Instrumentation:

Figure 4.3A shows a schematic diagram of a single-beam spectrophotometer. The light enters the instrument through an entrance slit, is collimated and focused on to the dispersing element, typically a diffraction grating. The light of desired wavelength is selected simply by rotating the monochromator and impinged on the sample. The intensity of the radiation transmitted through the sample is measured and converted to absorbance or transmittance (discussed later). Double beam spectrophotometers overcome certain limitations of the single beam spectrophotometers and are therefore preferred over them. A double beam spectrophotometer has two light beams, one of which passes through the sample while other passes through a reference cell (Figure 4.3B). This allows more reproducible measurements as any fluctuation in the light source or instrument electronics appears in both reference and the sample and therefore can easily be removed from the sample spectrum by subtracting the reference spectrum. Modern instruments can perform this subtraction automatically. The most commonly used detectors in the UV/Visible spectrophotometers are the photomultiplier tubes (PMT). Modern instruments also use photodiodes as the detection systems. These diodes are inexpensive and can be arranged in an array so that each diode absorbs a narrow band of the spectrum. Simultaneous recording at multiple wavelengths allows recording of the entire spectrum at once. The monochromator in these spectrophotometers is placed after the sample so that the sample is exposed to the entire spectrum of the incident radiation and the transmitted radiation is dispersed into its components.

Figure 4.3 Schematic diagram showing a single beam (A) and a double beam (B) spectrophotometer