Evaluation of specific high-performance liquid-chromatographic determinations of urinary metanephrine and normetanephrine by comparison with isotope dilution mass spectrometry

Abstract

SUMMARY.A method for the determination of metanephrine (MN; also known as metadrenaline), normetanephrine (NMN; also known as normetadrenaline) and 3-methoxytyramine (3-MT) in human urine using high-performance liquid chromatography followed by electrochemical detection (ECD) was validated primarily by comparing the results with those obtained by a gas chromatography±mass spectrometry (GC±MS) reference method. Correlation coef®cients of 0.93, 0.94 and 0.91 were obtained for MN, NMN and 3-MT, respectively, in a group of healthy controls consisting of 30 women and 30 men. A systematic difference was detected only for 3-MT (2 16%). Further tests of accuracy (linearity and recovery) and precision demonstrated that the described method must be considered to be a reliable approach to assess urinary metanephrines in the diagnosis of phaeochromocytoma. At lower concentrations (MN, 248 nmol/L; NMN, 434 nmol/L; 3-MT, 402 nmol/L), within-assay coef®cients of variation were close to 5% or less (5´3, 4´6 and 2´2%, respectively) and between-assay coef®cients of variation were 8´9, 11´2 and 12´3%, respectively, for the same low levels. This raises the possibility that this method can also be applied to assess urinary free, unconjugated metanephrines. Sex differences were detected for MN and NMN excretion when expressed in nmol per 24 h and nmol/mmol creatinine, respectively, by both ECD and GC±MS methods.

Plasma levels of free metanephrines and 3-methoxytyramine indicate a higher number of biochemically active HNPGL than 24-h urinary excretion rates of catecholamines and metabolites

“…These values were for urinary excretion: free norepinephrine 0.06-0.47 mmol/24 h, epinephrine !0.16 mmol/24 h, dopamine 0.46-3.40 mmol/24 h, VMA !30 mmol/24 h (deconjugated), MN (deconjugated) 33-99 mmol/mol creatinine, NMN (deconjugated) 64-260 mmol/mol creatinine, and 3MT (deconjugated) 45-197 mmol/mol creatinine (12).…”

Pheochromocytomas detected by biochemical screening in predisposed subjects are associated with lower prevalence of clinical and biochemical manifestations and smaller tumors than pheochromocytomas detected by signs and symptoms

“…Vanillylic mandelic acid (VMA) in urine was measured using HPLC with fluorometric detection with inter-and intra-assay CV of 2.4-9.1%. Reference ranges (expressed as mmol/24 h) obtained in healthy subjects was 0.06-0.47 for norepinephrine, !0.16 for epinephrine, 0.46-3.40 for dopamine and !30 for VMA (12). In order to ascertain adequacy of collection, the urinary creatinine content was assessed as well.…”

Stability of Urinary Fractionated Metanephrines and Catecholamines during Collection, Shipment, and Storage of Samples

“…Na 2 EDTA and Na 2 S 2 O 5 were obtained from Merck. For measurements of the catecholamines and metanephrines, we used the same reagents and materials as previously described (16,17 ), but the reagent set from Bio-Rad Laboratories (17 ) has been replaced by a newer version.…”

“…Chromatographic data were processed with Empower Pro Workgroup, including Oracle9i ® database Software version 1154. Separation and detection were performed as previously described (16,17 ).…”

“…Analytical characteristics of assay performance have been described (16,17 ). Most relevant to the present experiments were the within-run CVs for measurements of the catecholamines and metanephrines, which were calculated from 24 duplicate measurements of the 1st sample of each volunteer.…”