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Ultra-Violet Spectroscopy (UV)

Ultraviolet-visible (UV-vis) spectroscopy is used to obtain the absorbance spectra of a
compound in solution or as a solid. What is actually being observed spectroscopically
is the absorbance of light energy or electromagnetic radiation, which excites
electrons from the ground state to the first singlet excited state of the compound or
material. The UV-vis region of energy for the electromagnetic spectrum covers 1.5 6.2 eV which relates to a wavelength range of 800 - 200 nm. The Beer-Lambert
Law, Equation 1, is the principle behind absorbance spectroscopy. For a single
wavelength, A is absorbance, is the molar absorptivity of the compound or molecule
in solution (M-1cm-1), b is the path length of the cuvette or sample holder, and c is
the concentration of the solution (M).

There are three types of absorbance instruments used to collect UV-vis spectra
which are single beam spectrometer, double beam spectrometer and simultaneous
spectrometer. All of these instruments have a light source, a sample holder and a
detector, but some have a filter for selecting one wavelength at a time. The single
beam instrument has a filter or a monochromator between the source and the sample
to analyze one wavelength at a time. The double beam instrument has a single
source and a monochromator and then there is a splitter and a series of mirrors to

get the beam to a reference sample and the sample to be analyzed, this allows for
more accurate readings. In contrast, the simultaneous instrument does not have a
monochromator between the sample and the source; instead, it has a diode array
detector that allows the instrument to simultaneously detect the absorbance at all
wavelengths. The simultaneous instrument is usually much faster and more efficient,
but all of these types of spectrometers work well.

To determine the amount of caffeine in the soft drink sample.

A Preparation of Standard solution
1. 250 mg of caffeine was poured into 250 ml volumetric flask and diluted
to mark with distilled water to make 1000 ppm stock solution.
2. A series of concentration was prepared by diluting 12.5 ml, 10 ml, 7.5
ml, 5 ml and 2.5 ml by using a burette into 50 ml volumetric flask. The
solution was diluted to mark with distilled water.
B Caffeine extraction
1. 50 ml of aliquot was took from the standard solution and placed in
separating funnel.
2. 25 ml of dichloromethane was added into the funnel.
3. The mixture was shook until the caffeine standard was fully extracted.

C Sample preparation
1. 10 ml of soft drinks (chrysanthemum tea) was put into the beaker and
ready to measure the absorabnce.

In this experiment, Caffeine is a stimulant that is commonly found in many foods and
drinks that we consume. Caffeine is a common organic molecule found in many
beverages such as coffee, tea and cola. Like many conjugated organic molecules,
caffeine absorbs radiation with a wavelength around 260 nm. Caffeine in beverages
such as chrysanthemum tea was analyzed and determined by using UV-Vis
A series of standard caffeine were prepared which concentration are 0.5, 1.5, 2.5, 3.5
and 4.5 mg/L respectively. The readings of the absorbance for each of the
concentration are as the following, 0.0864, 0.2152, 0.4591, 0.6367 and 0.8044. The
correlation coefficient of the caffeine standard is 0.99953. The readings of the
absorbance of the samples are 2.5432, 2.5385 and 2.5572. The average reading of
the samples is 2.5463.

The average reading of caffeine in the sample of Chrysanthemum tea is 2.5463.