Agilent InfinityLab LC Series Diode Array Detectors User Manual
88
5
Optimizing the Detector
Optimizing for Sensitivity, Selectivity, Linearity and Dispersion
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 88 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.
Figure 23 Optimization of Wavelength Setting
A wide bandwidth has the advantage of reducing noise by averaging over a
wavelength range — compared to a 4 nm bandwidth, the baseline noise is
reduced by a factor of approximately 2.5, whereas the signal is about 75 % of a
4 nm wide band. The signal-to-noise ratio for a 30 nm bandwidth is twice that for
a 4 nm bandwidth in our example.
Reference bandwidth 100 nm
Reference wavelength 360
Anisic acid
Wavelength (nm)
Absorbance (mAU)
sample wavelength 252 nm
30 nm
bandwidth
