Chemistry Reference
In-Depth Information
Anodes
Beam of
electrons
Light
Cathode
Quartz
window
Figure 7.15
A diagram of a photomultiplier tube.
absorbance. This is often done automatically during the 'warm-up' by modern
instruments.
The shutter is opened, the solvent (or 'blank') is placed in the light path and the
instrument is set to zero absorbance. The blank is usually just the solvent for
the assay but, strictly speaking, should be everything in the sample matrix except
the sample being measured. This means that in complex assays the blank solution
has to be made up to match exactly the composition of the solvent/medium in
which the sample will be measured, and has to be extracted or otherwise treated
in exactly the same way as the sample.
2.
The sample solution (or 'test') is placed in the light path and the absorbance is
read directly by the instrument.
3.
Dilutions
The most important part of any spectroscopic assay is not the performance
of the spectrophotometer (although the accuracy of the instrument is
checked periodically). The crucial part of any experiment is the accurate
preparation of the test and standard solutions. This often involves the ac-
curate dilution of a stock solution using the volumetric glassware introduced
in Chapter 6, namely the pipette and the volumetric flask.
A common procedure is to prepare a range of dilutions for use as a
calibration graph as in the worked example below.
