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Background of high value-added protein separation, purification, supercritical anti-solvent (SAS) precipitation technology with traditional organic solvent precipitation combined to form the with pressurized CO 2 from the mixed solvent precipitation novel protein separation methods. The method has a friendly environment, integration, in line with the trends of the biological separation techniques, has potential practical value. First, create a set for SAS precipitation separation of the experimental device, milk protein mock objects, effects of anti the solvent CO 2 precipitated protein in the mixed solvent the Super (near) critical cosolvent - - ethanol protein precipitation. The results show that: the precipitation of proteins in the milk in the presence of a critical temperature (32 ℃) and critical pressure (2.76MPa), and contribute to increased CO 2 the solubility when ethanol was added to the aqueous system, and enhance the ultra (nearly) critical CO 2 precipitation, therefore, ethanol and CO 2 milk protein exhibit double precipitation of the critical precipitation temperature and the critical precipitation pressure drop. When the ethanol concentration reached 30 to 40%, the critical precipitation temperature and the critical precipitation pressure is reduced to below 30 ° C and 0.98 MPa. This result also shows that the operability of the apparatus of our experiments, and SAS process is for the aqueous solution was mixed as the main solvent in the protein precipitation feasibility. The situation precipitated in ethanol - water mixed solvent cellulase, alpha-amylase, acid protease, alkaline protease super (nearly) the critical CO 2 visits and enzyme activity Determination of the changes that different enzyme precipitation phenomena and enzyme activity variation in the experimental conditions and different, and summed up containing protein (enzyme) mixture of ethanol - aqueous solution the ultra (near) the critical CO 2 the three preconditions processing, the the ultra (near) critical CO 2 processing temperature of cellulase, acid protease, alkaline protease CO 2 pressure The operating range of the concentration of ethanol. Third, through the milk, cellulase, acid protease, alkaline protease, etc. in the mixed solvent for ultra (near) supercritical CO 2 precipitated, obtained under experimental conditions of the total solution data on the solubility of the protein. Fourth approximation of the experimental system for the CO 2 -C 2 H 5 OH-H 2 O three yuan The Department, through the ternary phase equilibrium studies can be used for practical engineering calculations MESV equations and corresponding methods of checking credible under the experimental conditions of each component of the phase equilibrium composition data. First established under the super critical conditions of the (near) the solubility of the protein in the liquid phase mathematical model, and projection results using MESV Equation, successfully associated with the solubility data of the total protein in milk protein and cellulase the model associated with the dissolution rate of milk protein and cellulase system, the average relative error (AARD) were 11.5% (303.15K), 18.1% (308.15K) and 7.0% (303.15K), 7.7% (308.15K ). Finally, the first raised with pressurized CO 2 a new method to precipitate the protein from the mixed solvent used in separation and purification of cellulase and acidic protease, alkaline protease process of removing contaminating proteins. Analysis of the separation and purification of the fiber Zhejiang University doctorate the righteous Summary luciferase, the introduction of fractional precipitation chart, the results show that the maximum purification factor and yield up to 6.3 and 62%, respectively, eliminating the need for desalination , freeze drying process. The acid protease, alkaline protease better the effect of the removal of contaminating proteins. Therefore, the super-critical (near) * 0. Precipitation separation and purification of proteins, not only in theory, there is a further and explore the value, and also has a good prospect.
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