil and sesame oil and administered intraperitoneally or orally. TBA value and aminopyrine demethylase activity were determined in liver microsomal fraction and serum. The results were summerized as following. 1) Body weights of animals administered benzoyl peroxide suspension were decreased while that of oil administered group were increased. 2) The activity of aminopyrine demethylase was generally decreased in animals administered oil suspension of benzoyl peroxide. Furthermore, the marked reduction of the enzyme activity was observed in animals administered benzoyl peroxide intraperitoneally. 3) Generally, microsomal TBA values as well as serum TBA were significantly elevated in benzoyl peroxide group in comparison with the control group. However, the more remarkable increase of serum TBA than microsomal TBA was observed in animals administered orally for 6 days. 4) Specifically, the changing pattern of TBA value was notable in serum rather than in liver microsome by intraperitoneal administration of benzoyl peroxide.

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