Optimizing For Sensitivity, Selectivity, Linearity And Dispersion; Flow Cell Path Length - Agilent Technologies G1315B User Manual

Optimizing for Sensitivity, Selectivity, Linearity and Dispersion

Flow Cell Path Length

Lambert-Beer's law shows a linear relationship between the flow cell path
length and absorbance.
where
T is the transmission, defined as the quotient of the intensity of the
transmitted light I divided by the intensity of the incident light, I
ε is the extinction coefficient, which is a characteristic of a given substance
under a precisely-defined set of conditions of wavelength, solvent,
temperature and other parameters,
C is the concentration of the absorbing species (usually in g/l or mg/l), and
d is the path length of the cell used for the measurement.
Therefore, flow cells with longer path lengths yield higher signals. Although
noise usually increases little with increasing path length, there is a gain in
signal-to-noise ratio. For example, in
than 10 % but a 70 % increase in signal intensity was achieved by increasing
the path length from 6 mm to 10 mm.
When increasing the path length, the cell volume usually increases — in our
example from 5 – 13 µl. Typically, this causes more peak dispersion. As
Figure 26
separation in our example.
As a rule-of-thumb the flow cell volume should be about 1/3 of the peak
volume at half height. To determine the volume of your peaks, take the peak
width as reported in the integration results multiply it by the flow rate and
divide it by 3).
1200 Series DAD and MWD User Manual
Absorbance log T
= –
demonstrates, this did not affect the resolution in the gradient
I
0
ε ⋅ ⋅
log --- - C d
= =
I
Figure 26
How to optimize the Detector
,
0
the noise increased by less
5
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