【作者】 张欣；

【导师】 段玉峰；

【作者基本信息】 陕西师范大学 ， 食品科学， 2006， 硕士

【摘要】 小蓟Cephalanoplos segetum（Bge.） Kitam俗称刺儿菜、野红花、青青菜,为菊科植物,在我国分布广泛,资源丰富。它具有凉血止血,祛瘀消肿的功能,为传统的中药。小蓟同时也可以食用,作为药食两用的野生植物资源含有丰富的营养物质,为人们喜爱的民间野菜。多糖具有丰富的药理活性,且无毒副作用,已广泛应用于临床及保健品市场。为了进一步开发利用小蓟资源,本试验对小蓟多糖的提取分离、组成结构及其生物活性进行了研究。 本实验以陕西周至的小蓟干料为原料,进行了小蓟营养成分的测定;小蓟多糖的最佳提取工艺、分离纯化及对所得组分进行了纯度鉴定、分子量测定和组成分析;采用H₂O₂/Fe²⁺体系和Fe²⁺诱发卵黄脂蛋白PUFA过氧化体系研究小蓟多糖半纯品的体外抗氧化活性;通过动物实验进行了小蓟粗多糖生物学作用研究。通过以上研究,为综合开发利用小蓟资源提供参考和依据。 主要实验结果如下: 1.小蓟中灰分含量为8.760%、粗蛋白质含量为4.470%粗脂肪含量为0.430%。在已测定的17种氨基酸中,氨基酸总含量为10.302%,其中人体必需的氨基酸有7种,占氨基酸总含量的42.526%。在已测定的15种矿物元素中,含有9种人体必需元素,其中Ca、Mg的含量较高;有害元素Hg、As、Pb及Cd的含量较低。 2.热水浸提小蓟多糖的最佳工艺为:提取温度90℃,提取时间120min,料液比为1∶10。 3.超声波辅助提取小蓟多糖的最佳工艺参数为:功率250W,提取20min,料液比为1∶15。 4.木瓜蛋白酶+Sevag法为最佳脱蛋白方法,即可较完全的脱除粗多糖中的蛋白成分,又能大大降低多糖的损失率。蛋白脱除率达96.45%。 5.经过DEAE-52和Sephadex G-200柱层析分离到CSK-A’、CSK-B’、CSK-C’、CSK-D’四种多糖组分。 6.四种精制级分纯度鉴定:紫外光谱显示在260nm、280nm处均未发现核酸和蛋白质的吸收峰;醋酸薄膜电泳都得到单一色带;SephadexG-200柱洗脱后,四种多糖的洗脱峰基本均匀对称。 7.组成和结构分析:Sephadex G-200柱层析测定它们的分子量分别为20.7kD、28.6kD、148.0kD、501.2kD;气相色谱分析结果表明CSK-A’主要有木糖、半乳糖、葡萄糖组成,质量比为6.31∶8.52∶1.03,CSK-B’主要有鼠李糖、山梨糖和葡萄糖更多还原

【Abstract】 Cephalanoplos segetum（Bge.） Kitam called thorn vegetable, wild red flower, green vegetable, is one kind of feverfew and is extensively grow in China and rich in resource. As the traditional Chinese medicine, it can cool the blood and stop bleeding, and other pharmacological activities. Besides, this wild plant is fonded by people because of its abundant nutritional ingredient and can be consumed as food and medical materials as well. Cephalanoplos segetum（Bge.） Kitam is very active in pharmacological aspect while poses no poisonous effect to the body, so it is widely applied to clinical service and sold in health product market. This experiment studies its extraction, separation, components, and active functions.This experiment adopts dry Cephalanoplos segetum（Bge.） Kitam which grows in Zhouzhi within Shaanxi province as the raw material researchs its nutrition ingredients and studies the best way of extraction, separation, and also researches its purity, molecular weight, component. What’s more, this paper also focuses on its antioxidative activities by the means of H₂O₂/Fe²⁺ system and polyunsaturated fatty acid（PUFA） of the lipoprotein induced by Fe²⁺ system. This experiment is carried out by putting the polysaccharides of Cephalanoplos segetum（Bge.） Kitam in animal’s body to research its biology action. This study provides reference and gives evidence to exploit and utilize Cephalanoplos segetum（Bge.） Kitam.The main experiment result is as following:1. The content of ash in Cephalanoplos segetum（Bge.） Kitam was 8.760% , crude protein 4.470%crude fat 0.430%. The content of 17 determined total amino acids added up to 10.302%, which included 7 essential amino acids accounting for 42.526%of total amino acids. 15 mineral elements had been determined, including 9 essential elements among which the contents of Ca and Mg were higher. The contents of harmful elements Hg, As, Pb and Cd were lower.2. The best extracting technology is in hot water is: at 90℃, extracting 120min, and the ratio of material to liquid is 1:10.3. With the help of ultrasonic, the best technology parameter of extracting is: at the power of 250W, extracting 20 mins, and the ratio of material to liquid is 1:15.4. The best way of extracting protein is using enzyme combined with Sevag method, which can nearly extract the total protein in raw polysaccharide, while degrade the loss rate of polysaccharide. The protein extraction rate is 96.45%.5. Four kinds of polysaccharide as CSK-A’、 CSK-B’、 CSK-C’、 CSK-D’ areseparated by DE-52 and Sephadex G-200 chromatography.6. Four kinds of purification identification for the refined products: no nucleic acid and protein absorption peaks appear in the UV spectrum at the places of 260nm and 280nm. After the electrophoresis, there emerges a sole ribbon in the acetic acid membrane. After extracting by SephadexG-200 then, peaks of the four polysaccharides are basically and evenly symmetrical.7. Composing and Structure analysis: their molecular weights are 20.7kD, 28.6kD^ 1481dX 501.2kD respectively measured by Sephadex G-200 chromatography. The result in the spectrum indicates that CSK-A’ is mainly composed of xylose, galactose and glucose, with the mass ratio of 6.31:8.52:1.03;CSK-B’ is mainly made up of rhamnose, sorbose and glucose, with the mass ratio of 6.97:1.52:0.98;CSK-C is chiefly composed of xylose, fructose, sorbose and glucose, with the mass ratio of 7.01:8.93:3.04:1.03;CSK-D’ is formed mainly by xylose, fructose, sorbose and glucose, with the mass ratio of 2.10:9.85:1.05. Infrared spectrum reveals that the Infrared spectrums of these four polysaccharides are classical absorption spectrums, that is, at the place of 1400 cm"’-1200 cm"1 there is angle change vibration, at the place of 2930 cm" there appears C-H stretching out and drawing back vibration area, at the place of 3400 cm"1 there emerges absorption wide peak caused by O-H stretching out and drawing back vibration, at the place of 1200cm"’-1030 cm"1 there is absorption peak that is formed by C-0 stretching out and drawing back vibration. In addition, the absorption peak around the area of 1110 cm"1 shows that the monosaccharide in the polysaccharide is hexahydric heterocycle, with the point of 1640 cm"’-1600 cm"1 as the characteristical peak. At the place of 1075 cm"1-1000 in the Infrared spectrum there appears an absorption condition. So we can deduce that it is one type of B-pyrane polyoses that contains acetamide group.8. The test of antioxidative activities indicates:（1） CSK-B, CSK-C, CSK-D can remove OH that is produced by Fenton reaction through H2O2/Fe2+ system. With the increase of amount, the rate of reduction goes with the same trend, so there is a mutual relationship between reduction rate and quantity of polysaccharides, in which CSK-D poses the highest restraining effect, 1.8ml usage reaching 67.29%, with CSK-B coming as a close second. Although CSK-C has some reduction effect, it is comparatively weaker. While CSK-A almost has no such effect.（2） CSK-A, CSK-B, CSK-C has antioxidative effect to the polyunsaturated fatty acid （PUFA） of the lipoprotein induced by Fe2+ system. With the increase of amount, the rate of reduction goes with the same trend, so there is a mutual relationship between reduction rate and quantity of polysaccharides, in which CSK-C poses the strongest restraining effect, 2.5mg/ml usage reaching 53.64%, followed by CSK-A, CSK-B.While CSK-D almost has no such effect. 9. Animal experiment indicates:（1） The polysaccharides of Cephalanoplos segetum（Bge.） Kitam can help decrease the content of triglyceride（TG） and total cholesterol（TC）, increase the ratio of high-density lipoprotein （HDL-C） in the blood serum of mouse, and obviously regulate its metabolization of lipid.（2） The polysaccharides of Cephalanoplos segetum（Bge.） Kitam can stop mouse from getting old that is caused by D-galactose: it could increase the activities of blood superoxide dismutas（SOD）, glutathione peroxidase（GSH-PX）, catalase （CAT） of mice, liver and brain organization, decreased the content of the malondialdehyde（MDA）and the activity of monoamine oxidase（MAO）. Which indicate that it can reduce the speed of getting old from D-galactose.（3） The polysaccharides of Cephalanoplos segetum（Bge.） Kitam can improve the motion capability of mouse, lower the content of blood urea nitrogen（BUN） of post-exercise mice, decrease the consumption of liver glycogen and increase the activity of lactic dehydrogenase（LDH）. Which prove that it can effectively ease the hurt to mouse that is caused by over-motion.更多还原