Materials Included In Kit

Additional Materials Required

Prelab Preparation

Standard Dye Solutions

Blue No. 1

1. Mass 0.80 g of the Blue Dye No. 1 and transfer it to a 100-mL volumetric flask.
2. Fill the flask with about 50 mL of distilled or deionized water. Swirl to mix.
3. Fill to the mark with distilled or deionized water. Cap and mix thoroughly.
4. Use a 1.0-mL volumetric pipet to transfer 1.0 mL of the solution to a clean 1-L volumetric flask.
5. Add deionized water to the 1-L mark, cap and mix thoroughly.

Yellow No. 5

1. Mass 1.50 g of the Yellow Dye No. 5 and transfer it to a 100-mL volumetric flask.
2. Fill the flask with about 50 mL of distilled or deionized water. Swirl to mix.
3. Fill to the mark with distilled or deionized water. Cap and mix thoroughly.
4. Use a 1.0-mL volumetric pipet to transfer 1.0 mL of the solution to a clean 1-L volumetric flask.
5. Add deionized water to the 1-L mark, cap and mix thoroughly.

Red No. 40

1. Mass 2.33 g of the Red Dye No. 40 and transfer it to a 100-mL volumetric flask.
2. Fill the flask with about 50 mL of distilled or deionized water. Swirl to mix.
3. Fill to the mark with distilled or deionized water. Cap and mix thoroughly.
4. Use a 1.0-mL volumetric pipet to transfer 1.0 mL of the solution to a clean 1-L volumetric flask.
5. Add deionized water to the 1-L mark, cap and mix thoroughly.

Drink Mix Solutions

Cherry

1. Mass 0.90 g of the Cherry granular drink mix and transfer it to a 1-L volumetric flask.
2. Fill the flask with about 500 mL of distilled or deionized water. Swirl to mix.
3. Fill to the mark with distilled or deionized water. Cap and mix thoroughly.

Grape

1. Mass 1.00 g of the Grape granular drink mix and transfer it to a 1-L volumetric flask.
2. Fill the flask with about 500 mL of distilled or deionized water. Swirl to mix.
3. Fill to the mark with distilled or deionized water. Cap and mix thoroughly.

Lemon-lime

1. Mass 1.00 g of the Lemon-lime granular drink mix and transfer it to a 1-Liter volumetric flask.
2. Fill the flask with about 500 mL of distilled or deionized water. Swirl to mix.
3. Fill to the mark with distilled or deionized water. Cap and mix thoroughly.

Safety Precautions

The dye solutions and drink mix solutions are considered nonhazardous. Food items, once brought into a lab, are considered chemicals and, as such, should not be ingested. Wear chemical-splash goggles, aprons and gloves. Wash hands thoroughly with soap and water before leaving the laboratory. Please follow all laboratory safety guidelines. Remind students to wash their hands thoroughly with soap and water before leaving the laboratory. Please review current Safety Data Sheets for additional safety, handling and disposal information.

Disposal

Please consult your current Flinn Scientific Catalog/Reference Manual for general guidelines and specific procedures, and review all federal, state and local regulations that may apply, before proceeding. The leftover dye and drink mix solutions may be rinsed down the drain with plenty of excess water according to Flinn Suggested Disposal Method #26a.

Lab Hints

• Enough materials are provided in this kit for 30 students working in pairs or for 15 groups of students. Both parts of this laboratory activity can reasonably be completed in one 50-minute class period. The prelaboratory assignment may be completed before coming to lab, and the data compilation and calculations may be completed the day after the lab.
• Significant differences in dye concentration can occur between different lot numbers of specific granular drink mixes.
• If you use a data collection and analysis software technology, such as Vernier LabPro and LoggerPro, be sure to modify the procedure to reflect the specific steps required to use the system.

Teacher Tips

• Since passage of the Federal Food, Drug, and Cosmetic (FD&C) Act of 1938, color additives in the United States have been the responsibility of the Food and Drug Administration (FDA) and are labeled with FD&C names.
• The FDA divides coloring agents into two categories: certifiable and exempt from certification. The former are primarily derived from petroleum and the latter includes agents that are largely from mineral, plant, or animal sources. Certifiable color additives are available for use in food as either “dyes” or “lakes.” Dyes dissolve in water and are manufactured as powders, granules, liquids or other special-purpose forms. They can be used in beverages, dry mixes, baked goods, confections, dairy products, pet foods and a variety of other products. Lakes are the water insoluble form of the dye. Lakes are more stable than dyes and are ideal for coloring products containing fats and oils or items lacking sufficient moisture to dissolve dyes. Typical uses include coated tablets, cake and doughnut mixes, hard candies and chewing gums.

Answers to Prelab Questions

A sample of lemon-lime drink mix was analyzed to determine its content of the food dye FD&C Green No. 3. The λ_max was determined to be 625 nm and a calibration curve was constructed. Seven grams of the drink mix were diluted to 4.0 L with distilled water. This sample had an absorbance value of 0.263A at 625 nm.

1. What is the green food dye concentration of the drink mix solution?

   The sample absorbance value corresponds to a green food dye value of 3.0 x 10^–6 moles/liter.

2. The molar mass of FD&C Green No. 3 is 766 g/mol. Calculate the total grams of FD&C Green No. 3 in the 4 liters of solution.
   1. Total moles of FD&C Green No. 3 (3.0 x 10^–6 moles/liter) x 4 liters = 1.2 x 10^–5 moles
   2. Total grams of FD&C Green No. 3 1.2 x 10^–5 moles x 766g/mole = 9.2 x 10^–3 g of FD&C Green No. 3
3. What percent of the granular drink mix is FD&C Green No. 3?

   (9.2 x 10^–3 g of FD&C Green No. 3÷7 g of the drink mix) x 100 = 0.13%

Sample Data

Part A. Determining λ_max

Blue Dye No. 1

Yellow Dye No. 5 Red Dye No. 40 Part B. Calibration Curve Part C. Determining Drink Mix Solution Absorbances

Answers to Questions

1. Determine the concentration of Blue Dye #1, Yellow Dye #2 and Red Dye #40 in each of the drink mixes.

   (Values taken from calibration curves.)
   Cherry Drink Mix
   Blue Dye #1 – 1.4 x 10^–6 mol/L
   Red Dye #40 – 50 x 10^–6 mol/L
   
   Grape Drink Mix
   Blue Dye #1 – 3.7 x 10^–6 mol/L
   Red Dye #40 – 19 x 10^–6 mol/L
   
   Lemon-lime Drink Mix
   Blue Dye #1 – 1.2 x 10^–6 mol/L
   Yellow Dye #5 – 7.6 x 10^–6 mol/L

2. From your instructor, obtain the total volume of each drink mix solution and the mass of granular drink mix added to the solution. Calculate the concentration of the three dyes, in %, by weight, in each of the granular drink mixes. The molar mass of the dyes follow.

   Blue Dye #1 — 792.9 g/mol Cherry Drink Mix – 0.90 grams in 1 liter
   Yellow Dye #5 — 534.3 g/mol Grape Drink Mix – 1.00 grams in 1 liter
   Red Dye #40 — 496.4 g/mol Lemon-lime Drink Mix – 1.00 grams in 1 liter

   Cherry Drink Mix
   Blue Dye No. 1 grams = (1.4 x 10–6 mol/L) x (7.929 x 102 g/mol) x 1 liter = 1.1 x 10–3 grams of Blue Dye No. 1 %, by weight of Blue Dye No. 1 = (1.1 x 10–3 gram/0.90 gram) x 100 = 0.12%
   Red Dye No. 40 grams = (5.0 x 10–5 mol/L) x (0.4964 x 103 g/mol) x 1 liter = 2.5 x 10–2 grams of Red Dye No. 40 %, by weight of Red Dye No. 40 = (2.5 x 10–2 gram/0.90 gram) x 100 = 2.8%
   
   Grape Drink Mix
   Blue Dye No. 1 grams = (3.7 x 10–6 mol/L) x (7.929 x 102 g/mol) x 1 liter = 2.9 x 10–3 grams of Blue Dye No. 1 %, by weight of Blue Dye No. 1 = (2.9 x 10–3 gram/1.00 gram) x 100 = 0.29%
   Red Dye No. 40 grams = (1.92 x 10–5 mol/L) x (0.4964 × 103 g/mol) x 1 liter = 0.95 x 10–2 grams of Red Dye No. 40 %, by weight of Red Dye No. 40 = (0.95 x 10–3 gram/1.00 gram) x 100 = 0.95%
   
   Lemon-lime Drink Mix
   Blue Dye No. 1 grams = (1.2 x 10–6 mol/L) x (7.929 x 102 g/mol) x 1 liter = 0.95 x 10–3 grams of Blue Dye No. 1 %, by weight of Blue Dye No. 1 = (0.95 x 10–3 gram/1.00 gram) x 100 = 0.10%
   Yellow Dye No. 5 grams = (7.6 x 10–6 mol/L) x (0.5343 x 103 g/mol) x 1 liter = 4.1 x 10–3 grams of Yellow Dye No. 5 %, by weight of Yellow Dye No. 5 = (4.1 x 10–3 gram/1.00 gram) x 100 = 0.41%

References

Sigmann, S. B.; Wheeler, D. E. The Quantitative Determination of Food Dyes in Powdered Drink Mixes; J. Chem. Ed., 2004; 10, 1475–1478.

Introduction

Food dyes occur everywhere, from food and drink to cosmetics and are even used in theatrical blood! How much of food dyes are actually contained in products? In this experiment, determine how much of a series of food dyes are added to granular drink mixes to give these drinks their vivid colors.

Concepts

• Absorbance
• Percent
• by weight
• Consumer
• Conjugated bonding

Background

The three dyes used in this experiment appear as different colors under normal white light. They are each composed of different molecules—molecules that absorb different wavelengths of light. In general, a blue solution looks blue to the human eye because it is transmitting blue light. When white light is shined through this solution, the molecules in the solution absorb some of the wavelengths of the light and transmit others. All non-blue wavelengths of light will be absorbed by the blue solution to some extent, although yellow light will be absorbed the most. The yellow photons hit the solution and are absorbed by the molecules in the solution. They do not make it through the solution, and hence, we do not see a yellow color from this solution. In contrast, blue photons are not absorbed by the molecules in the blue solution. So, they pass right through the solution, and we see a blue color.

How do we know that the blue solution absorbs the yellow wavelengths of light? Yellow and blue are complementary colors—combined they make white light. The idea of complementary colors can be used as a first estimation of the wavelengths that are absorbed by a substance based on its color.

The following table lists the wavelengths associated with each of the colors in the visible spectrum and their complements. The representative wavelength can be used as a benchmark for each color. For example, instead of referring to green as light in the wavelength range 500–560 nm, one could simply say that green light is 520 nm.

Why do dyes absorb visible light energy? Many dye molecules are large, complex organic molecules. The structures of the three dyes you will analyze are shown below (see Figure 1). When molecules have a series of double bonds separated by single bonds, the bonding pattern is called conjugated, or joined together in pairs. This pattern of bonding results in a reduced separation between the ground state and the excited state of the electrons. This energy difference corresponds to the energy of photons in the visible region. As the amount of conjugation increases, the energy of the absorbed photon decreases. Based on their colors, the blue dye should absorb light that is least energetic of the three, followed by red and yellow with higher absorbed energies.

In this lab you will determine the amount of each dye contained in common granular drink mixes. You begin by finding the wavelength of light that results in the maximum absorbance value, λ_max, for each of the three dyes. For the blue dye, the absorbed energy lies in the yellow-orange region. This means the blue dye’s λ_max is between 580nm and 650nm. For the red dye, you will look for λ_max between 480nm and 500nm, for yellow, between 425nm and 480nm.

Once you have determined λ_max for each dye, you will create a calibration curve at that wavelength for each dye. Next, determine the absorption of known solutions of various drink mixes at these three λ_max values. From these data you will calculate the amount of each of the three dyes contained in the granular drink mixes.

Experiment Overview

The purpose of this experiment is to determine the amount of specific food dyes contained in several granular drink mixes. The wavelength of maximum absorbance for each of the dyes will be determined and a calibration curve created for each. By finding the absorbance of solutions of the drink mixes at these wavelengths the amount of each dye contained in the granular drink mixes may be calculated.

Materials

Prelab Questions

A sample of lime drink mix was analyzed to determine its content of the food dye FD&C Green No. 3 (see Figure 2).

The λ_max was determined to be 625 nm and a calibration curve was constructed (see Figure 3). Seven grams of the drink mix were diluted to 4.0 liters with distilled water. This sample had an absorbance value of 0.263 A at 625 nm.

1. What is the FD&C Green No. 3 concentration of the drink mix solution?
2. The molar mass of FD&C Green No. 3 is 766 g/mol. Calculate the total grams of FD&C Green No. 3 in the 4 liters of solution.
3. What percent of the granular drink mix is FD&C Green No. 3?

Safety Precautions

The dye solutions and drink mix solutions are considered nonhazardous. Food items, once brought into a lab, are considered chemicals and, as such, should not be ingested. Wear chemical-splash goggles, gloves and apron (more as protection from stains). Wash hands thoroughly with soap and water before leaving the laboratory. Please follow all laboratory safety guidelines.

Procedure

Part A. Determining λ_max
Follow the procedure for your colorimetric measurements of the solution as directed by the instructor. Generally, spectropho-tometers are used as follows: Turn the instrument on and allow it to warm up for 15 minutes. Set the wavelength at 450 nm. With no light passing through the instrument to the phototube, set the percent transmittance to zero with the “zero” control. Handle cuvets at the top so no fingerprints are in the light path. Polish cuvets with a tissue. Place a cuvet which is about ⅔ full of distilled water into the sample holder and set the percent transmittance to 100% with the appropriate control (not the zero control). Fill a cuvet about ⅔ full of a test solution, place it in the spectrophotometer and read the absorbance. Consult the instrument manual for details on its use.

Blue No. 1

1. Test for λ_max between wavelengths 610 nm and 655 nm for blue dye No. 1. Set the wavelength of the spectrophotometer to 610 nm.
2. Fill a cuvet ⅔ full with distilled or deionized water and place in the sample holder. Set the spectrophotometer to 100% transmission. Remove the cuvet.
3. Fill a cuvet about ⅔ full of a test solution, place it in the spectrophotometer and record the absorbance in the data table on the worksheet.
4. Repeat steps 1 to 3 for each wavelength listed in the data table.
5. Graph the data on the graph under the Blue No. 1 Data Table. From the graph, determine λ_max.

Yellow No. 5

1. Test for λ_max between wavelengths 410 nm and 475 nm for yellow dye No. 5. Set the wavelength at 410 nm.
2. Repeat steps 2 through 4 listed for Blue Dye No. 1.
3. Graph the data on the graph under the Yellow No. 5 Data Table.

Red No. 40

1. Test for λ_max between wavelengths 460 nm and 530 nm for Red Dye No. 40. Set the wavelength at 460 nm.
2. Repeat steps 2 through 4 listed for Blue Dye No. 1.
3. Graph the data on the graph under the Red No. 40 Data Table.

Part B. Calibration Curve

Blue Dye No. 1

1. Set up five clean test tubes in the test tube rack.
2. Prepare the five reference solution test tubes listed in the following table. Use a separate pipet to transfer the appropriate volumes of each reagent. Mix each solution using a stirring rod. Rinse the stirring rod and dry it between solutions. Label the test tubes with the corresponding reference solution number.
   {12164_Procedure_Table_2}
3. Set the spectrophotometer to λ_max of the Yellow Dye No. 5. Zero the instrument with distilled or deionized water.
4. Fill a cuvet about ⅔ full of a Yellow Dye solution 1, place it in the spectrophotometer and record the absorbance in the data table.
5. Repeat step 4 for the other Yellow Dye solutions.
6. Empty and clean the test tubes and cuvets.

Yellow Dye No. 5

1. Set up the five test tubes in the test tube rack.
2. Prepare the five reference solution test tubes listed in the following table. Use a separate pipet to transfer the appropriate volumes of each reagent. Mix each solution using a stirring rod. Rinse the stirring rod and dry it between solutions. Label the test tubes with the corresponding reference solution number.
   {12164_Procedure_Table_3}
3. Set the spectrophotometer to λ_max of the Yellow Dye No. 40. Zero the instrument with distilled or deionized water.
4. Fill a cuvet about ⅔ full of a Yellow Dye solution 1, place it in the spectrophotometer and record the absorbance in the data table.
5. Repeat step 4 for the other Yellow Dye solutions.
6. Empty and clean the test tubes and cuvets.

Red Dye No. 40

1. Set up the five test tubes in the test tube rack.
2. Prepare the five reference solution test tubes listed in the table below. Use a separate pipet to transfer the appropriate volumes of each reagent. Mix each solution using a stirring rod. Rinse the stirring rod and dry it between solutions. Label the test tubes with the corresponding reference solution number.
   {12164_Procedure_Table_4}
3. Set the spectrophotometer to λ_max of the Red Dye No. 40. Zero the instrument with distilled or deionized water.
4. Fill a cuvet about ⅔ full of a Red Dye solution 1, place it in the spectrophotometer and record the absorbance in the data table.
5. Repeat step 4 for the other Red Dye solutions.
6. Empty and clean the test tubes and cuvets.

Part C. Drink Mix Samples

1. Set up three clean test tubes in the test tube rack.
2. Prepare the three drink mix solution test tubes listed. Label the test tubes or beakers with the corresponding reference solution number.

Drink Mix Solutions

1. Cherry drink mix
2. Lemon-lime drink mix
3. Grape drink mix

3. Set the spectrophotometer to λ_max of the Blue Dye No. 1. Zero the instrument with distilled or deionized water.
4. Fill a cuvet about ⅔ full of drink mix solution 1, place it in the spectrophotometer and record the absorbance in the data table for Part C.
5. Repeat step 4 for the other drink mix solutions.
6. Set the spectrophotometer to λ_max of the Yellow Dye No. 5. Zero the instrument with distilled or deionized water.
7. Fill a cuvet about ⅔ full of drink mix solution 1, place it in the spectrophotometer and record the absorbance in the data table.
8. Repeat step 7 for the other drink mix solutions.
9. Set the spectrophotometer to λ_max of the Red Dye No. 40. Zero the instrument with distilled or deionized water.
10. Fill a cuvet about ⅔ full of drink mix solution 1, place it in the spectrophotometer and record the absorbance in the data table.
11. Repeat step 10 for the other drink mix solutions.

Student Worksheet PDF

12164_Student1.pdf