A simple, rapid and cost-effective UHPLC–MS/MS method for simultaneous quantitation of seven antiepileptic drugs/metabolites in human serum

Abstract

Background

Epilepsy affects approximately 50 million people worldwide. Antiepileptic drugs (AEDs) are the mainstream treatment. Therapeutic drug monitoring (TDM) of AEDs is necessary to maximize efficacy and minimize toxicity. We report a simple, rapid, and cost-effective ultra-performance liquid chromatography-mass spectrometry method that can simultaneously measure seven AEDs/metabolites, including levetiracetam (LEV), lacosamide (LCM), zonisamide (ZON), lamotrigine (LMT), 10-hydroxycarbazepine (OXC-M1), clobazam (CLO), and N-desmethyl clobazam (N-CLB) in serum.

Method

Only 20 µl of serum was used with simple protein precipitation and dilution. Analysis was performed on a SCIEX 6500 UHPLC–MS/MS in positive ion mode. Separation was performed on a C18 reversed-phase column using a gradient. The seven AEDs/metabolites were eluted in 4.5 min.

Results

The assay was linear over the concentration ranges 0.4–100 µg/mL for LEV, 0.12–30 µg/mL for LMT, 0.12–30 µg/mL for LCM, 0.32–80 µg/mL for ZON, 0.28–70 µg/mL for OXC-M1, and 7.82–2000 ng/mL for CLO, 78.2–20000 ng/mL for N-CLB, respectively, with correlation coefficient greater than 0.99. Recovery was from 88 to 108 %. Intra and inter assay precision for three levels of quality controls were from 2.1 to 6.8 % and 4.2 to 10.9 %, respectively. The accuracy was evaluated by comparing with the College of American Pathologists survey results, and a correlation coefficient greater than 0.96 was observed. The absence of matrix effects was also confirmed.

Conclusion

We have developed and validated a simple, rapid, and cost-effective UHPLC–MS/MS method for the simultaneous quantitation of seven AEDs/metabolites in serum within a 4.5-min analysis time. It has been implemented in our children’s hospital with same-day turnaround time.