Double-beam UV/Vis Spectrophotometer
- For a double-beam UV/Vis Spectrophotometer:
- • A beam of light from a visible and/or UV light source is separated into its component wavelengths by diffraction grating or monochromator.
- • Each monochromatic (single wavelength) beam in turn is split into two equal intensity beams by a half-mirrored device.
- • One beam, the sample beam, passes through a small transparent container (cuvette) containing a solution of the compound being studied in a transparent solvent.
- • The other beam, the reference, passes through an identical cuvette containing only the solvent.
- • The intensities of these light beams are then measured by electronic detectors and compared.
- • The intensity of the reference beam, which should have suffered little or no light absorption, is defined as I0.
- • The intensity of the sample beam is defined as I.
- • Over a short period of time, the spectrometer automatically scans all the component wavelengths in the manner described.
Samples for UV/Vis spectrophotometry are most often liquids, although the absorbance of gases and even of solids can also be measured. Samples are typically placed in a transparent cell, known as a cuvette. A cuvette can have either a circular or square cross section and can be made of either plastic, glass, or optical grade quartz glass (see Figure 6).
Plastic cuvettes dissolve in most organic solvents and absorb light in the ultraviolet region; therefore, they can only be used for aqueous solutions and measurements in the visible range of the spectrum (about 400 to 700 nm wavelengths). Glass cuvettes can be used with most solvents, however they too absorb in the UV range. Quartz cuvettes do not absorb in the UV-Vis spectrum and can be used with virtually all samples; however, they are very expensive.