Practical Issues - Agilent Technologies 1290 Infinity II 2D-LC User Manual

14 Theoretical Background

Practical Issues

Practical Issues

The table below gives an overview, which practical issues have to be considered
in 2D-LC.
Table 47 Practical issues in 2D-LC
Issue Theoretical base
Choice of first Has impact on trade off between optimum first dimension flow
dimension column rate and amount of sample injected into the second dimension
diameter column for each second column run
Ratio of column Causes significant analyte dilution effects
diameter in the two
dimensions
Goals of the analytical Chosen parameter depend on what is important in analysis:
method •
•
Selection of the For RPLC in both dimensions the retentivity of the second
stationary phases and dimension column must be much higher than that of the first
column formats dimension column required because:
•
•
Based on theory, in most cases following approaches to achieve best possible
2D-LC should be respected:
•
Take care when using any unconventional organic solvents that these are still
NOTE
compatible with the used instrumentation. In doubt, refer to the module
documentation or call Agilent.
1
In case the peak and time triggered operation of the second dimension separation, which is
optionally available with the Agilent 1290 Infinity 2D-LC solution, an UV-detector is required
between the first dimension column and the modulation valve.
2D-LC User Guide
separate as many constituents as possible or
focused on resolution and quantitation of a specific
constituent
a relatively large volume of the sample will be collected and
injected into the second column
to minimize peak broadening the sample should be focused
at the inlet of the second column
Methodology
As in Comprehensive 2D-LC is no direct need
dimension, other eluents than acetornitril or methanol are possible. This
implies the possibility to use unconventional organic solvents in the first
dimension.
Comment
True gradient elution in the second
dimension separation provides
better peak capacity than in
isocratic elution.
Gradient elution is the best
available mechanism for achieving
peak focusing.
1
for UV-detection in the first
380