Principles of Operation

The colour of every object we see is determined by a process of absorbtion and emission of the electromagnetic radiation (light) of its molecules. Colourometric analysis is based on the principle that many substances react with eachother and form a colour which can indicate the concentration of the substance to be measured. When a substance is exposed to a beam of light of intensity I₀ a portion of the radiation is absorbed by the substance's molecules, and a radiation of intensity I lower than I₀ is emitted.

The quantity of radiation absorbed is given by the Beer-Lambert Law: A=log I₀/I

Absorbance is also given by: A=ε_λ c d

Where:

ε_{λ: molar extinction coefficient of the substance at wavelength} λ;

c: molar concentration of the substance;

d: optical distance light travels through sample;

Therefore, the concentration (c) can be calculated from the colour of the substance determined by the emitted radiation (I), as the other factors are known. A typical block diagram of a photometer is shown below:

A monochromatic LED emits radiation at a single wavelength, supplying the system with the intensity (I₀). Since a substance absorbs the colour compliamentary to the one it emits (eg a substance appears yellow as it absorbs blue light), Hanna colourimeters use LEDs that emit the appropriate wavelegth to measure the sample.

The optical distance is measured by the dimension of the cuvet containing the sample. The photoelectric cell collects the radiation (I) emitted by the sample and converts it into an electric current, protducing a potential in the mV range. The microprocessor uses this potential to convert the incoming value into the desired measuring unit and to display it on the LCD.

In fact, the preparation of the solution to be measured occurs under known conditions, which are programmed into the meter's microprocessor in the form of a calibration curve. This curve is used as a reference for each measurement. It is then possible to determine unkown concentrations of a sample by inducing a colourimetric reaction, and thus obtain the mV related to the emitted intensity (I, the colour of the sample). By employing the calibration curve, one can determine the concentration of the sample that corresponds to the mV value.

The measurement process is done in two phases: setting the meter to zero and then the actual measurement. The first phase consists of collecting a sample of the substance in the calibration cuvet and inserting it into the meter. In this way, a reference value is set up, so that it is possible to establish how much the colour of the substance has varied in the next phase, after treating the sample in the measuring cuvet with a reagent.

The cuvet has a very important role because it is an optical element, and thus requires careful attention. First of all, it is important that boththe measurement and calibration cuvets are optically identical to provide the same measurement conditions. It is also necessary that the cuvet's surface is clean and not scratched, in order to avoid measurement interference due to unwanted reflection and absorbtion of light.