Chromatography Troubleshooting Tips - - Chromatography Online
Chromatography Troubleshooting Tips

  • Establish a benchmarking method (and representative data) BEFORE the system develops a fault. This can be either a method you regularly use or a method specifically designed as a system checkout, but it must be 100% reliable. Obtain a comprehensive set of data (back pressure, repeatability, sensitivity, and noise) from this method on the system when it is working properly.

  • Don’t allow problems to start. Be methodical and meticulous in the way you set up the HPLC instrument and where possible perform preventative maintenance routines..

  • Clearly define the problem:
    Compare a good chromatogram with the problem one.
    • Retention times: Do they change for all peaks (including T0) or only some?
    • Peak height: Are they smaller, large, show poor repeatabilit? Does this apply to all peaks or only some?
    • Elution order: Does this remain identical or does peak order change?
    • T0 peak: If it is the same (e.g. k’ changes) it is unlikely to be flowrate related, (as this affects all peaks including RI). If it is just retained peaks affected, could be temperature or gradient related?
    • Are there any extra peaks?
    • Has peak shape/width changed?

  • Don’t forget the obvious or assume anything. Check EVERYTHING.

  • Check the easy things first.

  • Check one thing at a time.

  • Keep a written note of every single thing you do as you will either quickly forget or doubt yourself.

  • Use your benchmarking method to establish if problem is due to either the instrument or the method (e.g. the sample prep, method conditions, column or analyst). For example, if your benchmarking method gives acceptable results, the problem is not related to an instrument fault..

  • Only take something to bits as a last resort or if you are reasonably sure this is causing the fault. 

  • Don't forget to use the CHROMacademy troubleshooters :-)
  • The complete article is available to CHROMacademy Lite and Premier members here >>

    CHROMacademy Lite Membership is FREE and it only takes two minutes to register.

    With a Lite Membership you are given access to:
    ∙ This month's webcast & tutorial
    ∙ Selected eLearning modules
    ∙ Featured CHROMacademy Content
    ∙ The CHROMacademy forum

    Test drive CHROMacademy and Check out more great content available FREE to our Lite members »


    blog comments powered by Disqus
    LCGC E-mail Newsletters
    Global E-newsletters subscribe here:



    Column Watch: Ron Majors, established authority on new column technologies, keeps readers up-to-date with new sample preparation trends in all branches of chromatography and reviews developments. LATEST: When Bad Things Happen to Good Food: Applications of HPLC to Detect Food Adulteration

    Perspectives in Modern HPLC: Michael W. Dong is a senior scientist in Small Molecule Drug Discovery at Genentech in South San Francisco, California. He is responsible for new technologies, automation, and supporting late-stage research projects in small molecule analytical chemistry and QC of small molecule pharmaceutical sciences. LATEST: HPLC for Characterization and Quality Control of Therapeutic Monoclonal Antibodies

    MS — The Practical Art: Kate Yu brings her expertise in the field of mass spectrometry and hyphenated techniques to the pages of LCGC. In this column she examines the mass spectrometric side of coupled liquid and gas-phase systems. Troubleshooting-style articles provide readers with invaluable advice for getting the most from their mass spectrometers. LATEST: Radical Mass Spectrometry as a New Frontier for Bioanalysis

    LC Troubleshooting: LC Troubleshooting sets about making HPLC methods easier to master. By covering the basics of liquid chromatography separations and instrumentation, John Dolan is able to highlight common problems and provide remedies for them. LATEST: How Much Can I Inject? Part I: Injecting in Mobile Phase

    More LCGC Columnists>>

    LCGC North America Editorial Advisory Board>>

    LCGC Europe Editorial Advisory Board>>

    LCGC Editorial Team Contacts>>

    Click here