Food waste (FW) is an abundant resource with great potential for lactic acid (LA) production. In the present study, the effect of storage time on FW characteristics and its potential for LA production was investigated. The largest part of sugars was consumed during 7 to 15 days of FW storage and the sugar consumption reached 68.0% after 15 days. To enhance the LA production, micro-aerobic conditions (13 mL air/g VS) and addition of β-glucosidase were applied to improve polysaccharides hydrolysis, resulting to increase of monosaccharides content to 76.6%. Regarding fermentative LA production, the highest LA titer and yield of hydrolyzed FW was 32.1 ± 0.5 g/L and 0.76 ± 0.01 g/g-sugar, respectively. Furthermore, L-LA isomer was higher than 70% when FW was stored for up to 7 days. However, attention should be paid on controlling the FW storage to approximately one week.