Health & Safety for microbes

The process for converting plant biomass into bioethanol is well known. It begins with a pretreatment step, in which the biomass is broken down with chemicals to release the cellulose and hemicellulose, which are then digested by enzymes into fermentable sugars. Finally, microbes such as yeast ferment these sugars into ethanol.

Also well known is the continuing challenge of making this process commercially viable. In part, this is because the pretreatment step is not compatible with the other steps. The strong acids and bases conventionally used to break down the biomass, together with some of the by-products generated as part of this step, are highly toxic, deactivating the enzymes and inhibiting or killing the microbes. Because of this, the chemicals and by-products need to be removed from the treated biomass, usually by washing and centrifugation, before the enzymatic digestion and fermentation steps, adding expense.

Even supposedly greener alternatives to the strong acids and bases for breaking down biomass, such as ionic liquids, suffer from these toxicity problems and thus need to be removed by washing. But a couple of recent studies show that a new range of milder solvents can effectively break down plant biomass while proving much less toxic to enzymes and microbes.

One of these solvents is known as a carboxylate-type liquid zwitterion. A zwitterion is a molecule that contains a functional group with a positive charge and a functional group with a negative charge, so that the molecule has no charge overall. With this ionic structure, zwitterions should theoretically make very effective solvents, just like ionic liquids. But although zwitterions are quite common, with amino acids being the best-known examples, they don’t tend to be liquid at room temperature.

As reported in a paper in the Journal of the American Chemical Society, Kenji Takahashi and his colleagues at Kanazawa University in Japan have now managed to develop just such a liquid zwitterion. They have also shown that it is just as effective at breaking down sugarcane bagasse as ionic liquids, while being far less toxic to microbes. They found that the concentration of their zwitterion required to reduce the growth of Escherichia coli by 50%, a common measure of toxicity, was 158g/L, indicating extremely low toxicity. For a model ionic liquid, by contrast, the concentration was just 9g/L.

As a consequence, when they tried passing treated bagasse straight to the enzymatic digestion and fermentation steps without first washing off the zwitterions, they were able to produce ethanol at a concentration of 1.4g/L. When they tried the same with the ionic liquid, however, they couldn’t produce any ethanol, because the ionic liquid completely inhibited fermentation by E. coli.

Meanwhile, a team of Italian chemists led by Alessandra Procentese at the Combustion Research Institute in Naples, Italy, chose to investigate the potential of deep eutectic solvents (DESs) for breaking down biomass. Like zwitterions, DESs have similar properties to ionic liquids, but they’re milder and cheaper. Procentese and her team thought they might prove particularly adept at breaking down agricultural food waste such as apple cores, potato peelings and the left-over residues from coffee and beer production, as these contain higher levels of sugar and lower levels of lignin than conventional plant biomass.

DESs are produced by mixing a quaternary ammonium salt, usually choline chloride, with a compound that can form hydrogen bonds with the salt. Mixing these two compounds together in just the right proportions produces a complex with a melting temperature that is much lower than either of the two compounds on their own, such that most DESs are liquid at room temperature. Procentese and her team tested two DESs: choline chloride–glycerol and choline chloride–ethylene glycol.

As they report in Biotechnology for Biofuels, both of these DESs could break down the food waste and release cellulose and hemicellulose for digestion into fermentable sugars, although choline chloride–glycerol was slightly more effective. They also found that the DES pretreatment didn’t produce any toxic by-products, leading them to conclude that no detoxification strategy is required for DESs.

Procentese and her team calculated that if this process was applied to all the agricultural food waste produced yearly in Europe, it could generate up 217,000 tonnes of fermentable sugars, all without harming any microbes.

The views represented here are solely those of the author and do not necessarily represent those of John Wiley and Sons, Ltd. or of the SCI.