Effects Of Humic Acids On Volatile Oil And Flavonoids In Lemon Leaves And Their Mechanisms

Humic acid is a kind of weak organic acids widely distributed in nature resource.According to the relative molecular weight and solubility,humic acid can be divided into fulvic acid,brown fulvic acid and black fulvic acid.As a natural organic substance,humic acid has been widely used in the field of agriculture.Lemon is the fruit with medicinal and edible values,which is rich in volatile oils,flavonoids and other bio-active substances.In this study,the effects of humic acids,such as sodium humate and fulvic acid,on volatile oils and flavonoids in lemon leaves and their mechanisms were systematically investigated,in order to provide a research basis for the mechanism of humic acids and the improvement of lemon quality.The main works are as follows:(1)Sodium humate and fulvic acid were extracted from lignite located in Deyang of Sichuan Province by alkali extraction method.Their contents of moisture,ash,total humic acids,free humic acids,water-soluble humic acids,fulvic acids and p H value were determined.(2)The volatile oils were extracted from lemon leave samples treated with different fertilizing methods(irrigation or foliar-spray)during 11 harvest periods by steam distillation,respectively.The comparison of the volatile oil contents in these samples indicated that 5g/L fulvic acid and 5g/L sodium humate could both increase the contents of volatile oils in lemon leaves at each collection point,and this positive effect of fulvic acid was better than that of sodium humate;In irrigated groups,the optimal concentration of fulvic acid was 5g/L and sodium humate was 1g/L;the positive effects in foliar-sprayed groups were better than that of irrigated groups,but the effects in foliar-sprayed groups showed some seasonal difference.(3)The contents of total flavonoids in lemon leave samples treated with different fertilizing methods(irrigation or foliar-spray)during 6 harvest periods were determined by UV spectrophotometry.The comparison of the total flavonoid contents in these samples indicated that fulvic acid and sodium humate could both increase the content of total flavonoids in lemon leaves at each collection point,but these positive effects showed the seasonal difference;in irrigated groups,the positive effect of 5g/L fulvic acid was better than that of 5g/L sodium humate,but in foliar-sprayed groups,that effect of 5g/L sodium humate was better than that of 5g/L fulvic acid;in irrigated groups,the optimal concentration of fulvic acid was 5g/L and that of sodium humate was 5g/L;the positive effects in foliar-sprayed groups were better than that of irrigated groups,but the effects in foliar-sprayed groups showed some seasonal difference.(4)The action mechanisms of sodium humate and fulvic acid were preliminarily discussed by transcriptome sequencing,combined with the metabonomic analysis.The comparison of transcriptome sequencing analysis between sodium humate or fulvic acid treated groups and the corresponding blank groups indicated that 765 differentially expressed genes(DEGs)were found between 5g/L fulvic acid treatment group and blank groups,of which 211 were annotated by using KEGG database,and they mainly play some parts in plant hormone signal transduction,starch and sucrose metabolism and amino acid biosynthesis,etc.Further,71 DEGs were found between10 g/L sodium humate treatment group and blank groups,of which 35 were annotated by using KEGG database,and they mainly play some parts in plant hormone signal transduction,phagosome,ubiquitin mediated proteolysis,etc.Additionally,307 DEGs were found between sodium humate leaf spray group and blank groups,of which 106 were annotatedby using KEGG database,and they mainly play some parts in amino acid biosynthesis,cysteine and methionine metabolism,phenylpropane biosynthesis,etc.Combined analysis of these DEGs and differential metabolites suggested that the influence of humic acids on the contents of terpenoids and flavonoids in lemon leaves was related to the regulation of humic acids on the key enzymes or intermediate metabolites in their bio-synthetic pathways,such as GGPP,PAL and 4CL.