A spectrophotometer is an instrument that measures the interaction of light with matter across different wavelengths of the electromagnetic spectrum. It works by passing a beam of light through a sample and comparing the intensity of the light before and after it interacts with samples including”solutions, transparent or opaque solids, such as polished glass, or gases,” as Wikipedia has noted. The spectrophotometer consists of a light source, a sample holder, a diffraction grating or prism to separate the light into its component wavelengths, and a detector to measure the intensity of the light at each wavelength.

Spectrophotometers are widely used in scientific research and industry to measure the intensity of light absorbed or transmitted by a sample. A search on Google scholar for “spectrophotometer applications” yields more than two million results. Fields where the device is used include chemistry, biology, physics, and materials science, by providing quantitative data on the concentration and properties of substances.

The importance of Beer–Lambert Law in spectrophotometry

Spectrophotometry’s roots can be traced back to the 19th century with the pioneering work of scientists like August Beer and Johann Heinrich Lambert, who laid the foundation for the Beer–Lambert Law (also known as Beer’s Law), which is a fundamental principle at work in spectrophotometers. The Law correlates the absorption of light to the properties of the material in which the light travels). The law states that the absorbance (A) is directly proportional to the concentration (c) of the absorbing species in the solution and the path length (l) of the sample, expressed as A = εcl. In the equation ε is the molar attenuation coefficient. In chemical analysis, for instance, this principle allows spectrophotometers to determine the concentration of copper(II) ions in water samples by measuring the absorbance of a blue copper-ammonia complex at 620 nm.

The Beer–Lambert Law is used in chemistry to measure the concentration of chemical solutions, analyze oxidation processes, and assess the degradation of polymers. It also explains how radiation is attenuated as it passes through the Earth’s atmosphere.

Sample spectrophotometer applications

Spectrophotometers find applications across various scientific disciplines:

• Pharmaceutical researchers use spectrophotometry to study drug-protein interactions, observing spectral changes when drugs interact with proteins, which can help understand drug efficacy and safety. The device has also been used in explore drug-protein interactions.
• In forensic science, spectrophotometry aids in analyzing evidence, such as estimating the age of bloodstains by tracking spectral changes over time.
• Physical and theoretical chemists apply spectrophotometry to study reaction kinetics, measuring absorbance changes over time to understand the dynamics of chemical reactions.
• In industrial manufacturing, spectrophotometry plays a crucial role in quality control, ensuring consistent production in processes such as paint color matching, thus maintaining accuracy in manufacturing.

Types of spectrophotometers

Spectrophotometers are grouped into a variety of classes. Single-beam spectrophotometers measure the intensity of light before and after it passes through the sample, while double-beam spectrophotometers split the light into two beams, one passing through the sample and the other through a reference, allowing for more accurate measurements. Different types of spectrophotometers operate across specific regions of the electromagnetic spectrum. UV-Vis spectrophotometers are commonly used for analyzing compounds in the ultraviolet and visible light regions, while IR spectrophotometers help researchers study molecular vibrations in the infrared region.