Interaction Of Two Phenolic Acids With DNA And Their Protection Of DNA Oxidative Damage

As a product of secondary metabolism,are widely used in many natural plants.They are an important class of high active substances,which have many pharmacological activities,such as anti-cancer,anti-inflammatory,antioxidation,antiaging,antiaging etc.Therefore,phenolic acids will have good application value and application prospect in many fields,especially in the field of medicine.DNA is an important genetic material,which is the target of the vast majority of anti-cancer and anti-inflammatory drugs in the organism.Through the specific combination of DNA,the drug molecules can interfere with the gene expression of replication and transcription of DNA,thus the study of the interaction between phenolic acids and DNA will help to understand the mechanism of action of phenolic acids at the molecular level,and provide a theoretical basis for the design and screening of new anticancer drugs.In this thesis,the interaction mechanism between two phenolic acids（caffeic acid and ferulic acid）and calf thymus DNA（ctDNA）were studied by combination of various spectroscopic methods and molecular simulation techniques,combined with melting point,viscosity measurement,and other means by using a variety of spectroscopic methods and molecular simulation techniques in the simulation of human physiological acidity（pH 7.4）.The protective ability and antioxidant capacity of caffeic acid and ferulic acid against oxidative damage of DNA were explored by UV scanning and gel electrophoresis.Meanwhile,the preparation of the inclusion compound of caffeic acid-hydroxypropyl-β-cyclodextrin as well as probing its interaction with ctDNA.The main results are as follows:1.The binding properties between two kinds of phenolic acids（caffeic acid and ferulic acid）and ctDNA were studied by UV–vis absorption and chemometrics（multivariate curve resolution-alternating least squares method,MCR-ALS）.The extended UV spectral matrix of caffeic acid and ferulic acid with ctDNA is decomposed by MCR-ALS.The concentration variation trend of each reaction component in the reaction process and the pure spectrum of each component are obtained.Thus,the monitoring of the reaction process can be realized.It is proved that caffeic acid and ferulic acid have formed a complex with ctDNA.The calculation results of thermodynamic parameters show that the combination of caffeic acid and ctDNA is mainly driven by hydrophobic forces and hydrogen bonds and van der Waals forces played a predominant role in the binding of ferulic acid to ctDNA.2.The binding modes of caffeic acid and ferulic acid with ctDNA were studied by competitive test,viscosity measurement,melting experiment and salt effect.All experimental results indicated that the two phenolic acids interacted with ctDNA through groove binding mode.Infrared spectroscopy showed that the main binding sites of caffeic acid and ferulic acid with ctDNA were A-T rich region,and circular dichroism spectroscopy analysis found that the conformation of ctDNA was slightly disturbed,but did not change the original conformation of ctDNA.The results of molecular simulation showed that caffeic acid and ferulic acid interacted with DNA（ID:453D）at A-T rich region of minor groove,which further supported the experimental results of FT-IR spectroscopy and shows the binding mode of caffeic acid and ferulic acid with DNA.3.The inclusion compound of caffeic acid-Hp-β-CD was obtained by ultrasonic method.The inclusion ratio was determined to be 1:1 and the stability constant was K_1:1=2048.3 L/mol.Characterization of caffeic acid-Hp-β-CD inclusion compound was proved by scanning electron microscopy.In addition,the interaction between caffeic acid-Hp-β-CD inclusion compound and ctDNA was further studied.The results showed that the binding constant between caffeic acid and ctDNA was decreased while caffeic acid included by Hp-β-CD,but the binding mode did not change,with the decrease of ctDNA binding constant,which makes caffeic acid more stable,but the binding mode does not change and still is groove binding.4.Using VE as a positive control,UV scanning and gel electrophoresis were used to explore the protective effects of caffeic acid and ferulic acid on DNA oxidative damage and its antioxidant capacity.The experimental results show that both caffeic acid and ferulic acid have good ability to protect the oxidative damage of DNA caused by hydroxyl radicals,and ferulic acid>VE>caffeic acid.Both caffeic acid and ferulic acid have effective ability to remove all kinds of free radicals,and have good antioxidant capacity.