Structural Motifs of Wheat Straw Lignin Differ in Susceptibility to Degradation by the White-Rot Fungus Ceriporiopsis subvermispora

Abstract

The white-rot fungus Ceriporiopsis subvermispora delignifies plant biomass extensively and selectively and, therefore, has great biotechnological potential. We previously demonstrated that after 7 weeks of fungal growth on wheat straw 70% w/w of lignin was removed and established the underlying degradation mechanisms via selectively extracted diagnostic substructures. In this work, we fractionated the residual (more intact) lignin and comprehensively characterized the obtained isolates to determine the susceptibility of wheat straw lignin’s structural motifs to fungal degradation. Using 13C IS pyrolysis gas chromatography–mass spectrometry (py-GC-MS), heteronuclear single quantum coherence (HSQC) and 31P NMR spectroscopy, and size-exclusion chromatography (SEC) analyses, it was shown that β-O-4′ ethers and the more condensed phenylcoumarans and resinols were equally susceptible to fungal breakdown. Interestingly, for β-O-4′ ether substructures, marked cleavage preferences could be observed: β-O-4′-syringyl substructures were degraded more frequently than their β-O-4′-guaiacyl and β-O-4′-tricin analogues. Furthermore, diastereochemistry (threo > erythro) and γ-acylation (γ-OH > γ-acyl) influenced cleavage susceptibility. These results indicate that electron density of the 4′-O-coupled ring and local steric hindrance are important determinants of oxidative β-O-4′ ether degradation. Our findings provide novel insight into the delignification mechanisms of C. subvermispora and contribute to improving the valorization of lignocellulosic biomass.

Evidence for ligninolytic activity of the ascomycete fungus Podospora anserina

“…1), we would have expected to see more prominent changes in the structural characteristics of the residual lignin. For several white-rot fungal-treated samples we, and others, have previously observed an approximate doubling of total C α -oxidized moieties at comparable extents of delignification [18,28,29], and C. subvermispora showed a 16-fold increase in diketone markers [20]. These observations would suggest that different (or additional) degradation routes underlie delignification, as elaborated below.…”

“…Note that even though in HSQC NMR spectra C β -H β correlation peaks are resolved for various substructures of β-O-4′ aryl ether linkages [20,33], the hemicellulosic substrate presented an overlapping peak with the β-O-4′-S/G-S signal at δ C /δ H 85.97/3.96 ppm in this experimental setup. Therefore, we quantified the C α -H α correlation peaks instead, as previously described by del Río et al [34].…”

“…Constituent monosaccharides released after acid hydrolysis were determined in duplicate as previously reported by Englyst & Cummings with reported modifications [20,62].…”

“…Recently, we have demonstrated that quantitative pyrolysis-GC-MS analysis is exceptionally useful for mapping the extent of fungal plant biomass delignification as well as for unravelling the underlying ligninolysis mechanisms [18][19][20]. Importantly, this technique can highlight minor structural changes in the remaining lignin after short periods of fungal growth, an unconditional requirement when studying the structural changes induced by micro-organisms with a short growth cycle [19,20].…”

Elucidation of ligninolysis mechanism of a newly isolated white-rot basidiomycete Trametes hirsuta X-13

“…Although ligninolytic enzymes have been extensively identified and characterized in white-rot fungi, the degradation pathways of lignin largely remain unknown. The only demonstrated ligninolysis routes in white-rot fungi were C α –C β and β-O-4 cleavages based on the detection of the corresponding benzoic acid derivatives during the lignocellulosic biomass degradation [ 17 – 20 ]. Furthermore, knowledge of the degradation routes of lignin by white-rot basidiomycete fungi when used as the sole carbon source were less studied.…”