CHEM 2425 (Organic Chemistry I)
EXPERIMENT 2
UV-Vis Spectroscopy Detection of Caffeine in Beverages
Introduction:
Caffeine is a common organic molecule found in many beverages such as coffee, tea, soft
drinks and energy drinks. It stimulates respiration, the heart, and the central nervous
system. That is why many students drink coffee or soda to help them feel alert. Caffeine
also can cause nervousness and insomnia and, like many drugs, can be addictive, making
it difficult to reduce the daily dose. A regular coffee drinker who consumes just four cups
per day can experience headache, insomnia, and even nausea upon with drawl from the
drug. On the other hand, it helps people to pay attention and can sharpen moderately
complex mental skills as well as prolong the ability to exercise.
Caffeine belongs to a large class of compounds known as alkaloids. These are of plant
origin, contain basic nitrogen, often have a bitter taste and complex structure, and usually
have physiological activity. Their names usually end in “ine”; many are quite familiar by
name if not chemical structure-nicotine, cocaine, morphine, strychnine.
CH3
N
N
N
N
H3C
O
CH3
O
In the experiment you will detect the content of caffeine in different coffee, tea, soft
drinks and energy drinks using UV-Vis Spectroscopy. UV-Vis spectroscopy is one of the
most common techniques in the detection of organic compounds. It is the study of the
interaction of electromagnetic radiation (EMR), 'light', with matter. EMR is a set of
oscillating electric and magnetic fields which is characterized by a frequency () and a
wavelength (λ), related through the speed of light, c, as shown in the following equation:
λ* = c
In the visible region of the spectrum the color of the sample indicates which regions of
the electromagnetic spectrum are interacting with the sample, and which regions are
passing through the sample with little or no interaction. The former are said to be
'absorbed' by the sample while the latter are said to be 'transmitted'. A sample that
appears to be blue in color does so because only the blue region of the visible spectrum is
passing through the sample; the red and green portions are interacting with the sample
and are absorbed. The color and amount of light absorbed can be detected and measured
using an instrument called a spectrophotometer. The major components of a
spectrophotometer operating in the visible portion of the spectrum are a white-light
source; a monochromator, a device that separates the white-light into its component
colors; a detector which measures the amount of light passing through the sample; and
various associated mirrors, lenses, and filters to change the light path within the
instrument. The instrument is constructed in such a way that the output signal, called the
absorbance, that it provides is directly proportional to the concentration of the light
absorbing species.
The spectrometer records the wavelengths at which absorption occurs, together with the
degree of absorption at each wavelength. The resulting spectrum is presented as a graph
of absorbance (A) versus wavelength (λ), as the caffeine spectrum shown below. This
spectrum have shown that in water (solvent), the maximum absorbance wavelength
max) of caffeine is around 273 nm, and this characteristic maximum wavelength can be
used to identify caffeine.
0.25
Absorbance
0.2
0.15
0.1
0.05
0
240
250
260
270
280
290
300
310
320
Wavelength (nm)
Materials and Apparatus
10 mg/L standard caffeine solution in water
Jasmine Tea
Green tea
Black Tea
Cocochai Tea
Nestle Coffee
Maxwell Coffee
Decaf Coffee
Coca-Cola
Sprite
Mountain Dew
Red Bull
Monster energy
Rockstar
Agilent Technology UV-Vis Spectrophotometer
Decaf Raspberry Tea
Pepsi
Full Throttle
UV quartz cells (10 mm, 3.0 mL)
Procedure
1. Obtained the standard caffeine spectrum:
 Get 2 UV cells, make sure they are clean
 Fill water near 3/4 full of each UV cell
 Add 2 drops of standard 10 mg/L caffeine solution to one of the cells
 Turn on the computer and UV instrument
 Click the ChemSation
 Turn on the light source of the UV and the visible.
 Place one cell containing water in number 1of the sample holder. (Wipe
the unfrosted sides with a Kimwipe. Place the cell with the unfrosted sides
facing the front and back. Close the lid)
 Place the cell containing caffeine sample in the number 2 sample holder
 In the ChemStation program, choose “Scan spectrum” and select the
region from 230 nm to 350 nm.
 Click to choose cell 1, then click “blank” to scan the solvent (water)
 Click to choose cell 2, then click “sample” to scan the sample
 Caffeine should have a peak at around 273 nm
Absorbance should ranges from 0.1~0.7. If absorbance is higher than 1.0,
dilute your sample with water until the absorbance drops below 0.7,
following the suggestions of your instructor. Mix the diluted solution well,
and run the spectrum again. When diluting your sample, assume that
absorbance will decrease in proportion to concentration.
 Print the plot of absorbance vs wavelength for the caffeine standard
sample, find out the maximum absorbance wavelength max).
2. Test the caffeine content in “Student Samples”:
Tea:
Jasmine Tea, Green tea, Black Tea, Cocochai Tea, Decaf Raspberry Tea
Coffee:
Nestle Coffee, Maxwell Coffee, Decaf Coffee
Soda:
Coca-Cola, Sprite, Mountain Dew, Pepsi
Sports drinks: Red Bull, Monster energy, Rockstar, Full Throttle
In the table above, pick up two samples from each group (totally 8) that you want to test,
and repeat the same procedure in Part 1 to test whether the samples have any caffeine in
them (whether the spectra of the tested samples have maximum absorbance wavelength
max at around 273 nm). Wash the cells of the samples thoroughly before each use.
Remember to label the printouts with sample name.
Report Form
Beverage
max (nm)
Absorbance
Caffeine
(Y/N)
Relative
Caffeine
Amount
Attach your UV-Vis spectra in below:
Pre-lab Questions
1. Are you caffeine user? What effect does caffeine have on you?
2. Briefly explain how UV-Vis is used as a detection technique for organic compounds?
3. What is the purpose of the caffeine standards?
Post-lab Questions
1. Why does the cell have to be carefully rinsed before each use?
2. For which of the samples was caffeine detected? Explain.
3. Roughly compare the caffeine content in each sample (absorbance is directly
proportional to the concentration of caffeine).Which one has the most amount? Which
one has the least amount? Is it reasonable based on our common sense?