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- Posts: 47
- Joined: Fri Sep 07, 2007 12:39 am
- Location: Adelaide, Australia
We are currently seeing some strange things happening on our Waters Alliance H-Class systems. We are new to the use of these systems and not very familiar with all the problems we may occur although we are finding out your margins for small “errors” are reduced. Our retention time is shifting without warning. We have been using this method for nearly 12 months and have not encountered this problem. Occasionally we had seen the peak shift out but only at certain times i.e.
The system had been left for a number of days
Mobile phase was aging
If mobile phase is prepared as a mixture of 80:20, pH 3.0 phosphate: Acetonitrile opposed to being used as a gradient with inline mixing
Following a good system purge the peak always shifts back to the typical retention time of 0.6 minute
Example:
A number of analyses performed on the system thought out the morning, retention time of the peak of interest is 0.6 minute, total run time of 1.0 minute. 3 hours later additional analysis performed and no peaks are observed. Upon investigating found that the peak had shifted from 0.6 minute to 1.1 minute. Nothing had changed and the system had not been touched between analyses.
Method details:
Gradient analysis – pH 3.0 phosphate buffer: Acetonitrile, mixing inline 80:20 at 0.6 mL/min
Wavelength at 280 nm, injection of 1.5 micro litres
Column is Acquity UPLC® BEH C18, 1.7 m, 2.1 x 50 mm at ambient temperature
Mobile phases are degassed and filtered through 0.2 filters, all reagents are of appropriate quality, samples are prepared in compatible reagents and good lab practice is employed.
Can anyone give any advice as to what might be causing this sudden shift? What can we do to eliminate this occurring or minimize the odds of this happening?
Any suggestions are appreciated and I thank you all for you time in advance. If you need any additional information please ask and I will provide.