Sample And Reference Wavelength And Bandwidth - Agilent Technologies 1200 Infinity Series User Manual

Sample and Reference Wavelength and Bandwidth

The detector measures absorbance simultaneously at wavelengths from 190 to
640 nm. A UV-lamp provides good sensitivity over the whole wavelength range.
If you know little about the analytes in your sample, store all spectra over the
full wavelength range. This provides full information but fills up your disk
space rather quickly. Spectra can be used to check a peak's purity and
identity. Spectral information is also useful to optimize wavelength settings
for your chromatographic signal.
The detector can compute and store at run time up to 8 signals with these
properties:
• sample wavelength, the center of a wavelength band with the width of
• reference wavelength, the center of a wavelength band with the width of
The signals comprises a series of data points over time, with the average
absorbance in the sample wavelength band minus the average absorbance of
the reference wavelength band.
Signal A in the detector default method is set to sample 254.0/4, reference
360.0/100, that is, the average absorbance from 252 – 256 nm minus the
average absorbance from 310 – 410 nm. As all analytes show higher
absorbance at 252 – 256 nm than at 310 – 410 nm, this signal will show you
virtually every compound which can be detected by UV absorbance.
Many compounds show absorbance bands in the spectrum.
page 90 shows the spectrum of anisic acid as an example. To optimize for
lowest possible detectable concentrations of anisic acid, set the sample
wavelength to the peak of the absorbance band (that is, 252 nm) and the
sample bandwidth to the width of the absorbance band (that is, 30 nm). A
reference of 360,100 is adequate. Anisic acid does not absorb in this range.
If you work with high concentrations, you may get better linearity above
1.5 AU by setting the sample wavelength to a valley in the spectrum, like
225 nm for anisic acid.
Agilent 1200 Infinity Series DAD User Manual
Optimizing for Sensitivity, Selectivity, Linearity and Dispersion
sample bandwidth (BW), and optionally
reference bandwidth.
Optimizing the Detector
Figure 20 on
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