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Study Notes: UV-Vis Sample Holder

The predominant use of UV-Vis spectroscopy is the measurement of samples that are in liquid or solution forms. The container which holds the liquid or solution is known as a cell or cuvette.

The cell is placed inside the cell compartment of the spectrophotometer after which the instrument’s lid is closed and monochromatic light is directed through the cell.

Simplified internal layout of spectrophotometer components from light source to monochromator to sample cell. White light is emitted from the source and light of different colours is transmitted to the cell.

Sample cells are normally a square or rectangular tube with two polished, optically smooth faces on opposite sides parallel to each other. The distance between the inside faces is the optical path length (remember Beer’s Law) and this can vary but the most popular size is 1cm (ie 1.00 cm, or very close to it!).

The cells are oriented in the cell compartment of the spectrophotometer with the polished faces directed towards the optical beam. If not, nonsense results will occur so beware.

The sample cell must be made from a material that allows the light beam to pass straight through with a constant low level of absorption (or maximum transmission). In effect, the cell must be transparent to the chosen wavelength of the monochromatic light. Diagrammatic representation of transparency in UV, Vis and IR ranges of cells of different materials of construction.

Glass is suitable for visible light and so too is NaCl but only with non aqueous samples. Quartz cells can be used for both visible and ultraviolet light (glass however is opaque to UV light and must not be employed for analysis in this part of the spectrum). Plastic cells can also be used for visible light - they are normally disposed of after a single use.

Glass and quartz cells are expensive and must be handled carefully to avoid scratches and finger marks that may absorb light or disperse it away from the detector thereby introducing inaccuracy. This is a key consideration.

The analyst should always check after filling a cell that there are no occluded air bubbles in the solution. This is another source of error.

Grip the cells on their non-optical surfaces - these can have a frosted appearance. Ensure that the cells are properly located in the cell holders with the optical faces towards the radiation. With age and use cell holders can become a little loose and this can lead to the cell being positioned slightly ‘off square’ thereby introducing error into readings.

Where different cells are used in the same analysis (for example, for blank and sample determinations) they must be spectrally matched. This means that they must behave identically, or almost so, as judged by an insignificant difference in absorbance (or transmission) from one cell to the other.

Proper cell management is the key to reliable results!

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