Anion Exchangers Prepared By Grafting Polymerization Of Cationic Monomers To Sepharose FF For Protein Adsorption And Chromatography

High binding capacity and uptake rate are desired for high performance ion exchange chromatography.For this purpose,we have respectively grafted dimethylamine ethyl methacrylate(DMAEMA),acrylamide(AAM),methyl acryloyl oxygen ethyl trimethyl ammonium chloride(DMAEMA-Q),dimethylamino propyl acrylamide(DMAPAA)and(3-acrylamide propyl)trimethyl ammonium chloride(DMAPAA-Q)onto Sepharose FF by atom transfer radical polymerization(ATRP).The ion-exchange chromatography resins with different grafting polymer chains were prepared.The basic physical properties of the resins and the adsorption properties of the protein were systematically studied.First,the DMAEMA-grafted resins and DMAEMA and AAM co-grafted resins were prepared.The results showd that uptake rate(D_e/D₀,defined as the ratio of effective diffusion coefficient(D_e)to free diffusivity of the protein(D₀))of bovine serum albumin(BSA)was significantly higher than traditional non-grafted resins and PEI grafted resins.Then,DMAPAA-grafted resins were prepared.The results showed that the adsorption capacity(q_m)and uptake rate(D_e/D₀)of the resins increased first and then decreased with the increase of IC value.When IC=458 mmol/L(FF-p DMAPAA458),the q_m and D_e/D₀ values reached the platform,which were 368 mg/m L and 2.45,respectively.Therefore,FF-p DMAPAA458 was selected to study the effect of ionic strength and p H on adsorption,as well as the chromatographic behavior of the resin.The q_m and D_e/D₀ of FF-p DMAPAA458 were much higher than other anion exchangers reported in the literature.When the salt concentration was as high as 100mmol/L or the p H was 6-9,the q_m and D_e/D₀ of FF-p DMAPAA458 remained above230 mg/m L and 2 respectively.Therefore,the dynamic binding capacity of FF-p DMAPAA458column in the range of salt concentration was significantly higher than that of Q Sepharose FF and two polymer graft anion exchange resins reported in the literature.Finally,four resins with similar IC values(FF-p DMAEMA,FF-p DMAPAA,FF-p DMAEMA-Q and FF-p DMAPAA-Q)were prepared.The results of BSA adsorption showed that the complete dissociation of the polymer chain at p H<7 and partial dissociation of the polymer chain at p H>7 were beneficial for protein adsorption capacity.In addition,the decrease of the dissociation degree and the increase of hydrophilicity of the polymer chain are beneficial to accelerate the uptake rate of the protein.This study shows that the high-performance protein chromatography can be prepared by ATRP method using monomers with high hydrophilicity,which lays a theoretical and experimental foundation for further development of high-performance chromatography resin.