Biorefinery, which utilizes carbon-neutral biomass as a resource, is attracting attention as a significant alternative in a modern society confronted with climate change. In this study, spent coffee grounds (SCGs) were used as the feedstock for lactic acid fermentation. In order to improve sugar conversion, alkali pretreatment was optimized by a statistical method, namely response surface methodology (RSM). The optimum conditions for the alkali pretreatment of SCGs were determined as follows: 75 °C, 3% potassium hydroxide (KOH) and a time of 2.8 h. The optimum conditions for enzymatic hydrolysis of pretreated SCGs were determined as follows: enzyme complex loading of 30-unit cellulase, 15-unit cellobiase and 50-unit mannanase per g biomass and a reaction time of 96 h. SCG hydrolysates were used as the carbon source for Lactobacillus cultivation, and the conversions of lactic acid by L. brevis ATCC 8287 and L. parabuchneri ATCC 49374 were 40.1% and 55.8%, respectively. Finally, the maximum lactic acid production by L. parabuchneri ATCC 49374 was estimated to be 101.2 g based on 1000 g of SCGs through the optimization of alkali pretreatment and enzymatic hydrolysis.