The Biochemistry Online – Free Online Lectures for Medical, Dental and Allied Health Sciences students by the Biochemistry Club.
Basic Separation Techniques
- Centrifugation
- Filtration
- Dialysis
1. Centrifugation
Centrifugation is a process in which centrifugal force is used to separate solid matter from a liquid suspension. The centrifuge carried out this action. It consists of a head (or rotor) and carries (or shields) which are attached to a vertical shaft of a motor and are enclosed in a metal covering with an ON/OFF switch. The speed is expressed in revolutions per minute (rom).
Few Applications of Centrifugation
- Separation of thick precipitates from solution.
- Separation of serum from clotted blood.
- Precipitating proteins from serum during colorimetric estimation of serum urea/creatinine. Trichloracetic acid is added to the serum sample, which denatures proteins and precipitates them. The precipitate is separated by centrifugation. The protein free filtrate is then used for further analysis.
2. Filtration
Filtration can be used instead of centrifugation for separation of solids from liquids. The filter material is made of paper. Cellulose and its derivatives, polyester fibers, glass etc. Traditionally, filter paper is folded in a manner that allows it to fit in a funnel by folding into its fourth.
The filter paper differs in pore size and should be selected according to the specific separation need and the associated flow rate for given liquids. The filter paper should not be used for strong acids or bases. The liquid that passes through the filter paper is called the filtrate. The precipitate that remains on the paper is called the residue.
3. Dialysis
Dialysis is the process of separating molecules in solution by the difference in their rates of diffusion through a semipermeable membrane. Typically, a solution of several types of molecules is placed into a semipermeable dialysis bag. Such as a cellulose membrane with pores. The sealed dialysis bag is placed in a container of a different solution or pure water. Molecules small enough to pass through the membrane (often water, salts and other small molecules) tend to move into or out of the dialysis bag. In the direction of decreasing concentration. Larger molecules (often proteins, DNA or polysaccharides) that have dimensions significantly greater than the pore diameter are retained inside the dialysis bag. One common reason for using this technique would be to remove the salt from a protein solution.