Estimation of catecholamines by ion-exchange chromatography on Asahipak ES-502C, using glycylglycine as the post-derivatizing agent.

The estimation of catecholamines in human urine was carried out by ion-exchange chromatography on a column of a weakly acidic ion exchanger with an hydrophilic matrix. The catecholamines were first adsorbed onto Amberlite CG-50 (buffered at pH 6.5 with 0.4 M phosphate buffer) and selectively eluted by 0.66 M boric acid solution. They were then separated from impurities that responded to fluorometric detection by isocratic elution from a column of Asahipak ES-502C, a cross-linked vinyl alcohol copolymer with carboxymethyl groups, at 60 degrees C. The mobile phase was 0.05 M sodium succinate buffer pH 5.25 containing 0.015 M borate and 0.5 mM ethylenediaminetetraacetate. Isoproterenol was used as the internal standard; epinephrine, norepinephrine, isoproterenol and dopamine were eluted in this order. One sample could be analyzed every 35 min. The detection limits were 0.2 ng for epinephrine and norepinephrine, 0.6 ng for dopamine. The elution pattern was quite reproducible; the elution volumes of the catecholamines had not changed after 500 determinations.