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Determination of Saikosaponins in Bupleuri Radix by Micellar Electrokinetic Chromatography with Experimental Design

ABSTRACT The important pharmacological components of Bupleuri radix include saikosaponins a–d. A multivariable approach with experimental design was employed in this study to determine four saikosaponins in different species of Bupleuri radix by cyclodextrin-modified micellar electrokinetic chromatography. Compared to traditional multiple-parameter investigations, the current experimental design can shorten the time necessary for method development, and the multivariable approach can be used to evaluate the separation capability of the method and to explore the interactions of the different parameters. The significant influences of two experimental variables, including the concentration of γ-cyclodextrin and the percentage of methanol, on the responses were investigated using chromatographic resolution statistics. The optimized conditions were as follows: 25 mM of borate (pH 9.18) containing 50 mM of sodium dodecyl sulfate, 5 mM of γ-cyclodextrin, and 15% methanol (v/v) as running buffer. After the separation conditions were optimized and good resolution was acquired, solid phase extraction was performed to reduce the interference of crude extracts of Bupleurum kaoi and Bupleurum chinense DC. The simple and rapid analytical method was successfully applied, allowing for the determination of four saikosaponins and the identification of species of Bupleuri radix.