Principles and Practical Aspects of Preparative Liquid Chromatography

[Norm.]
7.584
3000
2000
2.791
5.720
1000
0
1 2
3 4 5 6 7 8
Time [min]
Figure 3.20 High-volume injection with strong eluents using the alternative injection mode.
A broad solvent peak is obtained unless the DMSO has been washed out of the column.
The two compounds at 5.72 and 7.58 minutes have been retained on-column and eluted as
baseline separated peaks. No sample loss has been observed.
Sample: 50 mg acetamidophen and 50 mg caffeine in 5000 µL DMSO
Column: Agilent ZORBAX SB C18, 21.1 x 150 mm, 5 µm
Applied gradient profile for sandwich injection mode – a longer isocratic holding step
is needed to transfer the sample to the column prior to ramping up the gradient:
Time Flow %A %B
0 37 93 7
4.0 37 93 7
10.0 37 78 22
10.1 37 2 98
13.0 37 2 98
13.1 37 93 7
16.0 37 93 7
3.2.3.3 Injecting large
sample volumes
using an injection
pump
For separation of highly diluted samples in aqueous solvents, large
injection volumes that exceed the typical loop sizes of autosamplers
are often required. In these situations an injection pump can be used
to load the sample onto the column. Injection pumps can also be used
for applications such as enantiomer separation. Here, the same sample
solution must be loaded periodically and an injection pump can achieve
this much faster than an autosampler. After sample transfer to the column,
thoroughly purge the injection pump, the sampling and injection valves,
and all connection capillaries to avoid sample carryover. Figure 3.21
shows a typical system configuration for sample introduction using an
injection pump.
25