Clarinda clare linus

Diploma In Medical Sciences – January 2011

 At the end of this laboratory activity, the
students should be able to:
 Be introduced to the principles and
terminology of chromatography and
demonstrate separation of the dyes in
colored pens with paper chromatography.
 Paper chromatography is an analytical chemistry
technique for separating and identifying color
mixtures.
 In paper chromatography, substances are
distributed between stationary phase and a
mobile phase. Stationary phase is usually a piece
of filter paper and mobile phase is the colors that
travels up the stationary phase.
 Components of the samples will separate readily
according to how strongly they absorb on the
stationary phase vs. how readily they dissolve in
the mobile phase.
 6 beakers of jars
 6 covers of lids
 Distilled water
 Isopropanol
 Graduated cylinder
 6 strips of filter paper
 Different colors of water color pens
 Pencil
 Ruler
 Scissors
 tape
 Prepare 15 ml of isopropanol solutions in
labeled beakers
0%, 5%, 10%, 20%, 50% and 100%
 Cut 6 strips of filter paper
- Draw a line 1cm above the bottom edge of
the strip with the pencil
- Label each strip with its corresponding
solution
- Place a spot from each pen on your starting
line
 Place each strip in each beaker
- Cover the beakers
- Let the strips absorb the solutions about 2
cm from the top of the strips
- Remove strips and let them dry
 Observe the colors separated in low
concentrations of isopropanol and high
concentrations of propanol.
 Record data.
Distilled water 5% alcohol
10% alcohol 20% alcohol
50% alchohol 100% alcohol
 1st – black
 2nd – red
 3rd – green
 4th – blue
 5th – purple
 6th - pink
 According to the conducted experiment, the
results are as follows:

 Distilled water : Colors does not rise up to

stationary phase, dissolve
 Alcoholic solution: Colors rise up to stationary
phase and spread its colors
1. Black = purple and yellow color
2. Red = bright red and white
3. Green = blue and yellow
4. Blue = bright blue
5. Purple = bright purple and pink
6. Pink = Bright pink and white
 When a colored sample is placed on a filter
paper, the colors separate from the sample
by placing one of the paper in a low-leveled
solvent.
 The solvent diffuses up to the filter paper,
dissolving the various molecules in the
sample according to polarities of the
molecules and the solvent.
 If the sample contains more than one color, it
means that it have more than one kind of
molecule.
 Because of the different chemical structures
of each kind of molecules, the chances are
very high that each molecule will have at least
a slightly different polarity, giving each
molecule a different solubility in the solvent.
 The unequeal solubilities cause the various
color molecules to leave solution at different
places as the solvent continues to move up
the paper.
 Hence, the more soluble is a molecule, the
higher it will migrate up the paper.
 Chromatography has many uses. It is
commonly used in laboratories to isolate new
compounds, analyze subtle differences
between different environmental samples,
and even in the sequencing of DNA.
 To perform chromatography, one need two
things; a matrix and a color indicator. A
matrix is simply the materials to which a
sample is applied, the material is often
porous, action as filter paper that affects the
rate of flow of a sample. Special beads with
different pore sizes are commonly used to
separate proteins in biochemistry labs.
 The matrix may interact with a sample, and
depending on the chemical properties of
both, and is commonly used to analyze
mixtures of sugars.
 The color indicator may be the compound
itself. Some organic compounds are bright
yellow or orange, which makes them easy on
identifying on a chromatogram. Others need
to react with another chemicals to become
colored.
 Dyes specific for particular chemical
substances properties can be used to identify
samples. For examples, rhodamine dye can
be applied to visualize fats and oils.