Functional Preparation Of Multicolor Carbon Quantum Dots And Their Application In Biological Imaging,Foodborne Bacteria Detection And Combating

Food safety is an important issue confronting the global public health system.Foodborne diseases caused by foodborne pathogens or their toxins comprise a public health burden,significantly hindering global social and economic development.Highly sensitive detection methods,as well as strong and safe elimination of foodborne bacteria,are primary approaches to preventing and controlling foodborne bacteria contamination.One of the advantages of the development of nanomaterials is that methods based on fluorescent nanomaterials can be used for biological imaging and rapid detection of bacteria.Meanwhile,compared with traditional antibiotics,nanomaterials are being widely studied as new antibacterial agents due to the lack of their efflux pump of bacteia,the ability to increase the permeability of bacterial cell membrane,and the difficulty in causing bacterial drug resistance.However,these nanomaterials are mainly metal,semiconductor,rare earth nanomaterials and cationic conjugated polymers,and their potential environmental and biological toxicity limit their application.Carbon quantum dots?CDs?perfectly solve this problem because of their excellent optical properties,good antibacterial properties and biocompatibility.The detection and elimination of foodborne bacteria has been greatly improved.Thus,CDs exhibiting multicolor fluorescence emission and their multifunctional nano-assemblies were prepared in this study for biological imaging as well as rapid,highly specific,and highly sensitive detection of foodborne bacteria.Efficient and safe antibacterial CDs were synthesized by further surface functionalization to eliminate foodborne bacteria.To improve their antibacterial performance in vivo,p H-responsive antibacterial CDs releasing hydrogel was further constructed for in vivo antibacterial applications.This study presents an idea for the establishment of CD-based early monitoring and elimination of potential foodborne bacteria in food.1.Controlled synthesis and identification of CDs exhibiting multicolor fluorescent emission.Three isomers of phenylenediamine and graphite rods were used as precursors by using the hydrothermal method?“bottom-up”approach?and electrolysis?“top-down”approach?to synthesize multicolor fluorescent CDs and N,S-doped carboxyl–graphene quantum dots?GQDs??N,S/GQDs?,respectively.The optimal conditions for phenylenediamine during the synthesis of CDs exhibiting multicolor fluorescence emission were determined as follows:concentration,2 mg/m L;volume,30 m L;reaction time,3 h;and reaction temperature,120°C.Meanwhile,N,S/GQDs were synthesized by the electrolysis of graphite rods in 0.1 M Ts ONa acetonitrile solution at+3.0 V potential for 2 h.The synthetic CDs were finally characterized and identified in detail,providing conditions to establish biosensors and nano-assemblers for bioimaging and detection of foodborne pathogens using the fluorescence properties of the CDs.2.Optical properties and biological imaging applications of CDs exhibiting multicolor fluorescence emission.The luminescence mechanism of previously synthesized CDs exhibiting multicolor fluorescence emission was further discussed.The redshift in the fluorescence emission of CDs was related to the gradual increase in graphitization and graphitic nitrogen content.The CDs exhibiting multicolor fluorescence emission had long fluorescence lifetime,high fluorescence stability,and absolute fluorescence quantum yield.The fluorescence lifetime and absolute fluorescence quantum yield of B-CDs?Blue fluorescence emission CDs?were 5.57ns and 3.36%,respectively.R-CDs?Red fluorescence emission CDs?were irradiated with the highest excitation light for 1 h.The fluorescence intensity of B-CDs was stable,and more than90%of the initial intensity was maintained.To enhance the application of GQDs in cell imaging and detection,the aptamer fluorescence sensor based on nanomaterial surface energy transfer?NSET??Aptamer@Fe₃O₄@GQDs@Mo S₂?was further constructed,together with aptamers Mo S2 and Fe₃O₄,for the biological imaging and detection of tumor cells.The sensor exhibited good linearity at 2–64 n M,the linear equation was y=5.202x+0.744,R² was 0.9928,and the limit of detection?LOD?was 1.19 n M.The aptamer sensor showed good specific recognition for the epithelial cell adhesion molecule and good biocompatibility.The NSET biosensor was used to label and detect tumor cells in actual blood samples.The cell capture efficiency of all samples reached about 80%–90%.Finally,with the excellent red fluorescence emission performance and bio-penetration ability of R-CDs considered,the folic acid@R-CD nano fluorescent probe was further constructed based on in vitro cell imaging and successfully used for in vivo tumor imaging in mice.3.Construction of an“ON–OFF–ON”fluorescence sensor based on CD/Fe₃O₄ to detect foodborne pathogenic bacteria.To achieve rapid,highly specific,and sensitive detection of foodborne pathogens,B-CDs were modified with the c DNA?B-CD+c DNA?,on the basis of previous synthetic B-CDs.The Staphylococcus aureus?S.aureus?aptamer was modified on the Fe₃O₄ surface?Fe₃O₄+Aptamer?,the Fe₃O₄+Aptamer and B-CD+c DNA self-assembled into a nano-dimer?B-CD/Fe₃O₄?via hydrogen bonding.B-CDs acted as fluorescence donors,and Fe₃O₄ was the acceptor to form an“ON–OFF–ON”sensor based on fluorescence resonance energy transfer.This magnetic fluorescence aptasensor was used to separate and detect S.aureus.The ratio of the fluorescence donor and acceptor that affects the quenching efficiency of the sensor was explored.When the ratio of the donor to the acceptor was 2:1,the optimal quenching efficiency was achieved,and when the incubation time of the fluorescence sensor and target bacteria was 30 min,the highest system fluorescence recovery was reached.The B-CD/Fe₃O₄aptamer fluorescence sensor maintained a good linear relationship at a bacterial concentration of50–10⁷ CFU/m L.The linear regression equation was Y=5327X-5145,R² was 0.9818,and LOD was 8 CFU/m L.The aptamer fluorescence sensor showed exhibited specificity and reproducibility,could efficiently distinguish different kinds of bacteria,and the recovery rate of actual samples was 95%–106%.4.Synthesis and antibacterial properties of ultrahigh positive charge antibacteial CDs based on CD-based surface functional modification.On the basis of the previous real-time detection of foodborne pathogenic bacteria,functional modification CDs?antibacterial CDs?were innovatively introduced to realize the application of CDs in the antibacterial aspect of foodborne pathogenic bacteria.The ligands spermidine and lipoic acid were thus introduced based on previously synthesized multicolor fluorescent CDs to further synthesize new antibacterial CDs?La-Y-CDs and Spe-Y-CDs?.The addition of ligands resulted in a redshift?La-Y-CDs?and a blueshift?Spe-Y-CDs?in the fluorescence emission spectra of the synthesized CDs.By X-ray photoelectron spectroscopy,infrared spectroscopy,and other analyses,the red shift of the fluorescence emission spectra of La-Y-CDs was related to the doping of the S element and the increase in carboxyl content.Meanwhile,the doping of spermidine endowed the surface of Spe-Y-CDs with abundant amino groups,resulting in a zeta potential as high as+51.20 m V.The antibacterial properties of each CDs were further evaluated in vitro,and the minimum inhibitory concentration?MIC?of Spe-Y-CDs was 781 times lower than that of its precursor?spermidine?.Compared with existing antibiotics,Spe-Y-CDs had broader antibacterial spectra and even can combat drug resistance bacteria.Meanwhile,Spe-Y-CDs had a better antibacterial ability than that of antibiotics.This difference might be attributable to the abundance of amino groups on the surface of Spe-Y-CDs.The ultrahigh positive charge and small particle size facilitated the binding with bacteria?negative surface charge?,destroyed the surface charge of the bacteria,and entered the bacteria,leading to bacterial metabolism disorder and death.5.Synthesis of ultrahigh positive charge antibacterial CDs releasing hydrogels based on p H response and their antibacterial mechanism and applications in vivo and in vitro.To further enhance the application of the antibacterial properties of previously synthesized antibacterial CDs in vivo,antibacterial CDs were combined with acrylic acid,pectin,ammonium persulfate and poly?ethylene glycol?dimethacrylate to construct p H-responsive antibacterial CDs releasing hydrogel?SCDs-AP hydrogel?.CDs from the SCDs-AP hydrogel in buffer solutions with p H=5.5 and 8.5 showed release rates of about 39%and 36%,respectively,whereas those in the buffer solution with p H=7.4 had a release rate of only 21%.An acidic or alkaline environment can accelerate the release of antibacterial CDs,and the released CDs can exert antibacterial effects.The antibacterial activities of the SCDs-AP hydrogel against Methicillin-resistant S.aureus and Multidrug-Resistant Salmonella Typhimurium were 108.5 and88.5 times higher than that of the control group,respectively.Antibacterial mechanism studies have indicated that bacterial metabolism renders the culture medium acidic,and the SCDs-AP hydrogel achieves an antibacterial effect by releasing antibacterial CDs and destroying bacterial cell walls or membranes.The biocompatibility of the p H-responsive antibacterial CDs releasing hydrogel was explored.Compared with the control group,the SCDs-AP hydrogel exhibited stronger cell adhesion and higher cell viability.The antibacterial activity of the SCD-AP hydrogel in vivo was further studied.Skin tissues around the SCDs-AP hydrogel were in good condition,and the wound was well-repaired.The SCDs-AP hydrogel exhibited an antibacterial effect in vivo and could be used to safely treat bacterial wound infection in vivo.