高效液相色谱-串联质谱法测定家蚕血液中β-蜕皮激素

建立了高效液相色谱-串联质谱(HPLC-MS/MS)测定家蚕血液中β-蜕皮激素的方法。实验以高油菜素内酯为内标,用乙腈去除家蚕血液样品中的蛋白质;C18反相柱作为分析色谱柱,以乙腈和含0.5%乙酸的水溶液作为流动相,使用梯度洗脱的方法进行液相色谱分离。在优化的仪器条件下,血液中蜕皮激素的加标回收率在83.56%~97.11%之间,相对标准偏差小于5.4%,在3.13~50μg/L浓度范围内,线性相关系数R2为0.996 3,方法的检出限为0.2μg/L。该方法能较好地满足测定家蚕血液中β-蜕皮激素含量的要求,同时也可被推广测定其他昆虫血液中的β-蜕皮激素。

Determination of 20-Hydroxyecdysone in Hemolymph of Bombyx mori by HPLC-MS/MS

A method for the determination of 20-hydroxyecdysone in hemolymph of Bombyx mori was developed by high performance liquid chromatography-tandem mass spectrometry（HPLC-MS/MS）. 22(S), 23(S) homobrassinolide was used as an internal standard, and acetonitrile was used to precipitate the proteins in the hemolymph of Bombyx mori to eliminate the interference. The gradient elution program was performed on a C18 column using a mobile phase consisting of acetonitrile and 0.5% acetic acid solution. Under the optimum working conditions of the instrument，the analytical results show that the recoveries of 20-hydroxyecdysone in hemolymph of Bombyx mori are in the range of 83.56%-97.11%， and the relative standard deviation is below 5.4%. Linearity of the method in the range 3.13-50 μg/L is good with the correlation coefficients(R²) of 0.9963. The detection limit of the method is 0.2 μg/L. The method can meet the need for the determination of 20-hydroxyecdysone in the hemolymph of Bombyx mori, which may also be applied to determine 20-hydroxyecdysone in the hemolymph of other insects.