Reference intervals of nine steroid hormones over the life-span analyzed by LC-MS/MS: Effect of age, gender, puberty, and oral contraceptives.

New reference intervals need to be established for a new analytical method with improved sensitivity and specificity. We aimed to establish the new reference intervals from infancy to senescence of nine steroid hormones (cortisol, cortisone, progesterone, 17-hydroxyprogesterone (17-OHP), androstenedione, testosterone, estradiol, DHEAS, and aldosterone) for LC-MS/MS method. Serum samples from 4678 reference individuals (age range: 0.3-79 years) were measured with LC-MS/MS. Samples were collected between 7 a.m. and 10 a.m. Exclusion criteria were concomitant endocrine diseases and body mass index ≥ 33. Generalized additive model for location, scale and shape, the nonparametric or robust method was applied. We established the reference intervals of the nine steroid hormones by sex, age, and pubertal stage. Below the age of one, we observed the surge of androgen and estrogen which implied mini-puberty. At the same period of life, aldosterone and cortisone levels were very high reflecting physiological hyperaldosteronism. An increase of steroid hormones during the pubertal development and slow decrease towards senescence after the peak at early adulthood were observed. Due to the increase of CBG synthesis, cortisol levels were increased under oral contraceptives (OC) significantly (p < 0.0001), while OC suppressed progesterone, 17-OHP, androstenedione, and estradiol (p < 0.0001). Our results will facilitate the interpretation of patient data in routine diagnostics with the use of LC-MS/MS method. Since LC-MS/MS methods have shown good comparability among the different laboratories, our reference intervals can be further adopted in other laboratories equipped with LC-MS/MS, once the validation with a small number of reference samples is performed.