Direct enzyme adsorption from an unclarified microbial feedstock using suspended bed chromatography

• Published in: Journal of Chromatography A 2003-03, Vol.989 (1), p.109-118
• Main Authors: Ling, Tau Chuan, Lyddiatt, Andrew, Carmichael, Ian, Purdom, Geoff, Hathi, Prit, Levison, Peter R
• Format: Article
• Language: English
• Subjects: Adsorption; Biomass; Chromatography, Ion Exchange - methods; Electrophoresis, Polyacrylamide Gel; Enzymes; Glyceraldehyde-3-phosphate dehydrogenase; Glyceraldehyde-3-Phosphate Dehydrogenases - chemistry
• ISSN: 0021-9673
• DOI: 10.1016/S0021-9673(02)01839-3

Suspended bed chromatography (SBC) is a new hybrid technique concomitantly benefiting from batch adsorption, the process advantages of an enclosed system, and its compatibility with established commercial chromatographic contactors and adsorbents. SBC was evaluated in the anion-exchange capture and chromatographic fractionation of native glyceraldehyde-3-phosphate dehydrogenase (G3PDH) from the complex mixture of molecular and particulate moieties that constitute wet-milled bakers’ yeast. Method scouting established operating conditions exploiting Whatman Express-Ion Exchanger Q at pH 7.5 and a disrupted biomass concentration equivalent to 3.5% (wet mass/v original intact cells). Partially purified G3PDH was recovered directly from the yeast disruptate in a scaled-down process developed at 1/756 process scale. This was used to establish operating parameters to facilitate process scale-up to exploit a 44 cm I.D. Millipore IsoPak column, 18 kg (swollen mass) of Express-Ion Q anion-exchange cellulose and 275 l of 3.5% (wet w/v) bakers’ yeast disruptate. The generic utility of SBC was demonstrated for direct product adsorption from feedstocks characterised by a modest content of bioparticulates (equivalent to