【作者】 杨蓉；

【导师】 梁冰；

【作者基本信息】 四川大学 ， 生物化工， 2007， 硕士

【摘要】 观音苋是一种有药用价值的无公害野生蔬菜，其营养成分丰富，在保健品、医药、天然色素等方面均有着良好的开发前景。但迄今观音苋仅作为蔬菜在利用，未经深加工，经济效益较低。为了综合开发利用观音苋资源，本研究以四川成都观音苋植物茎叶为原料，对其主要成分的提取、分离和分析鉴定方法进行系统、基础性研究，为今后观音苋化学成分及药理作用的深入研究提供理论依据和实验参考。考察酸性染料有机相分光光度法、剩余酸性染料（水相）分光光度法和本文首次提出的剩余酸性染料（水相）—薄层扫描法在观音苋生物碱含量测定中的应用，结果，酸性染料有机相分光光度法干扰大，测定结果不准确；剩余酸性染料（水相）分光光度法测得观音苋干品中总生物碱为0.013％（RSD 4.4％，n=3）；剩余酸性染料（水相）—薄层扫描法测得结果为0.018％（RSD 8.5％，n=3）。后两者测定结果准确、稳定，适用于观音苋中总生物碱的测定。通过一系列条件的选择，提出了一条观音苋总生物碱的提取工艺路线，总生物碱得率为0.072％。对得到的总生物碱用制备薄层色谱分离后，再用薄层色谱和高效液相色谱法对分离得到的生物碱进行分析，以盐酸罂粟碱为对照品，确定其中含有五种生物碱，其中四种是与罂粟碱不同的生物碱。在“氨水碱化原材料、95％乙醇提取生物碱”的流程中黄酮类成分被一并提取出来，为了综合利用此资源，采用饱和正丁醇提取黄酮类成分。该工艺粗黄酮得率2.87％，采用NaNO₂—Al（NO₃）₃—NaOH体系分光光度法检测观音苋干粉中黄酮含量，结果为0.339％。为了充分利用资源，将用95％乙醇提取过生物碱和黄酮类的观音苋残渣烘干后，用“水提醇沉”法提取多糖类物质，采用Sevag法除蛋白后多糖得率为2.99％，而采用苯酚-硫酸法检测观音苋干粉中多糖含量约为0.76％。将得到的粗多糖用葡聚糖凝胶（Sephadex G-150）进一步分离、纯化，得到三种多糖。采用薄层色谱法分析水解多糖的单糖成分，以乙酸乙酯：甲醇：乙酸：水=13：3：3：1.5为展开体系，表明观音苋多糖由D-半乳糖、D-葡萄糖、D-果糖和D-木糖组成，还有一种对显色剂显黄色的成分未能鉴定。采用体外细胞试验法检测观音苋粗黄酮和粗多糖对宫颈癌Hela细胞的抑制作用，用MTT法评价其效果，结果二者对Hela细胞均不具有杀伤作用。另外，在研究过程中还得到了几种晶体，未能确证其为何物质，有待进一步分析、研究。更多还原

【Abstract】 Gynura bicolor （Roxb.） DC., which had medicinal values, was an pollution-freewild vegetable. Its nutrition components were very rich, and the developmentprospects of it in health products, medicines, natural pigments and so on were verygood. But up to now, it’s just utilized as the vegetable. The economic benefit waslow, because it’s not be deep processed. For comprehensively developing andutilizing Gynura bicolor （Roxb.） DC, with the stems and leaves of Gynura bicolor（Roxb.） DC, which planted in Chengdu Sichuan, as raw material, systematicly andbasicly studied the extraction, separation, analysis and identification methods of itsmain constituents. Supplied the theoretic foundation and experimental references forthorough researching its chemical constituents and pharmacological actions.Studied three different methods for determination of alkaloids’ content inGynura bicolor （Roxb.） DC, the first one was Acidic Dye organic phaseSpectrophotometry, the second was Residual Acidic Dye （water phase）Spectrophotometry and the third was Residual Acidic Dye （water phase）-TLCScanning method. The first method was interferenced seriously and gave unaccurateresults; By the second method, the content of alkaloids in Gynura bicolor （Roxb.）DC.was 0.013%（RSD 4.4%, n=3）. By the third one, the content was 0.018%（RSD8.5%, n=3）. So the measurement results of the latter two methods were accurate andimmobile, and both suitable for the determination of alkaloids’ content in Gynurabicolor （Roxb.） DC.By a series research, brang up a routing to extract the total alkaloids in Gynurabicolor （Roxb.） DC. and its yield was 0.072%. The total alkaloids were separated byPreparation TLC, then analyzed by TLC and HPLC. Papaverini Hydrochloridum was the reference substance, five different alkaloids were identified, and the fourkinds of them were different to Papaverine.In the routing of "Alkalization treatment of raw materials by ammonia liquor,extraction of alkaloids by 95% enthanol", the flavonoids were extracted together.For comprehensive utilization, used n-Butanol to extract flavonoids from the acidicwater. The yield of flavonoids was 2.87% in this technology. By Spectrophotometryin the NaNO₂—Al（NO₃）₃—NaOH system, the content of flavonoids in Gynurabicolor （Roxb.） DC. was 0.339%.Polysaccharides were extracted by water extract-ethanol precipitation methodfrom the residue of Gynura bicolor （Roxb.） DC.. The yield of polysaccharides afterremoving protein by Sevag method was 2.99%. They were separated and purifiedinto three fracions by dextran gel（Sephadex G-150） chromatography. The content ofpolysaccharides was detected by phenol-sulfuric acid method, the result was 0.76%in Gynura bicolor （Roxb.） DC..The components of polysaccharides’s hydrolyzates were measured by means ofTLC, and the developing system was ethyl acetate: methanol: acetic acid: water=13:3:3:1.5. The result showed that it’s composed of D-galactose, D-glucose,D-fructose, D-xylose. And there was one component still couldn’t be identified,which was a yellow spot.Studied the inhibitory effect on cell proliferation of cervical carcinoma Helacells by flavonoids and polysaccharides in Gynura bicolor （Roxb.） DC.. Cervicalcarcinoma Hela cells were detected with MTT assay. As a result, both of them didn’thave inhibitory effect on cell proliferation. Specially, in the other processes ofextraction, many kinds of crystals were obtained, they couldn’t be identified. So allof these were need to further study.更多还原