Fungal Enzymes for Saccharification of Gamma-Valerolactone-Pretreated White Birch Wood: Optimization of the Production of Talaromyces amestolkiae Cellulolytic Cocktail

Abstract

Lignocellulosic biomass, the most abundant natural resource on earth, can be used for cellulosic ethanol production but requires a pretreatment to improve enzyme access to the polymeric sugars while obtaining value from the other components. γ-Valerolactone (GVL) is a promising candidate for biomass pretreatment since it is renewable and bio-based. In the present work, the effect of a pretreatment based on GVL on the enzymatic saccharification of white birch was evaluated at a laboratory scale and the importance of the washing procedure for the subsequent saccharification was demonstrated. Both the saccharification yield and the production of cellulosic ethanol were higher using a noncommercial enzyme crude from Talaromyces amestolkiae than with the commercial cocktail Cellic CTec2 from Novozymes. Furthermore, the production of extracellular cellulases by T. amestolkiae has been optimized in 2 L bioreactors, with improvements ranging from 40% to 75%. Finally, it was corroborated by isoelectric focus that optimization of cellulase secretion by T. amestolkiae did not affect the pattern production of the main β-glucosidases and endoglucanases secreted by this fungus.

Keywords: bioethanol; cellulases; fungi; green solvent; lignocellulosic biomass.

FIGURE 1

Saccharification (72 h) of untreated and GVL‐pretreated white birch wood with the commercial cocktail Cellic CTec2. The pretreatments applied to the samples were (A) GVL fractionation (there is no bar because the saccharification of this material was not successfully); (B)–(D) as treated in (A), followed by different washing steps: (B) 1 × H₂O; (C) 15 × H₂O; (D) 15 × H₂O + screening (800 mesh).

FIGURE 2

Time course of the saccharification yields of white birch wood pre‐treated with GVL and washed with Hot GVL and then with H₂O, using Cellic CTec2 or a T. amestolkiae enzymatic crude as saccharification cocktails.

FIGURE 3

Time evolution of β‐glucosidase, endoglucanase, and FPU activities secreted by T. amestolkiae in 2 L bioreactor. (A) Before optimization (initial pH 4.5, 200 rpm, Mandels broth with 1% Avicel and 0.1% bacto peptone); (B) After optimization (pH controlled at 4.5, 400 rpm, Mandels broth modified with 2% Avicel and 0.4% CSS).