What do you do when the peak shape changes?
A change in peak shape is one of the most common observations of problems with a liquid chromatography (LC) method. Because of this, most system suitability tests include a measure of peak shape, so a quantitative value of peak shape can be tracked over time. Poor peak shape can compromise the results of an analysis by degrading resolution between closely eluted peaks and reducing precision and accuracy of measuring peak area, especially for small peaks. A change in peak shape is one of the first signs that the column is failing, but there are other causes of peak tailing, as well. This month we look at several aspects of peak tailing as we continue our "Troubleshooting Basics" series of column installments (1–3).
Measuring Peak Tailing
Most LC peaks tail or front a bit, so column manufacturers typically set their column-release specifications at 0.9 < TF < 1.2 as normal performance. As can be seen in Figure 2, when tailing increases, several practical problems can arise. The peaks are harder to integrate because the transition from the baseline to the peak or peak to baseline is much more gradual, and on noisy or sloping baselines the peak limits are difficult to determine. Generally, the peak area stays constant, so increased peak tailing translates into shorter peaks, and peak height is the limiting factor in determining detection limits, so method limits can suffer with tailing peaks. Tailing peaks also take a larger time window to be eluted, so to achieve baseline resolution between peaks, the run time must be longer. And tailing peaks are aesthetically less pleasing. You can see that all these factors favor symmetric peaks. From a practical standpoint, peak tailing is difficult to eliminate, however, for many applications peaks with TF ≤ 1.5 are acceptable. When TF ≥ 2, usually corrective action should be taken to identify and eliminate the source of tailing.
When peak tailing occurs, it usually shows up for one or just a few peaks in the chromatogram, but sometimes all the peaks in the run tail. The appearance of peak fronting is much less common. Most often, these three behaviors are caused by three different sources. We will look at each of the three problems next.