The microfungi are a group microorganisms which obtain their energy by aerobic oxidation or by anerobic fermentation of organic materials and release organic acid metabolite as an end product, which is corrosive in nature. The metabolites could be detected online by microbial biosensor. An amperometric microbial biosensor for the direct measurement of metabolites was developed on a modified oxygen electrode, with immobilized Acetobacter sp. on acetylcellulose membrane. The sensor signal and response time were optimized with respect to pH, temperature, concentration and weight of the immobilized microbial cells. The best sensitivity and response time by microbial biosensor were obtained with 1.2 mg of microbial cells at 4.3 pH of the corrosive environment at room temperature (29degreesC). The reproducibility of current and response time difference were examined, using, the same corrosive environment.