Degradation of ochratoxin A by Acinetobacter calcoaceticus

Microorganisms were screened for their capability to degrade ochratoxin A (OTA). Among test microorganisms, Acinetobacter calcoaceticus was found to degrade OTA. The degradation of OTA by A. calcoaceticus was studied in an ethanol-minimal salts medium with an initial OTA concentration of 10-50 /μg/mL at 25°C and 30°C. The hydrolysis of OTA by the crude enzyme isolated from A. calcoaceticus also was studied in a 0.1 M NaCl-0.02 M Tris buffer (pH 7.5) at 25°C. A. calcoaceticus was able to degrade OTA in ethanol-minimal salts medium with initial concentrations of OTA of up to 40 μg/mL. At 50 μg/mL, OTA inhibited the growth of A. calcoaceticus and no degradation of OTA occurred. The OTA degrading activity was found in the cell-free growth medium and in the cells. The rate of OTA removed by A. calcoaceticus was concentration dependent. As the OTA concentration increased, the rate of OTA removed by A. calcoaceticus increased. The average amounts of OTA removed by A. calcoaceticus in medium with an initial OTA concentration of 10, 20, 30, and 40 μ/mL were 0.1005, 0.1226, 0.1374, and 0.2262 μg/mL/hr at 30°C, respectively. At 25°C, the average amounts of OTA removed by A. calcoaceticus were 0.0636, 0.1172, 0.1264, and 0.2232 μg/mL/hr in medium containing 10, 20, 30, and 40 μg OTA/mL, respectively. At 25°C and 30°C, the growth of A. calcoaceticus in medium containing 50 μg OTA/mL was inhibited, and OTA degradation did not occur. The rate of OTA removed by A. calcoaceticus in medium containing the same initial OTA concentrations of 20, 30, and 40 μg/mL was not significantly different (p<0.05) at 25°C and 30°C. OTA was degraded significantly by A. calcoaceticus during and after the log phase of cell growth at incubation temperatures of 25°C and 30°C. At 25°C, the initial rate of OTA hydrolyzed by crude enzyme of 0.2 mg protein/mL in 0.1 M NaCl-0.02 Tris buffer containing 10-60 jug OTA/mL in 10 μg/mL increments was 0.1526, 0.3215, 0.4423, 0.5903, 0.6158, and 0.6198 μg/mL/hr, respectively. The initial rate of OTA hydrolyzed by crude enzyme of 0.15 mg protein/mL was 0.1194, 0.2454, 0.3593, 0.5126, 0.5362, and 0.5581 μg/mL/hr, respectively. The initial rate of OTA hydrolyzed by crude enzyme of 0.1 mg protein/mL was 0.0550, 0.1176, 0.1502, 0.1905, 0.3087, and 0.4592 μg/mL/hr, respectively. Generally, the initial rate of OTA hydrolyzed by crude enzyme increased as the concentrations of crude enzyme and/or OTA increased. However, the initial rates of OTA hydrolyzed by crude enzyme of 0.15 and 0.2 mg protein/mL at OTA concentrations of 40, 50, and 60 μg/mL were not significantly different (p>0.05). Hydrolysis of OTA by the cells and crude enzyme of A. calcoaceticus yielded ochratoxin α. A bioassay showed decreased toxicity of OTA toward Bacillus cereus mycoides as OTA was hydrolyzed to ochratoxin α by the cells and crude enzyme of A. calcoaceticus.