HPLC法测定饲料中的炔雌醇

Ethinylestradiol in Feeds Determined by HPLC

为了制定中国适用于饲料中炔雌醇的高效液相色谱（HPLC）测定的标准方法。研究通过添加回收方法，以三大类饲料基质（配合饲料、浓缩饲料、添加剂预混料）的代表样品为研究对象，建立了HPLC法测定饲料中炔雌醇的方法，即以乙腈-0．025％磷酸f1：1，v／v）组成混合提取液，超声辅助提取，经液液萃取和C18固相萃取净化后，以反相液相色谱荧光法测定。实验结果表明：该方法线性范围0．05～5mg／L，线性相关系数O．9992；检出限浓度0．04mg／kg（添加剂预混料为0．06mg／kg），定量限浓度0．1mg／kg（添加剂预混料为0．2mg／kg）；配合饲料在0．05、0．25、0．5mg／kg添加水平的回收率和精密度分别为83％～87％和5．9％～7．0％；三大类饲料平均回收率在81％～86％，相对标准偏差（RSDs）实验室内为3_3％～9．6％，实验室间为5．7％～6．6％。该方法操作简单、环保、灵敏，适用于各类饲料样品中炔雌醇的快速筛查。

To establish the domestic standard method, a high performance liquid chromatography （HPLC） assay was developed for ethinylstradiol determination in three key types of feeds （complete formula feeds, concentrated feeds, and vitamin premixes）. In this assay, ethinylstradiol was extracted from feeds by acetonitrile-0.025% phosphate （I：I, v/v） solution, then the extraction solutions were purified by liquid-liquid partitioning, and further purified by the reverse phase extraction C- 18 cartridge, and later, ethinylstradiol in the purified solutions was determined by reverse phase HPLC coupling fluorescence detector. The results showed that the linear ranges measured by this assay were 0.05-5 mg/L ethinylstradiol, and the linear correlation coefficient was 0.9992. The LOD of this assay for feeds and vitamin premixes was 0.04 mg/kg and 0.06 mg/kg, respectively. The LOQ was 0.1 mg/kg for feeds and 0.2 mg/kg for vitamin premixes. The average recoveries in complete formula feed at three spiked level of 0.05, 0.25, 0.5 mg/kg were between 83% and 87%, with RSD ranged from 5.9% to 7.0%. The average recoveries in three key types of feeds were 81%-86%, and the relative standard deviations （RSDs） of intra-lab and inter-lab were 3.3%-9.6% and 5.7%-6.6%, respectively. The estalished method was environment-friendly, sensitive, and convenient for fast determination of ethinylestradiol in different types of feeds.