Determination of catecholamines and methoxycatecholamines excretion patterns in pig and rat urine by ion-exchange liquid chromatography with electrochemical detection

Abstract

A simplified liquid chromatographic method for the simultaneous determination of free or total catecholamines and methoxycatecholamines in rat and pig urine is presented. The extraction procedure involves a two-stage batch extraction, with successive adsorption on cation- (catecholamine elution) and anion-exchange columns (methoxycatecholamine elution). The column eluates are successively monitored by reversed-phase high-performance liquid chromatography (HPLC) with electrochemical detection. The proportion of conjugates for each compound was assessed in both species, through the comparison of concentrations with or without hydrolysis pretreatment. Conjugates were found to account for a small fraction of total catecholamines and methoxycatecholamines excretion (0 to 35%). The free fraction of each compound was highly correlated with the total amount. Furthermore, the hydrolysis procedure leads to partial degradation of metanephrine (25%) and to the production of compounds giving artefactual peaks. Thus, we do not recommend hydrolysis of rat and pig urines for catecholamine and methoxycatecholamine determination.