Abstract
A liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI/MS/MS) method was developed for the determination and confirmation of clenbuterol in bovine muscle and milk. Clenbuterol and clenbuterol-D9 using as an internal standard in samples were extracted with ethyl acetate after hydrolysis and evaporated to dryness. The extracts were dissolved in 20% methanol and cleaned using HLB solid-phase extraction cartridge. The analytes were detected by LC-ESI/MS/MS on a $C_{18}$ column. Mass spectral acquisition was done in selected reaction monitoring (SRM) in positive ion mode to provide a high degree of sensitivity. Using MS/MS with SRM mode, the transitions (precursor to product) monitored were m/z 277${\rightarrow}$203 for clenbuterol, and m/z 286${\rightarrow}$204 for internal standard. The limits of quantitation (LOQ) and mean recoveries of clenbuterol in bovine muscle were $0.2{\mu}g/kg$ and 84.3~91.1%, respectively. The LOQ and mean recoveries in milk were $0.05{\mu}g/kg$ and 87.7~98.3%, respectively.
본 연구에서는 소고기와 우유에서의 클렌부테롤의 확인과 정량을 위하여 LC-ESI/MS/MS을 이용한 방법을 개발하였다. 클렌부테롤과 내부표준물질로 클렌부테롤-D9을 사용한 시료를 가수분해 한 후 에틸아세테이트로 추출하고 농축하였다. 그리고 추출물을 20% 메탄올에 용해하여 HLB 카트리지로 정제한 후 $C_{18}$ 칼럼을 이용한 LC-ESI/MS/MS로 분석하였다. 높은 감도를 얻기 위하여 positive ion mode에서 SRM(selected reaction monitoring)방법을 이용하였다. MS/MS의 SRM방법으로 클렌부테롤과 클렌부테롤-D9의 선구이온(precursor ion), 생성이온(product ion)을 각각 m/z 227${\rightarrow}$203, m/z 286${\rightarrow}$204로 분석한 결과 소고기에서의 클렌부테롤의 정량한계와 회수율은 각각 $0.2{\mu}g/kg$과 84.3~91.1%이었고, 우유에서의 정량한계와 회수율은 각각 $0.05{\mu}g/kg$과 87.7~98.3%이었다.
