• Proceedings of the Korean Society of Toxicology Conference
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• 2002.05b
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• pp.106.2-132
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• 2002

형질전환 (유전자 결핍; Knockout) Min 마우스를 이용하여 대장암 발생에서 배추, 양배추 주성분인 indole-3-carbinol (I3C)의 대장암 예방효과를 규명하고자 하였다. 실험동물로는 C57BL/6J-Apc$^{min/+}$(Min 마우스)계의 5내지 6주령의 수컷 이형접합체 형질전환 마우스 70마리와 C57BL/6J계의 동일 산자, 동일 주령의 수컷 wildtype 비형질전환 마우스 10kfl를 The Jackson Laboratory 사 (Bar harber, ME, USA)로부터 직접 구입하였다. C57BL/6J-Apc$^{min/+}$계 수컷 이형접합체 형질전환 (Min)마우스 70마리를 각 군 20내지 25마리씩 세군으로 나누었다. Group 1에는 20마리, Group 2에는 25마리, Group 3에는 25마리를 배치하고, I3C 투여 실험군 (Group 1과 2)에는 실험시작시에 AIN-76A 분말사료에 I3C가 각각 100 및 300ppm이 함유되도록 조제하여 공급하였다. 그리고 실험군(Group 3)에는 실험시작부터 종료시까지 AIN-76A 정제고형사료(Teklad사, WI, USA)를 자유로이 급이하였다. 각군간의 체중, 사료 및 음수소비량을 매 2주마다 측정하였고, 10주간 (16주령)의 실험종료시에는 최종체중과 간장, 신장, 비장 등의 장기무게를 측정하여 상대장기 무게비를 산출하였다. 대조군으로서 C57BL/6J계의 동일 산자, 동일 주령의 수컷 wildtype 비형질전환 마우스 10마리는 같은 조건의 사육실에서 AIN-76A 정제고형사료를 33주간 자유로이 급이하였다. 실험동물은 부검전에 하룻밤 동안 절식하고 이산화탄소 흡입 마취하에서 흉대동맥을 절단하여 방혈하고 각 장기(심장, 폐, 위)를 적출하여 생리심염수에 넣어 장기무게를 측정하고 포르말린에 고정하였다. 소장과 대장의 검사를 위하여 위의 식도부위와 직장을 실로 결찰하여 적출하고 생리심염수를 주입하여 팽창시켜, 십이지장, 공장, 및 회장, 그리고 대장으로 나누어 여과지에 펼친 후 포르말린에 고정하였다. 소장과 대장은 육안 및 자동 영상분석길ㄹ 이용한 분석이 끝난 후에 각 부위별로 4-6개의 절편을 작제하여 포르말린에 재고정하고, 통상적인 조직처리과정, 파리핀 포매 및 3-4$\mu$m 두께의 조직절편을 제작하여 H&E 염색을 실시하여 현미경으로 검경하였다. 약 1주일간의 포르말린 고정이 끝난 소장 및 대장을 부위별, 별 종양개수 및 분포를 자동영상분석기(Kontron Co. Ltd., Germany)로 분석하였다. 체의 변화, 장기무게, 사료소비량 및 마리당 종양의 개수에 대한 통계학적 유의성 검증을 위하여 Duncan's t-test로 통계처리 하였고, 종양 발생빈도에 대하여는 Likelihood ration Chi-square test로 유의성을 검증하였다. C57BL/6J-Apc$^{min/+}$계 수컷 이형접합체 형질전환 마우스에 AIN-76A 정제사료만을 투여한 대조군의 대장선종의 발생률은 84%(Group 3; 21/25례)로써 I3C 100ppm 및 300ppm을 투여한 경우에 있어서는 각군 모두 60%(Group 1; 12/20 례, Group 2; 15/25 례)로 감소하는 경향을 나타내었다. 대장선종의 마리당 발생개수에 있어서는 C57BL/6J-Apc$^{min/+}$계 수컷 이형접합체 형질전환 마우스에 AIN-76A 정제사료만을 투여한 대조군은 1.40$\pm$0.24(100%)에 비하여 I3C 저농도 투여 실험군(Group 1; 0.85$\pm$0.23; 61%, P<0.01), 그리고 I3C 고농도 투여 실험군(Group 2 ; 1.32$\pm$0.29 ; 94%)의 순으로 감소하였다. 선종의 크기별 종양의 발생개수의 분포에 있어서 I3C 저농도 투여 실험군에 있어서는 선종의 크기가 3mm이하의 수가 현저하게 감소하였다. C57BL/6J-Apc$^{min/+}$계 수컷 이형접합체 형질전환 마우스에 AIN-76A 정제사료만을 투여한 대조군의 부위별 소장선종의 발생수는 십이지장부위를 제외하고 각 군에서 유의한 변화는 관찰되지 않았다. 십이지장 종양의 발생개수에서만 I3C 저농도 투여 실험군(Group 1 ; 3.11$\pm$0.85)이 대조군 (Group 3: 1.48$\pm$0.35) 및 I3C 고농도 투여 실험군(Group 2: 1.56$\pm$0.47)에 비하여 유의성 있게 증가하였다. (P<0.05). 따라서 I3C은 소장에서는 암예방 효과가 뚜렷하지 않으나, 대장에 대한 암에방 효과가 있을 것으로 생각된다. 소장 및 대장을 제외한 간장, 신장, 비장, 심장, 폐 그리고 위 등의 기타 장기에서의 조직병리학적 변화는 관찰되지 않았다. 소장 및 대장의 종양은 선종(polyps)으로 관찰되었다. 지난 10여년간 형질전환 및 유전자 결핍 실험동물의 종류가 기하 급수적으로 증가하여 이용되고 있다. 가족성 대장 선종성 용종증(FAP)의 대표적인 모델로 이용되고 있는 C57BL/6J-Apc$^{min/+}$계 수컷 이형접합체 형질전환 마우스를 사용하여 배추나 양배추의 주요성분인 Indole-3-carbinol(I3C)의 대장암 예방효과가 있는지를 검색하여 본 결과 AIN-76A정제사료만을 투여한 대조군의 대장선종의 발생률 84%에 비하여 I3C 100 및 300ppm을 투여한 실험군에 있어서 각군 모두 60%로서 감소하는 경향을 나타내었으며, 대장선종의 마리당 발생개수에 있어서는 대조군의 1.40$\pm$1.041를 100%로 환산하였을 경우 I3C 저농도 및 고농도 투여 실험군에서는 각각 약 61%와 94%를 나타내여 감소하였다. 특히 대장선종의 크기별 분포에 있어서 선종의 크기가 3mm이하의 수가 현저하게 감소하였다. 따라서 저농도 I3C의 투여는 실험적 유전성 가족성 대장 선종성 용종증 모델에 있어서 어느정도 암 예방효과가 있는 것으로 생각된다. 그러나 소장 선종의 발생에는 별 영향이 없는 것으로 생각된다. 그러나 본 실험에 사용된 C57BL/6J-Apc$^{min/+}$계 수컷 이형접합체 형질전환 마우스는 실험개시 시점이 7내지 8주령이 경과하여 이미 태생기부터 소장 및 대장의 선종 발생이 진행되어 온 것을 감안하고 특히 비스테로이드계 항염증 소염제(NSAIDS)와 같은 강력한 COX-2억제제가 아님을 고려하면, 상당한 선종의 발생을 억제할 수 있는 가능성이 매우 높다고 생각한다. 또한 이제까지 배추나 양배추 성분의 복합성분들에 대한 실험적 대장암 모델에서의 촉진효과 등에 대한 보고들이 있어 온 점을 고려할 때 위암(Kim 등, 1994) 간암(Kim 등, 1994), 유방암(Grubbs, 등, 1995; Bradlow 등, 1995)에 대한 예방효과가 있을 것으로 생각된다. 앞으로 이러한 종양조직내에서의 COX-2 및 iNOS mRNA와 단백질의 발현정도를 분자병리학적으로 연구중에 있으며, 향후 십자화과식물 성분인 indole-3-carbinol이 마우스뿐 만 아니라 랫드의 화학발암물질에 의한 대장종양에 대한 억제효과 있는지 연구 필요가 있다. Min 마우스와 같은 형질 전환(유전자결핍;knockout) 실험동물을 이용한 새로운 중기 발암성 시험범의 확립을 통한 각종 환경 유해물질의 발암성 유무 및 COX-2 억제작용이 있는 식품인자의 암예방 후보물질을 체계적으로 검색하는데 유용하게 활용될 수 있을 것으로 생각한다.

• Maxillofacial Plastic and Reconstructive Surgery
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• v.31 no.3
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• pp.243-248
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• 2009

Purpose: Rehabilitation of the edentulous posterior maxilla with dental implants often poses difficulty because of insufficient bone volume caused by pneumatization of the maxillary sinus and by crestal bone resorption. Sinus grafting technique was developed to increase the vertical height to overcome this problem. The present study was designed to evaluate the sinus floor augmentation with anorganic bovine bone (Bio-$cera^{TM}$) using histomorphometric and clinical measures. Patients and methods: Thirteen patients were involved in this study and underwent total 14 sinus lift procedures. Residual bone height was ${\geq}2mm$ and ${\leq}6mm$. Lateral window approach was used, with grafting using Bio-$cera^{TM}$ only(n=1) or mixed with autogenous bone from ramus and/or maxillary tuberosity(n=13). After 6 months of healing, implant sites were created with 3mm diameter trephine and biopsies taken for histomorphometric analysis. The parameters assessed were area fraction of new bone, graft material and connective tissue. Immediate and 6 months after grafting surgery, and 6 months after implantation, computed tomography (CT) was taken and the sinus graft was evaluated morphometric analysis. After implant installation at the grafted area, the clinical outcome was checked. Results: Histomorphometry was done in ten patients.Bio-$cera^{TM}$ particles were surrounded by newly formed bone. The graft particles and newly formed bone were surrounded by connective tissue including small capillaries in some fields. Imaging processing revealed $24.86{\pm}7.59%$ of new bone, $38.20{\pm}13.19%$ connective tissue, and $36.92{\pm}14.51%$ of remaining Bio-$cera^{TM}$ particles. All grafted sites received an implant, and in all cases sufficient bone height was achieved to install implants. The increase in ridge height was about $15.9{\pm}1.8mm$ immediately after operation (from 13mm to 19mm). After 6 months operation, ridge height was reduced about $11.5{\pm}13.5%$. After implant installation, average marginal bone loss after 6 months was $0.3{\pm}0.15mm$. Conclusion: Bio-$cera^{TM}$ showed new bone formation similar with Bio-$Oss^{(R)}$ histomorphometrically and appeared to be an effective bone substitute in maxillary sinus augmentation procedure with the residual bone height from 2 to 6mm.

• Journal of Life Science
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• v.24 no.5
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• pp.505-514
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• 2014

To examine the anti-obese activity of miscellaneous cereal grains, 80% ethanol extracts from eight selected miscellaneous cereal grains were compared for their cytotoxic effects on 3T3-L1 murine preadipocytes. The ethanol extract of proso millet exhibited the highest cytotoxicity. Further fractionation of the ethanol extract with methylene chloride, ethyl acetate, and n-butanol showed that the cytotoxicity of the ethanol extract was mainly partitioned into the butanol fraction. As compared with differentiated mature adipocytes, 3T3-L1 preadipocytes were more susceptible to the cyctotoxicity of the butanol fraction. When each organic solvent fraction (25 ${\mu}g/ml$) was added during the differentiation period for 6 days, the cell viability was not affected significantly except for the butanol fraction, but the intracellular lipid accumulation declined to a level of 81.5%~50.3% of the control. The Oil Red O staining data also demonstrated that the ethanol extract as well as the butanol fraction could inhibit the differentiation of 3T3-L1 preadipocytes into mature adipocytes. The presence of the butanol extract during the induced adipocytic differentiation also resulted in a significant reduction in the expression levels of critical adipogenesis mediators $(C/EBP{\alpha}$, $PPAR{\gamma}$, aP2, and LPL) to a barely detectable or undetectable level and the cells retained the fibroblast-like morphology of 3T3-L1. In 3T3-L1 cells, the cytotoxicity of the butanol fraction (50-100 ${\mu}g/ml$) was accompanied by mitochondrial membrane potential (${\Delta}{\psi}m$) loss, caspase-3 activation, and PARP degradation. Taken together, these results indicate that proso millet grains possess pro-apoptotic and anti-adipocytic activities toward adipocytes, which can be applicable to prevention of obesity.

• Journal of The Korean Society of Grassland and Forage Science
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• v.30 no.2
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• pp.179-190
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• 2010

The experiment was conducted to observe the effects of dried whole crop barley treated with cellulolytic microorganisms (Aspergillus niger KCCM 60357 and Bacillus licheniformis KCCM 40934) on the chemical composition, in vitro colonic fermentation and whole tract digestibility in swine. Whole crop barley were fermented with no microorganism addition (control), A. niger, B. licheniformis and co-culture of A. niger and B. licheniformis (Mixture) for 3 days at $30^{\circ}C$. In the feed chemical composition, CP contents of whole crop barley treated with A. niger (7.52%) and B. licheniformis (7.77%) were significantly higher than control (6.81%) (p<0.05). The in vitro colonic fermentation of dried whole crop barley fermented with control showed significantly higher $CH_4$ contents than A. niger, B. licheniformis and Mixture at 18h incubation (p<0.05). Dry matter (DM) digestibilities of A. niger (55%) and Mixture (57.42%) treatments were significantly higher than control (43.74%) (p<0.05). Ammonia-N was significantly increased in A. niger, B. licheniformis and Mixture relative to control at 24 hour incubation (p<0.05). Xylanase activities in A. niger, B. licheniformis and Mixture treatments were significantly higher than control at 24 hour incubation (p<0.05). Concentrations of total VFA were significantly increased in B. licheniformis (12.61 mM) at 24hour incubation (p<0.05). In vitro whole tract digestibility was significantly increased in B. licheniformis (49.61%) compared with the control (45.65%) (p<0.05). In conclusion, whole crop barley treated with cellulolytic microorganisms improved whole tract digestibility and colonic fermentation for swine.

• Journal of Physiology & Pathology in Korean Medicine
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• v.16 no.4
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• pp.724-728
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• 2002

Angiotensin-converting enzyme (ACE) gene polymorphism, which consists of presence (insertion, I) or absence (deletion, D) of a 250-bp fragment, is associated with ischemic heart disease, renovascular disease, systemic lupus erythematosus. Subjects with the DD genotype have higher levels of circulating ACE than subjects with the II genotype and show an increased tendency towards vascular wall thickness and contribute to the development of vascular disease. But the association between I/D polymorphism of the ACE gene and cerebrovascular disease is still controversial. The aim of this study was to determine whether the DNA polymorphism of the ACE are associated with cerebrovascular disease in Korean population. The study group comprised 377 Korean patients admitted to Kyunghee Oriental Medical Center in the year of 2000 for the treatment of brain infarction or brain hemorrhage. Magnetic resonance imaging(MRI) was performed for each patient to determine the stroke phenotype, infarction or hemorrhage. The 183 subjects without evidence of brain infarction or brain hemorrhage were selected from the some ethnical population(control group). Venous blood samples were drawn from each subject for the extraction of DNA. Genotypes of ACE were determined by polymerase chain reaction amplification of the genomic DNA. Case and control genotype frequencies were compared by chi-square testing. Both the patients and the controls were classified respectively into 4 groups: age less than forty years, age forty one to fifty, age fifty one to sixty, age greater than sixty years. There were no significant differences in the distributions of ACE genotypes among the patients with infarction, with hemorrhage and controls (Infarction: D/D 15.8%, I/D 46.7%, I/I 37.5%, Hemorrhage: D/D 15.1%, I/D 46.5%, I/I 38.4%, Control: D/D 18.6%, I/D 50.3%, I/I 31.2%). There was a significant difference in the distribution of ACE genotypes between the age greater than sixty year subgroup of patient with brain hemorrhage and the control (Hemorrhage: D/D 0%, I/D 55.6%, I/I 44.4%, Control: D/D 13.0%, I/D 63.0%, I/I 23.9%; Pearson Chi-Square value 5.956, P<0.05). Furthermore, the frequency of the ACE D/D type declined with increasing age both in the patient and control group (Patient group: age < 50 D/D 21.5%, age > 50 D/D 14.42%; Control group: age < 50 D/D 21.0%, age > 50 D/D 14.2%). In conclusion there is no clear association between ACE polymorphism and cerebrovascular disease in Korean population. Although, there was a tendency for the frequency of the ACE D/D type declined with increasing age in both patients and controls.

• The Journal of Korean Society for Radiation Therapy
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• v.21 no.2
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• pp.67-74
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• 2009

Purpose: To evaluate the results of absorbed and effective doses using two different modes, standard mode (A-mode) and low-dose mode (B-mode) settings for prostate cancer IGRT from CBCT. Materials and Methods: This experimental study was obtained using Clinac iX integrated with On Board Imager (OBI) System and CBCT. CT images were obtained using a GE Light Speed scanner. Absorbed dose to organs from ICRP recommendations and effective doses to body was performed using A-mode and B-mode CBCT. Measurements were performed using a Anderson rando phantom with TLD-100 (Thermoluminescent dosimeters). TLD-100 were widely used to estimate absorbed dose and effective dose from CBCT with TLD System 4000 HAWSHAW. TLD-100 were calibrated to know sensitivity values using photon beam. The measurements were repeated three times for prostate center. Then, Evaluations of effective dose and absorbed dose were performed among the A-mode and B-mode CBCT. Results: The prostate absorbed dose from A-mode and B mode CBCT were 5.5 cGy 1.1 cGy per scan. Respectively Effective doses to body from A mode and B-mode CBCT were 19.1 mSv, 4.4 mSv per scan. Effective dose from A-mode CBCT were approximately 4 times lower than B-mode CBCT. Conclusion: We have shown that it is possible to reduce the effective dose considerably by low dose mode(B-mode) or lower mAs CBCT settings for prostate cancer IGRT. Therefore, we should try to select B-mode or low condition setting to decrease extra patient dose during the IGRT for prostate cancer as possible.

• Journal of Korean Society of Environmental Engineers
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• v.28 no.8
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• pp.866-871
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• 2006

The nutrient recovery in phosphate crystallization process was investigated by using laboratory scale uptlow reactors, adopting sequencing batch type configuration. The industrial waste lime was used as potential cation source with magnesium salt($MgCl_2$) as control. The research was focused on its successful application in a novel integrated sludge treatment process, which is comprised of a high performance fermenter followed by a crystallization reactor. In the struvite precipitation test using synthetic wastewater first, which has the similar characteristics with the real fermentation effluent, the considerable nutrient removal(about 60%) in both ammonia and phosphate was observed within $0.5{\sim}1$ hr of retention time. The results also revealed that a minor amount(<5%) of ammonia stripping naturally occurred due to the alkaline(pH 9) characteristic in feed substrate. Stripping of $CO_2$ by air did not increase the struvite precipitation rate but it led to increased ammonia removal. In the second experiment using the fermentation effluent, the optimal dosage of magnesium salt for struvite precipitation was 0.86 g Mg $g^{-1}$ P, similar to the mass ratio of the struvite. The optimal dosage of waste lime was 0.3 g $L^{-1}$, resulting in 80% of $NH_4-N$ and 41% of $PO_4-P$ removal, at about 3 hrs of retention time. In the microscopic analysis, amorphous crystals were mainly observed in the settled solids with waste lime but prism-like crystals were observed with magnesium salt. Based on mass balance analysis for an integrated sludge treatment process(fermenter followed by crystallization reactor) for full-scale application(treatment capacity Q=158,880 $m^3\;d^{-1}$), nutrient recycle loading from the crystallization reactor effluent to the main liquid stream would be significantly reduced(0.13 g N and 0.19 g P per $m^3$ of wastewater, respectively). The results of the experiment reveal therefore that the reuse of waste lime, already an industrial waste, in a nutrient recovery system has various advantages such as higher economical benefits and sustainable treatment of the industrial waste.

• The Korean Journal of Pesticide Science
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• v.21 no.1
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• pp.97-105
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• 2017

This study was carried out to determine residual characteristics and to evaluate safety of bifenthrin, carbendazim and metconazole in minor crop, Angelica gigas Nakai. The test pesticides were sprayed onto the crop according to the combinations of spray days before harvest and frequency and then sampling was performed at the harvest day. Non-systemic pesticide bifenthrin was not detected in all samples, whereas systemic pesticides, carbendazim and metconazole, were detected from the crop in the range of from 0.14 to 0.49 and from 0.19 to 0.26, respectively. These results indicated that residual characteristics of the test pesticides in Angelica gigas Nakai were affected by their systemic properties. Also, residual concentration of carbendazim and metconazole in the crop was increased as more spraying frequency and spraying more closer to harvest day. And residual concentration of carbendazim in the crop on the last spraying day was 2.1 times higher than that of metconazole because nominal spraying dose of carbendazim was higher than that of metconazole by 1.9 times. The residue levels of the test pesticides in the crop were evaluated to be safe, considering their %ADIs were less than 0.9%.

• Journal of Chest Surgery
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• v.31 no.12
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• pp.1147-1158
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• 1998

Background: Portable cardiopulmonary bypass(CPB) technique has been used increasingly as a potent and effective option for emergency cardiopulmonary resuscitation(CPR) because it can maintain more stable hemodynamics and provide better survival than conventional CPR techniques. This study was designed to develop a prototype of Korean portable CPB system and, by applying it to CPR, to discriminate whether it would be superior to standard open-chest CPR. Material and Method: By using adult mongrel dogs, open-chest CPR(OCPR group, n=4) and portable-CPB CPR(CPB group, n=4) were compared with respects to restoration of spontaneous circulation(ROSC), hemodynamics, effects on blood cells, blood gas patterns, biochemical markers, and survivals. Ventricular fibrillation-cardiac arrest(VF-CA) of arrest(VF-CA) of 4 minutes followed by basic life support(BLS) of 15 minutes was applied in either group, which was standardized by the protocol of American Heart Association. Then, advanced life support(ALS) was applied to either group under the support of internal cardiac massage or CPB. ALS was maintained until ROSC was achieved but not longer than 30 minutes regardless of the presence of ROSC. All of the measured values were expressed as means±SD percent change from baseline. Result: During the early ALS, higher mean arterial pressure was maintained in CPB group than in OCPR group(90±19 vs. 71±32 %; p<.05) and lower mean pulmonary arterial pressure was also maintained in CPB group than in OCPR group(105±24 vs. 146±6%; p<.05). ROSC was achieved in all dogs. Post-ROSC levels of hematocrit, RBC, and platelet were decreased and plasma free hemoglobin was increased significantly in CPB group compared to OCPR group(p<.05). Changes in blood gas patterns, lactate, and CK-MB levels were not different between groups. Early mortality was seen in 3 dogs in OCPR group(survival time 31±36 hours) and 2 in CPB group(228±153 hours, p=ns). The remainders in both groups showed prolonged survival. Conclusion: These findings indicate that portable CPB can be effective to maintain stable hemodynamics during cardiac arrest, to achieve ROSC and to prolong survival. Further study is needed to refine the portable CPB system and to meet clinical challenges.

• Food Science and Preservation
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• v.18 no.2
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• pp.236-243
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• 2011

We evaluated the nutritional value of 70% ethanol extracts (EE) from fermented cudrania tricuspidata fruit (ECT: EE of C. tricuspidata, EFCT: EE of fermented C. tricuspidata, EFCTL: EE of fermented C. tricuspidata by B. licheniformis, EFCTS: EE of fermented C. tricuspidata by B. subtilis) by determined the total polyphenol and flavonoid contents, antioxidant activities, and inhibitory actions on elastase and tyrosinase. The yields of freeze-dried powder of ECT, EFCT, EFCTL, and EFCTS were 54.22%, 54.43%, 57.51%, and 57.23% (each w/w), respectively. The color of $L^*,\;a^*,\;b^*$ values decreased of fermented cudrania tricuspidata. The fermented EFCTL and EFCTS had relatively higher contents of total polyphenol and flavonoid than those of other extracts. ABTS radical scavenging ability were 94.61%, 95.85%, 94.36%, and 96.69%, respectively. SOD (superoxide dismutase)-like activities were in the order EFCT (32.60%) > EFCTS (27.10%) > EFCT (23.30%) > ECT (22.00%), and nitrite scavenging activity was in order of EFCTS (51.18%) > EFCTL (45.61%) > EFCT (41.93%) > ECT (27.76%), respectively. Ferrous ion chelating activity of EFCTL (67.34%) and EFCTS (60.36%) was significantly the highest, whereas ECT (52.34%) and EFCT (51.73%) had not different significantly. Xanthine oxidase, elastase and tyrosinase inhibitory activities at 0.5% (w/v) solutions of EFCTL and EFCTS were somewhat higher than those of non-fermented ECT. In conclusion, we provide experimental evidence that extracts of fermented cudrania tricuspidata of B. subtilis and B. licheniformis exhibited higher antioxidant activities and inhibitory actions on elastase and tyrosinase compared with non fermented cudrania tricuspidata.