Systematic study on QAMS method for simultaneous determination of triterpenoid saponins in Ilex pubescens by HPLC and UPLC

Abstract

The quantitative analysis of multi-components with single marker (QAMS) method was firstly established for simultaneous determination of six triterpenoid saponins (ilexgenin A (C₁), ilexsaponin A1 (C₂), ilexsaponin B1 (C₃), ilexsaponin B2 (C₄), ilexsaponin B3 (DC₁), ilexoside O (DC₂)) in Ilex pubescens by Ultra Performance Liquid Chromatography (UPLC) and High Performance Liquid Chromatography (HPLC). Using C1 as the internal reference, the relative correction factors (RCF) of the other five triterpenoid saponins were calculated and statistically evaluated. The durability of the method was verified with five different LC instruments and five different C₁₈ columns under various chromatographic conditions. The external standard method after methodology verification was chosen to check the accuracy and feasibility of the QAMS method. The results showed that the RCFs of all compounds were obtained with good reproducibility (RSD < 5.0%), whether on different chromatography instruments or under various chromatographic conditions. The relative retention value method could be adopted for accurately positioning the chromatographic peak of the six constituents, with their values of RSD ranging between 0.5% and 1.6%. Meanwhile, no significant differences were found in the quantitative results of the six saponins in nine batches of medicines calculated by the QAMS method and external standard method, whether on a HPLC system or UPLC system. The QAMS method established in our research for simultaneous determination of the six saponins is accurate and can be feasibly used to evaluate the quality of Ilex pubescens, especially when the standard substance is inconvenient to obtain.