The flocculation of bacteria using cationic synthetic flocculants and chitosan

Abstract

Bacteria are flocculated with high molecular weight cationic synthetic flocculants and chitosan. High charge density polymers are the most effective of the synthetic flocculants. Only chitosan is effective in flocculating the E. coliand B. subtilis cultures in complex broth. The difference in effectiveness between the synthetic flocculants and chitosan for flocculating E. coli, B. subtifisand Z. mobilismay be attributed to hydrogen bonding between the polysaccharide flocculant and cell surface polymers in addition to electrostatic interactions, and, in complex media, complexation of synthetic polymers with anionic polyelectrolytes.

PDADMAC flocculation of Chinese hamster ovary cells: Enabling a centrifuge-less harvest process for monoclonal antibodies

“…4,[7][8][9][10][11][12] Also, product recovery for a harvest method utilizing flocculation will be dependent on the volume of cell culture fluid (CCF) recovered and any potential interaction of the product with the flocculent (flocculated cell/cellular debris particle) or the polycationic polymer. The parameters investigated in the development of a CHO cell culture PDADMAC flocculation harvest method are discussed in the following sections.…”

“…This simple flocculant dosing method does not require pre-harvest testing for flocculant dose determination as described by other flocculation methods. 7,9,10,12,13 The flocculant dose was affected by the submicron particle level generated during the cell culture and harvest process for optimum flocculation performance. Increasing the flocculant dose resulted in an optimum flocculation performance for cell broth containing both typical and elevated levels of submicron cellular debris (Fig.…”

“…4,[7][8][9][10][11][12] Since PDADMAC binds to the negatively-charged cells (both viable and non-viable cells) and cellular debris in the cell culture broth, a high cell density cell culture process requires a larger flocculation dose per volume compared to a lower cell density culture process to achieve the optimum flocculation performance (clarity, product recovery, and settling period). To account for cell culture process variability from process to process and run-to-run, a dosing regimen based on the cellular mass is desired.…”

“…Polycationic polymers bind the negatively-charged cells and cellular debris leading to the formation of larger particles that easily settled out or removed by centrifugation. A number of polycationic polymers have been used to flocculate cells, including polyethyleneimine, poly (diallyldimethylammonium chloride) or PDADMAC, chitosan, polycationic polyacrylamides, and partially benzylated poly(allylamine), e.g., smart Polymer E. [7][8][9][10][11][12][13] PDADMAC has several advantages over the other polycationic polymers that have been used to clarify cell broth. In contrast to some of the other polycationic polymers, PDADMAC is supplied as a chloride salt solution and requires no sample conditioning prior to use.…”

“…12 In comparison, chitosan and the polycationic polyacrylamides flocculate is explained by the bridging model which involves a multitude of interactions that include electrostatic, hydrophobic, or hydrogen bonding. 10,12 Flocculent particles produced by the patch/neutralization flocculation model are more resistant to shear fragmentation compared to flocculent particles produced by bridging. 12 Shear resistant flocculent particles are desirable from a commercial manufacturing standpoint because it is known that shear forces due to high mixing rates or prolonged mixing during the addition of the flocculant into the cell culture broth may inhibit flocculent growth or fragment the flocculent producing smaller and slower settling flocculent particles.…”