Measurement of urinary metadrenaline and normetadrenaline by liquid chromatography tandem mass spectrometry for the diagnosis of phaeochromocytoma.
BACKGROUND
Measurement of metadrenalines has been recommended in the investigation of phaeochromocytoma. Urinary assays remain the most common; however, drug interference is still one of the main challenges for analytical systems. We have developed a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the quantification of total urinary normetadrenaline and total urinary metadrenaline, which does not require extraction procedures prior to analysis.


METHODS
Total urinary normetadrenaline and total urinary metadrenaline were individually quantified by electrospray ionization in the multiple-reaction monitoring mode. Deuterated internal standards were used and an acid hydrolysis step was used to convert conjugated metabolites into free metadrenalines. Chromatographic separation was achieved using a Phenomenex Luna 3 micro PFP column followed by analysis on an API 3200 LC-MS/MS.


RESULTS
Linearity was exhibited across the calibration range for both normetadrenaline (r = 1, P < 0.0001) and metadrenaline (r = 1, P < 0.0001) with the limit of quantification of 0.05 and 0.02 micromol/L, respectively. Intra-assay imprecision for both normetadrenaline and metadrenaline was less than 5.5% with % coefficient of variations of less than 4%. Inter-assay imprecision was less than 13%. Neither noradrenaline or adrenaline interfere with the assay as determined by the spiking of samples with high concentrations of noradrenaline or adrenaline (P > 0.05). Acceptable analytical performance was seen with comparison to a high-performance liquid chromatography method and on External Quality Assessment returns.


CONCLUSIONS
An analytically simple and sensitive method has been developed and evaluated for the analysis of total urinary normetadrenaline and total urinary metadrenaline which is now in routine use.
Marrington
Rachel
R
Department of Biochemistry and Immunology, Birmingham Heart of England NHS Foundation Trust, Bordesley Green East, Birmingham B95SS, UK. rachel.marrington@heartofengland.nhs.uk
Johnston
Jane
J
Knowles
Sarah
S
Webster
Craig
C
D000310:Adrenal Gland Neoplasms / Q000175:diagnosis* / Q000652:urine; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D016014:Linear Models; D008676:Metanephrine / Q000652:urine*; D009647:Normetanephrine / Q000652:urine*; D010673:Pheochromocytoma / Q000175:diagnosis* / Q000652:urine; D053719:Tandem Mass Spectrometry / Q000379:methods*; D016482:Urinalysis / Q000379:methods*
D009647:Normetanephrine; D008676:Metanephrine