Agilent Technologies 1200 Infinity Series User Manual page 102

5
Optimizing the Detector
Optimizing the Detector Regarding to the System
Wavelength and Bandwidth
The detector measures absorbance simultaneously at wavelengths from
190 nm to 640 nm using diode-array detection. A UV-lamp provides good
sensitivity over the whole wavelength range. The diode-array detector (DAD)
can simultaneously compute and send to the data system up to eight
chromatographic signals and the full-range spectra at every time point.
A UV chromatogram or signal is a plot of absorbance data versus time and is
defined by its wavelength and bandwidth.
• The wavelength indicates the center of the detection band.
• The bandwidth defines the wavelength range over which the absorbance
For example, a signal at wavelength 250 nm with a bandwidth of 16 nm will be
an average of the absorbance data from 242 nm to 258 nm. Additionally, a
reference wavelength and reference bandwidth can be defined for each signal.
The average absorbance from the reference bandwidth centered on the
reference wavelength will be subtracted from its equivalent value at the signal
wavelength to produce the output chromatogram.
The signal wavelength and bandwidth can be chosen so that they are
optimized for:
• Broad band universal detection
• Narrow band selective detection
• Sensitivity for a specific analyte.
Broad band or universal detection works by having a wide bandwidth to
detect any species with absorbance in that range. For example, to detect all
absorbing molecules between 200 nm and 300 nm set a signal at 250 nm with a
bandwidth of 100 nm. The disadvantage is that sensitivity will not be optimal
for any one of those molecules. Narrow band or selective detection is used
most often. The UV spectrum for a particular molecule is examined and an
appropriate absorbance maximum is selected. If possible, the range where
solvents absorb strongly should be avoided (below 220 nm for methanol, below
210 nm for acetonitrile). For example, in
has a suitable absorbance maximum at 252 nm. A narrow bandwidth of 4 nm
to 12 nm generally gives good sensitivity and is specific for absorbance in a
narrow range.
The narrow band can be optimized for sensitivity for a specific molecule. As
the bandwidth is increased the signal is reduced but so is the noise and there
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values are averaged to give the result at each time point.
Figure 29 on page 104, anisic acid
Agilent 1200 Infinity Series DAD User Manual