Biosensing Methods Based On Specific Deoxyribozyme And Used For Uranyl Ion Detection In Water

The production of nuclear products and the subsequent treatment of nuclear waste can lead to the discharge of uranium into the environment in the form of ions,causing pollution of surface water and groundwater,and then causing great harm to the human body.Uranium exists stably in the form of uranyl ions(UO₂²⁺)in aqueous environments.Deoxyribozyme（DNAzyme）is widely concerned by researchers because of their superior specificity and sensitivity.It is often used as excellent DNA probes for the detection of trace or even trace metal ions.In this paper,three biosensing systems based on specific DNAzyme combined with fluorescence and colorimetry were designed and successfully applied to the detection of UO₂²⁺in actual water samples.Chapter 1 of this paper introduces the pollution status and detection methods of uranium,the research status of DNAzyme,and the basic properties and applications of materials such as molybdenum disulfide,gold nanoparticles and G-quadruplex.Chapter 2 of this paper presents an ultrasensitive fluorescence sensing method for UO₂²⁺detection based on the principle of entropy-driven cyclic amplification.The method is designed with the strategy of cyclic amplification.UO₂²⁺-specific DNAzyme was cleaved under the action of UO₂²⁺to release DNA fragments.A cyclic amplification reaction is initiated by the DNA fragment and the FAM-DNA contained in the DNA complex was released.Mo S₂ has a high affinity for DNA single strands and a low affinity for DNA double strands.DNA single strands with fluorophores bind to Mo S₂,and the fluorescence intensity was greatly reduced.There was a linear relationship between fluorescence intensity and UO₂²⁺in the range of 10～100 pmol/L（r=0.9932）,and the detection limit（LOD）was 3.60 pmol/L.The method was successfully used for the detection of UO₂²⁺in water samples,the recovery was in the range of 97.0%～103.0%,and the relative standard deviation（RSD）of the method was in the range of 1.0%～2.6%.These results demonstrated that we have successfully developed a fluorescence sensing method based on an entropy-driven cyclic amplification mechanism for ultrasensitive detection of UO₂²⁺in water.Chapter 3 of this paper proposes a biosensing system for UO₂²⁺detecting based on gold nanoparticle-modified DNA probes（Au NPs/DNA）.The modified fluorophore on the substrate chain of DNAzyme bind firmly to the DNA probe modified by Au NPs after the substrate chain was broken and detached.Au NPs have a quenching effect on fluorophores.The concentration of UO₂²⁺detecting was characterized according to the quenching degree of fluorescence in the system.The change of fluorescence intensity was linearly related to UO₂²⁺in the range of 2～100 nmol/L（r=0.9991）,and LOD in the method was 1.38nmol/L.The method was successfully used for the detection of UO₂²⁺in actual water samples,the recovery was in the range of 95.9%～103.6%,and the RSD was in the range of 0.8%～2.7%.These results demonstrated that we have successfully developed a fluorescent sensing method based on Au NPs/DNA for highly sensitive detection of UO₂²⁺in water.Chapter 4 of this paper presents a colorimetric biosensor based on G-quadruplex and hemin（G4s/hemin）.Under the action of UO₂²⁺,the DNAzyme was cleaved,and the detached substrate strand combined with a DNA double-strand with a naked part,and a G4s was formed by the unbound part due to the presence of G-rich sequences.The G4s combined with hemin to catalyze the oxidation of TMB by peroxide and the system appeared blue.An absorption peak appeared at a wavelength of 650 nm.The concentration of UO₂²⁺was characterized according to the height of the absorption peak.The experimental results showed that the method had a linear relationship in the range of 5～150 nmol/L（r=0.9972）,and LOD in the method was 3.06 nmol/L.The method was successfully used for the detection of UO₂²⁺in water samples,the recovery was in the range of 99.4%～106.0%,and RSD was in the range of 2.5%～4.2%.These results demonstrated that we have successfully developed a fluorescent sensing method based on G4s/hemin for highly sensitive detection of UO₂²⁺in water.