Effect Of Selenium-sulfur Interaction On The Biosynthesis Of Sulforaphane In Broccoli

Broccoli(Brassica oleracea L.var.italica Plench)originated from the eastern coast of the Mediterranean,which has the characteristics of strong adaptability,was widely planted in China.Sulforaphane,as an important secondary metabolite in broccoli,which has a strong effect of promoting the detoxification of carcinogens and inhibiting the growth of cancer cells.Selenium and sulfur are elements of the sixth main group,their related metabolic process are similar in plants,and the metabolic pathways have competitive synergistic relationship.It has important reference significance for high quality broccoli breeding to utilize selenium-rich characteristics of broccoli,to study the regulation mechanism of selenium-sulfur interaction on sulfur-containing secondary metabolite sulforaphane anabolism in broccoli.In this study,two sulphur levels(1 mM,4 m M)and four selenium levels(0,50,100,150μM)were used to treat with broccoli in budding stage.To study the characteristics of related genes expression and related metabolites change in sulforaphane synthesis pathway of broccoli by selenium-sulfur interaction.The specific results are as follows:1.The total selenium and total sulphur content in broccoli florets and leaves were detected by selenium and sulphur combined treatment.It was found that the absorption of sulphur has tissue difference,in broccoli leaves,under low sulphur treatment,low selenium(50 mM)dose could promote the absorption of sulphur element,thus no significant effect on sulfur absorption with selenium application shown under high sulphur treatment;exogenous selenium had a slight inhibition effect on the absorption of sulphur element in broccoli florets.The selenium content in leaves and florets increased linearly with the concentration of exogenous selenium application.2.The content of methionine and Se-methylselenocysteine were detected.It was found that the content of methionine decreased with the increase of exogenous selenium dosage under low sulfur treatment,but had little effect under high sulfur treatment.Exogenous selenium enhanced the content of Se-methylselenocysteine under low sulfur treatment,and the content increased first and then decreased under high sulfur treatment,which reached the highest level at 100 μM selenium treatment.3.The content of glucosinolates was determined and it was found that under low-sulfur treatment,50μM and 100μM selenium treatment had little effect on glucoraphanin,and 150μM selenium treatment significantly inhibited the accumulation of glucoraphanin,but no significant effect exhibited under high-sulfur treatment;under low sulfur treatment,selenium application significantly inhibited the synthesis of indolic and aromatic glucosinolates,while under high sulfur treatment,exogenous selenium promoted the accumulation of aromatic glucosinolates;increased the concentration of sulphur can effectively alleviate the inhibition of selenium on the synthesis of sulphur-containing compounds-glucosinolates;the content of glucoraphanin was positively correlated with that of methionine(r=0.498*),suggested that exogenous selenium treatment could affect the synthesis of amino acid,the precursor of glucosinolate,that are involved in the synthesis of glucosinolates.4.The analysis results of myrosinase activity showed that myrosinase activity first increased and then decreased under exogenous selenium treatment,and reached the highest value under exogenous selenium treatment of 100 μM both high sulfur and low sulfur treatment,when compared with the blank group that the myrosinase activity increased significantly after selenium treatment.Also the content of sulforaphane was detected which find that sulforaphane content increased first and then decreased under exogenous selenium treatment,and reached the peak value of sulforaphane content under different sulfur application conditions when treated with 100 μM of selenium,and sulforaphane accumulation was the highest value under the combination of 4mM sulfur and 100 μM selenium treatment(5.3 μmol/g).Significant positive correlation(r=0.453*)was shown which between the activity of myrosinase and the content of sulforaphane,suggested that selenium treatment mainly affected the activity of glucoraphanin hydrolase-myrosinase,thus affecting the synthesis of sulforaphane.5.The best combination of selenium and sulphur(Se:100μM;S:4mM)was selected to treat,analyzed the spatiotemporal characteristics of sulforaphane accumulation and the expression characteristics of genes related to sulforaphane synthesis in broccoli.It was found that exogenous selenium could induce stress on related genes and inhibit their expression,with the prolongation of treatment time that the inhibiting action gradually weakened,which performed temporal and spatial characteristics;however the expression of related genes was up-regulated in broccoli root;the accumulation of sulforaphane in florets increased afterexogenous selenium treatment,and the increased reached the maximum value at 12 h,but the expression of MY gene was significantly increased at 6 h,indicated that there was a temporal and spatial difference between the accumulation of sulforaphane in broccoli and the expression of related genes.