Identifying the Safe Concentration Level of Dye Blue No. 1

Introduction

Almost any beverage that can be purchased, with the exception of water, contains dyes. Companies add food dyes to consumer products in an attempt to make them more attractive for the consumer. The only problem with this is that the chemicals used to dye consumer products can sometimes be harmful, especially in higher concentrations. A study conducted by Swanson and Kinsbourne concluded that children diagnosed with hyperactivity performed poorly on tests after being exposed to food dyes compared to when they were given a diet free of food dyes [1]. Another study conducted by the Journal of the American Academy of Pediatrics found that many dyes used in commercial products could have adverse health affects for humans [2]. This is very important information for the consumer public to know. It is extremely necessary for a consumer to know if the products they are buying are safe. Without this information consumers could unknowingly purchase products that will harm them or their loved ones. The goal of this experiment was to determine if the concentrations of food dyes in certain commercial products were at safe levels for consumer use. The researchers chose to test products containing the dye Blue No. 1. This dye was chosen because it is a very commonly used dye that can be found in almost any blue colored beverage. A study from the UK shows that the maximum allotted concentration for Blue 1 is 100 mg/L for non-alcoholic flavored beverages [3]. Based on this information the researchers were looking for levels below 100mg/L in the consumer products being tested. The experimenters chose to test the concentrations of Blue 1 in three consumer products, blue Powerade, blue Gatorade and blue Scope mouthwash. The researchers chose to determine these concentrations using spectrophotometry. Spectrophotometry uses a machine called a spectrophotometer to determine the absorbance of a substance at particular wavelengths. A calibration curve is simply a line that plots concentrations against their respective wavelengths. Using a calibration curve a researcher can determine the unknown concentration of a substance in a solution by determining that substance’s absorbance and then comparing it to the calibration curve for that substance. By creating a calibration curve for the absorbance of Blue 1 at different concentrations and comparing this curve to the absorbencies of the commercial products the researchers were able to determine the concentrations of Blue 1 in the three consumer products. Alternatively from a calibration the use of Beer’s Law can be utilized to find the concentration of Blue 1 in each product.

Beer’s Law

A=ebc

Where A is absorbance, e is molar absorbtivity, b is the path length of the sample and c is concentration. After finding the absorbance via the spectrophotometer research can be done to find the molar absorbtivity of Blue No. 1. After this has been found the equation could be manipulated to find c.

C=A/eb

This would thus use Beer’s Law to find the concentration of Blue 1 in each product.

Methods

The procedure of this experiment was broken up into three parts. Part 1-Determining the concentration of food dye that would yield an absorbance reading below 1 and determining the wavelength of peak absorbance for the dye. Part 2- Creating a calibration curve for the absorbance of Blue 1. Part 3- Testing the absorbance of each consumer product and using the calibration curve to determine each product’s concentration of Blue 1.

Part 1:

Before the researchers chose to work with Blue 1, three different dyes were provided to them to find concentrations that would yield absorbance readings below 1 as well as find each dye’s wavelength of peak absorbance; the three dyes were Yellow No. 6, Blue No. 1 and Green No. 3. This part was conducted so that when the researchers did choose a certain dye they would have an idea of what concentrations to use for the calibration curve, and also what wavelength to create the curve with.

Yellow No. 6 (C16H10N2Na2O7S2) [3] Blue No. 1 (C37H34N2Na2O9S3) [3] Green No. 3 (C37H37N2O10S3+) [3]

The researchers chose to create their stock solution using .04g of each solid dye and mixing it with enough distilled water to fill a 50mL volumetric flask. This amount was chosen because it would create enough stock solution to be used later in the experiment and it also would not use too much of the solid food dye. The following calculations determined the molarities of the stock solutions.

Yellow No. 6 (MM- 452.38g/mol)

(.04g Yellow 6)/ x (1 mole)/452.38g=8.8x10-5mol

(8.8×〖10〗

(-5))/.05L=1.768x〖10〗

(-3)m/L

Blue No. 1 (MM- 792.89g/mol)

(.04g Blue 1)/ x (1 mole)/792.89g=5.045x10-5mol

(5.045×〖10〗

(-5))/.05L=1.01x〖10〗

(-3)m/L

Green No. 3 (MM- 765.90g/mol)

(.04g Green 3)/ x (1 mole)/765.90g=5.223x10-5mol

(5.223×〖10〗

(-5))/.05L=1.054x〖10〗

(-3)m/L

Knowing these solutions were far too concentrated to be below an absorbance of 1 the researchers diluted the stock solutions. The researchers decided to mix 5mL of each stock solution with distilled water in a 100mL volumetric flask. The following calculations show the molarities of the solution after the dilution.

M1V1=M2V2

Yellow No. 6

((1.768×〖10〗

(-3) m/L).005L)/.1L=8.842×〖10〗

(-5) m/L

Blue No. 1

((1.01x〖10〗

(-3) m/L).005L)/.1L=5.05×〖10〗

(-5) m/L

Green No. 3

((1.054x〖10〗

(-3) m/L).005L)/.1L=5.225×〖10〗

(-5) m/L

These diluted solution were then placed into cuvettes, which were then evaluated using the spectrophotometer. Each of the three solutions yielded an absorbance well above 1 meaning the researchers had to dilute the solutions further. The researchers decided to place 5mL of the first diluted solution into a 100mL volumetric flask and mix with distilled water. The following calculations show the molarities of these solutions.

M1V1=M2V2

Yellow No. 6

((8.842×〖10〗

(-5) m/L).005L)/.1L=4.421×〖10〗

(-6) m/L

Blue No. 1

((5.05×〖10〗

(-5) m/L).005L)/.1L=2.525×〖10〗

(-6) m/L

Green No. 3

((5.225×〖10〗

(-5) m/L).005L)/.1L=2.613×〖10〗

(-6) m/L

These solutions were then tested using the spectrophotometer as the previous solution was. However, these solutions all yielded an absorbance that was too low, at around approximately .1. The researchers then realized they needed to make solutions of concentrations somewhere in between the previous two. With this information the researchers decided to add 5mL of the first dilution to a 50mL volumetric flask and then add distilled water. The following calculations describe the concentrations that were created.

M1V1=M2V2

Yellow No. 6

((8.842×〖10〗

(-5) m/L).005L)/.05L=8.842×〖10〗

(-6) m/L

Blue No. 1

((5.05×〖10〗

(-5) m/L).005L)/.05L=5.05×〖10〗

(-6) m/L

Green No. 3

((5.225×〖10〗

(-5) m/L).005L)/.05L=5.225×〖10〗

(-6) m/L

These three solutions were then tested using the spectrophotometer as the other two concentrations were tested. This tested yielded absorbance readings under 1 but not too low. The researchers then knew what concentrations to work with. After this was completed the researchers decided to conduct the rest of the test using Blue No. 1. The researchers used the spectrophotometer again and determined that the peak absorbance for Blue 1 was at a wavelength of 628nm, this was the wavelength the researchers would use to create the calibration curve.

Part 2:

The researchers chose to create the calibration curve for Blue 1 at 628nm using the concentrations 1.0x10-6, 2.0x10-6, 3.0x10-6, 4.0x10-6, 5.0x10-6, 6.0x10-6, 7.0x10-6, 8.0x10-6, 9.0x10-6 and 1.0x10-5. These concentrations were decided upon because they create a realistic range for the possible concentrations in the three consumer products. The following calculations determine how much of the first diluted solution (5.05x10-5m/L) was needed to create a 100mL solution of each different concentration.

M1V1=M2V2

V_1=((1.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=1.98mL

V_1=((2.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=3.96mL

V_1=((3.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=5.94mL

V_1=((4.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=7.92mL

V_1=((5.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=9.90mL

V_1=((6.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=11.88mL

V_1=((7.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=13.86mL

V_1=((8.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=15.84mL

V_1=((9.0×〖10〗

(-6) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=17.82mL

V_1=((1.0×〖10〗

(-5) m/L)×.1L)/(5.05×〖10〗

(-5) m/L)=19.8mL

After conducting these calculations the researchers mixed each amount of the first diluted solution with the appropriate amount of water to fill a 100mL volumetric flask. These solutions of different concentrations were then placed in to the spectrophotometer and their absorbencies were determined at 628nm. The absorbance readings were then plotted on a graph against their respective concentrations to create the calibration curve.

Part 3:

The final part of this experiment simply involved placing each of the three consumer products, blue Powerade, blue Gatorade and Scope mouthwash into the spectrophotometer and recording their absorbencies at 628nm. After each product’s absorbance was determined the absorbance readings were compared to the calibration curve in order to determine each product’s concentration of Blue 1.

Results/Discussion

The following graph represents the results of Part 1 of this experiment. The graph shows how the peak absorbance of Blue 1 was determined to be 628 nm.

The following data represents the results of the spectrophotometer readings for the different concentrations of Blue 1.

Concentration (M) Absorbance

0.000001 0.001

0.000002 0.04

0.000003 0.197

0.000004 0.266

0.000005 0.334

0.000006 0.462

0.000007 0.597

0.000008 0.714

0.000009 0.791

0.00001 0.918

Clearly it is shown by this data that the higher the concentration or molarity of the substance the higher the absorbance. This data however is not the data that was used to create the calibration curve. Three point included in the data represented above were not used in the calibration curve. These concentrations are 1.0x10-6, 2.0x10-6 and 5.0x10-6. The reason the first two points were discarded is simply because they were far too low of readings to be beneficial on the calibration curve. The third reading was discarded because the researchers felt that it was inaccurate based on the readings surrounding it, and the curve can be created with out it. The following data represents the actual data used as well as the calibration curve that was created.

Concentration (M) Absorbance

0.000003 0.197

0.000004 0.266

0.000006 0.462

0.000007 0.597

0.000008 0.714

0.000009 0.791

0.00001 0.918

Concentration (Molarity)

This calibration curve can be used to find the concentration of Blue 1 in a substance if the absorbance of this substance at wavelength 628nm is known. Using the equation of the line on the graph, y=86245x, it easy to determine the concentration of each product. The following data shows the absorbance readings for each of the commercial products that were tested.

Product Name Absorbance

Gatorade 0.446

Powerade 0.466

Scope 0.279

Using this data the following calculations determine the concentrations off Blue 1, in molarity, of each product.

y=86245x

Gatorade

×=.446/86245=5.17×〖10〗

(-6) m/L

Powerade

×=.466/86245=4.28×〖10〗

(-6) m/L

Scope

×=.279/86245=3.23×〖10〗

(-6) m/L

These calculations show that of the three products Gatorade has the highest concentration of Blue 1 while Scope has the lowest concentration, and Powerade is in between the two. The research conducted by the experimenters concluded that safe levels for consumption of Blue No. 1 are at or below 100mg/L. Because the concentrations calculated above are in moles/L another calculation had to conducted to convert from moles of Blue 1 to milligrams of Blue 1, the following calculations show this conversion.

Gatorade

(5.17×〖10〗

(-6) moles Blue 1)/×(792.89g Blue 1)/(1 mole Blue 1)×1000mg/1g=4.1mg Blue 1

Powerade

(4.28×〖10〗

(-6) moles Blue 1)/×(792.89g Blue 1)/(1 mole Blue 1)×1000mg/1g=3.4mg Blue 1

Scope

(3.23×〖10〗

(-6) moles Blue 1)/×(792.89g Blue 1)/(1 mole Blue 1)×1000mg/1g=2.6mg Blue 1

The above calculations explicitly show that the concentration of Blue 1 in each of these three products is well below the recommended safe levels. In order to improve the procedure used in this experiment, future researchers could use more concentrations for the calibration curve to make it more accurate. Also researchers could test other products besides the ones tested here. Many other everyday commercial products contain Blue No. 1 and it is important that these products do not contain unsafe amounts of the chemical. For further testing researchers could test other dyes besides Blue 1. Most commercial beverages and some food products contain food dyes of all different colors so it is important to be sure that safe amounts of these chemicals are being used in said products.

Conclusion

To conclude, the purpose of this experiment was to determine if the concentration of dye Blue No. 1 was at a safe level in three commercial products, blue Gatorade, blue Powerade and Scope mouthwash. The researchers found that safe levels of Blue No. 1 are considered to be 100mg/L or less. Through experimentation the researchers determined that all three of the products tested contained Blue No. 1 in concentrations well under 100mg/L. This information means that these particular products are safe to use, as far as food dye is concerned. However, this does not mean that any product with Blue No. 1 is a safe product to use. It is certainly possible that other products contain much higher and potentially unsafe concentrations of Blue No. 1. Also this information is not meant to imply that all products with food dyes are safe. It is possible that other food dyes are much more harmful than Blue No. 1 and concentrations of these dyes may be unsafe when the same concentrations of Blue No. 1 would be safe.

Research Connection

At the State University of Maringa in Maringa, Brazil, Eliane C. Vidotti runs a team that conducted research on determining food dyes in products. The purpose of their research was to determine if another method besides spectrophotometry could be more efficient in determining food dyes [4]. This research is very important because it did conclude that there is a cheaper and just as efficient way of determining food dyes. The researchers started by determining food dyes using spectrophotometry as a constant and then test their alternative method. The researchers concluded that the alternative method of determining food dyes is just as useful and efficient as spectrophotometry, as well as much cheaper. The fact that research was conducted to find a better way of determining food dyes shows how important it is to know what dyes are in consumer products and the concentrations of those dyes.