高效液相荧光检测法测定柑橘中5-甲基四氢叶酸的含量

Quantitative Determination of 5-Methyltetrahydrofolic Acid in Citrus by High Performance Liq uid Chromatography-fluorescence Detection

利用5-甲基四氢叶酸自身产生荧光的特性,建立高效液相荧光检测法来测定柑橘中5-甲基四氢叶酸的含量。使用ODS-C18反相色谱分离柱,流动相为6%乙腈,94%0.05mol/L磷酸二氢钾溶液(pH3.0)作为流动相,流速1.0mL/min,柱温23℃,进样量为20μL,激发波长为290nm,发射波长为360nm。结果表明,5-甲基四氢叶酸在0.003μg/mL-0.1μg/mL有良好的线性关系,相关系数为0.9999,检出限为0.01ng/mL,样品测定的精密度为0.99%,样品加标平均回收率为96.6%。建立的高效液相荧光检测法具有良好的精密度和回收率,适合用于柑橘中5-甲基四氢叶酸的测定。

A sensitive and specific high performance liquid chromatography-fluorescence detection （RP-HPLC-FLD）） method was developed for the quantitative determination of 5-methvltetrahvdrofolic acid （5-MTHF） in citrus, using the acid＇s fluorescence-emitting characteristics. The determination was performed on a ODS C18 reverse-phase column en- closed in a heater maintained at 23℃, the mobile phase was a mixture of 94% 0.05mol/L aqueous potassium phosphate buffer （pH 3.0 ） containing 6% acetonitrile, and the flow rate was 1 mL/min. The loop size was 20μL, the optimized exci- tation wavelength was 290nm and the emission wavelength was 360nm. The results showed that the detection limits were 0.01ng/mL, the linear range was 0.003μg/mL - 0.1μg/mL, the correlation coefficient was 0.9999, the recovery was 96.6%, and the precision of the method was 0.99%. It was concluded that this method, being simple, fast and accurate, had high sensitivity, recovery and good repeatability and was suitable for the determination of 5-MTHF in citrus.