Boronic acid-functionalized agarose affinity chromatography for isolation of tropomyosin in fishes

Tropomyosin is now receiving increasing attention because of its significant allergenic activity in various fishery products but its simple and effective isolation still remains a challenging task.

An agarose-based boronate affinity chromatography was produced for the first time to isolate tropomyosin in various fishery products using 3,5-difluoro-4-formyl-phenylboronic acid as the functional monomer, tris(2-aminoethyl)amine as the multi-branched ligand, and agarose gel particles as supporting materials. The agarose concentration, binding pH, and the concentration of elution buffers demonstrated significant effects on separation performance. Under optimized conditions, the purity of the isolated tropomyosin was higher than 90%, with the column adsorption capacity over 1.85 mg mL^-1 and the enrichment efficiency over 65%. Such efficiency was also validated with different fish samples including Paralichthys olivaceus, Thunnusthynnus, Oreochromis spp., and Lophius litulon.

In comparison with conventional methods, the established affinity chromatography demonstrated excellent biocompatibility (without involving any organic solvent), better speed (from at least 1-2 days to 3-4 h), and simplicity (from at least five steps to three steps). This suggests that it is a novel and promising technique for the isolation of tropomyosin and other glycoproteins (including most allergens) in foodstuffs. © 2019 Society of Chemical Industry.