• Title/Summary/Keyword: in-vitro

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Developmental Capacity of Bovine Follicular Oocytes after Ultra-Rapid Freezing by electron Microscope Grid II.Cryopreservation of In Vitro Matured Bovine Oocytes (Electron Microscopic Grid를 이용한 초급속 동결이 소 난포란의 발달능에 미치는 영향. II. 체외 성숙된 소 미수정란의 동결에 관한 연구)

  • 김은영;김남형;이봉경;윤산현;박세필;정길생;임진호
    • Korean Journal of Animal Reproduction
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    • v.22 no.1
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    • pp.1-9
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    • 1998
  • This study was carried out to confirm whether the developmental capacity of bovine mature oocytes frozen ultra-rapidly using electron microscopic(EM) grids and EFS30 can be obtained, and whether the cryoprotectants and the freezing method used in this study effect detrimentally to the bovine oocytes by indirect immunocytochemistry. As freezing solution, we used EFS30 which consisted of 30% ethylene glycol, 0.5 M sucrose, 18% ficoll and 10% FBS added in D-PBS. The results obtained in this experiment were summarized as follows: When the effects of cryoprotectant and freezing procedure on the microtuble, micrfilament and chromatin morphology of oocytes were evaluated using indirect immunocytochemistry, the results of freezing as well as exposure group were not different with that of the control oocytes. When the fertilization abnormality after ultrarapid freezing of bovine mature oocytes was examined by Hoechst staining, the rates of total penetration(96.7, 9.0%), normal two pronuclei formation(74.6, 68.9%) and mean number of sperm / oocyte(1.50, 1.44) were not different between control and freezing group. In addition, when the developmental capacity of frozen-thawed of oocytes(85.5%) was survived, 74.5% of them were cleaved and 31.4% of cleaved embryos were developed to blastocyst. These data were similar to those of the control(76.0%, 34.6%) and exposure(74.5%, 33.0%) except survival rates. Also, when the total cell number of blastocysts produced from the each treatment at day after IVF was examined by hoechst staining, there were not different among groups. There results demonstrate that developmental capacity of frozen-thawed bovine mature oocytes can be successfully obtained by ultra-rapid freezing method using EM grid and EFS30 solution.

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The application of alternative methods for skin irritation evaluation on pesticides (농약에 대한 피부자극성 대체시험법 적용연구)

  • Jeong, Mi-Hye;Kim, Mi-Kyoung;Park, Soo-Jin;You, Are-Sun;Hong, Soon-Sung;Park, Kyung-Hun;Park, Jae-Eup
    • The Korean Journal of Pesticide Science
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    • v.16 no.3
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    • pp.261-266
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    • 2012
  • It is common to use many experiment animals to evaluate the toxicity of chemicals including pesticides. For protecting animal, the concepts of 3R (Reduction, Replacement, Refinement) were introduced and in vitro alternatives methods actively have been developed all over the world. Many experimental animals for toxicological tests have been used, so that it is important to establish the alternative methods. In this study, the alternative method using reconstituted human skin model (Keraskin$^{TM}$) was conducted for classification of skin irritation on pesticides. Sixteen formulations selected on the basis of the degree of irritation were treated by Keraskin$^{TM}$ test. The percent of cell viability was measured into the culture medium collected after treatment of the pesticides for 24-72 hrs. The skin irritations of formulations were evaluated by the cell viability. In this study, The 4 formulations with mild irritation in rabbits were evaluated as nonirritant, the 6 formulations with moderate and severe irritation were evaluated as irritant in human skin model test. We suggest that the alternative test using Keraskin$^{TM}$ model could be used as toxicity evaluation for primary irritation index (P.I.I.) score of greater than or equal to 2.1 of pesticides. The further studies should be required to apply for hazardous assessment of pesticides on alternative skin irritation methods because of the interindividual variability of the sensitivity of skin irritation on pesticides.

Survival Rate of Lactic Acid Bacteria and the Change of ${\beta}-Galactosidase$ Activity in Commercial Yogurts Under the Acidic Conditions (산성조건하에서 시판요구르트의 유산균 생존률과 ${\beta}-galactosidase$의 활성도)

  • Shin, Yong-Seo;Sung, Hyun-Ju;Kim, Dong-Han;Lee, Kap-Sang
    • Applied Biological Chemistry
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    • v.37 no.3
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    • pp.143-147
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    • 1994
  • Four samples of commercially manufactured yogurts (plain, drinking type) were purchased and evaluated their physico-chemical properties, buffering capacity. And the survival rate of lactic acid bacteria and their ${\beta}-galactosidase$ activity under the acidic conditions (in vitro) were investigated. The values of pH, titratable acidity, viscosity and viable cell counts of yogurts were $3.71{\sim}4.08$, $0.990{\sim}1.045%$, $256{\sim}3164\;cps.$ and $10^8{\sim}10^9\;cfu/ml$, respectively. The volume of 1.0 M-HCl required to reduce the pH of yogurt (50 ml) to minus 2 value was $3.58{\sim}4.33\;ml$. When commercial yogurts were incubated at $37^{\circ}C$ for 120 minutes under the acidic conditions (pH 3.5, 2.5, 1.5), the survival rates of lactic acid bacteria in yogurt were $3.5{\times}10^{-2}{\sim}3.6{\times}10^{-1}%$ at pH 2.5, $8.3{\times}10^{-5}{\sim}4.2{\times}10^{-3}%$ at pH 1.5, respectively, but there was no significant difference at pH 3.5. The remaining activities of ${\beta}-galactosidase$ were $9.4{\sim}36.2%$ at pH 2.5, $4.2{\sim}19.0%$ at pH 1.5, respectively. These results suggested that a significant number of lactic acid bacteria in yogurt might be destroyed in the hostile environment of the stomach, but ${\beta}-galactosidase$ activity from yogurt might be somewhat maintained probably due to the protecting effect by its cell wall and membrane.

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Effect of Varieties and Seeding Date on Over Winter and Dry Matter Yield of Italian Ryegrass in Paddy Field (논에서 이탈리안 라이그라스 품종별 파종시기가 월동 및 건물수량에 미치는 영향)

  • Kim, Meing-Jooung;Choi, Ki-Jun;Kim, Jong-Geun;Seo, Sung;Yoon, Sei-Hyung;Lim, Young-Chul;Im, Seok-Ki;Kwon, Eung-Gi;Chang, Sun-Sik;Kim, Hyeong-Cheol;Kim, Tae-Il
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.29 no.4
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    • pp.321-328
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    • 2009
  • This experiment was carried out to investigate the effect of cultivar and seeding date on the winter survival rate, quality and DM yield of Italian ryegrass on paddy field for three years in Suwon. Seeding started from 30th Sep. 2003. at intervals of five days and finished 20th Oct. New varieties of Italian ryegrass used in this experiment were "Kospeed", "Kowinmaster" and "Hwasan 101". The winter survival average rate of 5th Oct. seeding plot was 89.8% and it decreased with delayed seeding date. The heading date of "Kospeed" was 7th~13rd May, "Kowinmaster" was 16th May, but "Hwasan 101 ho" didn't show heading until 17th May. Dry matter (DM) yields of 30th Sep. seeding plot were Kospeed 7,909kg/ha, Kowinmaster 6,398 kg/ha and Hwasan 101 ho 5,204 kg/ha. DM yield was decreased with delayed seeding date. Total digestible nutrient (TDN) yield was also decreased with delayed seeding date. Crude protein (CP) content was the highest in Kospeed. seeding plot as 18.3% and in vitro dry matter digestibility (IVDMD) was not showed significant difference among seeding dates.

Effect of $CO_2$ Concentration, NAEH and Light Intensity on the Photoautotrophic Growth of Campanula punctata 'Rubriflora' Plantlets In Vitro (자주초롱꽃의 기내 자가영양배양시 $CO_2$농도, 환기횟수 및 광도가 생장에 미치는 영향)

  • Shim, Jae-Nam;Kim, Gyeong-Hee;Jeong, Byoung-Ryong
    • Journal of Bio-Environment Control
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    • v.14 no.4
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    • pp.233-238
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    • 2005
  • Growth of Campanula punctata 'Rubriflora' plantlets, as affected by three levels of photosynthetic photon flux (PPF), 70, 110, and $220{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}$, two levels of $CO_2$ concentration, 500 and $1,500{\mu}mol{\cdot}m^{-1}$, and two levels of number of air exchanges per hour (NAEH), 0.1 and $2.8 h^{-l}$, was studied. Explants were obtained from photomixotrophically-micropropagated plantlets. Four explants were planted in each $3.7{\times}10^{-4}m^3$ polycarbonate box containing MS basal medium and no added sucrose. Explants were cultured under cool-white fluorescent lamps for $16h{\cdot}d^{-1},\;at\;25\pm1^{\circ}C$ temperature, and $70\~80\%$ relative humidity In treatments of $2.8h^{-1}$ NAEH, a 10mm round hole made on the vessel cap was sealed with a microporous filter. For higher $CO_2$ concentrations in the culture room, $CO_2$ gas was provided from a tank of liquefied $CO_2$. Fresh and dry weights, length of the longest root, and number of leaves significantly increased with increasing PPF and especially $CO_2$ concentration. Length of the longest root, number of leaves, fresh and dry weights, and chlorophyll concentration were enhanced with increased NAEH. However, leaf area was the smallest in the $220{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}\;PPF\;2.8h^{-1}$ NAEH and especially, $1,500{\mu}mol{\cdot}mol^{-1}\;CO_2$ concentration treatment. Treatment effect became more produced with time. Overall, treatment with $220{\mu}mol{\cdot}m^{-2}{\cdot}s^{-1}\;PPF\;and\;1,500{\mu}mol{\cdot}mol^{-1}\;CO_2$ gave the most vigorous growth.

Hypoglycemic Effects of Fruits and Vegetables in Hyperglycemic Rats for Prevention of Type-2 Diabetes (고혈압쥐의 과일과 야채의 섭취에 따른 저혈당 효과)

  • Survay, Nazneen Shaik;Ko, Eun-Young;Upadhyay, Chandrama Prakash;Jang, Mi;Park, Se-Won;Lee, Dong-Ha;Jung, Yi-Sook;Yoon, Do-Young;Hong, Sae-Jin
    • Horticultural Science & Technology
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    • v.28 no.5
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    • pp.850-856
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    • 2010
  • An in vivo oral glucose tolerance test (OGTT) was performed on hyperglycemic male Sprague-Dawley rats to assess the effect of fruits and vegetables ($1g{\cdot}kg^{-1}$ body weight) on blood glucose levels (${\Delta}BGLs$) at different time intervals of 0, 5, 15, 30, 60, 90 and 120 min. The areas under glucose curve (${\Delta}AUCs$) were calculated at 120 min of OGTT by trapezoid method. Total phenolic content (TPC) and anti-oxidant activity (AOA) of fruits and vegetables were assayed in vitro by Folin Ciocalteu and DPPH (2, 2-diphenyl-1-picrylhydrazyl) methods, respectively. At the end of the experiment the correlations among the parameters TPC, AOA and ${\Delta}AUC$ was estimated by Pearson's correlations. Among fruit crops, tangerine, plum, grape and pear and among vegetables, blue leaf mustard, cabbage, chicory, broccoli and others exhibited significant hypoglycemic effects by reducing ${\Delta}BGLs$ with significant ${\Delta}AUC$. The effective ${\Delta}AUC$ ranged from $5548.2{\pm}462.1$ to $3823.3{\pm}282.0mg-min{\cdot}dL^{-1}$. The TPC and AOA ranged from $0.063{\pm}0.00$ to $0.913{\pm}0.14mg{\cdot}g^{-1}$ GAE and $01.05{\pm}0.08$ to $75.46{\pm}0.06%$, respectively. Overall, six fruits and fifteen vegetables exhibited higher TPC and one fruit and four vegetables exhibited higher AOA. There was a better correlation among TPC, AOA and ${\Delta}AUC$ of fruits and TPC & AOA of vegetables. We report that hypoglycemically significant fruits and vegetables investigated in this study have pharmacological importance which reduced ${\Delta}BGLs$ through insulin like activity and AOA in prevention of type-2 diabetes.

Enhancement of the Thermostability of a Fibrinolytic Enzyme from Bacillus amyloliquefaciens CH51 (Bacillus amyloliquefaciens CH51이 생산하는 혈전용해효소의 열안정성 개선)

  • Kim, Jieun;Choi, Kyoung-Hwa;Kim, Jeong Hwan;Song, Young-Sun;Cha, Jaeho
    • Journal of Life Science
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    • v.23 no.1
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    • pp.15-23
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    • 2013
  • AprE51 from Bacillus amyloliquefaciens CH51 is a 27 kDa subtilisin-like protease with fibrinolytic activity. AprE51-6 showing increased catalytic activity was produced previously. To enhance the thermostability of AprE51-6, 2 residues, Gly-166 and Asn-218 based on B. subtilis subtilisin E were mutated by site-directed mutagenesis. The results of the mutational analysis showed that substitution of arginine for Gly-166 (AprE51-7) increased the fibrinolytic activity 1.8-fold. An N218S mutant (AprE51-8) also increased the fibrinolytic activity up to 4.5-fold in a fibrin plate assay. Purified AprE51-7 and AprE51-8 mutants had a 1.9- and a 2.5-fold higher $k_{cat}$, respectively, and a 2.1-1.9-fold lower $K_m$, respectively. This resulted in a 3.8- and a 4.7-fold increase in catalytic efficiency ($k_{cat}/K_m$), respectively, relative to that of wild-type AprE51. AprE51-8 had a broader pH range than AprE51-6 and nattokinase, especially at an alkaline pH value. In addition, AprE51-8 showed higher thermostability than AprE51-6 at $60^{\circ}C$. The half-lives of AprE51-7 and AprE51-8 at $50^{\circ}C$ were 21.5 and 27.3 min, respectively, which are 2.0 and 2.6 times longer, respectively, than that of the wild-type AprE51.

Removal of Dissolved Organic Matter by Ozone-biological Activated Carbon process (오존처리와 생물활성탄 공정에 의한 상수원수 중의 용존유기물 제거)

  • 이상훈;문순식;신종철;최광근;심상준;박대원;이진원
    • KSBB Journal
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    • v.18 no.3
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    • pp.211-216
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    • 2003
  • The removal yield of dissolved organic matter in drinking water by biological activated carbon (BAC) process was investigated. The tested processes wer raw water-AC process (BAC1), raw water-ozonation-BAC process (BAC2), and raw water-ozonation-coagulation/sedimentation-BAC process (BAC3). The amounts of organic matter was measured as dissolved organic carbon (DOC), ulta-violet radiation at 254 nm wavelength ($UV_{254}$), total nitrogen (T-N), ammonia nitrogen (NH_3$-N), and total phosphate (T-P). As a results, 30.7% DOC was removed by BAC2 process, which showed higher removal efficiency than BAC1 or BAC3 processes. The removal yield of $UV_{254}$ in BAC1, BAC2, and BAC3 processes were observed as 45.3%, 44.6%, 58.4%, respectively. And the removal yield of ammonia nitrogen were 66%, 81%, 29% in each BAC processes. The optimal empty bed contact time (EBCT) of BAC processes was estimated as 10 minute. This study has shown that BAC process combined with ozone treatment was efficient for removing dissolved organic matter in water.

EFFECT OF HYDROGEN PEROXIDE CONCENTRATION ON THE WHITENING AND PHYSICAL PROPERTIES OF HYDROXYAPATITE DISCS (Hydrogen Peroxide 농도와 적용시간이 Hydroxyapatite Discs의 미백과 물리적 성질에 미치는 영향)

  • Yang, Yeon-Mi;Lee, Doo-Cheol;Baik, Byeong-Ju;Kim, Jae-Gon;Shin, Jeong-Geun
    • Journal of the korean academy of Pediatric Dentistry
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    • v.34 no.1
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    • pp.1-12
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    • 2007
  • The purpose of this study was to evaluate the effect that various concentration and application time of hydrogen peroxide had on tooth whitening and physical properties. The hydroxyapatite (HA) discs of $12mm({\Phi}){\times}1.2mm(t)$ in dimensions were made by compression $(100kg/cm^2)$ and sintering (at $1350^{\circ}C$ for 2 hours) All specimens were polished sequentially with '240 through '2000 emery paper and one side of each specimen was polished finally with $0.3{\mu}m$ alumina paste. The discs were placed in sterile whole stimulated saliva overnight at $37^{\circ}C$ in order to form an in vitro pellicle layer. Then the discs were rinsed with distilled water and soaked into staining broth at $37^{\circ}C$ for 7 days. These stained specimens were bleached with hydrogen peroxide according to the change of concentration $(3{\sim}30%)$ and application time ($3{\sim}10$ days). The specimens were analyzed with a spectrophotometer, X-ray diffractometer (XRD), scanning electron microscope (SEM), surface roughness tester, microhardness tester and biaxial flexural strength. The results of present study can be summarized as follows : 1. The bleaching effect was increased with the increased concentration and the extended application time of hydrogen peroxide. 2. The surface roughness was significantly increased from the specimen bleached with 15% hydrogen peroxide for 10 days and with 30% for 7 and 10 days respectively (p<0.05). 3. The changes of crystal phase observed by XRD between before and after bleaching weren't shown of any difference, but microporous structure of surface observed by SEM was shown of increase with the increased concentration and the extended application. 4. The biaxial flexural strength was significantly decreased from bleaching of specimen with 30% hydrogen peroxide for 7 and 10 days respectively (p<0.05) 5. Microhardness was significantly decreased from bleaching with 15% hydrogen peroxide for 10 days and with 30% for 3, 7 and 10 days respectively (p<0.05). Although the tooth bleaching effect was greater when the high concentration was applied, further in vivo experiment will be needed to prove it's safety.

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Effect of Dexamethasone on Gene Expression of Surfactant Protein B and Surfactant Protein C (스테로이드제가 백서 폐의 Surfactant B와 C 유전자 발현에 미치는 영향)

  • Park, Ik Soo;Sohn, Jang Won;Yoon, Ho Joo;Shin, Dong Ho;Park, Sung Soo
    • Tuberculosis and Respiratory Diseases
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    • v.54 no.4
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    • pp.439-448
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    • 2003
  • Background : Surfactant protein B(SP-B) and surfactant protein C(SP-C) are important in accelerating surface spreading of surfactant phospholipid. The glucocorticoids accelerate the morphologic differentiation of epithelial cells into type II cells and increase the rate of phosphatidylcholine synthesis. The hydrophobic surfactant protein has been shown to be upregulated by glucocorticoids in vitro, however, its regulation in vivo is not well established. Methods : The authors investigated the effects of glucocorticoid on the accumulation of mRNA encoding SP-B and SP-C protein content of the lung. Adult rats were given different doses of subcutaneous dexamethasone and sacrificed at 24 hours and 1 week. SP-B and SP-C mRNA were measured by a filter hybridization method. Results : 1) The accumulation of SP-B mRNA at 24 hours after 0.2 mg/kg dexamethasone treatment was increased by 23.7%. 2) The accumulation of SP-B mRNA at 1 week after 2 mg/kg dexamethasone treatment was significantly increased by 96.6%(P<0.001). 3) The accumulation of SP-C mRNA at 24 hours after 0.2 mg/kg dexamethasone treatment was significantly increased by 42.7%(P<0.01). 4) The accumulation of SP-C mRNA at 1 week after 2 mg/kg dexamethasone treatment was significantly increased by 60.0% (P<0.01). Conclusion : The authors concluded that dexamethasone treatment in vivo resulted in increased levels of SP-B mRNA and SP-C mRNA. These results suggested that dexamethasone stimulates the synthesis of hydrophobic proteins associated with surfactant.