Determination of rat brain kynurenic acid by column-switching HPLC with fluorescence detection

Kynurenic acid (KYNA), one of the tryptophan metabolites, serves as an endogenous antagonist of N-methyl-D-aspartate and the alpha 7 nicotinic receptors in mammalian brains. In the present study, the column-switching high-performance liquid chromatography (HPLC) method vie developed for plasma KYNA was extended and validated for the determination of brain KYNA. Rat cerebrum, cerebellum and brainstem homogenates were deproteinized with acetone, and the extracts reconstituted with the mobile phase were injected onto the HPLC. In spite of the facile pretreatment, the fluorescence peak of KYNA in the cerebrum, cerebellum and brainstem was clearly observed with no interfering peaks. Intra- and inter-day precisions [relative standard deviation (%)] and accuracies [relative mean error (%)] were satisfactory (< +/- 5.8%). The concentrations of KYNA in rat cerebrum, cerebellum, and brainstem were 224 +/- 65.8, 606 +/- 191, and 323 +/- 114 fmol/mg protein (n = 5), respectively. The proposed HPLC method will be a useful tool for pharmacokinetic and pharmacological researches on brain KYNA. Copyright (c) 2007 John Wiley & Sons, Ltd.