engleski

Deep eutectic solvents aqueous two-phase system based protein extraction in a microextractor

Nowadays, clean, sustainable and efficient extraction methods for downstream processing of biomolecules are of particular interest. The basic idea is to develop an effective and versatile green technology that allows high extraction efficiency at minimal cost and least negative impact on the environment. The aqueous two-phase system (ATPS) is one of these technologies. ATPSs are liquid-liquid systems usually formed by mixing certain polymers (e.g., PEG) with salts, detergents, ionic liquids, and short-chain alcohols. Recently, the use of deep eutectic solvents (DES) instead of polymers to promote extraction in ATPS has been reported. The preparation of DESs is very simple and cheap. Moreover, DES is referred to as a green solvent because it can be composed of natural components such as sugars, alcohols, and amides, and is biodegradable, nontoxic, and low-volatile. To use DESs for efficient extraction of various biomolecules, their composition must be optimized. Moreover, in order to intensify the extraction process, batch extraction is increasingly replaced by continuous extraction, and the extraction process itself is carried out in microsystems. In this study, ATPS based on salt and DES was used for the extraction of proteins (laccase and lipase). At the beginning of the study, different DES were prepared and characterized. In the next step, the influence of ATPS composition and salt concentration on extraction efficiency was investigated in a batch extraction performed for 30 minutes. The highest extraction efficiency of 94.70% was obtained for lipase by using the ATPS composed of K2HPO4 and betaine:urea DES. The highest extraction efficiency for laccase of 67.21% was achieved for the ATPS composed of K2HPO4 and choline chloride:poly(ethylene glycol):zinc chloride DES. For selected ATPSs additional process intensification was achieved by extraction performed in a microsystem. The influence of residence time, temperature and channel diameter on extraction efficiency was investigated. For the enzyme lipase, an extraction efficiency of 98.50% was achieved for a residence time of 30 s. Once the best process conditions were defined, the method was used for protein extraction from crude samples. Finally, an integrated system with continuous protein extraction and phase separation on a microscale was developed.

extraction ; protein ; deep eutectic solvent ; aqueous two-phase system ; aqueous two-phase system ; microextractor

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