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Development and Validation of an Ultra-Performance Liquid Chromatography–Tandem Mass Spectrometry Method for the Concurrent Measurement of Gabapentin, Lamotrigine, Levetiracetam, Monohydroxy Derivative of Oxcarbazepine, and Zonisamide Concentrations in Serum in a Clinical Setting

Palte, Michael J., MD, PhD*; Basu, Sankha S., MD, PhD*; Dahlin, Jameson L., MD, PhD*; Gencheva, Ralitsa, BS; Mason, Donald, PhD; Jarolim, Petr, MD, PhD*; Petrides, Athena K., PhD*

doi: 10.1097/FTD.0000000000000516
Original Article
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Background: Therapeutic drug monitoring of antiepileptic drugs (AEDs) is often necessary to prevent associated destructive toxicities. Tandem mass spectrometry (MS/MS) with stable-isotope–labeled internal standards is considered the gold standard for the measurement of AEDs. This study presents the development and validation of a clinical ultra-performance liquid chromatography–MS/MS method for the concurrent measurement of gabapentin, lamotrigine, levetiracetam, monohydroxy derivative of oxcarbazepine, and zonisamide in human serum.

Methods: To determine the optimal assay analyte range, one year of AED therapeutic drug monitoring results (n = 1825) were evaluated. Simple protein precipitation with acetonitrile containing isotopically labeled internal standards was used. Reverse-phase ultra-performance liquid chromatography chromatographic separation was used, having a total run time of 3 minutes. Quantification of analytes was accomplished using electrospray ionization in positive ion mode and collision-induced dissociation MS. Assay parameters were evaluated per Food and Drug Administration bioanalytical guidelines.

Results: After evaluating internal patient data, the analytical measuring range (AMR) of the assay was established as 0.1–100 mcg/mL. All AEDs were linear across the AMR, with R2 values ranging from 0.9988 to 0.9999. Imprecision (% coefficient of variation) and inaccuracy (% difference) were calculated to be <20% for the lower limit of quantitation and <15% for the low, mid, and high levels of quality controls across the AMR. All AEDs demonstrated acceptable assay parameters for carryover, stability under relevant storage conditions, matrix effects, recovery, and extraction and processing efficiency. In addition, the assay displayed acceptable concordance to results obtained from a national reference laboratory, with Deming regression R2 of 0.99 and slope values ranging from 0.89 to 1.17.

Conclusions: A simple, cost-effective, and robust ultra-performance liquid chromatography–tandem mass spectrometry method for monitoring multiple AEDs was developed and validated to address the clinical needs of patients at our institution.

*Department of Pathology, Brigham and Women's Hospital and Harvard Medical School;

Department of Pathology, Brigham and Women's Hospital, Boston, MA; and

Waters Corporation, Milford, MA.

Correspondence: Athena K. Petrides, PhD, Department of Pathology, Brigham and Women's Hospital, 75 Francis St, Boston, MA 02115 (e-mail: apetrides@bwh.harvard.edu).

D. Mason is an employee of Waters Corporation, Milford, MA. The remaining authors declare no conflict of interest.

Supplemental digital content is available for this article. Direct URL citations appear in the printed text and are provided in the HTML and PDF versions of this article on the journal's Web site (www.drug-monitoring.com).

Received January 04, 2018

Accepted March 04, 2018

Copyright © 2018 Wolters Kluwer Health, Inc. All rights reserved.