A series of experiments was conducted to improve the degradation of winery biomass waste by a fungal mixture comprising cultures of Trichoderma herzianum, Aspergillus niger, Penicillium chrysogenum and Penicillium citrinum in submerged and solid state (SSF) fermentation processes. Lignin degradation, cellulase, xylanase and β-glucosidase activities were analysed to quantify degradation. In the optimization process, the highest activities observed were 43.7 U/mL, 1414 U/mL and 181.4 U/mL for cellulases, xylanases and β-glucosidases, respectively. Higher cellulase activities were observed in SSF, while β-glucosidase activities were higher under submerged fermentation. Statistical modelling predicted the fungal percentage ratio of 60:14:4:2 for A. niger: P. chrysogenum: T. herzianum: P citrinum with a substrate medium ratio of 0.39:1. Under the optimized conditions, a considerable increase to 78.5 U/mL, 3544.7 U/mL and 250.9 U/mL was observed in activities of cellulases, xylanases and β-glucosidases, respectively, with cellulases and xylanases activity increased more than two-fold. A greater amount of lignin was degraded by mixed cultures (17.9% degradation) compared to 8% degradation under submerged fermentation using P. chrysogenum. Gas chromatography-mass spectrometry (GC-MS) analysis of derivatized degradation products indicated the presence of 220 metabolites, of which 78 were categorized as significant by statistical analyses. Although, a majority of metabolites were observed to be utilized during the process, stigmasterol, glycerol, maleic acid, xylitol and citric acid were observed to be generated by the fungi during the degradation process. The research demonstrated enhanced degradation abilities of winery-derived biomass using mixed fungal cultures.