Brewer’s Spent Grain (BSG) is renewable lignocellulosic biomass generated from the beer brewing process. It serves as a substrate for various biotechnological applications. BSG was used as the main substrate for bioethanol production with Saccharomyces carlsbergensis in submerged fermentation. Saccharification and fermentation studies were performed for the production of bioethanol. A sterilized fermenter was loaded with 50 g L-1 of BSG at 29±2 °C and an agitation speed of 180 rpm. pH was adjusted to 6.0 before the addition of 500 mL of yeast culture for 7 days under submerged and optimized conditions. The fermented product was concentrated using a rotary evaporator at 66±1 °C, and ethanol was qualitatively determined by the dichromate method. Bioethanol yield was 22%, with a specific gravity of 0.8 at 28 °C. Fourier-Transform Infrared Spectroscopy (FTIR) confirmed the presence of -CH3 stretch, -OHstretch and -CH2stretch in bioethanol. For the preservative test, Staphylococcus spp., Erwinia spp., Lactobacillus spp., Bacillus spp., Xanthomonas spp., Pseudomonas spp., Micrococcus spp. and Corynebacterium spp. were the bacteria isolated from fruits examined from different regions of Osun State. The genera of fungi isolated were Aspergillus, Colletotrichum, Penicillium, Fusarium, Alternaria, Rhizopus, Candida, Saccharomyces, Geotrichium and Pichia. Bioethanol produced from BSG inhibited the growth of microorganisms with zones of inhibition range from 7.0 mm to 11.5 mm, and thus, selected fruits were preserved. Hence, the fermentation technology of agro-industrial wastes with microorganisms can be adopted to convert waste biomass to useful resources.