Aqueous biphasic systems based on deep eutectic solvents for extraction of biopharmaceuticals in downstream processing

The increasing global demand for biopharmaceuticals calls for scalable and cost-efficient purification strategies. Downstream processing remains a major challenge, accounting for up to 80 % of total production costs and high solvent-related energy consumption. In this context, the present work investigates aqueous biphasic systems (ABS) based on deep eutectic solvents (DES), to reduce both purification costs and the environmental impact of purification methodologies. A series of DESs composed of tetraalkylammonium-based salts and choline chloride were prepared, characterized, and evaluated for their efficiency in extracting γ-globulins, employed here as monoclonal antibody (mAb) surrogates. The system was validated for rituximab (RTX) extraction in PBS (pH 7.4) with extraction efficiencies > 87.0 % and relative standard deviations < 6.4 %, demonstrating the accuracy and repeatability of the method. The [N₄₄₄₄⁺][Cl^-] : 3[Gly]-based ABS also enabled efficient RTX extraction directly from cell culture media, highlighting the DES-based ABS potential as a sustainable alternative for downstream biopharmaceutical purification.