Size Exclusion PEGylation Reaction Chromatography Modelling

Abstract

Size exclusion PEGylation reaction chromatography was investigated using a model developed by Fee (2005). Column dispersion was neglected and the PEGylation reaction was modelled as second order. The model allowed up to four PEG groups to be attached to a protein and accounted for succinic acid hydrolysis from activated PEG. The model was adapted to simulate a-lactalbumin PEGylation and succinic acid hydrolysis from activated PEG in a batch stirred tank so rate parameters from stirred tank kinetic experiments could be obtained and the model verified. The model was solved using finite differences and simulations run in Matlab. The effect of reaction parameters such as timing, length and concentration of PEG and protein injection, reaction rates, and model resolution on model simulation results was explored. In the size exclusion PEGylation simulations it was found that increasing protein concentration increased MonoPEG concentrations and increased the ratio of MonoPEG to starting protein feed concentration. Increasing PEG pulse length and starting PEG concentration initially increased MonoPEG concentration and product ratio until all protein had been PEGylated at which point MonoPEG concentration the product ratio levelled out. Increasing PEG hydrolysis rates did not affect the amount of MonoPEG produced but reduced the activated PEG concentration and increased succinic acid concentration. Optimal conditions for producing MonoPEG were found to be equal concentrations of PEG and protein, with the PEG injection length twice as long as the protein injection, and the PEG injection done immediately after the protein injection.