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Crude mushroom solution to degrade micropollutants and increase the performance of biofuel cells

Sabine Sané, a doctoral student in the Department of Microsystems Engineering (IMTEK) at the University of Freiburg, has developed a concept that shows how micropollutants can be degraded in wastewater and how the latter can serve as a valuable source of raw materials. She is one of four researchers who have been awarded the 2014 Huber Technology Prize “Future Water” with a purse of 10,000 euros. Her concept is based on an enzyme that is secreted by the turkey tail fungus Trametes versicolor. This enzyme, known as laccase, has been shown to efficiently degrade pollutants and increase the performance of biofuel cells.

Sabine Sané, doctoral student in the Department of Microsystems Engineering (IMTEK) at the University of Freiburg, is working on the improvement of the energy efficiency of wastewater treatment plants. © Conny Ehm

Wastewater contains a wide range of pollutants, including heavy metals, synthetic organic compounds, viruses and bacteria. Micropollutants are pollutants that occur in concentrations of less than a few micogrammes per litre. Technological progress has improved the detection of such small concentrations of substances. “The degradation of micropollutants has not previously been an issue, particularly because technologies that can detect them have only recently become available,” says Sané, doctoral student in the IMTEK’s Bioelectrochemical Systems research group. 

Drug residues or hormones from private households are among those compounds that wastewater plants are unable to degrade. “Many women take hormone-based contraceptives, which are partially excreted with urine,” says Sané. Ibuprofen and diclofenac are synthetic chemicals that cannot be broken down, and instead accumulate in the ecosystem. The same thing happens with X-ray contrast agents used in hospitals and antibiotics used in industrial livestock farming. 

The turkey tail fungus comes to the rescue

The turkey tail fungus Trametes versicolor grows on trees and is able to degrade lignin, a complex polymer derived from wood that is very resistant to degradation. © IMTEK, University of Freiburg

Compounds that cannot be broken down in wastewater plants accumulate in aquatic ecosystems, and threshold limits need to be put in place in order to reduce the negative effects on the environment. Antidepressant drugs are known to impact the metabolism and behaviour of fish: it makes them more aggressive rather than happier. Oestrogens have been shown to render fish infertile. 

However, a fungus known as Trametes versicolor or turkey tail fungus has been shown to change the situation. This particular fungus is a polypore fungus commonly found on European beech. The fungus degrades fallen trees and is also able to break down lignin. These processes are achieved by the fungal enzyme laccase, which catalyses the oxidation of aromatic compounds concomitantly with the reduction of oxygen to water. The enzyme is extremely stable and withstands high temperatures and solvents. The fungus has been used in research for a number of years, and can be cultivated in a similar way to bacteria.

Like mould on apple juice

Trametes versicolor grown in synthetic culture medium looks quite like the type of mould that grows on the surface of apple juice. However, Trametes is able to degrade micropollutants. © IMTEK, University of Freiburg

The fungal laccase enzyme can be used to break down pollutants into their constituents. “Laccase oxidises micropollutants and is far from selective,” says Sané. “It is completely non-specific and can use all kinds of substrates.” Compared to ozone or activated charcoal for the treatment of wastewater, the fungus appears to be the smarter alternative. Activated charcoal is produced from carbonaceous materials and needs to be disposed of after use; the method involving ozone consumes a lot of energy and requires trained personnel to deal with the large number of dangerous products generated. Nevertheless, the production of laccase tends to be rather expensive as it needs to be expressed in and isolated from microorganisms. 

Sané’s project therefore involved looking for a cheaper way to produce laccase. She decided to use the crude culture supernatant without further treatment. “When the fungus is grown on a liquid nutrient culture, it floats on top of it just like mould on apple juice.” In this case, Trametes appears to secrete more than just laccase. “The fungus secretes the entire enzyme complex into the liquid medium and we are using the crude supernatant for the degradation of pollutants without further purification.” 

This is only part of the concept for which Sané and three other up-and-coming scientists were recently awarded the 2014 Huber Technology Prize: Future Water for projects involving “Energy and Resources from Wastewater”. The objective of this new prize is to encourage up-and-coming scientists to develop concepts that contribute to protecting the environment and saving resources. The second part of Sané’s concept foresees the use of the fungus for improving the performance of a hybrid microbial-enzymatic fuel cell. 

Cost-efficient energy efficiency with Trametes

Schematic showing the potential integration of Trametes cultivation into the wastewater treatment plant concept: The fungal laccase provides the energy for the biofuel cell at the same time as degrading micropollutants in the wastewater. © IMTEK, University of Freiburg
In a hybrid microbial-enzymatic fuel cell, microorganisms that are found in wastewater can attach to the anode and transfer electrons resulting from the degradation of compounds. “These microorganisms are able to ‘breathe’ with the anode rather than with oxygen,” explains Sané, going on to add “at the same time, they metabolise the organic carbon in the wastewater. We do not need to feed them additional organic carbon compounds.” An electric circuit is established when the electrons migrate towards the cathode where the laccase enzyme catalyses the transfer of electrons to oxygen. However, laccase has a lifespan of no more than two weeks, and laccase-based fuel cells can currently only be used in the laboratory. The isolation and purification of the enzyme is a rather complex process, even in the laboratory. Sané had the idea of skipping the isolation and purification steps and using the crude supernatant without further treatment. This solution contains the enzyme as well as many other molecules. Sané found no significant difference in the fuel cell performance when using crude supernatant that contains laccase compared to purified laccase in culture medium. “This was totally unexpected,” says Sané. “We have been able to show that the lifespan of the cathode can be greatly prolonged if we repeatedly exchange the crude culture supernatant.” Sané ended up with a cathode that had a lifespan of four months, and was convinced that this could be prolonged further. “Unfortunately, my reactor dried out,” she says. Sané’s vision is to combine two methods based on the laccase enzyme: a wastewater fungus farm combined with a wastewater treatment plant, which would then be an autonomous wastewater plant that produces the energy it consumes at the same time as degrading micropollutants. “The smartest possible solution would be to grow the fungus on the surface of the wastewater that contains the fuel cell and in which the drug residues are broken down,” says Sané. “We would be able to purify water at the same time as generate electricity.” The next step will now be to turn the concept into reality.

Further information:
Sabine Sané
Department of Microsystems Engineering - IMTEK
University of Freiburg
Georges-Koehler-Allee 103
79110 Freiburg
Tel.: +49 (0)761 / 203-73262
E-mail: Sabine.Sane(at)imtek.uni-freiburg.de

Website address: https://www.biooekonomie-bw.de/en/articles/news/crude-mushroom-solution-to-degrade-micropollutants-and-increase-the-performance-of-biofuel-cells/