Implementation of a Liquid Chromatography Tandem Mass Spectrometry Assay for Eight Adrenal C-21 Steroids and Pediatric Reference Data

BACKGROUND: Sensitive and accurate determination of steroids is essential for diagnosing congenital and acquired adrenal diseases. Since plasma concentrations change during childhood, age-specific reference ranges are the prerequisite for clinical interpretation. The objectives of this study were to develop a sensitive and reliable method for simultaneous detection and quantification of progesterone, 17-hydroxyprogesterone, deoxycorticosterone (DOC), 11-deoxycortisol, 21-deoxycortisol, corticosterone, cortisol (F) and cortisone (E) by ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) and to establish age- and sex-specific reference ranges from birth to adulthood. METHODS: All eight steroids were measured simultaneously in 0.1 ml plasma by UPLC-MS/MS. Samples of 905 children were measured and grouped in five age groups. RESULTS: The assay was linear up to 70 ng/ml (700 ng/ml for F; r(2) > 0.992). The limit of detection ranged between 0.01 ng/ml for DOC and 0.07 ng/ml for E. Correlations with radioimmunoassays yielded a coefficient of determination between 0.82 and 0.99. Reference data are reported as a function of age and sex. CONCLUSIONS: The UPLC-MS/MS method presented here for the simultaneous detection of eight C-21 adrenal hormones together with the detailed reference ranges for children provides a valuable methodology for assessing adrenal steroids in clinical routine and research.