棉花植保素与桔梗皂甙的诱导生成研究
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摘要
本文采用多种类型的诱导子外源处理棉花叶片和桔梗悬浮培养细胞系，探讨了外源诱导子对植物次生代谢物的诱导生成的效果和机理，结果表明：诱导子外源处理植物生体和植物悬浮培养细胞系均有诱导新的活性物质产生或诱导原有活性成分增加的作用。这一研究成果对相关植物次生产物(如农药活性物质和医药活性物质)的代谢机理及其利用研究具有重要的参考价值。
1、以棉花叶片为对象，采用了病原菌直接侵染和生物诱导子处理两种方法。选取已经受到棉黄萎菌侵染的棉花叶片的病斑作为实验材料，对其化学成分进行分析，并与对照(物感染的健康棉花叶片)作比较，发现：在棉黄萎菌侵染的叶片中生成了多种植保素，其中可确定的一种植保素的分子量为280。选取健康的棉花叶片其作为实验材料，用人工合成寡糖(六糖)制备的诱导子处理离体棉花叶片，对其化学成分进行分析，发现在人工合成寡糖处理的棉花叶片中生成了多种植保素，其中可确定的有分子量为354、268、282、111和156的5种植保素。因此，不论用病原菌直接侵染还是用生物诱导子外源处理植物生体，都能有效诱导植保素的生成。研究结果可以作为棉花病虫害生物防治的理论依据。人工合成寡糖的诱导活性也进一步得到验证，为其在生物防治上的应用提供了理论依据。
2、以桔梗悬浮培养细胞系为对象，采用了来源于不同真菌的生物诱导子处理的方法。用三种霉菌菌株黑曲霉、刺囊毛酶及少根根酶制备真菌源多糖诱导子，并用同种浓度的三种诱导子分别处理桔梗悬浮培养细胞系，发现三种来源的诱导子均能不同程度的增加桔梗细胞的生物量和诱导桔梗皂甙的生成。当在桔梗悬浮培养细胞接种当天即加入诱导子处理时，发现用黑曲酶和刺囊毛霉制备的诱导子处理的桔梗细胞的生物量和总皂甙含量均有大幅度提高，其中用刺囊毛霉制备的诱导子处理的桔梗细胞的总皂甙含量接种后的第16天可达到对照的1.5倍。当在桔梗悬浮培养细胞接种后第9天加入诱导子处理时，发现用黑曲酶和刺囊毛霉制备的诱导子处理的桔梗细胞的生物量和总皂甙含量也能大幅度提高，其中用黑曲酶制备的诱导子处理的桔梗细胞的总皂甙含量在接种后的第20天可达到对照的1.6倍。但用少根根酶制备的诱导子处理桔梗悬浮培养细胞，却对桔梗细胞的生物量和总皂甙含量并没有明显变化。因此，用真菌诱导子作为提高植物悬浮培养细胞次生代谢物的产量的一种手段是很有效的，但需要大量的筛选。
3、本文还分析了诱导子的作用效果及其影响因素，并阐述了诱导子作用的机理及其应用前景。诱导子的作用效果与诱导子的种类、诱导活性成分的浓度、诱导处理的时间长短、诱导子加入的时期及诱导处理的系统种类等多种因素密切相关。诱导子引起植物代谢过程的一系列变化，最终使植物产生防卫反应，是一个连续的、信号逐级传导的过程，其作用机理十分复杂，其理论研究和应用研究均对生物学和医学具有重要意义。
In this thesis, kinds of elicitors were applied to cotton leaves and suspensive cultured cells of Platycodon grandiflorum., the effect and mechanism of the induction of plant secondary metabolites by elicitors were explored. The result was elicitor can induce new active composition in the plant tissue and increase the content of the active composition in suspensive cultured plant cells. This result is a reference data for studies on metabolic mechanism and utility of plant secondary metabolites.
1, As to cotton leaves, two method were applied: infected by pathogen directly and treated with biotic elicitors. Analysizing the chemical composition of the cotton leaves infected by Verticillium dahliae and treated by artificially synthesized oligosaccharide, many phytoalexins were found induced in both leaves. In tne leaves infected by Verticillium dahliae phytoalexin MW 280 was induced. In the leaves treated by oligosaccharide, five phytoalexins MW 354, 268, 282, 11 land 156 were induced. Therefore, infected by pathogen directly and treated with biotic elicitors both can induce the phytoalexins effectively. The result can be a theoretical foundation. The action of the artificially synthesized oligosaccharide was varified furtherly.
2, As to suspensive cultured cells of Platycodon grandiflorum., treated with biotic elicitors from different fungus. Treated suspensive cultured cells of Platycodon grandiflorum. with elicitors prepared from Aspergillus nige, Mucor spinosus and Rhizopus arrhizus in the same consistency, cell fresh weight and total saponins content were increased in varying degrees. The total saponin content can be 1.5 times of the contract in the cells treated by Mucor spinosus on 0 day after inoculated for 16 days. The total saponin content can be 1.6 times of contractin the cells treated by Aspergillus nige on 9 day after inoculated for 20 days. Therefore, increasing the content of the secondary metabolites in suspensive cultured plant cells by treated with fungal elicitors is a effective method.
3, This thesis also analysized the effect and influent factors and clarified the inducing mechanism and utility prospect of elicitor. The effect of elicitor was related to many factors such as the type of elicitor, the consistency of elicitor, time of the system treated by elicitor, time of the elicitor applied and the type of the system treated by elicitor. The changes of plant metabolism induced by elicitor induced defensive actions in plant. It is a successive signal transduction process. The mechanism is very difficult, but theoretical and practical studies on it are both important to biology and medical science.
1、以棉花叶片为对象，采用了病原菌直接侵染和生物诱导子处理两种方法。选取已经受到棉黄萎菌侵染的棉花叶片的病斑作为实验材料，对其化学成分进行分析，并与对照(物感染的健康棉花叶片)作比较，发现：在棉黄萎菌侵染的叶片中生成了多种植保素，其中可确定的一种植保素的分子量为280。选取健康的棉花叶片其作为实验材料，用人工合成寡糖(六糖)制备的诱导子处理离体棉花叶片，对其化学成分进行分析，发现在人工合成寡糖处理的棉花叶片中生成了多种植保素，其中可确定的有分子量为354、268、282、111和156的5种植保素。因此，不论用病原菌直接侵染还是用生物诱导子外源处理植物生体，都能有效诱导植保素的生成。研究结果可以作为棉花病虫害生物防治的理论依据。人工合成寡糖的诱导活性也进一步得到验证，为其在生物防治上的应用提供了理论依据。
2、以桔梗悬浮培养细胞系为对象，采用了来源于不同真菌的生物诱导子处理的方法。用三种霉菌菌株黑曲霉、刺囊毛酶及少根根酶制备真菌源多糖诱导子，并用同种浓度的三种诱导子分别处理桔梗悬浮培养细胞系，发现三种来源的诱导子均能不同程度的增加桔梗细胞的生物量和诱导桔梗皂甙的生成。当在桔梗悬浮培养细胞接种当天即加入诱导子处理时，发现用黑曲酶和刺囊毛霉制备的诱导子处理的桔梗细胞的生物量和总皂甙含量均有大幅度提高，其中用刺囊毛霉制备的诱导子处理的桔梗细胞的总皂甙含量接种后的第16天可达到对照的1.5倍。当在桔梗悬浮培养细胞接种后第9天加入诱导子处理时，发现用黑曲酶和刺囊毛霉制备的诱导子处理的桔梗细胞的生物量和总皂甙含量也能大幅度提高，其中用黑曲酶制备的诱导子处理的桔梗细胞的总皂甙含量在接种后的第20天可达到对照的1.6倍。但用少根根酶制备的诱导子处理桔梗悬浮培养细胞，却对桔梗细胞的生物量和总皂甙含量并没有明显变化。因此，用真菌诱导子作为提高植物悬浮培养细胞次生代谢物的产量的一种手段是很有效的，但需要大量的筛选。
3、本文还分析了诱导子的作用效果及其影响因素，并阐述了诱导子作用的机理及其应用前景。诱导子的作用效果与诱导子的种类、诱导活性成分的浓度、诱导处理的时间长短、诱导子加入的时期及诱导处理的系统种类等多种因素密切相关。诱导子引起植物代谢过程的一系列变化，最终使植物产生防卫反应，是一个连续的、信号逐级传导的过程，其作用机理十分复杂，其理论研究和应用研究均对生物学和医学具有重要意义。
In this thesis, kinds of elicitors were applied to cotton leaves and suspensive cultured cells of Platycodon grandiflorum., the effect and mechanism of the induction of plant secondary metabolites by elicitors were explored. The result was elicitor can induce new active composition in the plant tissue and increase the content of the active composition in suspensive cultured plant cells. This result is a reference data for studies on metabolic mechanism and utility of plant secondary metabolites.
1, As to cotton leaves, two method were applied: infected by pathogen directly and treated with biotic elicitors. Analysizing the chemical composition of the cotton leaves infected by Verticillium dahliae and treated by artificially synthesized oligosaccharide, many phytoalexins were found induced in both leaves. In tne leaves infected by Verticillium dahliae phytoalexin MW 280 was induced. In the leaves treated by oligosaccharide, five phytoalexins MW 354, 268, 282, 11 land 156 were induced. Therefore, infected by pathogen directly and treated with biotic elicitors both can induce the phytoalexins effectively. The result can be a theoretical foundation. The action of the artificially synthesized oligosaccharide was varified furtherly.
2, As to suspensive cultured cells of Platycodon grandiflorum., treated with biotic elicitors from different fungus. Treated suspensive cultured cells of Platycodon grandiflorum. with elicitors prepared from Aspergillus nige, Mucor spinosus and Rhizopus arrhizus in the same consistency, cell fresh weight and total saponins content were increased in varying degrees. The total saponin content can be 1.5 times of the contract in the cells treated by Mucor spinosus on 0 day after inoculated for 16 days. The total saponin content can be 1.6 times of contractin the cells treated by Aspergillus nige on 9 day after inoculated for 20 days. Therefore, increasing the content of the secondary metabolites in suspensive cultured plant cells by treated with fungal elicitors is a effective method.
3, This thesis also analysized the effect and influent factors and clarified the inducing mechanism and utility prospect of elicitor. The effect of elicitor was related to many factors such as the type of elicitor, the consistency of elicitor, time of the system treated by elicitor, time of the elicitor applied and the type of the system treated by elicitor. The changes of plant metabolism induced by elicitor induced defensive actions in plant. It is a successive signal transduction process. The mechanism is very difficult, but theoretical and practical studies on it are both important to biology and medical science.
引文
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1．赵俊云，李文新，盛桂莲。棉黄萎菌侵染诱导 PA 的分离及特性。华中师范大学学报(自然科学版)2001，6
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1．赵俊云，李文新，盛桂莲。棉黄萎菌侵染诱导 PA 的分离及特性。华中师范大学学报(自然科学版)2001，6
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