Bioprocess Parameters and Oxygen Transfer Characteristics in β‐Lactamase Production by Bacillus Species

After screening potential β‐lactamase producers in a medium containing penicillin G, an inducible ( Bacillus subtilis NRS 1125) and a constitutive ( Bacillus licheniformis 749/C ATCC 25972) β‐lactamase producer were selected. As the highest enzyme activity was obtained with B. licheniformis 749/C, the effects of the concentration of carbon sources, i.e., glucose, fructose, sucrose, citric acid, and glycerol, and nitrogen sources, i.e., (NH 4 ) 2 HPO 4 , NH 4 Cl, yeast extract, casamino acids and peptone, pH, and temperature on β‐lactamase production were investigated with B. licheniformis 749/C in laboratory scale bioreactors. Among the investigated media, the highest volumetric activity was obtained as 270 U cm −3 in the medium containing 10.0 kg m −3 glucose, 1.18 kg m −3 (NH 4 ) 2 HPO 4 , 8.0 kg m −3 yeast extract, and the salt solution at 32 °C and pH 0 = 6.0. By using the designed medium, fermentation and oxygen transfer characteristics of the bioprocess were investigated at V = 3.0 dm 3 bioreactor systems with a V R = 1.65 dm 3 working volume at Q O / V R = 0.5 vvm and N = 500 min ‐1 . At the beginning of the process the Damköhler number was <1, indicating that the process was at biochemical reaction limited condition; at t = 2–5 h both mass‐transfer and biochemical reaction resistances were effective; and at t = 6–10 h ( Da ≫1) the bioprocess was at mass transfer limited condition. Overall oxygen transfer coefficients ( K L a) varied between 0.01 and 0.03 s −1 , enhancement factor ( K L a/ K L a O ) varied between 1.2 and 2.3, and volumetric oxygen uptake rate varied between 0.001 and 0.003 mol m −3 s −1 throughout the bioprocess. The specific oxygen uptake and the specific substrate consumption rates were the highest at t = 2 h and then decreased with the cultivation. The maximum yield of cells on substrate and the maximum yield of cells on oxygen values were obtained, respectively, as Y X/S = 0.34 and Y X/O = 1.40, at t = 5 h, whereas the highest yield of substrate on oxygen was obtained as Y S/O = 6.94 at t = 3.5 h. The rate of oxygen consumption for maintenance and the rate of substrate consumption for maintenance values were found, respectively, as m O = 0.13 kg kg −1 h −1 and m S = 3.02 kg kg −1 h −1 .