Chem 414
UV Spectroscopic Analysis of Caffeine & Benzoic Acid in Soft Drinks
An informal lab report is due on this experiment one week after completing the lab
In this experiment, we use ultraviolet absorbance to measure two major species in soft drinks. Caffeine is
added as a stimulant and sodium benzoate is a preservative. Benzoic acid is a food preservative and it is
widely used in acidic foods (pH 2.5 - 4). It is normally added as the salt, sodium benzoate. Although
benzoic acid is a man-made food additive, benzoic acid naturally occurs in several fruits, often in amounts
that exceed the recommended allowance set by the FDA.
In this analysis we will use non-diet soft drinks because the sugar substitute, aspartame, found in diet
drinks, also absorbs ultraviolet radiation. This slightly interferes in the analysis. We also avoid drinks
containing carmelized sugar, as this colorant absorbs in the UV region of interest. Therefore, in this
experiment we shall analyze Mountain Dew, a highly caffeinated soft drink, and Sprite, which does not
contain caffeine. There will be some UV absorbance from the matrix of these drinks. This does cause a
small systematic error, but this error is very small compared to the levels of caffeine and benzoic acid
being measured.
As seen in the spectrum below, both caffeine and benzoic acid absorb UV light at many of the same
wavelengths.
However, as seen in the previous experiment, Beer’s law can be applied to solutions containing more
than one kind of absorbing substance. Provided there is no interaction among the various species (side
reactions), the total absorbance for a multicomponent system is given by:
Atotal = A1 + A2 + .….. + An (Equation 1)
Atotal = ε1bc1 + ε 2bc2 + .….. + ε nbcn
where the subscripts refer to absorbing components 1, 2, …n.
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The above equation indicates that the total absorbance of a solution at a given wavelength is equal to the
sum of the absorbances of the individual components present. This relationship makes possible the
quantitative determination of the individual constituents of a mixture, even if their spectra overlap. If
enough spectrometric information is available, all of the components of mixtures can be quantified without
separation. For a two-component mixture (compound X and Y) with overlapping absorbances, you can
solve for the concentration of each species, [X] and [Y], by measuring the absorbances at two different
wavelengthts, λ' and λ". The problem is mathematically equivalent to having two simultaneous equations
with two unknowns.
A1= ε x,1bcx + ε Y,1bcY (total absorbance at λ') (Equation 2)
A2= ε x,2bcx + ε Y,2bcY (total absorbance at λ") (Equation 3)
The four molar absorptivities, ε x,1 , ε y,1 , ε x,2 , ε y,2 , can be evaluated from individual standard
solutions of X and Y, or better, from the slopes of their Beer’s law plots. The problem becomes simpler
when one of the compounds has no interference with the other compound. If there is substantial
interference then you must solve the simultaneous equations.
Using UV spectroscopy, you will determine the concentrations of caffeine and sodium benzoate
(determined as benzoic acid), in the soft drink Mountain Dew. The UV spectra of caffeine and benzoic
acid overlap at certain wavelengths, thus you will need to measure the absorbance of the unknown
mixtures using two different wavelengths, and apply equations 2 and 3 to evaluate the concentrations of
caffeine and benzoic acid. Use your text and the previous lab for help in carrying out the calculations.
Background Information
Text chapters 13 and 14.
Prelab Requirements: Will count towards your data section of your lab report
1. Write out the procedure that you will use, starting with solid sodium benzoate and caffeine, to
produce 100mL of 10mg/L stock solution of sodium benzoate and 20 mg/L of caffeine. Then
prepare each of the standards listed in the laboratory procedures section of the experiment from
these stock solutions. Use the same equipment you have used in the previous experiments.
2. Give your calculations to your TA to be graded at the start of the lab.
Chemical Hazards
The calibration standards for both benzoic acid (10mg/L) and caffeine (20mg/L) solutions are prepared in
a 0.010M solution of HCl. Wear a labcoat, safety eyewear and gloves. Wash off any solution
immediately.
Laboratory Procedures
Preparation of 10ml Calibration Standards
1. Prepare a set of benzoic acid solutions containing 2, 4, 6, 8, and 10 mg/L in 0.010 M HCl.
2. Prepare caffeine standards containing 4, 8, 12, 16, & 20 mg/L in 0.010 M HCl.
Preparation of Soft Drink Solutions
1. Warm ~ 20 mL of soft drink in a beaker on a hot plate to expel CO2. (Do not boil!)
2. Filter the warm liquid through filter paper to remove any particles.
3. After cooling to room temperature, pipette 2.00 mL into a 50-mL volumetric flask.
4. Add 10.0 mL of 0.010 M HCl and dilute to the mark.
Constructing a Calibration Curve
1. Record the ultraviolet spectrum of each of the 10 standards with water in the reference cuvette.
2. Record the wavelength of peak absorbance for benzoic acid (λ′) and the wavelength for the peak
absorbance of caffeine (λ′′) in your laboratory notebook.
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Determination of Caffeine and Benzoic Acid in Soft Drinks
1. Measure the ultraviolet absorption spectrum of the diluted soft drink samples.
Calculations
Constructing a Calibration Curve
1. Prepare a calibration graph of absorbance versus concentration for each compound at each of
the two wavelengths. You should make 4 different graphs.
2. Each graph should go through 0. The slope of the graph is the absorptivity at that wavelength.
Determination of Caffeine and Benzoic Acid in Soft Drinks
1. With the absorbance at the wavelengths λ′ and λ′′ determine the concentrations of benzoic acid
and caffeine in each sample.
2. Copy the data from the spectrometers in electronic form. This data can then be converted into a
UV spectrum using Microsoft Excel.
Data and Results
1. 4 calibration curves with correlation coefficients and equation for the line included. All axes must
be labeled and a title on each graph
2. A single table containing the following information:
a. Absorptivities of caffeine and benzoic acid found from the slope of the graphs.
b. Concentrations of benzoic acid and caffeine in each diluted soft drink sample.
c. Amount of benzoic acid consumed in a single serving (12 ounces) of a soft drink.
d. Amount of caffeine consumed in a single serving (12 ounces) of a soft drink.
References
V.L. McDevitt, et al., J. Chem. Ed. 1998, 75, 625
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