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Amylose based chiral HPLC method for enantioselective separation of ondansetron was developed, optimized and validated on Chiral Pak AS-3R analytical column (150 mm × 4.6 mm i.d., 3 μm). Mobile phase comprising of Acetonitrile: 0.2% Triethylamine: Methanol (15.49:30:54.51) and at a flow rate of 1.33mL/min. The separation was detected at a wavelength of 254 nm and run time was achieved within 6.0 min. Applied Central composite design (CCD) to validate the impact of three independent variables: 0.2% triethylamine (TEA), Acetonitrile (ACN) and flow rate (FL) keeping the methanol concentration constant and the responses selected were capacity factor (k1), resolution (Rs2,1), and retention time (tR2). Using this approach, a mathematical model was elucidated and response surface plots were acquired for the separation. CCD results were integrated into a multi-criteria decision making approach in order to prevail a set of optimal experimental conditions directing to the most desirable compromise between resolution and analysis time. This method can be applied for the routine analysis.
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