Using Beers Law

For a compound to be analyzed by spectrophotometry, it must absorb electromagnetic radiation, and this absorption should be distinguishable from that of other species in the sample. Biochemists assay protein solutions in the ultraviolet region at 280 nm, where the aromatic amino acids tyrosine, phenylalanine, and tryptophan (Table 11-1) have maximum absorbance. Common salts, buffers, and carbohydrates have little absorbance at this wavelength. In this section, we use Beer s law for a simple analysis and then discuss the measurement of nitrite in an aquarium. 

Spectrophotometric analysis with visible radiation is called colorimetric analysis. Box 18-2 gives an example of the rational design of a colorimetric analysis. 

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The concentrations of all oligonucleotide stock solutions are determined by their UV absorbance. Values of molar absorbtivity coefficients for the various bases at neutral pH are taken from CRC Press Handbook of Biochemistry and Molecular Biology. For heteropolymers, concentrations are determined at 260 nm by summing the absorbtivity coefficients for all bases in the oligomer and using Beers Law, Homopolymers are similarly quantitated at their respective wavelength of maximum absorbance. 

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