11.2: Exercise

Part I: Identifying Food Dyes in Candies

Many foods, drugs and cosmetics are artificially colored with federally approved food dyes (FD & C dyes). These dyes include Red 40, Red 3, Yellow 5, Yellow 6, Blue 1, and Blue 2. Since each dye has an identifiable absorption spectrum and peak, a spectrophotometer may be used to identify the types of FD & C dye used in a product.

Pigments may be extracted from foods and drinks that contain one or more of these dyes. An absorption spectrum of that extract can then determine what dyes are in that food or drink by comparing the peaks of maximum absorbance with information in the table below. If the absorption spectrum of a food extract has a peak at 630 nm and one at 428 nm, you can assume the food contains both Blue #1 and Yellow #5. The following table gives the wavelength of peak absorbance for each of these dyes.

Table 1. Wavelength of Maximum Absorbance of Commonly Used FD & C Dyes

FD & C Dye	Name	Wavelength (nm) of Maximum Absorbance
Blue #1	Brilliant Blue FCF	630
Green #3	Solid Green FCF	625
Blue #2	Indigo Carmine	610
Red #3	Erythrosine	527
Red #40	Allura Red AC	502
Yellow #6	Sunset Yellow FCF	484
Yellow #5	Tartrazine	428

Materials

• Spectrophotometer
• Cuvette
• Skittles
• KimWipes
• Test tubes

Procedure

A. Extracting Dye from Candy

You will need one test tube and one cuvette for each color to be tested. Measure 4 mL water into one tube. Place 2-4 candies of the same color in a test tube with the water. Gently swirl, and wait one minute. After, pour approximately 1 mL of liquid into a microcentrifuge tube. Spin the microcentrifuge tube at max speed for 60 seconds. Make sure the centrifuge is balanced before spinning. Transfer the clear liquid (supernatant) into a cuvette. Make sure to leave behind the particulates (pellet).

Figure 4. Curve of absorbance

B. Measuring Absorbance with Spectrophotometer

1. Turn on the spectrophotometer. Let it warm up for 15 minutes.
2. Select wavelength scan.
3. Fill a cuvette 2/3 full with DI water to serve as the “BLANK” cuvette.
4. Calibrate the Spectrometer

1. 1. Wipe the outside of the BLANK cuvette with a KimWipe
   2. Place cuvette into the machine so that the clear portions of the cuvette are oriented left to right. (The light needs to pass through clear area on cuvette).
   3. Press "Zero"
   4. Remove the blank.

5. Determine optimum wavelength absorbance and set up data collection mode.

1. 1. Place a cuvette with a sample into the spectrophotometer
   2. Press "read"
   3. Set cursor mode to peak and valley under "Options" > "More" > "Cursor Mode"
   4. Press the left and right arrows to the right of the graph to select the highest peak.
   5. Record the wavelength and absorbance in Table 6.2.

6. Decide which dyes were used to make each color. Enter the Wavelength of that dye in the last column of table 6.2. Explain your reasoning for each choice in your lab notebook.

Results

Table 2.

Color skittle	Maximum Absorbance	Wavelength (nm) at Maximum Absorption	Proposed Dye	Wavelength (nm) of Maximum Absorption for Proposed Dye
Red
Orange
Yellow
Green
Purple

Instructions for Cleaning Cuvettes

1. Discard solutions to sink
2. Rinse with tap water once
3. Rinse with DI water two times
4. Place in tube rack, allow to air dry

Study Questions

1. Name the parts of the spectrophotometer and identify their function.
2. What is the difference between % transmittance and absorbance?
3. How did you determine which wavelength was absorbed at the highest level? How is this process useful in determining the identity of a molecule?
4. How do you clean a cuvette?