High Sensitivity Analysis of Aldosterone in Human Serum Using LCMS™-8060NX

Significance of the Topic

Aldosterone is a key mineralocorticoid regulating electrolyte balance, blood volume and pressure. Accurate measurement of its serum levels is critical in diagnosing conditions such as primary hyperaldosteronism, diabetic nephropathy and interstitial nephritis. Conventional immunoassays may overestimate concentrations due to cross-reactivity with structurally similar steroids. High-selectivity LC-MS/MS methods overcome these limitations and enable reliable quantitation at ultralow levels.

Study Objectives and Overview

This work demonstrates a high-sensitivity assay for aldosterone in human serum using the Shimadzu LCMS-8060NX triple quadrupole mass spectrometer. Calibration standards ranged from 0.5 to 10 000 pg/mL. Key goals included establishing linearity, precision and accuracy at the lower limit of quantification in both neat and serum-spiked samples.

Sample Preparation

Standard solutions were serially diluted in 50 % methanol to yield 0.5-10 000 pg/mL calibration levels. Steroid-free human serum was spiked at 1 pg/mL aldosterone. Each aliquot (100 µL) was combined with internal standard, extracted with hexane:ethyl acetate (9:1), centrifuged for 10 min, the organic layer evaporated and residues reconstituted in 50 % methanol for injection.

Analytical Methodology

• HPLC: Shim-pack Scepter C18-120 (50 × 2.1 mm, 1.9 µm), mobile phase A = 0.05 mM NH4F in water, B = methanol; gradient elution; flow rate 0.3 mL/min; injection volume 25 µL.
• MS: ESI positive mode, MRM transitions m/z 361.20→343.30 for aldosterone and m/z 365.25→346.25 for the D4 internal standard; interface voltage 1 kV; ion focus 3.5 kV; nebulizing gas 3 L/min; drying gas 10 L/min; heating gas 5 L/min; DL temp 250 °C; block heater 300 °C; interface 400 °C.

Results and Discussion

The calibration curve exhibited excellent linearity over 0.5-10 000 pg/mL (R2 = 0.9998). The %RSD at the LOQ (0.5 pg/mL) was 3.96 %, with accuracy across calibration points between 95.6 % and 109.8 %. Serum samples spiked at 1 pg/mL yielded a mean recovery of 106 % (n=3). Representative MRM chromatograms confirmed clear peak separation and signal-to-noise ratios suitable for trace quantitation.

Practical Benefits and Applications

This LC-MS/MS assay delivers high sensitivity and selectivity for aldosterone, enabling detection at sub-pg/mL levels and robust performance in complex serum matrices. It is well suited for clinical research, diagnostic support and quality control in pharmaceutical or clinical laboratories.

Future Trends and Applications

• Integration into multiplex steroid panels for comprehensive endocrine profiling.
• Automation and high-throughput workflows to support routine clinical diagnostics.
• Application to biomarker discovery and metabolic studies requiring ultra-trace quantitation.

Conclusion

The Shimadzu LCMS-8060NX method offers a reliable, sensitive and accurate platform for aldosterone quantitation in human serum across a broad dynamic range. Its performance in precision, linearity and recovery underscores its applicability in both research and diagnostic settings.

Instrumentation Used

• Shimadzu LCMS-8060NX triple quadrupole mass spectrometer
• Nexera X3 UHPLC system
• Shim-pack Scepter C18-120 column (50 × 2.1 mm, 1.9 µm)