Capture of bacteria from fermentation broth by body feed filtration: a solved problem?
The direct capture of bacteria produced in high cell density fermentation by filtration is not possible once the milliliter-scale has been surpassed. Filtration in the presence of a filter aid (body feed filtration) constitutes a putative and scalable alternative, but only if conditions proposed by industry for large-scale filtration processes, namely, flow rates (for aqueous solutions) in the range of 500-1,500 L/(m(2) x h) and a filter aid concentration of <or=20 g/L, can be met. To this end several filter aids (grades of cellulose, diatomaceous earth, mixes thereof) were tested, albeit with no success. Capture and retention of bacteria was low (not possible in the case of cellulose), and the produced filter cakes were unstable and of low permeability. In the case of the diatomaceous earth close to 100% bacterial capture was possible, but only with filter aid concentrations that were 1 order of magnitude above the proposed limit. By using flocculating agents (Fe(3+)/Al(3+), PEI) either alone or in combination with diatomaceous earth and also the use of positively charged cellulose, bacteria capture could be achieved from several liters of cell suspension (OD(600) <or= 15) in filtration experiments adhering to the industrial recommendations. However, the bacteria were only weakly retained in such cakes and easily released by sudden pressure pulses. Satisfactory results were only obtained by filtration of the bacteria suspension at slightly alkaline pH (8.0, 50 mM Tris buffer) after treatment with sodium-activated bentonite (5 g/L) and PEI (50 mg/L) in the presence of 250 mM NaCl. In such cases 100% of the bacteria could reproducibly be captured in a filtration using 20 g/L of diatomaceous earth as filter aid. The thus produced filter cakes strongly retained the bacteria and showed good filtration performance. The procedure is at present limited to E. coli culture with biomass contents of OD(600) <or= 15. Cultures with higher OD(600) have to be diluted prior to filtration.
O'Mahony
Kevin
K
Laboratory of Chemical Biotechnology, Swiss Federal Institute of Technology, Lausanne, Switzerland.
Freitag
Ruth
R
Dhote
Bernhard
B
Hilbrig
Frank
F
Müller
Patrick
P
Schumacher
Ivo
I
D001546:Bentonite; D002482:Cellulose; D003432:Cross-Linking Reagents / Q000737:chemistry; D003470:Culture Media; D004926:Escherichia coli / Q000302:isolation & purification*; D005285:Fermentation; D005374:Filtration / Q000295:instrumentation* / Q000379:methods*; D006863:Hydrogen-Ion Concentration; D007097:Imines; D011095:Polyethylenes; D012965:Sodium Chloride; D013696:Temperature
D003432:Cross-Linking Reagents; D003470:Culture Media; D007097:Imines; D011095:Polyethylenes; C505405:poly(ethylene imine); D001546:Bentonite; D012965:Sodium Chloride; D002482:Cellulose