Retention-time drift, temperature control, and broad peaks from a new column are the topics of this month's discussion.
As many of you know, a major part of my job is teaching liquid chromatography (LC) training courses around the world. Besides getting to see some pretty fascinating places, I get to meet lots of people and have a chance to help solve some LC problems that are troubling the attendees. One thing I have found is that chromatographers everywhere tend to have the same struggles with LC problems. Sometimes I feel a bit like the detective on Garrison Keillor's A Prairie Home Companion, whose radio dramas always end with the tag line, ". . . one man is trying to find answers to life's persistent questions . . . Guy Noir, Private Eye." In this month's installment I'd like to share a few of the questions that came up in recent classes.
Retention-Time DriftOne attendee mentioned a problem related to a small, but noticeable increase in retention time that continued to grow over several weeks. In an earlier installment (1) we looked at tricks to help determine the cause of retention drift. One of the ways to differentiate between hardware (flow rate) problems and chemical (column, mobile phase, or temperature) problems was to see if the disturbance at the column dead time changed in the same proportion as the retention change. If it did, a hardware problem was indicated; otherwise it was a chemical problem. However, the current problem was observed on an LC system with a mass spectrometry (MS) detector. In contrast to ultraviolet (UV) detectors, where a solvent-front disturbance is almost always seen, unless the MS system is set to specifically look for ions at the dead time, the baseline is flat in this region. As a result, the dead-time diagnostic didn't help.
The first step taken was the easy one — replace the column. When this approach didn't help, a new batch of mobile phase was made, but it didn't solve the problem either. Next came cleaning the pump check valves and finally replacement of the pump seals. None of these changes arrested the drift.
Then one day, the user was inspecting the autosampler and noticed a reflection from a drop of liquid on the injection valve. This led him to service the valve. He discovered that there was a scratch between two of the passages in the valve rotor, creating a situation called cross-port leakage. A tiny amount of mobile phase was leaking out the waste line from the injector, effectively lowering the flow rate to the column and thus increasing the retention times. As the leak became worse over time, the loss of flow increased and retention times drifted to even longer times. Replacement of the rotor seal corrected the problem. Because the leak was small, likely only in the tens of microliters per minute, and a piece of opaque tubing was used to route the injector waste to the waste container, the leaking solvent was not noticed. Replacement of the valve rotor corrected the problem.