Alleviative Effects And Mechanism Of Exogenous Ascorbic Acid On Chromium(Cr⁶⁺) Toxicity In Wheat

In recent years,due to the rapid development of tannery,iron-making,dyeing and other industries,chromium pollution of waste water has become increasingly prominent.Wheat is one of the most important food crops of mankind,and its quality is well received.In this study,Yangmai 16(Cr6+-tolerance)and Yumai 51(Cr6+-sensitivity)were used as materials to analyze the effects of Cr6+ toxicity on transporting,antioxidant systems,photosynthetic characteristics of genotypic differences were investigated.Besides,the alleviation effect and physiological mechanism of ascorbic acid(AsA)as exogenous remission substances on Cr6+ toxicity were investigated.The main study were as follows:Under Cr6+ Stress,wheat dry weight,total root length,root surface area were significantly inhibited,while Yangmai 16 depressed less than Yumai 51;The Cr content in root was significantly higher than that in shoots.In addition,Cr is mainly distributed in the root cell walls;The contents of K and Fe in wheat roots decreased significantly,while the contents of Ca and Zn were significantly increased,and the contents of P,Ca and Mg in the shoots were significantly increased,but the Fe content decreased significantly.Simultaneous addition of 500 ?mol.L-1AsA treatment,wheat growth was improved,especially in dry weight,total root length and root surface area;Cr transport rate decreased significantly,indicating that exogenous AsA mainly through inhibiting Cr transport to shoot to reduce the Cr poisoning effect on shoot.Moreover,exogenous AsA treated can promote the roots absorb K,Fe and Mn from the nutrient solution,and increase the accumulation of Mn on the shoot.The effects of exogenous AsA treatment on the metabolism of active oxygen in roots and leaves of two cultivars underCr6+ stress were studied by hydroponics experiment.The results showed that the contents of O2·-,H2O2 and MDA in the roots and leaves of the two cultivars under 100?mol.L-1Cr6+ were significantly higher than control treatments,among them,the O2·-,H2O2 and MDA accumulated in Yumai 51 were higher than those of Yangmai 16,which indicated that Yumai 51 were more severely affected by Cr6 + than Yangmai 16;The total antioxidant capacity of roots and leaves decreased,The contents of GSH and AsA,the ratio of AsA/DHA and GSH/GSSG in roots and leaves were decreased,especially in the leaves decreased more seriously;The activities of APX,MDHAR,DHAR and GS in the roots and leaves of AsA-GSH decreased,which indicated that the AsA-GSH cycle in wheat roots and leaves was inhibited.Under Cr6+ treatment,The addition of ascorbic acid synthesis inhibitor(Lycorine Hydrochloride)treatment will further inhibit the activity of APX,MDHAR,DHAR,GS and AsA and GSH in AsA-GsH cycle.The activities of SOD,POD and CAT were increased,the contents of O2·-,H2O2 and MDA were significantly decreased,and the contents of AsA and GSH contents and APX,MDHAR,DHAR,GS enzyme activity in root and leaf were significantly increased by adding 500 m mol.L-lAsA under Cr6+ treatmnet.When the ascorbic acid direct premise substance(L-galactose-1,4-lactone)was added under Cr6+ treatment,the activity of APX,MDHAR,DHAR,GS increased in AsA-GSH cycle,and the content of AsA and GSH increased too.These results indicate that AsA plays an important role in remove of reactive oxygen species under Cr6+ stress.Leaf stomatal conductance(Gs),net photosynthetic rate(Pn),maximum photochemical efficiency(Fv/Fm),actual photochemical efficiency(?PSII),photochemical quenching coefficient(qP)decreaseing,the intercellular carbon dioxide concentration(Ci)decreased firstly and then increased with extended Cr6+ exposure time,which suggested that the net photosynthetic rate of wheat leaves was affected by stomatal and non-stomatal factors,and the photosynthetic system was inhibited obviously,the inhibition of these parameters were more pronounced in Yumai 51 than in Yangmai 16.Cr6+ stress caused the decrease of chlorophyll a and b in leaves of two cultivars,chloroplast photosynthetic phosphorylation levels decreased,ATP synthesis blocked,the activity of Ca2+-ATPase and Mg2+-ATPase was decreased,plasma membrane Ca2+andMg2+ transport disorder,eventually hindering photosynthesis.The efficiency of deoxidized epoxidation of wheat leaves and the energy dissipated by the xanthophyll cycle accounts for the amount of energy dissipated in the form of heat(qE/NPQ)decreased;The activities of Gs,Pn,Fv/Fm,?PSII,qP and qE/NPQ,Ca2+-ATPase and Mg2+-ATPase were increased by the addition of exogenous AsA treatment under Cr6+ stress,and the activity of photosynthetic phosphorylation Level increased,ATP content increased.These changes indicate that exogenous AsA is more effective for the action of xanthophyll loops under Cr6+ stress,timely dissipating energy that can not be used for photosynthetic electron transport,avoid the occurrence of photoinhibition,which play an important role in restoring photosynthesis.The effects of exogenous ascorbic acid(AsA)on the pH value of wheat rhizosphere,H+-ATPase activity in root plasma membrane,NH4+ and NO3-ion absorption,the species and content of organic acids secreted by roots,scavenging effect of secreted organic acids on DPPH free radicals and chelating effect on Cr6+.The results showed that the PH value of rhizosphere of two wheat cultivars increased gradually under 100 umol.L-1 Cr6+ treatment,to the 10th day exposed to Cr6+,the treatment group was significantly higher than that of control,And the increase of PH value in Yumai 51 was higher than that of Yangmai 16,which indicated that the anion-cation absorption imbalance caused by Cr6+ stree was more seriously.The H+-ATPase activity of the apical plasma membrane was significantly decreased,which indicated that Cr6+ stress can inhibit H+-ATPase activity,hinder the intracellular H+ pump out to the extracellular process,eventually leading to rhizosphere PH value increased.Under Cr6+ stress,the content of organic acids secreted by wheat roots was significantly increased,among the five organic acids,oxalic acid and citric acid are the main organic acids secreted by wheat roots.Adding exogenous AsA treatment,the increase of PH value in rhizosphere of two cultivars was significantly smaller than that of C6+treatment,to the 10th day treatment,the pH value of Yangmai 16 was close to the control level.Exogenous AsA alleviation treatment can enhance the H+-ATPase activity in root tip membrane and the uptake rate of NH4+-N in the environment under Cr6+ stress.Under different pH conditions,the reduction rate of Cr6+ by organic acids that secreted by roots was enhanced with the decrease of pH value and the prolongation of treatment time.In addition,exogenous AsA treatment,the wheat root mainly secrete citric acid,tartaric acid.Organic acids can complex Cr6+ in the environment and convert Cr6+ into a chelate that is less toxic to plants.The organic acids secreted by roots also have significant scavenging effects on DPPH ' free radicals In summary,the exogenous AsA alleviation treatment can increase the H+-ATPase activity of wheat root membrane and enhance the absorption rate of NH4+-N,maintain the acidic environment required for the normal growth of wheat,roots secreted more citric acid and tartaric acid and other organic acids,these secreted organic acids can reduce Cr6+ in acidic environments,chelating Cr6+ and participate in the scavenging of free radicals,these changes play an important role in alleviating the toxicity of Cr6+ stressThe effects of exogenous AsA on root elongation,root lignin content,cell wall polysaccharide content,pectin methylesterase and POD activity,APX activity and AsA content in apoplast and symplasm of wheat root under Cr6+ stress were studied.The results showed that the root elongation of two wheat cultivars was significantly inhibited under Cr6+ stress,and the inhibition degree of Yumai 51 was more serious than that of Yumai 51.The contents of pectin,cellulose and hemicellulose in the root cell wall increased,and the activities of pectin methylesterase and POD were significantly increased,and the activity of APX and the content of AsA in apoplast and symplasm were increased.With the addition of exogenous AsA,the root elongation of the two cultivars was significantly recover;Root lignin content decreased significantly,between them,Yangmai 16 is close to the control level;The contents of pectin,cellulose and hemicellulose in the cell wall of the root were higher than those of Cr6+ independent treatment,the content of hemicellulose and pectin in the cell wall of Yangmai 16 increased significantly(P<0.05).The activity of pectin methylesterase in the cell wall was significantly increased and the activity of POD was decreased significantly.the activity of APX in apoplast and the content of AsA in apoplast and symplasm were significantly higher than those treated with Cr6+ alone(P<0.05).In addition,the addition of exogenous AsA treatment could significantly increase AsA content and APX activity in apoplast of wheat root under Cr6+ stress.APX can remove H2O2 from AsA as substrate,and avoid the accumulation of H2O2 and recovery POD activity,timely removal of lignin synthesis precursor material,ultimately reduced the synthesis of root lignin,reducing the Cr6+ stress caused by root lignification on root elongation growth inhibition.On the other hand,exogenous AsA can promote the accumulation of polysaccharides such as pectin,the increase of pectin methylesterase activity increases the number of free carboxyl groups on pectin,which enhances the fixation ability of root cell wall to Cr,Increased the effect of cell wall area isolation,avoid more of the Cr into the protoplasts,reducing the root cells by Cr toxicity.