| Objective:(1)The synthesis and biological activity evaluation of Baicalin-2-ethoxyethyl ester(BAE).(2)To establish a rat model of acute kidney injury(AKI)induced by gentamicin,and to explore the therapeutic effect and mechanism of baicalin-2-ethoxyethyl ester on acute kidney injury in rats.Method:(1)Using baicalin as raw material,baicalin-2-ethoxyethyl ester was synthesized by acid chloride method,direct esterification method and halogenated hydrocarbon substitution method;and its structure was characterized by nuclear magnetic resonance and mass spectrometry.The type and amount of the base used in the synthesis of baicalin sodium salt and the type and amount of the crystallization solvent were optimized.Cross-experimental synthesis was optimized to improve yield.(2)Six SD rats were randomly selected as blank control group,and the remaining rats were injected with gentamicin(100 m/kg)intraperitoneally every day for 7 consecutive days to establish acute kidney injury rats.On the 8th day,orbital blood was collected to detect the changes of CRE and BUN levels by plasma creatinine and blood urea nitrogen kit;and 24-hour urine was collected to detect the levels of NAG and LZM to judge the establishment of the model.Thirty rats with successfully established acute kidney injury were divided into model group,baicalin-2-ethoxyethyl ester low,medium and high dose groups and 5groups of verapamil: model group,baicalin-2-Ethoxyethyl ester low,medium,high and verapamil groups,6rats in each group,were given intragastric administration to the rats,while the blank group and model group were given solvent,and all rats were sacrificed after 14 days of continuous intragastric administration.The blood and kidney tissues of the rats in each group were collected,and the appearance changes of baicalin-2-ethoxyethyl ester on the kidneys of the rats in each group were observed,the kidney coefficient was calculated by weighing,and the pathological changes of the kidneys were detected by HE staining;The levels of oxidative stress-related indexes of MDA,SOD,GSH-Px,NO,and NOS in kidney tissue,and the levels of IL-6,IL-1β,and TNF-α inflammatory factors in serum of rats were analyzed by enzyme-linked immunosorbent assay(ELISA);The protein expression levels of Bax,Bcl-2,NF-κB p65 and Caspase3 in the kidney tissue of rats in each group were detected by immunoblotting.Result:(1)In this study,baicalin and sodium carbonate are used to generate salt,and DMF is used as solvent to react with 2-bromoethyl ether to generate baicalin-2-ethoxyethyl ester.The optimal reaction conditions for the reaction of baicalin-2-ethoxyethyl ester are: the ratio of materials is 1:5.5,the reaction temperature is90 ℃,the reaction time is 12 h,and the solvent multiple is 50 times.(2)In this study,rats were injected intraperitoneally with gentamicin 100 mg/kg for 7 consecutive days.The results showed that compared with the blank group,the serum levels of BUN,CRE,NAG and LZM in the model group were significantly increased.A model of acute kidney injury was successfully established;HE staining showed that baicalin-2-ethoxyethyl ester could significantly reduce renal tubular epithelial cell necrosis,tubular vacuolization and inflammatory cell infiltration in AKI rats;compared with the model group,baicalin-2-ethoxyethyl ester can significantly reduce the serum creatinine,blood urea nitrogen and KIM-1content in rats;compared with the model group,baicalin-2-ethoxyethyl ester can significantly reduce the rat The contents of MDA and NO in renal tissue significantly increased the activities of SOD,NOS and GSHPX;compared with the model group,baicalin-2-ethoxyethyl ester could significantly reduce the serum inflammatory factors IL-1β,The levels of IL-6 and TNF-α;Western Blot results showed that baicalin-2-ethoxyethyl ester could significantly reduce the protein expression levels of Bax,Caspase3 and NF-κB p65 in rat kidney tissue.Conclusion:(1)In this experiment,baicalin-2-ethoxyethyl ester was synthesized,which is a new compound that has not been reported.The yield was improved by reaction optimization.(2)Baicalin-2-ethoxyethyl ester protects rats with acute kidney injury by resisting oxidative stress,inhibiting apoptosis of tissue cells,and reducing the level of inflammatory factors in serum. |
