Optimization Of The Extraction Process And Anti-BVDV Mechanism Of Polysaccharide From Baitiao-Codonopsis Pilosula

As the local medicinal material of Gansu Province,Codonopsis pilosula is one of the main traditional Chinese medicine preparations.Numerous studies have shown that traditional Chinese medicine polysaccharides have good pharmacological activities.In this paper,both single factor and response surface methods were used to optimize the extraction process of Baitiao-Codonopsis pilosula polysaccharides(BCPPS).According to the Fick's second law,the kinetics and thermodynamics of water extraction of crude polysaccharides from Baitiao-Codonopsis pilosula were studied.Subsequently,crude polysaccharides'purification,physicochemical properties,structure and antioxidant properties were studied.Finally,this study focus on the antiviral mechanism of crude polysaccharides.The main findings are as follows:(1)The optimal process conditions for extracting Baitiao-Codonopsis pilosula crude polysaccharides by response surface method were as follows:the ratio of material to liquid was 1:26.1 g/m L,the extraction temperature was 348.5 K,and the extraction time was 2.61 h.Under this condition,the yield of BCPPS was 9.719%.This optimal result has been verified that it is consistent with the actual extraction yield.The crude polysaccharide has protein content of 0.36%,sulfate content of 13.89%and uronic acid content of 0.35%.After the crude polysaccharide of Baitiao-Codonopsis pilosula was separated and purified by DEAE-52cellulose column and Sephadex G-100 gel column chromatography,one neutral polysaccharide fraction was obtained.The rheological properties of BCPPS showed that the viscosity increased with the increasing of polysaccharide concentration,and the viscosity of the solution decreased with the increasing of shear force.As the temperature of polysaccharide solution increased,the viscosity decreased;When the p H of the polysaccharide solution increases,the viscosity of the solution increases first and then decreases.In different salt solutions,as the molecular weight of the salt solution increases,the viscosity of the solution also increases.The results of FT-IR showed that the crude polysaccharide was?-pyranose.The crude polysaccharide showed the apperance like porous sponge under scanning electron microscopy.Thermogravimetric analysis revealed that there were two stages of water loss during the heating of the polysaccharide from 30°C to 800°C,and the whole process was accompanied by three exothermic reactions with enthalpy changes of 10.4 J/g,68.51 J/g and 15.297 J/g.(2)Water extraction process of BCPPS meets Fick's second law,according to the kinetic model's formula,the activation energy was increased firstly and then decreased with the increasing of material to liquid ratio,the half-life decreased with the increase of temperature,and the effective diffusion coefficient increased with increasing temperature.The thermodynamic calculation results showed that?Gm>0,entropy change?Sm<0,enthalpy change?Hm>0,indicating that the extraction process is a reverse reaction spontaneous process.And the elvated temperature makes the extraction process more chaotic and accompanied by endothermic reaction.(3)The in vitro antioxidant activity of BCPPS showed that the ability to scavenge DPPH free radicals increased with the increase of polysaccharide concentration,and the EC50 was0.6 mg/m L.The removal ability of the mass peroxide gradually decreased after it increased to0.4 mg/m L,and the EC50 was 0.2 mg/m L.The H₂O₂ scavenging capacity increased with the increase of the concentration,and the clearance rate was up to the highest,which was 70%,and the EC50 was 0.52 mg/m L.(4)The antiviral effects of ribavirin,Baitiao-Codonopsis pilosula polysaccharides and standard Codonopsis pilosula polysaccharides on the BVDV were studied.The relative expression of m RNA and protein in core and Npro of BVDV were determined.And the relative expression levels of m RNA and protein of inflammatory regulatory cytokines(IL-8,TNF-?and IFN-?)in host MDBK cells infected by BVDV.The results expressed that BCPPS had good preventive effect on BVDV virus.