Development and Validation of a Rapid LC-MS/MS based Method to Quantitate Gabapentin and Buprenorphine in Human Serum

Rationale Gabapentin has shown initial promise as an opioid-sparing medication in pain patients as well as a treatment for opioid withdrawal and LC-MS/MS is often used for clinical monitoring. Despite reports of validated tandem masspectormetric methods for the determination of gabapentin and buprenorphine, mechanisms for the collision-induced fragmentation have not been adequetly described. Methods A rapid analytical method has been developed to determine the gabapentinoid, gabapentin, and partial opioid agonist, buprenorphine in 20 microliters of human serum using liquid chromatography-tandem mass spectrometry (LC-MS/MS) with a chromatographic run time of 2 minutes. A simplified sample cleanup procedure using methanol precipitation of serum proteins/lipids followed by evaporation and reconstitution in mobile phase was demonstrated. Gabapentin and buprenorphine were detected following positive ion electrospray ionization using multiple-reaction-monitoring. The internal standard approach was used for quantitation with labeled gabapentin-D10 and buprenorphine-D4 serving as internal standards. Using organic reaction principals and stable isotope labels, collision-induced fragmentation mechanisms for both gabapentin and buprenorphine are proposed. The method was validated according to the FDA Guidance for Industry - Bioanalytical Method Validation. Results Accuracy was demonstrated by error values ≤15% for buprenorphine and ≤6% for gabapentin. The inter-day precision was ≤4.88% and 15.59% for gabapentin and buprenorphine and the intra-day precision was ≤5.20% and 11.65% for gabapentin and buprenorphine. The lower limit of quantitation corresponded to 10 ng/mL for gabapentin and 1 ng/mL for buprenorphine in serum. Recoveries were 104% ± 2.55 and 85% ± 2.03 for gabapentin and buprenorphine, respectively. Conclusions Concentrations of gabapentin and buprenorphine were determined for 5 authentic human serum samples to further validate the utility of the method and applicable to therapeutic drug monitoring beyond its use as a drug screening assay. Furthermore, new mechanisms for the collision-induced dissociation of gabapentin and buprenorphine have been proposed.