Exp 11 Report

1.
0 Title: Spectrophotometric Analysis of Caffeine and Benzoic acid in Soft Drink

2.0 Objective: To obtain the absorbance of caffeine and benzoic acid in soft drinks.
3.0 Introduction:
In this experiment we use ultraviolet absorbance (Figure 1) to measure two major
species in soft drinks. Caffeine is added as a stimulant and sodium benzoate is a
preservative.
All solutions will contain 0.010 M HCl, so the sodium benzoate is protonated to make
benzoic acid. Caffeine has no appreciable basicity, so it is neutral at pH 2.
Figure 1. Ultraviolet absorption of benzoic

acid, caffeine, and a 1:50 dilution of Mountain Dew soft drink. All solutions contain 0.010 M HCl.
We restrict ourselves to non-diet soft drinks because the sugar substitute aspartame
in diet soda has some ultraviolet absorbance that slightly interferes in the present
experiment. We also avoid darkly colored drinks because the colorants have ultraviolet
absorbance. Mountain Dew, Mello Yello, and, probably, other lightly colored drinks are
suitable for this experiment. There is undoubtedly some ultraviolet absorbance from
colorants in these beverages that contributes systematic error to this experiment.
The procedure we describe includes the construction of calibration curves. The
experiment could be shortened by recording just one spectrum of caffeine (20 mg/L) and
one of benzoic acid (10 mg/L) and assuming that Beer's law is obeyed. The experiment could
be expanded to use high-performance liquid chromatography (HPLC) and/or capillary
electrophoresis to obtain independent measurements of caffeine and benzoic acid (and
aspartame in diet drinks).
4.0 Apparatus:
 10ml pipette
 100ml volumetric flask
 10ml graduated cylinder
 5ml graduated cylinder
 UV/Vis Spectrophotometer
 Filter paper
5.0 Materials:
 Benzoic acid standard (100mg/l)

 Caffeine standard (200mg/l)
 0.1M HCl
 Soft drinks
6.0 Procedures:
A. Calibration of Standards
First, benzoic acid solution containing 2, 4, 6, 8 and 10 mg/ml in 0.1M HCl are prepared.
2.00ml of benzoic acid standard plus 10.0ml of 0.10M HCl are mixed in a 100ml volumetric
flask and is diluted with distilled water to the mark. 4, 6, 8 and 10ml of benzoic acid are used
to prepare the other standards. In a similar manner, caffeine standards containing 4, 8, 12,
16 and 20 mg/ml in 0,10M HCl are prepared.
B. Soft Drink
20ml of soft drink is warmed in a beaker on a hot plate to expel CO2 and the warm liquid is
filtered through filter paper to remove any particles. 4.00ml of soft drink is pipetted into a
100ml volumetric flask after cooling to the room temperature. 10.0ml of 0.10M HCl is added
and diluted to the mark. A second sample containing 2.00ml of soft drink is prepared instead
of 4.00ml.
C. Spectrophotometer Analysis
An ultraviolet baseline from 350 to 210nm are recorded with water in the sample and
reference cuvettes (1.00cm pathlength)The ultraviolet spectrum of each of the 10
standards is recorded with water in the reference cuvettes. The wavelength of peak
absorbance of benzoic acid (λ’) and the wavelength of peak absorbance of caffeine (λ’’) are
noted. A calibration graph of absorbance versus concentration (M) for each compound is
prepared at each of the two wavelengths. At least three replicate readings are taken for
each standards. The least-squares slope of the graph is the molar absorptivity at that
wavelength. The ultraviolet absorption spectrum of the 2:100 and 4:100 dilutions of the soft
drink are measured for unknowns. With the absorbance at the wavelengths λ’ and λ’’, the
concentration of benzoic acid and caffeine are found in the original soft drink. At least three
trials are ran for each of the soft drink prepared samples. Results from both dilute solutions
and original soft drink are reported.
7.0 Result:
Concentration of Benzoic acid Absorbance Value Wavelength (λ)

(ppm)
2 0.459 230
4 0.921 230
6 1.399 230
8 1.851 230
10 2.306 230
Concentration of Caffeine (ppm) Absorbance Value Wavelength (λ)

8 0.421 275
16 0.776 275
24 1.165 275
32 1.545 275
40 2.020 275
Mountain Absorbance Value Wavelength

Dew/ml Benzoic acid Caffeine Benzoic acid Caffeine
2 0.286 0.185 230 275
4 0.485 0.325 230 275
Sprite/ml Absorbance Value Wavelength

Benzoic acid Caffeine Benzoic acid Caffeine
2 0.313 0.300 230 275
4 0.584 0.566 230 275
8.0 Calculations:
From Graph 1:
To calculate the concentration of benzoic acid in 2ml of Mountain Dew
M ₁ V ₁=M ₂V ₂
M ₁( 2)=(1.300)(100)
M ₁=65 ppm
To calculate the concentration of benzoic acid in 4ml of Mountain Dew
M ₁ V ₁=M ₂V ₂
M ₁( 4)= ( 2.1 )( 100)
M ₁=52.5 ppm
To calculate the concentration of benzoic acid in 2ml of Sprite
M ₁ V ₁=M ₂V ₂
M ₁( 2)= (1.4 ) (100)
M ₁=70 ppm
To calculate the concentration of benzoic acid in 4ml of Sprite
M ₁ V ₁=M ₂V ₂
M ₁( 4)= ( 2.6 ) (100)
M ₁=65 ppm
From Graph 2:
To calculate the concentration of caffeine in 2ml of Mountain Dew
M ₁ V ₁=M ₂V ₂
M ₁( 2)=(4.0)(100)
M ₁=200 ppm
To calculate the concentration of caffeine in 4ml of Mountain Dew
M ₁ V ₁=M ₂V ₂
M ₁( 4)=(6.8)(100)
M ₁=170 ppm
To calculate the concentration of caffeine in 2ml of Sprite
M ₁ V ₁=M ₂V ₂
M ₁( 2)=(6.0)(100)
M ₁=300 ppm
To calculate the concentration of caffeine in 4ml of Sprite
M ₁ V ₁=M ₂V ₂
M ₁( 4)=(11.6)(100)
M ₁=290 ppm
The Average Concentration of Benzoic Acid in Mountain Dew:
( 65+52.5 ) ppm
Average concentration=
2
¿ 58.75 ppm
The Average Concentration of Benzoic Acid in Sprite:

( 70+65 ) ppm
2
¿ 67.5 ppm
The Average Concentration of Caffeine in Mountain Dew:

( 200+170 ) ppm
2
¿ 185 ppm
The Average Concentration of Caffeine in Sprite:

( 300+290 ) ppm
2
¿ 295 ppm
9.0 Discussion:
In this experiment, the absorbance of benzoic acid and caffeine are obtained using
spectrophotometric analysis. Spectrophotometry is one of the quantitative analysis in many
fields such as Chemistry, Physics, Chemical engineering and more. It is a method to measure
the amount of chemical substance absorb light by measuring the intensity of light as a beam
of light passes through the solution. The basic concept and principle behind
spectrophotometry is each compound absorbs and transmits light over a certain range of
wavelength. (Chemistry LibreTexts, 2019) A spectrophotometer is an instrument that used
to measure to absorbance of solution as light specified wavelength is passes through it.
I0 It
Figure 1.The differences between the incident and transmitted light indicates the
absorbance.
To start the absorbance measurement process, the initial irradiance is measured by
placing a solvent in the sample holder, cuvette. This also known as blank measurement. The
purpose of obtain blank measurement is to calibrate the spectrophotometer to take into
account the natural absorbance of the diluents. Then, the solvent is replaced with the real
samples and the irradiance are measured. The amount of protons that passes through the
cuvette and into the detector is dependent to the length of cuvette and the concentration of
the sample. Transmittance can be relate to the fraction of light passes through the sample.
Iₜ
Transmittance= Transmittance is then related to the absorption by using this expression:
I₀
Iₜ
Absorbance=−log
I₀
With the absorbance value that obtained, the unknown concentration of the sample
by using Beer-Lambert Law. This law states that the concentration states that the
concentration is the sample is directly proportional to the absorbance Beer’s Law can be
written as
A=εcl
where A is absorbance (no units), ε is the molar absorptivity, c is the concentration and l is
the path length. Experimentally, this can be shown the graph of absorbance (y-axis) against
concentration (x-axis). (Helmenstine, 2019)
In this experiment, the amount of benzoic acid and caffeine in Mountain Dew and
Sprite were determined. Benzoic acid is widely used as a food preservative and in the
manufacture of various cosmetics, dyes and more. (Eds, 2008) Caffeine is a chemical
compound that used to improve mental illness, which act as a stimulant and it can be found
commonly in coffee, tea, cola and many. Benzoic acid will exhibits the absorbance at the
wavelength, 230nm and caffeine exhibits absorbance at 275nm.
From Graph 1, the graph of absorbance against concentration for benzoic acid is
plotted. The relationship of graph can be shown seen to be linear scattered around the
straight line. It can be seen that there is a positive relationship between the variables. As the
concentration of benzoic acid increases, the absorbance increases. From the line of best fit,
when the absorbance is 0.286, the predicted concentration is 1.3ppm. When the
absorbance is 0.485, the predicted concentration is 2.1ppm. When the absorbance in the
2ml and 4ml of Sprite are 0.313 and 0.584, the predicted concentration for each is 1.4ppm
and 2.6ppm. By substituting the predicted concentration value in the formula,
M ₁ V ₁=M ₂V ₂ the concentration of benzoic acid in 2ml and 4ml of Mountain Dew is
65ppm and 52.5ppm and the average is 58.75ppm. The concentration of benzoic acid in 2ml
and 4ml of Sprite is 70ppm and 65ppm and the average is 67.5ppm.
From Graph 2, the graph of absorbance against concentration for caffeine is
plotted. The relationship of graph can be shown seen to be linear scattered around the
straight line. It can be seen that there is a positive relationship between the variables. As the
concentration of caffeine increases, the absorbance increases. From the line of best fit,
when the absorbance is 0.185, the predicted concentration is 4.0ppm. When the
absorbance is 0.325, the predicted concentration is 6.8ppm. When the absorbance in the
2ml and 4ml of Sprite are 0.300 and 0.566, the predicted concentration for each is 6.0ppm
and 11.6ppm. By substituting the predicted concentration value in the formula,
M ₁ V ₁=M ₂V ₂ the concentration of caffeine in 2ml and 4ml of Mountain Dew is 200ppm
and 170ppm and the average is 185ppm. The concentration of benzoic acid in 2ml and 4ml
of Sprite is 300ppm and 290ppm and the average is 295ppm.
During the experiment, there are also may have some errors occur during
spectrophotometric analysis. Mistakes such as the cuvettes are accidentally touch with
fingers and the fingerprints are on it or improper positioning of the cuvettes. These mistakes
could be introduce the errors.
10.0 Precaution steps:

There are few precaution steps have been taken in this experiment. First, HCl is
handled in the fume hood. If any acids or bases accidentally splashes onto the skin,
immediately rinse off with water. Next, safety goggles and gloves are worn when conducting
the experiment. Thirdly, the volumetric flask are labelled to prevent any mistaken
concentration used. Lastly, the chemical solution are disposed into the disposal container
prepared by the lab officer.
11.0 Conclusion:
The spectrophotometry is used to obtain the absorbance of benzoic acid and
caffeine in soft drink, Mountain Dew and Sprite. From graph 1, the concentration of benzoic
acid in 2ml and 4ml of Mountain Dew and Sprite are 65ppm, 52.5ppm, 70ppm and 65ppm
respectively. From graph 2, the concentration of caffeine in 2ml and 4ml of Mountain Dew
and Sprite are 200ppm, 170ppm, 300ppm and 290ppm respectively.
12.0 References:
Chemistry LibreTexts, 2019. Spectrophotometry. [online] Available at:
<https://chem.libretexts.org/Bookshelves/Physical_and_Theoretical_Chemistry_Textbook_
Maps/Supplemental_Modules_(Physical_and_Theoretical_Chemistry)/Kinetics/Reaction_Ra
tes/Experimental_Determination_of_Kinetcs/Spectrophotometry> Accessed 22 July 2019
Helmenstine, A, M., 2019. Beer’s Law Definition and Equation. [online] Available at: <
https://www.thoughtco.com/beers-law-definition-and-equation-608172> Accessed 22 July
2019
The Editors of Encyclopaedia of Britannica, 2008. Benzoic Acid. [online] Available at:
<https://www.britannica.com/science/benzoic-acid> Accessed 23 July 2019

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1.

0 Title: Spectrophotometric Analysis of Caffeine and Benzoic acid in Soft Drink

Figure 1. Ultraviolet absorption of benzoic

 Benzoic acid standard (100mg/l)

Concentration of Benzoic acid Absorbance Value Wavelength (λ)

Concentration of Caffeine (ppm) Absorbance Value Wavelength (λ)

Mountain Absorbance Value Wavelength

Sprite/ml Absorbance Value Wavelength

The Average Concentration of Benzoic Acid in Sprite:

The Average Concentration of Caffeine in Mountain Dew:

The Average Concentration of Caffeine in Sprite:

10.0 Precaution steps:

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