Ezhu, one of traditional Chinese medicines, is derived from three species of Curcuma including Curcuma phaeocaulis Val., C. kwangsiensis S. G. Lee et C. F. Liang and C. wenyujin Y. H. Chen et C. Ling according to Chinese Pharmacopoeia. The essential oil of Ezhu was reported to possess anti-tumour and antivirus activities. Previous studies showed that the three species of Ezhu had variation in their chemical composition. However, only curcumol was usually quantitated for quality control of Ezhu because of the absence of other chemical standards. Actually, germacrone was used as marker for quality control of Ezhu in China Pharmacopoeia (2005). However, it is thought that interaction of multiple chemical compounds contributes to the therapeutic effects of Chinese medicines. In this paper, 11 chemical compounds were separated from Rhizoma Curcuma and two transformative compounds of curzerene were obtained, which were confirmed by UV, MS and NMR data. GC-MS and HPLC-DAD were developed to control the quality of Ezhu, and the chemical components in Ezhu originated from three different plant species were also compared. This thesis consists of five chapters. Chapter 1 is the literature review on Ezhu, which includes the recent development on chemical compositions and pharmacological studies of Ezhu. The advance of quality control for Ezhu has also been reviewed in this chapter. Chapter 2 focuses on the separation of chemical compounds in Ezhu. Thirteen pure compounds, including curcumenone, curzerene, furanodienone, curcumol, isocurcumenol, furanodiene, germacrone, neocurdione, curcumenol curdione, beta-elemene, callitrin and callitrisin were obtained and their chemical structures were confirmed by UV, MS and NMR data. Purity test showed that the compounds could be used as chemical standards and they were suitable for quality control of Ezhu. The qualitative and quantitative analysis of Ezhu by using GC-MS was showed in Chapter 3. For optimization of GC conditions, some heat unstable compounds in Ezhu were studied. The pathway of transformation of those compounds, including furanodiene, germacrone, curdione, curcumol and curzerene were found. And GC-MS condition was optimized. The results showed that some heat labile components degraded with the increasing temperature. Chapter 4 showed the qualitative and quantitative determination of Ezhu by using HPLC-DAD, The content of eleven compounds in Ezhu samples was determined and HPLC fingerprint of Ezhu was developed. Furthermore, GC-MS and HPLC-DAD for analysis of Ezhu was compared and evaluated in Chapter 5. Based on the work, thirteen compounds derived from Ezhu were obtained. Our results showed that curcumol, which was identified as active component in Ezhu, was undetectable in Ezhu of three species of plants. However, curcumol could be formed from degradation of curcudione at high temperature. The phenomena could be found for several compounds such as furanodiene, furanodienone, germacrone, neocurdione. Therefore, GC-MS condition for analysis of Ezhu should be optimized to avoid the degradation of heat sensitive compounds. But furanodiene and furanodienone still degraded at the optimized condition, which suggested that GC-MS should not be used for analysis of these compounds. In addition, HPLC-DAD was developed to analyze the multiple components in Ezhu. The methods of GC-MS and HPLC-DAD for quality control of Ezhu were compared and evaluated. 21 Ezhu samples originated from three species of plants were qualitative and quantitative determined by GC-MS and HPLC-DAD, and fingerprint of Ezhu was also established by using GC-MS and HPLC-DAD. In sum, the origin of curcumol in Ezhu was discovered for the first time in this thesis, and the transformative pathway of 5 heat liable compounds in Ezhu was elucidated. Simultaneous determination of eight and eleven components in Ezhu by GC-MS and HPLC-DAD were developed, respectively. Fingerprint of Ezhu was also established by GC-MS and HPLC-DAD. In addition, the difference of chemical components of Ezhu from three species was also compared, which should be helpful to the further development of quality control standards for Ezhu. Key words: Ezhu, Curcuma phaeocaulis, C. kwangsiensis, C. wenyujin, GC-MS, HPLC-DAD, heat-liable compound, transformation, cluster analysis, fingerprint, quality control, chemical standards

我術為溫郁金(C. wenyujin Y. H. Chen et C. Ling)、蓬莪術(C. kwangsiensis S. G. Lee et C. F. Liang)以及廣西装術(Curcuma. phaeocaulis Val.)的乾燥根莖。據報導, 我術油具有抗腫瘤及抗病毒等功效。由於化學對照品的缺乏, 術的品質評價研究 通常只有我術醇的定量測定, 2005 年版藥典則以吉馬酮為品質控制指標。由於中 藥為多成分的複雜體系,其效應為其多種成分綜合作用的結果,因此,以單一化學 指標難以客觀反映其品質,多指標品質控制的關鍵之一是必須具有中藥化學對照 品。為此,我們首先對莪術油中的主要化合物進行了分離純化,得到11個化合物和 2個莪術烯的轉化產物,並以UV,MS 和NMR 確證了其結構。在此基礎上,建立 了中藥義術 HPLC-DAD和GC-MS 的定性定量分析方法,並比較了三種基原的術 藥材化學成分的差異。 全文共分五章,第一章是有關莪術研究的文獻綜述,包括莪術的化學成分和藥 理學研究,以及莪術品質控制方法研究進展。第二章著重於我術主要化學成分的分 離,共分離得到十三個化合物,經 UV,MS 和NMR 確認為:β-欖香烯(β-elemene)、 我術烯(curzerene)、義術醇(curcumol)、異莪術烯醇(isocurcumenol)、味喃二烯酮 (furanodienone)、 喃二烯(furanodiene)、吉馬爾(germacrone)、 術二酮(curdione)、 義術烯醇(curcumenol)、新莪術二 (neocurdione)、我術烯酮(curcumenone)、卡利春 (callitrin)、卡利春辛(callitrisin)。所有分離得到的化合物純度均在98%以上,適合作 為品質控制的對照品。第三章是莪術的GC-MS 定性定量研究。在進行 GC 條件優化 之前,首先對莪術中部分熱敏感成分進行研究,以闡明這些成分轉化途徑。進而基 於對熱敏感成分的分析結果進行 GC條件的優化,並對三種基原21個 術樣品中八 個成分(β-欖香烯、莪術醇、異莪術烯醇、吉馬酮、莪術二酮、莪術烯醇、新莪術 二酮、莪術烯酮)進行了 GC-MS 定量測定,同時對我術的GC-MS 指紋圖譜進行了 研究。第四章是我術的HPLC-DAD 定性定量研究,包括分析條件的優化,三種基原 21 個莪術樣品中11 個成分(喃二烯、我術烯、味喃二烯酮、我術醇、異莪術烯 醇、吉馬酮、莪術二酮、莪術烯醇、新莪術二酮、莪術烯酮、β-欖香烯)的HPLC-DAD 定量測定,以及義術的HPLC-DAD 指紋圖譜的研究。第五章在比較 GC-MS 和 HPLC-DAD 分析 術的基礎上,對莪術品質控制方法進行了評價,以供莪術研究時 參考。 本文首次發現中藥莪術中莪術醇的來源,揭示了莪術中5種熱不穩定成分的轉 化規律;分別建立了同時測定義術中8種和11 種化學成分含量的 GC-MS 和 HPLC-DAD方法;比較了3種基原莪術化學成分的差異,為全面控制莪術藥材品質 提供了科學客觀的理論依據。 關鍵詞:莪術,溫郁金,蓬莪術,廣西莪術,GC-MS,HPLC-DAD,熱不穩定化合物,轉化,聚類分析,指紋圖譜,品質控制,化學對照品