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Should I run a gradient from 5-95% organic solvent or from 0-100%?

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When running a gradient, it is important to remember that the total time for each run includes both the run time, and the necessary re-equilibration before the next run.

When running from 0-100% organic solvent the re-equilibration step is far longer than for 5-95% such that the majority of users sacrifice the small restriction in terms of method flexibility for a much larger gain in productivity.

There is speculation on the mechanistic reason behind the lengthy equilibration time required for methods utilizing 0% aqueous and 100% organic solvent in the gradient.

Phase wetting/dewetting and phase collapse have been proposed causes. Either way, in practice, variations in retention can be observed with alkyl phases using 100% aqueous conditions.

If necessary for retention of certain highly polar compounds, SiliCycle offers columns designed to be stable under such conditions, such as our SiliaChrom dt HPLC columns.