Korean Journal of Food Science and Technology (한국식품과학회지)

Korean Society of Food Science and Technology (한국식품과학회)
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Changes in Color Response of MTT formazan by Zinc Protoporphyrin

MTT formazan의 발색에 미치는 zinc protoporphyrin의 영향

  • Park, Kyung-A (Department of Food Science and Technology, College of Natural Science, Seoul Women's University) ;
  • Choi, Hyun-A (Department of Food Science and Technology, College of Natural Science, Seoul Women's University) ;
  • Kim, Mi-Ri (Department of Food Science and Technology, College of Natural Science, Seoul Women's University) ;
  • Choi, Yoo-Mi (Department of Food Science and Technology, College of Natural Science, Seoul Women's University) ;
  • Kim, Hyun-Jung (Department of Food Science and Technology, College of Natural Science, Seoul Women's University) ;
  • Hong, Jung-Il (Department of Food Science and Technology, College of Natural Science, Seoul Women's University)
  • 박경아 (서울여자대학교 자연과학대학 식품공학과) ;
  • 최현아 (서울여자대학교 자연과학대학 식품공학과) ;
  • 김미리 (서울여자대학교 자연과학대학 식품공학과) ;
  • 최유미 (서울여자대학교 자연과학대학 식품공학과) ;
  • 김현정 (서울여자대학교 자연과학대학 식품공학과) ;
  • 홍정일 (서울여자대학교 자연과학대학 식품공학과)
  • Received : 2011.08.31
  • Accepted : 2011.10.31
  • Published : 2011.12.31
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Abstract

Zinc protoporphyrin (ZnPP) is produced endogenously during heme metabolism and treated in cells as a heme oxygenase inhibitor. In the present study, the effects of ZnPP on the color response of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, a commonly-used method for analyzing cell viability, were investigated. ZnPP induced rapid decolorizaion of MTT formazan under light; the degradation rates were 10- and 20- folds faster in the presence of 5 and $10{\mu}M$ ZnPP, respectively. Methylene blue (MB), another type of photosensitizer, also accelerated degradation of formazan under light. Butylated hyroxytoluene did not inhibit ZnPP- or MB-induced formazan degradation. The color degradation of formazan dye was signficantly delayed in the presence of N-acetylcysteine or ${\beta}$-carotene. The present results suggest that certain photosensitizing compounds may affect the color and stability of MTT formazan, which should be carefully considered when conducting the MTT assay.

본 연구는 세포의 사멸 및 성장변화 등의 평가에 널리 이용되는 MTT assay에서 생성된 formazan dye에 미치는 ZnPP의 영향을 조사하였다. ZnPP는 생체 내에 자연적으로 생성되거나 다양한 관련 실험에 인위적으로 첨가해주는 물질로서, MTT formazan dye의 빛에 의한 탈색을 가속화시키는 것으로 밝혀졌다. Formazan dye의 분해는 5와 $10{\mu}M$ ZnPP 존재 시 반감기를 기준하여 각각 10 및 20배 가량 가속화되었으며, 빛이 차단된 조건에서는 영향을 미치지 않았다. ZnPP 구조 중 Zn는 formazan dye의 탈색에 영향을 미치지 않았으나, porphyrin 구조와 공통적인 감광체 성질을 나타내는 MB에 의해서 ZnPP 존재 시와 유사하게 dye의 탈색을 가속화 시켰다. 이러한 ZnPP와 MB에 의한 formazan dye의 탈색반응은 NAC와 ${\beta}$-carotene에 의해 지연되었으나, BHT에 의한 저해효과는 나타나지 않았다. 본 결과는 세포 중에 존재하는 ZnPP 등의 감광체류가 MTT tetrazolium으로부터 환원된 formazan dye의 빛에 의한 신속한 분해를 유도하며, 관련 실험에서 빛의 차단 등의 조치와 함께 각별히 유의해야함을 시사한다.

Keywords

References

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