How to deal with variable retention times?

a) Leak. Solution: Check system for loose fittings. Check pump for leaks, salt build-up, and unusual noises. Change pump seals if necessary. b) Change in mobile phase composition. Small changes can lead to large changes in retention times. Solution: Check make-up of mobile phase. If mobile phase is machine mixed, hand mix and supply from one reservoir. c) Air trapped in pump. Retention times increase and decrease at random times. Solution: Purge air from pump head or check valves. Change pump seals if necessary. Be sure mobile phase is degassed. d) Column temperature fluctuations. Solution: Use reliable column oven or insulate column. e) Column overloading. Retention time usually decreases as mass of solute injected on column exceeds
column capacity. Solution: Inject smaller volume or dilutions of the sample. f) Sample solvent incompatible with mobile phase. Solution: Adjust solvent. Whenever possible, inject samples in mobile phase. g) Column problem. Not a common cause of erratic retention. As a column ages, retention times gradually decrease. Solution: Substitute new column of same type to confirm column as cause. Discard old column if restoration procedures fail.


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Based in Berlin, KNAUER has been serving the sciences since 1962. We develop and manufacture scientific instruments of superior quality for liquid chromatography systems and components, including:

Analytical HPLC / UHPLC
Preparative HPLC
Fast protein liquid chromatography (FPLC)
Multi-Column Chromatography / SMB

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