# Urinary Normetanephrine for the Diagnosis of Canine Pheochromocytoma via Liquid Chromatography–Mass Spectrometry

## TL;DR

A new urine test using advanced mass spectrometry helps diagnose rare adrenal tumors in dogs by measuring specific hormone metabolites.

## Contribution

Development and validation of a non-invasive LC–MS/MS method for measuring urinary normetanephrine in dogs to improve pheochromocytoma diagnosis.

## Key findings

- Dogs with pheochromocytoma had significantly higher urinary normetanephrine-to-creatinine ratios compared to other groups.
- Using a cut-off of 203.7 nmol/mmol for uNMN:CR achieved 78.6% sensitivity and 83.3% specificity for PHEO detection.
- The LC–MS/MS method demonstrated high diagnostic accuracy with an overall accuracy of 85%.

## Abstract

Pheochromocytomas are rare adrenal tumours in dogs producing catecholamines that derive from the chromaffin cells and often cause non-specific clinical signs, making diagnosis challenging. This study describes the development and validation of a non-invasive urine test using liquid chromatography–tandem mass spectrometry to measure hormone metabolites. Reference ranges were established in healthy dogs, and affected dogs showed higher values than dogs with other conditions. This analytical approach provides accurate and stable results and supports veterinarians in the biochemical identification of pheochromocytoma, improving diagnostic confidence and clinical decision-making.

Pheochromocytomas (PHEO) are rare adrenal medullary tumours characterised by catecholamine secretion. Accurate diagnosis relies on the sensitive measurement of metanephrine (MN) and normetanephrine (NMN); however, the application of liquid chromatography–mass spectrometry (LC–MS/MS) in veterinary medicine remains limited. This study aimed to develop and analytically validate an LC–MS/MS method for the simultaneous quantification of urinary MN and NMN in dogs, establish reference intervals (RIs) for MN- and NMN-to-creatinine ratios (uMN:CR and uNMN:CR), and preliminarily assess the diagnostic utility of uNMN:CR for PHEO detection. The RIs were 2.2–78.9 nmol/mmol for uMN:CR and 4.4–77.4 nmol/mmol for uNMN:CR. Dogs with PHEO showed significantly higher uNMN:CR values compared with dogs with other adrenal masses, non-adrenal disease, or healthy controls. Using the upper RI limit as a threshold, sensitivity and specificity were 91.7% and 35.7%, respectively. Receiver operating characteristic analysis identified an optimal uNMN:CR cut-off of 203.7 nmol/mmol, yielding 78.6% sensitivity, 83.3% specificity, and an overall diagnostic accuracy of 85% (95% CI: 0.68–1.00). These findings support LC–MS/MS as a reliable approach for urinary metanephrine analysis and uNMN:CR as a valuable biochemical tool for PHEO identification in dogs.