Synthesis Of Caffeic Acid Sulfonamide Derivatives And Study Of Their Biological Activity

Caffeic acid has good antioxidant activity.The main antioxidant mechanisms include free radical scavenging,inhibiting the formation of free radicals and lipid hydrogen peroxide.Caffeic acid can not only affect the nutritional value and color of food,but also prevent peroxidative damage related to skin aging,cancer,diabetes,Alzheimer’s disease,Parkinson’s disease,cardiovascular and other diseases.However,the solubility and stability of caffeic acid in many common solvent systems are relatively low,lipophilicity is poor,and in alkaline environments,hydroxyl groups are easily oxidized and lose their antioxidant capacity.Amide bond is ubiquitous in proteins and peptides in organisms,and it is also the basic unit for the synthesis of many compounds.Compounds containing amide bond usually have a variety of biological activities.Synthesis of compounds with amide structure can not only improve the solubility,lipophilicity and stability of the original compounds,but also reduce the irritation of the compounds to organisms and improve their pharmacokinetic properties in vivo.The sulfonamide bond can not only make the hydrogen on the nitrogen atom more active,but also accept the attack of the nucleophile.The property of this special structure makes the sulfonamide bond become a hotspot in the study of drug synthesis.In this paper,caffeic acid was selected for structural modification,and a series of caffeic acid amide derivatives with amide bond and sulfonamide bond were synthesized using sulfonamide drugs with sulfonamide bond as the synthon.Then,characterize the structure of the synthesized derivative,detect the ultraviolet absorption peak and lipophilicity of the derivative,evaluate the biological activity of the derivative in terms of antioxidant,antibacterial,and cytotoxicity,and study and analyze the derivative’s antioxidative stress on A549 cells in vitro Ability and mechanism of anti-oxidative stress to seek antibacterial and anti-oxidant drugs with high efficiency and low toxicity.The details are as follows:(1)Synthesis: Structure modification of caffeic acid by three-step synthesis method,purification by extraction,washing and separation,recrystallization,etc.to prepare 20 caffeic acid amide derivatives with amide bond and sulfonamide bond were prepared and named CS1-CS20.The structure of the synthesized derivatives was identified by IR,MS,1H-NMR,13C-NMR and UV-Vis methods,among which 4 were new compounds.(2)Lipophilicity: Firstly,a standard curve of caffeic acid amide derivative in PBS saturated n-octanol solution was prepared,and then the shake flask method was used to detect the oil-water distribution coefficient(Log P)of the derivative in n-octanol-PBS system.It has been proved that the synthesized derivatives have good lipophilicity,and the Log P of these derivatives can reach the range of optimal drug absorption.(3)Antioxidant activity: The DPPH free radical scavenging method was selected to evaluate the antioxidant activity of caffeic acid amide derivatives,and it was proved that these derivatives have antioxidant activity,among which the three compounds of CS8,CS18 and CS20 have the strongest antioxidant activity(4)Antibacterial activity: Two Gram-positive bacteria such as Staphylococcus aureus and Pneumococcus and three Gram-negative bacteria such as Bacillus subtilis,Escherichia coli and Bacillus cereus were selected as the test strains,and the antibacterial activity of derivates were analyzed by the test tube method There are 8 compounds have broad spectrum,among which CS6,CS13 and CS15 have the highest antibacterial activity.(5)Cytotoxicity: The cytotoxicity caffeic acid derivatives were evaluated by MTT assay.It was proved that these derivatives have no cytotoxicity when the concentration was below 50 μmol/L.When the concentration is between6.25-12.5 μmol/L,the derivatives have obvious proliferation promoting effect.(6)In vitro antioxidant activity: Firstly,A549 cells were treated with 800μmol/L H2O2 as a model drug for 4 hours to establish an oxidative stress damage model.Caffeic acid amide derivative with 6.25 μmol / L was selected to pretreat A549 cells.Then,the ROS and MDA levels in A549 cells,as well as the activity of the endogenous enzymes SOD,CAT and GSH-PX were evaluted.It has been proved that these derivatives can reduce the oxidative damage of cells by H2O2,among which CS6 and CS20 have the best protective effect.(7)Expression of antioxidant stress gene: The caffeic acid amide derivatives with better anti-oxidative stress effect on A549 cells were selected,and the gene expression of Nrf2,HO-1,NQO1,TXNRD1 in oxidative stress cells was evaluated by q RT-PCR technology.Our study have shown that these derivatives can not only remove reactive oxygen species,block lipid peroxidation,and inhibit the production of reactive oxygen species,but also regulate the Nrf2 signaling pathway,further up-regulate the expression of genes such as Nrf2,HO-1,NQO1,TXNRD and other genes,promote the production of endogenous antioxidant enzymes,and improve the ability to protect the body from oxidative stress damage.