【作者】 梅虎；

【导师】 谈锋；

【作者基本信息】 西南师范大学 ， 植物学， 2002， 硕士

【摘要】 本试验以紫苏(Perilla frutescens(L.)Britte)为材料，进行了紫苏花芽生理分化机理以及紫苏子α-亚麻酸提取和纯化工艺的研究。 一、紫苏花芽生理分化机理研究 紫苏花芽生理分化期分为成花诱导期和花芽孕育期。成花诱导完成于六月中下旬，花芽孕育则从六月中下旬开始，完成于七月下旬。在此基础上，我们研究了土壤水分胁迫和弱光胁迫两个抑花因子对紫苏花芽生理分化的影响，以探讨紫苏花芽生理分化机理。主要研究结果如下： 旺盛的DNA复制、RNA合成为花芽生理分化所必需。花芽孕育期中期DNA和RNA含量的升高对于花芽生理分化有明显的促进作用。RNA／DNA值急剧降低为花芽生理分化的一个明显特征。 激素对花芽生理分化的作用，可能通过时间和空间上调节激素之间比例或平衡以及核酸的代谢来完成对花芽诱导和孕育的调控。主要表现在：①时间上的控制。4种内源激素峰值顺序出现：IAA，GA(成花诱导期)→ZRs(花芽孕育前期)→ABA(花芽孕育中期)。②空间上的控制。在成花诱导期中，IAA／GA值的变化可能起着关键的主导作用；在花芽孕育期中ZR_s／GA值与成花率具有明显的正相关性。 各碳水化合物、可溶性蛋白含量的增加为紫苏花芽生理分化所必需。这可能与花器原始体形成所需碳水化合物和酶的合成有关。总氮含量在成花诱导期后持续下降，至花芽孕育期中期达到最低值。C／N值在诱导期至孕育期前期呈上升趋势，孕育期后期开始回落，相对高的C／N值可能有利于花芽诱导和孕育的完成。此外我们还研究了α-淀粉酶(α-AMY)、硝酸还原酶(NR)活性的变化以及淀粉酶同工酶谱的变化。 中英文摘要 水分对紫苏花芽生理分化影响最为显著时期为花芽孕育期的中后期。持续的土壤水分胁迫在此期引起膜系统的伤害，严重影响花芽生理分化的进行和最终的成花率。而光照对紫苏花芽生理分化影响最显著的时期为孕育期前期，持续的弱光胁迫在此期导致净同化作用速度明显低于对照，最终对成花率产生显著的影响。二、紫苏子a．亚麻酸提取与纯化工艺研究 （一）苏子油的提取 紫苏子经预处理后，我们对提取方法、提取溶剂、以及影响提取效率的其它参数进行了比较和研究。结果显示：在本次试验中苏子油的最佳提取条件为：4倍质量体积的石油醚：乙酸乙酯Oj）为提取溶剂，提取温度 60 oC，提取时间12h，提取二次。得油率为32．87％，经气相色谱分析苏子油中a－亚麻酸含量为42石4％。 （二）a．亚麻酸的提取和纯化1苏子精油的制备：用 15％NaOH溶液除去少量的游离脂肪酸、磷脂、色素、蛋白质、胶质等成分，所得苏子精油中a－亚麻酸的含量为46刀1％，提纯倍数为1．079，a．亚麻酸收率为93．32％。2 改良碱解：在比较酸解法和碱解法的基础上，我们对碱解法进行了改进。苏子精油经4倍质量体积的3．7％NaOH乙醇溶液皂化后，所得滤液分别置于室温、－5．ZC、－9．soC、－15．loC、－19。7℃下保存12h。最后所得混合脂肪酸的分析结果显示：随处理温度的降低，各组a－亚麻酸含量和提纯倍数均明显上升，u．亚麻酸收率则呈降低趋势。3尿素包合：在对影响腺包反应的腺包温度、腺包比例、腺包时间、腺包次数、腮包客体、腺包溶剂研究的基础上，我们得出本次试验中腺包反应条件的最佳组合即：以混合脂肪酸为腺包客体，甲醇或乙醇为溶剂，脂肪酸：甲醇（或乙醇）：尿素二1：8：2，包合温度．10刀℃，包合时间12h，包合1次。4改良碱解和尿素包合对a－亚麻酸提取纯化的综合效应：将苏子精油在乙醇溶液中皂化后，所得滤液分别置于室温、－5．It、－10．OC和－15．ZoC下处理 12h。最后所得的混合脂肪酸分别在最优条件下进行包合。结果表明：在－5loC下进行改良碱解，然后以最优条件进行包合为最佳组合，此时液相中的a－亚麻酸含量为88．16％，得率为23．14％，提纯倍数为1．916。5硝酸银柱层析和减压蒸馏：将豚包所得混合脂肪酸甲酯化后，应用硝酸银柱层析技术或减压蒸馏技术，可使a－亚麻酸含量达到了98％以上。 （三）本次试验的主要成果和创新①应用正交试验设计得出了苏子油的最佳提取条件。 2 中英文摘要②首次应用硝酸银薄层色谱和硼酸薄层色谱对a－亚麻酸的测定进行了比较和研究。这为在常规实验条件下测定a－亚麻酸含量提供了一个可行的方法。③利用脂肪酸在乙醇溶液中随温度降低溶解度变化的规律，对苏子油碱解法进行了改进。经改良碱解使a－亚麻酸含量由常规碱解的52．45％提高到63．50儿提纯倍数由 1．140提高到l．380。④采用尿素包合法对苏子油a－亚麻酸进行了纯化研究，并对包合条件作了深入细致的研究。⑤在研究改良碱解法和尿素包合法的基础＿．〔，探讨了二者对a．亚麻酸提取和纯化的综合效应。③应用硝酸银柱层析和减压蒸馏技术对Q－亚麻酸的进一步纯化进行了尝试和比?更多还原

【Abstract】 Physiological flower bud differentiation mechanism of Perrilla frutescens(L.) Britte and technology of extracting and purifying a -Linolenic acid were studied in this paper. 1 Physiological flower bud differentiation mechanismThe results showed that the phase of physiological flower bud differentiation could be divided into two phases: floral induction and floral initiation. The floral induction accomplished in the last ten-day period of June, and from which floral initiation began and accomplished in the last ten-day period of July.On this foundation, the author studied the effect of soil water stress and low-light stress on physiological flower bud differentiation of Perilla Frutescens(L.) Britte. The major results as follows:Copying DNA and transcribing RNA vigorously were essential for the physiological flower bud differentiation. The contents of DNA and RNA went up in the middle period of floral initiation, which obviously promoted physiological flower bud differentiation. The sharply reduce in value of RNA/DNA was a clear character for the physiological flower bud differentiation.Endogenous hormones regulated and controlled physiological flower bud differentiation by the way of adjusting, probably from time and space, the proportion or balance in themselves as well as nucleic acids metabolism.The major control as follows:Controlling by the time: peak value of endogenous hormones appeared in sequence: IAA and GA’s (floral induction), ZRs’s (early in floral initiation), ABA’s(later in floral initiation).Controlling by the space: the change of IAA/GA’s value probably dominated inthe floral induction. ZRs/GA’s value possessed clear positive correlation with flower bud formation.The increase in carbohydrate and soluble protein were necessary in floral initiation, as might be related with synthesis of carbohydrate and enzyme needed in the formation of floral primordia. Total nitrogen content went on to descend after floral induction phase, and the lowest value of it came in middle period of floral bud initiation. C/N’s value tended to rise between floral induction phase and early phase of floral initiation, and reached the lowest in the middle period of floral initiation. Relative high C/N’s value probably was favor of accomplishment of the physiological flower bud differentiation. In addition, we also studied changes of the activity of a - AMY and NR as well as change of amylase isozyme band.The most evident effect of soil water stress on physiological flower bud differentiation was in the intermediary and later stage of floral initiation. The lasting soil water stress caused the harm of membrane system in this phase and restrained physiological flower bud differentiation. The most evident effect of low-light stress on physiological flower bud differentiation was in early stage of floral initiation. The lasting low-light stress obviously caused the speed of NAR lower than that of the control, and at the same time affected frequency of flower bud formation heavily.2 Technology of extracting and purifying a -Linolenic acidBased on studying the technology of extracting and purifying a -Linolenic acid, the results as follows:For extracting perilla seed oil, the best solvent was composed of petroleum ether and ethyl acetate. After perilla seed oil being extracted twice, each extraction lasting 12 hours at 60 癈, the average yield of perilla seed oil was 32.87%. Analyzed by gas chromatogram, the results showed that the content of a -Linolenic acid was 42.64%.In order to take-off the compositions such as a few dissociating fatty acids, phosphatides, pigments, proteins and so on., the perilla seed oil was refined by the treatment with NaOH solution. The results showed that the content of a -Linolenic acid, a -Linolenic acid yield and purification multiple were respectively 46.01%, 93.32% and 1.079 in refined perilla seed oil.On the base of comparing acid hydrolysis method and alkaline hydrolysis method, We improved the alkaline hydrolysis method.After being ref更多还原