Enzymatic Oxidation of Ca-Lignosulfonate and Kraft Lignin in Different Lignin-Laccase-Mediator-Systems and MDF Production

Abstract

Laccase-mediator-oxidized lignin offers replacement for conventional chemical binders to produce fiberboards. Compared to the previously reported laccase–mediator system (LMS), a lignin-laccase-mediator-system (LLMS) has an advantage in that it requires much shorter fiber-enzyme incubation time due to significantly increased redox reactions. However, the cost of regularly applying laccase on an industrial scale is currently too high. We have employed CcLcc5 from cultures of the basidiomycete Coprinopsis cinerea as a novel basi-laccase (a CAZy subfamily AA1_1 laccase) in medium-density fiberboard (MDF) production, in comparison to the commercial formulation Novozym 51003 with recombinantly produced asco-laccase MtL (a CAZy subfamily AA1_3 laccase-like multicopper oxidase from the ascomycete Myceliophthora thermophila). With the best-performing natural mediator 2,6-dimethoxyphenol (DMP), unpurified CcLcc5 was almost as good as formulated Novozym 51003 in increasing the molecular weight (MW) of the technical lignins tested, the hydrophilic high-MW Ca-lignosulfonate and the hydrophobic low-MW kraft lignin (Indulin AT). Oxygen consumption rates of the two distantly related, poorly conserved enzymes (31% sequence identity) with different mediators and lignosulfonate were also comparable, but Indulin AT significantly reduced the oxidative activity of Novozym 51003 unlike CcLcc5, regardless of the mediator used, either DMP or guaiacol. Oxygen uptake by both laccases was much faster with both technical lignins with DMP than with guaiacol. In case of lignosulfonate and DMP, 20–30 min of incubation was sufficient for full oxygen consumption, which fits in well in time with the usual binder application steps in industrial MDF production processes. LLMS-bonded MDF was thus produced on a pilot-plant scale with either crude CcLcc5 or Novozym 51003 at reduced enzyme levels of 5 kU/kg absolutely dry wood fiber with lignosulfonate and mediator DMP. Boards produced with CcLcc5 were comparably good as those made with Novozym 51003. Boards reached nearly standard specifications in internal bond strength (IB) and modulus of rupture (MOR), while thickness swelling (TS) was less good based on the hydrophilic character of lignosulfonate. LLMS-bonded MDF with Indulin AT and DMP performed better in TS but showed reduced IB and MOR values.

Enhanced Enzyme Activity of Thermoalkaliphilic Laccase (CtLac) of Caldalkalibacillus thermarum Strain TA2.A1 by Random and Site-directed Mutagenesis and Its Application to Lignin Degradation

“…For example, it could be applied to develop a laccase-mediator-system (LMS) to improve the low redox potential of laccase itself [28][29][30]. Euring et al evaluated the performance of different natural mediators in enzymatic oxidation of lignin by comparing oxygen consumption rates in different LMS [29]. It found that the mediator, 2,6-DMP, signi cantly enhanced the oxidation of lignosulfonate and Indulin AT with a more decrease in oxygen saturation.…”

“…Therefore, comparing oxygen consumption might be an obvious criterion for evaluating enzyme activity. For example, it could be applied to develop a laccase-mediator-system (LMS) to improve the low redox potential of laccase itself [28][29][30]. Euring et al evaluated the performance of different natural mediators in enzymatic oxidation of lignin by comparing oxygen consumption rates in different LMS [29].…”

Improvement of thermoalkaliphilic laccase (CtLac) by a directed evolution and application to lignin degradation

“…Therefore, comparing oxygen consumption might be an obvious criterion for evaluating enzyme activity. For example, the technique could be applied to develop a laccase-mediator-system (LMS) to improve the catalytic efficiency of the enzyme (Kirsch et al 2015 ; Euring et al 2022 ; Hilgers et al 2018 ). On the other hand, Euring et al ( 2022 ) evaluated the performance of different natural mediators in the enzymatic oxidation of lignin by comparing oxygen consumption rates in different LMS and found that the 2,6-DMP mediator significantly enhanced the oxidation of lignosulfonate and Indulin AT with a big decrease in oxygen saturation.…”

“…For example, the technique could be applied to develop a laccase-mediator-system (LMS) to improve the catalytic efficiency of the enzyme (Kirsch et al 2015 ; Euring et al 2022 ; Hilgers et al 2018 ). On the other hand, Euring et al ( 2022 ) evaluated the performance of different natural mediators in the enzymatic oxidation of lignin by comparing oxygen consumption rates in different LMS and found that the 2,6-DMP mediator significantly enhanced the oxidation of lignosulfonate and Indulin AT with a big decrease in oxygen saturation. On the other hand, the oxygen consumption of LMS with the help of ABTS was faster than that of 1-hydroxy-benzotriazole (HBT) supported-LMS in the oxidation of GGGE (Hilgers et al 2018 ).…”

Effect of the Adhesive System on the Properties of Fiberboard Panels Bonded with Hydrolysis Lignin and Phenol-Formaldehyde Resin

“…Depending on the method by which they are obtained, the residual lignin products, i.e., technical lignins, are sulfate (Kraft) lignin, sulfite lignin (lignosulfonate), organosolv lignin and hydrolysis lignin. Although Kraft lignin is the most widespread globally, much of it is burned in the factories where it is obtained, which regenerates some of the chemical reagents used and produces heat and energy [30,31]. The main drawbacks for using lignosulfonates in wood adhesive formulations are related to the higher number of impurities, e.g., high sulfur and ash content, compared to the Kraft lignin, and the deteriorated hydrophobic properties of the wood-based panels, so it is recommended to be used in combination with synthetic binders with or without additional cross-linking [32][33][34][35][36][37][38][39][40][41][42].…”