A COMBINED LC-MS/MS METHOD FOR THE ANALYSIS OF ALDOSTERONE AND PLASMA RENIN ACTIVITY FOR CLINICAL RESEARCH USING THE XEVO TQ ABSOLUTE MASS SPECTROMETER

Importance of the Topic

The renin–angiotensin–aldosterone system (RAAS) plays a central role in cardiovascular homeostasis by regulating blood pressure and fluid balance. Accurate quantification of aldosterone and plasma renin activity (PRA) is essential for evaluating novel antihypertensive therapies and understanding pathophysiological mechanisms in clinical research.

Objectives and Study Overview

This study aimed to develop and validate a single LC-MS/MS method for simultaneous measurement of aldosterone and PRA in human plasma. The approach was designed to improve efficiency by combining two analyses into one workflow while meeting stringent sensitivity and precision requirements for clinical research applications.

Methodology

Sample preparation involved:

• Thawing 125 µL plasma and incubating with generation buffer at 37 °C for 3 h to convert prorenin to active renin.
• Adding stable isotope internal standards and ammonia for protein precipitation.
• Performing solid-phase extraction on Oasis™ MAX 96-well µElution plates with sequential washes and elutions to recover aldosterone and angiotensin I.
• Sealing, mixing and centrifuging extracts prior to analysis.

Used Instrumentation

• ACQUITY™ UPLC™ I-Class FL system for chromatography.
• XBridge™ Premier BEH™ C18 column (3.2 min run time) with ammonium fluoride and acetonitrile mobile phases.
• Xevo™ TQ Absolute triple quadrupole mass spectrometer with polarity switching in MRM mode.
• Oasis™ MAX µElution plates for extraction.

Main Results and Discussion

Functional sensitivity tests demonstrated signal-to-noise ratios >10:1 at 28 pmol/L aldosterone and 0.08 nmol/L/hr PRA, with precision <20%. Quality control assessments across three concentration levels showed reproducibility and repeatability CVs ≤7.5% for both analytes. External quality assessment (UK NEQAS) of 39 samples gave a Deming fit of y = 0.93x + 2.14 and a mean bias of –6.1% versus scheme LC-MS mean. Method comparison with independent LC-MS/MS assays yielded a Passing-Bablok fit y = 0.96x – 2.49 and Bland–Altman bias of –6.0%, confirming excellent agreement.

Benefits and Practical Applications

This combined LC-MS/MS method reduces sample volume and turnaround time by analyzing aldosterone and PRA in a single run. It offers high sensitivity, robust precision, and compatibility with clinical research workflows. Multiplex capability enhances data richness while lowering operational costs.

Future Trends and Potential Applications

Advances may include full automation of sample preparation, integration with high-throughput platforms, expansion to additional RAAS biomarkers, and adoption of next-generation mass spectrometers for even greater sensitivity. Such developments could improve patient monitoring, personalized therapy assessment, and large-scale epidemiological studies.

Conclusion

The validated LC-MS/MS assay on the Xevo TQ Absolute platform provides a reliable, efficient, and precise approach for simultaneous measurement of aldosterone and PRA. It meets key analytical performance criteria for clinical research and demonstrates strong agreement with existing reference methods.

References

• Foley D., Wardle R., Calton L.J. A Combined LC-MS/MS Method for the Analysis of Aldosterone and Plasma Renin Activity for Clinical Research Using the Xevo TQ Absolute Mass Spectrometer. Waters Corporation; 2023.