Engineering Research Center of Pharmaceutical Process Chemistry, Ministry of Education, School of Pharmacy, East China University of Science and Technology, Shanghai 200237, PR China.
Engineering Research Center of Pharmaceutical Process Chemistry, Ministry of Education, School of Pharmacy, East China University of Science and Technology, Shanghai 200237, PR China.
J Chromatogr A. 2020 Jun 7;1620:461010. doi: 10.1016/j.chroma.2020.461010. Epub 2020 Mar 6.
In this paper, adsorption mechanism of triterpenoid saponins in reversed-phase liquid chromatography (RPLC) and hydrophilic interaction liquid chromatography (HILIC) was proposed based on the study of the retention behavior of mogroside V as test substance. The change of peak shape of mogroside V and its influencing factors was first investigated. As the increase of sample loading, a tailing peak of mogroside V was observed in MeOHHO of both two modes. It was the fronting peak in ACNHO of HILIC while there was a transition from fronting peak to tailing peak in ACNHO of RPLC that was largely affected by column temperature and ACN concentration. The adsorption isotherm of mogroside V in ACNHO of RPLC was fitted by Moreau model, where a monolayer adsorption with large inter-molecular interaction was formed on the C18 surface. While in ACNHO of HILIC, the adsorption of mogroside V was in accordance with BET model, showing multilayer adsorption behavior. In MeOHHO of both HILIC and RPLC, there was always monolayer adsorption, which was fitted by Langmuir model. At last, by choosing the suitable chromatographic mode, controlling the key factors such as the solvent concentration and column temperature, and predicting the broadening trend of peak, three methods were screened out, namely, C18 column with 22% ACN (30 °C), Click XIon column with 90% MeOH or 70% ACN, to get mogroside V of purity greater than 98% from Siraitia grosvenorii extract. Among them, the RPLC method of 22% ACN that showed the highest loading sample per hour (1.92%) and the lowest solvent consumption emerged as the best approach.
本文以罗汉果苷 V 为研究对象，探讨了反相液相色谱（RPLC）和亲水作用液相色谱（HILIC）中三萜皂苷的保留机理。首先考察了罗汉果苷 V 峰形变化及其影响因素。随着进样量的增加，在两种模式的 MeOH-H2O 中均观察到罗汉果苷 V 的拖尾峰。在 HILIC 模式的 ACN-H2O 中为前伸峰，而在 RPLC 模式的 ACN-H2O 中则从前伸峰转变为拖尾峰，该转变受柱温和 ACN 浓度的影响较大。RPLC 模式的 ACN-H2O 中，罗汉果苷 V 的吸附等温线符合 Moreau 模型，表明罗汉果苷 V 在 C18 表面形成了单层、分子间相互作用较大的吸附。而在 HILIC 模式的 ACN-H2O 中，罗汉果苷 V 的吸附符合 BET 模型，表现出多层吸附行为。在 HILIC 和 RPLC 的 MeOH-H2O 中，均为单层吸附，符合 Langmuir 模型。最后，通过选择合适的色谱模式、控制溶剂浓度和柱温等关键因素，并预测峰展宽趋势，筛选出三种方法，即 22% ACN（30℃）的 C18 柱、90% MeOH 或 70% ACN 的 Click XIon 柱，可从罗汉果提取物中得到纯度大于 98%的罗汉果苷 V。其中，22% ACN 的 RPLC 方法显示出最高的每小时进样量（1.92%）和最低的溶剂消耗，是最佳方法。
