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Ultra-Performance Liquid Chromatography Coupled with a Triple Quadrupole Mass Spectrometric Method for the Quantification of Anti-epileptic drugs Methsuximide and Normesuximide in Human Plasma and its Application in a Pharmacokinetic Study



Extensive therapeutic drug monitoring needs an analytical method for efficient and sensitive quantification of analytes of interest in clinical pharmacology. 



 A rapid, robust, sensitive and simple UPLC-MS/MS method to quantify Methsuximide (Ms) and N-desmethyl methsuximide/Normesuximide (MsMET) in human plasma was optimized, developed, and validated for application in a pharmacokinetic study. 



 Reverse phased chromatography was performed using Zorbax SB-C18, 4.6 x 75 mm., 3.5 µm as stationary phase, methanol and 0.1% formic acid (60:40 v/v) as mobile phase which was delivered isocratically at a flow rate of 0.9 mL/min. The sample injection volume was 5 µL. Mass spectrometric quantification of the analytes was performed using positive electrospray ionization as mass interface along with multiple reaction monitoring (MRM) as acquisition mode. 



 The selected mass transition ions for analyte, metabolite and its respective internal standards are as follows, precursor ion (m/z) and product ion (m/z): Ms (204.06 and 119.02), MsMET (190.05 and 119.82), Ms internal standard (MsIS) (209.17 and 124.02), and MsMET internal standard (MsMETIS) (195.09 and 124.16), respectively. The current method was found to be linear for Ms (60.72-5920 ng/mL) and MsMET (60.38-6010 ng/mL) with r2 values not less than 0.999. The mean recoveries of all analytes ranged between 71.37 and 86.38 percentage. 




 This method was validated in accordance with USFDA’s bioanalytical guidelines. This method could be applied for a routine analysis of Ms and MsMET in clinical pharmacological practice.

