Electrochemical DNA Sensors Based On Functional Materials For Lead(Ⅱ) And Mercury(Ⅱ) Detections In Aqueous Solution

Heavy metal pollutants possess the characters of high toxicity and difficult degradation,and enrich in the human body via food chain with low concentration.It is harmful to humans and environment for the occuraences of the heavy metal accidents in recent years.The detection technology should be enhanced in great demand for heavy metal pollutants real time monitoring and laying foundation for heavy metal treatment.The mian way for heavy metal detections are spectral and chromatographic methods in the world.That obtains the shortcomings of high cost,difficult mai ntain,cumbersome operation,low interference and low accuracy.However,sensors not only obtain the features of low cost,simple operation,but also can quantitatively and sensitively detect pollutants.Comparing with the specific biomolecules,such as enzyme and immunoglobulin,DNA is more easily to be synthesized and modified,more specific for targets,more stable for long-term storage and more resistant to complex environment.Sensors developed with DNA have aroused widespread concerns in environmental detection field.Different configuations of DNA sensors applied for heavy metal ions(as Pb²⁺and Hg²⁺)detections were developed in this paper.With the completed work,Pb²⁺sensor based on G-quadruplex and triplex DNA,Hg²⁺sensor based on"Y-shape"DNA,Pb²⁺sensor based on DNAzyme with signal amplification were fabricated.This paper includes the following four parts:The first section was about that G-quadruplex which was composed of guanine（G）-rich nucleic acid sequence could specificly react with Pb²⁺and form the stable accumulational configuration with four G-quarter.G-rich probe（S1）combined on GCE which was modified with ultrathin g-C₃N₄ nanosheets and electrodeposition gold nanoparticles（EAu）via Au-S bond and hybridized with the complementary probe S2.The DNA duplex broke and S1 transformed into G-quadruplex with the existence of Pb²⁺.Methylene blue（MB）had an affinity with G could bind with G-quadruplex via electrostatic adsorption and intercalation.Pb²⁺was detected by square wave voltammetry（SWV）with the linear detection between 2.0×10^-33 M and 1.0×10^-6 M and detection limitation of 6.23×10^–8 M.Also,the DNA sensor was tested in real water samples with excellent selectivity.According to the research of section（1）,DNA sensor was fabricated based on triplex DNA probe which was synthesized with ssDNA by PCR and composed of the G-rich circle and triple helix with thymine·adenine-thymine（T·A-T）.The triplex DNA combined on the modified GCE based on reduced graphene oxide（rGO）and EAu via Au-S.The triplex DNA broke and G-quadruplex formed when Pb²⁺was captured by the G-rich loop.MB acted as the signal indicator in the research.The sensor possessed high sensitivity to Pb²⁺in the linear detection between 0.01 and 10μM,and the limitation of detection was 3.62×10^–10 M.In addition,"Y-shape"DNA,comparing with the ordinary of DNA duplex structure,can be more facilely synthesized with one step.Sensors developed with that strategy could minimize utilization of reagents and multiple washing steps for potentially maintaining the stability and reproducibility of sensors.As we known from the section of（1）,g-C₃N₄ obtained lager surface area,higher electrical conductivity and better water solubility with long time ultrasound.In order to further enhance the properties of g-C₃N₄,carbon was self-doping into it.C-g-C₃N₄ was dealt with ultrasonic treatment for a long time.And ultrathin C-g-C₃N₄ nanosheets was produced with lager surface area,better electrical conductivity and higher dispersibility in water.Glass carbon electrode（GCE）orderly modified with ultrathin C-g-C₃N₄ nanosheets and EAu combined with S1 via Au-S bond.In the present of Hg²⁺,S2 and S3,partial bases complementary occurred between S1 and S2 via thymine-Hg²⁺-thymine(T-Hg²⁺-T)mismatch,the remaining bases complementary accomplished with each other via the principle of Watson-Crick base pairing.Therefore,the stable Y-shape DNA structure fabricated with simply one-pot method.MB was the electrical indicator for Hg²⁺detection.S3 containing poly adenine（polyA）could be stably bound on nanoAu for the enhancement of electrical conductivity.It indicated that the sensor could be fabricated with simple process,the linear detection ranged from 5.0 nM to 5000.0 nM with the detection limitation of 1.62×10^–11 M.Also,the sensor was tested with tap water,Xiangjiang River and medical wastewater samples for Hg²⁺detection.Moreover,some nucleic acids（DNA and RNA）possess the ability of biocatalysis in the present of heavy metals.And based on the researches of section 1,2 and 3,the sensors were just carried on the reactions between DNAs and heavy metal ions with limited current response.From the above considerations,sensor based on signal amplication with DNAzyme could be developed for better detection performance.Therefore,Pb²⁺sensor based on ordered mesoporous carbon nitride（MCN）and EAu and the electrical signal amplification strategy was fabricated.The electrical signal amplification was composed of MCN,nanoAu,MB and substrate DNA via thiol.8-17DNAzyme was the probe S2 of metalloenzyme and the substrate probe S1 was the complementary strand of S2 and the DNA/RNA chimera with a ribonucleotide adenosine（rA）.The metalloenzyme of 8-17 DNAzyme could catalyze hydrolysis in the present of Pb²⁺,DNA duplex of S1 and S2 was broken and S1 was splited at rA.The residual strand of S1 combined on the electrical signal amplification could react with the complementary DNA anchored on MCN and EAu modified elec trode.Pb²⁺was detected with the indirectly electrical responses.The linear detection of the high performance sensor ranged from 1.0×10^-3 M to 1.0×10^-1414 M.The sensor also carried in real water samples for testing.The performance of the sesor was much higher than other Pb²⁺sensors.It indicated that MCN and EAu can improve the the electrical conductivity and specific surface area,also,the electric signal amplification strategy based on MCN,nanoAu and MB could enhance the performance of detection.