Investigation On The Interaction Between Several Synthetic Food Additives And DNA And The Effect Of Food Additives On DNA Conformation

Food additives are importance in improving the quality of food including color,aroma,taste and shape,improving food quality and prolong the storage life of food,which are important part of food industry.In recent years,some enterprises,especially small food workshops used the additives without following the national standard especially in small food processing to pursue the color and taste of food in recent years,causing damage to human health.Therefore,in accordance with national standards of food safety strickly controlling in the dosage of food additives.Calf thymus DNA(ctDNA)as a carrier of human genetic information,involved in the synthesis,replication and transcription process of protein and enzyme.As the same time,DNA is deemed as the target of many chemical harmful substances.Thus,in this paper,the binding mode,site,property and the effect of DNA conformation between synthetic food additive(maltol,vanillin,ethyl vanillin and benzoyl peroxide as soon)and ctDNA were investigated by multiple spectroscopy methods with chemometrics and molecular docking under simulative physiological condition(pH 7.4),to provide theoretical basis for further understanding the mechanism of action and toxicity of these synthetic food additives and ctDNA.The main contents of these investigations were summarized as follows:1.The binding properties and mode associated with calf thymus DNA(ct DNA)upon interaction with maltol(MAL)in vitro was investigated in physiological buffer(pH 7.4)by multi–spectroscopy technology coupled with melting and viscosity experiments of DNA.The results indicated that the melting temperature and relative viscosity of DNA didviscosity of DNA did not changed significantly,and with increasing concentration of NaCl,the absorbance of MAL–ctDNA had no significant change,which indicated that the mode between MAL and ctDNA was groove and no electrostatic.The calculated themodynamic parameters by the temperature experiment of ultraviolet indicated that the force was hydrogen bonding and van der Waals.Results from CD and FT–IR analysis manifested that the specific binding site was T base,and MAL induced B–form of ctDNA vary from compact morphology to loose morphology.Then the molecular docking further verified the results of experiments and intuitively showed the binding pose of MAL with ctDNA.The results of agarose gel electrophoresis indicated MAL didn't induce DNA damage.2.In this study,the interaction between VAN or EVA and calf thymus DNA(ctDNA)were characterized by multi-spectroscopic methods,multivariate curve resolution-alternating least-squares(MCR–ALS)chemometrics algorithm and molecular simulation.The concentration profiles for the components(VAN or EVA,ctDNA and VAN–ctDNA or EVA–ctDNA complex)by the MCR–ALS analysis showed that VAN or EVA interacted with ctDNA and formed VAN–ctDNA or EVA–ctDNA complex.The groove binding of VAN or EVA to ctDNA was supported by the results from viscosity measurements,melting studies,denaturation experiments,and competitive binding investigations.Analysis of the FT–IR spectra corroborated the prediction by molecular docking that VAN and EVA preferentially bound to thymine bases region of ctDNA.The CD and DNA cleavage assays indicated that both VAN and EVA induced conformational change(from B-like DNA structure toward to A-like form),but didn't lead to a significant damage on DNA.The fluorescence quenching of Hoechst 33258–ct DNA complex by VAN or EVA was a static quenching,and the main forces was hydrogen bonding and van der Waals forces.3.Influence of ionic liquid(1-butyl-3-methylimidazolium chloride,BMIM–Cl)on the interactions between calf thymus DNA(ctDNA)and VAN or MAL were investigated in the physiological buffer(pH 7.4)by applying UV-vis absorption,CD and FT–IR,combined with viscosity measurements and melting point effect of DNA.The results showed that BMIM–Cl increased the stability of ctDNA without altering its structure.VAN and MAL could cause little change in meilting temperature and viscosity of ctDNA,and the binding constant between VAN(MAL)and ssDNA was higher than that of VAN(MAL)and dsDNA in the presence of BMIM–Cl,which indicated that VAN and MAL bound to the groove of ctDNA.The calculated enthalpy and entropy changes suggested that the driving force in the binding of VAN(MAL)to ctDNA was hydrogen bonding and van der Waals forces interaction in the present of BMIM–Cl.In addition,the changes in the CD and FT-IR spectra showed that BMIM–Cl have intervention effects on binding sites and comformation between VAN(MAL)and ctDNA.4.The binding characteristics of benzoyl peroxide with 2-hydroxypropy1-?-cyclodextrin(Hp–?CD–BPO)and calf thymus DNA(ctDNA)in pH 7.4 Tris-HCl buffer was investigated by multi–spectroscopic techniques.The results showed the 1:1 inclusion compound was formed between BPO with Hp–?CD,and its inclusion constant was determined to be 2.40 × 104 L·mol–1.MCR–ALS was used to analyze the spectral matrix between BPO and ctDNA.The concentration profiles of the reaction components(BPO?ctDNA and BPO–ctDNA)were obtained.The results of molecular docking indicated that the binding mode between BPO and ctDNA was intercalative.The calculated thermodynamic parameter indicated that the binding of Hp–?CD–BPO inclusion compound to ctDNA was driven mainly by hydrogen bonding and van der Waals interaction.It was found that Hp–?CD–BPO bound to ctDNA with groove and electrostatic binding with no obvious change in melting and viscosity of ctDNA as well as in iodide quenching effect and the fluorescence intensity of Hp–?CD–BPO–ctDNA varied in the presence of Na3PO4.The results from FT–IR and CD analysis manifested that Hp–?CD–BPO preferential bound to the guanine(G)base of ctDNA,and led to a transformation from B-like DNA structure to A-like conformation.