Bioconversion of starches into maltotetraose using Pseudomonas stutzeri maltotetraohydrolase in a membrane recycle bioreactor: Effect of multiple enzyme systems and mass balance study

A simplified single-step method involving simultaneous production and purification of maltotetraose (G₄) by employing ultrafiltration (UF) membranes was previously proposed. The addition of a pretreatment step using pullulanase and then the G₄-amylase was expected to increase the yield of G₄. The single-enzyme system, however, showed 0.42 g higher total product output than the successive dual-enzyme system throughout 6 h reaction. The G₄ yield using the successive dual-enzyme system could be improved after removing the unwanted side product with UF. Experiments were conducted with membranes of larger pore size, but this did not significantly increase the total product output. The membrane unit with a molecular weight cutoff of 1,000 was the most appropriate membrane pore size for the G₄-exo-α-amylase membrane recycle bioreactor system. The total amount of substrate fouled in the membrane during a 6-h reaction was estimated as 69 mg glucose equivalent when substrate concentration was 0.25% (w/v). The mass balance equation indicated that the percent conversion of soluble starch to G₄ at steady state was 65%.