Simultaneous determination of tryptophan, kynurenine and 5-hydroxytryptamine by HPLC: Application in uremic patients undergoing hemodialysis

Abstract

Objectives: To develop a reliable HPLC method for the simultaneous determination of plasma tryptophan, kynurenine and 5-hydroxytryptamine to analyze tryptophan metabolism.

Design and methods: Separation was carried out on a C8 column with the mobile phase composed of acetate buffer (pH 4.5) and acetonitrile using theophylline as internal standard. The eluates were monitored by ultraviolet detection with programmed wavelength.

Results: Analysis was achieved in less than 8.0min. The limits of quantification were 3.97μmol/L, 4.36nmol/L and 0.421μmol/L for tryptophan, 5-hydroxytryptamine and kynurenine, respectively. Reproducibility and recovery were satisfactory. Twenty healthy adults and 20 uremic patients undergoing hemodialysis were analyzed using the present method. Tryptophan metabolism was found to be disturbed in uremic patients and was improved obviously after hemodialysis.

Conclusions: The developed HPLC method is simple, reliable and suitable for monitoring tryptophan metabolism in uremic patients undergoing hemodialysis.