Development and validation of a method using supported liquid extraction for aldosterone determination in human plasma by LC-MS/MS.

Abstract Background Accurate quantitation of aldosterone is essential for screening, diagnosis and subtype classification in primary aldosteronism. A simple, sensitive method for aldosterone in human plasma using supported liquid extraction (SLE) in combination with liquid chromatography tandem mass spectrometry (LC-MS/MS) was developed and validated. Methods Plasma samples were diluted with water containing d 7 -aldosterone as internal standard. The samples were extracted with methyl-tert-butyl-ether (MTBE) on SLE cartridges. Separation was carried out on a Luna C18 (2) column using a methanol–water gradient. Detection was performed in the negative electrospray multiple reaction monitoring (MRM) quantitation. The use of water-based calibrators was evaluated against calibrators prepared in steroid-free serum. Results The assay was linear up to 3265 pmol/L with an LOQ of approximately 40 pmol/L. Within-run and between-run precision for plasma aldosterone were less than 10% except at low level near LOQ but were still less than 14.7% (Westgard's desirable specification). The mean recovery of the analyte added to plasma was greater than 97.7% and matrix effects were less than 4%. Comparison with another LC-MS/MS method was performed on a more sensitive instrument (ABSciex TQ 5500) and gave the equation API 3000 = 0.957 × TQ 5500 + 12.6, linear regression r 2  = 0.974 ( n  = 43). An estimation of the reference interval for adults was established on a group of healthy volunteers ( n  = 53). Calibration with water-based calibrators was validated and can be used for measurement of aldosterone by LC-MS/MS. Conclusions This method is reliable, easy to perform on plasma specimens in a clinical environment and is attractive because of its simplicity.