Column chromatography is a versatile purification method used to separate compounds in a solution. A solution mixture is carried by a solvent through a column containing an adsorbent solid, called the stationary phase. Thus, each compound exits the column at a different time.

In column chromatography, the stationary phase, a solid adsorbent, is placed in a vertical glass (usually) column. The mobile phase, a liquid, is added to the top and flows down through the column by either gravity or external pressure.

The stationary phase or adsorbent in column chromatography is a solid. The most common stationary phase for column chromatography is silica gel, the next most common being alumina. Cellulose powder has often been used in the past.

Chromatography is used to separate mixtures of substances into their components. All forms of chromatography work on the same principle. They all have a stationary phase (a solid, or a liquid supported on a solid) and a mobile phase (a liquid or a gas). The silica gel (or the alumina) is the stationary phase.

If the eluent is very polar relative to your compound, it will dissolve your sample and the sample will move with the mobile phase. Overall, the eluent and your sample will compete for a space (an active site) on the adsorbent (stationary phase) coated on the TLC plate.

There are four main types of chromatography. These are Liquid Chromatography, Gas Chromatography, Thin-Layer Chromatography and Paper Chromatography.

Chromatography is based on the principle where molecules in mixture applied onto the surface or into the solid, and fluid stationary phase (stable phase) is separating from each other while moving with the aid of a mobile phase.

Top 12 Types of Chromatographic Techniques | Biochemistry

• Type # 1. Column Chromatography:
• Type # 2. Paper Chromatography:
• Type # 3. Thin Layer Chromatography:
• Type # 4. Gas-Liquid Chromatography (GLC) or Simply Gas Chromatography (GC):
• Type # 5. High Performance Liquid Chromatography:

What is the principle of TLC? TCL is based on the principle of separation through adsorption type. The separation relies on the relative empathy of compounds towards the mobile phase and stationary phase.

TLC uses two different phases, stationary and mobile, where the stationary phase is the very very polar silica gel and the less polar mobile phase.

Advantages And Disadvantages Of Thin Layer Chromatography

• An easy method of separation of the components.
• In this technique, fewer types of equipment are used.
• All components of UV light is achievable to visualize.
• The non-volatile compounds can be separated by this method.

Silica gel is by far the most widely used adsorbent and remains the dominant stationary phase for TLC. The surface of silica gel with the highest concentration of geminal and associated silanols is favored most for the chromatography of basic compounds because these silanols are less acidic.

Rf is a fraction. It is the ratio of how far a substance travels up the chromatography paper compared to the distance the solvent has travelled. This means that it must be less than 1. [If this sounds general, it is because changing any of the experimental conditions will change the Rf value.

Silica and alumina are both polar adsorbents so the more polar components in the mixture to be separated are retained more strongly on the stationary phase and are therefore eluted from the column last. Typically, 70–230 silica gel is used for gravity columns and 230–400 mesh for flash columns.

Silica gel is a polar adsorbent. This allows it to preferentially adsorb other polar materials. When it comes to polarity, materials interact more with like materials. This principle is particularly important to many laboratories, which use silica gel as the stationary phase for column chromatography separations.

It should be noted that silica gel is highly polar and is capable of hydrogen bonding. Consider the side-on view of the development of a TLC plate below. As the solvent travels up the plate, over the spot, an equilibrium is set up, as development solvent competes with the TLC plate for the solute.

Benzene is a nonpolar molecule due to the presence of many nonpolar carbon-hydrogen bonds spaced out in equal proportion around the molecule ring. It is a symmetrical molecule in which all the bond dipoles cancel.

The stationary phase i.e. silica is very polar in nature, while the solvent is less polar compared to silica.

Stationary phase, in analytical chemistry, the phase over which the mobile phase passes in the technique of chromatography. Typically, the stationary phase is a porous solid (e.g., glass, silica, or alumina) that is packed into a glass or metal tube or that constitutes the walls of an open-tube capillary.

For example, the positive side is attracted to the negative side of another molecule (opposites attract). The larger the charge difference, the more polar a molecule is. You will find that as you increase the polarity of the solvent, all the components of the mixture move faster during your chromatography experiment.

Silica gel, the most commonly used stationary phase, has the empirical formula SiO2. However, at the surface of the silica gel particles, the dangling oxygen atoms are bound to protons. The presence of these hydroxyl groups renders the surface of silica gel highly polar.

The mobile phase flows through the stationary phase and carries the components of the mixture with it. The silica gel (or the alumina) is the stationary phase. The stationary phase for thin layer chromatography also often contains a substance which fluoresces in UV light – for reasons you will see later.

In general, the adsorptivity of compounds increases with increased polarity (i.e. the more polar the compound then the stronger it binds to the adsorbent). Non-polar compounds move up the plate most rapidly (higher Rf value), whereas polar substances travel up the TLC plate slowly or not at all (lower Rf value).

Aspirin contains one carboxylic and one ester group. Ibuprofen contains one ester group only. As Caffeine contains most number of heteroatoms in the functional group and it has lowest Rf value. So it is most polar.

Acetaminophen has two hydrogen bonds and one dipole-dipole making it more polar than Aspirin, Phenacetin, and Ibuprofen.

1 Expert Answer. The difference between the two is that aspirin is salicylic acid with the free OH of salicylic acid having been esterified. Both compounds still have the -COOH group. The -OH group is more polar than the ester so salicylic acid is more polar than aspirin.

Therefore, the aspirin and caffeine in the filtrate can be separated by extraction either with acid, which will remove the caffeine as a water-soluble salt, or by extraction with base, which will remove the aspirin as a water-soluble salt.

Acetaminophen has two hydrogen bonds and one dipole-dipole making it more polar than Aspirin, Phenacetin, and Ibuprofen. Lastly, caffeine has the lowest R F value because it has the capability to ionize and form a hydrogen bond with the lone pair of electrons on the nitrogen.

Aspirin belongs to the group of medicines known as salicylates and to the group of medicines known as anti-inflammatory analgesics. The sodium bicarbonate in aspirin, sodium bicarbonate, and citric acid is an antacid. It neutralizes stomach acid by combining with it to form a new substance that is not an acid.

The solubility of caffeine decreases in the order of chloroform, dichloromethane, acetone, ethyl acetate, water, methanol, ethanol, and carbon tetrachloride. Thus, chloroform is a better solvent to separate and purify caffeine from solutions.