Hydrodynamic chronoamperometry for probing kinetics of anaerobic microbial metabolism – case study of Faecalibacterium prausnitzii

摘要

Monitoring in vitro the metabolic activity of microorganisms aids bioprocesses and enables better understanding of microbial metabolism. Redox mediators can be used for this purpose via different electrochemical techniques that are either complex or only provide non-continuous data. Hydrodynamic chronoamperometry using a rotating disc electrode (RDE) can alleviate these issues but was seldom used and is poorly characterized. The kinetics of Faecalibacterium prausnitzii A2-165, a beneficial gut microbe, were determined using a RDE with riboflavin as redox probe. This butyrate producer anaerobically ferments glucose and reduces riboflavin whose continuous monitoring on a RDE provided highly accurate kinetic measurements of its metabolism, even at low cell densities. The metabolic reaction rate increased linearly over a broad range of cell concentrations (9 × 10⁴ to 5 × 10⁷ cells.mL⁻¹). Apparent Michaelis-Menten kinetics was observed with respect to riboflavin (KM = 6 μM; kcat = 5.3×10⁵ s⁻¹, at 37 °C) and glucose (KM = 6 μM; kcat = 2.4 × 10⁵ s⁻¹). The short temporal resolution allows continuous monitoring of fast cellular events such as kinetics inhibition with butyrate. Furthermore, we detected for the first time riboflavin reduction by another potential probiotic, Butyricicoccus pullicaecorum. The ability of the RDE for fast, accurate, simple and continuous measurements makes it an ad hoc tool for assessing bioprocesses at high resolution.