A single-run liquid chromatography mass spectrometry method to quantify neuroactive kynurenine pathway metabolites in rat plasma.

By Orsatti, Laura; Speziale, Roberto; Orsale, Maria Vittoria; Caretti, Fulvia; Veneziano, Maria; Zini, Matteo; Monteagudo, Edith; Lyons, Kathryn; Beconi, Maria; Chan, Kelvin; et al
From Journal of Pharmaceutical and Biomedical Analysis (2015), 107, 426-431. Language: English, Database: CAPLUS, DOI:10.1016/j.jpba.2015.01.030

Neuroactive metabolites in the kynurenine pathway of tryptophan catabolism are assocd. with neurodegenerative disorders. Tryptophan is transported across the blood-brain barrier and converted via the kynurenine pathway to N-formyl-L-kynurenine, which is further degraded to L-kynurenine. This metabolite can then generate a group of metabolites called kynurenines, most of which have neuroactive properties.The assocn. of tryptophan catabolic pathway alterations with various central nervous system (CNS) pathologies has raised interest in anal. methods to accurately quantify kynurenines in body fluids. We here describe a rapid and sensitive reverse-phase HPLC-MS/MS method to quantify L-kynurenine (KYN), kynurenic acid (KYNA), 3-hydroxy-L-kynurenine (3HK) and anthranilic acid (AA) in rat plasma. Our goal was to quantify these metabolites in a single run; given their different physico-chem. properties, major efforts were devoted to develop a chromatog. suitable for all metabolites that involves plasma protein pptn. with acetonitrile followed by chromatog. sepn. by C18 RP chromatog., detected by electrospray mass spectrometry. Quantitation range was 0.098-100 ng/mL for 3HK, 9.8-20,000 ng/mL for KYN, 0.49-1000 ng/mL for KYNA and AA. The method was linear (r > 0.9963) and validation parameters were within acceptance range (calibration stds. and QC accuracy within ±30%).