[Simultaneous determination of tryptophan and its key metabolites by high performance liquid chromatography with...

A high performance liquid chromatographic (HPLC) method with programmed wavelength ultraviolet (UV) detection was established to simultaneously determine tryptophan (Trp) and its key metabolites kynurenine (Kyn) and 5-hydroxytryptamine (5-HT). A BDS-Hypersil-C8 column (150 mm x 4.6 mm, 5 microm) was used for the analysis at 25 degrees C. The separation was carried out with the mobile phase consisting of 10 mmol/L sodium acetate-acetic acid (pH 4. 5) and acetonitrile (94: 6, v/v) using theophylline as internal standard (IS) at a flow rate of 0.6 mL/min. The eluates were monitored by the programmed wavelength UV detection at 360 nm for Kyn and IS, 220 nm for 5-HT and 302 nm for Trp. The mean recoveries were in the range of 87% to 113%. The linearities were from 3.97 to 400 micromol/L for Trp, 0.421 to 20.2 micromol/L for Kyn and 4.36 to 980.5 nmol/L for 5-HT. The detection limits were 0.134 micromol/L for Trp, 0.016 micromol/L for Kyn and 2.03 nmol/L for 5-HT. Fifteen plasma samples of patients with depression and fifteen plasma samples of healthy controls were tested, and the results demonstrated that the metabolism of Trp in the plasma from the depression group was significantly different from that of the control group.