Thin-layer chromatography-an image-processing method for the determination of acidic catecholamine metabolites

摘要

A sensitive and convenient method for acidic catecholamine metabolites (including homovanillic acid, vanillylmandelic acid, 3,4-dihydroxymandelic acid, and 3,4-dihydroxyphenylacetic acid) determination was developed based on thin-layer chromatography and image-processing analysis. The metabolites were separated without a prederivatization step using reversed phase RP-18W high-performance plates. The mobile phase composition, detection, and quantification conditions were systematically investigated through several trials. The reaction with 2,2-diphenyl-1-picrylhydrazyl radical allowed specific detection of acidic catecholamine metabolites with a high sensitivity and a wide linear range. The limit of detection and the limit of quantification were in the range of 13-103 and 18-120 ng/spot, respectively, in all cases. Mean recoveries determined were in the range 95-106% for all of the investigated compounds. The proposed method allowed rapid simultaneous determination of acidic catecholamine metabolites from spiked human urine sample.