On-Demand Cannabis Webinar: Chromatography Foundations for Cannabis Labs

In a recent SelectScience webinar we were joined by Scott Grossman, content development specialist for Restek, to discuss the concepts of high-performance liquid chromatography (HPLC) and gas chromatography (GC) for cannabis testing. Now available to watch on demand, this webinar is ideal for both those new to analytical testing, and the experienced user looking for a refresher.

With the rise in use of medicinal and recreational cannabis in the USA, quality control is becoming increasingly important to ensure consistent product quality and consumer safety. HPLC and GC are currently the primary techniques used in the development of safe and effective products. The concepts learned in this webinar will help develop a working knowledge of the separations methodologies in cannabis testing, and how to overcome challenges in workflows. You can catch the webinar on demand here, or read on to discover the Q&A highlights from the live event.

Question and Answer Session

Q: You said that a broad peak between narrow peaks wouldn’t happen because of normal chromatographic processes, but that we might see it. Why might it happen?

SG: It’s true that you won’t normally see a broad peak in between narrow peaks, but there are some reasons why you might see that. The first reason is that you have very closely co-alluded peaks where you have retention but not selectivity, so two co-alluded peaks might appear as one broader peak. This can occur frequently if, during a GC run, your analytical run conditions are not long enough to elute all the compounds in your sample. This is especially the case in temperature-controlled runs since by the time your run ends, your oven begins to cool down, and compounds still left in the column will be trapped until your next analysis. The longer they are trapped, the broader that band is going to get. When the next run occurs, and the oven begins to heat up again, that peak is going to elute at a point that is not its normal retention time. It’s going to appear as a much broader and quite possibly misshapen peak, among all of your beautifully formed chromatographic peaks.

Q: If you were to change one method or column variable, would you have to consider changing others as well or is it better to just change one thing and keep the others constant?

SG: Changing just one thing at a time and keeping everything else constant is a very scientific approach, but it can have some consequences when you are changing column parameters. Let me use gas chromatography as an example again, where something called phase ratio is relevant. When changing inner diameter and keeping film thickness the same, you are changing your phase ratio; the phase ratio is the relationship between inner diameter and film thickness for a wall coated open tubular column. This can have effects on your selectivity. So, if you are going to a narrower column, for efficiency gains, you may want to change your film thickness at same time to maintain the same phase ratio.

In a HPLC column, if you are going from a larger particle size to a smaller particle size there are many method translation considerations that may require you to scale your methods. If your ultimate goal is to get the same chromatography in less time you are going to need to scale more than just one of those column features that gives you better efficiency. By just changing inner diameter for example, we would have created more plates, but we wouldn’t have got the same chromatography results. You need to scale your change across many variables therefore.

Q: You gave the specific example of a tailing peak situation where some of your molecules are of arriving later than they should, but how would you describe a split peak?

SG: If you think of the chromatogram as this story of the journey of your molecules through the system, a tailing peak is a representation of some of those molecules being dragged out, and made a little late. If we are talking about a split peak, the first consideration is, “is it truly a split peak?”. We should make sure that we are not actually talking about two different compounds which have not been resolved, but if we are confident that it’s the same compound and that the peak’s shape is split, we would begin to ask questions.

A bimodal population might occur because we didn’t introduce our sample into the instrument as one distinct band, for example, when you’ve got imperfect injection due to delay. Another way that split peaks might be produced is through separation of the molecules. In gas chromatography, this scenario can occur where there is a mismatch between your solvent and stationary phase; if you have a very non-polar stationary phase and a very polar solvent, that very polar solvent won’t evenly wet the surface of the stationary phase and may actually bead up on the stationary phase, like water on a waxed car. That mismatch can cause the sample to collect in distinct populations which move through the chromatographic system as distinct bands.

Early eluters may never reform during both of these processes, and you may see split peaks, while for later eluting compounds, they may have the time for other chromatographic properties to take over and they’ll merge back together. If you see a split peak and you’re sure it’s one compound, the main question to ask is, “what would make that one population of molecules move through my chromatography system as two?”

Q: Which LC column from Restek’s catalogue will you recommend for analysis of cannabis oil?

SG: For the analysis of cannabis oil, the method that we showed uses Raptor Arc-18, which is a column that has been seeing a lot of use in our labs for the analysis of cannabinoids, and in the industry at large. I demonstrated one application that shows the Arc-18 in a small particle format, but there are options depending on the instrument that you have, or what your other considerations are. Our Raptor line of LC columns is Restek’s version of the superficially porous particle, so for those of you out there that have fully porous particles, superficially porous particles, and shell particles. Raptor is our superficially porous particle line, in which Arc-18 is the phase that comes in a variety of dimensions. Of course, we have apps on our website that show different phase dimensions that you might want to use.

Q: What is a good column for low concentration ppb level for cannabis in drinking water?

SG: A good column would be catalogue number 931421E from the Raptor Arc-18 catalogue, since you still need separation of isobars.

Q: Can a pre-column with a filter on a LC column be the reason for peaks tailing?

SG: Yes, this could be the cause if all peaks are tailing.

Find out more on this topic by watching the full webinar on demand, or visit our Separations community>>

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