Background Eighteen‐hydroxycortisol (18‐OHF) is a potential biomarker for differential diagnosis of the two major primary aldosteronism subtypes, aldosterone‐producing adenoma, and idiopathic hyperaldosteronism. Methods Urine samples were processed, and the 18‐OHF in urine samples were successfully quantified by in‐house established dilute‐and‐shoot liquid chromatography–tandem mass spectrometry (LC–MS/MS) method. Separation was accomplished on a Sigma Ascentis Express C18 column with a gradient mixture of phase (A) 0.2% formic acid in water and phase (B) 0.2% formic acid in methanol at a flow rate of 0.4 ml/min. Mass spectrometric detection was performed in positive electrospray ionization mode via a mass spectrometer. Results The linearity of urinary 18‐OHF ranged from 4.28 to 8.77 × 103 nmol/L, with a lower limit of quantification at 4.28 nmol/L. The intra‐ and inter‐precision were both below 3%. The range of analytical recovery was 97.8%–109.2%. The validated dilute‐and‐shoot LC–MS/MS method was compared with the SPE LC–MS/MS method modified from the one reported in 2013. The results by Passing–Bablok regression analysis and Bland–Altman plotting demonstrated a good agreement between the two methods. The presented method was then applied to establish sex‐specific reference intervals from 62 males and 62 females, respectively. The calculated 2.5%–97.5% reference intervals for 24‐h urinary 18‐OHF were 113–703 nmol/day for males and 71.2–450 nmol/day for females. Conclusion The presented dilute‐and‐shoot LC–MS/MS method for 18‐OHF quantification showed a good performance in the clinical application. Furthermore, the sex‐specific reference intervals for 24‐h urinary 18‐OHF were first established and quite important for its application in primary aldosteronism subtyping.
Urinary and plasma 18-hydroxycortisol (18-OHF) have been investigated for primary aldosteronism (PA) subtyping. However, there is no research exploring the impact of sample types on the diagnostic performance of 18-OHF in PA subtyping. In this study, 18-OHF levels in both urine and plasma were determined in patients with idiopathic adrenal hyperplasia (IHA), aldosterone-producing adenoma (APA), and essential hypertension (EH) by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Urinary18-OHF was determined using an established LC-MS/MS method, whereas plasma18-OHF was measured by a modified LC-MS/MS method. Differences in urinary and plasma 18-OHF levels between APA, IHA, and EH patients were investigated by Kruskal-Wallis test for non-parametric analysis. The LC-MS/MS method yielded a lower limit of quantitation (LLOQ) of 18-OHF in urine of 4.28 nmol/L and 0.190 nmol/L in plasma. The intra- and inter-precision for urine and plasma methods were < 6%, with accuracies between 95.9% and 110.3%. Urinary and plasma 18-OHF in 12 IHA, 18 APA, and 91 EH patients were quantified and analyzed. Non-parametric analysis by Kruskal-Wallis test revealed that urinary 18-OHF levels in IHA and APA patients were significantly different (P < 0.05) while plasma 18-OHF levels were not (P > 0.05), indicating that urinary 18-hydroxycortisol outperformed plasma 18-hydroxycortisol for primary aldosteronism subtyping.
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