THE SPECTROPHOTOMETER Background

A spectrophotometer is an instrument designed to detect the amount of radiant light energy absorbed by molecules. To do this, the instrument must have five basic components:

1. a light source; 2. a prism or diffraction grating; 3.an aperture or slit;/ monochromator 4. a detector (a photoelectric tube); 5.a digital meter to display the output of the phototube. The arrangement of these parts is shown below. When light is reflected from a diffraction grating, it is split into its component colors or wavelengths, which then diverge. Sections of the projected spectrum can be either blocked or allowed to pass through the slit so that only one wavelength will pass to the other sections of the spectrophotometer (The position of the grating is adjustable so that the region of the spectrum projected on the slit can be changed.). Light that passes through the exit slit travels to the

phototube/sample, where it creates an electric current proportional to the number of photons striking the phototube. THE SAMPLE IN A CUVET , ABSORB A PORTION OF LIGHT , AND THE REST IS TRASMITED THROUGH THE SAMPLE. THE TRANSMITTED BEAM STRIKES A DETECTOR, WHICH CONVERT RADIANT ENERGY INTO ELECTRICAL ENERGY PROPORTIONAL TO THE AMOUNT OF LIGHT. A READ OUT DEVICE PRESENTS THE ELECTRICAL SIGNAL IN USEFUL UNITS. Note that the relationship between absorbance and concentration is linear. As concentration increase the absorbance also increases. This relationship allows one to convert an absorbance value into a concentration.