ABSTRACT
Puerarin is the main active component of flavonoids in Puerariae Lobatae Radix. In this study, agar gel microspheres bonded with β-cyclodextrin (AG-β-CD) were synthesized by using economical agar, and then high-purity puerarin was obtained with AB-8 through high-yield separation. With purity and yield of puerarin, and chromatographic purity of related impurities as indexes, four macroporous resins of different properties, namely ADS-7 (high polarity), ADS-17 (medium polarity), ADS-21 (polarity) and AB-8 (weak polarity), were selected for separation of puerarin and technological optimization. In addition, the AG-β-CD purification process was optimized and verified. The results showed that, AB-8 resins showed the best effect and selected as the pre-treatment resins for crude puerarin, and puerarin with the purity of 87.68% showed a recovery rate of 89.66%. The optimized purification process parameters of AG-β-CD included mobile phase (15% ethanol), loading capacity (the ratio of loading amount to column volume) (1.33 g•L⁻¹), sample concentration (8 g•L⁻¹) and flow rate (1 mL•min⁻¹), puerarin with the purity of 95% showed a recovery rate of more than 97%.
ABSTRACT
Objective: Daidzein was efficiently purified by agar gel microspheres bonded β-cyclodextrin (AG-β-CD). Methods: Using agar as raw material, after emulsification, crosslinking, and bonding β-CD as functional group, AG-β-CD was synthesized for the purification of daidzein, and the purification process was determined and proved with mobile phase, flow rate, and loading capacity of microspheres. The structure of daidzein was identified by MS and NMR, AG-β-CD was chromatographically evaluated with daidzein, EGCG, and puerarin as the following tripartition such as difference of retention behavior on C18 reversed phase column chromatography, molecular simulation by autoDOCK4.0, and retention time curves on AG-β-CD with different contents of acetonitrile. Results: The main component of soybean isoflavone was daidzein (57.14%). The loading quantity of AG-β-CD was 1.33 mg/mL, flow rate was 2 BV/h, eluted by 2 BV of 20% ethanol, 1.33 BV of 40% ethanol, and 6-7 BV of 70% ethanol, the content was ≥ 95%, purity of daidzein (96.98%) was obtained with 97.86% yield. Chromatographic mechanism research showed that AG-β-CD had hydrophilic interaction chromatography and reversed-phase chromatography. Conclusion: AG-β-CD is capable of highly efficient purification of daidzein.