Establishment and Validation of Sensitive Liquid Chromatography-Tandem Mass Spectrometry Method for Aldosterone Quantification in Feline Serum with Reference Interval Determination

Abstract

Aldosterone, a mineralocorticoid hormone synthesised in the adrenal cortex, is essential for maintaining electrolyte balance and fluid homeostasis. Its role in feline physiology remains underexplored, despite its importance in regulating sodium reabsorption and potassium excretion via mineralocorticoid receptors in renal tubules. This study is warranted given aldosterone's importance in cats, particularly in light of their unique physiological traits, including highly concentrated urine and sensitivity to hydration status. Primary hyperaldosteronism, the most common feline adrenocortical disorder, contributes to arterial hypertension and chronic kidney disease, yet often remains underdiagnosed due to overlapping symptoms like hypokalaemia and hypertension. This research aimed to validate a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to measure serum aldosterone and to establish a reference interval in a population of healthy cats across a broad age range. The method demonstrated high precision and accuracy, with inter-assay coefficients of variation under 15%. Analysis of 49 healthy cats (40 young, 9 old) revealed a reference interval of 5.0-78.4 pg/mL (13.8-217.2 pmol/L). These findings provide a robust framework for diagnosing aldosterone-related disorders in cats and underscore the need for species-specific diagnostic tools. Improved understanding of aldosterone's role could refine treatment strategies and enhance outcomes for affected feline patients.

Keywords: LC-MS/MS; adrenal gland; cat; mineralocorticoid hormone.

Figure 1

The top section of the figure displays the transitions used for aldosterone quantification along with their corresponding qualifiers. Below, the transitions utilised for quantification by the internal D4-aldosterone standard are shown, accompanied by their respective qualifiers. Image obtained from Agilent MassHunter Quantitative Analysis software, version 6.0 (Agilent Technologies, Santa Clara, CA, USA).

Figure 2

The chromatogram illustrates the separation and detection of three analytes along with their corresponding deuterated internal standards. From left to right, the peaks are assigned as follows: at a retention time of 3.6 min, the blue peak corresponds to aldosterone, while the red peak represents D4-aldosterone. At 4.1 min, the pink peak corresponds to cortisone, with the purple peak indicating D8-cortisone. At 4.6 min, the dark green peak denotes cortisol, and the light green peak corresponds to D4-cortisol. Image obtained from Agilent MassHunter Qualitative Analysis software, version 6.0 (Agilent Technologies, Santa Clara, CA, USA).

Figure 3

Distribution and reference intervals for aldosterone with Reference Value Advisor (V2.1) software [18]. CI denotes confidence interval.