LC-MS/MS结合分子网络技术快速鉴定生三七和砂烫三七的三萜皂苷类成分

Rapid Identification of Triterpenoid Saponins from Panax Notogin⁃seng and Processed Panax Notoginseng with Hot Sand Based on LC-MS/MS Combined with Molecular Network Technology

采用液相色谱-串联质谱(LC-MS/MS)和分子网络技术,快速鉴定了生三七和砂烫三七中的三萜皂苷类成分。通过170~220℃高温河砂炒制获得砂烫三七,70%乙醇-水回流提取获得生三七和砂烫三七浸膏,利用D101大孔树脂以70%乙醇-水洗脱获得三七总皂苷。采用Agilent C_(18)色谱柱(100 mm×2.1 mm,3.5µm),以0.05%甲酸-水和乙腈为流动相进行梯度洗脱,在负离子模式下采集三萜皂苷对照品、生三七总皂苷和砂烫三七总皂苷的二级质谱信息。将经格式转换后的MS/MS数据上传至全球天然产物社会分子网络(GNPS)平台计算分析,借助Cytoscape 3.9.1软件构建可视化分子网络,获得生三七和砂烫三七三萜皂苷的分子网络图。根据对照品、保留时间、特征碎片离子、MS/MS裂解规律和文献报道等信息,从生三七中鉴定出50个三萜皂苷,从砂烫三七中鉴定出60个三萜皂苷,二者共有23个皂苷类成分;生三七单独含有27个成分,包括11个丙二酰基取代的皂苷;砂烫三七单独含有37个成分,包括19个乙酰基取代的成分。其中有11个成分通过对照品进行验证。采用高效液相色谱法测定了生三七和砂烫三七总皂苷中人参皂苷Rb1、Rg1、Rg3、Rh4、Rk3和三七皂苷R1的含量,发现生三七中原生皂苷的含量较高,砂烫三七中转化皂苷的含量较高。

Triterpenoid saponins from Panax notoginseng and processed Panax notoginseng with hot sand were fast identified based on liquid chromatography-tandem mass spectrometry(LC-MS/MS)combined with molecular network technology.The processed Panax notoginseng was obtained using sand roasting at 170-220℃.Panax notoginseng and processed Panax notoginseng were extracted with 70%ethanol-water under reflux and the extractions were subjected to D101 macroporous resin eluting with 70%ethanol-water to afford the total saponins.The Agilent C_(18) column(100 mm×2.1 mm,3.5µm)was selected for gradient elution with 0.05%formic acid-water and acetonitrile as the mobile phases,and mass spectrometry data of the reference substances and total saponins were col⁃lected in negative ion mode.After format conversion,the MS/MS data were uploaded to the Global Natural Products Social Molecular Network(GNPS)to establish a molecular network and the visualiza⁃tion was performed in Cytoscape 3.9.1.According to the similarity of reference substances,reten⁃tion times,characteristic fragment ions,MS/MS fragmentation rules,and literatures,50 triterpe⁃noid saponins were identified from Panax notoginseng and 60 triterpenoid saponins were character⁃ized from processed Panax notoginseng with hot sand in which 23 saponins were found in both of them.There were 27 components in Panax notoginseng alone,including 11 malonyl-glucoside sub⁃stituted saponins.The processed Panax notoginseng with hot sand contained 37 saponins alone,in⁃cluding 19 acetyl-glucoside substituted compounds.Eleven of these saponins were verified by refer⁃ence substances.The quantitative analysis of ginsenoside Rb1,Rg1,Rg3,Rh4,Rk3,and no⁃toginsenoside R1 in the total saponins of Panax notoginseng and processed Panax notoginseng with hot sand were determined by high performance liquid chromatography.The results showed that Panax notoginseng mainly contained primary saponins and the processed Panax notoginseng with hot sand mainly contained transformed saponins.