Metabolic analysis on synthesis of polyhydroxyalkanoates (PHAs) from volatile fatty acids by Ralstonia eutropha

This work investigates the PHA production from mixtures of volatile fatty acids by Ralstonia eutropha. Polyhydroxyalkanoates (PHAs) are a family of biodegradable polymers and naturally accumulated in some microorganisms as carbon and energy storage materials under unbalanced growth conditions. The major problem which hinders the industrial production and wide applications of PHAs is its relatively high production cost. Using cheap sources, such as fatty acids digested from food and agriculture byproduct, for PHAs production is one of the attracting ways to reduce its production cost. PHAs synthesis from short-chain fatty acids in batch culture was investigated with both nutrient rich medium and chemical defined medium. It was observed that Ralstonia eutropha could use fatty acids containing 2 to 4 carbon atoms such as acetic, propionic or butyric acids as the sole carbon source to grow and accumulate PHAs. The yield of PHAs from acetic acid is relatively low, and the high yield, about 0.40 g PHA/ g substrate was obtained with butyric acid as the only carbon source. It was found that the inhibitory effect of acids to bacteria increased with increasing acid concentration and carbon number of the acids. The initial biomass concentration strongly affects the acid detoxification by R. eutropha and the efficient production of PHA on fatty acids. PHA production by Ralstonia eutropha in mixtures of volatile fatty acids in batch culture was investigated. With a binary mixture of acetic acid and propionic acid, a quick acids consumption and accumulation of PHA were observed. When R. eutropha grew on the mixture of propionic acid and butyric acid, the specific propionic and butyric acids consumption rate were quite low compared with other acid mixtures, but the PHA content in cells was the highest. The mixture of propionic and butyric acids was the efficient substrates for PHA production. Acetic acid and its relative concentration to other acids affect the utilization of propionic and butyric acid by R. eutropha. The metabolic pathways have been described in detail with reactions occurred intracellularly for the synthesis of PHA and biomass in mixtures of acids. Metabolic flux analysis on the growth of cells and synthesis of polymer showed the carbon distribution among the three major carbon utilizations (biomass synthesis, polymer synthesis and due to energy generation). A small flux entering gluconeogenesis is due to the nitrogen limitation conditions of the controlled cultures. Most of the input carbon was incorporated into polymer and/or excreted as CO2. The generation of cofactor NADPH affects the relative fluxes of acetyl-CoA to TCA cycle and acetyl-CoA to glyoxylate shunt