Abstract:In this manuscript,the separation and purification process of hesperidin from orange peel were developed,through investigating the effect of crude extract volume,pH and temperature on the variation of hesperidin partition and its recovery. Firstly two organic compounds with low molecular weight and three inorganic salts were selected to construct the aqueous two-phase system,whose performance was characterized by cloud point method. The chosen systems with good property in phase forming were applied to extract hesperidin from its crude ethanol extract,meanwhile the dependence of partition coefficient on the concentration of phase-forming substance and the type of system were also researched here. Therefore the best two-phase system was obtained,which is Acetone-K2HPO4and applied to extract hesperidin from the peels. The recovery of hesperidin could reach to 96.69% under the optimization condition. The extract was refined further with prep-HPLC and purer hesperidin product was received. Gradient elution strategy made the refining process more effective in prep-HPLC system. The purity of hesperidin samples obtained from aqueous two-phase extraction was about 71.36%. It increased to 96.09% ca. after pre-HPLC refining. The total recovery of hesperidin was 83.54%.
