Screening And Synthesis Of Higher Brightness DNA-AgNCs And Application In Detection Of Health Food Adulterants

Long-term consumption of food containing antibiotic residues or illegal addition of antibiotic drugs in health products may lead to a number of diseases,such as cardiovascular failure,interstitial pneumonia,skin allergy and so on.Captopril is one of the important drugs in the clinical treatment of hypertension and congestive heart failure,while bleomycin is a glycopeptide antibiotic used in the treatment of various cancers.In recent years,it has been reported that captopril and bleomycin are illegally added to proprietary medicines or health products because of their significant effects.Therefore,it is of great significance to quantitatively monitor captopril and bleomycin.DNA-templated silver nanoclusters（DNA-AgNCs）have been widely used in food safety,environmental hazard analysis,medical diagnosis,cell imaging and other fields due to its simple synthesis method,good optical performance and low biotoxicity,which integrates target recognition and signal output.The fluorescence characteristics of DNA-AgNCs are highly dependent on their templates,and the synthesis of DNA-AgNCs is particularly important.However,the screening of DNA templates mostly adopts mass random screening method,which is time-consuming and laborious,and the random process limits the acquisition of high-brightness DNA templates.Therefore,in this dissertation,a DNA template with high brightness and quantum yield of silver nanoclusters was obtained by directional screening,and low-cost and highly sensitive fluorescence detection methods were established by using the synthesized DNA-AgNCs as the signal probe for detecting captopril and bleomycin.The specific work contents are as follows:（1）Twenty-one DNA sequences based on the regular changes of C bases and G bases were independently designed as templates for the synthesis of silver nanoclusters,and the fluorescence excitation/emission spectra and fluorescence intensity values of corresponding DNA-AgNCs were measured.The double-stranded DNA template sequence CCCGGG24 and single-stranded DNA template sequence CCCGGG-3 were obtained by two-step screening.The fluorescence quantum yields of the synthesized CCCGGG24-AgNCs and CCCGGG-3-AgNCs reached 26.16%and27.99%,respectively.（2）By exploring the three synthesis conditions of temperature,silver nitrate addition and sodium borohydride addition,the optimal synthesis conditions of DNA:AgNO₃:Na BH₄=1:18:6,40 degrees and 4 hours were obtained.Compared with traditional one-pot synthesis of silver nanoclusters,the new synthesis conditions greatly shortens the time of synthesis,which is beneficial to the detection and application of silver nanoclusters.（3）A novel method for rapid detection of captopril was developed using synthesized CCCGGG24-AgNCs.The thiol groups in captopril molecules can compete with DNA for silver atoms,resulting in fluorescence quenching of DNA-AgNCs.The results of methodology validation showed that the fluorescence quenching degree of the system was linearly related to the concentration of captopril in the range of 0.1-0.8μg/m L,the linear equation was y=1.11647x-0.00397（R²=0.99467）,and the limit of detection was 0.0447μg/m L.The results of selectivity test,accuracy and recovery showed good specificity,stability and repeatability for detecting captopril.（4）A new method for rapid detection of bleomycin was developed using synthesized CCCGGG-3-AgNCs.The activated bleomycin can induced irreversible strand breaks in DNA templates,resulting in fluorescence quenching of DNA-AgNCs.The results of methodology validation showed that the fluorescence quenching degree of the system was linearly related to the concentration of bleomycin in the range of 25-400 nmol/L,the linear equation was y=-59.857x+46124（R²=0.9973）,and the limit of detection estimated to be 4.27 nmol/L.The results of selectivity test,accuracy and recovery showed good specificity,stability and repeatability for detection of bleomycin.