|AN INTRODUCTION TO COLORIMETRIC ANALYSIS & SPECTROSCOPY
Most test substances in water are colorless and undetectable to the human eye. To test for their
presence we must find a way to "see" them. A colorimeter or
spectrophotometer can be used to measure any test substance that is itself colored or can be reacted
to produce a color. In fact a simple definition of colorimetry is "the measurement of color" and a colorimetric method is "any technique
used to evaluate an unknown color in reference to known colors". In a colorimetric chemical test the intensity of the color from the
reaction must be proportional to the concentration of the substance being tested. Some reactions have limitations or variances inherent
to them that may give misleading results. Most limitations or variances are discussed with each particular test instruction. In the
most basic colorimetric method the reacted test sample is visually compared to a known color standard. However, the eyesight of the
analyst, inconsistencies in the light sources, and the fading of color standards limit accurate and reproducible results.
To avoid these sources of error, a colorimeter or spectrophotometer can be used to photoelectrically
measure the amount of colored light absorbed by a colored sample in reference to a colorless sample (blank). A
colorimeter is generally any tool that characterizes color samples to provide an objective measure of
color characteristics. In chemistry, the colorimeter is an apparatus that allows the absorbance of a solution at a particular frequency
(color) of visual light to be determined. Colorimeters hence make it possible to ascertain the concentration of a known solute, since it is
proportional to the absorbance.
A spectrophotometer is a photometer (a device for measuring light intensity) that can measure intensity as a
function of the color, or more specifically, the wavelength of light. There are many kinds of spectrophotometers. Among the most
important distinctions used to classify them are the wavelengths they work with, the measurement techniques they use, how they acquire a
spectrum, and the sources of intensity variation they are designed to measure. Other important features of spectrophotometers include
the spectral bandwidth and linear range. The most common application of spectrophotometers is the measurement of light absorption.
White light is made up of many different colors or wavelengths of light. A colored sample typically
absorbs only one color or one band of wavelengths from the white light. Different chemical substances absorb varying frequencies of the
visible spectrum. Only a small difference would be measured between white light before it passes through a colored sample versus after
it passes through a colored sample. The reason for this is that the one color absorbed by the sample is only a small portion of the total
amount of light passing through the sample. However, if we could select only that one color or band of wavelengths of light to which the
test sample is most sensitive, we would see a large difference between the light before it passes through the sample and after it passes through
the sample. Colorimeters rely on the principle that the absorbance of a substance is proportional to its concentration i.e., a more
concentrated solution gives a higher absorbance reading.
Global Water's spectrophotometers use either a tungsten
or xenon flashlamp as the source of white light. The white light passes through an entrance slit and is focused on a ruled grating consisting
of 1200 lines/mm. The grating causes the light to be dispersed into its various component wavelengths. The monochromator design allows
the user to select which specific wavelength of interest will be passed through the exit slit and into the sample. The use of mirrors and
additional filters prevents light of undesired wavelengths (diffraction of higher order, stray light) from making it to the sample. A photodetector measures
the amount of light, which passes through the sample.
Global Water's colorimeters pass a colored light beam through an optical filter, which transmits only one
particular color or band of wavelengths of light to the colorimeter's photodectector where it is measured. The difference in the amount of
monochromatic light transmitted through a colorless sample (blank) and the amount of monochromatic light transmitted through a test sample
is a measurement of the amount of monochromatic light absorbed by the sample. In most colorimetric tests the amount of
monochromatic light absorbed is directly proportional to the concentration of the test factor producing the color and the path length through the
sample. However, for a few tests the relationship is reversed and the amount of monochromatic light absorbed is inversely proportional to
the concentration of the test factor.
The choice of the correct wavelength for testing is important. It is interesting to note that the wavelength that
gives the most sensitivity (lower detection limit) for a test factor is the complementary color of the test sample. For example the
Nitrate-Nitrogen test produces a pink color proportional to the nitrate concentration in the sample (the greater the nitrate concentration, the
darker the pink color). A wavelength in the green region should be selected to analyze this sample since a pinkish-red solution absorbs
mostly green light.