One-step Purification Method of MC-LR Fab Fragment by Gel Filtration Chromatography and Preliminary Study on Its Adsorption Mechanism

For the efficient preparation of MC-LR Fab fragment,the purified antibody was digested with papain and the purification method was explored based on a special phenomenon that the Fab fragment could bind to the gel filtration chromatography(GFC)medium. The effects of ionic strength and pH in mobile phase on the behavior of the Fab fragment in GFC column were investigated through the single factor experiments,and the binding mechanism was considered from these experiments and the properties of the Fab fragment. The results showed that,the purity of IgG purified by protein G was more than 90%,and the digestion products contained the Fab fragment,the Fc fragment and a small amount of undigested IgG. After the one-step purification,the purity of the Fab fragment was 94.5% with the recovery rate of 51% and the inhibition rate of MC-LR reached 94% for the Fab fragment. The pI of the Fab fragment was 6.02 and it had the strong hydrophobicity on the surface which caused the aggregation. Hydrophobic interaction was the reason why the Fab fragment bound to the GFC medium. Decreasing the ionic strength of mobile phase which could decrease the hydrophobic interaction and increasing pH which could increase the ion exclusion could eliminate the interaction,then the efficient preparation method of the Fab fragment was developed.