Ocean Optics CHEM2000 - Manual (Page 27)

3: Experiment Tutorial
2. Select Scope under Mode of Operation in the software display area. Make sure the signal is on
scale by adjusting acquisition parameters. Take a reference spectrum of your reference lamp.
Take the reference reading by clicking the Reference button in the software display area. (This
command merely stores a reference spectrum. To save a spectrum, you must select File | Save
Spectral Values from the menu.) Storing a reference spectrum is requisite before the software
can calculate relative irradiance spectra.
3. While still in Scope Mode, take a dark spectrum by first completely blocking light from going to
your spectrometer. Take the dark reading by clicking the Dark button in the software display
area. (This command merely stores a dark spectrum. To save a spectrum, you must select File |
Save Spectral Values from the menu.) Storing a dark spectrum is requisite before the software
can calculate relative irradiance spectra.
4. Begin a relative irradiance measurement by first positioning the fiber at the light or emission
source you wish to measure. Then select Relative Irradiance under Mode of Operation in the
software display area. Click on the Scan button in the display area to take a scan. If Single is
selected, only one scan will be taken. If Continuous is selected, the spectrometer will
continuously take scans until you click on the Stop button. To save the spectrum, select File |
Save Spectral Values from the menu.
Note
If at any time any sampling variable changes – including integration period, averaging,
boxcar smoothing, distance from light source to sample, etc. – you must store a new
reference and dark spectrum.
Concentration Experiments
The absorbance of a solution is related to the concentration of the species within it. The relationship,
known as Beer’s Law , is:
A
λ
= ε
λ
c l
Where:
A
= Absorbance at wavelength
λ,
ε
= Extinction coefficient of the absorbing species at wavelength
λ
c = Concentration
l
= Optical pathlength
Concentration is the amount of a specified substance in a solution. Graphs of absorbance vs.
concentration are known as Beer’s Law plots. These are prepared by measuring the light absorbed by a
series of solutions with different known concentrations. The length of the sample -- such as the path
length of our cuvette holder -- and the wavelength chosen for monitoring the amount of light absorbed are
constants. A linear plot from taking scans of these standard solutions with known concentrations is then
obtained. The plot is then used to determine the unknown concentrations of substances in solutions.
105-00000-CHM-02-0405 PRELIMINARY DRAFT 21