A new PHB producer of Ralstonia eutropha KHB 8862 was developed and high cell density fed-batch fermentation was carried out to investigate the mass production possibilities of poly(3-hydroxybutyrate) (PHB), using firstly a lab scale and secondly a pilot scale 200L fermentor.

During the fermentation process, fructose syrup was used as the sole carbon source and pH and nitrogen sources were controlled with NH₄OH solution. PHB accumulation was almost totally associated with nitrogen, oxygen and phosphate limitation during the fed-batch culture. We investigated the effect of initial volume on the cell growth, PHB accumulation and productivity in a 5 L jar fermentor and found that this small initial volume was very effective in increasing productivity. The importance of this work is that we could achieve, given an initial volume of 1.0 liter and a final volume of 3 L. a final cell concentration of 202 g/L, a PHB concentration of 156 g/L in 76hr, and hence a PHB productivity of 2.1 g/L/h. PHB content, cell yield from fructose syrup, and PHB yield from fructose syrup were 78%, 0.4, and 0.32 (w/w), respectively.

Following these lab scale fermentation resets, we scaled up the size to a pilot scale fermentor, using the same power input per unit volume (P/V＝N3D5) and the same impeller tip speed (πND), in order to investigate the possibilities for PHB mass production. A 200L pilot fermentation run resulted in a final cell concentration of 168 g/L, with PHB content, PHB yield from fructose syrup and productivity being 74%, 0.27 (w/w) and 1.6 g/L/h, respectively.

Shake-flask experiments in o니r laboratories for the production of P(3HB-co-3HV) with R. eutropha KHB 8862 showed that growth was inhibited by propionate as a precursor for copolymer production at concentrations higher than 0.5 g/L. The final percentage of 3HV in the polymer did not vary significantly relative to the concentration of supplied sodium propionate.

The PHB production cost from bacterial fermentation was analyzed and economic evaluation was performed. The effect of various costs such raw materials, utility, labor, maintenance and depreciation were examined. In the case of new investment being implemented or not, the production cost of PHB was $3.15/kg and $2.41/kg, respectively.

PHB productivity and PHB yield on a carbon substrate were both important factors to optimized. Among these, PHB yield on a carbon substrate was found to be the most important because it has multiple effects on the PHB content, recovery efficiency and cost of consumed carbon sources. The increase of PHB yield on a carbon sources significantly decreased the PHB production cost but the increase in productivity had a relatively slight effect on the decrease in PHB production cost because the cost of carbon sources (37%) for PHB was larger in proportion to total cost than the depreciation cost (17%).

Finally, we assumed that the increased PHB yield from carbon sources and the development of new cheaper substrates would be more effective in decreasing PHB production cost than the increase in productivity.

It was presumed that the PHB production cost would be $4/kg. Furthermore, it was demonstrated that PHB is not in competition with consumable plastics such as PET in present market. Therefore, it is essential to lower production cost to be used as a bulk product and desirable to develop new application fields for PHB such as biomedical and cosmeceuticals. However, PHB production cost of $4/kg in this study was considerable result compared with the price of Biopol ($15/kg).

An efficient process for the preparation of poly(3-hydroxybutyrate) (PHB) microspheres with a narrow size distribution was developed. PHB was produced by a fed-batch culture of Ralstonia eutropha KHB 8862 using fructose syrup as the sole carbon source. After autoclaving the cells, PHB granules which had accumulated in the cells were isolated by a detergent-hypochlorite treatment and then spray dried in order to obtain the microspheres. The diameters of the PHB microspheres ranged from 0.6 to 1.1 ㎛ and the weight-average molecular weights were approximately 50,000 with polydispersity indexes of 5.0. The microspheres had a porous internal structure with an average porosity value of 71.5% and efficiently blocked UV light shorter than 220 nm. When isosorbide dinitrate was used as a model drug, the optimal drug loading concentration of the microspheres for controllable retardation was 3% (w/w). The drug release rates in vitro were very fast Almost 80% of the loaded drug was released within 12 h exhibiting typical sustained drug release behaviors.