• 생명과학회지
• /
• 제9권6호
• /
• pp.653-658
• /
• 1999

A novel marine microorganism, Vibrio sp. YE-101, was isolated from pufferfish and investigated for its ability to synthesize tetrodotoxin (TTX). Various strains isolated from the intestine of pufferfish were grown on TCBS agar plate, and then cultured on Ocean Research Institute (ORI) medium supplemented with 3% NaCl at 23$^{\circ}C$ for 3days. The cells were harvested, disrupted, fractionated by Bio-Gel P-2 column chromatography and then TTX-producing strain, Vibrio sp. YE-101, was identified using mouse bioassay. The isolated TTX from Vibrio sp. YE-101 was also analyzed and identified by HPLC and gas chromatography-mass spectrometer (GC-MS). The mass fragmentation of trimethylsilyl derivatives of C9-base of TTX from Vibrio sp. YE-101 was interpreted and the pattern of fragmentation was same with that of authentic standard. The purfied TTX was also positive to the mouse bioassay, which clearly represents that Vibrio sp. YE-101 can synthesize TTX.

• 한국어업기술학회:학술대회논문집
• /
• 한국어업기술학회 1998년도 수산관련공동학술대회발표요지집
• /
• pp.348.2-349
• /
• 1998

• 한국수산과학회지
• /
• 제41권5호
• /
• pp.309-314
• /
• 2008

The toxicity of five species of puffer fish, Arothron firmamentum (Byeolbok), Lagocephalus gloveri (Heukmilbok), L. wheeleri (Eunmilbok), L. inermis (Minmilbok) and L. lunaris (Milbok), collected from fish markets in Korea, was determined using a mouse bioassay. In A. firmamentum, the proportion of toxic specimens containing >10 MU/g was 87.5% in the ovaries, and 10.0% in the skin; no toxicity was detected in the muscle, fin, liver, intestine and gallbladder using the mouse bioassay. The highest toxin levels were found to be 87 MU/g in the ovaries, and 13 MU/g in the skin. Toxic specimens containing >10 MU/g were not detected from samples taken from any of the organs in L. wheeleri and L. inermis. In L. gloveri, most specimens were found to be non-toxic, but toxin levels of 11-72 MU/g were detected from within the skin, fins, and intestines in one specimen. In L. lunaris, the proportion of toxic specimens was 50.0% in the ovaries, and 7.1% in the gallbladder; no toxicity was detected in the other organs by the mouse bioassay. The highest toxin levels were 75 MU/g in the ovaries, and 14 MU/g in the gallbladder. Therefore, the toxicities of edible muscle and skin in the five species of puffer fish marketed in Korea were found to be within acceptable levels for human consumption.

• Fisheries and Aquatic Sciences
• /
• 제16권3호
• /
• pp.159-164
• /
• 2013

The mouse bioassay and high performance liquid chromatography (HPLC) post-column oxidation method are different methods of quantifying paralytic shellfish poisoning toxins. In this study, we compared their ability to accurately quantify the toxicity levels in two types of field sample (oysters and mussels) with different toxin profiles for routine regulatory monitoring. A total of 72 samples were analyzed by both methods, 44 of which gave negative results, with readings under the limit of detection of the mouse bioassay ($40{\mu}g/100g$ saxitoxin [STX] eq). In 14 oysters, the major toxin components were gonyautoxin (GTX) 1, -2, -3, -4, -5, decarbamoylgonyautoxin-2 (dcGTX2), and decarbamoylsaxitoxin (dcSTX), while 14 mussels tested positive for dcSTX, GTX2, -3, -4, -5, dcGTX2, neosaxitoxin (NEO), STX, and dcSTX. When the results obtained by both methods were compared in two matrices, a better correlation ($r^2=0.9478$) was obtained for mussels than for oysters ($r^2=0.8244$). Additional studies are therefore needed in oysters to investigate the differences in the results obtained by both methods. Importantly, some samples with toxin levels around the legal limit gave inconsistent results using HPLC-based techniques, which could have a strong economic impact due to enforced harvest area closure. It should therefore be determined if all paralytic shellfish poisoning toxins can be quantified accurately by HPLC, and if the uncertainties of the method lead to doubts regarding regulatory limits.

• Toxicological Research
• /
• 제21권1호
• /
• pp.1-14
• /
• 2005

Transgenic mouse models have been introduced and accepted by regulatory bodies as an alternative to carcinogenicity assay models to predict and evaluate chemical carcinogens. The recent research outcomes in transgenic mouse models have made progressive advances in the understanding of chemical carcinogenesis and the evaluation of potential human carcinogens. However, these models still remain to be insufficient assay systems although the insufficiencies have been recognised and are being resolved. Based on up to date information from literature, this review article intends to understand currently accepted transgenic mouse models, issues arising from study design, interpretation of the study, results of validation project and their cancer prediction rate, and further perspectives of cancer assay models from the regulatory view point.

• 한국식품위생안전성학회지
• /
• 제27권1호
• /
• pp.37-41
• /
• 2012

현재 복어독의 표준 시험법은 다른 나라들과 마찬가지로 마우스를 이용한 동물시험법으로 명시되어 있다. 그러나, 살아있는 동물에게 고통을 주는 동물실험의 규제 확대로 인해 기기분석 시험법으로의 개선이 요구되고 있다. 또한, 동물시험법의 감도나 정밀성 및 정확성의 한계로 최근에 동물시험법을 대체할 수 있는 시험법에 대한 연구가 활발히 진행되고 있다. 본 연구에서 LC/MS/MS 시험법은 시료에서 테트로도톡신을 추출한 후 SPE(Solid phase extraction) 정제칼럼을 이용하여 정제하고 양이온 모드에서 MRM(multiple reaction monitoring)방법으로 분석하는 시험법으로서 밸리데이션 결과 검출한계(LOD)는 부위에 따라 $0.03{\sim}0.08{\mu}g/g$이었고, 정량한계(LOQ)는 $0.10{\sim}0.25{\mu}g/g$이었다. 검량선의 상관계수($r^2$)는 0.9986~0.9997이고, 회수율은 80.9%~103.0%이었으며 상대표준편차(RSD)는 4.3%~13.0%로서 적합한 시험법임을 확인하였다. 이 시험법을 이용하여 복어 검체의 부위별로 분석을 실시하였으며, 동물시험법의 시험결과와의 상관관계를 분석한 결과, 상관계수는 0.95이상의 상관성을 확인하였다.

• The Korean Journal of Physiology and Pharmacology
• /
• 제10권6호
• /
• pp.349-354
• /
• 2006

The aromatic hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates many of the biological and toxicological effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD, dioxin) and related chemicals. The application of recombinant reporter plasmid such as the firefly luciferase gene has proven to be a very effective method to detect these chemicals. The bioassay system, CALUX, is sensitive in directly detecting AhR-agonists from a variety of environmental and biologic materials. However, responses of the AhR-dependent bioassays are dependent on the cell types used. Thus, we developed a sensitive bioassay using the recombinant mouse hepatoma cell (Hepa1c1c7) for the determination of dioxins. The recombinant cell line was stably transfected with firefly luciferase reporter gene (pGudLuc1.1). The transfected cells showed the highest induction of luciferase activity at 4.5 hr and a decrease beyond this time point. The system showed the highest sensitivity of detection ever reported. Upon TCDD exposure cells showed 2 fold increase at 10 pM and 7 fold increase at 100 pM, respectively. The passage number after the transfection played an important role in the sensitivity. The increase of passage number tended to increase the sensitivity of the cells up to 15. The media without phenol red showed a higher induction rate than with phenol red, suggesting the preferable use of phenol red-free media for the bioassay. Since each of the assays has unique characteristics that make them suitable for some screening applications and not others, development of sensitive bioanalytical methods based on a variety of cellular systems in a key to the successful determination of dioxins. The bioassay system developed in this study will contribute to further development of successful screening the AhR agonists among the environmental mixture. In addition, the rapid and sensitive nature of this cellular system can be applied as a valuable tool to screen the dioxin-like moieties among the prodrugs at the initial stage, thereby expediting the new drug discovery.

• KSBB Journal
• /
• 제18권4호
• /
• pp.255-260
• /
• 2003

현재는 재조합 Erythropoietin의 생물학적 활성을 mouse를 이용한 in vivo bioassay로 실시하고 있으나, 이 방법의 여러가서 불편함으로 인하여, 이를 대체하기 위해 Ba/F3 세포주를 이용한 in vitro assay 방법을 확립하였으나, 위에 상술한 바와 같이 in vitro assay는 erythropoietin의 당분포에 의한 차이를 보이지 않게 때문에, 이를 보완하기 위해서는 in vivo bio-activity와 정량적인 상관관계가 있는 당단백질의 isoform 분석법인 Capillary zone electrophoresis (CZE) 와 sialic acid 함량 결과를 동시에 분석해야 했다. 위의 결과 sialic acid 함량은 erythropoietin 원액에서 10 mol/mol,EPO 이상의 sialic acid 함량을 가져야 되며, CZE 결과는 재조합 erythropoietin이 isoform 분포가 isoform 3의 경우 5.594～0.593％, isoform 4의 경우 31.598～11.704％, isoform 5의 경우 37.033～29.301％, isoform 6의 경우 27.837～18.807％, isoform 7의 경우 17.085～7.824％, isoform 8의 경우 7.642～1.964％을 보여줌과 동시에 isoform 4의 면적은 isoform 5의 면적보다 항상 작아야한다. 이상 두 가지 시험결과와 in vitro assay 결과를 combine해서 in vivo assay를 대체할 수 있다는 좋은 실험적 data를 얻었으므로 이상 위의 3가지 분석법을 활용한 combined in vitro bioassay 법의 기초를 확립하였다.

• 대한임상검사과학회지
• /
• 제50권4호
• /
• pp.382-390
• /
• 2018

임신진단검사는 고대 이집트에서 밀과 보리를 이용한 발아시험으로 시작하였다. 이후 Hippocrates와 Galen을 거쳐 uroscopy를 활용하던 중세를 넘어 1800년대 초반까지도 비과학적 방법들이 사용되었다. 그러나 1800년대 중반 이후 과학적 실험방법과 근거들이 나타나기 시작하면서 임신의 생리학적 특성에 대한 과학적 연구들이 시작되었다. 이와 같은 과학적 연구결과를 활용하려는 최초 시도는1927년 Aschheim와 Zondek의 rat와 mouse를 이용한 bioassay로 시작되었고, 이후 토끼와 개구리를 이용하는 실험으로 발전하였다. 1960년대부터 시작된 면역검사방법은 bioassay의 문제점을 개선하면서 임신진단검사를 일반화하는데 크게 기여하였다. 1976년에는 가정에서 사용할 수 있는 임신진단키트가 출시되어 임신진단을 대중화하는데 기여하였다. 1980년대 이후 진단검사의 기술적 진보는 임신진단검사에도 활용되어 임신진단의 검사신뢰도는 더욱 향상되었으며, 2000년대에는 가정용 임신진단키트의 정확도와 사용편리성이 혁신적으로 향상되었다. 본 조사에서는 임신진단검사의 역사와 과학적 발전과정을 고찰하였다.

• 한국환경성돌연변이발암원학회지
• /
• 제23권1호
• /
• pp.30-34
• /
• 2003

Recent industrial society has human widely exposed to PAHs (polynuclear aromatic hydrocarbons) that are comming from the incomplete combustion of organic material as wider spread environmental contaminants. Biological activities of PAHs are not known although PAHs are considered as carcinogens. Our laboratory have been studied the effect of PAHs in the mouse liver hepa 1 cells. In this study, we examined the mouse liver hepa-l cells as a new bioassay system to evaluate bioactivity of PAHs. We have selected 13 PAHs to examine bioassay using cyp1a1-luciferase reporter gene expression system where cyp1a1 1.6 Kb 5flanking region DNA was cloned in front of luciferase reporter gene and this plasmid was transfected into hepa 1 cells transiently. This cells then used for the study to observe the effect of PAHs. We demonstrated that PAHs induced the CYP1A1 promoter and 7-ethoxyresolufin O-deethylase (EROD) activities in a concentration-dependant manner. Some of PAHs showed stronger stimulatory effect on CYP1 gene expression than TCDD. Acenaphthene, anthracene, fluorine, naphthalene, pyrene, phenanthrene, carbazole were weak responders to cyp1a1 promoter activity stimulation and EROD induction in hepa 1 cells and these chemicals seemed to respond less to EROD than cyp1a1 promoter activity. Benz(a)anthracene, benzo(b)fluoranthene, benzo(k)fluoranthene, chrysene, and dibenzo(a,h)anthracene showed strong response to cyp1a1 promoter activity stimulation and also EROD induction in hepa 1cells. Results of dose response study suggested that four strong responding PAHs, such as benzo(a)anthracene benzo(k)fluoranthene, chrysene, and dibenzo(a, h)anthracene might be mediated through arylhydrocarbon receptor system in hepa1 cells.