title: A liquid chromatography-tandem mass spectrometry assay for detection and quantitation of the dipeptide Gly-Gln in rat brain.
abstract: The enzymatic cleavage products of β-endorphin (β-endorphin1-27 and Gly-Gln) reduce voluntary alcohol consumption in alcohol-preferring (P) rats. Gly-Gln also inhibits the reward-benefiting effects of morphine and nicotine. It would be useful for the investigation of these effects to have an analytical method suitable for Gly-Gln detection and quantitation. Given the now widespread availability of liquid chromatography-tandem mass spectrometry (LC-MS/MS) instruments, the development of an LC-MS/MS-based approach seemed a viable option. An LC-MS/MS method for Gly-Gln quantitation was developed based on derivatization with Marfey's reagent. The Marfey's adduct of Gly-Gln (Mar-Gly-Gln) was chromatographically resolved and readily detected and quantitated by LC-MS/MS. Precursor/product positive ions of 456.2/366.2, 456.2/237.2, and 456.2/147.0 were used for detection and quantitation. This method shows good linearity from 1 to 500 pmol of Mar-Gly-Gln (R2 > 0.99). The assay also demonstrated good accuracy and precision, with an average percentage standard deviation for Gly-Gln over the range of the assay of less than 5%. A combination of multiple reaction monitoring (MRM) fragment ratio normalization and chromatographic peak shifting was used to ensure that the LC-MS/MS peak for Mar-Gly-Gln was free from possible isobar interferences. This assay was then demonstrated for the determination of in vivo Gly-Gln levels in P and Sprague-Dawley rat cortex and nucleus accumbens samples.
authors.affiliation: Division of Pharmaceutical Sciences, School of Pharmacy, University of Missouri-Kansas City, Kansas City, MO 64108, USA.
authors.forename: Sudheer
authors.initials: S
authors.lastname: Bobba
authors.forename: Garth E
authors.initials: GE
authors.lastname: Resch
authors.forename: William G
authors.initials: WG
authors.lastname: Gutheil
mesh_terms: D000818:Animals; D001921:Brain / Q000187:drug effects / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D004151:Dipeptides / Q000032:analysis*; D000431:Ethanol / Q000494:pharmacology; D008297:Male; D051381:Rats; D017207:Rats, Sprague-Dawley; D053719:Tandem Mass Spectrometry
chemical_list: D004151:Dipeptides; C039788:glycylglutamine; D000431:Ethanol