【作者】 陈胡兰；

【导师】 董小萍；

【作者基本信息】 成都中医药大学 ， 中药学， 2009， 博士

【摘要】 紫花地丁为堇菜科(Violaceae)植物紫花地丁Viola yedoensis Makino.的干燥全草,收载于《中国药典》2005年版一部,具有清热解毒,凉血消肿的功效,主治疔疮肿毒,痈疽发背,丹毒,毒蛇咬伤等症。紫花地丁为药用历史悠久的常用中药材,具有分布广、价廉易得、临床应用广泛等特点,具有较高的研究和开发价值。目前国内外文献报道的紫花地丁药效学研究、化学成分研究较少,其有效部位或有效成分尚不明确；现有的质量控制方法尚不完善。为了进一步合理开发利用紫花地丁药材,寻找其有效成分,揭示其清热解毒,凉血消肿作用物质基础,并为药材质量标准的制订奠定基础,本课题以药效学为先导,结合现代化学分离分析技术和手段对紫花地丁有效部位的化学成分、药材质量控制方法进行了研究。1紫花地丁有效部位的化学成分研究(1)紫花地丁有效部位的筛选通过采用二甲苯致小鼠耳肿胀试验及体外抑菌试验对紫花地丁抗炎及抑菌作用的有效部位进行了筛选,结果表明,紫花地丁乙酸乙酯部位对二甲苯引起的小鼠耳肿胀有明显的抑制作用,对大肠杆菌、沙门氏菌、金色葡萄球菌、表皮葡萄球菌均有抑制作用,与紫花地丁传统水煎剂作用相似。根据该结果确定紫花地丁乙酸乙酯部位是其抗炎抑菌作用的主要有效部位。(2)紫花地丁抗炎抑菌有效部位化学成分研究根据药效学试验的结果,采用多种现代化学分离手段对紫花地丁的乙酸乙酯部位进行了化学成分分离和纯化,共分离得到16个化合物；通过1H-NMR、13C-NMR等现代波谱分析技术,鉴定出他们的结构,分别为木犀草素(luteolin)(1)、芹菜素(apigenin)(2)、槲皮素(quercetin)(3)、5,7-二羟基-3,6-二甲氧基黄酮(5,7-dihydroxy-3,6-dimethoxyflavone)(4)、柚皮素(Naringenin) (5)、芦丁(Rutin)(6)、7-羟基-8-甲氧基香豆素(7-hydroxy-8-methoxycoumarin)(7)、秦皮乙素(esculetin)(8)、咖啡酸(caffeic acid)(9)、2-羟基-1-(4-羟基-3-甲氧基)苯基-1-丙酮(2-hydroxy-1-(4-hydroxy-3-methoxyphenyl)propan-1-one)(10)、3,4-二羟基苯甲酸(3,4-dihydroxy benzoic acid)(11)、3-羟基-4-甲氧基苯甲酸甲酯(methyl-3-hydroxy-4-methoxybenzoate)(12)、奎宁酸(quinic acid)(13)、4-Muurolene-3,10-diol.(14)、Nigranoic acid.(15)、β-谷甾醇(p-sitosterol)(16)。其中,除化合物8和16外,其余14个化合物均为首次从该植物中分到。除化合物1,2,3,6,8和16外,其余10个化合物为首次从该属植物中分到。分离得到的16个化合物,以秦皮乙素含量最高,且文献研究表明,该成分具有抗炎抑菌的作用。故将其作为进一步研究紫花地丁药材质量控制方法的指标成分。2紫花地丁药材质量的控制方法研究(1)紫花地丁药材的指纹图谱研究采用反相高效液相色谱法,对来自全国多个地区的以紫花地丁名称出售的38批药材(其中29批为紫花地丁,9批为混淆品)进行了指纹图谱的研究,建立了紫花地丁药材的指纹图谱方法；以质量一致的15批药材生成的共有模式为基准,对29批紫花地丁药材进行相似度评价,结果显示紫花地丁指纹图谱具有一定的专属性,能较好区分紫花地丁药材质量的差异；将所建立的方法用于鉴别紫花地丁的混淆品,结果显示,该方法能很好区别紫花地丁与其混淆品。(2)紫花地丁药材质量标准的研究以秦皮乙素为指标成分,对紫花地丁药材质量标准的内容进行了深入研究。研究了21批药材的鉴别和检查项下相关内容；采用高效液相色谱法,建立了紫花地丁药材中秦皮乙素含量的测定方法,结果表明所建立的高效液相含量测定方法简便,准确,重复性好。本研究结果为建立科学的紫花地丁质量评价方法奠定了重要基础。综上,本课题确定了紫花地丁的乙酸乙酯提取部位是其抗炎、抑菌作用的有效部位,并对该部位的化学成分进行了研究；在此基础上,建立了紫花地丁药材专属性指纹图谱,对药材的质量标准进行了研究。研究结果为最终揭示紫花地丁药效物质基础,建立科学、完善的紫花地丁药材质量评价方法,对药材的进一步开发利用奠定了重要基础。更多还原

【Abstract】 Viola yedoensis Makino. is the dried whole grass of perennial herbage Viola yedoensis Makino which belongs to the genus Viola. Viola yedoensis Makino. is commonly used in TCM. According to the theory of Chinese medicine, Viola yedoensis Makino. has the functions of heat-clearing and detoxification, cooling the blood and detumescence, it isapplied to furunculosis, swelling, superficial infection, carbuncle of the back, erysipelas, venomous snake bite, and all kinds of inflammation. For further reasonable development and application, we have systematically studied on the chemical constituents of effective parts, and quality control methods of Viola yedoensis Makino..The models of xylene-induced mice ear swelling and inhibiting bacteria in vitro were used to study anti-inflammatory and inhibiting bacteria in vitro effects of different extracted parts of Viola yedoensis Makino.. The results showed that the ethyl acetate extracted part of Viola yedoensis Makino. had prominent biological activities of anti-inflammatory and inhibiting bacteria in vitro, which is similar to the water decoction.These results showed that the ethyl acetate extracted part is the effective parts of Viola yedoensis Makino.In order to find the effective substance basis of Viola yedoensis Makino., we studied the chemical constituents of Viola yedoensis Makino. by phytochemical researches and spectroscopic analysis.16 compounds were isolated from the ethyl acetate extracted part of Viola yedoensis Makino. and were identified as follow: luteolin (1), apigenin(2), quercetin (3),5,7-dihydroxy-3,6-dimethoxyflavone (4), Naringenin (5), Rutin(6),7-hydroxy-8-methoxycoumarin (7), esculetin (8), caffeic acid (9),2-hydroxy-1-(4-hydroxy-3-methoxyphenyl)propan-1-one (10),3,4-dihydroxy benzoic acid (11),methyl-3-hydroxy-4-methoxy benzoate(12),quinic acid(13), 4-Muurolene-3,10-diol(14), Nigranoic acid(15),2-hydroxyoleanolic acid((16), (3-sitosterol(17). They were first obtained from this plant for the first time besides compound 8 and 17,11 compounds were isolated from the genus of viola for the first time.The research on HPLC fingerprint chromatogram of Viola yedoensis Makino. was done. Total 38 batches of Viola yedoensis Makino. samples and their adulterants were analyzed. The mutual mode to HPLC fingerprints was set up, The similarities among Viola yedoensis Makino. samples and their adulterants of different sources were obtained. Simple, rapid, stable and reproducible. It can be uesd to identify Viola yedoensis Makino. and its adulterants.In aspect of the quality standard for Viola yedoensis Makino., the identification of characteristics, micro-construction and TLC was carried out; The water contents, ash contents and esculetin contents of 21 batch samples were determined by RP-HPLC for the first time.In short, through the studies mentioned above on Viola yedoensis Makino., it partially revealed the pharmacodynamic material basis and supplied a gap of identification of TLC, quantitation determination, fingerprint chromatogram. Our study provided the theoretical basis for the further study, development and application of Viola yedoensis Makino., and had great referential significance of the modifying and consummating of the quality standard for Viola yedoensis Makino. Meanwhile, these researches highly promote modernization of Viola yedoensis Makino.更多还原