The rapid and practical determination of lactic acid concentration during anaerobic digestion and in acidification samples is extremely important to ensure the digester is running properly and to avoid build-up of lactic acid. Thus, developing a simple and fast method for analyzing lactic acid in anaerobic digestion samples is important. The results of the present study revealed that lactic acid is an intermediate product of anaerobic digestion, which could be quickly measured using a portable lactate biosensor. We obtained linear standard curves from the relationship between various lactic acid concentrations of the standard solution and the lactate biosensor reading, which were tested at different temperatures (30, 45 and 55EC). A high value of coefficient determination, -0.98–0.99, was obtained from all standard curves. This suggests that the evaluated system was accurate, reliable, and reproducible.