澳门大学硕士学位论文 ABSTRACT Modern analytical techniques, including thin layer chromatography (TLC), high performance liquid chromatography (HPLC), gas chromatography (GC) and high performance capillary electrophoresis (HPCE), play a more and more important role in the quality control of Chinese medicines and/or natural medicines. Especially, capillary electrophoresis, a new technique developed in the 1980's, has been considered the most powerful and advanced branch in the field of analytical chemistry. It can be applied in different fields because of its outstanding advantages, such as high resolution, efficiency, autoimmunization, diverse separation models, little sample needed and etc. The thesis consists of two parts: 1. 2. Analysis of nucleosides and bases in Cordyceps using capillary electrophoresis (CE) was performed. The main parameters, such as boric acid concentration, pH and the proportion of acetonitrile used as the modifier for CE, were optimized by using orthogonal test and central composite design (CCD). Resolution (Rs) was used as the marker for evaluating the response function. The results showed that CCD is more powerful than orthogonal test on the optimization of CE conditions. Six nucleosides including adenine, adenosine, uracil, uridine, guanosine and inosine in Cordyceps were determined using adenosine-5'-phosphate as internal standard. The data performed the cluster analysis using the SPSS software, which showed CE was a simple method for discrimination of natural and cultured Cordyceps. Furthermore, adenosine and inosine were optimized as the markers for quality control of Cordyceps. On the other hand, an attempt for analysis of nucleosides in Cordyceps by capillary electrochromatography (CEC) was also performed. The results showed bubble was the problem. A capillary zone electrophoresis (CZE) method was developed for analysis of 5 anthraquinones including rhein, emodin, aloe-emodin, chrysophanol and physcion in Rhubarb. The optimized condition was: running buffer, 0.5M boric acid-sodium hydroxide buffer (pH 8.2) with 25% acetonitrile and 25% isopropyl alcohol as organic modifier; voltage, 25KV; temperature, 25°C. The established method was accurate, simple and rapid. 3 毛细管电泳在中药质量控制中的应用研究 Key words: capillary electrophoresis (CE), Chinese medicines, Cordyceps, nucleosides, Rhubarb, antharquinones 4

澳门大学硕士学位论文 中文摘要 現代分析方法在中藥質量控制中發揮了越來越重要的作用。高效的色譜技術實現 了多種化學成分得以有效分離,使天然藥物從定性分析為主發展到以定量為主,定量 分析方法從以薄層色譜(TLC)為主逐步發展到以高效液相色譜(HPLC)、氣相色 譜(GC)、高效毛細管電泳(HPCE)等現代分析技術為主。高效的色譜技術增強了 中藥質量控制的力度,使中藥的研究水準不斷提高,進而形成了我國藥物分析的特色 領域。 毛細管電泳技術,是20世紀80年代發展起來的一種新型分離分析技術,被認為 是上世紀末分離分析領域最有影響力的分支之一,也是該領域的公認前沿。高效毛細 管電泳具有分離效率高、分離速度快、分離模式廣泛、所需樣品少、應用範圍廣、自 動化程度高等優點,在各個不同領域顯示了極其重要的應用前景。 本論文利用毛細管電泳技術進行中藥質量控制研究,主要有兩個方面的工作: 一、毛細管電泳在冬蟲夏草質量控制中的應用研究。首先,以分離度為考察指標,應 用星點設計(central composite design)優化影響毛細管電泳的主要因素(緩衝液濃度、 pH 和有機溶劑比例);採用優化後的毛細管電泳法測定天然和人工蟲草中6個核苷 類成分(腺 哈、腺苷、尿 啶、尿苷、鳥苷和肌苷)含量;並對實驗資料進行了聚 類分析。結果:優化的毛細管電泳條件是500mM 硼酸氫氧化鈉緩衝,pH8.6,乙腈 的比例為12.2%,電壓20kV,溫度20℃;測定的6種核苷類成分,腺 哈、腺苷、 尿 啶、尿苷、鳥苷的含量均以人工冬蟲夏草為高,但天然冬蟲夏草中肌苷的含量卻 遠遠高於人工冬蟲夏草;核苷類成分毛細管電泳譜可區別天然和人工冬蟲夏草及其產 品。通過對天然和人工冬蟲夏草核苷類成分毛細管電泳譜的分析和優化,首次提出, 腺苷和肌苷應是天然和人工冬蟲夏草的質量控制指標。 在上述研究的基礎上,對毛細管電色譜分析冬蟲夏草的核苷類成分進行了初步探 索,為進一步研究奠定了基礎。 二、毛細管電泳在大黃質量控制中的應用研究。首次建立了大黃酸、大黃素、蘆 薈大黃素、大黃酚、大黃素甲醚5種蔥 類成分分離的毛細管自由區帶電泳(CZE) 法,並成功地應用於大黃藥材中此類成分的含量測定。優化的CZE 條件為:500mM 1 毛细管电泳在中药质量控制中的应用研究 硼酸緩衝溶液(pH8.2)含25%乙腈,25%異丙醇,操作電壓為25kV,溫度為25℃。 此方法簡單快捷,重現性好,結果穩定可靠。 關鍵字:毛細管電泳,中藥,質量控制,冬蟲夏草,大黃 2