spectrophotometers

A spectrophotometer is an analytical instrument that measures light intensity as a function of wavelength to determine the concentration of substances in solution. Light passes through a sample and the transmitted light is measured; absorbance is inferred relative to a reference. Under suitable conditions, absorbance follows the Beer-Lambert law, being proportional to concentration and path length.

Most instruments are single-beam or double-beam. Single-beam models compare the sample to a baseline; double-beam models use a reference channel to compensate for source fluctuations. They are commonly configured for ultraviolet, visible, or near-infrared light (UV/Vis, NIR).

Key components include a light source (tungsten or deuterium lamp; LEDs in some models), a monochromator or filter, a sample holder or cuvette, a detector (photodiode or photomultiplier), and a readout system. Some units include temperature control and software for data analysis.

Applications include quantitative analysis of solutions, nucleic acids and proteins, enzyme kinetics, pharmaceutical quality control, and environmental monitoring. Spectral data can also aid material identification through wavelengths of maximum absorption.

Calibration involves wavelength and photometric accuracy checks and baseline correction. Limitations include sample turbidity, color interference, and deviations from Beer-Lambert behavior at high concentrations. Modern instruments offer good sensitivity, speed, and user-friendly software.