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黄芪种子脂肪酸在幼苗形态建成中的代谢研究

The Study on Metabolism of Fatty Acids of Astragalus Membranaceus Seeds during Seedling Morphology

【作者】 杨楠

【导师】 郭晓瑞;

【作者基本信息】 东北林业大学 , 植物学, 2019, 硕士

【摘要】 黄苗(Astragalus membranaceus)是一种传统中药材,其药用历史已有2000多年。黄芪从种子萌发到幼苗形态建成的过程是其完成从种子供养到光合作用供养转变的过程,也是其生活史中面对环境变化表现较为脆弱的阶段。本文以黄芪种子为材料,利用GC-MS研究了种子萌发以及形态建成过程中包括脂肪酸在内的贮藏物质代谢变化,同时利用UPLC-MS还研究了形态建成过程中黄芪幼苗黄酮和皂苷等活性物质含量变化,以期为深入了解黄芪种子萌发和形态建成的养分代谢规律提供参考。得到如下主要结果:在黄芪种子萌发及后生长过程,发现子叶贮藏的淀粉被优先利用,而脂质则主要在萌发后生长阶段被动员。脂肪酸组分分析结果显示,子叶中主要饱和及单不饱和脂肪酸所占比例降低,多不饱和脂肪酸则与它们相反,但总体变化幅度都较小。外源脱落酸和茉莉酸甲酯处理均抑制了黄芪种子萌发及后生长。同时,脱落酸和茉莉酸甲酯抑制了贮藏物质动员,降低了其利用效率,还引发了早期贮藏物质碳的高消耗,阻止了子叶脂肪酸的去饱和及碳链延长。在黄芪幼苗形态建成过程,发现子叶脂肪酸组分发生了较大幅度的变化。与暗形态建成过程相比,光照条件下子叶中α-C18:3比例增加了 28.62%,而C18:1则降低了30.96%。同时,子叶脂肪酸去饱和酶基因FAD7和FAD2的转录水平在光形态建成过程中均有明显上调,表明它们可能参与了 α-C18:3和C18:1比例转变的过程。在细胞器膜脂组分研究中,与暗形态建成相比,光照诱导了质体膜C16:1、α-C18:3、线粒体膜C22:0、C24:0、C16:0、α-C18:3以及过氧化物酶体膜C16:0、C18:0的大幅度增加。同时,相关性分析结果显示,质体和线粒体中α-C18:3与子叶α-C18:3表现出显著正相关,表明光照诱导的子叶α-C18:3增加可能与质体和线粒体中α-C18:3增加密切相关。与子叶不同,新生组织中α-C18:3、C18:2和C18:1三种主要不饱和脂肪酸比例在形态建成过程中逐渐下降,而C16:0和C18:0比例则持续升高,并且光照条件下α-C18:3的下降幅度以及C16:0和C18:0的升高幅度均小于黑暗下。此外,在黄芪幼苗形态建成过程中,光照均诱导了异黄酮和皂苷的积累,并且其主要是由新生组织中芒柄花黄素和黄芪甲苷的含量变化引起的。与暗形态建成相比,子叶和新生组织中H2O2含量和POD活性在光形态建成中明显增加。异黄酮和皂苷等次生代谢产物的积累变化可能通过调控组织细胞氧化还原水平参与了黄芪幼苗光形态建成中的脂肪酸氧化还原代谢过程。

【Abstract】 Astragalus membranaceus(A.membranaceus),a traditional Chinese medicinal plant,has more than 2000 years medicinal history.The process of A.membranaceus from seed germination to seedling morphogenesis is the process of its transformation from seed feeding to photosynthesis support,and it is also a relatively fragile stage in its life history in the face of environmental changes.In this paper,the seeds of A.membranaceus were used as material to study the metabolic changes of storage reserves including fatty acids during seed germination and morphogenesis.At the same time,the changes of active substances such as flavonoids and saponins of A.membranaceus seedlings during morphogenesis were also studied.It is expected to provide a reference for further understanding of seed storage reserves metabolism of A.membranaceus on seed germination and morphogenesis.The main results are as follows:During the seed germination and post-germination growth of A.membranaceus,it was found that the starch stored in the cotyledons was preferentially utilized,while the lipid was mainly mobilized during the post-germination growth.The analysis of fatty acid compositions showed that the proportion of the main saturated and monounsaturated fatty acids in the cotyledons was reduced,whereas the polyunsaturated fatty acids were the opposite,but the overall change was small.Exogenous abscisic acid and methyl jasmonate inhibited the germination and later growth of A.membranaceus seeds.At the same time,abscisic acid and methyl jasmonate inhibited the mobilization of storage reserves,reduced its utilization efficiency,led to high consumption of carbon in storage reserves at early stage,and prevented the desaturation of fatty acids in cotyledons and the elongation of carbon chain.In the process of morphogenesis of A.membranaceus seedlings,it was found that the fatty acid compositions in the cotyledons had undergone significant changes.Compared with skotomorphogenesis,the proportion of alpha-linolenic acid in cotyledons increased by 28.62%under light,while oleic acid decreased by 30.96%.At the same time,the transcriptional levels of fatty acid desaturase genes FAD2 and FAD7 in cotyledons were obviously up-regulated during photomorphogenesis,suggesting that they may be involved in the transformation of alpha-linolenic acid and oleic acid.In the study of organelle membrane lipid compositions,compared with skotomorphogenesis,light induced a large increase of palmitoleic acid and alpha-linolenic acid in the plastid membrane,docosanoic acid,tetracosanoic acid,palmitic acid and alpha-linolenic acid in mitochondrial membrane,and palmitic acid and stearic acid in the peroxisome membrane.At the same time,the correlation analysis showed that the alpha-linolenic acid in the plastids and mitochondrias was positively correlated with the alpha-linolenic acid in cotyledons,indicating that the increase of alpha-linolenic acid in cotyledons induced by light may be closely related to the increase of alpha-linolenic acid in the plastids and mitochondrias.Unlike the cotyledons,the proportions of the three main unsaturated fatty acids including alpha-linolenic acid,linoleic acid,and oleic acid in newly differentiated tissues gradually decreased during morphogenesis,while the proportions of palmitic acid and stearic acid continued to increase.Under light conditions,the decrease of alpha-linolenic acid and the increase of palmitic acid and stearic acid were less than those under dark conditions.In addition,the accumulation of isoflavones and saponins was induced by light in the morphogenesis of A.membranaceus seedlings,and it was mainly caused by the changes of the content of formononetin and astragaloside in the newly differentiated tissues,Compared with skotomorphogenesis,H2O2 content and POD activity in cotyledons and newly differentiated tissues increased significantly during photomorphogenesis.The accumulation of secondary metabolites such as isoflavones and saponins may be involved in the redox metabolism of fatty acids in the photomorphogenesis of A.membranaceus seedlings by regulating the redox levels of histiocytes.

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