The effect of Polyhydroxyalkanoates (PHAs) in Microbial Fuel Cells (MFCs)

Abstract

Βioremediation technologies are eco-friendly and usually nature-based solutions to combat environmental pollution. Among them, bioelectrochemical systems (BES), such as microbial fuel cells (MFCs), can be used to accelerate the process. In MFCs, the degradation of pollutants can result in generation of electricity and/or other added-value products contributing to a circular economy concept while producing non-toxic, biodegradable compounds and renewable energy. Waste cooking oil (WCO) is an agro-industrial wastewater stream of severe environmental concern. MFCs could be an effective treatment way to manage WCO with concurrent production of electricity and added-value products. Furthermore, the effect of added-value products, such as polyhydroxyalkanoates (PHAs) which are considered as biopolymers to replace the petrochemical based polymers used in plastics, on electricity generation is examined. Firstly, enrichment experiments, with activated sludge (AS) as inoculum and acetate, which is known to be favorable for PHAs production, as carbon source was used. Six samples were isolated fed with different initial sodium acetate concentration. Next generation sequencing was performed to identify the microorganisms in the six mixed cultures. It was realized that as sodium acetate concentration was increased, an increased abundance of microorganisms belonging Actinobacteria was observed in the samples. Considerable amounts of the genus Corynebacterium of the family Corynebacteriaceae were found in S4 (6 g/L acetate) (18.90%), S5 (8g/L acetate) (49.75%) and S6 (10g/L acetate) (65.82%). The species Corynebacterium sputi was found in S5 (63.24%) and S6 (63.24%) in high concentrations. 24h cultures were performed for all samples. Gas Chromatography-Mass spectrometry (GC/MS) intracellular analysis revealed that all samples had the same PHAs, at different abundance. In particular, 3-hydroxydecanoate, 3-hydroxydodecanoate and 3-hydroxytetradecanoate. Many fatty acids (FAs) were also observed indicating microbial lipids production. As Corynebacterium seems to be correlated with PHA production and is electroactive, the mixed culture with high abundance of this genus was chosen for further studies (10g/L acetate). So MFCs were set up using this microorganism and WCO in the anode to check the effect of PHAs. MFCs using AS and WCO in the anode and were also set up, for comparison. In the MFCs with AS and WCO which operated for 183 days, the highest % COD removal was noted at day 80, an average of 49%. Extracellular production of a white compound was obvious. The product formed was a mixture of biosurfactants produced by the mixed microbial community of AS. Biosurfactants are amphiphilic molecules with hydrophobic and hydrophilic portions that reduces the surface and interfacial tensions between fluids of different polarities, enhancing the solubility, bioavailability and biodegradation of hydrophobic substrates. In total, 12 FAMEs were detected. The specific biosurfactants’ lipid profile indicates an enhanced degradation of hydrophobic substances, such as hydrocarbons and fats of WCO due to the many fatty acids detected in it. The MFCs operation of mixed cultures as inoculum and WCO as fuel lasted 97 days. The initial resistance was 4.5 kΩ and gradually decreased to 3 kΩ. On the 97th day of operation and 7 days after the last feed replacement of the anode with WCO, several analyses were performed. Intracellularly, twenty-two FAs ranging from C5-C25, were detected in all MFCs at similar concentrations. In total, 6 medium chain length PHAs were observed which are the double amount compared to 24h-aerobic cultures. Comparing the MFCs found here and under 24h-aerobic conditions, the culture here shows itself to be in survival mode. On average 56% COD removal was observed. Extracellularly, total petroleum hydrocarbons (TPHs) and FAs of WCO before and after treatment were measured. The mean hydrocarbons removal rate was 75.4%, with some of them, especially the n-alkanes, reaching removal rates > 60%. Both aromatic and aliphatic hydrocarbons were biodegraded. In total, 7 FAs were detected in WCO. Mixed cultures in MFCs appear to have effectively diminished the unsaturated FAs of methyl elaidate and methyl linoleate, as well as methyl behenate. However, the concentrations of saturated ones to MFCs were significantly higher. This outcome may be explained by the gradual growth of intra- and extracellular metabolites, such as polyhydroxyalkanoates (PHAs) and biosurfactants, in the anodic compartment. In addition, the Emulsification Index test was performed indicating that this mixed culture possibly produces biosurfactants. Comparing the best MFCs from each condition (AS and mixed cultures), the MFC with AS and WCO reached maximum current at 0.145 mA and power was measured at 0.105 mW, while in the MFC with mixed culture and WCO power was measured as high as 0.163 mW and current reached 0.233 mA. In the latter MFC, when the fuel is depleted, microorganisms can then use PHAs as electron donors to continue the process of electron transfer which was also observed in this thesis. After 7 days with no feed, where the WCO’s components were significantly removed, PHAs were accumulated while electricity generation remained at high levels. On top, comparing PHAs accumulation at aerobic conditions (after enrichment) and PHAs under MFCs conditions, more PHAs, in terms of variety, were produced showing the capability of the microorganisms to survive and perform under adverse conditions (i.e. no WCO-fuel feed). This is a novel and promising circular-based approach for wastewater treatment with concurrent added-value products and renewable energy generation.