Detection Of Sulfadimethoxine In Aquatic Products Based On Aptamer Flurescence Sensors

As a sulfonamide widely used in aquaculture,sulfadimethoxine(SDM)is easily accumulated in aquatic animals,causing some adverse reactions and side effects to human health.Hence,it is important to detect the residue of SDM.In this dissertation,based on the specific recognition performance of nucleic acid aptamer,several label-free fluorescence sensing methods were established for the detection of SDM in aquatic products.The dissertation mainly includes three parts:Section 1,a method for detecting SDM was established based on the specific recognition of SDM-aptamer and the inner filter effect of CdTe QDs and AuNPs.In the absence of target,AuNPs dispersed in salt solution because of the aptamer protection,which could effectively quench the fluorescence emission of CdTe QDs.While in the presence of SDM,AuNPs aggregated due to the specific recognition of SDM-aptamer to SDM,which resulted in the fluorescence recovery.According to the degree of fluorescence recovery of CdTe QDs,the SDM content could be detected.The optimized detection conditions were described as follows:150 mmol L-1 NaCl,0.25 μmol L-1 of SDM-aptamer and 1.5 μmol L-1 of CdTe QDs,and the binding time of aptamer to SDM was 20 min.Under this condition,the linear range of detection of SDM was 10 ng mL-1-250 ng mL-1,and the detection limit was 1.54 ng mL-1.This method was applied to the detection of SDM with recoveries ranging from 97.3%to 100.6%in fish samples and from 104.1%to 112.3%in water samples.Section 2,a method for detecting SDM was established based on the specific recognition of SDM-aptamer and the quenching effect of PDDA on CdTe QDs fluorescence.The positive charge of poly(diallyldimethylammonium chloride)(PDDA)caused the fluorescence quenching of negatively charged CdTe QDs.However,aptamers can restore the CdTe QDs fluorescence due to the negatively charged phosphate backbone that bound to PDDA to form a "double-stranded"structure.When SDM was present in the solution,the aptamer specifically bound to SDM so that it no longer interacted with PDDA,resulting in CdTe fluorescence being quenched by PDDA.This method had the advantages of simple system and short time.Under the optimized conditions,the linear range of detection of SDM was 25 ng mL-1-300 ng mL-1,the detection limit was 22.38 ng mL-1.This method was applied to the detection of SDM with recoveries ranging from 104.2%to 117.7%in fish samples and from 94.2%to 112.6%in water samples.Section 3,based on the competitive binding of aptamers to complementary strand DNA and SDM,the SGI staining fluorescence of dsDNA,a sensitive detection method for trace SDM was established.AuNPs bound with DNA-aptamer against SDM can be centrifuged with a red color in residue,and the complementary strand DNA(cDNA)and SYBR Green I(SGI)subsequently mixed in supernatant stand alone with no fluorescent emission.However,in the presence of SDM,the aptamer specifically binds to SDM in supernatant,leading to a blue color of AuNPs deposited in PBS solution after centrifugation.After adding cDNA and SGI in supernatant,cDNA and aptamer hybridize because of the higher affinities of aptamer to cDNA than that of SDM,and the double stranded DNA can be dyed by SGI producing a fluorescence emission at 530 nm.Therefore,SDM can be quantitatively detected by the fluorescent intensities.The optimized conditions were described as follows:87.5 nmol L-1 of aptamer,pH 8.0 for PBS,20 mmol L-1 MgCl2,1:15 molar ratio of aptamer to SGI,109.4 nmol L-1 of cDNA,and the response time was 4 min.Under the optimized conditions,the linear range of detection of SDM was 2 ng mL-1-300 ng mL-1,the detection limit was 0.62 ng mL-1.This method was applied to the detection of SDM with recoveries ranging from 99.2%to 102.0%in fish samples and from 99.5%to 100.5%in water samples.