Protein extraction using non-ionic reverse micelles of Span 60

In protein extraction using non-ionic Span 60 reverse micelles, the critica l micelle concentration (CMC) of a Span 60/IPA/hexane system was at Span 60 concentration of 3.5 g l(-1). At concentrations higher than the CMC, cytoc hrome c was extracted into the Span 60 reverse micelles as judged from UV s pectra of the Span 60/IPA/hexane solution after it contacted with the prote in aqueous solution. In extractions of cytochrome c and lysozyme using this system, forward extraction was found to be pH-dependent, with high extract ion percentages being obtained at neutral pHs. The forward extraction perce ntage was drastically reduced by only a small increase in the buffer concen tration (even up to 0.1 M). In cytochrome c extraction using various means of phase contacting, high extraction percentages were obtained with rotary and reciprocal shaking, but in well-mixed extraction by magnetic stirring, extraction percentages were much lower. Addition of alcohol, especially iso propyl alcohol, to the micellar organic phase enabled the backward extracti on of cytochrome c to be achieved without the formation of insoluble aggreg ates. (C) 1998 Elsevier Science S.A. All rights reserved.