The principle of chromatography using silica gel is based on the differential affinity of substances for the stationary phase (silica gel) and the mobile phase (eluent or solvent). Chromatography is a separation technique used to separate and analyze mixtures of different compounds based on their physical and chemical properties.
Silica gel is a porous and adsorbent material composed of silicon dioxide (SiO2). It has a high surface area with numerous small pores and a large internal surface, which allows it to adsorb various compounds. The principle of chromatography using silica gel involves the following steps:
1. Sample application: The sample mixture is applied as a small spot or line near the bottom of a solid support, such as a glass plate or a chromatography paper. In the case of column chromatography, the sample is loaded onto the top of the silica gel column.
2. Development of chromatogram: The solid support (e.g., chromatography paper) is then placed vertically in a container, and a suitable solvent (mobile phase or eluent) is allowed to move through the solid support via capillary action. As the solvent moves up the paper or down the column, it carries the sample compounds along with it.
3. Separation of compounds: As the solvent moves through the silica gel, the various components of the sample mixture interact differently with the stationary phase (silica gel) based on their chemical and physical properties. Compounds that have a higher affinity for the silica gel will be retained more strongly and will move more slowly through the stationary phase. On the other hand, compounds that have a lower affinity for the silica gel will be less retained and will move more rapidly.
4. Visualization and analysis: After the solvent has moved a certain distance (usually measured from the starting point of the sample application), the chromatogram is taken out, and the separated compounds are visualized. This can be done by exposing the chromatogram to ultraviolet (UV) light, using chemical reagents that react with specific compounds, or by staining the chromatogram. The distance traveled by each compound is then measured and can be used to calculate the retention factor (Rf), which is a characteristic value for each compound under specific chromatographic conditions.
By comparing the Rf values of the separated compounds with known standards or reference compounds, the components of the sample mixture can be identified. Silica gel chromatography is commonly used in analytical chemistry and is particularly useful for separating and purifying organic compounds based on their polarity and molecular interactions with the silica gel surface.