Principle of preparation of biological macromolecules
I. Dialysis and Ultrafiltration
1, dialysis
It is a method in which biomacromolecules such as proteins cannot be purified through a semipermeable membrane. Desalting dialysis is the most widely used dialysis method. The salt-containing biomacromolecule solution is placed in a dialysis bag, and the bag mouth is placed in a large container filled with distilled water, and stirring is continued to keep the distilled water flowing. After a period of time, the small molecule salts enter the distilled water through the semipermeable membrane to balance the salt concentration inside and outside the membrane. If the liquid in the large container is replaced several times during the dialysis process, the purpose of desalting the biomacromolecule solution can be achieved. Balanced dialysis is also one of the commonly used dialysis methods. The method comprises the steps of: loading a dialysis bag containing a biological macromolecule into a large container containing a certain concentration of a salt solution or a buffer solution, and performing dialysis, the salt concentration inside or outside the bag or the pH of the buffer is uniform, thereby controllingly changing the dialyzed solution. Salt concentration or pH. If the dialysis bag is placed in a high concentration water-absorbent polymer solution, the water in the solution in the dialysis bag is quickly absorbed by the extra-package polymer, thereby achieving the purpose of concentrating the liquid in the bag. This method is called "reverse dialysis." Polypolymers which can be used for reverse dialysis include polyethylene glycol (PEG), polyvinyl pyrrolidone (PVP), dextrose, sucrose and the like. There are many types of semi-permeable membranes for dialysis, and cellophane, cotton glue, animal film, and tissue paper can be used to make dialysis bags. Shanghai Chuangsai Technology provides polyvinylpyrrolidone 40000, PVP40000 (Polyvinylpyrrolidone), commodity number: B11000373-500g, the price is 400 yuan.
2, ultrafiltration method
The macromolecular solution is filtered (normal pressure, pressurized or decompressed) by using a microporous membrane having a certain pore size, so that the macromolecule remains in the solution above the ultrafiltration membrane, and the small molecular substance and water are filtered out. This achieves the purpose of desalting, buffer replacement or concentration. This method of separating large molecules and small molecules by ultrafiltration membrane filtration is called ultrafiltration.
Second, decompression concentrated freeze-drying
Because biomacromolecules are usually unstable to heat and highly variability, concentrated and dried biomacromolecules cannot be heated and evaporated. Therefore, concentration under reduced pressure and freeze-drying have become the concentrated drying techniques commonly used in the preparation of biomacromolecules. The lyophilization method is to freeze the protein solution on the wall of the container while allowing the liquid to sublime in a vacuum to obtain a lyophilized sample. The product obtained by freeze-drying can maintain the natural properties of the biomacromolecular material, and has the characteristics of being loose and easy to dissolve, and is convenient for preservation and application. This is the most common and best way to preserve biomacromolecules.
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