title: Development and validation of an analytical method based on high performance thin layer chromatography for the simultaneous determination of lamotrigine, zonisamide and levetiracetam in human plasma.
abstract: Methods based on HPLC technology are the most frequently adopted for monitoring blood levels of novel antiepileptics. Here a rapid method based on HPTLC was developed for quantitative determination of lamotrigine (LTG), zonisamide (ZNS) and levetiracetam (LVT) in human plasma and compared with HPLC and LC-MS/MS methods. Chromatographic separation was achieved on silical gel 60F(254) plates using ethylacetate:methanol:ammonia (91:10:15v/v/v) as mobile phase. Quantitative analysis was carried out by densitometry at a wavelength of 312, 240 and 210nm for LTG, ZNS and LVT, respectively. Calibration curves were linear over range of 0-200ng for LTG and ZNS and 0-400ng for and LVT. The limit of quantification of LTG, ZNS and LTV was found to be 3.69, 3.7 and 6.85μg/ml, respectively. Intra and inter-assay precision provided relative standard deviations lower than 10% for all three analytes. Correlation and Bland-Altman plot showed general agreement between HPTLC and LC-MS/MS quantification, with a mean bias of -0.25, -0.46 and 0.5μg/ml for LTG ZNS and LVT, respectively. Likewise, comparison between HPLC-UV and LC-MS/MS showed good agreement for all the three compounds analyzed. In conclusion, the proposed HPTLC method is simple, rapid, precise and accurate. It therefore is appropriate for the routine quantification of therapeutic levels of LTG, ZNS and LVT in human plasma.
authors.affiliation: Dipartimento di Fisiologia e Farmacologia, V. Erspamer - Sapienza University of Rome and Centro Antidroga and Farmacologia Clinica Policlinico Umberto I, Italy.
authors.forename: Letizia
authors.initials: L
authors.lastname: Antonilli
authors.forename: Valentina
authors.initials: V
authors.lastname: Brusadin
authors.forename: Francesca
authors.initials: F
authors.lastname: Filipponi
authors.forename: Renzo
authors.initials: R
authors.lastname: Guglielmi
authors.forename: Paolo
authors.initials: P
authors.lastname: Nencini
mesh_terms: D000927:Anticonvulsants / Q000097:blood*; D002138:Calibration; D002855:Chromatography, Thin Layer / Q000379:methods*; D016903:Drug Monitoring / Q000379:methods; D006801:Humans; D007555:Isoxazoles / Q000097:blood*; D000077213:Lamotrigine; D000077287:Levetiracetam; D016014:Linear Models; D013058:Mass Spectrometry; D010889:Piracetam / Q000031:analogs & derivatives* / Q000097:blood; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014227:Triazines / Q000097:blood*; D000078305:Zonisamide
chemical_list: D000927:Anticonvulsants; D007555:Isoxazoles; D014227:Triazines; D000077287:Levetiracetam; D000078305:Zonisamide; D000077213:Lamotrigine; D010889:Piracetam