A liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based assay for simultaneous quantification of aldosterone, renin activity, and angiotensin II in human plasma.

Abstract Accurate quantification of plasma aldosterone (ALD) and renin activity (PRA) is critical for the diagnosis of primary aldosteronism (PA). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is considered the “gold standard” method for the determination of ALD and PRA. The aim of this study is to develop a new LC-MRM/MS assay for quantifying plasma ALD, PRA, and angiotensin II (Ang II) simultaneously and validate its effectiveness. To be more specific, plasma samples were prepared by solid-phase extraction and separated in an ultra-performance reversed-phase column. MS detection was performed via a triple quadrupole mass spectrometer containing both positive and negative ion monitoring modes. The developed assay was then validated according to the standard guidelines and the influence of sample incubation on ALD and Ang II concentration was evaluated. In addition, the variation of endogenous Ang I was explored. The proposed LC-MRM/MS method was compared another LC-MS/MS method, which detects ALD, Ang I, and Ang II separately. Analytes were separated and quantified within 5 min. The assay was validated to be linear up to 5000 pg/ml for ALD and Ang II and 33.3 ng/ml/h for PRA. The lower limit of quantification (LLOQ) was 15 pg/ml, 15 pg/ml, and 0.1 ng/ml/h for ALD, Ang II, and PRA respectively. Specificity, precision, accuracy, and stability were tested to meet the requirements of the guidelines. Significant changes were not found in ALD and Ang II concentrations over the 3 h-incubation. In addition, it was demonstrated that the result of PRA was not strongly influenced by the endogenous Ang I. Comparison with another LC-MS/MS method was performed using the same apparatus and the proposed method was proved to be in good coincidence with the correlation coefficients ranging from 0.955 to 0.996. A sensitive and reliable method for simultaneous quantification of ALD, PRA, and Ang II has been developed and this study will significantly promote laboratory workflow efficiency and throughput.