A high sensitive simultaneous determination method of nine steroid hormones including Estrone (E1), estradiol (E2), testosterone (T), dihydrotestosterone (DHT), 17-hydroxyprogesterone (17OH-P4), androstenedione (AN), progesterone (P4), dehydroepiandrosterone (DHEA), and cortisol by liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) was established in a positive mode using recently developed picolinyl derivatization. Steroid hormones were derived with picolinic acid and 2-methyl-6-nitrobenzoic anhydride, then purified by solid-phase extraction with InterSep SI cartridge. The LC-ESI-MS/MS method enhanced the specificity and sensitivity for E1, E2, T, DHT, DHEA, and cortisol. The method validation indicated that the limits of quantification for E1, E2, T, DHT, 17OH-P4, AN, P4, DHEA, and cortisol were 1, 1, 1, 2, 2, 2, 2, 20, and 100 pg/tube, respectively with acceptable accuracy and precision within ± 15%. The present method was applied to the measurement of nine steroid hormones in children’s serums with high reliability and reproducibility.

Keywords: LC-ESI-MS/MS, steroid hormone, picolinyl derivatization, human serum, solid-phase extraction.

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