Simultaneous Determination of Kynurenine and Tryptophan in Serum by High Performance Liquid Chromatography

A method was established for the simultaneous determination of kynurenine (Kyn) and tryptophan (Trp) in serum by high performance liquid chromatography-ultraviolet detection (HPLC-UV). It employed a Symmetry Shield RP-C18 column (150 mm×3.9 mm i.d., 5 μm) and a mobile phase of 15 mmol/L sodium acetate-acetic acid solution containing 2.7% (v/v) acetonitrile (pH 3.6) at a flow rate of 1.0 mL/min. The ultraviolet detector was operated at 225 nm. Serum samples were first precipitated with a 5.0% perchloric acid solution, then centrifuged to remove protein residue and finally analyzed by HPLC. The retention time of Kyn was 3.5 min, the linear range of the method was from 0.098 to 49 μmol/L, and the detection limit was 0.02 μmol/L. The recoveries of Kyn were from 90.82% to 93.45%; the intra-day and inter-day variations were 2.37% and 3.66%, respectively. The retention time of Trp was 8.1 min, the linear range of the method was from 4.9 to 490 μmol/L, and the detection limit was 0.20 μmol/L. The recoveries of Trp were from 95.51% to 98.67%; the intra-day and inter-day variations were 1.50% and 2.65%, respectively. The method is simple, fast, accurate, and suitable for routine analysis.