DoE Screening and Optimization of Liquid Chromatographic Determination of Nicotinic Acid and Six Statins: Application to Pharmaceutical Preparations and Counterfeit Detection.

Abstract

An isocratic reversed-phase high performance liquid chromatographic method has been developed and validated to simultaneously determine nicotinic acid, pravastatin sodium, rosuvastatin calcium, atorvastatin calcium, pitavastatin calcium, lovastatin sodium and simvastatin sodium in focus on counterfeit drug detection. Thin-layer chromatography, nuclear magnetic resonance and mass spectrometry have been additionally performed to verify the identification of adulterants of counterfeit herbal medicines. Chromatographic separation was carried out on Inertsil® ODS-3 C18 (4.6 × 150mm, 5μm) with isocratic mobile phase elution containing a mixture of acetonitrile: methanol: 25mM potassium dihydrogen phosphate buffer, pH2.86 adjusted with 0.1M o-phosphoric acid (48: 30: 22, v/v/v), at a flow rate of 1mL/min and with UV detection at 238nm. The design of experiment methodology, Plackett-Burman and Box-Behnken designs, was used to screen and optimize the mobile phase composition. The validation of the method was also carried out under the International Conference on Harmonization guidelines. The developed method was sensitive, accurate, simple, economical and highly robust, in addition to the comprehensiveness and novelty of this method for separating the seven drugs. The results were statistically compared with the reference methods used Student's t-test and variance ratio F-test at P < 0.05.