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In this paper, a chemometrical approach is applied for the development of a reversed-phase high-performance liquid chromatography method for the simultaneous determination of mycophenolate mofetil and its degradation product mycophenolic acid in dosage form. The fractional factorial design is used in screening experiments for selecting the variables that significantly influence the chromatographic procedure. The investigated variables are column type, temperature of the column, and composition of the mobile phase (with respect to pH and the percentage of organic modifier). Investigation is performed using two columns, XTerra (RP 18, 150 mm × 3.9 mm) and Chromolith (RP-18e, 100 mm × 4.6 mm). Because the column type shows no influence on separation process, the Chromolith column is further used due to its ability to achieve a high-speed separation without loss of column efficiency. Total analysis time is reduced from 8.34 min on XTerra to 1.27 min on Chromolith. The columns' efficiency, analysis cost, and peak symmetries are briefly compared. For both substances, only two variables are found significant: percentage of acetonitrile and pH of the water phase. Afterward, the main variables are optimized using response surface methodology for visualization and easier identification of optimal conditions. The optimal conditions are obtained with mobile phase composition of acetonitrile-15mM phosphate buffer (pH adjusted to 4.0 with 85% orthophosphoric acid) (35:65, v/v) at the flow rate of 5 mL/min. The temperature of the column is adjusted to 25°C and detection is performed at 254 nm.

journal of chromatographic science

Development of Liquid Chromatographic Method for Simultaneous Determination of Mycophenolate Mofetil and its Degradation Product Mycophenolic Acid in Dosage Form