Diazotization of catecholamines and their analogs and metabolites for urinary screening tests: chemical aspects

Coupling of diazotized p-nitroaniline to catecholamines and their metabolites in urine has been proposed for use in screening for secreting neuroblastoma in childhood. We have coupled diazotized p-nitroaniline to catecholamines, derivatives, and metabolites and examined the reaction products by thin-layer chromatography and physico-chemical methods (ultraviolet spectra, mass spectroscopy, nuclear magnetic resonance). We conclude that during diazotization, products containing a p-hydroxybenzyl alcohol or a p-hydroxybenzoic acid structure (e.g., vanillic acid, vanilmandelic acid, 3-methoxy-4-hydroxyphenyl-ethyleneglycol, metanephrine, normetanephrine, synephrine, and isoproterenol) react with a diazonium cation, with release of their alcohol or acid moiety. Therefore the mentioned qualitative screening methods are very nonspecific. In contrast, thin-layer chromatographic screening methods provide complete separation and unambiguous identification of those metabolites and are to be preferred for use in detecting secreting neuroblastoma in childhood.