ORGANIC cHEMISTRY - III

Experiment 2

APPARATUS/CHEMCIALS REQUIRED

Ethyl acetate, Petroleum ether, Mixture and their corresponding standard compounds, Silica Gel GF 254, Filter Paper, TLC

sheet, TLC chamber, Capillary, Iodine chamber, UV Chamber for visualisation of developed TLC

THEORY-

TLC is a simple, quick, and inexpensive procedure that gives the chemist a quick answer as to how many components are there in a mixture. TLC is also used to support the identity of a compound in a mixture when the Rf of a compound is compared with the Rf of a known compound (preferably both run on the same TLC plate).

A TLC plate is a sheet of glass, metal, or plastic which is coated with a thin layer of a solid adsorbent (usually silica or alumina). A small amount of the mixture to be analyzed is spotted near the bottom of this plate. The TLC plate is then placed in a shallow pool of a solvent in a developing chamber so that only the very bottom of the plate is in the liquid. This liquid, or the eluent, is the mobile phase, and it slowly rises up the TLC plate by capillary action.

As the solvent moves past the spot that was applied, an equilibrium is established for each component of the mixture between the molecules of that component which are adsorbed on the solid and the molecules which are in solution. In principle, the components will differ in solubility and in the strength of their adsorption to the adsorbent and some components will be carried farther up the plate than others. When the solvent has reached the top of the plate, the plate is removed from the developing chamber, dried, and the separated components of the mixture are visualized. If the compounds are colored, visualization is straightforward.

Usually the compounds are not colored, so a UV lamp is used to visualize the plates. (The plate itself contains a fluorescent dye which glows everywhere except where an organic compound is on the plate.)

Interactions Between the Compound and the Adsorbent

The strength with which an organic compound binds to an adsorbent depends on the strength of the following types of interactions: ion-dipole, dipole-dipole, hydrogen bonding, dipole induced dipole, and van der Waals forces. With silica gel, the dominant interactive forces between the adsorbent and the materials to be separated are of the dipole-dipole type. Highly polar molecules interact fairly strongly with the polar Si-O-H groups at the surface of these adsorbents, and will tend to stick or adsorb onto the fine particles of the adsorbent while weakly polar molecules are held less tightly. Weakly polar molecules generally tend to move through the adsorbent more rapidly than the polar species. Roughly, the compounds follow the elution order given above.

The Rf value

The retention factor, or Rf, is defined as the distance travelled by the compound divided by the distance travelled by the solvent. For example, if a compound travels 2.1 cm and the solvent front travels 2.8 cm, the Rf is 0.75:

The larger an Rf of a compound, the larger the distance it travels on the TLC plate. When comparing two different compounds run under identical chromatography conditions, the compound with the larger Rf is less polar because it interacts less strongly with the polar adsorbent on the TLC plate. Conversely, if you know the structures of the compounds in a mixture, you can predict that a compound of low polarity will have a larger Rf value than a polar compound run on the same plate.

PROCEDURE –

STEP 1 Prepare the TLC plate

-TLC plates used in the organic chemistry teaching labs are purchased as 20 cm x 20 cm sheets.

-Each large sheet is cut horizontally into plates which are usually 5 cm tall having width as per requirement; the more samples you plan to run on a plate, the wider it needs to be.

STEP 2 Spotting the samples:

• Mark a reference line about 1 – 2 cm from the bottom edge of the plate with the pencil gently

• The standards provided to you must be applied as tiny dots on the starting line and well-spaced from each other.

• You need to put individual spots on the starting line corresponding to the standard, unknown mixture along with a co-spot of the standard as well as unknown mixture.

• To put a spot, draw a small amount of solution in a capillary, say 1 – 2 cm and lightly touch the capillary to Silica surface so that sample is soaked up by Silica and forms a tiny spot.

• Do not use the capillary for another sample to avoid contamination

STEP -3 TLC solvent run:

• After spotting, TLC plate is transferred to a glass chamber containing the solvent to be used as mobile phase (ethyl acetate & petroleum ether).

• The solvent level in the chamber must be few mm below the starting line.

• Allow the solvent to rise along the Silica surface and as soon as the solvent front reaches the finish line, remove the plate from the solvent chamber and allow the solvent to dry out.

• Repeat this procedure using mobile phases of different polarity, until the mixture is separated properly on TLC plate.

STEP-4 Visualization of spots:

• Transfer the plate to Iodine / UV chamber and watch the spots appear on the plate.

• In the Iodine chamber the sample spots absorb iodine and develop a yellow-brown color, the density of which depends on solubility of I2 in the given sample.

• After withdrawing the plate from the iodine chamber, circle the spots with pencil and measure the distance travelled by each spot.

• Draw the chromatogram in your journal sheet and calculate the Rf value for each spot and use it to identify the constituent in mixture.

• The components can also be identified by comparison of the color of spots with those given by standards.

CALCULATIONS

• Distance travelled by the solvent = _____________________________(X) cm

• Distance travelled by Compound 1 = _____________________________(Y) cm

• Distance travelled by Compound 2 = _____________________________(Z) cm

• Rf of compound 1 = _____________________

• Rf of compound 2 = _____________________

RESULT

• Rf of compound 1 = _______________________

• Rf of compound 2 = _______________________