Novel Strategies For Signal Conversion And Signal Amplification Of Electrochemical Biosensor And Their Applications

With the rapid development of modern industry and agriculture,more and more toxic contaminants are spilled into the environment,which becomes a serious threat to human health and ecological safety.Microbial whole-cell biosensors are able to effectively detect toxic pollutants.However,conventional biosensors usually use fluorescence or enzyme activity as the output signal,which requires complex instruments for signal conversion to achieve quantitative detection.Previous study in our laboratory demonstrated that it is possible to construct a whole-cell electrochemical biosensor with current as the direct output signal by adopting the electroactive microorganisms and its extracellular electron transfer capability.In this study,Shewanella oneidensis MR-1 was used as the typical electroactive microorganisms to develop a new signal conversion strategy by converting physiological signals of cellular metabolic activity into electrochemical signals and was applied to detect water toxicity.Furthermore,a signal amplification strategy that utilizes the redox of electroactive microorganisms to enhance the electrochemical signal is proposed and applied for the ultra-sensitive detection of environmental bio-virulence factor.The main research achievements are as following:1)The effect of toxic contaminants in water on the inhibition of the electron transport activity of electroactive microorganisms was studied.It was found that there is a linear response between the electronic output efficiency of electroactive microorganisms and the concentration of toxic substances.Based on this,a biological toxicity signal conversion strategy was proposed which was further converted to the current output profile of the microorganism and the the degree of magnitude on current alteration was used to access the overall water toxicity.At the same time,it has been found that the extracellular electron transfer capacity of electroactive microorganisms can be used as an electron generation system,which can continuously provide the redox recyclability of electrons that retain redox activity.An electrochemical signal amplification strategy for reduction/reactivation cycles was proposed in order to achieve the redox cycle of the small molecule,and enhance its electrochemical response signal.2)Based on the signal conversion strategy,a novel bio-electrochemical dual signal for bio-toxicity of heavy metals was developed which was assessed by the inhibition ratio of maximum output current and lag time for the maximum current evolution.The half maximal inhibitory concentration of Cu²⁺,Cr⁶⁺,Cd²⁺and Zn²⁺determined by this biosensor were about 0.35mg/L,2.48 mg/L,6.52 mg/L and 22.63 mg/L.3)A novel bioelectrochemical method for the detection of pyocyanin（Pseudomonas aeruginosa biomarkers）was developed based on the signal amplification strategy,which can use the forward electron transfer ability of S.oneidensis MR-1 to provide electrons for the reactivation/recycling of pyocyanin.This enabled sensitive detection of pyocyanin with low LOD was 47±1 pM which 3 orders of magnitude lower than other electrochemical methods and successful application to the analyses clinical samples.4)Furthermore,based on the basic principle of bio-electrochemical signal amplification,the bidirectional electron transfer ability of S.oneidensis MR-1 was utilized to simultaneously activate the redox of pyocyanin and 1-hydroxyphenazine（another biomarker of Pseudomonas aeruginosa）.A new bioelectrochemical method was developed for the simultaneous detection of pyocyanin and 1-hydroxyphenazine,which enabled the sensitive detection of pyocyanin and1-hydroxyphenazine.