SCRIPT: ORGANIC CHEMISTRY 3 (PAPER CHROMATOGRAPHY)

 Basic Procedure
 Applications

BASIC PROCEDURE
TYPES OF PAPER:

1. Whatman Filter paper: These are filters papers bala na mga retentive bala haw or In general
the paper contains 98-99% of α-cellulose, 0.3 – 1% β -cellulose.
2. Modified papers -These are acid or base washed filter paper, glass fiber type paper
3. Hydrophilic papers- Papers modified with methanol, formamide, glycol, glycerol,etc.
4. Hydrophobic Paper- Used in reverse phase chromatography. OH groups are acetylated
resulting to a hydrophobic nature in paper
Chromatographic paper should not be touched since it is t is important to keep your hands
clean and dry, as the oils and residue from your fingers can interfere with the results.cause the
developing solvent to take a different path producing very irregular spots when the
chromatogram is developed. Rubber gloves, finger cots or tweezers are therefore used when
working with the paper

Solvents: Most of commonly employed solvents are polar solvents but still but the choice
depends on the nature of the components to be separated. You can use non polar solvents or
moderately polar solvents. Using polar solvents naga increase the SEPARATION between
components sang mixture and Polar components (such as polar dyes or pigments) dissolve
well in polar solvents.

Mixture of solvent gina himu man para ang solubility sang substance can easily be adjusted
by changing the ratio of solvents. *shows slide sa rules* A useful solvent pair mixtures for
mobile phases include water-saturated with phenol, butanol-aqueous ammonia, acetone-water.

Preparation of the sample: Broadly speaking, the purpose of sample preparation is to

improve the quality or suitability of the sample and, in turn, improve the subsequent
analytical results.

Spotting: Diba ma draw ka pencil line ka sang imo starting point sa filter paper. You gotto
spot it onto the same line. And don’t forgot to label it. Sa pag spot you can apply one to two
microliters portions on the top of each other. Do it repeatedly until your desired amout is
reached.

Chromatogram Development: immerse the paper for about 1-2 hours. When immersing the
paper sa mobile phase It is important that the solvent level is below the line with the spots on
it. As for the development chamber any tight air container or sufficient nga size ok nana .Test
tubes, large reagent jars or aquaria can also serve as excellent chambers.

Detection of Compounds: They examine the spots sa UV bc many organic compounds can
be seen using this technique. Kay for example if the spots are not obvious holding paper
under an UV lamp can help to make the spots visible sa naked eye. Then you outline them.

APPLICATIONS:

Separation of Amino Acids: A mixture of unknown amino acids can be separated and
identified by means of paper chromatography. Ang position of the amino acid sa
chromatogram can be detected by spraying with ninhydrin, (a chemical compound with the
formula C9H6O4; IUPAC name: 2,2-dihydroxyindane-1,3-dione) which reacts with amino
acids to yield highly coloured products which is purple. And yellow-orange for secondary
amines.
Forensic Analysis: paper chromatography is useful in the field of forensic science, to identify
presence of alcohol or chemicals in blood. This methods is useful as it can be successfully
carried out with even the very small quantities of material. Samples from crime scenes and for
DNA and RNA fingerprints can be identified using this technique.

Food Analysis: paper chromatography has been primarily used for analysis of food colors in
ice creams, sweets, drinks and beverages, jams and jellies. To ensure that no non-permitted
coloring agents are added to the foods, only edible colors are permitted for use.

Pharmaceutical Analysis: This process is cost-effective and hence used as an alternative

method in monitoring the active ingredients present in the drug forms. Paper chromatography is
also applicable in colour identifications of pharmaceutical formulations.