Production of two phase polyhydroxyalkanoic acid granules in Ralstonia eutropha

To synthesize layered granules consisting of selected phases of polyhydroxy butyrate (PHB) homopolymer and PH(B-co-V) copolymer, Ralstonia eutropha was grown on fructose and limited quantities (1 g/l) of valeric acid. Exhausti on of the valerate resulted in a carbon source shift and a shift in the com position of polyhydroxyalkanoate (PHA) being synthesized within the cell. T he synthesis rates were 0.030 g PH(B-co-V)/l per h and 0.033 g PHB/l per h, giving a copolymer composition of 48% HV. The valerate was exhausted at ap proximately 12 h at a rate of 0.0894 g/l per h after which only PHB was pro duced through the remaining 12 h at 0.033 g PHB/l per h from the remaining fructose, which was utilized at a constant rate of 0.0861 g/l per h through out all 24 h of the experiment. Differential scanning calorimetry (DSC) of isolated granules showed two glass transitions, confirming the presence of two distinct polymer phases within the layered granules. Transmission elect ron microscopic images stained with RuO4 revealed a heavily stained copolym er core within a lighter stained PHB shell, confirming the expected morphol ogy of granule composition. Thus, biosynthesis can be exploited for the con trol of domain sizes in layered granules, potentially providing metabolic c ontrol over the physical properties of the resultant polymer. (C) 1999 Else vier Science B.V. All rights reserved.