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Objective-To establish normal, trimester-specific reference intervals for serum 17β-estradiol, progesterone (P), 17α-hydroxyprogesterone, cortisol, 11-deoxycortisol, androstenedione, DHEA, and DHEAS, measured simultaneously using isotope dilution tandem mass spectrometry. Design-Sequential cohort study.Patient(s)-Healthy women undergoing a normal pregnancy (age, 25-38 years; mean, 30 years) attending a prenatal well clinic at gestation weeks 12, 22, and 32 and approximately 1 year postpartum.Main Outcome Measure(s)-Trimester-specific reference intervals of endogenous steroid hormones analyzed using an isotope dilution tandem mass spectrometer equipped with an atmospheric pressure photoionization source with deuterium-labeled internal standards.Result(s)-Serum estradiol, P, 17α-hydroxyprogesterone, and 11-deoxycortisol increased throughout pregnancy; cortisol increased up to the second trimester and then remained steady, while androstenedione increased by 80 percent by gestation week 12, then remained constant. Serum DHEA-S decreased by 50% by the third trimester. Normal pregnancy depends on pronounced adaptations in pregnancy-related hormone concentrations, characterized by elevated levels of several circulating steroid hormones, which normally increase with the progression of pregnancy (1-6). Endogenous steroid hormone exposure during pregnancy has been of interest in studies of duration of gestation, fetal size, twin pregnancies, control of labor, nausea and vomiting in pregnancy, pregnancyinduced hypertension, and other disease states (5-9). The studies have generally produced weak and inconsistent findings because the hormone levels were either not available or lacked specificity or because surrogate measures of exposure to altered steroid hormone levels were often used to estimate these hormones. Conclusion(s)- NIH Public AccessSteroid hormones are derived from cholesterol. Binding proteins facilitate their transport and increase their half-life but limit their entry into target cells, thereby regulating their biological activity. Binding proteins make precise determination of serum steroid hormone concentrations difficult by interfering with different steroid hormones immunoassays (IAs). Furthermore, the lack of specificity of IAs due to cross reactivity with structurally related molecules is a well-known phenomenon (10-12).In contrast, isotope dilution liquid chromatography-tandem mass spectrometry (LC/MS/MS) is a specific detection method that allows the quantification of the analyte of interest. It also allows for a simpler approach to sample preparation without employing lengthy and timeconsuming extraction and sample derivatization steps. This has been reported in a previous publication, and steroid hormone analysis using isotope dilution LC/MS/MS was compared with the analysis of the same samples using IA techniques (14). Generally, tandem mass spectrometry provides lower values than IA, no doubt because of improved specificity.The reasons for the improved specificity in our tandem mass sp...

This method allows for the simultaneous measurement of four estrogens in human serum within 8 min. It can be routinely employed in a clinical environment and is attractive because of its simplicity in sample processing, micro sample requirement, and high throughput.

The steroid hormones are synthesized in the adrenal cortex, the gonads, and the placenta; are all derived from cholesterol and many are of clinical importance. Steroid hormones are synthesized in the mitochondria and smooth endoplasmic reticulum. Because they are lipophilic, they cannot be stored in vesicles from which they would diffuse easily and are therefore synthesized when needed as precursors. Upon stimulation of the parent cell, steroid hormone precursors are converted to active hormones and diffuse out of the parent cell by simple diffusion as their intracellular concentration rises.Because all steroid hormones are derived from cholesterol, they are not soluble in plasma and other body fluids. As a result, steroids are bound to transport proteins that increase their half-life and insure ubiquitous distribution. The protein-bound steroids are in equilibrium with a small fraction of free steroids, which are 'active.' Steroids can act quickly, by binding to cell surface receptors, or slowly, by binding to cytoplasmic or nucleic receptors and ultimately activate gene transcription.The adrenal glands are composed of the adrenal medulla and the adrenal cortex. The adrenal cortex is divided into three major anatomic zones: the zona glomerulosa, which produces aldosterone; and the zonae fasciculata and reticularis, which together produce cortisol and adrenal androgens. The medulla synthesizes catecholamines. More than 30 steroids are produced in the adrenal cortex; they can be divided into three functional categories: mineralocorticoids, glucocorticoids, and androgens.The steroids that are made almost exclusively in the adrenal glands are cortisol, 11-deoxycortisol, aldosterone, corticosterone, and 11-deoxycorti-costerone. Most other steroid hormones, including the estrogens, are made by the adrenal glands and the gonads [ MineralocorticoidsThe mineralocorticoids are formed in the zona glomerulosa. The main function of the mineralocorticoids is to promote tubular reabsorption of sodium and secretion of potassium and hydrogen ions at the collecting tubule, distal tubule, and collecting ducts [2]. When sodium is reabsorbed, water is absorbed simultaneously. The absorption of sodium and water increases fluid volume and arterial pressure.Aldosterone is the most potent mineralocorticoid and accounts for about 90% of the total mineralocorticoid activity. Mineralocorticoid potency in descending order is: aldosterone, 11-deoxycorticosterone, 18-oxocortisol, corticosterone, and cortisol [1]. Although cortisol has mainly glucocorticoid activity, it also has some mineralocorticoid activity. Cortisol has 1/400 of the potency of aldosterone, but its concentration is about 80 times greater than that of aldosterone [4]. The adrenal production of cortisol is approximately 25 mg/day and that of aldosterone is 100μg/day. Corticosterone has mainly glucocorticoid activity and some mineralocorticoid activity.Aldosterone secretion is regulated primarily by the renin-angiotensin system; it also is stimulated by increased serum potas...

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Hypothesis-A cortisol response to adrenocorticotropin injection is the standard test for diagnosing adrenal insufficiency. Multiple steroid hormones can now be accurately measured by tandem mass spectrometry in a single sample. The study objective was to determine whether a steroid profile, created by simultaneous measurement of ten steroid hormones by tandem mass spectrometry, would help determine the cause of adrenal insufficiency.Design-A ten-steroid profile was measured by tandem mass spectrometry during the performance of a standard high dose cortrosyn stimulation test. The steroids were measured at baseline, and 30 and 60 minutes following synthetic adrenocorticotropin injection. Adrenal insufficiency was defined as a peak cortisol level of less than 20 μg/dL. Testing was conducted in the General Clinical Research Center of a University Medical Center. Normal volunteers, patients suspected of having adrenal insufficiency, and patients with known adrenal insufficiency participated.Results-Our results showed that adrenal insufficiency of any cause was adequately diagnosed using the response of 11-deoxycortisol, dehydroepiandrosterone, or these analytes combined in a two-steroid profile. A three-steroid profile yielded a test with 100% accuracy for discriminating primary adrenal insufficiency from normal status. Primary adrenal insufficiency was well separated from secondary adrenal insufficiency using only a single aldosterone value. 11-deoxycortisol, dehydroepiandrosterone, and a two-steroid profile each provided fair discrimination between secondary adrenal insufficiency and normal status.Conclusions-We conclude that stimulated levels of aldosterone, 11-deoxycortisol, dehydroepiandrosterone, and a two-or three-steroid profile provided additional discrimination between states of adrenal sufficiency and insufficiency. It is proposed that a steroid profile measuring

This is the first study to provide pediatric reference intervals of FT(4) and FT(3) for children from birth to 18 years of age using LC/MS/MS. Analysis using LC/MS/MS provides more specific quantification of thyroid hormones. A comparison of the ultrafiltration tandem mass spectrometric method with equilibrium dialysis showed very good correlation.

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