Ionic Liquids for Natural Product Extraction Adam Walker Bioniqs Ltd.

Ionic Liquids - Introduction NaCl (mp 801°C) BMIm PF6 (mp -79°C) Ionic liquids are salts, with melting points below ambient temperature They are composed solely of anions and cations – no water is present Their regular, ordered structure gives them very different properties to conventional liquids Ionic liquids based on imidazole have been well studied as novel solvents

Ionic Liquids - Properties Extremely good solvents for many compounds – organic, inorganic, mineral, polymeric and biological Non-volatile – do not evaporate Non-flammable Stabilize sensitive compounds Many possible combinations of ions – can design the solvent for a particular application

Applications Solvents for chemical and biochemical reactions Extraction media for natural product isolation Cleaning agents Storage media for unstable compounds Lubricants Electrochemical/battery materials

“First Generation” Ionic Liquids - Disadvantages Expensive in comparison with molecular solvents. Very few commercially available. Viscous Poorly biodegradable and potentially toxic.

“Second Generation” Ionic Liquids Bioniqs has developed new types of simple ionic liquids, based on ammonium salts. These retain the advantages of ionic liquids, but also offer: - reduced viscosity. - low cost. - straightforward preparation from sustainable precursors. - minimal toxicity and environmental impact. - recyclability and biodegradability.

Example Applications Stabilization of enzymes for biocatalysis Dissolution and processing of biomass Natural product isolation and purification Reactor decontamination

Stabilization of Biomolecules Activity of alcohol dehydrogenase enzyme after storage in water compared to standard (purple) and optimized (red) ionic liquids. H2O AEFF-390 AAFF-015 Enzyme in water deactivated after 10 days at room temperature. Enzyme in AEFF-390 retains activity after 90 days at room temperature.

Biomass Processing Ionic liquids can dissolve numerous polymers, including biological macromolecules such as:- Cellulose (up to 10% wt. solutions) Lignin Proteins DNA/RNA Bioprocessing requires solvent to dissolve substrate without denaturing enzyme. Ionic liquid can be designed accordingly.

Natural Product Extraction & Processing H2O 0.15 Et2O 1.2 CHCl3 3.3 BMIm PF6 77.6 Morphine (free base) – solubility (gL-1) Solubility of natural products in ionic liquids often very much higher than in conventional solvents. Can be used to extract high concentrations from natural sources, using small extractor volumes.

Natural Product Extraction & Processing Dissolution of penicillin in ionic liquids – varies between solvent formulations Solubility limit ranges from 20-270g/L Can design solvent to optimize extraction efficiency

Natural Product Extraction & Processing With support from MMV, Bioniqs has developed ionic liquids for artemisinin extraction, from both wet and dry plant material. Extraction up to 4 times faster than in hexane Solubility in ionic liquids up to 110 gL-1 @ 25°C. Artemisinin may be recovered by adding water, precipitating and filtering off. Residual solvent content of recovered artemisinin <0.001%. Solvent may then be recycled and re-used Image provided courtesy of www.pp.clinet.fi

Decontamination & Cleaning Solubility of two nitroaromatic pollutants in ionic liquids compared to water. Ionic liquid may be designed to dissolve one compound even in presence of structurally similar materials.

Decontamination & Cleaning ILs designed to remove specified contaminant (amoxicillin) All ILs tested dissolve contaminant to >300g/L; several to >500g/L

Ecotoxicity and Biodegradation ILs have been widely touted as “green” solvents recyclable non-flammable non-volatile BUT early imidazole-based ILs were poorly biodegradable toxic to micro-organisms may form hazardous hydrolysis products Biodegradability must be “built in” to an ionic liquid at the design stage.

Biodegradation of Ionic Liquids Ordinary soil organisms can grow on Bioniqs ionic liquids and completely degrade them within 48 hours (above). Degradation products are water, carbon dioxide and ammonium ion. No significant toxic effects at concentrations below 80mM.

Summary Ionic liquids are not a panacea, but can offer major benefits in certain key areas:- Can now compete on comparable cost footing with conventional solvents for bulk applications. Reduced pollution and waste Improved process safety Enhanced efficiency of reaction and extraction processes Facilitate improved processes. Increased solubilities – smaller volumes Improved turnover rates and yields Stabilization of sensitive molecules Application to poorly soluble substrates

Acknowledgements

Further Information Bioniqs Ltd. Biocentre York Science Park Heslington York YO10 5DG United Kingdom 0(+44)1904 561538 office@bioniqs.com www.bioniqs.com