Mechanism Of Copper Accumulation In Castor (Ricinus Communis L.) And Its Application On Remediation Of Copper Contaminated Soil

Phytoremediation technology is a measurement to purify pollutants in soil or water by the absorption,evaporation,root filtration,degradation or stabilization with plants.The key to this technology is to find a plant with large biomass or high concentration of metals in its tissues.Castor(Ricinus communis L.),found in the Tonglvshan,is a plant with a fast growth rate.It is strong tolerant to copper(Cu)in the soil,and it has resistance to poor soil due to its developed root system.Castor also has relatively high economic value.Castor seeds harvested from the Tonglvshan mine area were selected as the materials for this study.The hydroponic culture and pot experiment were conducted to analyze the composition of the root exudates from castor,the subcellular distribution of Cu in castor,the Cu contents and the Cu speciation in the castor tissues.Based on those results,the mechanism of the uptake,transportation,and accumulation of Cu in castor tissues was revealed primarily.Also,phytoremediation combined with the agronomic regulation measures such as the addition of phosphate fertilizer and chelating agents in pot experiment were carried out to evaluate their effectiveness to remediate the Cu contaminated soil.The research contents and relevant results are listed as follows:1.The results from the hydroponic culture and pot experiment showed that castor was tolerant to Cu and its roots accumulated the majority of Cu.Oxalic acid,succinic acid,tartaric acid,citric acid,and malic acid were the major organic acids in the root exudates from castor under Cu stress.Moreover,the acidification of root exudates on the rhizosphere soil increased the percentages of acid exchange Cu and reducible Cu,and the percentages of Cu forms decreased at the same time.The bioavailability of Cu in the soil was altered by root exudates,thus to affect the uptake and accumulation of Cu by castor.2.Citric acid,tartaric acid,and oxalic acid were added into the artificial Cu contaminated soil to investigate the effect of exogenous organic acid on the chemical speciation of soil Cu.Results revealed that all the three organic acids increased the concentration and proportion of acid exchangeable Cu,thus effectively activated the Cu in the soil.Compared with the control,the concentration of acid exchangeable Cu in the polluted soil increased by 24.1% when 20 mmol/kg citric acids were applied,and the increasing rate reached a maximum value.The activation effect increased with the increase of the concentrations of organic acids supplied.To sum up,the citric acid was better than other two organic acids on activating Cu in the soil.3.The hydroponic experiment was carried out to explore the effect of Cu stress on the growth of castor,antioxidant system,photosynthesis pigments in castor tissues.The accumulation of Cu in the castor tissues was determined.The results demonstrated that high concentration of Cu inhibited the growth of castor by reducing the fresh weight of the plant and the contents of photosynthetic pigments in leaves.Malondialdehyde(MDA)contents in leaves also increased.The sequence of the concentration of subcellular Cu in roots was: cell wall > organelle > cytoplasmic soluble fraction,while the sequence of that in leaves was: cytoplasmic soluble fraction > cell wall > organelle.4.The total amount of free amino acids in roots,leaves,and phloem increased with the increasing concentrations of Cu.The total amount of free amino acids in the roots,phloem,and leaves of castor treated with 24 mg/L Cu were 3.08,5.95 and 8.81 times of that in control,respectively.Aspartic acid(Asp),threonine(Thr),arginine(Arg),glutamine(Glu),and histidine(His)were the major amino acids in the castor tissues in term of their proportions,which might be involved in the detoxification and transportation of Cu in castor.The proportions of Asp,Thr and Arg in total free amino acids in the root of castor treated with 24 mg/L Cu were 23.9%,32.5%,and 25.8%,respectively;while their proportion in the phloem were 3.5%,41.0% and 27.8%% respectively.The Cu speciation in castor tissues also was investigated with the aid of XANES technology.The Cu most existed in castor tissues were in the form of Citrate-Cu,Arginine-Cu,Cu3(PO4)2 and Cu O.When castor treated with 24 mg/L Cu,Citrate-Cu,Arginine-Cu,and Cu O were the main speciation in the roots,accounting for 34%,35.8% and 18.1% of the total Cu.Moreover,Cu O and GSH-Cu were the most stable forms in the stems,accounting for 30.9% and 39.5% of the total Cu,respectively.While Citrate-Cu,Arginine-Cu,Cu O,and GSH-Cu were the main speciation in leaves,which accounted for 24.9%,25.6%,15.1% and 13.5% of the total Cu,respectively.In summary,the intracellular deposition was another way for castor to enrich Cu.5.The application of phosphate fertilizer affected significantly on the uptake of Cu by castor in the Cu-contaminated soil.The application of oxalic acid activated phosphate rock(APR),sodium dihydrogen phosphate,and calcium phosphate all significantly increased the biomass of castor and promoted the accumulation of Cu in the roots.Compared to control,the MDA content and antioxidant enzymes(CAT,SOD)activities in leaves both decreased after addition of phosphate fertilizer in the soil,while the GSH content increased significantly,indicating that the ability of antioxidant defense system was enhanced with the addition of phosphate fertilizer.The amount of extracted Cu from contaminated soil increased most significantly after application of 600 mg/kg P2O5 in the form of sodium dihydrogen phosphate.When the phosphorus fertilizer level was 600 mg/kg,the total amount of Cu extracted by castor in Ca(H2PO4)2,APR and Na H2PO4 treatments reached 2.98,3.08 and 3.65 mg,respectively.The application of phosphate fertilizer for one time can be effective in providing phosphorus for a long term.Therefore,the addition of phosphate fertilizer with appropriate amount is an effective method to increase the biomass of castor and the extraction of Cu from the soil,which provides a theoretical basis for the future practice of phytoremediation.6.The addition of citric acid,oxalic acid,and tartaric acid increased the castor's biomass in the two Cu-contaminated soils and promoted the transport of Cu from root to shoot.The addition of organic acids did not alter the soil p H significantly,but it increased the activity of soil enzymes,i.e.,urease,catalase,and alkaline phosphatase to some extent.The addition of exogenous organic acids has no much influence on the concentration of acetic acid-extractable Cu in the soil(LC-Soil)with a low concentration of Cu,a high concentration of phosphorus and organic matters thus reduced the accumulation of Cu in castor.However,the addition of exogenous organic acids increased the concentration of acetic acid-extractable Cu in the soil(XC-Soil)with a high concentration of Cu,a low concentration of phosphorus and organic matters thus promoted the accumulation of Cu in castor.