A method for fast chiral separation of cetirizine and quantitation of levocetirizine in human plasma using subcritical fluid chromatography with tandem mass spectrometry was developed and validated. The chromatographic separation was performed using a Chiralpak IE column (2.1 mm×150 mm, 5 μm) with an isocratic elution of CO2/organic modifier (55/45, v/v) at a flow rate of 0.85 mL/min. The organic modifier was composed of water/methanol (5/95, v/v). The makeup flow was optimized at water/methanol (10/90, v/v) and 0.2 mL/min. The most influential parameters on the separation of cetirizine affecting resolution, retention time and sensitivity were selected by fractional factorial design. The 3 selected factors were optimized by response surface methodology. Tandem mass spectrometry was used at electrospray ionization, positive ion mode, and multiple-reaction monitoring mode. Isotope-labeled cetirizine-d4 was used as the internal standard. The sample preparation of human plasma was conducted by solid phase extraction of hydrophilic-lipophilic balance (HLB) type. The developed method was validated for selectivity, linearity, precision, accuracy, recovery, limit of quantitation (LOQ), and limit of detection (LOD). The real human plasma samples were analyzed and the pharmacokinetic results were compared with results of previous research. The developed method was found to be reliable based on the similarity between the results of the current and previous methods. The chiral separation for cetirizine and economic feasibility were compared with those of previous studies using normal phase-HPLC or reversed phase-HPLC. The established analytical method could be successfully applied to pharmacokinetic study with reduction in the analysis time and costs.
Keywords: Cetirizine; Chiral separation; Enantiomer; Mass spectrometry; Response surface methodology; Subcritical fluid chromatography.
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