An automated magnetic-bead-assisted liquid chromatography-tandem mass spectrometry method for analyzing angiotensins, ALD, and 18-OH derivatives: Application of primary aldosteronism early screening and subtyping.

Angiotensin I and II (Ang I, Ang II), aldosterone (ALD), 18-hydroxycorticosterone (18-OHB), 18-hydroxycortisol (18-OHF), and 18-oxocortisol (18-OXOF) are critical biomarkers for the screening, confirming, and subtyping of primary aldosteronism (PA). Liquid chromatography-tandem mass spectrometry (LC-MS/MS) enables simultaneous quantification of multiple analytes; however, obstacles such as labor-intensive manual procedures, complex biological matrices, structural analog interference, and the physicochemical diversity of target analytes limit its application and promotion in clinical laboratories. This study established and validated a robust LC-MS/MS method based on automated magnetic-bead-assisted sequential extraction (MBASE) for the simultaneous detection and quantitation of Ang I, Ang II, ALD, 18-OHB, 18-OHF, and 18-OXOF. The method achieved complete chromatographic resolution with no interference from endogenous hormones or exogenous medications. The analytical recoveries ranged from 86.14 % to 112.89 %, with total imprecisions between 2.93 % and 12.89 %. The MBASE demonstrated a strong correlation with traditional SPE for all six analytes (R: 0.967-0.997), with biases ranging from -4.0 % to 5.1 %. Compared to traditional SPE process, the MBASE workflow reduced total processing time by 46 % and manual handling time by 77 %. The clinical application of this LC-MS/MS method indicated that Ang I and Ang II are effective screening biomarkers, and ALD, 18-OHB, 18-OHF, and 18-OXOF could be used to differentiate PA subtypes. Overall, the newly developed MBASE-LC-MS/MS method exhibits excellent performance and significant advantages for PA diagnosis. It can be a promising tool in the early screening and subtyping for PA and enhance automated mass spectrometry processes in clinical application.