Effects Of Exogenous Selenium On Nutritional Quality And Flavonoid Accumulation In Wheat Grain(triticum Aestivum L.)

Selenium is one of essential trace elements for human body and has multiple functions such as anti-cancer,preventing cancer,improving immunity and eliminating harmful free radicals in human body.Common wheat(Triticum aestivum L.)is one of the most widely cultivated staple crops in the world.Selenium biofortification in wheat is an effective way to improve human selenium intake.However,the cultivation techniques for selenium biofortification of wheat under specific environmental and soil conditions need to be standardized,the optimal application stage and dosage of selenium fertilizer in the process of production practice are not clear,and there are few reports on the effects of selenium fertilizer on nutritional quality of wheat grains.In this study,different wheat varieties were treated with different amounts of selenium fertilizer at different stages under field cultivation conditions.The objects are to explore the optimal application stage and dosage of selenium fertilizer and to investigate the effects of selenium fertilizer on agronomic characters and nutritional quality of wheat.Then,the effect of exogenous selenium on the types and contents of secondary metabolites in wheat grains was studied by metabolite profiling analysis.Integrated analysis of metabolome and transcriptome was carried out to reveal the molecular mechanism by which exogenous selenium caused changes of secondary metabolites in wheat grains.The main conclusions of this study are as follows:(1)The results of analysis of variance showed that application stage and amount of selenium fertilizer had highly significant effect on the selenium content of common wheat grains,and the differences in selenium contents were highly significant between wheat varieties.In general,foliar spray of selenium fertilizer significantly increased the selenium contents of mature wheat grains.When the application amount of selenium was 40 g·hm-2,selenium was effectively enriched in the grains of the tested wheat varieties and did not exceed the safety limit in our country.Under the same amount of supplemented selenium fertilizer,the selenium contents of wheat grains were higher with foliar spray at filling stage than those at heading stage.The concentrations of chromium,arsenic,lead and cadmium in wheat grains decreased significantly after selenium application,and there were significant differences between wheat varieties.(2)Different amount of selenium fertilizer had no significant effect on the numbers of panicles and grains per panicle of black wheat(Jizi496 and Linhei145),however,the1000-grain weight increased significantly and grain yield also increased in varying degrees.After foliar application of selenium fertilizer,the selenium contents and selenium accumulation amount of black wheat grains increased significantly.The selenium contents in grains increased with increase in application amount of selenium,and the differences between treatments were significant.The utilization efficiency of selenium for Linhei145 was significantly higher than that of Jizi496.In this study,the suitable application amount of selenium for Jizi496 was 30 g·hm-2 and 20 g·hm-2 for Linhei145.Selenium in black wheat grains mainly existed in the form of selenomethionine.The proportion of selenomethionine in total selenium in grains increased after applying selenium fertilizer.The contents of chromium,arsenic,lead and cadmium in black wheat grains decreased in varying degrees after foliar spray.The results of element correlation analysis showed that selenium was negatively correlated with chromium,arsenic,lead and cadmium.The anthocyanin content of black wheat grains increased significantly after foliar spray,and there was a positive correlation between the selenium content and the anthocyanin content of grains.(3)The results of two-year yield investigation showed that the grain yield of Tai113,Zhongmai175 and Changzhi451 increased in varying degrees after foliar spray,and the differences were significant between wheat varieties.Selenium treatment had no significant effect on the numbers of panicles and grains per panicle,however,the 1000-grain weight increased significantly,and the 1000-grain weight of Tai113 increased significantly.The length of the flag leaf increased significantly after selenium treatment,and the increase was more obvious when foliar-spraying at filling stage.The dry matter mass and the chlorophyll content increased slightly after selenium application at filling stage.The plant height of Zhongmai175 increased significantly after selenium application,and the increase was more obvious when foliar-spraying at filling stage.The width of the flag leaf for Changzhi451 increased significantly after selenium application at filling stage.The total selenium content and selenium accumulation amount in grains increased significantly after foliar application of selenium,and most of selenium existed in the organic form.The selenium utilization efficiency for application at filling stage was significantly higher than that at heading stage,and the selenium utilization efficiency of Tai113 was significantly higher than those of the other two varieties.After foliar application,the contents of essential amino acids in wheat grains increased.The effects of foliar selenium on the contents of total starch,amylopectin,amylose,soluble sugar and sucrose in wheat grains depended on wheat varieties.However,the anthocyanin contents in wheat grains of three varieties were significantly increased,and the increase was more obvious when foliar-spraying at filling stage.(4)After selenium treatment,the albumin content of wheat grains increased significantly and the proportion of essential amino acids increased.The contents of total anthocyanins and total flavonoids in selenium-enriched wheat grains were significantly higher than those in the control.Metabolite profiling analysis showed that 611 metabolites were identified in wheat grains with selenium treatment and the control,including 59 significantly changed metabolites(57 up-regulated and 2 down-regulated),of which 27 significantly changed metabolites belonged to flavonoids and were significantly up-regulated in selenium-enriched wheat grains.Selenium application contributed to the enrichment of flavonoids(tricin,dihydroquercetin,anthocyanins and catechins)in wheat grains.Significantly changed metabolites were mainly enriched in the pathways of flavonoid biosynthesis and flavone and flavonol biosynthesis.In addition,glycetin,an isoflavone was firstly detected in wheat grains.(5)A total of 1733 differentially expressed genes were identified by transcriptome sequencing,of which 1193 genes were up-regulated and 540 genes were down regulated.KEGG pathway enrichment analysis showed that the differentially expressed genes were mainly enriched in the flavonoid biosynthesis pathway,and other enrichment pathways included flavone and flavonol biosynthesis,glutathione metabolism and sulfur metabolism.The differentially expressed genes enriched in the above pathways were significantly up-regulated.In addition,a total of 182 differentially expressed transcription factor genes were identified,most of which were significantly up-regulated.These differentially expressed genes might mediate selenium-induced flavonoid biosynthesis.The transcriptome sequencing results were then verified by quantitative real-time PCR.(6)Integrated analysis of metabolite profiling and transcriptome sequencing showed that the pathways of flavonoid biosynthesis,flavone and flavonol biosynthesis were closely related to selenium.Exogenous selenium enhanced the biosynthesis of flavonoids(tricin,dihydroquercetin,anthocyanins,chrysoeriol and catechins)in wheat grains by increasing the expression levels of seven key genes in flavonoid biosynthesis(2 F3 Hs,2 DFRs,1 F3’5’H,1OMT and 1 ANR).The present study revealed the molecular mechanism by which exogenous selenium enhanced the biosynthetic pathway of tricin,anthocyanins and catechins in wheat grains.In conclusion,this study explored the effects of exogenous selenium on selenium enrichment and nutritional quality of wheat grains,and determined the optimal application stage and amount of selenium fertilizer.Meanwhile,this study revealed the changes of secondary metabolites in selenium-enriched wheat grains,and proposed the possible molecular mechanism by which exogenous selenium enhanced flavonoid accumulation in wheat grains.The present study confirms that selenium biofortification is an effective way to improve functional nutrients such as selenoproteins and flavonoids in wheat grains,and lays a theoretical foundation for further development of selenium-enriched wheat health products.