A novel technique for the measurement of disruption in high-pressure homogenization: Studies on E. coli containing recombinant inclusion bodies.
The high-pressure homogenization of Escherichia coli, strain JM101, containing inclusion bodies of recombinant porcine somatotropin was investigated. A novel technique employing an analytical disc centrifuge was used to monitor the disruption. This a direct technique which measures cell disintegration rather than soluble protein release. The technique is particularly suited to measurements where the disruption approaches 100%. The disk centrifuge provides a size distribution of the homogenate, and furnishes evidence for the preferential disruption of larger cells. For E. coli containing inclusion bodies, and increase in the cell feed concentration from 145 g/L (wet weight) to 330 g/L resulted is poorer homogenization. Poorer disruption was also obtained by lowering the feed temperature from 20 degrees C to 5 degrees C. Only slight variations in performance were obtained by increasing the feed pH from 7.5 to 9.0 or by storing the feed at 4 degrees C for 24 h prior to disruption. Comparison with uninduced E. coli strain JM101, showed that the disruption obtained is higher for bacteria containing a recombinant inclusion body.
Middelberg
A P
AP
Department of Chemical Engineering, The University of Adelaide, Adelaide, Australia.
O'Neill
B K
BK
L Bogle
I D
ID
Snoswell
M A
MA