Analysis of glutamine, glutamate, pyroglutamate, and GABA in cerebrospinal fluid using ion pairing HPLC with positive electrospray LC/MS/MS.
A simple and sensitive method for the separation and quantitation of glutamine, glutamate, pyroglutamate, and gamma-aminobutyric acid (GABA) in cerebrospinal fluid (CSF) is presented. The method utilizes ion pairing with heptafluorobutyric acid (HFBA) to achieve HPLC separation with detection by positive ESI LC/MS/MS. The method does not require extraction or derivatization, utilizes a heavy labeled internal standard for each analyte, and allows for rapid throughput with a 5 min run time. The method was developed with particular attention taken to prevent conversion between analytes known to occur under certain conditions. The lower limit of quantitation is 7.8 ng/ml for all analytes, and the intra-day and inter-day accuracy (%RE) and precision (%R.S.D.) are defined for all analytes. The method was developed as a sensitive, selective, and robust method to investigate the excitatory and inhibitory neurotransmitters (glutamate and GABA) as biomarkers in drug development.
Eckstein
James A
JA
Eli Lilly and Company, Greenfield Laboratories, Greenfield, IN 46140, USA. Eckstein_Jim_A@Lilly.com
Ammerman
Gina M
GM
Reveles
Jessica M
JM
Ackermann
Bradley L
BL
D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D018698:Glutamic Acid / Q000134:cerebrospinal fluid*; D005973:Glutamine / Q000134:cerebrospinal fluid*; D006801:Humans; D011761:Pyrrolidonecarboxylic Acid / Q000134:cerebrospinal fluid*; D051381:Rats; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D005680:gamma-Aminobutyric Acid / Q000134:cerebrospinal fluid*
D005973:Glutamine; D018698:Glutamic Acid; D005680:gamma-Aminobutyric Acid; D011761:Pyrrolidonecarboxylic Acid