澳門大學碩士學位論文 Abstract: This task is one subtask of the modifications for the China Pharmacopoeia (2005 edition). The object of our research is to supply a reference standard of the chemical qualitative and quantitative method for quality control of Fritillariae cirrhosae materials. This thesis can be divided into four sections. In section I, the progress on researches of chemical constituents and their pharmacological activities of Fritillariae cirrhosae materials was reviewed. Based on the previous studies, our researching schemes were fetched out. In section II, the plant morphology, drug characters and the micrograph properties of Fritillariae cirrhosa materials were described. At the same time, a UV scan method and thin layer chromatography (TLC) methods were studied for the identification of those different bulbus Fritillaria species. In our current studies, firstly, we compared the UV scan spectra and their 1st derivative spectra of the extracts with 7 different extraction solvents among 3 Fritillaria species including Fritillaria cirrhosa D. Don., F. thunbergii Miq. and F. ussriensis Maxim. According to the results, four solvents including water, ethanol, n-hexane and cyclohexane were selected. Moreover, the UV scan spectra and their 1st derivative spectra of the extracts with those 4 solvents were further studied among 9 Fritillaria species which were recorded in China Pharmacopоeia (2000 edition). Secondly, 5 main active alkaloids were selected according to the previous studies with our research group as the index points for the TLC differentiation of Fritillaria cirrhosa species. Furthermore, those 9 Fritillaria species were differentiated with our optimization chromatographic conditions. Based on our researching results, a chemical differentiation method for Fritillaria materials was established and supplied as a qualitative method. In section III, a HPLC-ELSD method was established to detect the main active components in different sources of Fritillariae cirrhosae materials. Peimissine was selected among five alkaloids as the index point to control the quality of Fritillariae cirrhosa materials according to the analysis of 4 species of standard materials. In the further studies of 33 batches of different sources of Fritillariae cirrhosae materials, the content of peimissine was detected and the lower limit of its content in that material was ΙΠ 川貝母質量標準及貝母藥材中主要活性生物鹼的比較研究 stipulated and the quality control standard was established. This standard was supplied as a reference to the China pharmacopoeia to control the quality of the Fritillariae cirrhosa materials. The supplied standard filled the gap which there is no quality control method in the current China pharmacopoeia. In section IV, we used the established HPLC-ELSD method to compare the amount and distribution of 5 active alkaloids in 9 different species of Fritillaria materials which recorded in the current China Pharmacopoeia (2000 Edition). The corresponding peaks were identified with HPLC-MS method to ensure its veracity. This study supplied a basal chemical data to the differences among those 9 Fritillaria species. In our present studies, a chemical qualitative method was recommended to differentiate the Fritillariae cirrhosae materials from some other Fritillaria materials recorded in current China Pharmacopoeia, and a quantitative method was established to detect the main active alkaloid in 9 species of Fritillariae cirrhosae materials. This established method was confirmed with a HPLC-MS method by identifying the corresponding peaks. These studied supplied an exclusive, repeated and feasible method to detect the content of the active component in Fritillariae cirrhosa materials. Key Words: Fritillariae cirrhosae; Fritillaria; plant modality; medicinal materials characters; micrographs; UV scanning spectra; thin layer chromatography (TLC); high performance liquid chromatography- evaporating light scattering detection (HPLC-ELSD); HPLC-MS; qualitative analysis; quantitative analysis IV

澳門大學碩士學位論文 摘要 本課題為2005年版中國藥典標準修訂的子課題,其研究的目的在於為2005年中 國藥典川貝母項下增加定性鑒別的化學方法以及增加含量測定方法提供參考標準。整 個論文研究包括4個部分:第一部分 國內外研究概況;第二部分 川貝母藥材定性分 析方法研究;第三部分川貝母定量標準分析方法研究;第四部分 5種生物鹼在9種 貝母中的分佈和含量差異的比較分析。 在第一部分中,我們總結和綜述了國內外學者對川貝母有效成分的化學分析及藥 理活性的研究進展,總結了川貝母的定性、定量分析方法的研究現狀,為建立川貝母 質量標準的進一步研究提供了參考資料。 在第二部分中,我們從植物形態、藥材性狀、顯微特徵上對川貝母進行了描述, 同時應用紫外掃描、薄層色譜等化學方法對川貝母的定性鑒別進行了研究。在我們的 研究中,我們從整體組分的角度出發,首先應用紫外掃描的方法對貝母類藥材進行了 鑒別分析。通過對比分析川貝母、浙貝母和平貝母的7種不同溶劑提取液的紫外掃描 光譜及其一階導數光譜,從中選取了其中差異較大的4種溶劑作為提取溶劑,進一步 對比分析了9種貝母的4種不同溶劑提取物的掃描光譜和一階導數光譜。其次,在本 課題組多年對貝母有效成分的藥理研究基礎上,選取了5种主要活性生物 為指標, 建立了川貝母的薄層色譜分析方法,並應用優化的薄層色譜條件對2000年版中國藥 典收載的5類9種貝母類藥材進行了區分。通過對研究結果的分析,初步制定了川貝 母的化學鑒別方法,為2005年中國藥典川貝母項下增加化學鑒別提供了參考方法。 在第三部分中,我們選取了5種主要活性生物鹼成分為指標,通過對4個基源川 貝母藥材的分析,從中選取了能代表川貝母特徵的主要成分貝母辛作為定量指標,建 立了川貝母活性成分的含量測定方法。進一步根据33 批次不同產地、不同來源的川 貝母樣品中貝母辛的含量分析結果進行總結,規定了川貝母中貝母辛的含量下限,建 立了川貝母質量控制的標準,填補了中國藥典中川貝母項下一直以來沒有含量控制的 空白,為2005年中國藥典川貝母項下增加含量測定提供了參考。 在第四部分中,我們以貝母類藥材臨床應用的中醫理論為基礎,對比分析了 5 种主要活性生物鹼在9种貝母藥材中的含量分佈情況。通過 HPLC-ELSD 含量測定方 法的建立以及對9种貝母藥材的成分分析,從化學物質基礎上為這些貝母藥材的臨床 用藥差異提供了研究數據。為了驗證該方法的準確性,我們同時應用標準對照法,採 川貝母質量標準及員母藥材中主要活性生物鹼的比較研究 用HPLC-MS 對HPLC-ELSD 檢測下的主要色譜峰進行了確證,結果表明我們所建立 的HPLC-ELSD 方法檢測得到的主要活性成分的色譜峰與對照品相一致,該方法用於 同時測定貝母藥材中5中活性生物鹼含量的準確性良好。 本論文通過對川貝母及貝母類藥材的成分分析,最終建立了川貝母的定性鑒別參 考方法及含量測定參考標準,從物質基礎研究上對不同貝母藥材的成分差異進行了 述。本研究在分析研究多批貝母藥材的基礎上,制定了川貝母品質控制的參考標準, 並通過HPLC-MS對主要色譜峰進行了歸屬,以保證制定的含量測定標準具備專屬性、 重現性和可行性。 關鍵詞:川貝母,貝母,形態描述,藥材性狀,顯微鑒別,紫外掃描,薄層色譜, 高效液相-蒸發光散射,液質連用,定性分析,定量分析 II