Investigations On The Toxic Mechanism Of CdTe Quantum Dots On Lysozyme,DNA And Hepatocytes

Nanomaterials(carbon nanotubes,gold nanoparticles,quantum dots(QDs),etc)have been widely used in various important biomedical and biotechnology field,such as biomedical labeling,cellular imaging and tumor therapy.As important nanomaterials,quantum dots present a broad potential in the biological application because of their unique optical and electronic properties including high fluorescent quantum yield,narrow/symmetric emission spectra,broad excitation spectra and high photostability.Quantum dots are semiconductor nanocrystals with a diameter of 2-100 nm which could enter human bodies,tissues or even cells.With the extensive application of QDs,the potential toxicity of these nanoparticles on human health have raised concern.The potential toxicity of QDs can be affectd by the environment conditions and the physicochemical properties including the size,charge,surface coating materials and so forth.Numerous studies have shown that cadmium-based quantum dots accumulate in the main target organs of liver when enter human bodies,so it is necessary to investigate the toxicity of CdTe QDs to humans.We chose different sized NAC-CdTe QDs as exogenous substances and lysozyme as target to explore the relationship between toxicity of CdTe QDs with particle size.Furthermore,we selected hsDNA and liver cell as target material to investigate the interaction mechanisms between DNA and NAC-CdTe QDs from the molecular and cellular perspectives.The major works and results are as follows:In the first part:NAC-CdTe quantum dots(QDs)were synthesized following the one-pod method with minor modifications.Different sized QDs were obtained at 30 min,60 min,90 min,respectively.Then,the as-prepared CdTe QDs was purified and characterized.In the second part:herein,the effect of NAC-CdTe quantum dots(QDs)with different sizes on lysozyme were investigated by isothermal titration calorimetry,enzyme activity assays,and multispectroscopic methods including fluorescence,synchronous fluorescence,three-dimensional fluorescence,light scattering,UV-vis absorption and circular dichroism techniques.Multi-spectroscopic measurements revealed that all of the three sized QDs caused strong quenching of the lysozyme's fluorescence.Moreover,the changes of secondary structure and microenvironment of the tryptophan residues in lysozyme caused by the QDs was higher with bigger size.ITC and enzyme activity assays results proved that NAC-CdTe QDs can spontaneously bind with lysozyme by hydrophobic force and the formation of QDs-lysozyme complex inhibited the activity of lysozyme.Based on these results,we conclude that NAC-CdTe QDs with larger particle size have a larger impact on the structure and fuction of lysozyme.In the third part:the toxic effect of NAC-CdTe quantum dots(QDs)on DNA were investigated by UV-vis absorption,fluorescence,fluorescence lifetime,isothermal titration calorimetry and circular dichroism techniques.We found that DNA effectively quenched the fluorescence of NAC-CdTe QDs by the static quenching mode in a concentration-dependent manner.According to multispectroscopic measurements and ITC result,intercalative binding was the most possible mode when binding NAC-CdTe QDs with DNA and hydrophobic force were the main combining modes.In the fourth part:the cytotoxicity of NAC-CdTe quantum dots(QDs)were explored using the Cell Counting Kit-8(CCK-8)assay,ROS measurement and cell apoptosis assay.(1)CCK-8 assay was used to analyse the cytotoxicity of NAC-CdTe QDs by assessing the cell viability of mouse primary hepatocytes.The cell viability decreased significantly with the exposure concentration,so the cytotoxicity of NAC-CdTe QDs to mouse primary hepatocytes was a dose-dependent manner.(2)The results of ROS measurement showed that the content of reactive oxygen species(ROS)increased with the exposure dose of NAC-CdTe QDs,which caused the oxidative damage to mouse primary hepatocytes.(3)The NAC-CdTe QDs induced apoptosis in mouse primary hepatocytes.Therefore,we speculated that the DNA damage in mice hepatocytes may cause by oxidative stress of NAC-CdTe QDs.In this paper,the toxicity of NAC-CdTe QDs were explored and evaluated from two aspects of molecular and cellular level,which could provide basic data for the toxicity evaluation of CdTe QDs.In addition,it provides a scientific basis for assisting in the design of biocompatible and stable quantum dots by analyzing the correlation between the size and the toxicity of quantum dots.