For practical application of autotrophic production of P(3HB) from CO2 by A. eutrophus, the loss of substrate gas utilization efficiency concomitant with the exhaustion of the gas from fermenter, and the potential of explosion of the substrate gas mixture are very serious problems to be solved. A recycled-gas closed-circuit culture system attains high gas utilization efficiency by reusing the exhausted gas. Figure 2 shows the gas-recycling culture system in bench scale. This system could also eliminate the potential for the gas mixture to explode by maintaining the oxygen concentration in the gaseous phase below the lower limit for explosion(6.9 %(v/v)) and introducing several other safety measures. Oxygen consumption by the cells in autotrophic synthesis of P(3HB) is very large as shown in the following equations, the decrease in the driving force for oxygen from the gas phase into the liquid phase then results in the serious decrease in P(3HB) productivity.

Exponential cell growth; 21.36 H2 + 6.21 O2 + 4.09 CO2 + 0.76 Nffi

-» C4.09H7.1 ЗО 1.89N0.76

P(3HB) formation; 33 H2O + 12 O2 + 4 CO2 —> C4H6O2 + ЗО H2O

Hence, a doughnut-shaped agitation system to attain a KLa of 2,970 h_1 was used in the bench plant to compensate for the decrease in oxygen transfer. As a result, cell and P(3HB) concentrations increased to 91.3 gednr3 and 61.9 gedm’3 respectively, under 02-limited condition after 40 h of cultivation (Fig.3). While the 62 concentration in the gas phase was maintained at very low level, the overall productivities of biomass and P(3HB) obtained in this cultivation were 2.28 g*dnr3* h_1 and 1.55 g-dnr^lr1, respectively, which were much higher than those reported for other autotrophic cultivation of hydrogen-oxidizing bacteria (Table I).