Liquid Chromatography - The Refractive Index Detector 1

In its simplest form, light from a fixed wavelength UV lamp passes through a cell, through which the column eluent flows and acts as the excitation source. Any fluorescent light that is emitted is sensed by a photo electric cell positioned normal to the direction of exciting UV light. The photo cell senses fluorescent light of all wavelengths but the wavelength of the excitation light can only be changed by use of an alternative lamp. This simple type of fluorescence detector was the first to be developed, it is relatively inexpensive and for certain compounds can be extremely sensitive. Typical specifications for a fluorescence detector are as follows:-

Typical Specifications for a Fluorescence Detector


Sensitivity (Anthracene)

1x 10-9 g/ml

Linear Dynamic Range

1 x 10-9 to 5 x 10-6 g/ml

Response Index

0.96 - 1.04

A more elaborate form of fluorescence detector uses a monochromator to select the excitation wavelength and a second monochromator to select the wavelength of the fluorescent light. This instrument gives the maximum versatility and allows the maximum sensitivity to be realized for any type of solute. The system can also provide a fluorescence spectra by arresting the flow of mobile phase when the solute resides in the detecting cell and scanning the fluorescent light.

The Refractive Index Detector

The refractive index detector is one of the least sensitive LC detectors. It is very sensitive to changes in ambient temperature, pressure changes, flow-rate changes and can not be used for gradient elution. Despite these many disadvantages, this detector is extremely useful for detecting those compounds that are nonionic, do not adsorb in the UV, and do not fluoresce. There are many optical systems used in refractive index detectors (9) but one of the most common is the differential refractive index detector shown diagramatically in figure 24.