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No single technique is perfect and UV-Vis spectroscopy is no exception. But the technique has a few key strengths that make it popular.
=>> The technique is non-destructive, allowing the sample to be reused or to proceed to further processing or analysis.
=>> Measurements can be made quickly, allowing easy integration into experimental protocols.
=>> The devices are easy to use, requiring little user training before use
=>> Data analysis typically requires minimal processing, again meaning little user training is required.
=>> The instrument is generally inexpensive to acquire and operate, making it accessible to many laboratories.
Although the strengths of this strategy seem overwhelming, there are also some weaknesses:
Stray light – In a real instrument, wavelength selectors are not perfect and small amounts of light from a wide wavelength range may still be transmitted from the light source, 1 potentially causing serious measurement errors. 9 Stray light can also come from the environment or a loosely fitted compartment within the instrument
Light scattering – Light scattering is often caused by suspended solids in liquid samples, which can cause serious measurement errors. The presence of bubbles in the cuvette or sample will scatter light, resulting in irreproducible results.
Interference of multiple absorbing species – A sample, for example, may contain multiple types of the green pigment chlorophyll. Different chlorophylls will have overlapping spectra when tested together in the same sample. For an accurate quantitative analysis, each chemical species must be separated from the sample and tested separately.
Geometrical Considerations – Misalignment of any component of the instrument, especially the cuvette holding the sample, can lead to irreparable and inaccurate results. Therefore, it is important that each component of the instrument is aligned in the same orientation and placed in the same position for each measurement. Some basic user training is therefore generally recommended to avoid misuse.
