Study On Biosynthesis And Antioxidant Effect Of Natural Plant Product Caffeic Acid

Nowadays,we are facing two major environmental problems:environmental pollution and energy depletion.With the intensification of the greenhouse effect,the ability to turn carbon dioxide into treasure has become a research hotspot.Algae play a unique role in this field,the research and development of biological resources with cyanobacteria as the research target has received extensive attention.The use of cyanobacteria as a host for the production of bioenergy has both economic and environmental protection value.As a natural phenolic compound,caffeic acid is a typical representative of hydroxycinnamic acid.It is widely present in plants and has good biological activity.In this study,the engineering design concept of synthetic biology was adopted,and Synechocystis sp.PCC6803 was selected as the genetically engineered host algae.By expressing exogenous genes phenylalanine enzyme?pal?,cinnamic acid 4-hydroxylation enzyme?c4h?and p-coumarate-3-hydroxylase?ref8?,caffeic acid was directly synthesized from carbon dioxide using photosynthesis.This study further adopted adriamycin-induced cardiomyopathy as a research model to confirm the antioxidant effect of caffeic acid and its important application value.The main contents and results of this research are as follows:?1?Using bioinformatics technology to systematically analyze the complex metabolic network data of Synechocystis sp.PCC6803,we designed exogenous synthetase genes phenylalanine enzyme?pal?,cinnamic acid 4-hydroxylase?c4h?and the caffeic acid anabolic pathway composed of p-coumarate-3-hydroxylase?ref8?.Using homologous recombination technology to insert the caffeic acid synthesis gene into the cyanobacteria6803 genome,and knock out the gene encoding slr2081 of the cyanobacterium arogenate dehydrogenase to eliminate the competition of endogenous metabolic pathways,driving the synthesis of caffeic acid under the induction of the strong promoter PrbcL gene expression,so as to realize the directed biosynthesis of cyanobacteria that uses photosynthesis to convert the greenhouse gas CO₂ into natural plant products.?2?On the basis of successful screening to obtain mutant Synechocystis sp.PCC6803containing the caffeic acid synthesis gene,the growth activity of mutant and wild-type Synechocystis sp.PCC6803 was further analyzed,and the yield of caffeic acid was analyzed by HPLC.The results showed that there is no significant difference in growth status between the wild type and the mutant type.When the cell growth OD₇₃₀=1.0,the production of caffeic acid in the mutant medium reached 5.603 mg/L,while the target product caffeic acid was not detected in the wild type medium.?3?Caffeic acid has a significant improvement on adriamycin cardiomyopathy,mainly in inhibiting oxidative stress and protecting mitochondrial damage.Caffeic acid can reduce the oxidative effect of adriamycin on cardiomyocytes through the combination of adriamycin and caffeic acid.It can also protect the mitochondria of cardiomyocytes,which can improve the viability of myocardial cells and inhibit the oxidation of cardiomyocytes,this response works by regulating NF-?B p65.It has potential application value in the field of adriamycin cardiomyopathy treatment and biopharmaceutical research and development.