High Performance Liquid Chromatography Method Validation and Forced degradation Studies of Pterostilbene

Abstract

A novel, rapid, selective, precise and accurate stability-indicating reversed phase high-performance liquid chromatography (RP-HPLC) method was developed for Pterostilbene. The method was validated accord¬ing the Q2(R1) guidelines of International Conference on Harmonization (ICH) with respect to system suitability, linearity, range, LOD, LOQ, accuracy, precision. The forced degradation was validated according to International Conference on Harmonization (ICH). The chromatographic analysis was performed on Agilent 1260 Infinity HPLC instrument using an ACE C-18 Column (150 x 4.6mm, 3um) and mobile phase comprising water: ACN (35:65 v/v) at the flow rate of 1 ml/min. The column eluent was monitored at 306nm. The total run time was 6 min and the average retention time of Pterostilbene was found to be 3.19 min. The method showed excellent linear response with correlation coefficient values (R2) of 0.999 which were within the limit of correlation coefficient (R2 >0.995). Average percentage recovery of Pterostilbene was 98.65 found within acceptable limits (97.0-103%). The LOD and LOQ were 0.006875ng and 0.020626 ng respectively. Percent RSD values of intra-day precision study were below 2%. Pterostilbene showed significant degradation when exposed to water, acid (0.1N HCL), base (0.1N NaOH), oxidizing agent (10% H2O2), and UV light. Pterostilbene content of the aqueous extract (extractive yield: 5.25%) of Pterocarpus marsupium powder by using proposed HPLC method was found to be 1.732µg. Similarly, for aqueous-ethanolic extract (extractive yield: 10%) was found to be 30.540µg. Ninety five % ethanolic extract (extractive yield: 3.33%) was found to contain 34.663µg Pterostilbene.