UVVis

UV-Vis spectroscopy is a technique that measures the absorption of ultraviolet and visible light by a sample. It relies on electronic transitions within molecules, with color arising from light that is not absorbed. The typical spectral range spans roughly 200 to 800 nanometers, encompassing both UV and visible regions.

In a measurement, a beam of light passes through a sample and the transmitted light is detected. The absorbance A is related to the incident and transmitted intensities and is commonly interpreted through the Beer-Lambert law: A = εlc, where ε is the molar absorptivity, l is the path length, and c is the concentration. This relation enables quantitative analysis over a practical concentration range, provided the sample remains within the instrument’s linear response.

Instrumentation often includes a light source (deuterium lamps for UV, tungsten-halogen for visible), a monochromator or filter to select wavelengths, a sample holder, and a detector (such as a photodiode, photomultiplier tube, or a diode array). Modern instruments frequently use a double-beam design to compensate for source fluctuations and may scan a full spectrum with a diode-array detector.

Applications are diverse: measuring concentrations of colored compounds, determining nucleic acid and protein content (commonly at 260 nm for nucleic acids and around 280 nm for proteins), and monitoring reaction kinetics. UV-Vis is widely used in chemistry, biochemistry, environmental analysis, and pharmaceutical testing. Limitations include dependence on chromophores, potential interferences, and the need for proper blanks, calibration, and consideration of solvent effects and scattering in turbid samples.