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Practical HPLC Troubleshooting and Maintenance practical course
2 Day Course | HPLC Level 2

A logical approach to HPLC Troubleshooting and Maintenance is fully explored within this one-day course. Commonly encountered problems and best practice are reviewed for all major system components including mobile phase, pump, injection system, column and detector.

Examination of separation chemistry, interrogation of poor peak shape and retention problems are a fundamentally important part of the course, leading to increased awareness of troubleshooting principles.

Practical exercises are fundamental to this course and familiar equipment will be used to carry out a wide range of laboratory-based tasks.

  • We limit numbers to 6 per course so that each delegate gets the opportunity to ask questions and fully participate in practical exercises
  • When delivered on-site we can design the course material to suit your specific training needs
  • Customisable written assessments are available if required

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download course pdfAll components of the HPLC system and associated problems are systematically studied. The use of chromatograms as diagnostic tools is explored in depth, including baselines, peak shape and retention time inconsistencies, spurious peaks, etc.

Laboratory based sessions are included to make an all-encompassing two day course.

Who is this course for

This course is designed for those relatively new to HPLC, or for those users of HPLC systems who want to gain a deeper insight of their instruments. Ideally will follow-on from attendance to the fundamental HPLC training course.

Previous knowledge

Some previous knowledge of chromatography and basic experience are recommended. A good grounding in Chemistry is also beneficial.

What you will learn

  • Instrument problems - identifying and defining the issues
  • Common maintenance schedule operations
  • Investigative operations
  • Hands-on pump, auto sampler and detector maintenance procedures
  • Common calibration protocols
  • Putting it all together

Common Maintenance Schedule Operations

  • Pumps: pistons, mixers, check-valve replacement, filters, etc.
  • Autosamplers: injection valves, seals, metering devices, syringes etc.
  • Detectors: flow cell clean, lamp replacement

Common Calibration

  • Calibrating detector wavelength and response
  • Testing lamp intensity
  • Testing flow cell cleanliness
  • Calibrating column temperature

Investigative Operations

  • Log-keeping
  • Calibrating pump flow
  • Calibrating autosampler volume
  • Writing a maintenance process document

Chromatographic Troubleshooting

  • Baseline problems
  • Peak shape problems
  • Retention time drift
  • Peak area irreproducibility


  • Installation and conditioning
  • Column chemistry
  • Efficiency loss
  • pH operating range/bleed
  • Proper column management
  • Loss of sensitivity

Putting it All Together

  • Maintenance records and schedules

Training Calendar

Click on a title below to download a detailed course description or click a date and book your course.

Can't find a suitable training course? Call 01357 522 961 or email us.

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Experimental Determination of System Dwell

Gradient dwell volume is the total system volume in a gradient system between the point where the gradient is formed and the inlet of the column. In a high-pressure-mixing system, this includes the mixing chamber, connecting tubing, and injector. In a low pressure mixing system this ALSO includes the volumes of the pump head, pulse damper and other connective tubing. Typical dwell volumes range from 1 ml to 5 ml, but values can be as low as 0.5 ml or higher than 13 ml.

The dwell volume can be converted into a dwell time by multiplying by the eluent flow rate. The dwell volume has a significant impact when transferring methods between different HPLC systems. It is important to account for differences in dwell volume (time) between systems in order to accurately reproduce a separation. This is usually achieved by having an ‘isocratic hold’ section at the beginning of the gradient profile which can lengthened or shortened according to the differences in dwell time between the two HPLC systems.

Dwell volume can be easily measured using your UV detector to trace the gradient profile generated by your system – this is explained in more detail opposite.

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