Clarinda clare linusDiploma 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 beakers0%, 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.
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 colors1. Black = purple and yellow color2. Red = bright red and white3. Green = blue and yellow4. Blue = bright blue5. Purple = bright purple and pink6. 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.
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