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目的:建立衍生化-毛细管气相色谱法测定白念珠菌中羊毛甾醇和麦角甾醇等甾醇的含量。方法:白念珠菌经破碎、离心、皂化、提取出甾醇部分后再经1-三甲基甲硅烷咪唑(TMSI)硅烷化后,其TMSI衍生物采用HP 50毛细管柱进行气相色谱分析,柱温:程序升温150～290℃,起始温度150℃,按15℃·min~(-1)升至290℃恒定;检测器温度280℃;进样口温度270℃。柱头压35kPa;载气:氮气,流速82mL·min~(-1);每次进样量:1μL;分流比为100:1。结果:在0.1862～1.862mg·mL~(-1)衍生物的氯仿溶液范围羊毛甾醇和麦角甾醇线性关系良好(r分别为0.9996和 0.9969),回收率分别大于97％和93％(RSD<5％),最低检测限分别为1.86ng·mL~(-1)和2.54ng·mL~(-1),日内精密度良好(RSD均在5％以内)。结论:本方法准确、灵敏、专属性强,结果可靠,为抗真菌药物研究提供了有力的工具。
OBJECTIVE: To establish a derivatization-capillary gas chromatographic method for the determination of lanosterol and ergosterol in Candida albicans. Methods: Candida albicans was broken up, centrifuged and saponified. After the sterol fraction was extracted, the TMSI derivatives were silylated by 1-trimethylsilyl imidazole (TMSI). The column was analyzed by gas chromatography with HP 50 capillary column. : Temperature 150 ~ 290 ℃, initial temperature 150 ℃, constant at 15 ℃ · min ~ (-1) to 290 ℃; detector temperature 280 ℃; inlet temperature 270 ℃. Column pressure 35kPa; carrier gas: nitrogen, flow rate 82mL · min -1; each injection volume: 1μL; split ratio of 100: 1. Results: There was a good linear relationship between lanosterol and ergosterol (r = 0.9996 and 0.9969, respectively) in the range of 0.1862 ~ 1.862 mg · mL -1 of chloroform. The recoveries were higher than 97% and 93% respectively %). The detection limits were 1.86ng · mL -1 and 2.54ng · mL -1, respectively. The intra-day precision was good (RSD within 5%). Conclusion: The method is accurate, sensitive, specific and reliable. It provides a powerful tool for the study of antifungal agents.