| Non-invasive micro-test technology（NMT）is a technology that can be used to detect physiological functions of living materials,which can detect information such as the flux and three-dimensional motion direction of specific ions or molecules.This technology does not cause any damage to the sample during the detection process,and it can ensure that the data obtained from the test is generated under the normal physiological state of the sample.NMT is now widely used to explore physiological characteristics that are difficult to measure by other technologies because of its non-destructive,high spatio-temporal resolution,and real-time dynamic detection.It has important significance for the study of life activities pattern of organisms,especially for the interaction between plants and the environment.Microelectrode is an important part of NMT.With the development of heavy metal microelectode,this technology is widely used in the study of ecotoxicology and bioaccumulation mechanism of heavy metals in plants.Nevertheless,at present,there is only one Cd2+microelectrode that was generally applied,which greatly limits the application of NMT in the field of heavy metal ecotoxicology.Therefore,this research aimed to study the development,performance characterization and optimization of other heavy metal ion（Cu,Pb,Zn）microsensors used in NMT,and to explore the application of various heavy metal microsensors in the detection of heavy metal flux in the rhizosphere of plants based on NMT.（1）Firstly,a Cu2+selective microelectrode for NMT was constructed with N,N,Nμ,Nμ-Tetracyclohexyl-2,2μ-thiodiacetamide as the ionophore.The linear response range of the microelectrode in DI water is 10-110-6 mol/L,the detection limit is 10-6.4 mol/L,and its Nernst slope is 27.29 mV/dec;While in simple simulated soil solution,it displayed a Nernstian response within Cu2+concentration range of 10-110-5 mol/L with slope of 26.37 mV/dec.The microelectrode can work well within a wide pH range of 2.06.5 and it maintain good stability in actual detection of Cu2+flux at plant root surface.（2）Then,a Pb2+selective microelectrode for NMT was constructed Based on tert-butylcalix[4]arene-tetrakis（n,n-dimethylthioacetamide）as an ionophore,The linear response range of the microelectrode in DI water is 10-210-7 mol/L,and its Nernst slope is 29.25 mV/dec;While in simple simulated soil solution,it displayed a Nernstian response within Pb2+concentration range of10-210-6 mol/L with slope of 26.37 mV/dec.The microelectrode can work well within a wide pH range of 1.07.0 and it maintain good stability in actual detection of Pb2+flux at plant root surface.（3）At the same time,a Zn2+selective microelectrode for NMT was constructed with Tetra-n-butylthiuram Disulfide as an ionophore and for the first time it was used for real-time monitoring Zn2+ion flux at plant rhizosphere under living condition.The linear response range of the developed microelectrode to Zn2+in DI water is 10-610-1 mol/L,the Nernst slope is 30.21 mV/dec.The response time of the microelectrode is less than or equal to 1 s and the microelectrode can work well within a wide pH range of 3.57.0;While in simple simulated soil solution,it displayed a Nernstian response within Zn2+concentration range of 5.0×10-510-1 mol/L with slope of 28.08mV/dec.This indicated that the microelectrode has good anti-interference performance to the coexisting cations in soil solution.（4）At last,the developed Cu2+,Pb2+,Zn2+selective microelectrodes and a commercially available Cd2+selective microelectrode were then used todetect rhizosphere ion flux on a Zn/Cd hyperaccumulation plant Sedum plumbizincicola and non-hyperaccumulation ectotype plant Sedum alfredii Hance.The results found that the Sedum plumbizincicola and Sedum alfredii Hance have different characteristics of heavy metal ion flux in the rhizosphere.The Sedum plumbizincicola showed significant Zn2+and Cd2+efflux in the mature area,which may be an important reason for its ability to tolerate heavy metals in the environment. |
