Isolation of Pseudomonas putida BM01 Accumulating High Amount of $PHA_{MCL}$

A Pseudomonas putida strain able to accumulate high amount of polyesters of medium-chain-length 3-hydroxyalkanoic acids ($PHA_{MCL)$) was isolated from soil in a landfill site using an enrichment technique. Culture condition of the isolated strain for polyester production in a one-step culture was optimized in a mineral-salts medium against pH and concentrations of ammonium sulfate, carbon source(e.g., octanoate), and phosphate. The optimal values for maximal cell growth and PHA accumulation were: pH; 7$\sim$8, $(NH_4)_2SO_4$; 8 mM, octanoate; 40 mM. The optimum temperature was in the range of $20\sim30^{\circ}C$, which was rather broader than in other bacteria. Cell growth was strongly inhibited by the phosphate limitation to less than 1 mM. An increase of phosphate concentration above 1 mM showed little effect on cell growth and polyester accumulation. When the strain was grown on octanoate under this optimized condition it produced 3.4 g dry biomass per liter and yielded 1.7 g PHA per liter amounting to 53 wt% of dry cells. The monomer units composing the polyester synthesized from octanoate were 3-hydroxyoctanoate (3HO), 3-hydroxycaproate (3HC), and 3-hydroxybutyrate (3HB) (85:13:2, mole ratio). Other low linear $C_3\simC_{10}$ monocarboxylic acids were also tested for polyester production.