目的从知母药材中快速分离制备知母皂苷BⅡ对照品。方法知母药材粗粉用80%乙醇提取,经大孔树脂D-101柱吸附,以30%乙醇洗脱,得到知母总皂苷。用制备RP-HPLC对总皂苷进行分离纯化,采用柱后分流,ELSD同时检测,收集含高质量分数知母皂苷BⅡ的组分。结果 制备产品经化学方法和13C-NMR鉴定;HPLC-ELSD检测,质量分数大于99%。结论该制备方法简单,重复性好,效率较高,所得化合物质量分数高,可用于知母皂苷BⅡ对照品的制备。 Objective To rapidly prepare the timosaponin BII reference substance from Zhimu herbs. Methods The crude powder of Rhizoma Anemarrhenae was extracted with 80% ethanol and adsorbed with macroporous resin D-101 column and eluted with 30% ethanol to obtain anemarrhena total saponins. The total saponins were separated and purified by preparative RP-HPLC. The post-column shunts were used for simultaneous ELSD detection and the fractions containing high quality fractions of timosaponin BII were collected. Results The prepared product was identified by chemical method and 13C-NMR. The mass fraction of HPLC was more than 99% by HPLC-ELSD. Conclusion The preparation method is simple, reproducible and efficient. The obtained compound has a high mass fraction and can be used for the preparation of timosaponin BII reference substance.