Retention time shift and co-elution after first injection on ACQUITY UPLC H-Class system - WKB4104

SYMPTOMS

• The results of the first injection are satisfactory
• All subsequent injections have shifted retention times and co-elution of peaks

ENVIRONMENT

• ACQUITY UPLC H-Class 
• Empower 3

CAUSE

• The method in use calls for a gradient.
• The instrument method does not provide enough time for the system to equilibrate before the next injection is made.

FIX or WORKAROUND

Extend the run time to allow for equilibration.

ADDITIONAL INFORMATION

An appropriate re-equilibration time, or the time between changing back to initial conditions at the end of one injection and the time of the next injection, should be long enough to allow enough mobile phase through the system and column to adequately ensure both proper pH and organic strength.  A good starting number would be about (3x system volume + 5x column volume) in terms of mobile phase pumped. This can be longer or shorter depending on specific method conditions.