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Author Topic: The instumentations, use and principles of operations of a spectrophotometer  (Read 15546 times)
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Francis Umeoguaju
Expert in Bioscience Issues
Posts: 657

« on: April 10, 2010, 07:39:01 am »

A spectrophotometer is a device used for measuring light absorption of a particular solution at different wavelengths. It’s a recent modification to a colorimeter which defers in the sense that it works on a preset wavelength values which is generated by light filters.

The spectrophotometer is  an indispensable instrument in a laboratory. It is used to determine the concentration of samples by comparing their absorbance with that of known concentration at particular wavelength.

Types of spectrophotometer

There are several types of spectrophotometer but one common feature amongst them is that they generally falls into two groups; those that operates within the visible spectrum. The one encountered on the course of my working in the lab are the spectronic 21, 20 and UV- visible spectronic 21.

The visible spectronic 20 and 21  operates in the visible spectrum capable of generating wavelength of between 320mm to 1000mm.
The glass curvette and the disposable plastic curvette are used with it.

The UV- Visible spectronic 21  operates in both the visible and UV spectrum. It is capable of generating wavelength that ranges between 200mm to 1000mm. Quartz or silica curvettes is used when taking readings in the UV region, this is because glass absorbs at this region.

Principle of operation

The use of spectrophotometer is based on Beer-Lambert law which relates the amount of light transmitted through a colored solution to be a function of the length of light path and the concentration of the absorbing media.

A spectrophotometer is made up basically of a light source which can be the visible or ultraviolet region, a monochromator, the sample cell and the detectors or photocells.

The light source is usually a tungsten lamp (for spectrophotometer that operates in the visible region such as spectronic 20 and visible spectronic  21) and a hydrogen or deuterium lamp (for spectrophotometers capable of operating within the UV spectrum) the UV- visible “spec” are equipped  with the two types of lamps.

The incident light of particular wavelength selected with the monochromator is absorbed by the test solution resulting in lowered intensity of transmitted light. The photomultiplier sends signal to the photometer to display the result. The monochromator are made of grating or prisms to produce light with a slit width wavelength range of 5mm.


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