Inside the Machine UV-Visible Spectrophotometer Components Quiz

The light source provides the initial radiation. In UV-Vis instruments a combination of a deuterium lamp for the UV range and a tungsten halogen lamp for the visible range is typically used to cover the entire spectrum from 190 to 800 nanometers.

The monochromator uses a prism or a diffraction grating to disperse the light from the source into its component wavelengths. It then uses an exit slit to select a single narrow band of light to pass through the sample.

Cuvettes must be made of materials that do not absorb light in the region being studied. Quartz cuvettes are required for UV work because standard glass and plastic absorb ultraviolet radiation which would interfere with the measurement.

A photomultiplier tube is a highly sensitive detector that converts incident photons into an electrical signal. It amplifies the signal internally making it possible to detect very low levels of light transmitted through a sample.

The width of the slits determines the range of wavelengths known as bandwidth that reaches the sample. Narrower slits provide better spectral resolution but decrease the intensity of light requiring a more sensitive detector.

The light must be selected for wavelength by the monochromator and then pass through the sample before reaching the detector. If the detector were first it would receive all wavelengths at once making it impossible to measure specific absorption.

The detector such as a photodiode or photomultiplier tube generates a current proportional to the intensity of the light hitting it. This current is then processed by the electronics to display an absorbance or transmittance value.

A diffraction grating is a reflective surface with thousands of closely spaced parallel grooves. It is more efficient and provides better linear dispersion than traditional glass prisms allowing for more precise wavelength selection.

The blank contains the solvent and all reagents except the analyte. By measuring the blank first the instrument can subtract the background absorption caused by the solvent and the cuvette walls ensuring the final reading is due only to the sample.

A photodiode array detector allows for extremely fast analysis because it captures the entire spectrum simultaneously. This is especially useful in liquid chromatography where the chemical composition of the flow is changing rapidly.

Fingerprints and bubbles scatter light leading to falsely high absorbance readings. Dust on optics reduces the intensity of light reaching the detector. Quartz is the correct material for UV work and would not cause an error.

Tungsten lamps are used for visible light while deuterium lamps are the standard for ultraviolet light. Most modern instruments automatically switch between these two sources depending on the wavelength selected.

A double beam instrument uses a beam splitter to send light through both the sample and a reference blank. This allows the instrument to automatically compensate for fluctuations in light source intensity or solvent absorbance.

For visible light above 340 nanometers inexpensive plastic or glass cuvettes are sufficient. Quartz is also suitable but is generally reserved for UV applications because it is more expensive. Lead is opaque and would block all light.

The collimator usually a curved mirror or lens takes the divergent light from the source and aligns it into a parallel beam. This ensures that the light enters the monochromator or sample compartment with a uniform path.