Introduction Spectroscopy is a very interesting field in chemistry and physics in which the interaction between radiation and matter

is studied as a function of wavelength. 1 Spectroscopy has many applications: from the analysis of chemicals in foods and pharmaceuticals to determination of the age and composition of distant starts. 2 In the case of this experiment, the focus was to examine the roll of food dyes and their concentration in food products through the use of spectroscopy as many times companies can lie about how much of an ingredient they put into their product. Spectroscopy is a very good method of analyzing concentration of the food dyes within the food product, as there is a correlation between the color of the food dye, wavelength given off by that color, absorbance of that color, and the concentration of the dye in the food. To begin, there has to be an understanding of each of the factors mentioned above and how the spectrometer will allude to the concentrations of the food dye. Light itself acts as both a wave and a particle and their for can be analyzed as either one. When being analyzed as a wave, the energy of the light will be determined by its wavelength with a long wavelength producing a low energy and a short wavelength producing a high energy.

Illustration of wavelength vs energy level of light 3

Visible Light, the light humans are capable of seeing, falls under a very small segment of the electromagnetic spectrum, with each color having a specific range of wavelength. Beginning with the longest wavelength; red has a wavelength of about 650nm, orange about 590 nm, yellow about 570 nm, green about 510 nm, blue about 475nm, indigo about 445 nm, and violet with about 400 nm. 4 Max wavelength is a very important factor in understanding the characteristics for each compound.5 Using a spectrometer just doesnt require the understanding of wavelengths, however, as it is not the only factor in analyzing a compound. The absorbance spectrum, the extent to which a sample absorbs light,5 is another factor of the spectrometer that depends strongly upon the wavelength of light. 5 With spectrometry the samples are analyzed for their absorbance spectrum first as the absorbance spectrum will show the correlation of absorbance against wavelength thusly revealing the max wavelength of the sample. 5 As spectroscopys idea is that matter interacts with radiation, the spectrometer reveals the interaction between light and the color sample thus illuminating upon the wavelength and absorbency of the color. By analyzing certain food dyes prior to analyzing the actual food product, the Wavelengths of the electromagnetic spectrum4

correlation between absorbency and concentration can be constructed almost as if puzzle pieces slowly add up to making the complete image. Food dyes, primarily yellow 5 yellow 6 and red 40, are used in almost all manufactured food products in order to give them that dazzling appeal which allows them to sell better, but what if these companies are putting more than they say they put into their products. Food dyes can result with problems including allergies, hyperactivity, learning impairment, irritability and aggressiveness6 so that is why it is crucial to monitor the amount of food dye in food products.

Procedure The experiment to determine the concentration in each of the three food products was a three step process which began with first analyzing the food dyes yellow 5, yellow 6, and red 40. Week 1 1. Week 1 began with making sense of the connection between colors and the wavelengths associated with them. The food dyes available were in powder formation and thus had to be made into a solution of the dye and water, which will be called the stock solution. For each of the dyes the stock solution consisted of 0.5 g of the dye and 10ml of D.I. water. The spectrometer, however, cannot measure the absorbency and wavelength of the stock solution because it was too dark and thus had to be diluted. 2. A serial dilution was thusly done with each of the stock dye solution until the spectrometer was able to measure the wavelength and absorbency of each of the

dyes. Yellow 5 was diluted once using 1ml of the stock solution and 9 ml of DI water. Yellow 6 was diluted twice: the first time with 1 ml from the stock solution and 9 ml with DI water and the second time with 1 ml from dilution #1 and 9 ml DI water. Red 40 was done in a similar manner to Yellow 6: the first time with 1 ml from the stock solution and 9 ml with DI water and the second time with 1 ml from dilution #1 and 9 ml DI water. 3. Finally a table of wavelength vs. solution color was made and then a graph comparing the absorbance spectrum against the wavelengths for the dyes were made. see results section Week 2 1. The goal of this week was to create the calibration curve for the three food dyes. To be able to obtain a graph that compared absorbance vs. concentration for each food dye, three dilutions that were readable by the spectrometer were needed. The following diagrams illustrate which solutions were used in the spectrometer for each dye. Yellow 5 Dilution #2 1ml Dil1, 9ml DI water Dilution #1 1mlStock 9ml DI water

Stock 0.5g Dye 9ml DI water

Successful #3 4ml Succ #2 6ml DI

Successfu l #2 2ml Dil 1 8ml DI

Successful #1 4ml Dil 1 6ml DI

Yellow 6

Successful Dilution #1 1ml Dil2, 9ml DI water

Dilution #2 1ml Dil1, 9ml DI water

Dilution #1 1mlStock 9ml DI water

Stock 0.5g Dye 9ml DI water

Successful #3 4ml succ #2 6ml DI Red 40

Successful #2 2ml Dil 1 8ml DI

Dilution #3 1ml Dil2, 9ml DI water

Dilution #2 1ml Dil1, 9ml DI water

Dilution #1 1mlStock 9ml DI water

Stock 0.5g Dye 9ml DI water

Successful #3 4ml Dil 2 6ml DI water

Successful #2 4ml succ #1 6ml DI

Successful #1 2ml Dil 2 8ml DI

2. Once each readable successful dilution was made the concentration had to be determined for each in order to be able to compare it to the absorbency of each of the three successful dilutions of the dyes. Results show the graphs of absorbency vs. concentration while the calculations portion shows how the concentration was obtained for each dye Week 3 1. The final step was to use all the data gathered in determining the concentration of food coloring in our three food products; powerade, minutemade lemonade, and minutemade pink lemonade. Both powerade and minutemade pink lemonade corresponded with the red 40 dye while minutemade lemonade corresponded with the yellow 5 dye. The first step was to determine the absorbance and wavelength of the colors within each of the food product by placing the drinks in the cubets and then placing them into the spectrometer for measurement. 2. Using the graph of absorbance vs. concentration the concentration of each of the food dyes within the products could be matched up by lining it with the line of best fit at a given abundance. To give more accuracy, however, it was decided that the formula, A=

, in which A represents absorption, represents the

absorptivity constant, c represents concentration, and l represents the length of the cubet. Calculations are provided in the calculation potion of the lab.

Calculations The following calculations are to determine the concentrations of each of the successful dilutions of each food dye as shown in the diagrams in the methods portion of this paper. The calculations will be done for yellow 5, however, all further calculations for Red 40 and yellow 6 are done in a similar manner.

Primary formula used to determine concentration: M1V1=M2V2 Molar mass of Yellow 5: 534.69 g/mol Concentration of Stock: [stock] =

= 0.0009356 mol
= 0.93569 M

Concentration of Dilution #1: (0.093569 M)(1ml) = M2 (10ml) M2 = 0.00935699 M Concentration of Dilution #2: (0.00935699 M)(1ml) = M2 (10ml) M2 = 0.000935699 M Concentration of Successful Dilution #1: (0.000935699 M)(4ml) = M2 (10ml)

M2 = 0.00037428 M Concentration of Successful Dilution #2: (0.000935699 M) (2ml) = M2 (10ml) M2 = 0.00018714 M Concentration of Successful Dilution #3: (0.00018714 M) (4ml) = M2 (10ml) M2 = 0.000074856 M

The following calculations are to determine the concentrations of the dyes in each of the food products. The calculations will be done for powerade below, however, all further calculations for minutemade lemonade and minutemade pink lemonade are done in a similar manner. Formula used: A= Equation is rearranged:

the length of the cubet is negligible as it cancels out during the rearranging of the equation Known information is entered into the equation: = 0.0005 M

Results Table 1: Week 1 Dye Color vs. Max Wavelength vs. Max absorbance Dye Color Yellow 5 Yellow 6 Red 40 Max Wavelength 421-427nm 502nm 506-510 nm Max Absorbance 0.218 0.242 0.370

Table 2: Week 2 Absorbance vs. Concentration For Each Successful Dilution Dye Color Yellow 5 successful dilution 1 Yellow 5 successful dilution 2 Yellow 5 successful dilution 3 Yellow 6 successful dilution 1 Yellow 6 successful dilution 2 Yellow 6 successful dilution 3 Red 40 successful Dilution 1 Red 40 successful Dilution 2 Red 40 successful Dilution 3 Concentration (M) 0.00037428 0.000.18714 0.000074856 0.001105314 0.000110531 0.000008842 0.00020144 0.00040288 0.000080576 Absorbance 0.370 0.787 0.152 0.242 0.618 0.323 0.721 0.314 0.167

Graph 1: Week 2 Absorbance vs. Concentration For Yellow 5 Successful Dilution

0.9 0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0 0.0001 0.0002 0.0003 0.0004 Series1 Linear (Series1) y = 1644.3x R = -0.397

Graph 2: Week 2 Absorbance vs. Concentration For Yellow 6 Successful Dilution

0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0 0.0005 0.001 0.0015 y = 274.43x R = -4.77

Series1 Linear (Series1)

Graph 3: Week 2 Absorbance vs. Concentration For Red 40 Successful Dilution

0.8 0.7 0.6 0.5 0.4 0.3 0.2 0.1 0 0 0.0001 0.0002 0.0003 0.0004 0.0005 Series1 Linear (Series1) y = 1362.1x R = -0.565

Table 3: Week 3 Food Products and their Wavelength, Absorbance and Concentration Food Product Powerade Minutemade Lemonade Minutemade Pink Lemonade Wavelength (nm) 502-503 385 371 Absorbance 0.661 0.677 0.680 Concentration (M) 0.0005 0.0025 0.0005

Discussion The results gathered were reflective of the procedure of the experiment. As stated before, the goal of week one was to obtain and understand how the spectrometer worked and the correlation between wavelength and absorbency. What was understood through week one was that the color through the spectrometer is seen as a wave or in other words it fluctuates a lot, however, at the max absorbance the wavelength is also max or at its highest point and this was the information that was sought out. The max wavelength was obtained through the max absorbency and their correlation with each other. Table 1 shows that yellow 5 has a 421-427nm max wavelength, yellow 6 has a 502nm max wavelength, and red 40 has a 506-510 nm max wavelength. These were found at the maximum absorbency of each of the colors and those are recorded in table one. Understanding how the spectrometer worked and the relationship between the max wavelength and absorbency allowed for further progression with calculating various dilutions for each color. The reasoning behind this process was that a graph could be made with a line of best-fit using three points (the three different dilutions for each color) to find the unknown concentrations of the food product. To clarify, by finding an array of absorbencies for each food dye and discovering their concentrations through the mathematical process provided above, a graph was made with those points. The line of best fit was then drawn to show the trend between each of the points as this acts as a correlation factor between absorbancy and concentration of the dye. The food products absorbency is then found using the spectrometer and now the graph can allude to the concentration of the dye in the food product. By matching were the absorbency is for

example 0.661 in powerade, the concentration that is intersecting the line of best-fit at that absorbancy would be the concentration of red 40 in powerade. The results showed that powerade had a 0.0005M concentration of red 40, Minutemade Lemonade had a concentration of 0.0025M, and the Minutemade Pink Lemonade had a concentration of 0.0005 as well. This method of finding concentration, however, leaves a massive room for error as it is done with the eye and not mathematically. Thusly, the absorbency equation was implemented to obtain the most accurate concentration for the food products. This allowed for a massive removal of error in the study as a whole, however, there were other errors in the experiment that prevented the final analysis of the concentration of the food product to be off. The biggest error after human error was the analysis of the concentration of the dilutions that occurred. As they did not have a proper proportion with each serial dilution the mathematics corresponding with the analysis has many chances to be askew leading to the assumptions that the dilutions themselves may have been preformed incorrectly. Secondly, the new program that was to be used had many errors in itself and could have disrupted the data of all data. Overall the design of the experiment was very hectic and allowed very little room for error yet gave many puzzles along the way. Looking at the graphs they themselves show that the data may not be accurate, as the line of best fit should mathematically have a negative correlation amongst the absorbance and concentration. The process in which the lab was preformed, however was the important factor and that the process was done correctly even if other factors had laid a brick wall in front of the door to success.

Research Connection The analysis of food dyes in food products has been going on for more than 30 years now. Food dye consists of chemicals used to add color to food.6 It has been noticed that children consuming food coloring have had hyperactive behavior and there is a correlation between food coloring consumption and ADHD. A study by the United Kingdoms Food Standards Agency in 2007 showed that the consumption of foods containing dyes could increase hyperactive behavior in children. 6 The study gave children various drinks some containing food coloring and some not and the study showed a positive correlation with hyperactive behavior by the 8-9 year olds with each beverage. This is just one of many studies that are being conducted another being Dr. Martins studies on this topic.

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Given project Packet for the lab

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"What Wavelength Goes With a Color?" What Wavelength Goes With a Color? N.p., n.d. Web. 30 Nov. 2012. <http://scienceedu.larc.nasa.gov/EDDOCS/Wavelengths_for_Colors.html>.
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"Spectrophotometry." : Absorbance Spectrum. N.p., n.d. Web. 30 Nov. 2012. <http://www.chm.davidson.edu/vce/spectrophotometry/absorbancespectrum.html>

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"The Hidden Health Risks of Food Dyes." Healthy Recipes, Healthy Eating, Healthy Cooking. N.p., n.d. Web. 30 Nov. 2012. <http://www.eatingwell.com/food_news_origins/food_news/the_hidden_health_risks_of _food_dyes>.
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"Food Dye and ADHD: Food Coloring, Sugar, and Diet." WebMD. WebMD, 28 Jan. 0000. Web. 30 Nov. 2012. <http://www.webmd.com/add-adhd/guide/food-dye-adhd>.