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J Pharm Biomed Anal. 2015 Mar 25;107:426-31. doi: 10.1016/j.jpba.2015.01.030. Epub 2015 Jan 24.

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

Author information

1
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: l.orsatti@irbm.it.
2
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: r.speziale@irbm.it.
3
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: m.orsale@irbm.it.
4
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: f.caretti@irbm.it.
5
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: m.veneziano@irbm.it.
6
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: m.zini@irbm.it.
7
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: e.monteagudo@irbm.it.
8
Freelance Consultant, P.O. Box 64, Holland, NY, USA. Electronic address: kathyalyons@gmail.com.
9
Retrophin Inc. (RTRX), 301 Binney Street, 3rd Floor, Cambridge, MA 02142, USA. Electronic address: Maria.Beconi@retrophin.com.
10
Alcon Research, Ltd., 6201 South Freeway, Fort Worth, TX 76134-2099, USA. Electronic address: kelvinchan2014@verizon.net.
11
CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: todd.herbst@chdifoundation.org.
12
CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: leticia.toledosherman@chdifoundation.org.
13
CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: ignacio.munoz@chdifoundation.org.
14
IRBM Science Park, Via Pontina km 30,600, 00040 Pomezia (Roma), Italy. Electronic address: f.bonelli@irbm.it.
15
CHDI Management/CHDI Foundation, 6080 Center Drive, Los Angeles, CA, USA. Electronic address: celia.dominguez@chdifoundation.org.

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%).

KEYWORDS:

Kynurenine; Liquid chromatography; Mass spectrometry; Plasma; Tryptophan metabolism

PMID:
25668794
DOI:
10.1016/j.jpba.2015.01.030
[Indexed for MEDLINE]

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