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Molecules such as FAD (flavin adenine dinucleotide), NADH and NADþare impor-

tantcoenzymesof proteins involved in electron transfer reactions (RedOx reactions).

They can be conveniently assayed by using their UV/Vis absorption: 438 nm (FAD),

340 nm (NADH) and 260 nm (NADþ).

Chromophores in genetic material

The absorption of UV light by nucleic acids arises fromn!p* andp!p* transitions of

the purine (adenine, guanine) and pyrimidine (cytosine, thymine, uracil) bases that

occur between 260 nm and 275 nm. The absorption spectra of the bases in polymers

are sensitive to pH and greatly influenced by electronic interactions between bases.

12.2.2 Principles

Quantification of light absorption

The chance for a photon to be absorbed by matter is given by anextinction coefficient

which itself is dependent on the wavelengthlof the photon. If light with the

Wavelength (nm)

250

Absorbance

300 350 400 450 500

Fig. 12.5The presence of larger aggregates in biological samples gives rise to Rayleigh scatter visible by a

considerable slope in the region from 500 to 350 nm. The dashed line shows the correction to be applied to

spectra with Rayleigh scatter which increases withl^4. Practically, linear extrapolation of the region from 500

to 350 nm is performed to correct for the scatter. The corrected absorbance is indicated by the double arrow.

485 12.2 Ultraviolet and visible light spectroscopy