【作者】 张海悦；

【导师】 周义发；

【作者基本信息】 东北师范大学 ， 生物化学与分子生物学， 2011， 博士

【摘要】 荨麻属植物狭叶荨麻（Urtica angustifolia）资源丰富,主要分布于东北、内蒙古、河北等地,在我国是传统的可药可食野生植物。具有袪风除湿,活血止痛,平肝定惊,消积通便,解毒,抗疲劳等功效。在东北地区民间常用于治疗风湿性关节炎、荨麻疹、糖尿病等症。在欧洲,狭叶荨麻的同属植物异株荨麻有悠久的药用历史,目前,国外已开发出多种用于治疗风湿性关节炎、良性前列腺增生的药物和保健品。而国内关于狭叶荨麻的研究很少,为充分利用东北地区丰富的植物资源,本文研究了狭叶荨麻降血糖和抗疲劳的有效活性成分。首先对狭叶荨麻的各部位的化学成分进行初步鉴定,通过小鼠糖耐量试验寻找狭叶荨麻叶中降血糖活性成分。然后通过水提醇沉得到狭叶荨麻根多糖,通过强迫小鼠游泳实验研究狭叶荨麻根多糖的抗疲劳活性。最后再经过离子交换和凝胶柱层析相组合的方法将狭叶荨麻根多糖全面系统地分级,获得狭叶荨麻的各种多糖级分,并对其进行纯度和均一性鉴定,分析了其单糖组成和结构。研究结果如下:狭叶荨麻根、茎、叶化学成分进行系统鉴定结果表明,根中含有黄酮、甾醇、苷类、酚类化合物及糖类、蛋白质等成分;茎中含有多糖类、甾体及萜类、有机酸等成分;叶中含有色素、黄酮、甾醇、苷类、糖类、蛋白质、油脂等成分。通过对狭叶荨麻不同部位根、茎、叶的提取物进行小鼠的糖耐量试验,初步筛选出狭叶荨麻叶具有降血糖的作用,对狭叶荨麻叶进一步筛选降血糖活性成分,通过对狭叶荨麻叶进行提取,所得4种提取物进行小鼠糖耐量试验,最终得到提取物3和提取物4具有降血糖作用,对提取物3和提取物4进行化学分析和结构鉴定,结果表明:提取物3为7位含有糖的山奈苷类化合物、提取物4为胡萝卜苷。对狭叶荨麻降血糖活性成分的研究中,发现用狭叶荨麻根多糖饲喂小鼠后没有降血糖效果,而显示出具有抗疲劳活性,因此对狭叶荨麻根多糖抗疲劳活性进行了研究。首先通过水提醇沉、Sevag法脱蛋白得到狭叶荨麻根多糖UA（1.6%）;然后用UA进行小鼠的抗疲劳实验,其结果表明:狭叶荨麻根多糖3个剂量组均可以延长负重小鼠的游泳时间,提高小鼠的运动能力,以高剂量组（300mg/kg/d）的剂量为佳（P<0.01）。游泳后低、中、高剂量给药组的血清尿素氮含量水平显著低于对照组,狭叶荨麻根多糖能使小鼠的运动负荷能力提高,不易发生疲劳。同时分别测试狭叶荨麻根多糖3个浓度剂量组对小鼠肝糖原的影响,结果表明,狭叶荨麻根多糖对维持肝糖原含量的高水平有积极作用,从而延缓疲劳的出现。同时对UA成分进行了初步分析,UA总糖含量为44.6%,糖醛酸含量为12.7%,淀粉含量为18.3%,蛋白含量为27.1%。经HPLC分析,狭叶荨麻根多糖主要是由葡萄糖（64.3%）,半乳糖醛酸（15.9%）以及少量的半乳糖（8.2%）和阿拉伯糖（5.9%）组成。将UA用DEAE-纤维素离子交换柱层析进行分级,先用H₂O和0.5M氯化钠水溶液分步洗脱,得到中性级分UAN和酸性级分UAA;将UAA再次用DEAE-纤维素离子交换柱层析分级,以0、0.1、0.3和0.5M的NaCl水溶液分步洗脱,得到相应的四个级分UAA-1、UAA-2、UAA-3和UAA-4。根据分子量分布情况,用凝胶柱层析将UAA-3进一步纯化分级得到酸性组分UAA-3-1和UAA-3-2。根据13C-NMR和高碘酸氧化和Smith降解结果分析,UAA-3-1中富含RG-I结构域,Gal和Ara存在多种复杂的键连方式。UAA-3-2富含HG结构域,以α-（1,4）-GalA连接方式为主。更多还原

【Abstract】 Urtica angustifolia is one species of Urtica. It is extremely rich in the northeast, Mongolia, Hebei and so on. Urtica is a kind of traditional medicine edible wild plant in China. It was reported that Urtica angustifolia has the function of blocking the wind dehumidification, promoting blood circulation and stopping pain, relaxing convulsions, antiinflammation, detumescence and anti-fatigue. In northeast of China, the people used Urtica angustifolia to care rheumatoid arthritis, urticaria, diabetes and so on. In Europe, the United States and other developed countries have researched on the nettle, especially the Urtica dioica. In particular, the development of medicine (Rheumatoid arthritis and Benign Prostatic Hyperplasia) had made remarkable achievements. In China, Urtica angustifolia has not been developed and utilized yet. In order to take full advantage of the northeastern region rich plant resources, this article studed on the hypoglycemic and anti-fatigue cmponents of the Urtica angustifolia. First,we conducted study of the chemical composition, and through oral glucose tolerance test in mice further selected hypoglycemic activity components from Urtica angustifolia; Secondly the crude polysaccharides were extracted from the Urtica angustifolia roots,and the crude polysaccharides were fractionated into fractions through the ion-exchange chromatography and Gel chromatography methods. Purity and uniformity were identificated, and using modern method to analysis composition and structure of polysaccharides. Meanwhile, the anti-fatigue activity of the polysaccharides was evaluated in mice. The investigation results are as follows:Studied on the Urtica angustifolia root, stem and leaf chemical composition, the identification results shown that the roots contained flavonoids, sterols, organic acids, glycosides, phenolic compounds and carbohydrates, protein, amino acids, fats and other ingredients; stem contained sugar and glycosides, steroids and terpenes, organic acids and other ingredients; leaf contained pigments, flavonoids, sterols, glycosides, sugars, proteins, amino acids, fats and other ingredients.Through the oral glucose tolerance test in mice, leaf decoration showed significantly hypoglycemic trend among root, stem, and leaf extracts from Urtica angustifolia. For further screening of leaf hypoglycemic active ingredients, four kinds of extractions were obtained from leaves and fed to mice. Through the oral glucose tolerance test in mice, which indicated extractions U3 and U4 had hypoglycemic effect. Also extraction U3 and U4 were flavonoids with 7’-Glc and daucosterol compounds, respectively which was identified by Chemical analysis and structural identification Study on hypoglycemic ingredients from Urtica angustifolia, after feding mice with Urtica angustifolia root polysaccharide, it was found without lowering blood sugar effect, but it shown anti-fatigue activity, therefore, the anti-fatigue activity of Urtica angustifolia roots polysaccharide was studied.The polysaccharides (UA) were extracted from the roots Urtica angustifolia with hot water, precipitated by 80% ethanol and deproteinated using the Sevag method. The polysaccharide mixture, referred to as UA(for Urtica angustifolia Polysaccharides), was obtained in a yield of 1.6%(w/w).The anti-fatigue experiment was studied in mice. It showed that three dose groups of UA were extend the swimming time of mice, improve the mouse athleticism, the high-dosage groups (300mg/kg/d) was perfect (P<0.01). After swimming, the content of serum urea nitrogen in the experimental groups were significantly lower than the control group, UA can extend exercise load capability of mice, it is not easy to occur fatigue. At the same time, glycogen content in mice were tested, the influence of the results shown that UA can maintain the high leve content of hepatic glycogen, it has positive function, thus to delay fatigue to appear. Meanwhile, the composition of of UA has been analysised, UA contained 44.6% total sugar, 12.7% uronic acid and less than 1% protein. Sugar composition analysised by HPLC indicated that UA consisted of Glucose (64.3%), Galacturonic acid (15.9%), Galactose (8.2%) and Arabinose (5.9%).UA was separated on a preparative DEAE-Cellulose column into two fractions first: an unbound fraction(UAN)by water elution and a bound fraction(UAA)by 0.5 M NaCl elution. Then, UAA was separated on a preparative DEAE-Cellulose column into four fractions UAA-1, UAA-2, UAA-3 and UAA-4 corresponding to 0.0, 0.1, 0.2, 0.3 and 0.5 M NaCl elutions, respectively. Finally, the acidic fractions UAA-3 was separated on a preparative DEAE-Cellulose column into two fractions UAA-3-1 and UAA-3-2.Using periodate oxidation, smith degradation, and 13C-NMR to analysis the structure of UAA-3-1 and UAA-3-2. According to the data analysis, UAA-3-1 might constitute type-I Rhamnogalacturonan (RG-I), Gal and Ara have several complex connections. The 13C-NMR result showed that UAA-3-2 might constitute Homogalacturonan (HG) linked toα-(1,4)-GalA.更多还原