人参次苷H滴丸原料药中皂苷含量测定及细胞膜色谱技术对效应成分筛选

Active ingredients screening by cell membrane chromatography and simultaneous quantitation of ginsenosides in bulk drug of secondary ginsenoside H dripping pills

目的建立高效液相色谱串联三重四级杆质谱法(HPLC-MS/MS)测定人参次苷H滴丸(SGHDP)原料药中拟人参皂苷RT5、20(S)-人参皂苷Rh1、20(S)-人参皂苷Rh2的方法并采用细胞膜色谱(CMC)技术筛选效应成分。方法采用Century SIL BDS C18色谱柱(250 mm×4.6 mm,5μm),以0.2%甲酸水溶液-乙腈为流动相,梯度洗脱,正离子多重反应监测(MRM)模式进行扫描定量,并对CMC技术中所得的红细胞膜固相化SGHDP的效应成分进行确定。结果拟人参皂苷RT5、20(S)-人参皂苷Rh1、20(S)-人参皂苷Rh2分别在0.095~0.235、0.042~0.168、0.105~0.419 mg/m L线性关系良好(r2≥0.999 0),3种成分的平均加样回收率分别为99.95%、100.12%、100.06%,RSD分别为1.06%、0.96%、0.91%。SGHDP原料药中拟人参皂苷RT5、20(S)-人参皂苷Rh1、20(S)-人参皂苷Rh2质量分数分别为21.24%、21.42%、29.70%。红细胞膜固相化SGHDP的效应成分为20(S)-人参皂苷Rh2。结论该方法对SGHDP原料药中3种人参皂苷含量的测定准确、可靠,为SGHDP的质量控制提供了新的参考依据。20(S)-人参皂苷Rh2为红细胞膜固相化成分,推测其可能是SGHDP的效应成分,与前期的抗癌及抗抑郁的研究结果一致。

Objective To establish a HPLC-MS/MS method for simultaneous determination and active ingredients screening of pseudoginsenoside RT5, 20（S）-ginsenoside Rh1 and 20（S）-ginsenoside Rh2 by cell membrane chromatography（CMC） in secondary ginsenoside H dripping pills（SGHDP）. Methods The samples were separated on Century SIL BDS C18 column（250 mm × 4.6 mm, 5 μm） eluted with 0.2% formic acid aqueous solution-acetonitrile in a gradient mode, and the target compounds were analyzed by positive ion multiple reaction monitoring（MRM） mode, and active ingredients of SGHDP obtained in solid-phase of biomembrane by CMC technology were determined at the same time. Results The linear ranges of pseudoginsenoside RT5, 20（S）-ginsenoside Rh1, and 20（S）-ginsenoside Rh2 were 0.095—0.235, 0.042—0.168, and 0.105—0.419 mg/m L; the extraction recoveries were 99.95%, 100.12%, and 100.06%; and RSD were 1.06%, 0.96%, and 0.91%, respectively. The contents of pseudoginsenoside RT5, 20（S）-ginsenoside Rh1, and 20（S）-ginsenoside Rh2 in SGHDP were 21.24%, 21.42%, and 29.70%, respectively. 20（S）-Ginsenoside Rh2 was the active ingredient obtained by biomembrane using as a new quality control maker for SGHDP. Conclusion The developed method is accurate and reliable for the determination of three ginsenosides in SGHDP, and provides a new reference for quality control of SGHDP. 20（S）-Ginsenoside Rh2 is a immobilization component of red cell membrane, speculated to be the active ingredient of SGHDP, which is in consistent with previous studies on antitumor and antidepression.