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Quantitative-Profiling Method of Serum Steroid Hormones by Hydroxylamine-Derivatization HPLC-MS |
| Qi Liu, Quan Chi, Ru-Ting Fan, Hui-Dong Tian, Xian Wang |
| Key Laboratory of Analytical Chemistry of the State Ethnic Affairs Commission, College of Chemistry and Materials Science, South-Central University for Nationalities, Wuhan 430074, Hubei, People's Republic of China |
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Abstract A sensitive and rapid high performance liquid chromatography-mass spectrometry (HPLC-MS) method was developed and validated for simultaneous quantification of ten steroid hormones, including estrogens, androgens, progesterones, and corticosteroids four classes of steroids. The following ten steroid hormones were analyzed:progesterone, 21-deoxycortisol, estrone, 4-androstenedione, testosterone, dihydro-testosterone, androstenone, dehydroepiandrosterone, corticosterone and cortisone. Stable deuterated isotopes were used as internal standards for quantification. Sample preparation with and without derivatization were performed after liquid-liquid extraction, and the corresponding results were compared according to sensitivity and selectivity. Hydroxylamine derivatization was found to improve the ionization efficiency of the analytes for electrospray ionization MS analysis. The gradient of mobile phase and experimental parameters for HPLC separation were optimized. The lower limits of quantification were in the range of 0.05-5 ng mL-1 with wide linear range for the ten steroid hormones. The intra-day precision < 11.1% and recovery of 84.5-120% with negligible matrix effect were achieved, where within the acceptance limits of the FDA guideline. Total HPLC-MS analysis time was 6 min. This method enables simultaneous quantification of steroids in human serum. It will be helpful for the serum steroid profiling in order to understand various endocrinology diseases.
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| Keywords
Steroids
Derivatization
High performance liquid chromatography-mass spectrometry (HPLC-MS)
Quantitative-profiling
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| Fund:The authors thank the National Natural Science Foundation of China (Grant No. 21275167), the Natural Science Foundation of Hubei Province (Grant No. 2014CFA025), and the Preferred Research Foundation for the Returned Overseas Scholars from Ministry of Human Resources and Social Security of the People's Republic of China for financial support. |
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Corresponding Authors:
Xian Wang
E-mail: xwang27@mail.scuec.edu.cn
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Issue Date: 05 June 2019
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