中压液相色谱法快速制备木犀草素-7-O-β-D-葡萄糖醛酸苷单体及鉴别

Preparation and Identification of Luteolin-7-O-β-Dglucuronide by Preparative Medium Pressure Liquid Chromatography

本文采用二元中压制备液相色谱系统,以装有反相C18（Chromatorex,MB 100-40/75）分离填料的两端具有锥形不锈钢接口的耐压玻璃柱（φ49 mm×H310 mm）作为制备色谱柱,流动相为甲醇（A）/含0.05%冰醋酸水溶液（B）,梯度洗脱程序：0～20 min,10%（A）;20～230 min,30%（A）;230～270 min,60%（A）;270～300 min,100%（A）,流速：50 mL/min,检测波长254 nm,进样量：200 mL,从抱茎苦荬菜（苦碟子）总黄酮中快速、高效地分离得到木犀草素-7-O-β-D-葡萄糖醛酸苷（L7GU）单体。所得单体经UV、IR、TOF-MS、1H NMR、13C NMR方法进行了结构确证。经硅胶和聚酰胺薄层色谱检查,在365 nm紫外灯下均呈单一的黄绿色荧光斑点;HPLC-UV分析表明,在210、254、290、348 nm四个波长下,采用色谱峰面积归一化方法计算产品纯度均在98%以上。所得产品可直接作为对照品用于苦碟子药材及制剂的质量控制。

To establish a rapid and efficient method for isolation and purification of lnteolin-7-O-β-D-glucuronide （LTGU） from herb of Ixeris sonchifolia （Bge） Hance, a preparative medium pressure liquid chromatography （MPLC） method was developed and used. The MPLC system consisted two-unit pumps and a glass column （φ49 mm×H310 ram） linked with cone-shaped stainless steel pert and packed with Cls particles （Chromatorex, MB 100-40/75）. The mobile, phase was methanol （A）-water （containing 0.05% acetic acid, B）. The gradient elution program was as follows ：0- 20 win, 10% A ;20-230 rain,30% A ;230-270 rain,60% A ;270-300 rain, 100% A. The flow rate was 50 mL/min, and the detection wavelength was set at 254 nm. The structure of isolated L7GU was subsequently confirmed by analyzing its UV,IR,TOF-MS,IH NMR and 13C NMR. Silica gel HPTLC,polyamide TLC and HPLC analyses were performed to test and verify the purity of isolated LTGU. One single yellowish green spot under UV at 365 nm was detected in both TLC analyses. The HPLC analyses at detection wavelength of 210 nm,254 nm,290 nm and 348 nm demonstrated.the purity of isolated L7GU was more than 98%. The high-purity L7GU prepared by this method can be used as reference substance for the quality control of herb of I. sonchifolia and its proprietary medicine.