HOW LOW CAN WE GO? ANALYSIS OF ALDOSTERONE USING A HIGHLY ANALYTICALLY SENSITIVE TANDEM QUADRUPOLE MASS SPECTROMETER FOR CLINICAL RESEARCH

Importance of the Topic

Aldosterone is a key mineralocorticoid hormone responsible for maintaining sodium/potassium balance and blood pressure. Accurate measurement at low concentrations is critical in clinical research on aldosterone synthase (CYP11B2) inhibitors. Traditional immunoassays can suffer cross-reactivity with structurally related steroids, whereas LC-MS/MS offers greater selectivity and sensitivity.

Study Objectives and Overview

This study compared the performance of a conventional Xevo TQ-S tandem quadrupole mass spectrometer and the new Xevo TQ-XS system, both coupled to UPLC, for quantifying aldosterone in human plasma down to 2 pg/mL using 200 µL sample volume. It also evaluated the novel UniSpray ionization against electrospray ionization (ESI) for enhancing analytical sensitivity.

Methodology

Sample Preparation:

• Automated on a Tecan Freedom EVO 100 liquid handler.
• Protein precipitation with zinc sulfate in methanol and 0.05 % phosphoric acid.
• SPE cleanup using Waters Oasis MAX µElution plates.

Chromatography:

• Waters ACQUITY UPLC I-Class system.
• CORTECS UPLC C18 column (2.1×100 mm).
• Water/methanol gradient at 0.3 mL/min.

Used Instrumentation

• Waters Xevo TQ-S and Xevo TQ-XS tandem quadrupole MS.
• UniSpray source with grounded probe and Coanda-effect gas flow.
• Waters ACQUITY UPLC I-Class.
• Tecan Freedom EVO 100.

Main Results and Discussion

Analytical Sensitivity and Linearity:

• LLOQ at 1 pg/mL (2.8 pmol/L) with S/N ≥10 on Xevo TQ-XS (350 fg on column).
• Over 5-fold S/N improvement and over 6-fold peak area increase versus Xevo TQ-S.
• Calibration range 15–1500 pg/mL (42–4161 pmol/L), r² > 0.9999.

Precision and Accuracy:

• QC levels at 21, 42, 174 and 669 pg/mL showed total precision ≤7.9 % RSD on Xevo TQ-XS versus ≤8.2 % on Xevo TQ-S.

System Comparison:

• Deming regression between systems: y = 0.98x – 5.90, no significant bias (p > 0.05).
• Altman-Bland mean bias of –4.2 %.
• Linear correlation r = 0.999.

UniSpray Evaluation:

• Provided >3× increase in S/N at 2 pg/mL versus ESI.
• Peak area reproducibility <20 % RSD at 60 fg on column.

Benefits and Practical Applications

• Enables reliable low-level aldosterone quantification for pharmacokinetic and pharmacodynamic studies.
• Reduces sample volume requirements and increases confidence in data at trace concentrations.
• UniSpray may lower maintenance downtime and extend analyte coverage.

Future Trends and Applications

• Broader validation of UniSpray for diverse steroid panels.
• Integration of high-throughput workflows for clinical bioanalysis.
• Application to additional low-abundance hormones and metabolites.
• Advancements in automation and miniaturized sample prep.

Conclusion

The Waters Xevo TQ-XS coupled to UPLC demonstrates superior sensitivity, precision and linearity for plasma aldosterone analysis compared to the Xevo TQ-S. UniSpray ionization shows promising gains in signal and robustness, offering a powerful platform for clinical research requiring low-level steroid quantification.

References

1. Häggström M, Richfield D. Diagram of the pathways of human steroidogenesis. Wikiversity Journal of Medicine. 2014;1(1).