Ensiling pretreatment fortified with laccase and microbial inoculants enhances biomass preservation and bioethanol production of alfalfa stems

Abstract

This study investigated the potential of ensiling pretreatment fortified with laccase and a lactic acid bacteria (LAB) inoculant on improving the utilization of alfalfa stems for bioethanol production. The alfalfa stems were ensiled with no additives (Con), 0.04 % laccase (LA), a LAB inoculant containing Pediococcus pentosaceus at 1 × 10⁶ fresh weight (FW) and Pediococcus acidilactici at 3 × 10⁵ cfu/g FW (PP), and a combination of LA and PP (LAP) for 120 days. By reshaping the bacterial community structure of alfalfa stem silages toward a higher abundance of Lactobacillus, the addition of laccase and LAB inoculant either alone or in combination facilitated lactic acid fermentation to reduce fermentation losses, as evidenced by low concentrations of ammonia nitrogen (53.7 to 68.9 g/kg total nitrogen) and ethanol (2.63 to 3.55 g/kg dry matter). All additive treatments increased lignocellulose degradation and soluble sugars concentrations of alfalfa stem silages. Due to delignification and polyphenol removal, glucan and xylan conversion (70.3 % vs. 35.7 % and 51.6 % vs. 27.9 %, respectively) and ethanol conversion efficiency (53.9 % vs. 26.4 %) of alfalfa stems were greatly increased by ensiling fortified with LA versus Con, and these variables (79.8 % for glucan, 58.7 % for xylan, and 60.1 % for ethanol conversion efficiency) were further enhanced with a synergistic effect of LA and PP fortification. The spearman correlation analysis revealed that bioethanol fermentation of silage biomass was closely related to ensiling parameters and total phenols. In conclusion, ensiling pretreatment with LA and PP combination offered a feasible way to efficient utilization of alfalfa stems for bioethanol production.

Keywords: Alfalfa stems; Bioethanol production; Biological additives; Biomass preservation; Silage.