Development of Enzyme-Protein Binding Assay for Rapid and Sensitive Analysis of Biotin

Biotin 정량분석틀 위한 효소-단백질결합 분석법(EPBA)의 개발

  • 이경애 (한국식품개발연구원) ;
  • 손동화 (한국식품개발연구원) ;
  • 고영태 (덕성여자대학교 식품영양학과)
  • Published : 1998.12.01

Abstract

Conditions for enzyme-protein binding assay (EPBA) were established in order to detect biotin more rapidly and reproducibly than traditional microbiological assay (MBA). EPBA with streptavidin and biotin-KLH conjugate showed cross-reactivities on biocytin, a derivative with biotin activity, at the rate of 109% $(IC_{50}=0.3\;ppb)\;and\;197%\;(IC_{50}=0.8\;ppb)$, respectively, but not on other derivatives with no biotin activities, such as desthiobiotin, diaminobiotin and 2-iminobiotin. Detection ranges of biotin by EPBA with streptavidin and biotin-KLH conjugates were $0.01{\sim}30\;ng/mL\;and\;0.01{\sim}1.0\;ng/mL(ppb)$, respectively. In the spike test with milk, fruit flake and pine-carrot juice, the correlation coefficience between MBA and EPBA with biotin-KLH conjugates was r=0.994. But MBA showed cross-reactivities both on biocytin and desthiobiotin at the rate of 80.1% and 66.7%, respectively. Detection range of biotin by MBA was $0.1{\sim}0.5\;ng/mL(ppb)$. These results strongly suggest that EPBA is efficient for biotin detection in sensitivity, detection range, cross-reactivity and time consuming.

Keywords

biotin;enzyme protein binding assay;microbiological assay