Determination of 6 Kinds of Sex Hormones in Fish Using Subcritical 1,1,1,2-Tetrafluoroethane Extraction-Gas Chromatography-Tandem Mass Spectrometry

Abstract A sample pretreatment method was developed for the analysis of six sex hormones in fish by subcritical 1,1,1,2-tetrafluoroethane (R134a) extraction coupled with gas chromatography-mass spectrometry(GC-MS). After the extraction process was completed, the fatty co-extracted interferent of the sample was removed by freezing filtration. Then the analytes were cleaned up by C 18 and NH 2 solid phase extraction (SPE) column. At last, the hormones were derivatived by heptafluorobutyric anhydride (HFBA) followed by GC-MS validation. The optima extraction conditions for the six sex hormones were as follows: extraction pressure, 4 MPa; extraction temperature, 30 °C; extraction cosolvent amount, 6 mL. Under these conditions, the calibration curves showed good linearity with correlation coefficients larger than 0.99 in the hormones concentrations of 5–1000 μg L −1 ; the detection limits were 0.2–1.0 μg kg −1 ( S / N = 3). At the spiked level of 1, 5 and 10 μg kg −1 , the mean recoveries were between 70.5% and 103.6%, and the relative standard deviations (RSDs) ranged from 2.1% to 12.5%. Finally, diethylstilbestrol was detected in real samples by the proposed method, and the residue content of which was 14.6 μg kg −1 .