Direct concurrent measurement of urinary vanillylmandelic acid, 5-hydroxyindoleacetic acid and homovanillic acid by HPLC. Three methodologies compared

Three different direct HPLC methods for the determination of 3-methoxy-4-hydroxymandelic acid (VMA, vanillylmandelic acid), 5-hydroxyindoleacetic acid (5-HIAA) and 3-methoxy-4-hydroxyphenylacetic acid (HVA, homovanillic acid) in urine were compared: two spectrofluorometric methods, applying discontinuous gradients, and one serial coulometric linear gradient method. The imprecision study (n = 6) revealed comparable coefficients of variation (CV), intra-assay ranging 1.4-11.1%, and inter-assay ranging 5.9-11.8% for physiological and moderately elevated levels of VMA, 5-HIAA and HVA. All methods showed good linearities up to 100 mumol/L for each of the three compounds studied. Analytical recoveries were 97-114% for VMA, 87-103% for 5-HIAA, and 80-95% for HVA. Recoveries were not dependent on urinary relative densities in the range 1.010-1.030 kg/L or on protein content (prior to acidification) in the range 0.1-3 g/L, or on the pH of conservation in the range 2-5 or on storage temperature in the range -20 - +22 degrees C for three weeks. The distributed-sample comparison revealed acceptable correlations and clinically unimportant accuracy differences between the methods. It is concluded that direct fluorometric and electrochemical HPLC methods can be used in the determination of major catecholamine and serotonin metabolites in human urine for clinical diagnosis and follow-up of neural crest and carcinoid tumours.