半制备型高效液相色谱法分离刺葡萄花色苷单体

摘要

经大孔吸附树脂HP-20纯化刺葡萄花色苷的粗提物后，以半制备型高效液相色谱法分离得到高纯度的刺葡萄花色苷单体。以XCharge C18柱（20 mm×250 mm，10μm）制备柱，考察梯度洗脱条件、流动相流速、进样量对刺葡萄花色苷分离的影响，确定了最佳制备条件为甲醇-3%甲酸溶液流动相梯度洗脱、流速15 mL/min、进样量1.2 mL，实现了2种主要花色苷单体的分离及制备。经超高效液相色谱-四极杆飞行时间质谱鉴定，2种花色苷单体分别是锦葵素-3,5-O-双葡萄糖苷和锦葵素-3,5-O-双葡萄糖苷-香豆酰，产品纯度分别达到了99.54%和98.28%。方法具有简单易行、经济快速、易于放大等特点，适用于刺葡萄花色苷标准品的大规模制备。%A semi-preparative high performance liquid chromatographic (HPLC) method using an XCharge C18 preparative column (20 mm× 250 mm, 10μm) was established for the preparation of anthocyanins monomers with high purity from crude extracts of Vitis davidiiFoex after purification with macroporous resin HP-20. The effects of experimental conditions including mobile phase gradient elution conditions, flow rate and injection volume were investigated on the separation efficiency of anthocyanins. The optimal chromatographic conditions were determined as follows: mobile phase, a mixture of methanol and 3% formic acid water for gradient elution; flow rate, 15 mL/min; and injection volume, 1.2 mL, which allowed the isolation and preparation of two major anthocyanins monomers in 18 min. The two anthocyanins were identified by MSE method using ultra-performance liquid chromatography quadrupole time of flight mass spectrometry (UPLC-QTOF MSE) as malvidin-3,5-O- diglucoside and malvidin-3,5-O-diglucoside-coumary with purities of 99.54% and 98.28%, respectively. The developed procedure was simple, effective, scalable and economic. It could be a promising alternative procedure for large-scale preparation of standard materials of anthocyanins fromVitis davidiiFoex.