Cannabis Potency by GC-FID with Carrier Gas Comparison (Helium, Hydrogen, Nitrogen)
Due to the legalization of cannabis in multiple countries for medicinal or recreational use, there has been an increased demand for quality control of hemp and marijuana products. This is to ensure the product labels accurately reflect the characteristics and quality of the flood of new products entering the market.
With governing bodies such as the FDA looking to review their regulatory guidelines on Cannabinoid products it is becoming increasingly crucial to be able to accurately quantify the active components in Cannabis products. 
Potency analysis identifies different cannabinoids and measures their concentration as an indication of the strength of the product. There are over 500 chemical compounds in Cannabis, in this application we look at the most commonly tested cannabinoids; these are listed in Table 1.
Cannabis potency can be determined using a number of analytical techniques. In this application note we will describe a method using Gas Chromatography with a Flame Ionisation Detector (GC-FID). SCION also has application notes discussing other techniques such as HPLC (AN145) and GC-MS (AN142), these can be found on our website.
Helium has been the standard carrier gas for GC applications however, due to recent global shortages many companies are keen to find alternatives. This application note will give a comparison between different carrier gases showing the variation in results between the two alternatives to Helium; Hydrogen and Nitrogen.
This application can be performed on either the SCION Instruments 8300 GC or 8500 GC platforms equipped with an 8400 Pro Autosampler, S/SL injector, and FID detector. These are shown below in Figure 1.
Figure 1 SCION Instruments 8300 GC and 8500 GC platforms equipped with 8400 PRO Autosampler
Marijuana usually contains high levels of Δ9-THC and a lower level of CBD, with hemp it is the opposite. The primary psychoactive component is Δ9-THC, while CBD is the primary therapeutic component.
Consumer hemp generally comes in the form of hemp oil which is used for medical purposes while marijuana is often smoked. Both sample types require different sample preparation before analysis due to the difference in sample matrixes.
The focus of the calibration is on six components: CBC, CBD, Δ8-THC, Δ9-THC, CBG and CBN. Several Cannabinoid components, CBDV, CBGA, THC-A and CBDA cannot be detected without extra sample preparation (derivatization) due to decarboxylation caused by the heat of the injector, this reduces the acid components to their respective parent cannabinoid (CBDV to CBD, CBGA to CBG, THC-A to THC and CBDA to CBD). Results will therefore be presented as total cannabinoid content for these components.
In SCION application note AN145 using HPLC these compounds can be detected without the need for derivatization, greatly reducing sample preparation time if these components are required to be quantified separately.
Two different product types, cannabis oil and medical marijuana were chosen for potency analysis in this application note.
The cannabis oil was diluted with methanol, for the medical marijuana more extensive preparation was necessary. In order to extract the cannabinoids from the medical marijuana, it was dried for 2 hours, then ground to a powder. 30 mL of methanol was added and the sample was placed in an ultrasonic bath for 30 minutes. The sample was blown dry with nitrogen and then reconstituted in methanol.
When designing the instrument parameters for the GC it is important that the injector temperature is high enough to prevent carry over. Carry over can occur from previous runs or extra material that is being introduced into the system.
Another solution for this problem could be a washing method between samples.
Using the method parameters shown in Table 2 we achieved analysis of the most commonly tested cannabinoids in less than 8 minutes without carry over.
Potency analysis was repeated using this method with Helium, Hydrogen and Nitrogen as carrier gas.
Results and Discussion
The method stated in Table 2 gave excellent specificity when using each carrier gas. This is demonstrated in figures 2 – 4 which clearly show good resolution between all the components as well as good peak shape (all peak tailing factors of each component between 0.5 – 1.5 which is the standard requirement of many analytical methods).
Table 3 gives the order of elution, which was consistent across all tested carrier gases. There was some variation noted in retention times, but this had no effect on the overall results.
Figure 2 Chromatogram of a potency standard using Helium carrier gas
Figure 3 Chromatogram of a potency standard using Nitrogen carrier gas
Figure 4 Chromatogram of a potency standard using Hydrogen carrier
The system precision was determined by injecting a standard containing ca 50 µg/mL of each component 10 times. This was performed for each carrier gas, a comparison of the results can be found below in Table 4.
The results for the system precision using each carrier gas are very similar to each other and all demonstrate RSD of <2%. Which is generally what is considered to be acceptable for method validation.
Linearity solutions were prepared containing all components at 2 – 100 µg/mL. These solutions were ran using each carrier gas and the potency calibration curves can be seen opposite in figures 5 – 7. All components demonstrated a correlation coefficient (R2) of >0.9990 which is an excellent result, with many regulations requiring an R2 value of only ≥0.98.
Figure 5 Calibration curves of the potency standards when using Helium as carrier gas
Figure 6 Calibration curves of the potency standards when using Nitrogen as carrier gas
Figure 7 Calibration curves of the potency standards when using Hydrogen as carrier gas
A quality control (QC) sample was prepared at ca 20 µg/mL for both Helium and Hydrogen and at ca 30 µg/mL for Nitrogen. The respective QC samples were injected 10 times for each carrier gas and the average % recovery calculated. The results can be seen below in Table 5.
The results of the QC samples show similar recovery using each carrier gas, with results well within the 70 – 130% required by many regulations.
The theoretical limit of detection (LOD) and limit of quantification (LOQ) was calculated for each component with each carrier gas type used. The LOD was based on 3x signal to noise ratio and LOQ 10x. The results can be seen in Table 6 below.
The LOD/LOQ for the components measured are very similar when using each carrier gas, showing that for cannabis potency by GC-FID both hydrogen and nitrogen are suitable for use, offering comparable sensitivity to Helium for this analysis.
Due to the timescales involved in the running of the 3 tests, one for each carrier gas, the cannabis oil and medical marijuana samples were different for each run. Therefore a direct comparison between the results cannot be drawn.
However, there were some interesting anomalies noted between the samples, especially the cannabis oil which stated a CBD content of 100mg for each sample, and no mention of any other cannabinoids. The medical marijuana samples came with no certificate of analysis and so the label claim cannot be compared to. The results can be seen in Table 7 below.
The results, although not directly comparable for the different carrier gases as they are from different samples, do however show the importance of quality control within the cannabis market. There are very large discrepancies between the label claim for example, of the cannabis oil samples which all stated 100mg and the actual results seen here.
As regulations inevitably tighten in the future it will become even more important to be able to accurately quantify the different cannabinoid components of these products
Below in figures 8 – 13 are example chromatograms from the cannabis oil and medical marijuana samples ran with each carrier gas during this application.
Figure 8 Chromatogram of CBD oil sample ran using He as carrier gas
Figure 9 Chromatogram of CBD oil sample ran using N2 as carrier gas
Figure 10 Chromatogram of CBD oil sample ran using H2 as carrier
Figure 11 Chromatogram of marijuana sample ran using He as carrier gas
Figure 12 Chromatogram of marijuana sample ran using N2 as carrier gas
Figure 13 Chromatogram of marijuana sample ran using H2 as carrier gas
The SCION 8300 GC and 8500 GC platforms equipped with a split/spitless injector, SCION Instruments column and FID is capable of analysing potency from cannabis products in a qualitative and quantitative way.
Due to the high injector temperature the acidic forms of certain cannabinoids (CBD, CBG and 9-THC) degrade into their non acidic forms during the analysis. This means that the acidic and non acidic forms of those compounds cannot be discriminated in this method without further sample preparation, and are therefore reported as a summation of both forms for each of these cannabinoids. When additional discrimination of these forms is needed the recommendation is to use the SCION LC6000 method for Cannabis Potency by HPLC as described in AN145.
It is worth noting that many national legislations allow for reporting of these components as a summation of the acidic and non acidic forms. It is advised to check local requirements before performing potency analysis.
The results from this application note have shown that both Hydrogen and Nitrogen are both suitable alternatives to Helium for use with GC-FID for this particular application.
Although it was necessary to adjust column flow parameters between each carrier gas, which lead to changes in the retention times of the components, this had no negative effect on the resolution of the method with all 3 carrier gases showing the required specificity for this analysis.
The analytical method validations (system precision, linearity, QC sample, LOQ and LOD) gave comparable results for all 3 carrier gases. Showing that cannabinoid potency by GC-FID can be replicated with Hydrogen and Nitrogen with no detriment to the analytical results for this particular application.
With the current global shortages of Helium and the need to move to greener technologies, such as the use of a hydrogen or nitrogen generator, it is important to be able to replicate methods using different carrier gases so that these alternatives can be considered.
The results from the different cannabis oil and medical marijuana samples shows the necessity of quality control within the cannabis market, as it is possible for products to vary significantly from their label claims. This will only become more important as regulations across the world inevitably become more stringent.
For ordering info on the SCION 8500 GC, which offers greater functionality with the option of up to 4 detectors (including MS), please contact your local SCION sales representative.
- FDA Statement, Janet Woodcock, M.D. January 26, 2023 https://www.fda.gov/news-events/press-announcements/fda-concludes-existing-regulatory-frameworks-foods-and-supplements-are-not-appropriate-cannabidiol
- FDA Consumer Update, May 03, 2020 https://www.fda.gov/consumers/consumer-updates/what-you-need-know-and-what-were-working-find-out-about-products-containing-cannabis-or-cannabis
- Innovation News Network, Helium Shortage 4.0, April 04, 2023 https://www.innovationnewsnetwork.com/helium-shortage-4-0-what-caused-it-and-when-will-it-end/29255/
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