The selective lignin-degrading white-rot fungi are regarded to be the best lignin degraders and have been widely used for reducing the saccharification recalcitrance of lignocellulose. However, the biological delignification and conversion of lignocellulose in biorefinery is still limited. It is necessary to develop novel and more efficient bio-delignification systems. Physisporinus vitreus relies on a new versatile peroxidase (VP)-based delignification strategy to remove enzymatic recalcitrance of corn stover efficiently, so that saccharification of corn stover was significantly enhanced to 349.1 mg/g biomass (yield of glucose) and 91.5% (hydrolysis yield of cellulose) at 28 days, as high as levels reached by thermochemical treatment. Analysis of the lignin structure using pyrolysis–gas chromatography–mass spectrometry (Py–GC/MS) showed that the total abundance of lignin-derived compounds decreased by 54.0% and revealed a notable demethylation during lignin degradation by P. vitreus. Monomeric and dimeric lignin model compounds were used to confirm the ligninolytic capabilities of extracellular ligninases secreted by P. vitreus. The laccase (Lac) from P. vitreus could not oxidize nonphenolic lignin compounds and polymerized β-O-4 and 5-5′ dimers to precipitate which had a negative effect on the enzymatic hydrolysis of corn stover in vitro. However, the VP from P. vitreus could oxidize both phenolic and nonphenolic lignin model compounds as well as break the β-O-4 and 5-5′ dimers into monomeric compounds, which were measured by high-performance liquid chromatography–electrospray ionization–mass spectrometry (LC–ESI–MS). Moreover, we showed that addition of purified VP in vitro improved the enzymatic hydrolysis of corn stover by 14.1%. From the highly efficient system of enzymatic recalcitrance removal by new white-rot fungus, we identified a new delignification strategy based on VP which could oxidize both phenolic and nonphenolic lignin units and break different linkages in lignin. In addition, this is the first evidence that VP could break 5-5′ linkage efficiently in vitro. Moreover, VP improved the enzymatic hydrolysis of corn stover in vitro. The remarkable lignin-degradative potential makes VP attractive for biotechnological applications.

Background: Irpex lacteus, a versatile lignin-degrading fungus with various extracellular enzymes, has been widely used for biological pretreatment. However, most studies have focused on the change of substrate structure after biological pretreatment, and the effect of these changes on the enzymatic hydrolysis, but the effect of byproducts from biological pretreatment process on subsequent enzymatic hydrolysis is not well understood. Methods: We developed a biological pretreatment process with I. lacteus that can produce stimulatory byproducts that enhance the enzymatic hydrolysis of cornstalks. Results: The maximum hydrolysis yield of glucan (82%) was obtained after pretreatment for 28 days. The maximum reducing sugar yield decreased from 313.5 to 200.1 mg/g raw cornstalks after water-soluble byproducts of biological pretreatment were removed from pretreated cornstalks. The effect of byproducts on enzymatic hydrolysis was also investigated. We found that the hydrolysis efficiency of commercial cellulase preparation on cornstalks could be improved by water extracts from bio-pretreated cornstalks with hydrolytic enzyme activity and iron-reducing activity. Conclusion: The key finding suggested that byproducts from biological pretreatment play important roles in enhancing downstream hydrolysis, which might be attributable to hydrolytic enzymes and iron-reducing compounds produced by I. lacteus.