Colorimeter and Flame Photometry Principle and Uses

Colorimeter Principle

In the colorimeter technique, a particular reagent is utilized which responds with the particular segment to colorimeter structure shaded complex. In a pencil of light is permitted to go through a shaded arrangement some measure of light saw to corresponding to the idea of the arrangement. The convergence of the solute or shading and the separation of the way of light is known as bear's law.

                              A=abc

Where

          A = Absorbance

          a = Nature of solution

          b = Distance of the path of light

          c = Concentration

The photodetector system, however, can only measure the transmitted lights. If the scale of the photometric electrometric colorimeter is in, it should be first be converted to absorbance before using the value for the calculation of the concentration of the unknown.

As Beer's law shows the immediate connection among absorbance and focuses the transmittance perusing is first changed over to be absorbance scale for setting up the standard bend.

Uses Of Colorimeter

Colorimeter use in emergency clinics and labs for the estimation of biochemical examples like the serum, plasma, cerebrospinal liquid, and pee. They are additionally utilized by the nourishment business and assembling of painting and textline organization.

Component Of Photometer

1. Light Source  

This provides radiant energy. Tungsten lamp is used for the UV range and lower visible.

2. Monochromatic Filter

These systems provide the desired quality of light by using filters or grits. The latter provides a narrow spectrum of light. The grits have partially replaced the prism because they are expensive. The complementary filter is used to expand the particularity and affectability of optical estimations, the particular nature of light is permitted to enter the shaded arrangement.

The quality of the light-regulated by monochromator in case of the spectrometer, which allows one to select a specific wavelength of light to pass through to test solution. In the common stage, an electrical photometer refers to colorimeters or spectrophotometer. The colorimeter use filter while spectrometer use prism or grit that yields a single selected wavelength of light.

3. Slit

The light emerging from the monochrometer is than pass through a narrow slit that improves the quality of light by choosing a single wavelength. This is found in all spectrophotometers.

4. Cuvette

The cuvette holds the solution whose absorbance is to be measured the cuvette must be optically transparent, scruple clean, devoid of any scratch, and free from contamination. The cuvette is either round like a test tube or rectangular.

The optical path within the cuvette is always" 1cm ". Glass cuvettes are utilized in noticeable range colorimetry or spectrophotometry while quartz or silica cells are utilized for UV radiation. This is because glass observes lights. Special plastic cuvettes are known available which are specially used for calorimetry.

Some the production gives a characteristic of the cuvette which ought to be by the imprint given on the colorimeter. Clearing of the cuvette must be done with lens paper or soft tissue paper. Never clean the cuvette with chronic acid, use only mildly good quality detergent.

Flame Photometry

The two important electrolytes of the body sodium and potassium are conveniently and accurately analyzed by flame photometry or flame emission photometry. The request for the analysis of sodium and potassium is most often a stat procedure where the speed and the accuracy of the operation is critical.

Flame photometry meets this requirement. In addition to Na and K flame photometer also analyze the concentration of lithium(Li). Lithium is used in therapy does for some mental patients and the level to be maintained in the serum is critical.

The recent introduction of the ion-selective electrode will eventually replace the flame photometer. The ion-selective electrode is more accurate and relatively simpler to use.

Principle of Flame Photometry

Several elements like Na, K, Ca, Li AND Cs(The earth metal) when place in the heat of flame become excited and emit light of a specific wavelength which is characteristics while they return to their ground state. The amount of light emitted is proportional to the number of excited atoms present or in other words concentration of the elements in solution.

Quantization of the emitted light is done by the photodetector system. The various elements that can be radically analyzed by flame emission photometry only sodium and potassium routinely done. Li only occasionally does on special request from mental hospitals. Analysis of Ca by flame emission photometry is not every popular as it requires a high temperature to volatilize(2000-2500 degree Celcius) the element as compared to Na and K(1500).

The specific wavelength for the clinically significant element is sodium(589 nm or yellow), potassium(1404 nm or violet and 767 nm or red), and Lithium (671 nm or red). By setting appropriate monochromators and corresponding, photocells for each element to be analyzed, the concentration of different elements can be determined simultaneously from the same flame. As the absorption maxima are wide apart for sodium(589nm) and potassium(766nm).

Siple filters are required as monochromators to provide specific wavelengths. The flame is the light source for flame emission photometry. It is burnt with a mixture of propane and butane. The aqueous specimen is sprayed into flame as a fine mist by means of a nebulizer or aerosol.

Components of Flame Emission Photometry

1. Nebulizer - This helps to spray the specimen into the burner. It is usually of the scent type. Whereby a forced stream of air passes over a capillary tube that dips into the test solution.

2. Flame - The flame generates heat that volatilizes the element that becomes luminous when it returns to the ground state. The most common gas mixture that provides an optimum temperature for the routine determination of sodium and potassium is propane, Butane, or natural gas.

The mixture of the fuel gas and air in the right proportions is ignited which provides the flame. The purpose of the flame is to generate heat and volatilizes the elements that become luminous when they return to the ground state.

3. Monochromators - These devices increase the specificity of the analysis. Simple filters are used that yield 589nm for Na and 767 nm for K. These are not complementary, filters against the color of the flame. Along with the monochromator filters, a heat filter is placed between the flame and monochromator filter to stop the passage of heat.

The monochromator screen out the wavelength of light except the specific one emitted by the element analyzed to separate the monochromator is used or each element analyzed. 

4. Photo Detector - The photodetector system quantifies the emitted light by converting it into an electrical impulse which is eventually transmitted to the readout galvanometer.

Procedure Of Photometer

1.     Dilution of the specimen and std (1:200) if the internal std is to be used, specimen and std are dilute with the internal std solution. The instrument may be provided with a combined automated dilution and feeding system this saves time in dilution. In most laboratories of developing countries, manual dilution is done.

2.     Turn off the electrical connection to the flame photometer. Permit it to balance out for a couple of moments. Ignite the flame by pressing the ignition button on the instrument panel.

3.     Put the deionizer water under the inlet tube. Water will be sucked in at a specific rate.

4.     Replace the water with the std solution and set the level for sodium and potassium.

5.     Put the dilution specimen and take the reading.

6.     Run the quality control sera occasionally to check the validity of the results

7.     To run off machine, first, close the gas value.