高效液相色谱-氢化物发生-原子荧光光谱联用技术测定富硒大米中的5种硒形态

Determination of 5 kinds of selenium species in rice by high performance liquid chromatography-hydride generation-atomic fluorescence spectrometry

目的建立高效液相色谱-氢化物发生-原子荧光光谱法(high performance liquid chromatographyhydride generation-atomic fluorescence spectrometry,HPLC-HG-AFS)检测富硒大米中硒代胱氨酸(Se Cys2)、甲基硒代半胱氨酸(Me Se Cys)、亚硒酸根(Se IV)、硒代蛋氨酸(Se Met)和硒酸根(Se VI)的方法。方法选择Hamilton PRP-X100阴离子交换色谱柱,对流动相浓度、p H值以及提取条件进行选择优化。样品采用链霉蛋白酶E水解超声提取方式,使用40 mmol/L的磷酸氢二铵溶液(p H 6.0)作为流动相。结果 20 min内可以完全分离5种硒形态,各种硒形态标准曲线线性相关系数(r)均大于0.999,检出限分别为3.7、1.7、1.2、4.2、7.2μg/L。加标回收实验显示富硒大米中硒的主要存在形态即硒代蛋氨酸回收率范围在95.1%~98.2%,其他4种硒形态回收率范围均在80.4%~102.5%。结论本方法可以快速、准确地测定大米中硒的5种形态。

Objective To establish a method for detecting of 5 kinds of selenium species such as selenocystine （SeCys2）, seleniet （SeIV）, methylselenocysteine （MeSeCys）, selenomethionine （SeMet） and selenate （SeVI） in selenium enriched rice by high performance liquid chromatography-hydride generation-atomic fluorescence spectrometry （HPLC-HG-AFS）. Methods A Hamilton PRP-X100 reversed-phase anion exchange column was used for separation, the concentration, pH value of the mobile phase and the extraction procedure were analyzed andoptimized. The Pronase E hydrolysis with ultrasonic extraction was taken as the sample pretreatment method, and 40 mmol/L （NHa）2HPO4 （pH 6.0） solution was used as mobile phase. Results Five species could be completely separated within 20 min, and all linear correlation coefficients of the 5 selenium species were better than 0.999. The detection limits of SeCys2, MeSeCys, SelV, SeMet, SeVI were 3.7, 1.7, 1.2, 4.2, 7.2 μg/L, respectively. The recoveries were in the range of 95.1%-98.2% for SeMet, which of other species were in the range of 80.4%-102.5%. Conclusion This method can be used to determine 5 kinds of selenium species in rice quickly and accurately.