The UV-Visible spectrophotometer is a sophisticated instrument designed to measure the intensity of light in the ultraviolet to visible light spectrum range after it passes through a sample.
The basic principle involves measuring how much a chemical substance absorbs light at different wavelengths.
Here's a breakdown of the primary components of a typical UV-Visible spectrophotometer
1. Sources of Radiation:
Radiation sources are responsible for emitting light, which is then analyzed after it interacts with the sample.
The type of source used depends on the range of wavelengths required for the analysis.
A. Deuterium Lamp:
Function: Produces UV radiation.
Construction: Contains deuterium (D2) gas. When an electric discharge is passed through the gas, it emits a continuous spectrum in the UV range (typically 190-380 nm).
Benefits: Offers a wide and almost continuous UV spectrum, making it ideal for various applications in UV spectroscopy.
B. Tungsten or Halogen Lamp:
Function: Emits radiation in the visible (VIS) and near-infrared (NIR) region.
Construction: Tungsten filament is the primary emitter, and in some cases, halogen gas is added to prolong the lamp's life. This lamp emits radiation from about 350 nm to 2500 nm.
Benefits: It provides a stable and intense source of light, with the halogen cycle ensuring a longer filament lifespan.
2. Wavelength Selectors:
Wavelength selectors, or monochromators, are used to select a specific range (or even a specific wavelength) of light from the broad spectrum emitted by the source.
A. Prisms:
Function: Disperse light based on the refractive indices of the prism material for different wavelengths.
Construction: Made of materials like quartz or specialized glass. As white light enters the prism, shorter wavelengths (blue/violet) are refracted more than longer wavelengths (red/yellow).
Benefits: Provide good separation of wavelengths and have been historically significant. However, they are less commonly used now due to the superior performance and flexibility of diffraction gratings.
B. Diffraction Gratings:
Function: Separate light based on interference patterns.
Construction: Consist of a surface with many closely spaced lines or grooves, which diffract and interfere with incident light to produce dispersed light.
Benefits: Offer high resolution, can handle high intensities of light without damage, and are more versatile than prisms. They're the most common monochromator in modern UV-VIS spectrophotometers.
3. Sample Cells:
Sample cells, or cuvettes, are containers in which the sample is placed for measurement. The light from the source passes through this cell, and the interaction of light with the sample is observed.
A. Material Selection:
Quartz: This is transparent in the UV and VIS regions. Ideal for measurements requiring wavelengths from the deep UV (around 190 nm) to the visible range.
Glass: Primarily transparent only in the visible region. Suitable for most visible light spectroscopy applications.
Plastic: Often used for disposable cuvettes, these are also generally transparent in the visible range. They might not provide the precision of quartz or glass but are useful for routine applications where contamination risks are high.
B. Shape and Design:
Most cuvettes are rectangular, allowing light to pass through the flat sides. This design ensures a uniform path length, which is crucial for quantitative measurements in spectroscopy.
The path length (typically the internal width of the cuvette where the light passes through) is usually standardized. Common path lengths are 1 cm, but other sizes are available for specialized applications.
Maintenance and Handling:
Cuvettes, especially quartz ones, need careful handling to prevent scratches or damages, as these imperfections can affect the accuracy of measurements.
It's essential to clean cuvettes thoroughly between measurements to prevent contamination and carryover effects.
4. Detectors:
Detectors are vital as they convert the light that emerges from the sample into an electrical signal that can be measured and analyzed.
A. Photo Tube (Phototube or Photocell):
Operates on the photoelectric effect.
Contains a photosensitive material that releases electrons when struck by photons.
The emitted electrons produce an electric current proportional to the light's intensity.
B. Photomultiplier Tube (PMT):
A highly sensitive version of the phototube.
It comprises a series of electrodes called dynodes.
Electrons emitted from the photosensitive material strike the first dynode, leading to the emission of more electrons. This process gets repeated across multiple dynodes, amplifying the signal at each step.
C. Photo Voltaic Cell (Solar Cell):
Converts light into electricity directly.
When photons strike the cell, they displace electrons, generating a direct current.
Commonly made of semiconductor materials like silicon.
D. Silicon Photodiode:
A type of photodetector capable of converting light into either current or voltage.
Operates similarly to the photovoltaic cell but is often used in a reverse bias condition, enhancing its sensitivity and response speed.
