Enhancing Methods for the Chemical Characterization of Cannabis Products

The medical and recreational use of Cannabis sativa and Cannabis indica by humans has a history dating back almost 30,000 years. Thought to originate in southern Siberia, Russia, the plant’s usefulness in medicine and textiles led it to be cultivated and spread by the Egyptian, Greek, and Roman Empires, and despite some modern day attempts to limit the plant’s usage and production in some countries, it has become the most widely used illicit drug worldwide.

Cannabis plants contain over 400 different chemical compounds, each with their own distinctive properties and structures. The largest class of these compounds are the cannabinoids, which are responsible for much of the plant’s bioactivity. The cannabinoids include: Cannabigerol, an anti-bacterial agent; Cannabidiol, the cannabinoid with the highest number of confirmed medical uses; and Δ9-Tetrahydrocannabinol (Δ9-THC), the main psychoactive component in cannabis. In addition to the cannabinoids, terpenes are also typically present, a class of compound responsible for the aromatic features of the plant. As well as aromatic properties, many terpenes may also have pharmacological properties, such as antiseptic and anti-inflammatory effects. By studying the chemical compounds present in a sample it is possible to determine the geographic origin of that specific plant, as well as evaluate the potential medical properties of the plant based on its biologically active components.

Factors affecting characterization methods

The geographical spread of cannabis and the way it is grown creates variations in the levels of pesticide use, heavy metal contamination, and the presence of microbial life on the plant. In addition to this, strains of cannabis from different origins often have different chemical compositions, properties, and derivatives. Methods aiming to characterize cannabis and its natural products must be sensitive enough to account for these factors and identify chemical differences down to the level of parts per million (PPM) and beyond.

The “gold standard” in chemical characterization

Gas chromatography in conjunction with a flame ionization detector (GC-FID) or a mass spectrometer (GC-MS) are two of the main techniques used for cannabis identification and are even recommended by the United Nations.  

GC-FID works by detecting ions that are created when the organic compounds from the gas chromatograph (GC) combust and using this to estimate the concentration of each organic component detected by the GC. Romano and Hazekamp, from the University of Siena and Leiden University, used GC-FID analysis for their work comparing different ways of preparing medical cannabis oil, where they found that the non-toxic and cheap olive oil was the safest approach. 

GC-MS combines the use of a gas chromatograph and a mass spectrometer and is commonly seen as the “gold standard” for substance identification. A study comparing hemp and cannabis seeds was able to use GC-MS to sensitively detect higher Δ9-THC levels in the cannabis seeds. The same study also proved that much of the cannabinoid detected was found on the surface of the seed, and not within it.

Despite its wide usage, GC analysis is far from a perfect analytical solution for cannabis analysis. Depending on the type of sample being analyzed, there may have to be multiple complicated sample preparation steps to create a derivative that will separate noticeably through the GC to give clear results. This is the only way to accurately distinguish between neutral and acidic cannabinoid forms.

Better methods for analysis and characterization

Liquid chromatography (LC) avoids the need for these sample preparation steps by passing the sample through an unheated system with a nonpolar column. For cannabinoids, the mobile phase in the column is typically an equal mixture of formic acid in water and formic acid in methanol. Similarly, to GC-MS, there is also a method of analysis that combines LC and MS. However, unlike GC-MS, it is possible to combine LC-MS with a hybrid quadrupole time-of-flight analyzer (QTOF) to reveal an extra level of information. LC-QTOF-MS introduces a way to study the mass-to-charge ratio of the ions and ionic fragments in the sample, in addition to the chromatography and mass spectrometry information already provided. The greater resolving power of the TOF component allows the LC-QTOF-MS method to be both extremely sensitive and accurate, which could be vital for detecting possible risks such as heavy metal contamination or distinguishing chemicals with a similar molecular weight and structure. Unfortunately, this also drives up the operating cost of the analysis. 

Cheaper options for liquid chromatography analysis than LC-MS are in regular use. High-performance liquid chromatography used in conjunction with an ultraviolet detector (HPLC-UV) is a much less costly approach. HPLC-UV identifies the composition of a sample primarily using retention time data from liquid chromatography, then by using the absorption profiles for the detected substances. This is a far simpler process than techniques like LC-QTOF-MS, though due to there being few cannabinoid absorption profiles available in the literature for comparison, it can sometimes be difficult to make qualitative assignments. This limitation means that HPLC-UV is often used in conjunction with another method, such as GC-FID, to perform a full analysis.

As well as cost and desired information, there are other reasons that one type of characterization may be preferred over another. Different cannabis products will have different properties, and these could influence which characterization methods are most suitable. For example, terpenes are far more volatile than cannabinoids and also have a more isomeric nature. To account for this, terpenes are usually characterized using a chiral GC column with a slow temperature ramp to maximize separation between different enantiomers of the terpenes and produce a more accurate analysis. Terpenes are also commonly analyzed using headspace analysis (HS), which exploits the volatility of terpenes by analyzing the material in the gas phase. This has the added benefit of minimizing the amount of sample preparation required, but it does require samples to be concentrated in the ppm to ppt range, so it isn’t suitable for all samples. 

The outlook for analysis

With a wealth of cannabis products and derivatives already proven to exhibit useful medicinal properties, the characterization of cannabis products will remain a topic of interest for years to come. One shared goal is the creation of a universal detection and characterization method that is cheap to use and sensitive enough to distinguish between many different types of compounds at differing concentrations, without the need for overly-laborious sample preparation. With many countries choosing to reform their views on cannabis use and research in light of cannabis’ medical benefits, scientists, medical professionals, and consumers alike all stand to benefit from fully characterizing the chemical composition of cannabis.