Optimization of a gas chromatography--mass spectrometry method using chemometric techniques for the determination of ezetimibe in human plasma

Authors: EBRU UÇAKTÜRK, NURAN ÖZALTIN

Abstract: A new, rapid, and sensitive gas chromatography--mass spectrometry (GC-MS) method was developed for the determination of ezetimibe (EZE) in human plasma. EZE was derivatized prior to GC-MS analysis. Various derivatization techniques such as acetylation, methylation, and silylation were tried. EZE was extracted from plasma with high recovery (94.39%--97.57%) using methyl tertbutyl ether and carbonate buffer (pH 9). Chromatographic conditions were optimized using chemometric methods. In the first step, optimization with factorial design, chromatographic variables (initial and final column temperature, oven ramp rate, and flow rate of gas) were screened to select important variables for the retention of EZE. In the second step, central composite design was applied to decide on the retention time of EZE. The analysis was achieved in a short period of time (<4 min). The developed method was validated for parameters including specificity, limit of quantitation, linearity, accuracy, precision, recovery, stability, robustness, and ruggedness. The limit of quantitation was found to be 10 ng mL^{-1}. The method was successfully applied to determine total EZE in the plasma of hypercholesterolemic patients.

Keywords: Ezetimibe, gas chromatography--mass spectrometry, human plasma, experimental design, validation

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