Stability-indicating Ultra-high Performance Liquid Chromatography Method for Determination of trans-Resveratrol Bulk and Tablets

Abstract

Introduction: t-Resveratrol, trans-3,5,4'-trihydroxystilbene, is a phenolic compound found in grapes, berries and peanuts. It shows many pharmacological actions such as cardioprotective effects, antioxidant and anti-inflammatory activity. Although a variety of analytical methods has been developed to quantify t-resveratrol in natural products, none of these methods has been applied to determine t-resveratrol in bulk and tablet dosage-form for quality control purpose. This research was therefore conducted to develop the stability-indicating method for determination of t-resveratrol by using Ultra-high Performance Liquid Chromatography (UPLC). The developed method was subsequently validated under ICH guidelines on validation of analytical procedures. Furthermore, accelerated stability, standard stock solution (SSS) stability and auto-sampler (AS) stability studies of t-resveratrol in bulk and tablets were also carried out. Methods: UPLC conditions were developed and then validated with the following criteria; accuracy, precision, specificity, limit of detection (LOD), limit of quantitation (LOQ), linearity, range and robustness. The content of t-resveratrol in bulk and tablets was analyzed after 1 month storage at 40^oC 75%RH. For the SSS and AS stability studies, the response of t-resveratrol between aged solution and freshly prepared solution was compared. Results: The optimum UPLC conditions were found to be as follows: reversed-phase C₁₈ column with gradient elution system comprised of acetronitrile and deionized water, run time 3 minutes, flow rate at 0.4 mL/min and UV detection at 306 nm. The degradants of t-resveratrol in bulk and tablets obtained from forced degradation study were well separated from the t-resveratrol peak, and the peak purity angle values of t-resveratrol peak in all stress conditions were less than peak purity threshold values. The linearity of the proposed method was found in a range of concentration 0.4 – 1.2 µg/mL. The coefficient of determination (r²) was 0.9999. Repeatability and intermediate precision showed a good reproducibility with %RSD less than 2.0. Recovery of method was found to be 99.04% and 99.93% for t-resveratrol in bulk and tablets, respectively. The LOD and LOQ were 0.01 µg/mL and 0.02 µg/mL, respectively. The content of t-resveratrol resulted from changing of flow rate and column temperature were within ±2% compared to developed analytical conditions. t-Resveratrol in bulk and tablets was stable for 1 month of storage at 40^oC 75%RH, standard solution was stable for 25 days at -20^oC and 2 days in auto-sampler compartment at room temperature. Conclusion: The newly developed method was precise, accurate and reliable. It is suitable to be a stability-indicating method for determination of t-resveratrol in bulk and tablets.