改性松香键合二氧化硅高效液相色谱固定相的制备及其对三七总皂苷的分离

三七中发挥药效的主要成分为三七皂苷R1、人参皂苷Rg1、人参皂苷Re、人参皂苷Rb1和人参皂苷Rd,用于贫血、冠心病、高血压、脑卒中后遗症等疾病的治疗,但其化学成分多且难分离。将氢化松香丙烯酸羟乙酯(HRHA)通过巯基-烯点击化学反应键合到烷基化硅胶表面,制备出一种新型的改性松香键合二氧化硅高效液相色谱固定相(SiO₂@HRHA),用于三七总皂苷的分离。对色谱固定相进行一系列表征,表明SiO₂@HRHA固定相具有球形度好和表面多孔的特点,比表面积为308.55 m²/g,平均孔径达到6.78 nm。将制备的固定相湿法装柱,色谱柱性能评价结果表明,SiO₂@HRHA柱具备反相色谱行为、较好的流通性及重复性。从色谱柱的综合评价Tanaka测试可知,SiO₂@HRHA柱具有较好的立体选择性及氢键容量。将其用于三七总皂苷的分离,SiO₂@HRHA柱在一定程度上优于C18柱,而SiO₂@HRHA柱的分离效果又优于实验室同期合成的色谱柱,对5种皂苷(R1、Rg1、Re、Rb1和Rd)的分离度依次为3.33、3.54、20.17和9.72,表明SiO₂@HRHA高效液相色谱柱对三七总皂苷具有优异的分离效果,为从实际样品中分离纯化三七总皂苷提供了新思路。

Preparation of modified rosin bonded silica high performance liquid chromatographic stationary phase and separation of Panax notoginseng saponins

The sanqi is the dried root of Panax notoginseng(Burk.)F.H.Chen.The main components responsible for the drug actions of sanqi are notoginsenoside R1,ginsenoside Rg1,ginsenoside Re,ginsenoside Rb1,and ginsenoside Rd,which account for about 80%of the saponin content in sanqi.It is widely used in the treatment of anemia,coronary heart disease,hypertension,stroke sequelae,and other diseases.However,sanqi has many chemical components with complex and similar structures,which are difficult to separate.In this study,alkylated silica gel bonded with hydrogenated rosin hydroxyethyl acrylate(HRHA)was prepared via mercapto-ene click chemistry.A new type of modified rosin-bonded silica stationary phase(SiO₂@HRHA)for high performance liquid chromatography was prepared for the separation of five saponins(R1,Rg1,Re,Rb1,and Rd).It was characterized by thermogravimetric analysis,Fourier-transform infrared spectroscopy,specific surface area and microporous physical adsorption and elemental analysis.The results showed that SiO₂@HRHA had a regular spherical shape with porous surfaces,along with a specific surface area of 308.55 m²/g and an average pore diameter of 6.78 nm.Performance evaluation of the column revealed that the SiO₂@HRHA column showed typical reversed-phase chromatographic behavior with better flowability and reproducibility.Results of the Tanaka test showed that SiO₂@HRHA column had good stereoselectivity and hydrogen bond capacity.Compared to other stationary phases,e.g.silica modified with acrylopimaric acid(16-hydroxyethyl-34-hydroxyethyl acrylate)ester(AAE)and dihydroterpineol(DTP),which were prepared in our laboratory at the same time,the SiO₂@HRHA column demonstrated better resolution(Rs)for the separation of the five saponins under optimal chromatographic conditions.The Rs values for R1,Rg1,Re,Rb1,and Rd were 3.33,3.54,20.17 and 9.72,respectively on the SiO₂@HRHA column.Rs between Rg1 and Re was also better than that obtained on a C18 column.Panax notoginseng saponins were separated on the SiO₂@HRHA column using acetonitrile and water as the mobile phases at the flow rate of 1.0 mL/min at 25℃.The optimal UV detection wavelength was 203 nm.It was found that the five saponins could be separated better using the SiO₂@HRHA column than the SiO₂@AAE and SiO₂@DTP columns.Because the ternary phenanthrene skeleton of the rosin group in SiO₂@HRHA had structural similarity and good stereoselectivity to the polycyclic compounds(Panax notoginseng saponins).In addition,according to the hydrophobicity evaluation,the SiO₂@HRHA column showed the best hydrophobicity among the three columns,which may be conducive to the separation of the five saponins.Thus,this study can provide a new avenue for the separation and purification of Panax notoginseng saponins from actual samples.