HPLC Doctors and Nurses

All HPLC practitioners will be aware of the need to overcome issues in both method development and routine analysis. Even the most sophisticated equipment can present unexpected chromatographic behavior. And though such issues certainly plague modern instruments less frequently, sooner or later, a problem will inevitably occur.

There are some preventative measures that can be taken to increase the chance of seamless operation and performance of HPLC instrumentation – the most common of these being:

• filtering of the mobile phase solvents,
• filtering of the samples prior to injection,
• using the right buffers and following instructions to remove or avoid precipitation of salts in the system due to organic solvent,
• preparing fresh buffer or aqueous solutions instead of storing in the fridge,
• proper washing of the column after use.

But issues occur – and recur – regardless of these measures, and chromatographic abnormalities can be difficult to spot. It is easier during analysis of standards than of unknown samples for sure, though, and injection of control samples can help us to spot aberrant outcomes.

Record keeping is a requirement in any analytical procedure, and it should be considered absolutely essential in HPLC. (It was actually one of the first lessons I learned in practicing HPLC, and it is one of the most important lessons I teach my students as they begin using the technique…). We should know how the system works when functioning properly; only then can we recognize any irregular signs and symptoms – allowing us to subsequently resolve the problem. Recording pressure and keeping typical chromatograms for comparison enable the recognition of a non-proper function.

Knowing how to confront the problems and solve them could be considered a prerequisite for efficient HPLC operators. HPLC is incredibly useful, but also highly complex. Analytical chemists must know the fundamental theory behind the simple act of sample injection. We are “analysts,” not “analyzers,” and – as practicing scientists – we must all have a firm grasp of this knowledge.

It is not easy to have the right answers to all problems in chromatographic analysis. HPLC troubleshooting manuals outline a vast number of potential irregular functions, and also many corrective actions – but not all lead to the right solution. And that’s not to mention the fact that, although we all know the rule that we have to change one thing at a time, when we are in hurry, we sometimes ignore it and change many more...

But what happens when more “invasive therapy” is required? Are all HPLC practitioners able to proceed and fix the most common problems? Of course, we can call for assistance and technical support, but this is often time-consuming and associated with financial cost – nevertheless, we often find ourselves seeking an expert technician. But shouldn’t chromatographers themselves be the expert technicians? Being “doctors” to our own HPLC systems – healing its symptoms when sick – could save us both time and money, allowing us to inject both of these resources back into our research, perhaps learning valuable lessons in the process.

I suggest that analytical scientists – especially novices – using HPLC instrumentation should attend hands-on workshops to learn how to confront the problems arising in routine operation. And, at the very least, surely it would be better to invest time in actually reading the HPLC system’s instruction manual rather than blindly paying an external technician to fix the problem.