Quantitation of Homovanillic Acid (HVA) and Vanillylmandelic Acid (VMA) in Urine Using Gas Chromatography-Mass Spectrometry (GC/MS)

Neuroblastoma, in most cases, is characterized by increased production of catecholamines and their metabolites. Laboratory diagnosis and clinical follow-up include the measurement of urinary homovanillic acid (HVA) and vanillylmandelic acid (VMA). In the following procedure, urine samples are diluted to give a creatinine concentration of 2 mg/dL. Deuterated internal standards are added to the diluted urine samples followed by acidification using HCl. Ethyl acetate is used to extract HVA and VMA from the acidified samples, and the extract is dried. The residue is treated with bis-(trimethylsilyl)trifluoroacetamide (BSTFA), 1% trimethylchlorosilane (TMCS), and pyridine to prepare trimethylsilyl derivatives of HVA and VMA. The derivatized samples are injected to into gas-chromatograph mass spectrometer. The concentration of HVA and VMA is determined by comparing responses of unknown sample to the responses of calibrators using selected ion monitoring.