Brewer’s spent grain (BSG) constitutes 85% of brewing byproducts and 30% of initial malt. In 2021, BSG production reached approximately 37.2 million tons. Owing to its high moisture and nutritional content, BSG is prone to biological deterioration, causing environmental issues when discarded as waste. It has limited application as low-quality animal feed owing to its high fiber and low protein content, making landfilling the primary disposal method. However, BSG's abundance of starch, cellulose, hemicellulose, lignin, and protein make it ideal for value-added product generation in a biorefinery. The aim of this research was to investigate BSG fractionation and identify valuable products from each fraction, ultimately establishing a BSG-based biorefinery for sustainable valorization. 

Fungal cultivation, anaerobic digestion, hydrothermal, and organosolv pretreatments were employed to establish a BSG-based biorefinery. Edible filamentous fungi (Aspergillus oryzae, Neurospora intermedia, and Rhizopus delemar) were cultivated on crude BSG to produce food and feed-grade biomass. Fungal growth increased the protein content of the BSG by up to 47%. However, entangled solids with fungal filaments negatively affected product digestibility, limiting its incorporation in food and feed. This problem was resolved by recovering a solid-free, starch- and protein-rich stream from BSG via hydrothermal pretreatment. 

Hydrothermal pretreatment effectively separated BSG's starch and protein components, with solubilizations reaching 82% and 48% of the initial content, respectively. Fungal assimilation of the liquid stream produced pure, high-protein biomass and high ethanol yield. However, most of the BSG cellulose and lignin remained in the solid fraction. Organosolv pretreatment was applied to further separate BSG polymers into valorizable fractions efficiently, yielding a cellulose-rich solid stream, polysaccharide-rich organosolv liquor, and high-purity lignin (~95%). This pure lignin product can enhance the biorefinery’s economy and be sold or converted into platform chemicals. 

Direct fungal cultivation on cellulose-rich pulp and liquor fractions from organosolv revealed that the liquor fraction was suitable for producing pure, high-protein fungal biomass, while the pulp fraction required further processing. Moreover, anaerobic digestion was employed to produce a diverse array of products improving the product flexibility of the biorefinery. Organosolv liquor produced biohydrogen and volatile fatty acids (VFAs) without methanogen inhibition, while BSG and BSG organosolv solid fractions generated biogas. Inhibiting methanogens shifted the BSG process towards VFAs production, while organosolv solid fractions showed limited potential for VFAs generation. 

These results illustrate that BSG can serve as the foundation for a multi-product biorefinery that generates food-grade fungal biomass and valuable co-products, including high-purity lignin, bioethanol, biogas, biohydrogen, and VFAs. This flexibility allows the biorefinery to adapt to market changes and ensure its economic viability.