A spectrophotometer is a scientific instrument used to measure the intensity of light at different wavelengths. The instrument is commonly used in fields such as chemistry, biochemistry and environmental science. It can analyse the concentration of a substance in a sample based on its absorbance or transmittance. But how does a spectrophotometer work?
What is a visible spectrophotometer?
The UV-visible spectrophotometer is a very important class of analytical instruments with many important applications in several fields.
It breaks down the light emitted from a light source into different wavelengths using a spectroscopic element and measures the degree of absorption of this light by the sample.
What does a spectrophotometer measure?
A spectrophotometer uses spectrophotometry to measure the transmittance and absorbance properties of any given material as a function of wavelength to determine the concentration of an analyte.
In short, a spectrophotometer measures the intensity of light by wavelength and can be measured to derive the concentration of a solution according to Beer-Lambert law.
Components of a spectrophotometer
- Light source: The wavelength of high-intensity light provided by a light source.
- Monochromator: A device that breaks down the continuous spectrum emitted by a light source into monochromatic light
- Sample chamber: Used to hold the sample to be analyzed
- Detector: Light passing through the sample to be analyzed the detector, which uses the photoelectric effect to convert the light energy into a current signal.
- Display: Shows the transmittance of the sample and the detection parameters.
How does a spectrophotometer work?
The core component of a spectrophotometer is the entrance slit. The size of the slit determines the amount of light that can enter and be measured. This affects the speed of the spectrometer’s engine and the optical resolution. The smaller the slit size, the higher the resolution.
A lamp provides a light source. The light beam hits a diffraction grating, which works like a prism to separate light into its constituent wavelengths. The grating is rotated so that only light of a specific wavelength reaches the exit slit. The light then interacts with the sample. From this point on, the detector measures the transmittance and absorbance of the sample.
Transmittance is the amount of light that completely passes through the sample and hits the detector. Absorbance is a measure of the amount of light absorbed by the sample. The detector senses the light transmitted through the sample and converts this information into a digital display.
Application of visible spectrophotometer
Determination of substances
According to some characteristic absorption on the absorption spectra, especially the maximum absorption wavelength λ max and the molar absorption coefficient ε, the common physical parameters for testing substances.
Comparison with standards and standard charts
The analytical and standard samples are prepared in the same solvent with the same concentration, and the UV-visible absorption spectra are measured under the same conditions. If the two are the same substance, the spectra of the two should be identical. If there is no standard sample, it can also be compared with the ready-made standard spectra. This method requires accurate instrumentation and high precision; measurement conditions should be the same.
Reaction kinetics study
Several chemical reaction rate constants can be derived with spectrophotometry. The reaction activation energy is derived from the velocity data obtained under two or more temperature conditions.
Purity test
UV absorption spectroscopy can determine the presence of trace amounts of impurities with UV absorption in a compound. Suppose the compound has no obvious absorption peaks in the UV-visible region, and its impurities have strong absorption peaks in the UV region. In that case, the impurities in the compound can be detected.
Determination of hydrogen bond strength
Different polar solvents produce different strengths of hydrogen bonding, which can be used to determine the strength of hydrogen bonding of compounds in different solvents using ultraviolet spectroscopy to determine which solvent to choose.
Determination of complex composition and stability constants
Metal ions often form complexes with organic matter, and most of the complexes in the ultraviolet-visible region are absorbed; we can use spectrophotometry to study the composition.
Conclusion
In conclusion, a spectrophotometer is a versatile and invaluable tool in the world of science. By measuring the absorption or transmission of light at specific wavelengths, this instrument can provide valuable information about the concentrations of substances in a sample. From determining the purity of a compound to analyzing the composition of a biological sample, the spectrophotometer plays a crucial role in a wide range of scientific applications.
Get Social