Spectrophotometry

Describe the theories behind the science of spectrophotometry. Differentiate spectroscopy with
spectrophotometry. Identify key applications of this kind of chemical analysis. Why is this analysis method
not as popular as with other chemical analysis methods?
In analyzing chemical solutions, spectrophotometry uses a light beam that passes through the sample held
in a cuvette, and each compound in the solution absorbs or transmits light over a specific wavelength.
Spectrophotometers are built to transmit light of narrow wavelength ranges. Different compounds do not
absorb wavelengths equally, which is why a spectrophotometer is a tool used to distinguish compounds by
examining the pattern of the wavelengths absorbed by a solution. In the schematic diagram of a
spectrophotometer shown in the lecture notes, the source provides the electromagnetic radiation that is
absorbed by the sample. The monochromator (can be a prism) separates the light from the light source into
one particular energy or wavelength/color. The photoresistor is the detector that measures the amount of
light that passes through the solution. Spectrophotometers report measurements as transmittance or
absorbance. The amount of light transmitted passed through the solution (transmittance) or absorbed by the
solution (absorbance) when light of a particular wavelength is passed through a solution inside a cuvette is
measured by a light meter. When absorbance is 100%, the transmittance is 0%. A spectrophotometer can
also be used to determine concentrations of compounds in solution. The amount of light absorbed is directly
proportional to the concentration of absorbing compounds in the solution. This relationship between
concentration and absorbance is expressed by Beer-Lambert Law [A = εcι] where A is absorbance, ε (m-
1
cm-1) is molar absorption coefficient, c (M) is molar concentration, and ι (cm) is the optical path length.
Spectroscopy is the study of the absorption and emission of electromagnetic radiation or light by matter
(Levine, 2009, p. 737). It is a general study of the interaction of matter with electromagnetic waves or the
whole spectra. In contrast, spectrophotometry is the measurement of absorption of radiation in a particular
spectral region (Atkins & de Paula, 2006, p. 792). It is a technique to measure light absorption or the number
of chemicals in a solution. Spectrophotometric analysis is typically used as a quantitative estimation and
identification of compounds. It is most useful when one substance in the reaction mixture has a strong
characteristic absorption in a conveniently accessible region of the electromagnetic spectrum. The key
difference is that spectrophotometry uses a device composed of a spectrometer and a photometer. The
spectrometer is responsible for producing the desired range of wavelength light, while the photometer is
the device that detects the number of photons absorbed. Depending on the range of wavelengths of the light
source, a spectrophotometer can be classified as UV-Vis or IR. A UV-Vis spectrophotometer uses a light
over the ultraviolet (UV) range of 185-400 nm and visible range of 400-700 nm of the electromagnetic
radiation spectrum. Spectrophotometry can be used in any application dealing with chemical substances or
materials. In chemistry, spectrophotometry is typically used to analyze analytes such as transition metal
ions, highly conjugated organic compounds, and biomolecules. Several variations of spectrophotometry
exist, such as atomic absorption spectrophotometry and atomic emission spectrophotometry. Although a
conventional and inexpensive technique, spectrophotometry has limitations, including low sensitivity and
selectivity. The Beer-Lambert law can only describe the absorption behavior of dilute solutions, preferably
solutions containing relatively low amounts of solutes dissolved in them. Another factor to consider is that
the solutions must be colored so the appropriate color of light or wavelength from the light source that is
best absorbed by the solution can be selected.

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