Kushen, one of the traditional Chinese medicines, is the dried root of Sophora flavescens Ait. Recent studies have demonstrated the multiple pharmacological actions of Kusgen such as anti-tumour, anti-inflammation, anti-bacterial and anti-viral activities. In this thesis, four flavonoids were isolated from Kushen. And HPLC methods were developed for quantification of alkaloids and flavonoids in Kushen, respectively. Near-infrared diffuse reflectance spectrometry (NIRDRS) was also first developed for identification and quantification of Kushen. Based on the studies above, quality control of Kushen was improved. The thesis is consist of four chapters. Chapter 1 is a review of recent researches on Kushen, including chemical components, quality control, bioactivities and clinical uses. Chapter 2 is for the preparation and identification of chemical compounds from Kushen. Four flavonoids (trifolirhizin, kurarinone, kushenol C and kushenol P) wereisolated by chromatography and characterized by UV, MS, H-NMR and 13C-NMR.The purity of the four compounds is more than 98%, which is suitable for quality control of Kushen.In chapter 3, HPLC methods were developed to quantitate the contents of alkaloids and flavonoids in Kushen, respectively. In brief, the contents of five activealkaloids such as matrine, oxymatrine, sophocarpine, oxy sophocarpine and sophoridine in Kushen were determined by using HPLC. A carbohydrate column(250mmx4.6mm 5µm) was used and eluted with acetonitrile-ethanol-water(pH2)(79:11:10). The detection was at 220 nm. Then, RP-HPLC method was first developed for simultaneous determination of three flavonoids ( kurarinone, kushenol C and kushenol P) in Kushen. Both methods are simple, accurate and repeatable. Using the methods, contents of the investigated compounds in Kushen from different places were determined. On the other hand, NIRDRS was first developed for identifying Kushen. Amodel was established based on the contents of oxymatrine and total alkaloids inKushen from different sources and their NIRDRS sperctra. Using this model, qualityof Kushen could be controlled. The predicted values of oxymatrine and total alkaloidsin Kushen by extra validation were well in accordance with the true values. This is a new, rapid, simple and inexpensive method for controlling the quality of Kushen..Chapter 4 is the study of the anti-tumour activities of Flavonoids in Kushen. In-vtro anti-tumour activities of the four isolated compounds from Kushen, Trifolirhizin, Kurarinone, Kushenol C and Kushenol P, were screened by using HL-60,BGC-823 and MDA-MB-435 tumour cell lines. The results showed that Kurarinone inhibited the HL-60 and MDA-MВ-435 сells at least 50%, while Trifolirhizin, Kushenol C and Kushenol P did not show any significant inhibition. Chapter 5 is the conclusions of this study. Keywords Sophora flavescens Ait Alkaloid Flavonoid Quanlity control

中藥苦參[Radix Sophorae flavescentis (Kushenj] 為豆科槐屬植物苦參(Sophora flavescens Ait.)的根,為中國藥典收載品種之一,具有清熱解毒,怯風燥濕,殺蟲止癢的功效。現代研究表明苦參中所含的生物鹼和黃酮類化合物具有抗腫瘤、抗菌、抗病毒等多種生物活性。本文針對中藥苦參質量控制的局限性,對苦參標準物質及兩類化學成分、多個指標的定量控制展開研究。首次以HPLC法測定了苦參黃酮類成分,建立了快速、簡便的近紅外漫反射定性、定量控制苦參質量的方法,填補了苦參在該項研究的空白,為中藥苦參的標準化、現代化定了一定基礎。 全文共分4章。第一章 文獻綜述作者總結和綜述了近年來苦參在化學成分、質量控制、藥理活性及臨床應用四方面的研究進展。第二章 苦參黃酮標準物質研究以研究並獲得苦參黃酮標準物質為目的,建立了對照品製備工藝,利用溶劑法和色譜法從苦參中分離得到4個化合物,經光譜技術和理化鑒定手段鑒定了化合物的結構,分別為三葉紫檀(Trifolirhizin),苦參酮(Kurarinone)、苦參醇 C(Kushenol C)和苦參醇 P(KushenolP), HPLC-ELSD 檢測含量均大於98%。第三章 苦參的質量控制研究以苦參質量控制為目的。收集了不同省、市、自治區的12份苦參商品藥材。以苦參險(Matrine)、氧化苦參鹼(Oxymatrine)、槐果 (Sophocarpine)、氧化槐果鹼(Oxysophocarpine)、槐定鹼(Sophoridine)、苦參酮、苦參醇C及P為苦參的定量測定指標,建立了 HPLC 測定苦參生物鹼和黃酮類成分含量方法。用 Carbohydrate Analysis column (250mm×4.6mm, 5um)以乙腈-乙醇-磷酸水溶液(pH=2) (79:11:10)為流動相;流速 1ml/mim,檢測波長 220nm;柱溫35℃,同時測定了槐果鹼、苦參鹼、氧化槐果鹼、槐定鹼、氧化苦參鹼等5個生物鹼的含量。苦參鹼、氧化苦參鹼、槐果鹼、氧化槐果鹼、槐定鹼測定方法的回收率分別為96.00%、98.56、96.90%,99.24%、97.19%; RSD分別為 1.70%、2.69%,1.32%、1.84%、1.33%。首次以 ZORBAX SB-C18 柱(250mm×4.6mm, 5um),以甲醇—乙腈-磷酸水溶液(pH=2.5)(66: 4:30)為流動相,在265mm 檢測波長下,同時測定苦參酮、苦參醇C和P等二氫黃酮及黃酮醇的含量 苦參酮 苦參醇C和P測定方法的回收率分別為98.08% 97.18% 98.13%, RSD分別為1.86% 1.91% 1.83%。用建立的苦參生物、黃酮的HPLC含量測定方法,測定了12份不同商品來源的苦參藥材,提出苦參中苦參鹼、氧化苦參鹼、槐果鹼、氧化槐果鹼、槐定鹼、苦參酮、苦參醇C和P的含量限度建議,方法簡便、準確,可控性強,可用於苦參的質量控制。運用近紅外漫反射光譜法(near-infrared diffuse reflectance spectrometry, NIRDRS),首次對不同產地苦參進行了初步的聚類分析定性鑒別研究。建立的圖譜庫,用聚類分析法對12 個不同產地的苦參通過 NIRDRS 原圖即可進行唯一鑒定;以HPLC法、酸鹼滴定法測得的數據為苦參藥材中氧化苦參鹼、總生物鹼的標準值,應用 BrukerOPUS / QUANT-2 定量分析軟件中 PLS 法進行數據處理,建立了苦參藥材NIRDRS 的數學模型,對未知苦參樣品中氧化苦參鹼及總生物鹼進行預測,預測數據與 HPLC 法、酸鹼滴定法測定結果比較,較為一致;表明 NIRDRS 技術對於中藥的鑒定及質量控制具有一定的意義。第四章苦參黃酮的抗腫瘤活性研究 對分離得到的三葉紫檀苷、苦參醇C、苦參醇P及苦參酮4個苦參黃酮類成分,進行體外抗腫瘤活性篩選,篩選結果表明三葉紫檀苷、苦參醇C、苦參醇P對HL-60、BGC-823 和 MDA-MB-435腫瘤細胞沒有表現有細胞毒活性,而苦參酮對 HL-60及MDA-MB-435 細胞的增殖有一定的抑制作用。第五章 結語 通過對苦參質量控制研究,作者首次研究建立了快速、簡便的近紅外漫反射定性、定量控制苦參質量的方法;為全面系統的評價苦參質量,建立了同時測定多種生物鹼及多種黃酮類化合物的HPLC法為使苦參質量控制體系更趨完善,提供了科學依據。 關鍵字 苦參 生物鹼 黃酮質量控制