Effects of storage on mixed-culture biological electrodes

Abstract

Storage methods are important to preserve the viability and biochemical characteristics of microbial cultures between experiments or during periods when bioreactors are inactive. Most of the research on storage has focused on isolates; however, there is an increasing interest in methods for mixed cultures, which are of relevance in environmental biotechnology. The purpose of this study was to investigate the effect of different storage methods on electrochemically active enrichment cultures. Acetate-oxidizing bioanodes generating a current density of about 5 A m(-2) were enriched in a microbial electrolysis cell. The effect of five weeks of storage was evaluated using electrochemical techniques and microbial community analysis. Storage by refrigeration resulted in quicker re-activation than freezing in 10% glycerol, while the bioelectrochemical activity was entirely lost after storage using dehydration. The results showed that the bioelectrochemical activity of bioanodes stored at low temperature could be retained. However, during the re-activation period the bioanodes only recovered 75% of the current density generated before storage and the bacterial communities were different in composition and more diverse after storage than before.

Figure 1. Current and acetate concentration with time in the reactor when the anodes were controlled at 0 V vs SHE.

Arrows show when the nutrient medium was replaced and vertical lines show when normal operation was stopped and electrochemical tests were carried out.

Figure 2. Cyclic voltammograms of anodes in the reactor.

The dashed lines show the average cyclic voltammograms for all eight anodes in the reactor obtained during the second test before storage (see Fig. 1 for the times of these tests). The solid lines shows the average cyclic voltammograms for duplicate electrodes exposed to the three storage methods: acetone dehydration, refrigeration, and freezing in 10% glycerol solution. New refers to new graphite rod anodes placed in the reactor when the system was restarted after storage. The cyclic voltammetry was carried out between −1.0 and 0.5 V vs SHE; however, only the region between −0.5 V and 0.2 V is shown in the figure.

Figure 3. Linear sweep voltammograms from six different tests of the anodes in the reactor.

Two tests (1^st–2^nd) were done before storage (see Fig. 1 for the times of these test) and the rest (3^rd–6^th) after storage. The figure legends refer to the three storage methods used: acetone dehydration, refrigeration, and freezing in 10% glycerol solution. New refers to new graphite rod anodes placed in the reactor when the system was restarted after storage.

Figure 4

(A) Kinetic factor (K), which corresponds to i₀F/RT, for different tests after storage. (B) Open circuit potential (OCP) of the eight different anodes placed in the reactor after storage. The grey regions show the variation in K and OCP of the eight anodes before storage. The figure legends refer to the three storage methods used: acetone dehydration, refrigeration, and freezing in 10% glycerol solution. New refers to new graphite rod anodes placed in the reactor when the system was restarted after storage.

Figure 5. Relative abundance of 16S rRNA gene sequences in the electrode biofilms.

(A) Distribution of phyla with relative abundance >0.5% in any of the samples. (B) Distribution of major taxa (relative abundance >5% in any of the samples) at highest possible taxonomic determination using the Greengenes taxonomy.