复杂发酵产物中黄豆苷元和雌马酚的定量分析

Quantitative Analysis of Daidzein and Equol in Complex Fermentation Products

为解决复杂发酵液背景下非目的代谢物干扰问题,建立一种适用于体外发酵条件下,黄豆苷元及其生物转化物雌马酚的高效液相色谱法分析方法。粪便菌群在脑心浸液肉汤培养基中转化黄豆苷元48 h,经乙酸乙酯提取,C18色谱柱(250mm×4.6 mm,5μm)分离,流速为0.8 mL/min,柱温25℃,进样量20μL,以水-甲醇-乙腈(体积比5∶3∶2)为流动相,在230nm和260 nm双波长处检测。结果表明:方法快速、准确,在6.25~100μg/mL范围黄豆苷元和雌马酚的线性相关系数(R2)分别为0.9999和0.9996,平均回收率为92.12%~104.34%,相对标准偏差为0.15%~0.24%,检出限分别为0.07μg/mL和0.05μg/mL,定量限分别为0.23μg/mL和0.16μg/mL。本研究所建方法具有较高的精确度且方法简便,可应用于粪便菌群发酵液中黄豆苷元和雌马酚的定量检测。

To solve the interference problem of undesired metabolites with complex background fermentation liquid,high performance liquid chromatography method for analyzing suitable for in vitro fermentation conditions,the measurement daidzein and equol.The fecal microbiota was transformed into daidzein in Brain Heart infusion(BHI)broth medium for48 hours.After extracted by ethyl acetate,the samples were analyzed on a C18 column(250 mm×4.6μm,5μm).The flow rate was 0.8 mL/min,the column temperature was 25℃,the injection volume was 20μL,and the mobile phase was water-methanol-acetonitrile(volume ratio,5∶3∶2).The detection was carried out at UV 230 nm and 260 nm.The linear correlation coefficients of daidzein and equol were R2=0.9999,0.9996,respectively,the average recovery were92.12%-104.34%,the relative standard deviation were 0.15%-0.24%,the limit of detection were 0.07μg/mL and 0.05μg/mL,and the limits of quantification were 0.23μg/mL and 0.16μg/mL,respectively.The method established in this study had high accuracy and precision and was simple.It has been applied to determine daidzein and equol in fecal microbiota fermentation.