Objective To illustrate the use of paper chromatography for the separation and identification of amino acids.

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Paper Chromatography

Objective

To illustrate the use of paper chromatography for the separation and identification of amino acids.

Results

Amino Acid

Distance travelled /cm

Rf value

by solvent

by amino acid

Aspartic acid

- alone

- in mixture

5.3

5.3

.2

.2

0.2264

0.2264

Leucine

- alone

- in mixture

5.3

5.3

3.95

4.3

0.7453

0.8113

Lysine

- alone

- in mixture

5.3

5.3

3.1

3.3

0.5849

0.6226

Q & A

.> For each amino acid, compare your two Rf values with each other and with the specimen results (you can obtain from your teacher)

Why do you think there is some variation?

[ANS] Both Rf values are the same for aspartic acid but there is some variation for leusine & lysine. This may due to:

1.> The grease on our fingers contaminated on chromatographic paper will affect greatly on the amino acids that are less

polar.

2.> The measurement error of the distance travelled by solvent and/or amino acid.

3.> The shape of chromatographic paper is not exactly square-shaped, so that one side of the paper may be lifted and the

speed of the solvent travel in one side is faster.

4.> The different proportion of chemicals (?measured by measuring cylinder) may be used between the researchers and us, and
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the temperatures may also different, thus Rf values obtained are different.

2.> Why do Rf values change when a different solvent is used?

[ANS] Each amino acid has different polarity and each solvent has different polarity, too. So, when a different solvent is used, amino

acids will travel at a different speed.

For example, if a more polar solvent is used, the more polar amino acid will travel at a faster speed

and has a larger Rf value.

On the contrary, if a less polar solvent is used, the non-polar amino acid ...

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