In recent years, a new class of ‘green’ solvents named deep eutectic solvents (DESs) has appeared¹. DESs are obtained by simple thermal mixing of two compounds, which results in the formation of a eutectic solution with a melting point lower than those of the individual components. Due to the properties of DESs such as high thermal stability, low toxicity, biodegradability, conductivity, and others, DESs or their mixtures with a buffer solution can be used in various fields of chemistry, including biocatalysis. However, there are only a few studies describing the use of oxidoreductases for the synthesis in DES-buffer mixtures.
Laccase (p-benzenediol:oxygen oxidoreductase, EC 1.10.3.2) belongs to the ‘blue’ oxidases; it catalyzes the oxidation of various organic compounds with molecular oxygen. This enzyme attracts much attention as a catalyst for fine organic synthesis. One of the promising laccase substrates is aniline, the oxidation of which results in the formation of polyaniline, the most important compound among conducting polymers. Polyaniline and its functionalized derivatives possess antimicrobial properties and could be used to create film coatings, protecting various surfaces from bacterial contamination².
In this work, DES betaine-glycerol (molar ratio 1:1) was used as a co-solvent for efficient laccase-catalyzed template copolymerization of aniline and 3-aminobenzoic acid. The fungal laccase Trametes hirsuta retained ~50% activity in a DES/buffer mixture (60/40 vol.%) after 120 h incubation. Enzymatic reactions conducted in DES/buffer mixtures met the requirements of sustainable chemistry. The synthesized copolymer was characterized by UV-Vis and FTIR spectroscopy, atomic force microscopy, and cyclic voltammetry.
The copolymer strongly inhibited the Staphylcocus aureus and Escherichia coli growth. The minimum growth inhibitory concentration (MIC, the lowest concentration of a compound that inhibits bacterial growth for 24 hours) was determined by the standard serial two-fold dilution method in LB nutrient medium. It was found that the MIC values of copolymer against gram-positive S. aureus and gram-negative E. coli bacteria were 0.125 and 1.0 mg/mL, respectively.
Thus, it has been shown that betaine-based deep eutectic solvents are promising media for the polymerization of various laccase substrates, and functionalized polyanilines synthesized enzymatically can be used to create antimicrobial coatings.