Page 254 - Vogel's TEXTBOOK OF QUANTITATIVE CHEMICAL ANALYSIS

8 COLUMN AN0 THIN4AVER LlîlUlO CHROMATOCRAPHV

Column
mobile phase

Mirror

solvent
Fig. 8.4 Refractive index detector.

index of the liquid changes. In this detector both the column mobile phase
and a reference flow of solvent are passed through small cells on the back
surface of a prism. When the two liquids are identical there is no difference
between the two beams reaching the photocell, but when the mobile phase
containing solute passes through the ce11 there is a change in the amount of
light transmitted to the photocell, and a signal is produced. The smaller ce11
volume (about 3 PL) in this detector makes it more suitable for high-efficiency
columns but, for sensitive operation, the ce11 windows must be kept
scrupulously clean.
Ultvaviolet detectous. The UV absorption detector is the most widely used in
HPLC, being based on the principle of absorption of UV visible light as the
effluent from the column is passed through a small flow ce11 held in the radiation
beam. It is characterised by high sensitivity (detection limit of about
1 x 10-'g mL-' for highly absorbing compounds) and, since it is a solute
property detector, it is relatively insensitive to changes of temperature and flow
rate. The detector is generally suitable for gradient elution work since many of
the solvents used in HPLC do not absorb to any significant extent at the
wavelengths used for monitoring the column effluent. The presence of air bubbles
in the mobile phase can greatly impair the detector signal, causing spikes on
the chromatogram; this effect can be minimised by degassing the mobile phase
prior to use, e.g. by ultrasonic vibration. Both single and double beam
(Fig. 8.5) instruments are commercially available. Although the original
detectors were single- or dual-wavelength instruments (254 and/or 280 nm),
some manufacturers now supply variable-wavelength detectors covering the
range 210-800 nm so that more selective detection is possible.
No account of UV detectors would be complete without mention of the diode
array (multichannel) detector, in which polychromatic light is passed through
the flow cell. The emerging radiation is diffracted by a grating and then falls
on to an array of photodiodes, each photodiode receiving a different narrow-
wavelength band. A microprocessor scans the array of diodes many times a