Puerarin palmitate was synthesized through the enzymatic acylation of puerarin and two acyl donors-palmitic anhydride and vinyl palmitate (Fig. 2). First, puerarin was dried in a vacuum desiccator (Gast Manufacturing, MI, USA) with silica gel for more than seven days. Puerarin (10 mM) and the acyl donors (150 mM) dissolved in an organic solvent were then added to a 50 mL serum bottle with a working volume of 10 mL. An equal amount of immobilized lipase was added to p-NPP (25 U/L). The inlet of the serum bottle was then sealed to prevent the evaporation of the organic solvent. The reaction proceeded for 6 h in a shaking incubator (Jeio Tech, Daejeon, Korea) with constant stirring at 180 rpm and 40 °C. All experiments were performed in duplicate; results are presented as average values.
The experimental procedure for reusing immobilized lipase in the puerarin palmitate synthesis reaction was as follows. First, the immobilized lipase was filtered from the reaction solution using a filter paper. The immobilized lipase was then washed with tert-amyl alcohol, used as a reaction solvent, washed with n-hexane for rapid drying, and dried in a vacuum desiccator for at least 12 h. The dried immobilized enzyme was
The experimental procedure for reusing immobilized lipase in the puerarin palmitate synthesis reaction was as follows. First, the immobilized lipase was filtered from the reaction solution using a filter paper. The immobilized lipase was then washed with tert-amyl alcohol, used as a reaction solvent, washed with n-hexane for rapid drying, and dried in a vacuum desiccator for at least 12 h. The dried immobilized enzyme was
