A Stability-Indicating Ultra Performance Liquid Chromato-Graphic (UPLC) Method for the Determination of a Mycophenolic Acid-Curcumin Conjugate and Its Applications to Chemical Kinetic Studies.

ABSTRACT

A simple, precise, and accurate reversed-phase ultra-performance liquid chromatographic (UPLC) method was developed and validated for the determination of a mycophenolic acid-curcumin (MPA-CUR) conjugate in buffer solutions. Chromatographic separation was performed on a C18 column (2.1 × 50 mm id, 1.7 µm) with a gradient elution system of water and acetonitrile, each containing 0.1% formic acid, at a flow rate of 0.6 mL/min. The column temperature was controlled at 33 °C. The compounds were detected simultaneously at the maximum wavelengths of mycophenolic acid (MPA), 254 nm, and curcumin (CUR), or MPA-CUR, at 420 nm. The developed method was validated according to the ICH Q2(R1) guidelines. The linear calibration curves of the assay ranged from 0.10 to 25 µg/mL (r2 ≥ 0.995, 1/x2 weighting factor), with a limit of detection and a limit of quantitation of 0.04 and 0.10 µg/mL, respectively. The accuracy and precision of the developed method were 98.4-101.6%, with %CV < 2.53%. The main impurities from the specificity test were found to be MPA and CUR. Other validation parameters, including robustness and solution stability, were acceptable under the validation criteria. Forced degradation studies were conducted under hydrolytic (acidic and alkaline), oxidative, thermal, and photolytic stress conditions. MPA-CUR was well separated from MPA, CUR, and other unknown degradation products. The validated method was successfully applied in chemical kinetic studies of MPA-CUR in different buffer solutions.