Development and Validation of Stability Indicating HPLC Method for Determination of Caffeic Acid, Vitexin and Rosmarinic Acid in Thunbergia laurifolia Leaf Extract

Thunbergia laurifolia has been a popular herb used in Thai traditional medicine for detoxification and as antipyretic. It contains rosmarinic acid (RA), caffeic acid (CA) and vitexin as major compounds. In order to control the herbal quality, the stability indicating high-performance liquid chromatography (HPLC) was developed and validated. The stability study of compounds in T. laurifolia leaf extract was investigated. The chromatographic separation was performed using a reversed-phase C18 column and mobile phase consisted of 0.5% acetic acid and methanol using a gradient elution with 1.0 mL/min flow rate. The detection wavelength was set at 330 nm. The method was validated for its linearity, precision, accuracy, limit of detection and limit of quantitation. Forced degradation of three compounds in extract showed that they were stable in oxidative condition, but highly labile under alkaline hydrolytic conditions. All three compounds in T. laurifolia leaf extract were stable at room temperature at least 3 months while a remarkable decrease of RA, vitexin and CA in the extract were found in accelerated condition. This finding could be applied for predicting the storage recommendation and expiry of T. laurifolia extract and its related pharmaceutical products.