A multistage system for poly(hydroxyalkanoate) (PHA) production consisting of five continuous stirred tank reactors in series (5-CSTR) with Cupriavidus necator DSM 545 as production strain was modelled using formal kinetic relations. Partially growth-associated production of PHA under nitrogen limited growth was chosen as modelling strategy, thus the Luedeking-Piret's model of partial growth-associated product synthesis was applied as working hypothesis. Specific growth rate relations adjusted for double substrate (C and N source) limited growth according to Megee et al. and Mankad-Bungay relation were tested. The first stage of the reactor cascade was modelled according to the principle of nutrient balanced continuous biomass production system, the second one as two substrate controlled process, while the three subsequent reactors were adjusted to produce PHB under continuous C source fed and nitrogen deficiency. Simulated results of production obtained by the applied mathematical models and computational optimization indicate that PHB productivity of the whole system could be significantly increased (from experimentally achieved 2.14 g L(-1) h(-1) to simulated 9.95 g L(-1) h(-1)) if certain experimental conditions would have been applied (overall dilution rate, C and N source feed concentration). Additionally, supplemental feeding strategy for switching from batch to continuous mode of cultivation was proposed to avoid substrate inhibition.

A metabolic network consisting of 48 reactions was established to describe intracellular processes during growth and poly-3-hydroxybutyrate (PHB) production for Cupriavidus necator DSM 545. Glycerol acted as the sole carbon source during exponential, steady-state cultivation conditions. Elementary flux modes were obtained by the program Metatool and analyzed by using yield space analysis. Four sets of elementary modes were obtained, depending on whether the pair NAD/NADH or FAD/FADH2 contributes to the reaction of glycerol-3-phosphate dehydrogenase (GLY-3-P DH), and whether 6-phosphogluconate dehydrogenase (6-PG DH) is present or not. Established metabolic network and the related system of equations provide multiple solutions for the simultaneous synthesis of PHB and biomass; this number of solutions can be further increased if NAD/NADH or FAD/FADH2 were assumed to contribute in the reaction of GLY-3-P DH. As a major outcome, it was demonstrated that experimentally determined yields for biomass and PHB with respect to glycerol fit well to the values obtained in silico when the Entner-Doudoroff pathway (ED) dominates over the glycolytic pathway; this is also the case if the Embden-Meyerhof-Parnas pathway dominates over the ED.

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