Simultaneous Detection Of Pb(?) And Ag(?) Ions Using Single-labeled Oligonucleotide Fluorescent Probes Based On Different Mechanisms

Due to the high toxicity from the accumulation of heavy metal ionswhich seriously harms the environment and human health,the detection of metal ions would be an important analytical task in biology and environment.To date,many methods based on the specific binding between metal ions and nucleic acids have been developed for the detection of metal ions.In this paper,single-labeled oligonucleotide fluorescent probes based on G-quadruplex,C-Ag(?)-C coordination and the inherent quenching ability of G-rich groups are used for trace Pb(?)and Ag(?)ions.The main results are as follows:(1)Based on Pb(?)can induce guanine to form G-quadruplex structure,Ag(?)and cytosine can form C-Ag(?)-C specific coordination and the inherent quenching ability of G-rich groups,a single-labeled nucleotide fluorescent probe rich in guanine and cytosine was designed with a fluorescent group hexachlorofluorescein(HEX)at one end in this subject.Two different specific structures of G-quadruplex and C-Ag(?)-C coordination can be formed under the induction of Pb(?)or Ag(?)ions,respectively,so that the G-rich group at the other end can occur near HEX fluorescence quenching to determine Pb(?)and Ag(?)ions.(2)In presence of Pb(?),as the concentration of Pb(?)increased,a G-rich oligonucleotide chain was induced to form G-quadruplex structure,making HEX gradually close to G-rich groups and further causing the fluorescence intensity decrease.The experimental results showed that the concentration of Pb(?)in the range of 1-20 nmol/L had a good linear relationship with the quenching rate of HEX with G-rich groups,and the detection limit was 92 pmol/L.The best optimized conditions were p H 6.0 and ionic strength 40.0 mmol/L.(3)In presence of Ag(?),as the concentrations of Ag(?)incresed,a C-rich oligonucleotide chain was induced to form a C-Ag(?)-C,making HEX gradually close to G-rich groups and further causing the fluorescence intensity decrease.The experimental results showed that the concentration of Ag(?)also in the range of 1-20 nmol/Lhad a good linear relationship with the quenching rate of HEX with G-rich groups,and the detection limit was82 pmol/L.The best optimized conditions were p H 6.0 and ionic strength 40.0 mmol/L.(4)By checking the sensitivity,selectivity and standard curve of this test method,the results showed that the established fluorescence probe method had high sensitivity and selectivity to both Pb(?)and Ag(?),and related cations were not easy to interfere with Pb(?)and Ag(?).The standardization test of Pb(?)and Ag(?)in pure water and wastewater samples showed that Pb(?)and Ag(?)can be detected and the detection results were more accurate,with a detection rate of 97%.Pb(?)detection was carried out on Salvia miltiorrhiza and honeysuckle in Chinese medicine samples,and Ag(?)detection was performed in licorice and honeysuckle,which the results were consistent with that of AAS.(5)The mechanism of the interaction between the quenching of G-rich group and the HEX in the HEX-OND probe mediated by Pb(?)and Ag(?)was studied using ultraviolet absorption spectroscopy and thermodynamic methods.The results showed that the number of binding sites for the interaction between the G-rich group and HEX was 0.8,and the binding ratio was close to 1: 1;the reaction can proceed spontaneously,which is an enthalpy driven electrostatic interaction force type.