THE EFFECT OF PECTINASE ON THE BUBBLE FRACTIONATION OF INVERTASE FROMALPHA-AMYLASE

Fermentation broth normally contains many extracellular enzymes of ind ustrial interest. To separate such enzymes on-line could be useful in reducing the cost of recovery as well as in keeping their yield at a m aximum level by minimizing enzyme degradation from broth proteases (ei ther the desired enzymes or the proteases could be removed selectively or both removed together and then separated). Several large-scale sep aration methods are candidates for such on-line recovery such as ultra filtration, precipitation, and two-phase partitioning. Another promisi ng technique for on-line recovery is adsorptive bubble fractionation, the subject of this study. Bubble fractionation, like ultrafiltration, does not require contaminating additives and can complement ultrafilt ration by preconcentrating the enzymes using the gases normally presen t in a fermentation process. A mixture of enzymes in an aqueous bubble solution can, in principle, be separated by adjusting the pH of that solution to the isoelectric point (pI) of each enzyme as long as the e nzymes have different pIs. The model system investigated here is compr ised of three enzyme separations and the problem is posed as the effec t of pectinase (a charged enzyme) on the bubble fractionation of inver tase (a relatively hydrophilic enzyme) from alpha-amylase (a relativel y hydrophobic enzyme). The primary environmental variable studied, the refore, is the pH in the batch bubble fractionation column. Air was us ed as the carrier gas. This prototype mixture exemplifies an aerobic f ungal fermentation process for producing enzymes. The enzyme concentra tion here is measured as total protein concentration by the Coomassie Blue (Bradford) solution method (1), both as a function of time and co lumn position for each batch run. Since, from a previous study (2), it was found that invertase and alpha-amylase in a two-enzyme system can be partially separated in favor of one vs the other at two different pHs (pH 5.0 and 9.0) with significant separation ratios, emphasis is p laced on the effect of pectinase at these pHs. In this study, the addi tion of pectinase reduced the total separation ratio of the alpha-amyl ase-invertase mixture at both pHs.