多花黄精组培体系建立及甾体皂苷成分提取

发布时间：2018-01-03 02:36

本文关键词：多花黄精组培体系建立及甾体皂苷成分提取　出处：《安徽大学》2017年硕士论文　论文类型：学位论文

【摘要】：多花黄精(Polygonatum cyrtonema Hua),隶属百合科(Liliaceae)黄精属(Polygonatum),性平、味甘,具有补肾益精、降血糖、降血压、降血脂、提高人体免疫力、抗衰老、抗肿瘤等功效,是药食同源的多年生草本植物,为中国传统大宗药材,具有很好的开发与应用价值。多花黄精中的甾体皂苷主要有薯蓣皂苷、薯蓣皂苷元等,具有降血糖、抗肿瘤、抗HIV的生物活性。在人工栽培中受种子发芽率低下、育苗周期长等因素影响,多花黄精通过自然繁殖的方式无法满足市场需求。较传统栽培方式,组织培养不受季节、取材数量等因素制约,可在短期内大规模繁殖性状稳定的优良种苗。同时毛状根具有生长速度快、合成次生代谢物质能力强且稳定等优点,因此,可以利用毛状根液体培养技术生产甾体皂苷。本研究建立UPLC法同时测定多花黄精野生植株和毛状根中薯蓣皂苷和薯蓣皂苷元的含量,为黄精资源的利用提供科学依据。研究结果如下:1.低温沙藏、培养基中添加不同浓度激素处理种子,找出萌发率较高的处理方法,即0℃沙藏100d,接种到MS+2.0mg/L6-BA培养基中,野生多花黄精种子发芽率最高,达到43.7%。2.通过三因素三水平正交实验,筛选出多花黄精最适快繁培养基配方为:MS+1.0 mg/L 6-BA +0.5 mg/L NAA +0.3 mg/L IAA,芽增殖系数达到 3.2,并且长势良好;最适生根培养基为1/2MS+1.0mg/LNAA,平均生根率达到86.92%;炼苗最适基质为泥炭土:沙子2:1。3.采用正交设计方法,探究毛状根最佳液体培养条件:培养基为液体1/2 MS,pH为5.8,摇床转速150 r/min,26℃暗培养,增殖倍数达到9.4。4.优化超声波法提取甾体皂苷的提取方法,以75%的乙醇作为提取剂,料液比1:15,超声2 h后,用正丁醇萃取。5.采用超高效液相色谱法(UPLC)对多花黄精野生植株和毛状根中薯蓣皂苷和薯蓣皂苷元含量进行统计分析,野生植株根茎中薯蓣皂苷和薯蓣皂苷元含量分别为106.27 μg/g、84.95μg/g,毛状根中薯蓣皂苷和薯蓣皂苷元含量分别为 89.13 μg/g、72.24μg/g。
[Abstract]:P.cyrtonema (Polygonatum cyrtonema Hua), Liliaceae (Liliaceae) Huang Jingshu (Polygonatum), flat, sweet, with a kidney Yijing, reducing blood sugar, reducing blood pressure, reducing blood fat, improve body immunity, anti-aging, anti-tumor effect, is a perennial herb medicinal and edible plant, as the traditional China the development and Application of medicinal materials, has a very good value. Steroidal saponins p.cyrtonema consist of dioscin, diosgenin, reducing blood glucose, anti tumor, anti HIV bioactivity. The low seed germination rate in artificial cultivation, breeding effect of long cycle and other factors, p.cyrtonema through natural reproduction mode can not meet the market demand. Compared with the traditional way of cultivation, tissue culture is not the season, the number of materials and other factors, can be large-scale excellent seed and seedling traits of stable propagation in a short period of time. At the same time, hairy root with fast growth, secondary synthesis The advantages, metabolism ability and stability, therefore, can make use of hairy roots in liquid culture technology to produce steroidal saponins. The establishment of simultaneous determination of dioscin and Diosgenin p.cyrtonema wild plants and hairy roots by UPLC, to provide a scientific basis for the use of Polygonatum resources. The results are as follows: 1. at low temperature sand storage, seeds of different hormone concentration added in culture medium, find a higher rate of germination processing method, namely 0 C sand 100D were inoculated into MS+2.0mg/L6-BA medium, wild p.cyrtonema seeds germination rate is highest, reaching 43.7%.2. through three factors and three levels orthogonal experiment, screening of Polygonatum cyrtonema optimal propagation medium: MS+1.0 mg/L 6-BA +0.5 mg/L NAA +0.3 mg/L IAA, bud growth coefficient reached 3.2, and grew well; the optimum rooting medium was 1/2MS+1.0mg/LNAA, the average rooting rate reached 86.92%; the optimum seedling Substrate was Peat: sand 2:1.3. by orthogonal design method, to explore the best conditions: hairy roots cultured in liquid medium for liquid 1/2 MS, pH 5.8, rotation speed of 150 r/min, 26 C dark culture, extraction of steroidal saponins extract proliferation rate reached 9.4.4. optimization by ultrasonic wave method, using 75% ethanol as extraction agent, liquid ratio 1:15, ultrasonic 2 h, extracted with n-butanol.5. by ultra performance liquid chromatography (UPLC) to carry on the statistical analysis of Dioscorea p.cyrtonema wild plants and hairy root saponin and Diosgenin content of wild plants in potato yam rhizome saponin and Diosgenin content were 106.27 mu g/g 84.95, g/g, the hairy roots of dioscin and Diosgenin content were 89.13 g/g, 72.24 g/g.

【学位授予单位】：安徽大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S567.239;R284.2

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