Simultaneous determination of alpha, beta and gamma cyclodextrins by LC

Cyclodextrins (CDs) can be synthesized from starch by cyclodextrin glycosyltransferase (CGTase). This enzyme produces alpha-, beta- and gamma-CDs in varying proportions. In the production of cyclodextrins, purity as well as yield are important factors. A precise and reproducible method was developed and validated for the simultaneous determination of alpha-, beta-, and gamma-CDs. Optimum separation between the three CDs was achieved using a Finepak amino column with a mobile phase consisting of acetonitrile-water (70:30, v/v) at a flow rate of 1 ml/min. Detection was carried out using a differential refractive index detector. The developed method gave good chromatographic resolution of the three components with retention times of 13.16, 16.83 and 21.74 min for alpha-, beta- and gamma-CDs, respectively. The polynomial regression data for the calibration plots exhibited good linear relationship (coefficient of correlation r = 0.9987 for alpha, r = 0.9986 for beta and r = 0.9998 for gamma-CDs) over a concentration range of 2-10 mg/ml. Statistical analysis proves that the proposed LC method is precise, reproducible and accurate for the estimation of alpha-, beta- and gamma-cyclodextrins. The method can be employed for determination of percent purity as well as estimation of process yields of the cyclodextrins during the enzymatic production.