Abhiram Arunkumar
(Coadvised by Prof. Mark R. Etzel)
Analysis and Design of charged ultrafiltration membranes for robust fractionation of therapeutic proteins
Fractionation of dairy proteins is currently performed using column chromatography. Chromatography offers excellent selectivity for the fractionation of dairy proteins and offers many modes. However, the sophisticated chromatography systems are expensive, not environmentally friendly and are unfamiliar to production personnel in the dairy industry. On the other hand, membrane ultrafiltration (UF) does not require extensive quantities of salts and buffers, and offers a high-throughput technology for downstream bioprocessing. Membranes offer easy regenerability, eliminating thus, the troublesome aspects of chromatography. However, conventional UF offers very low selectivity for protein fractionation. UF has traditionally been used in the dairy and biopharmaceutical industry to concentrate streams containing proteins but not fractionate them.
The purpose of this research is to make ultrafiltration selective for protein fractionation by basing the separation on both the hydrodynamic (steric) and electrostatic interactions and evaluate the use of membrane cascades for fractionating milk serum proteins. By adding a positive charge to the membrane and adjusting the solution pH, it is possible to selectively retain proteins that are similarly charged as the membrane and allow permeation of uncharged and smaller proteins. It was discovered that positively charged UF membranes with a pore size rating of 300 kDa can be used to fractionate proteins that are about 20 times smaller such as alphalactalbumin (ALA, 14.4 kDa) and betalactoglobulin (BLG, 18.6 kDa). Operating the ultrafiltration in the tangential-flow filtration (TFF) mode reduces membrane fouling and allows for easy scale-up.
Fabrication of novel positively charged TFF membranes, charged with salt-tolerant polycations to provide enhanced retention properties to the membrane will be examined. These membranes will be used for ALA fractionation from milk-serum microfiltration permeate (MFP). Apart from ALA, glycomacropeptide (GMP), which is used as the primary protein source for patients suffering from phenylketoneuria, will be fractionated at high throughputs and purities from cheese whey.
Thermodynamic and transport aspects of hindered transport of a charged solute in fluid filled charged cylindrical pores will be examined and the data obtained from experiments would be used to calculate the parameters of this model. This would help in understanding protein transport through charge modified filtration systems better, since most of these studies have been performed only for ideal systems of proteins in buffer, as opposed to a real bioprocess fluid.
Development of high performance charged ultrafiltration membranes and staged membrane processes would make ultrafiltration attractive for fractionation and bioprocessing of therapeutic macromolecules, giving products of chromatographic purity without chromatography.