2.8.1 Columns calculator
The columns calculator enables you to scale a method by calculating operating parameters that
give equivalent chromatographic performance. It can quickly define methods to test further in the
laboratory.
See also: The ACQUITY console online Help for additional details.
2.8.2
Transferring from HPLC to UPLC
Follow these guidelines to preserve a chromatographic profile when transferring from one system
to another:
• Consider the difference in dwell volume between the two systems.
• Pre-injector volume, specified in the instrument method, enables the gradient to start before
the injection is triggered. Use a pre-injection volume to maintain a constant dwell volume to
column volume ratio on both systems.
• For a target system, with a smaller volume, use an isocratic hold to account for the dwell
volume differences.
• Active preheating is the default configuration for the ACQUITY UPLC H-Class system. An
optional, passive, column stabilizer is available for existing chromatographic methods not
suitable for active preheating.
• Select the column with the most similar selectivity using the Interactive Waters Reversed
Phase Column Selectivity Chart, which you can download from the Waters Web site or by
double-clicking the shortcut icon on the desktop. The Waters columns are highlighted (larger
white dots).
• For the initial evaluation, keep conditions as consistent as possible. You can optimize the
separation later.
See also: Transferring methods in the ACQUITY Console online Help.
2.8.3
Transferring from UPLC to HPLC
Follow these guidelines to preserve the integrity of a chromatographic separation.
• Match the ratio of column length to particle size (L/dp), the measure of resolving power.
• Maintain the number of gradient column volumes for each step of the gradient, to preserve its
separation power.
• Calculate appropriate gradient hold volumes at initial gradient conditions when going from a
larger system volume to a smaller one.
August 8, 2016, 715005049 Rev. C
Page 44
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