Fermentation optimization for the production of poly(β-hydroxybutyric acid) microbial thermoplastic

Abstract

Batch culture of Alcaligenes latus, American Type Culture Collection 29713, was investigated for producing the intracellular bioplastic poly(β-hydroxybutyric acid) (PHB). A central, composite experimental design was used to optimize the composition of the culture medium for maximizing the productivity of PHB. Investigated were the effects of temperature, the initial culture pH, the ionic strength of the medium, the concentration of trace elements, the type of nitrogen source, and the carbon-to-nitrogen ratio. The optimal temperature for growth and PHB synthesis appeared to be 33°C; however, over the 25–37°C range, the effect of temperature was negligible. An initial pH value of 6.5 gave the best results; pH values that differed even slightly from the optimum reduced the culture performance. Typical culture characteristics were: 0.075/h maximum specific growth rate, 0.38 g/l h maximum specific sucrose consumption rate, and 0.15 g/l h maximum specific PHB production rate. PHB was lost because of hydrolysis in the stationary phase, suggesting critical importance of timing the harvest. Under the best conditions, PHB constituted up to 63% of dry cell mass after 93 h of culture. The average biomass yield coefficient on sucrose was about 0.4 kg/kg. Of the four nitrogen sources—ammonium chloride, ammonium sulfate, ammonium nitrate, and urea—used, only the first two supported the culture satisfactorily. The biomass and PHB showed clear yield maxima at 1.5 g/l ammonium chloride (C:N ratio = 21.5) and 1.4 g/l ammonium sulfate (C:N ratio = 28.3). The yields were higher with ammonium sulfate and were relatively more sensitive to changes in its concentration. Ionic strength had a strong negative effect on PHB productivity. The highest PHB yield occurred at 4 g/l phosphate buffer concentration. Iron appeared to have the potential to enhance the proportion of PHB in the cells.