Reaction kinetics of anodic biofilms under changing substrate concentrations: Uncovering shifts in Nernst-Monod curves via substrate pulses

Abstract

In the present study, it is shown that the concentration dependency of undefined mixed culture anodic biofilms does not follow a single kinetic curve, such as the Nernst-Monod curve. The biofilms adapt to concentration changes, which inevitably have to be applied to record kinetic curves, resulting in strong shifts of the kinetic parameters. The substrate concentration in a continuously operated bioelectrochemical system was changed rapidly via acetate pulses to record Nernst-Monod curves which are not influenced by biofilm adaptation processes. The values of the maximum current density j _max and apparent half-saturation rate constant K _s increased from 0.5 to 1 mA cm^-2 and from 0.5 to 1.6 mmol L^-1, respectively, within approximately 5 h. Double pulse experiments with a starvation phase between the two acetate pulses showed that j _max and K _s decrease reversibly through an adaptation process when no acetate is available. Pseudo-capacitive charge values estimated from non-turnover cyclic voltammograms (CV) led to the hypothesis that biofilm adaptation and the observed shift of the Nernst-Monod curves occurred due to changes in the concentration of active redox proteins in the biofilm. It is argued that concentration-related parameters of kinetic models for electroactive biofilms are only valid for the operating points where they have been determined and should always be reported with those conditions.

Keywords: Nernst‐Monod model; anodic biofilms; bioelectrochemical system; concentration dependency; reaction kinetics.

FIGURE 1

Schematic illustration of different Nernst‐Monod curves which result from microbial adaptation processes of a single electroactive biofilm, and an apparent Nernst‐Monod trajectory which may be constructed from experimental steady state data points

FIGURE 2

Schematic drawing of the experimental setup. Blue lines indicate acetate substrate solution, green lines nitrogen. WE1‐3 are the three working electrodes, RE is the reference electrode, CE is the counter electrode

FIGURE 3

(A) Current density, acetate concentration, and CDWD over cultivation time during an acetate pulse experiment. Continuous flow operation was started at 45 h, flow rate is 0.4 mL min⁻¹, acetate feed concentration is 4 mmol L⁻¹, reactor volume is 0.25 L. (B) Nernst‐Monod plot for the acetate pulse experiment. The measured data points and the Nernst‐Monod curves before (1, orange, short term response) and after (2, green, long term response) the pulse are shown

FIGURE 4

Current density, acetate concentration, and CDWD over cultivation time during two acetate pulses with (A) continuous acetate supply between two acetate pulses, and (B) starvation phase without acetate supply between two acetate pulses. Continuous flow operation was started at 45 h, flow rate is 0.4 mL min⁻¹, acetate feed concentration is 4 mmol L⁻¹, reactor volume is 0.25 L. The letters A‐H relate the data points to the Nernst‐Monod curves in Figure 5. CV1 to CV3 are related to CV discussed in Figure 7

FIGURE 5

Nernst‐Monod plot for two consecutive acetate pulses with (A) continuous acetate supply between two acetate pulses (at A and C) and (B) a starvation phase without acetate supply between the two acetate pulses (at F and H). The measured data points are displayed as markers, the solid lines indicate Nernst‐Monod curves that were fitted to the data at different points in cultivation time. Nernst‐Monod curves before (curve 1, orange) and after (curve 2, green) the acetate pulses as well as after the starvation phase (curve 3, purple) are shown. The letters A–H relate the data points to the data over cultivation time from Figure 4

FIGURE 6

(A) CDWD over total charge before anode extraction for seven experiments. To assess the deviations between the electrodes, two anodes were extracted at the same time in experiment 4 and all anodes were extracted at the same time in experiments 6 and 7. (B) Current density delivered by the individual anodes relative to an even distribution measured before extraction of an anode. Measurement No. 1 is from the experiment in Figure 4B, No. 2 and 3 from the experiment in Figure 4A. Measurement No. 3 was done after one anode had already been extracted. Measurements No. 4–6 are from a set control experiments. (C) CDWD of anodes from the control experiments which were extracted at the same time relative to an even distribution of the CDWD

FIGURE 7

(A) CV under non‐turnover conditions recorded before (CV1), and after the starvation phase (CV2) and after the second acetate pulse (CV3) (compare also to Figure 5B). The CV curves are corrected for the residual (turnover) current. (B) Areas enclosed by the CV obtained by integration

FIGURE 8

Nernst‐Monod plot for acetate pulse experiments with an acetate feed concentration of 4 (blue crosses) and 2.5 mmol L⁻¹ (green circles). Data for 4 mmol L⁻¹ is identical to Figure 5A