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Several features of this process should be considered. The nature of the salt is all-important.
One must consider physical characteristics of the salt such as solubility, besides its
effectiveness in causing precipitation. The most effective salts are those with multile-charged
anions such as sulfate, phosphate, and citrate; the cation is relatively less important. Saltingout ability of anions follows the hofmeister series, which for some common anions is SCN-,
CLO4-, NO3-, Br-, Cl-, acetate-, SO42-, PO43-, Although phosphate at neutral pH consists of
a mixture of HPO42- and H2PO4- ions, which are less effective than PO43-, As far as cations
are concerned, monovalent ions should be used, with NH4+ >K+ >Na+ in precipitation
effectiveness (36). Of the common inexpensive salt that are effective in causing precepitation,
sodium, potasium and ammonium sulfates, phospates, and citrates are attractive candidates.
Further considerations of the suitability are the cost of kilogram quantities of sufficient pure
salt, the solubility (several molar solutions are required), and the heat of solution. It is
undersirable to get heating while dissolving in the salt; also, a large Ho of solution means
large changes in solubility with temperature. Finally, the density of concentrated solution
should be considered the aggregate and the solvent (ef. Section 1.2).
Most potassium salts are ruled out on solubility grounds, though potasium phosphates can be
used. Sodium citrate is very soluble, and can be used also, but rarely offers any advantage
other than the possibility of use above pH 8- which is not possible with ammonium salts,
because of the buffering action of ammonia. Citrate cannot be used so effectively below pH 7
for the same reason-the strong buffering action of citrate ions. Sodium sulfate forms several
hydrates and has a complex solubility phase diagram, but it is not highly soluble at low
tempeature. It has been used in the purification of egg albumi, keeping the temperature
around 300C so that the salt is sufficiently soluble. Except when it is required to operate at a
high pH, one salt has all the advantages and no disadvantages for a typical protein;
ammonium sulfate.
The solubility of ammonium sulfate varies very little in the range 0-300C; a saturated
solution in pure water is approximately 4M. The density of this saturated solution is 1,235
g.cm-3, compared with a protein aggregate density in this solution is 1,29 g.cm-3 (ef. Section
1.2). in contrast, 3M. Potassium phosphate at pH 7,4 has a density of 1,33 g.cm-3, which
makes it, impossible to centrifuge off protein, since this is also the density of protein
aggregates in this solution. However, filtration is possible, as is adsorption to amphiphilic
matrices such as agarose (ef. Section 5,7). Because of extra density caused by other salts and
compounds present in crude extracts, precipitates in the range 3-4 M ammonium sulfate also
can sometimes be difficult to sediment
bila diperlukan untuk beroperasi pada pH tinggi, satu garam memiliki semua keuntungan dan
tidak ada kerugian untuk protein khas; amonium sulfat.
Kelarutan amonium sulfat bervariasi sangat sedikit di 0-300C kisaran; larutan jenuh dalam air
murni adalah sekitar 4M. Kepadatan larutan jenuh ini adalah 1,235 g.cm -3, dibandingkan
dengan protein kepadatan agregat dalam solusi ini adalah 1,29 g.cm -3 (ef. Bagian 1.2).
Sebaliknya, 3M. Kalium fosfat pada 7,4 pH memiliki kerapatan 1,33 g.cm -3, yang
membuatnya, mungkin untuk centrifuge off protein, karena ini juga kepadatan agregat protein
dalam larutan ini. Namun, filtrasi mungkin, seperti adsorpsi untuk matriks amphiphilic seperti
agarosa (ef. Bagian 5,7). Karena kepadatan ekstra yang disebabkan oleh garam lainnya dan
senyawa hadir dalam ekstrak minyak mentah, endapan di kisaran 3-4 M amonium sulfat juga
kadang-kadang bisa sulit untuk sedimen