Download About UV-Vis Molecular Absorbance Spectroscopy

Spectroscopy Pre-Lab
In molecular absorbance spectroscopy a beam of ultraviolet or visible light is directed through a sample.
Some of the light may be transmitted through the sample. Light that was not transmitted through the
sample was absorbed. Transmittance (T) is defined as the ratio of P/Po. Absorbance (A) is defined as log(T).
A molecule can absorb some of the light only if it can accommodate that additional energy by promoting
electrons to higher energy levels. The energy of the light being absorbed must match the energy required
to promote the electron. Therefore, not all wavelengths of light are absorbed equally by a sample. An
absorbance spectrum depicts what wavelengths of light are absorbed by a sample. The UV-Vis
absorbance spectrum below was obtained by passing different wavelengths of light through a solution of
fabric dye, and measuring the intensity of light (P) passing through the solution. One can readily see what
wavelengths of light are absorbed (peaks), and what wavelengths of light are transmitted (troughs).
One very important relationship in absorbance spectroscopy is Beer's Law:
A=abc
Where: A is absorbance; a describes the ability of a molecule to absorb radiation at a particular
wavelength; b is the length of sample through which the light beam passes
c is the concentration of the absorbing species. This relationship is the basis of all quantitative work in
absorbance spectroscopy. It allows one to determine the concentration of an absorbing species simply by
measuring its absorbance.
Increasing Energy
Intoxilyzer
This device uses infrared (IR) spectroscopy, which identifies molecules based on the way they absorb IR light.
Molecules are constantly vibrating, and these vibrations change when the molecules absorb IR light. The changes in
vibration include the bending and stretching of various bonds. Each type of bond within a molecule absorbs IR at
different wavelengths. So, to identify ethanol in a sample, you have to look at the wavelengths of the bonds in
ethanol (C-O, O-H, C-H, C-C) and measure the absorption of IR light. The absorbed wavelengths help to identify
the substance as ethanol, and the amount of IR absorption tells you how much ethanol is there.
Diagram of the Intoxilyzer
Pre-Lab Questions:
1.) What is the relationship between chemicals and spectrum?
2.) What kind of mathematical relationship does Beer's Law represent?
3.) Which has greater energy—red or green light?
4.) Under what condition(s) would a chemical absorb light?
5.) Under what condition(s) would a chemical allow light to be transmitted through?
6.) If you transmitted red light through a chemical which resulted in zero absorbance, what might you do next to test
the chemical?
7.) Energy plays some kind of role in spectroscopy, what is that role (think in terms of identifying chemicals and
types of spectrum)?
8.) What statement can you make about chemicals and spectrum? What can toxicologists learn from your
statement?
9.) Suppose you are trying to identify the concentration of an unknown chemical: Using UV/IV spectrometry, how
would you design the experiment (or describe the steps you might take—be specific)?
10.) What questions do you have about spectrum or spectroscopy?