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Clinical Chemistry 50: 603-611, 2004. First published January 15, 2004; 10.1373/clinchem.2003.024703
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Right arrow Endocrinology and Metabolism
(Clinical Chemistry. 2004;50:603-611.)
© 2004 American Association for Clinical Chemistry, Inc.

Endocrinology and Metabolism

Measurement of Plasma Free Metanephrine and Normetanephrine by Liquid Chromatography–Tandem Mass Spectrometry for Diagnosis of Pheochromocytoma

Susan A. Lagerstedt1, Dennis J. O’Kane1 and Ravinder J. Singh1,a

1 Department of Laboratory Medicine & Pathology, Mayo Clinic and Foundation, 200 First Street SW, Rochester, MN 55905.

aAuthor for correspondence. Fax 507-284-9758; e-mail singh.ravinder{at}mayo.edu.

Background: Quantification of plasma free metanephrines is usually accomplished by HPLC with electrochemical detection, but sample preparation is labor-intensive and time-consuming, run times are long, and interfering substances sometimes obscure the relevant peaks. The aim of this study was to develop a sensitive and specific LC-MS/MS method for plasma free metanephrines.

Methods: After solid-phase extraction, chromatographic separation of normetanephrine (NMN) and metanephrine (MN) was accomplished by use of a cyano analytical column. NMN, MN, d3-NMN, and d3-MN positive ions were detected in the multiple-reaction monitoring mode using the specific transitions m/z 166->134, 180->148, 169->137, and 183->151, respectively, with an atmospheric pressure chemical ionization source.

Results: Multiple calibration curves exhibited consistent linearity and reproducibility. Interassay imprecision values (CV; n = 20) for NMN at 0.64, 1.9, and 2.7 nmol/L were 6.6%, 7.8%, and 13%, respectively. Interassay CV for MN at 0.60, 1.2, and 2.1 nmol/L (n = 20) were 9.2%, 6.8%, and 9.8%, respectively. The mean recoveries of NMN and MN relative to the internal standard were 100% and 96%, respectively. The assays were linear between 0.20 and 10.0 nmol/L. Deming regression of HPLC and LC-MS/MS results yielded slopes of 0.93 (95% confidence interval, 0.89–0.98) and 0.89 (0.85–0.93) and y-intercepts of -0.16 and 0.03 nmol/L for NMN (n = 132) and MN (n = 92), respectively.

Conclusions: This novel LC-MS/MS approach provides a precise, rapid, and specific alternative method to HPLC for the quantification of the low nanomolar concentrations of free metanephrines in plasma.




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