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Graphical abstract Highlights • GFP recovery and LPS removal on C 10 E 4 /buffer + salts-based ATPMS were studied. • C 10 E 4 /buffer + salts promoted GFP partitioning to the micelle-poor phase (K GFP > 7). • C 10 E 4 /buffer + salts-based ATPMS removed LPS to the micelle-rich phase (REM LPS > 98%). • High loads of LPS (>106 EU/mL) increased the GFP partition into micelle-rich phase. • C 10 E 4 /buffer + salt are able to separate selectively the GFP from contaminant LPS. Abstract Lipopolysaccharide endotoxins (LPS) are the most common contaminant pyrogenic compounds found in intracellular recombinant biomolecules purified from Gram-negative bacteria, such as Escherichia coli. Thus, the purification downstream processing should guarantee the effective removal of LPS from the final bioproduct, particularly, therapeutic biopharmaceuticals. Aqueous two-phase micellar systems (ATPMS) appear to be an excellent strategy to purify recombinant biopharmaceuticals from the cell lysate of E. coli , reducing high LPS concentrations. In order to demonstrate the effectiveness of ATPMS as a biopharmaceutical purification platform, the influence of inorganic salt electrolytes (NaCl, Li 2 SO 4 , KI, or KNO 3) on the partitioning of green fluorescent protein (GFP) and LPS removal using ATPMS composed of n -decyl tetraethylene oxide (C 10 E 4) was evaluated. The impact of different LPS concentrations on GFP partitioning was also studied. The addition of electrolytes (i.e., NaCl or Li 2 SO 4) to the C 10 E 4 -based ATPMS have reduced the phase forming temperatures to very mild conditions (ca. 17.00 and 13.00 °C, for NaCl and Li 2 SO 4 , respectively). The selective partitioning ability of the proposed ATPMS was further demonstrated, where a complete removal of the LPS from the micelle-poor phase (REM LPS > 98%) and a preferential GFP recovery (REC GFP = 97%, K GFP > 7) to the micelle-poor phase was obtained. The GFP partitioning was even enhanced by increasing LPS loading (104–106 EU/mL), probably due to the formation of mixed micelles between LPS and C 10 E 4. It is here demonstrated that a C 10 E 4 /buffer + salt-based ATPMS can be a useful and straightforward platform for the removal of endotoxin contaminants and the purification of recombinant biopharmaceuticals from E. coli cell lysates. [ABSTRACT FROM AUTHOR]

• Partition of curcumin (CCM) using ATPMS composed of Pluronic F68 + ILs was studied. • Cholinium ILs decreased the phase separation temperature of Pluronic F68/buffer. • The phase separation decreased with the increase of the IL anion hydrophobicity. • CCM was preferentially partitioned into polymeric micelles (PMs)-rich phase. • ILs-based ATPMS were very selective to recover CCM from crude extract of C. longa L. Novel aqueous two-phase micellar systems (ATPMS) composed of Pluronic F68, a triblock amphiphilic copolymer, and cholinium-based ionic liquids (ILs) were formulated and applied for separation/purification of curcumin (CCM). CCM stability in the presence of ATPMS components was also evaluated. CCM is stable up to 24 h in copolymer (1.0 – 10.0 wt%) and ILs (0.1 – 3.0 M) aqueous solutions. Very mild phase separation conditions (close to room temperature) were achieved by adding cholinium ILs to the Pluronic F68 + McIlvaine buffer at pH 6.0 solution. The decrease of cloud-point temperature is dependent on the relative hydrophobicity of IL anion, [Hex]− > [But]− > [Pro]− > [Ac]− > Cl−. ATPMS composed of more hydrophobic ILs ([Ch][Hex] > [Ch][But] > [Ch][Pro]) are most efficient in the partition of commercial CCM into polymeric micelles (PMs)-rich phase. The best ATPMS (0.70 M [Ch][But] and 0.60 M [Ch][Hex]-based ATPMS) were then used to purify CCM from a crude extract of Curcuma longa L. Both systems were very selective to separate CCM from protein-based contaminants (selectivity values ≥ 25; purification yields ≥ 12-fold). Pluronic F68-based ATPMS are promising for selective separation of hydrophobic biomolecules by using cholinium-based ILs as adjuvants to adjust phase separation temperatures and biomolecules' partition. [ABSTRACT FROM AUTHOR]