Effect of the spacer of gemini surfactants on reverse micellar extraction of bovine serum albumin

Abstract

Reverse micellar extraction has the potential to overcome the hurdles associated with the conventional separation and purification procedures for proteins and hence has been considered as an alternative to conventional techniques. The surfactants used in protein extraction so far have been almost exclusively di-2-ethylhexyl sodium sulfosuccinate and cetyl trimethyl ammonium bromide. In the present paper, we extracted bovine serum albumin (BSA) by using a kind of novel surfactant – gemini surfactant C_m-s-C_m·2Br (m = 12 and 16, and s = 2, 5, 8, and 12). Compared with conventional surfactant reverse micelles, all the gemini surfactant reverse micelles show superior performance. The length and flexibility of spacer s of C_m-s-C_m·2Br contribute to the advantages of gemini surfactant reverse micelles, and increasing hydrophobic alkyl chain length weakens the effect of the spacer. As a result, the minimum content ([surfactant]_min) of the surfactant needed for transferring protein into an organic phase is increased in the order of s being 5, 8, 12 (2) sequentially and [surfactant]_min for C₁₆-s-C₁₆·2Br is higher than that for C₁₂-s-C₁₂·2Br. The loading capacity of C₁₆-s-C₁₆·2Br reverse micelle is lower than that of C₁₂-s-C₁₂·2Br reverse micelle and C₁₂-12-C₁₂·2Br reverse micelle can load the maximum amount of protein. In the backward extraction process, gemini surfactants with s being 8 and 12 are much more efficient than the other gemini surfactants at pH 4.3. With C₁₂-8-C₁₂·2Br and C₁₂-12-C₁₂·2Br reverse micelles, BSA can be recovered under neutral or basic conditions given a sufficient amount of salt present (while a pH of ca. 4.3 is prerequisite for the recovery of BSA from conventional surfactant reverse micelles) and the efficiency with C₁₂-12-C₁₂·2Br reverse micelle is higher. Since so far the reports about the effect of surfactant structure on protein extraction have been limited, this study should help us know more about how to optimize the surfactant structure.