Spectrophotometry/Catalog of methods in biophysics

When optical radiation passes through a substance, its intensity gradually decreases due to absorption. The quantity characterizing the degree of absorption in a substance is called absorbance. An instrument for measuring absorption spectra of radiation that has passed through an environment is called an absorption spectrophotometer. It records the ratio of the intensity of the radiation absorbed by the measured sample to the intensity of the reference (standard) radiation depending on the wavelength of the radiation incident on the sample. Relative intensity values ​​ranging from 0 to 100% or directly absorbance values ​​are usually recorded. The optical part contains a radiation source (light bulb for the visible region, hydrogen discharge lamp for the ultraviolet region), monochromator (prism or grating), detector (photocell, photomultiplier), and optical elements. The electromechanical part includes the monochromator tuning drive, detector signal meter and output device (computer). The polychromatic radiation of the source is split into spectral components by a prism or monochromator grating, and the desired radiation is isolated by a slit, the width of which determines the spectral purity of the radiation. Unabsorbed radiation hits the detector. Modern registration spectrophotometers draw an absorption spectrum in a few minutes.

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