Dexamethasone suppression test: development of a method for simultaneous determination of cortisol and dexamethasone in human plasma by liquid chromatography/tandem mass spectrometry

Abstract

Background: Dexamethasone is a synthetic glucocorticoid and is analogous to cortisol. It is used in the low-dose overnight dexamethasone suppression test (LDODST) to diagnose hypercortisolism in patients suspected to be suffering from Cushing's syndrome (CS). Measuring plasma dexamethasone in conjunction with measuring the amount of cortisol following the LDODST may allow clinicians to improve the diagnosis of CS. Methods: Plasma samples were cleaned up by solid-phase extraction before analysis. Liquid chromatographic separation was carried out under reversed-phase conditions prior to detection by tandem mass spectrometry. The analytes were determined in the presence of deuterated internal standards cortisol-d4 and dexamethasone-d4. Results: Limit of quantitation (LOQ) was 1.89 nmol/L for dexamethasone and ,0.02 mmol/L for cortisol. Recoveries of both analytes ranged from 80.2% to 114.4%. Intra-and interassay coefficients of variation were ,15%. The concentration of dexamethasone and cortisol was determined in 62 patients after performing LDODST. Dexamethasone concentrations ranged from 3.0 to 21.5 nmol/L (median 7.4 nmol/L) for 57 of these samples. For five patients the concentration was ,LOQ. Cortisol concentrations were ,0.08 mmol/L (median 0.037 mmol/L, n ¼ 54) except for eight patients (.0.22 mmol/L). Conclusions: A method for the simultaneous measurement of dexamethasone and cortisol in human plasma by liquid chromatography/tandem mass spectrometry has been developed and validated. The method is suitable for controlling the compliance to the LDODST and for determining the cortisol plasma concentration after the test. The interpretation of LDODSTs was improved by the simultaneous determination of both analytes.

In the overnight dexamethasone suppression test, 1.0 mg loading is superior to 0.5 mg loading for diagnosing subclinical adrenal Cushing’s syndrome based on plasma dexamethasone levels determined using liquid chromatography-tandem mass spectrometry

“…DEX levels after a low-dose DST in the current study did not attain a predictably effective value (2.2 ng/mL) in a higher proportion of patients than previously reported [21]. One possible explanation for the unexpected results is that the Endocrine Society recommendations regarding DEX levels may be inadequate.…”

“…Nonetheless, plasma DEX levels determined using LC-MS/MS at 0800 h following 1-mg DST were similar to those determined using an RIA method, with plasma levels in most subjects within the range of 1.0 ng/mL to 10 ng/mL [8,[21][22][23].…”

Steroid metabolome analysis reveals prevalent glucocorticoid excess in primary aldosteronism

“…We used validated and established liquid chromatography-tandem MS assays to measure serum cortisol (51), serum dexamethasone (52), and salivary cortisol (53). Cosyntropin test results were compared to those obtained in 82 healthy controls measured with the same assay; the diagnostic cut-off for diagnosis of adrenal insufficiency was defined as the 5th centile of the healthy control cohort and determined as 15.6 μg/dl (430 nmol/l).…”

Drugs as diagnostic tools

“…In this test, 1 mg of dexamethasone, an orally administered corticosteroid medication, is given to the patient at bedtime, and serum cortisol is measured the next morning. 1,4,8,9 This is an example of a unique subset of tests where medications are used to mimic endogenous compounds to elicit expected results. In a healthy patient, the dexamethasone should act like cortisol to suppress CRH and ACTH, which should result in reduced cortisol secretion overnight, and thus a serum cortisol level of < 50 nmol/L the next morning.…”

“…If no suppression occurs, the source is more likely to be autonomous and non-ACTH-dependent. 1,4,8,9 Caution must be used when performing the LDDST, as common medications such as estrogen supplementation (in oral contraceptive pills) can affect total cortisol levels, and drugs such as carbamazepine and rifampin can increase metabolism of dexamethasone itself, such that it insufficiently suppresses cortisol. Use of these medications during the test or in the weeks prior to it could result in an inaccurate test.…”