Fukushima Takeshi, Mitsuhashi Shogo, Tomiya Masayuki, Kawai Junko, Hashimoto Kenji, Toyo'oka Toshimasa
Division of Bio-Analytical Chemistry, School of Pharmaceutical Sciences, University of Shizuoka, 52-1 Yada, Shizuoka 422-8526, Japan.
Biomed Chromatogr. 2007 May;21(5):514-9. doi: 10.1002/bmc.786.
Kynurenic acid (KYNA), one of the tryptophan metabolites, serves as an endogenous antagonist of N-methyl-d-aspartate and the alpha7 nicotinic receptors in mammalian brains. In the present study, the column-switching high-performance liquid chromatography (HPLC) method we 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.
犬尿喹啉酸（KYNA）是色氨酸代谢产物之一，在哺乳动物大脑中作为N-甲基-D-天冬氨酸和α7烟碱受体的内源性拮抗剂。在本研究中，我们开发的用于测定血浆中KYNA的柱切换高效液相色谱（HPLC）方法得以扩展并经过验证，用于测定大脑中的KYNA。大鼠大脑、小脑和脑干匀浆用丙酮进行脱蛋白处理，用流动相复溶后的提取物注入HPLC。尽管预处理简便，但在大脑、小脑和脑干中仍能清晰观察到KYNA的荧光峰，且无干扰峰。日内和日间精密度[相对标准偏差（%）]和准确度[相对平均误差（%）]均令人满意（<±5.8%）。大鼠大脑、小脑和脑干中KYNA的浓度分别为224±65.8、606±191和323±114 fmol/mg蛋白质（n = 5）。所提出的HPLC方法将成为大脑中KYNA的药代动力学和药理学研究的有用工具。
