M&m dyes for each color

Chemistry Lab Report?

Chromatography of M&M candies and food dyes.
(Minor changes to Supplemental Material for Online Publication from Birdwhistell and Spence. This handout can be cited as indicated below)
objective
Use a physical separation technique to separate and identify the food dyes used to give M&M’s their color.
Hazards
Avoid exposure to UV radiation from hand-held UV lamp. Do not look into the lamp with the lamp on.

introduction
Almost all substances we come into contact with on a daily basis are impure; that is, they are mixtures. Similarly, compounds synthesized in the chemical laboratory are rarely produced in a pure state. They are almost always produced with impurities including reaction byproducts and leftover reactants. As a result, a major focus of research in chemistry is designing methods of separating and identifying the various components of mixtures.

Many of these separation methods rely on physical differences between the components of a mixture. Undoubtedly, you are already familiar with several means chemists use to effect separations based on physical differences. These techniques include: Filtration, where separation may be effected because substances are present in different states (solid vs. liquid); Centrifugation, where separation is effected by differences in density; and Distillation, where separation is effected by taking advantage of differences in boiling temperatures of the various components. In this laboratory exercise, we will effect a separation of a mixture of food dyes using paper chromatography.

All chromatography techniques have three important components: the analyte or mixture of species being separated, a mobile phase, and a stationary phase. The mobile phase is a flowing liquid or gas used to push the analyte over or through a stationary porous material (the stationary phase). Because of physical interactions between the analyte and the stationary phase, the analyte moves through or over the stationary phase more slowly than the mobile phase. Furthermore, because physical interactions between the analyte and the stationary phase can be different for each component of the mixture, the different components transit the stationary phase at different speeds. Those that strongly interact with the stationary phase lag behind those that interact only weakly. As a result, the components of the mixture may be separated.

Figure 1 shows the separation of a two-component mixture using a typical liquid column chromatography apparatus. The analyte, dissolved in a liquid, is injected into a flowing mobile phase continuously pumped through the apparatus. The mobile phase sweeps the mixture into a chromatography column containing the stationary phase. As the two components are pushed through the column by the mobile phase, one interacts with the stationary phase more strongly and lags behind. As a result, the two components come out or elute from the column at different times allowing them to be collected in separate vials.

There are several different types of physical interactions that can occur between the analyte and the stationary phase which slow the progress of the analyte across the stationary phase. Under the right experimental conditions, each component of a mixture will exhibit a unique binding interaction with the stationary phase and, therefore, elute from the chromatography column at a unique time.

Paper chromatography is a form of liquid chromatography using a piece of paper as the stationary phase rather than a packed chromatography column. In paper chromatography, analyte is applied directly onto the bottom of the stationary phase (the chromatography paper) which is then placed on edge in a developing tank containing the mobile phase so that the bottom edge of the paper is submersed. Capillary action draws the mobile phase up the sheet of paper carrying along the different components of the mixture. Due to physical interactions between the different components and the stationary phase, the components move up the paper at different speeds. The paper is removed from the tank before the solvent front reaches the top of the paper and the position of each component of the mixture is marked as well as the position of the solvent front itself. Unlike in liquid column chromatography, the components of the mixture are not allowed to elute off of the top of the stationary phase. Rather, each component is characterized by the distance it traveled up the paper. The ratio of this distance to the distance the solvent front traveled, denoted Rf, is characteristic of a particular substance and remains constant regardless of the other components present in the mixture and, therefore, can be used to qualitatively identify the substance. The Rf of each component is dependent on the type of stationary phase used and the composition of the mobile phase.

image4.jpg

Figure 1 Liquid Column Chromatography. A two component mixture is injected into the flowing mobile phase and swept into a chromatography column containing the stationary phase. Because the molecules interact more strongly with the stationary phase than the component, they lag behind. As more mobile phase is pumped through the column, the two components are eventually separated and removed from the column.

Identification of the components of a mixture is aided if each substance in the mixture exhibits a unique color (as with most food dyes). However, for dyes with similar colors or for substances exhibiting no color, identification may be aided by examining the fluorescence exhibited under illumination with an ultraviolet (UV) lamp. Fluorescence is the phenomenon in which molecules absorb and immediately re-emit light. The emitted light is usually a different color than the absorbed light due to loss of energy between the absorption and emission events. Some substances re-emit absorbed radiation on longer time scales, seconds or minutes after the absorption event. Such substances are said to be phosphorescent. As with fluorescence, phosphorescence can be used to identify substances appearing similar under normal lighting.

In this laboratory, you will use liquid chromatography to determine Rf values of several common food dyes; you will then run a second chromatogram to separate dyes used in the colorful coatings of M&M candies. By comparing Rf values, color, and fluorescence characteristics of the dyes you will identify the individual dyes used in the mixtures found in the M&M coatings.

Procedure
A. Characterization of Standard Food Dyes.
1. Prepare a developing tank by pouring 10 mL of the mobile phase (0.10% wt/v solution of NaCl in H2O) into a 400 mL beaker and covering the beaker with Parafilm. It is important that the air above the mobile phase become saturated with solvent vapor so that solvent does not evaporate from the stationary phase as the chromatogram develops. Therefore, be sure to keep the developing tank covered at all times.

2. [Check with your TA for the size and type of filter paper to be used] With a pencil , draw a horizontal line 1.5 cm from the bottom edge of the chromatography paper. Draw vertical tick marks along this line every 2 cm (see fig 2(a)). Using a capillary and one of the standard dye solutions, make a spot on the chromatography paper at one of the marks. Keep the capillary with the correct dye to avoid cross contamination and waste of capillaries. Try to keep your spots less than 4 mm in diameter. Allow the dye to dry and reapply the same dye in the same spot 1 or 2 times or until a sufficiently dark spot has been achieved. With a pencil, note the name of the dye below the spot.

3. Repeat steps 2 and 3 for the remaining dyes across the bottom of the chromatogram.

4. When the spots have been applied, put your name in pencil across the top of the chromatography paper, form the chromatography paper into a cylinder, and staple the edges of the paper together making sure to leave a gap between the edges as shown in fig 2(b). If the edges come into contact, solvent will not travel at a uniform speed up the chromatography paper and the components of the mixture will not move in a straight line.

5. Place the chromatography paper into the developing tank, do not let it touch the sides of the tank and quickly replace the Parafilm cover. Make sure the level of the mobile phase is below the line of dyes on your paper. Allow the chromatogram to develop.

6. When the solvent front is approximately 1 cm from the top of the chromatography paper, remove the chromatogram and lay it flat on a paper towel. Immediately mark the position of the solvent front with a pencil. The front will continue to move as the paper dries so it is important that you mark this position now. Measure and note the distance the solvent front traveled (Dsolvent, see fig 3.).

7. Draw an ellipse around each spot on the developed chromatogram and draw a horizontal line through the center of each spot. If a spot shows significant “tailing” make your horizontal line through the darkest part of the spot (see fig 3). Use the distance from the starting line (not the bottom of the paper!) to these horizontal lines to determine Ddye for each dye. Record distances and Rf values in your notebook. Recall:

Rf = Ddye / Dsolvent

8. Place the chromatogram on edge, in the drying oven for 5 min or until dry. Take your dry chromatogram to the UV lamp area and observe the spots under illumination with a hand-held UV lamp on long-wavelength irradiation. Note the color of the observed fluorescence.

WARNING: UV light can damage your eyes. Always point lamps away from you. Do not look into the UV lamps

B. Separation and Identification of Dyes Used to Coat M&M Candies.
1. Do the M&M candies contain any of the standard dyes above? To begin to answer this question, take one M&M of each color to the UV lamp and see if they fluoresce. Note the color of any fluorescence you observe in your notebook.

2. [Three pairs of students will share making two solutions each.] To separate the dyes used to coat M&Ms, you must first prepare a solution of the coating dyes. To do this, place 4 M&M’s of the same color into a small beaker. Add 3 mL of a 50/50 mixture of water and ethanol and swirl the solvent until the candy coating has dissolved. Remove the M&M’s from the solvent before the chocolate center is exposed. You and your partner may coordinate with two other pairs of experimenters so that each pair only has to prepare 2 solutions that can be shared with the group.

3. Prepare a second chromatogram like that in Part A using the 6 M&M solutions. The dye solutions prepared with the M&M candies are not as concentrated as the solutions of the standard dyes. You will need to spot these dye mixtures several (>5) times to obtain a sufficiently dark spot. [7-9 times may be necessary] Again, dry each spot between applications of the mixture to maintain as small and concentrated a spot as possible.

4. Develop the chromatogram. Characterize the Rf value, color, and fluorescence characteristics of each dye observed. Remember that unlike the previous chromatogram, you are now separating mixtures of dyes and there may be as many as four components each with its own Rf value, color and fluorescence characteristics. Use all three characteristics to identify all components found in each M&M coloring.

5. Are any of the fluorescing substances you observe on your chromatogram actually phosphorescent? Place your chromatogram on a table and place the UV lamp, on “short-wavelength” on top of your chromatogram. Allow your eyes to adjust to the dark room. Quickly draw your chromatogram out from under the UV lamp and note any substances that continue to glow once out from under the lamp. (hint: pay close attention to the solvent front!)

[Record you data in your notebook in tables similar to these but not on these pages. For your JAR you can copy and paste these into word as a template for your results tables.
Characterization of Standard Dyes
Distance solvent front traveled (Dsolvent) (mm):

Dye

Color

Distance Dye Traveled (Ddye)(mm)

Rf

Fluorescence Color/Intensity

FD&C Red #3

FD&C Red #40

FD&C Blue #1

FD&C Blue #2

FD&C Yellow #6

FD&C Yellow #5

FD&C Green #3

[Record you data in your notebook in tables similar to these but not on these pages. For your JAR you can copy and paste these into word as a template for your results tables.
Characterization of Dyes Used to Coat M&M Candies
Distance solvent front traveled (Dsolvent, mm):

Beside each M&M color note the color, Ddye, Rf value, and fluorescence color / intensity for each spot observed. If possible, identify each spot as one of the standard dyes above. (Most mixtures will not have four components.)

Before Identifying dyes based on Rf values it is useful to have a good idea what the relative error is in your determination of Rf values. Using what you know about how difficult it is to identify the center of a dye spot, estimate the error in your determination of Rf values.

M&M Color

Dye Color

Ddye (mm)

Rf

Fluorescence Color/Intensity

F, D&C Food Dye

Red

Yellow

Green
M&M Color

Color

Ddye (mm)

Rf

Fluorescence Color/Intensity

F, D&C Food Dye

Blue

Orange

Brown

Post-Lab Questions – Answer these as a list in your JAR you can copy these and type your answers below them.

1. In chromatography what do the mobile and stationary phases do?

(a) What is the mobile phase in this experiment?

(b) What is the stationary phase in this experiment?

2. Why was a pencil used to mark the paper rather than a pen or marker?

3. Why is it important for the initial dye “spots” to be small?

4. Define Fluorescence

5. Define Phosphorescence

6. In chemistry or other sciences, what is chromatography used for?

7. What is the possible range of values for Rf? (Lowest and highest)

Figure 2 Paper Chromatogram. (a) Schematic for laying out spots. (b) Chromatogram ready for developing tank.

Figure 3 Developed Chromatogram. Immediately mark solvent front when paper is removed from the developing tank. The spot on the right exhibits significant tailing and the distance the spot has traveled has been correctly identified.

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� Adapted by Lary Sanders, Wright State University form supplemental online material of: Birdwhistell, K.R; Spence T. G. A New Glow on the Chromatography of M&M Candies J. Chem. Educ., 2002, 79 (7), p 847 DOI: 10.1021/ed079p847 Publication Date (Web): July 1, 2002 Supplemental Information online.

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