Abstract
Background5-Hydroxymethylfurfural (HMF) is a highly valuable platform chemical that can be obtained from plant biomass carbohydrates. HMF can be oxidized to 2,5-furandicarboxylic acid (FDCA), which is used as a renewable substitute for the petroleum-based terephthalic acid in polymer production.ResultsAryl-alcohol oxidase (AAO) from the white-rot fungus Pleurotus eryngii is able to oxidize HMF and its derivative 2,5-diformylfuran (DFF) producing formylfurancarboxylic acid (FFCA) thanks to its activity on benzylic alcohols and hydrated aldehydes. Here, we report the ability of AAO to produce FDCA from FFCA, opening up the possibility of full oxidation of HMF by this model enzyme. During HMF reactions, an inhibitory effect of the H2O2 produced in the first two oxidation steps was found to be the cause of the lack of AAO activity on FFCA. In situ monitoring of the whole reaction by 1H-NMR confirmed the absence of any unstable dead-end products, undetected in the HPLC analyses, that could be responsible for the incomplete conversion. The deleterious effect of H2O2 was confirmed by successful HMF conversion into FDCA when the AAO reaction was carried out in the presence of catalase. On the other hand, no H2O2 formation was detected during the slow FFCA conversion by AAO in the absence of catalase, in contrast to typical oxidase reaction with HMF and DFF, suggesting an alternative mechanism as reported in some reactions of related flavo-oxidases. Moreover, several active-site AAO variants that yield nearly complete conversion in shorter reaction times than the wild-type enzyme have been identified.ConclusionsThe use of catalase to remove H2O2 from the reaction mixture leads to 99% conversion of HMF into FDCA by AAO and several improved variants, although the mechanism of peroxide inhibition of the AAO action on the aldehyde group of FFCA is not fully understood.
Highlights
Interest in the production of new chemicals and materials from renewable resources has exponentially grown in the last years with biomass-derived compounds as a future alternative to fossil-based compounds
Analysis of HMF oxidation by Aryl-alcohol oxidase (AAO) in the NMR tube Taking the previous results into account, we looked into the possible formation of a dead-end formylfurancarboxylic acid (FFCA) product, which might be unstable under the HPLC analysis conditions
The addition of catalase enables the complete oxidation of HMF to furandicarboxylic acid (FDCA) by P. eryngii AAO, as reported for other reactions catalyzed by glucose-methanol-choline oxidase/dehydrogenase (GMC) oxidases [26, 27]
Summary
Interest in the production of new chemicals and materials from renewable resources has exponentially grown in the last years with biomass-derived compounds as a future alternative to fossil-based compounds. FDCA production from lignocellulose biomass involves first the production of 5-hydroxymethylfurfural (HMF). This compound is obtained by dehydration of monosaccharides (generally fructose) that can originate from cellulose hydrolysis, followed by glucose isomerization [5,6,7]. The chemical methods to obtain FDCA from HMF require high temperature, high pressure, metal catalysts, and organic solvents, which render the process polluting and expensive. For this reason, enzymatic conversion appears a suitable alternative, since enzymes are selective and act under mild conditions [8, 9]
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