Published: Journal of Analytical Toxicology, ISSN 0146-4760, Volume 31, Number 9, November/December, pp.555-565

Gabapentin in Horses: Validation of an Analytical Method for Gabapentin Quantitation
A.F. Lehner[1], J. Stewart[1], A. Dafalla[2], K.J. Ely[2], A.L. Connerly[2], C.N. Jones[2], H. ElkHoly[2], K. Thompson[3], T. Tobin[4], and L. Dirikolu[5],
[1]Livestock Disease and Diagnostic Center, College of Agriculture, University of Kentucky, Lexington, Kentucky 40512-4125;
[2]Department of Biomedical Sciences, College of Veterinary Medicine, Nursing and Allied Health, Tuskegee University, Tuskegee, Alabama, 36088;
[3]Department of Large Animal Clinics, College of Veterinary Medicine, Nursing and Allied Health, Tuskegee University, Tuskegee, Alabama, 36088;
[4]128C Maxwell Gluck Equine Research Center, Department of Veterinary Science, University of Kentucky, Lexington, Kentucky 40546-0099; and
[5]Department of Veterinary Biosciences, University of Illinois, College of Veterinary Medicine, 3830 VMBSB, 2001 S. Lincoln Avenue, Urbana, Illinois 61802

Gabapentin [1-(aminomethyl)cyclohexaneacetic acid, Neurontin“], is a new anticonvulsant used as adjunctive therapy in the treatment of partial seizures in humans not controlled with standard antiseizure drugs, and it has also been used in veterinary medicine. In performance horses, gabapentin is listed as a class 3 performance-enhancing substance by the Association of Racing Commissioners International, and thus is considered to have the potential to influence the outcome of races. Therefore, we developed and validated a sensitive gas chromatographic–mass spectrometric (GC–MS) method for gabapentin detection. Gamma-aminobutyric acid-d2 (GABA-d2) was used as an internal standard during solid-phase extraction; lacking the cyclohexyl ring of gabapentin, GABA-d2 formed a lactam structure to only a minor extent. Gabapentin, on the other hand, readily formed a lactam on thermal exposure during trimethylsilyl-derivatization and/or GC analysis; electrospray-ionization MS was employed to verify that the original compound was present as the expected 171 m.w. compound. Extraction efficiency for the assay was about 60%, and a curvilinear standard curve ranging from 50 ng/mL to 3000 ng/mL provided excellent within-run and between-run coefficients of variation and accuracies over a range of low, medium, and high values. The limit of detection, defined as the concentration calculated from the mean response at zero concentration plus two times the standard deviation, was calculated at 7.6 ng/mL; the limit of quantitation, defined as the concentration calculated from the mean of the zero responses plus five times the standard deviation, was calculated at 17 ng/mL. This method will enable accurate quantification of gabapentin in equine biological fluids for use in both pharmacokinetic and forensic studies.

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