Innovative separation methods in bioprocessing 187 Reverse ·““ Protein Fig. 7.6. Protein partition into reverse There are two techniques for transferring proteins into the micellar phase. The m widely used method involves extraction of the protein with a biphasic liquid system,i.e iquid-liquid extraction. One phase is the aqueous solution of the protein, and the other the organic micellar solution, usually in equal volume. By gently shaking the two phases, he protein partitions from the aqueous into the micellar phase. In the second method solid state extraction of the protein, the protein powder is suspended in the micellar phase and gently stirred The protein solubilised in the reverse micellar solution can be transferred back into an aqueous solution, by contacting the micellar solution with an aqueous solution containing a high concentration of a particular salt(KCl, CaCl2), which has the capability to ex change with the protein in the micelles The basic idea is that the process of protein extraction by reverse micelles can be made pecific (i.e. tailored to a specific protein) and efficient (i.e. high extraction yield) by ontrolling the micellar parameters such as the water content, the type and concentration of surfactant, the type and concentration of salt, and the pH Leser et al.(1986)examined the transfer of ribonuclease-A, lysozyme, trypsin an pepsin, monitoring the protein concentration and the concentration of water found in the organic phase. It was observed that the transfer of water is generally moderate(beloy 4%), whereas, under certain conditions, the protein is quantitatively transferred. This fact demonstrated that the transfer of the protein into the micellar phase is not a passive process, i.e. is not simply due to the fact that water is transferred and with it the protein The conclusion was that there is a thermodynamic driving force for the hydrophilic protein to leave the aqueous environment and to transfer into the reverse micelles. In other words, it seems that under certain conditions the protein-reverse micelle complex is energetically favoured above the free protein and empty reverse micelles. Interactions can be electrostatic, when surfactants with charged head groups are used, or hydrophobic with the surfactant interface or the apolar solventInnovative separation methods in bioprocessing 187 Organic phase 4V& - .t nv Reverse 4 ' L micelle ?fiC%4k/ h 3 nc L.UVVaUL. V UdU UU!iiCUY u Protein 0 a a Aqueous phase Fig. 7.6. Protein partition into reverse micelles. There are two techniques for transferring proteins into the micellar phase. The most widely used method involves extraction of the protein with a biphasic liquid system, i.e. liquid-liquid extraction. One phase is the aqueous solution of the protein, and the other the organic micellar solution, usually in equal volume. By gently shaking the two phases, the protein partitions from the aqueous into the micellar phase. In the second method, solid state extraction of the protein, the protein powder is suspended in the micellar phase and gently stirred. The protein solubilised in the reverse micellar solution can be transferred back into an aqueous solution, by contacting the micellar solution with an aqueous solution containing a high concentration of a particular salt (KC1, CaC12), which has the capability to ex￾change with the protein in the micelles. The basic idea is that the process of protein extraction by reverse micelles can be made specific (Le. tailored to a specific protein) and efficient (Le. high extraction yield) by controlling the micellar parameters such as the water content, the type and concentration of surfactant, the type and concentration of salt, and the pH. Leser et al. (1986) examined the transfer of ribonuclease-A, lysozyme, trypsin and pepsin, monitoring the protein concentration and the concentration of water found in the organic phase. It was observed that the transfer of water is generally moderate (below 4%), whereas, under certain conditions, the protein is quantitatively transferred. This fact demonstrated that the transfer of the protein into the micellar phase is not a passive process, i.e. is not simply due to the fact that water is transferred and with it the protein. The conclusion was that there is a thermodynamic driving force for the hydrophilic protein to leave the aqueous environment and to transfer into the reverse micelles. In other words, it seems that under certain conditions the protein-reverse micelle complex is energetically favoured above the free protein and empty reverse micelles. Interactions can be electrostatic, when surfactants with charged head groups are used, or hydrophobic with the surfactant interface or the apolar solvent