在线净化液相色谱-串联质谱法快速测定牛奶中的6种孕激素

建立了在线净化液相色谱-串联质谱同时测定牛奶中炔诺酮、17α-羟基孕酮、甲羟孕酮、乙酸甲地孕酮、孕酮和醋酸美伦孕酮6种孕激素的方法。本方法采用乙腈为提取溶剂提取目标化合物。提取液经在线净化柱Cyclone-P净化，经Phenyl-Hexyl色谱柱分离，流动相采用0.5%（v/v）甲酸水溶液-乙腈，在电喷雾正离子模式下以多反应监测（MRM）方式测定，内标法定量。方法在0.1~50 μg/L范围内呈线性关系，线性相关系数均大于0.999。6种分析物的测定低限为0.5 μg/kg，在牛奶中3个水平的添加回收率在90.8%~107.5%之间，相对标准偏差在6.3%~11.8%之间。该方法快速简便，灵敏度高，选择性好，可用于牛奶样品中孕激素的快速定性定量分析。

Determination of six progesterones in milk by on-line cleanup liquid chromatography-tandem mass spectrometry

A new method using TurboFlow on-line cleanup liquid chromatography combined with tandem mass spectrometry (MS/MS) has been developed for simultaneous determination of six progesterones including 19-norethindrone, 17 α-hydroxyprogesterone, medroxyprogesterone, megestrol acetate, progesterone and melengestrol acetate in milk. Samples were extracted by acetonitrile. The analytes in extract were on-line purified directly on Cyclone-P purification column where the sample matrices were washed away. Subsequently, the analytes which were eluted from the extraction column onto Phenyl-Hexyl column were separated with a gradient elution, and detected with electrospray ionization mass spectrometry in the positive scan mode using multiple reaction monitoring (MRM). The isotope internal standards were employed for quantification. As a result, the linearities were satisfactory with the correlation coefficients of >0.999 at concentrations ranging from 0.1 μg/L to 50 μg/L. Based on the repeated analysis of a known blank sample, the limit of quantification (LOQ) is 0.5 μg/kg. Average recoveries of the analytes fortified at three levels (1, 5 and 25 μg/kg) ranged from 90.8% to 107.5%, and the relative standard deviations (RSDs) ranged from 6.3% to 11.8%. This proposed method is simple, rapid, sensitive and highly selective, and can be applied to simultaneous identification and quantification of the six progesterones in milk.