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LC-MS/MS-Based Quantification of 9 Antiepileptic Drugs From a Dried Sample Spot Device

D'Urso, Annachiara BiolSciD*; Cangemi, Giuliana PhD; Barco, Sebastiano BSc; Striano, Pasquale MD, PhD; D'Avolio, Antonio MSc, SM§; de Grazia, Ugo PhD*

doi: 10.1097/FTD.0000000000000600
Original Article

Background: Therapeutic drug monitoring (TDM) of antiepileptic drugs (AEDs) is commonly performed on plasma or serum. The use of dried plasma spots (DPSs) could represent a useful tool to facilitate sample shipment to reference laboratories. In this article, the authors describe the application of a commercially available UHPLC-MS/MS method for the determination of 9 commonly prescribed AEDs (levetiracetam, lacosamide, topiramate, ethosuximide, lamotrigine, rufinamide, zonisamide, primidone, and oxcarbazepine and its active metabolite 10-OH-monohydroxycarbazepine) to DPS collected on dried sample spot devices (DSSDs).

Method: Fifty microliters of plasma were spotted on DSSD. After being air-dried at room temperature, they were extracted using an organic extraction solution containing the appropriate deuterated internal standards. The chromatographic separation was performed on a UHPLC reversed-phase C-18 column, and the analytes were quantified using a triple quadrupole mass spectrometer (LC-MS/MS).

Results: The assay was linear over the concentration ranges tested with a total runtime of 10.3 minutes. Recovery ranged from 93.7% to 106.8%. Intraday and interday precision for all quality control levels, including lower limit of quantification, ranged from 2.1% to 18.4% and 2.1% to 13.2%. Intraday and interday accuracy biases ranged from −11.7% to 14.3% and −9.2% to 8.0%. The absence of matrix effects was also tested and confirmed. Real samples derived from patients under therapy were also analyzed, and the comparison of results obtained from DSSD with those obtained from plasma showed that the 2 matrices were interchangeable. Stability tests performed on both quality controls, and real samples demonstrated that DSSDs can be easily stored and shipped at room temperature for 15 days.

Conclusions: The application of the LC-MS/MS method allowed the authors to obtain a very specific, sensitive, and rapid (total runtime = 10.3 minutes) quantification of 9 AEDs starting from very low volumes of plasma samples. The main advantage of DPS over wet samples is room temperature storage and shipment, which lowers shipment costs and makes it suitable for routine TDM. Moreover, in comparison with other alternative matrices, DPS allows for the use of the same therapeutic ranges on which routine TDM is based. DPS on DSSD can thus be considered as a useful and cheap tool for the broader application of TDM.

*Laboratory of Neurological Biochemistry and Neuropharmacology, Fondazione IRCCS Istituto Neurologico Carlo Besta, Milano;

Area Aggregazione Servizi e Laboratori Diagnostici, IRCCS Istituto Giannina Gaslini;

Pediatric Neurology and Muscular Diseases Unit, Department of Neurosciences, Rehabilitation, Ophthalmology, Genetics, Maternal and Child Health, University of Genoa, IRCCS Istituto Giannina Gaslini, Genova; and

§Laboratory of Clinical Pharmacology and Pharmacogenetics, Department of Medical Sciences, University of Turin, Ospedale Amedeo di Savoia, Torino.

Correspondence: Ugo de Grazia, PhD, Laboratory of Neurological Biochemistry and Neuropharmacology, Fondazione IRCCS Istituto Neurologico Carlo Besta, via Celoria 11, 20133 Milano, Italy (e-mail:

A. D'Urso was supported by the Italian Ministry of Health Ricerca Corrente project 2017.

The authors declare no conflict of interest.

Received August 02, 2018

Accepted December 20, 2018

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