Abstract: Immobilized lignin peroxidase has been investigated using a flow system in the steady state and by flow injection analysis (FIA). In the steady state, the extreme sensitivity of the enzyme towards inactivation by H2O2 resulted in a stable response only in the presence of saturating levels of organic substrate and at very low (10 µM) peroxide concentrations. By contrast, the low contact time during FIA led to a stable response to injections of 100 µM H2O2. At higher peroxide concentrations a reproducible inactivation was observed, allowing a study of factors affecting both activity and stability. Lignin peroxidase substrates that undergo at least semi-reversible oxidation/reduction, including high-molecular-weight lignin fractions, could be detected by electrochemical reduction of the oxidation products. With this detection system it was possible to demonstrate the role of veratryl alcohol as mediator. This mediated oxidation of lignin functioned only when all components were present simultaneously, and was not observed when lignin was separated from the site of veratryl alcohol oxidation.

Abstract: A simple membrane reactor housing with negligible dead volume was constructed to take commercially available membranes. Three immobilized-enzyme systems were investigated: glucose oxidase, glutaminase and laccase for the determination of glucose, glutamine and phenolics, respectively. The response of reactors with Zetaffinity AM membranes was comparable to that of packed bed reactors containing controlled-pore glass or Biosynth VA Epoxy, but the membrane reactors had less pressure build-up and thus increased lifetime compared with the packed beds (which were prone to clogging) in continuous monitoring. Their lifetime was determined by the loss of enzyme activity. Membrane-type reactors were prepared using a commercially available support. In three test systems, involving glucose oxidase, glutaminase and laccase, they compared favorably with packed-bed reactors in a flow injection set-up. Although similar responses were observed for all reactor types, the membranes are easier to handle than microparticles. When making continuous inline measurements of bioprocesses at high sampling frequencies, the lifetimes of packed beds of controlled pore glass were determined by pressure build-up within the reactor. When replaced with membrane reactors, no clogging was observed, the lifetimes being determined by loss of enzyme activity.