title: A single-run liquid chromatography mass spectrometry method to quantify neuroactive kynurenine pathway metabolites in rat plasma.
abstract: Neuroactive metabolites in the kynurenine pathway of tryptophan catabolism are associated 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 association of tryptophan catabolic pathway alterations with various central nervous system (CNS) pathologies has raised interest in analytical 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-chemical properties, major efforts were devoted to develop a chromatography suitable for all metabolites that involves plasma protein precipitation with acetonitrile followed by chromatographic separation by C18 RP chromatography, 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 standards and QC accuracy within ±30%).
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: l.orsatti@irbm.it.
authors: Laura
authors: L
authors: Orsatti
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: r.speziale@irbm.it.
authors: Roberto
authors: R
authors: Speziale
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: m.orsale@irbm.it.
authors: Maria Vittoria
authors: MV
authors: Orsale
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: f.caretti@irbm.it.
authors: Fulvia
authors: F
authors: Caretti
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: m.veneziano@irbm.it.
authors: Maria
authors: M
authors: Veneziano
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: m.zini@irbm.it.
authors: Matteo
authors: M
authors: Zini
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: e.monteagudo@irbm.it.
authors: Edith
authors: E
authors: Monteagudo
authors: Freelance Consultant, P.O. Box 64, Holland, NY, USA. Electronic address: kathyalyons@gmail.com.
authors: Kathryn
authors: K
authors: Lyons
authors: Retrophin Inc. (RTRX), 301 Binney Street, 3rd Floor, Cambridge, MA 02142, USA. Electronic address: Maria.Beconi@retrophin.com.
authors: Maria
authors: M
authors: Beconi
authors: Alcon Research, Ltd., 6201 South Freeway, Fort Worth, TX 76134-2099, USA. Electronic address: kelvinchan2014@verizon.net.
authors: Kelvin
authors: K
authors: Chan
authors: CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: todd.herbst@chdifoundation.org.
authors: Todd
authors: T
authors: Herbst
authors: CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: leticia.toledosherman@chdifoundation.org.
authors: Leticia
authors: L
authors: Toledo-Sherman
authors: CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: ignacio.munoz@chdifoundation.org.
authors: Ignacio
authors: I
authors: Munoz-Sanjuan
authors: IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: f.bonelli@irbm.it.
authors: Fabio
authors: F
authors: Bonelli
authors: CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: celia.dominguez@chdifoundation.org.
authors: Celia
authors: C
authors: Dominguez
mesh_terms: D000818:Animals; D001812:Blood-Brain Barrier / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D007736:Kynurenic Acid / Q000097:blood / Q000737:chemistry; D007737:Kynurenine / Q000097:blood / Q000737:chemistry* / Q000378:metabolism*; D010949:Plasma / Q000737:chemistry*; D051381:Rats; D014364:Tryptophan / Q000097:blood / Q000737:chemistry; D062367:ortho-Aminobenzoates / Q000097:blood / Q000737:chemistry
chemical_list: D062367:ortho-Aminobenzoates; C031385:anthranilic acid; D007737:Kynurenine; D014364:Tryptophan; D007736:Kynurenic Acid
keywords: Kynurenine; Liquid chromatography; Mass spectrometry; Plasma; Tryptophan metabolism