A novel high-throughput method for kinetic characterisation of anaerobic bioproduction strains, applied to Clostridium kluyveri

Hexanoic acid (HA), also called caproic acid, can be used as an antimicrobial agent and as a precursor to various chemicals, such as fuels, solvents and fragrances. HA can be produced from ethanol and acetate by the mesophilic anaerobic bacterium Clostridium kluyveri , via two successive elongation steps over butyrate. A high-throughput anaerobic growth curve technique was coupled to a data analysis framework to assess growth kinetics for a range of substrate and product concentrations. Using this method, growth rates and several kinetic parameters were determined for C . kluyveri . A maximum growth rate (µ max ) of 0.24 ± 0.01 h −1 was found, with a half-saturation index for acetic acid (K S,AA ) of 3.8 ± 0.9 mM. Inhibition by butyric acid occurred at of 124.7 ± 5.7 mM (K I,BA ), while the final product, HA, linearly inhibited growth with complete inhibition above 91.3 ± 10.8 mM (K HA of 10.9*10 −3 ± 1.3*10 −3 mM −1 ) at pH = 7, indicating that the hexanoate anion also exerts toxicity. These parameters were used to create a dynamic mass-balance model for bioproduction of HA. By coupling data collection and analysis to this modelling framework, we have produced a powerful tool to assess the kinetics of anaerobic micro-organisms, demonstrated here with C . kluyveri , in order further explore the potential of micro-organisms for chemicals production.