Brewer's Spent Grain (BSG) is a byproduct of brewing in the food industry. However, few studies have addressed the functional features of protein from BSG and how it can be enhanced through optimization studies. The present study is centered on extracting protein using an alkaline treatment to improve recovery. This extraction process was systematically carried out under different alkaline concentrations (0.1, 0.5, 2, 4, and 8 M KOH), with 5 mM sodium metabisulfite added to each of the concentrations. The samples were incubated at room temperature (30 °C; 30 rpm) for 24 h. Precipitation with saturated citric acid to a pH of 3 and freeze-drying for 24 hr were performed. This then represented a fixed process variable. Bradford assay was utilized for the quantification of the protein extract. The result of the quantification showed a protein content of 614.6 mg/100 g of BSG at a concentration of 0.5 M KOH, which was significantly different (p<0.05) from other concentrations. While 4 M KOH was also significantly different, it recorded the lowest protein content of 33.36 mg/100 g. Variation in the alkaline concentration impacts the protein yield. This highlights the potential for optimized protein recovery using the alkaline method for upscaling and sustainable biomass conversion. It is pertinent to explore innovative methods for protein extraction from agro-industry byproducts like BSG, as this will assist in addressing the demand for suitable protein sources.