关键词: 亲水作用液相色谱, 人参皂苷, 农药残留, 人参提取物, 脱除技术

Abstract:

Ginseng extracts are rich in a variety of ginseng monomer saponins, which have pharmacological functions of retarding aging, enhancing immunity, stimulating blood circulation, and lowering blood pressure. Ginseng is widely used in health products and dietary supplements in the domestic and foreign market. However, the amount of pesticide residues is an important index for measuring the quality of ginseng and ginseng extracts. Therefore, studies focused on methods for the removal of pesticide residues in ginseng extract are of great significance. Hydrophilic interaction liquid chromatography (HILIC) is used to improve the retention and separation selectivity of strongly polar substances, and it is widely employed in drug analysis, metabolomics, proteomics, etc. In this study, a method for the removal of pesticide residues was developed based on the difference in the retention behavior of pesticide residues and ginsenosides on the HILIC column. Using commercially available ginsenoside extracts, the retention behaviors of pesticide residues and ginsenosides on reverse chromatography and hydrophilic chromatographic columns were evaluated by high performance liquid chromatography. The results proved that on the reversed-phase liquid chromatography (RPLC) stationary phase, in addition to the strong retentions of quintozene and pentachloroaniline, which could be clearly separated from the saponins, the retentions of the other five pesticide residues including carbendazim, azoxystrobin, procymidone, iprodione and propiconazole were similar to total ginsenosides. The seven ginsenosides showed strong retention due to the formation of hydrogen bonds between the hydroxyl groups on the sugar chain and the carboxyl groups on the HILIC stationary phase. However, the pesticide residues were not well retained because of their poor hydrophilicity and small molecular weights. For this reason, the pesticide residues and ginsenosides could be completely separated on the HILIC column. Thus, enrichment of the seven ginsenosides and removal of the 14 pesticide residues was realized in one step on the HILIC column. In addition, the effects of loading amount, loading volume, and washing volume on the removal of pesticide residues in ginsenosides were investigated using the Click XIon SPE column. Then, taking the ginsenoside recoveries and pesticide residue removal rates into account, we confirmed the following: the ratio of the maximum sample loading mass to the filler mass was 1∶10; the optimal elution volume was twice the column volume; and the optimal loading volume was twice the column volume. The ginseng extracts were solvated with a 95% ethanol solution and loaded onto an HILIC column. The sample was subjected to pesticide residue removal, and ginsenoside purification and enrichment under the optimum removal conditions. Gradient elution was carried out using ethanol and water as the mobile phases. The total ginsenoside content in the final extracts was increased to 69.61%. The recovery of the total ginsenosides was 94.4%. The pesticide residues in the samples were quantitatively detected by gas chromatography-triple quadrupole mass spectrometry (GC-MS/MS) in the multiple reaction monitoring (MRM) mode. The 14 pesticide residues in the original ginsenoside extracts were effectively removed. The amounts of five residues were reduced to below 0.05 mg/kg, while the other nine residues were completely eliminated. This study demonstrates the application of HILIC to pesticide residue removal in traditional Chinese medicine extracts and reveals a new technique for the purification of natural products. The proposed method shows a high removal rate of pesticide residues and a high recovery of total ginsenosides. It is safe, efficient, and environment-friendly, and can aid the development of high-quality ginsenoside extracts.

Key words: hydrophilic interaction liquid chromatography (HILIC), ginsenosides, pesticide residues, ginseng extracts, removal method