Radix Notoginseng (Sanqi) is a valuable Chinese Medicine (CM) and widely used in clinical. Some previous studies showed that saponins are the main active ingredients in this CM. Xuesaitong injection, a sterilized injection, was prepared by extracting the total saponins from the crude materials of Radix Notoginseng. Previous pharmacological studies indicated that this CM has many cardiovascular activities such as anti-platelet aggregation, anti-thrombosis and anti-arrhythmias. According to the standard set by the Ministry of Health P. R. China(The pharmaceutics of Chinese medicine prescription, volume 19), a double wave scan of thin layer chromatography (TLC) was recommended for the detection of the content in Xuesaitong injection. However, factors such as structural diversity, large molecule structure, lack of chromophores of different saponins make analysis of individual saponin very difficult. In addition, TLC is an opening system which means the results produced by this method are highly affected by the environmental conditions and therefore show poor reproducibility. In some previous studies, a quantitative method was developed to analyze the Sanqi saponins by using HPLC coupled with UV or diode array detector (DAD). The main problem for this method is due to the lack of ultraviolet (UV) absorption in most of the saponins. Although evaporating light scattering detector (ELSD) can solve the problem of no UV absorption, it has a drawback of not able to detect nost trace components within the detectable limits. Therefore, in this study, HPLC-MS-MS method was employed as a key technique for the qualitative and quantitative analysis of the injection. In the present studies, Agilent 1100 Series LC/MSD VL Trap System and LC/MSD Trap Software Version 4.2 were used for data analysis. A gradient elution system consisted of water and acetonitrile was used for mobile phase. Additionally, 8 mM ammonium acetate was added in the water phase as electrolyte solution. The flow rate of the mobile phase was set at 0.4 ml/min and the total analytical time was 40 min. At the same time, an electro-spray ionization (ESI) source and negative ion mode were used in the mass spectrometer and the scan range was from 200 to 1400u. Full scan mode (auto MS/MS) and selected reaction monitoring (SRM) were used for the qualitative and quantitative analysis. In the test results of limit of detection (LOD) and limit of quantification (LOQ), the detection limit was found to reach pg level for most components in the injection. The result showed a high sensitivity for this detection method. In the qualitative analysis, 27 peaks could be identified with the help of 9 reference standards and the fragmentation patterns in the mass spectra. Comparing 10 different batches and/or sources of Xuesaitong injection and their corresponding crude material (Sanqi), the results showed that all samples were with very similar saponin components. In the quantitative analysis, notoginsenoside R4, ginsenoside Rb1, Rb2, Rb3, Rc, Rd, Re, Rf and Rg were quantified. The results obtained from the 10 batches of Xuesaitong injection showed that the notoginsenoside and ginsenoside content varied a lot from batch to batch, even for those injections manufactured by the same factory. The content of the components was found to vary as follows: notoginsenoside R from 6.63% to 10.50%, ginsenoside Rg from 21.30% to 39.00%, Re from 1.99 to 4.89%, Rf from 0.04 to 0.26%, Rb1 from 37.49 to 50.37%, Rc from 0.03 to 0.04%, Rb2 +Rb3 from 0.52 to 0.93% and Rd from 8.61 to 16.62 %. From these findings, it is better to ensure the quality of Xuesaitong injections by multi-ingredient evaluation or fingerprinting method. In conclusion, HPLC/MS/MS provides a quick, sensitive and highly effective method for the qualitative and quantitative analysis of those components without UV absorption. Moreover, it may also be used as a potential method for the quality control of crude CM material and its related products. Key words:HPLC-MS, panax notoginseng, Xuesaitong (Sanqi) injection, notoginsenoside, ginsenoside, quality control

三七是五加科植物三七 [Panax notoginseng. (Burk.)F.H. Chen] 的乾燥根,為常用名貴中藥,皂苷類成分一直被視為三七的主要活性成分。血塞通注射液是以三七總皂苷為原料製成的滅菌水溶液,具有很好的活血祛瘀,通脈活絡等心血管藥理作用;現有標準(衛生部药品标準,中藥成方製劑第十九冊) 採用雙波長薄層色譜掃瞄法作為血塞通注射液中皂苷類成分含量測定方法。然而,皂苷這種分子量大、極性大、沒有紫外特徵吸收的成分,給分離檢測帶來極大的困難;加上薄層色譜這種開放式的分析方法,易受外界因素的影響。近年來,利用高效液相色譜法测定三七皂苷類成分時有報導,但皂苷類成分缺乏紫外吸收,紫外檢測器應用受到限制;蒸發光散射檢測技術雖較好地解決了無紫外吸收成分的檢測問題,但因其靈敏度較低,不能檢測痕量成分。因此,本研究利用高效液相色譜-質譜-質譜聯用技術,對血塞通注射液進行質量評價研究,旨在為中藥注射劑質量控制提供方法學的借鑒。本研究應用 Agilent 1100 Series LC/MSD VL Trap System 和 LC/MSD Trap SoftwareVersion 4.2 進行在綫操控及資料處理,含8mM 醋酸铵水和乙腈作為流動相,梯度洗脫分離樣品,流速每分鐘 0.4ml;電噴霧離子源,負離子模式檢測,採用全掃瞄(質荷比從200-1400u) 自動質譜/質譜及選擇反應監測對血塞通注射液中的皂苷類成分進行定性及定量分析。結果發現:多種皂苷類成分最低檢測限及最低定量限均可達到pg級,顯示了質譜檢測的靈敏度極高;定性方面,根據9個標準對照品及其質譜裂解規律,檢識了血塞通注射液中27峰的歸屬,不同來源的10批血塞通注射液比較顯示,其所含成分基本一致,且這些成分均來自於藥材本身固有成分;定量方面,利用標準曲線,測定了血塞通注射液中三七皂苷 R,人參皂苷 Rgi, Re, Rf Rbi, RC, Rb2+Rb3,Rd 等9種成分的含量,發現10批血塞通注射液中各皂苷類成分含量差異極大,即使同廠家不同批號的血塞通注射液,亦存在明顯的差異,其中三七皂苷 R6.63-10.50%,人參皂苷 Rg 21.30-39.00%, Re 1.99-4.89%, Rf 0.04-0.26%, Rbi37.49-50.31%, Rc 0.03-0.04%, Rb2 +Rb3 0.52-0.93%, Rd 8.61-16.62 %,綜合評價或指紋圖譜才是中藥注射液質量控制的有效方法。高效液相色譜-質譜-質譜聯用技術為皂苷等無紫外吸收成分提供了快速,靈敏,高效的定性及定量分析方法,將在中藥產品質量控制中發揮極其重要的作用。 關鍵詞:高效液相色譜-質譜,三七,血塞通(三七)注射液,三七皂苷,人參皂苷,質量控制。