Measurement of reference intervals for urinary free adrenal steroid levels in Japanese newborn infants by using stable isotope dilution gas chromatography/mass spectrometry

Abstract

BackgroundIn newborn infants, there are no reference intervals for urinary free steroids, which are thought to reflect the bioavailable fraction of steroids in the blood. We establish a method for simultaneous measurement of urinary free adrenal steroids such as pregnenolone, progesterone, 16α-hydroxyprogesterone, 17α-hydroxyprogesterone, 21-deoxycortisone, 21-deoxycortisol, dehydroepiandrosterone, androstenedione, and 11β-hydroxyandrostenedione by using stable isotope dilution gas chromatography/mass spectrometry (SID-GC/MS) and determined the reference intervals for urinary levels of free adrenal steroids in Japanese newborn infants. MethodsNewborn pooled urine was used for validation. Spot urine samples were collected from 67 full-term Japanese newborn infants (34 male and 33 female infants) at 3–4days of age to determine reference intervals. The extracted and purified free steroids were delivered with heptafluorobutyric anhydride and analyzed by SID-GC/MS. ResultsWe validated a SID-GC/MS method with good repeatability and recovery rate. The preliminary reference intervals (median [range], μmol/mol creatinine) were as follows: pregnenolone, 4.2 (0.7–31.6); progesterone, 0.5 (not detected (n.d.)–0.6); 16α-hydroxyprogesterone, 1.4 (n.d.–10.3); 17α-hydroxyprogesterone, 1.1 (n.d.–1.9); 21-deoxycortisone, n.d. (n.d.–n.d.); 21-deoxycortisol, n.d. (n.d.–n.d.); dehydroepiandrosterone, 2.2 (0.6–27.3); androstenedione, 0.7 (n.d.–5.2); and 11β-hydroxyandrostenedione, 2.9 (n.d.–26.7). ConclusionsWe established a reliable SID-GC/MS method and were able to determine preliminary reference intervals for 9 urinary free adrenal steroids in newborn infants.