Microplate enzyme assay for screening lipoxygenases to degrade wood extractives

Wood extractives are known to cause a range of problems in pulp and paper manufacturing. Soybean lipoxygenase enzyme treatments to degrade wood extractives present in thermomechanical pulp (TMP) samples resulted in a 15% reduction in total extractives. However, commercially available lipoxygenase is currently not cost effective for this application. A 96-well UV microplate assay was developed to screen enzymes from fungal species for a more cost-effective alternative lipoxygenase. The substrate used for this assay is linoleic acid, a predominant fatty acid in wood extractives. The enzyme activity and reaction kinetics determined by this microplate assay were compared to those obtained from a conventional bench scale assay. A number of hydrolytic enzymes, and other types of oxidases were also tested by this protocol to examine the specificity of the assay. The results from this study show that the developed microplate assay can provide an inexpensive method for accelerated screening of a large number of enzymes to identify potential oxidative enzymes with specific action in degrading wood extractives.