Electrophoresis
- Electrophoresis is an analytical technique which separates ions by placing them in an electrical field
- This method is often used in biochemical analysis to identify and purify proteins
- A sample of amino acids is placed between two oppositely charged electrodes
- The positively charged ions will move towards the negative electrode
- The negatively charged ions will move towards the positive electrode
- The rate (how fast) at which the ions move towards the electrodes depends on:
- The size of the ions: larger ions move more slowly
- The charge of the ions: highly charged ions move more quickly
- An electropherogram is the series of bands which are observed on the paper or gel after electrophoresis has occurred
- Each band in the electropherogram corresponds to a particular species
Separating mixtures of amino acids by varying the pH
- The charge on the amino acid ions depends on the pH of the solution
- The movement of the ions to the electrodes during electrophoresis will therefore be affected by the pH
- Consider a sample which consists of a mixture of three amino acids at pH 7
- Amino acid A: lysine, side-chain is positively charged
- Amino acid B: glycine, side chain is neutral
- Amino acid C: glutamic acid, side chain is negatively charged
The different amino acids within the sample
The sample consists of a mixture of three amino acids which are separated using electrophoresis
- The amino acids in this mixture can be separated by electrophoresis
- Amino acid C will move towards the positive electrode
- Amino acid B will remain in the well where the sample is applied to the gel
- Amino acid A will move towards the negative electrode
- Since glutamic acid is larger than lysine, it will travel towards the positive electrode at a slower rate compared to lysine
Separation of a mixture of amino acids by electrophoresis
During electrophoresis, positive amino acids move to the negative electrode and negative amino acids move to the positive electrode
