Determination of urinary 5-S-cysteinyldopa by high-performance liquid chromatography.

ABSTRACT

A high-performance ion-pair liquid chromatographic method with electrochemical detection is described, which is suitable for routine determination of urinary 5-S-cysteinyldopa. The clean-up procedure includes a first purification step on the cation exchanger AG 50 W (H +). After desorption from the resin at moderately raised pH the catecholic amino acid is adsorbed on alumina at pH 8.6, washed and finally desorbed by elution with perchloric acid. By the combined clean-up procedures, easily oxidized compounds are eliminated, which otherwise cause a number of interfering peaks in the chromatography. The synthesis of 5-S-cysteinyl-L-3,4-dihydroxyphenyl [2,3-3H]alanine is described, and this tritium-labelled 5-S-cysteinyldopa is used to determine the recovery in the sample. The precision (C.V. = 5.7% at low and C.V. = 4.9% at high 5-S-cysteinyldopa concentration) and recovery (105.0 +/- 8.6%) were satisfactory. The mean urinary excretion was 0.34 +/- 0.13 (S.D.) mumol per 24 h (range 0.02-0.58 mumol per 24 h) in healthy subjects (n = 24) and in patients with melanoma metastates (n = 13) the excretion ranged from 0.9 to 4.8 mumol per 24 h.