Scientists studying the chemistry of Cannabis leaves have uncovered something that had never been documented in the plant before: evidence of a rare group of natural compounds known as flavoalkaloids.
Analytical chemists at Stellenbosch University (SU) made the discovery while examining the complex mixture of chemicals found in commercially grown Cannabis from South Africa. Their findings suggest that the leaves, which can be treated as low value plant material or waste, may contain a much richer collection of potentially useful compounds than previously recognized.
Rare Compounds Hidden in Cannabis Leaves
Flavoalkaloids belong to a broader family of plant chemicals known as phenolic compounds. Phenolics are produced naturally by plants and include substances involved in pigmentation, defense, and protection from environmental stress.
One major subgroup, flavonoids, is especially well known. These compounds have drawn considerable interest from pharmaceutical researchers because many are associated with antioxidant, anti-inflammatory, and anti-carcinogenic properties.
Flavoalkaloids, however, are much less common in nature.
In the new analysis, the researchers detected 79 phenolic compounds across three strains of Cannabis grown commercially in South Africa. Of those, 25 had never previously been reported in Cannabis.
Sixteen of the compounds were tentatively identified as flavoalkaloids, providing the first evidence that this rare class of chemicals occurs in Cannabis leaves. Even more strikingly, most of the flavoalkaloids appeared primarily in just one of the three strains.
The findings were published in the Journal of Chromatography A.
Cannabis Contains Hundreds of Different Metabolites
Dr. Magriet Muller, an analytical chemist in the LC-MS laboratory of the Central Analytical Facility (CAF) at Stellenbosch University and first author on the paper, says studying plant phenolics can be especially difficult.
These compounds are often present at very low concentrations, and their chemical structures can vary enormously. That makes it challenging to separate, identify, and distinguish one compound from another within an already complex plant sample.
“Most plants contain highly complex mixtures of phenolic compounds, and while flavonoids occur widely in the plant kingdom, the flavoalkaloids are very rare in nature,” she explains.
The chemical complexity of Cannabis made the results particularly surprising. The plant is already known to contain hundreds of metabolites, a broad term for small molecules produced through biological and chemical processes inside living organisms.
“We know that Cannabis is extremely complex – it contains more than 750 metabolites – but we did not expect such high variation in phenolic profiles between only three strains, nor to detect so many compounds for the first time in the species. Especially the first evidence of flavoalkaloids in Cannabis was very exciting.”
Advanced Chemistry Reveals What Earlier Tests Could Miss
The discovery grew out of analytical techniques Muller developed during her postgraduate studies in SU’s Department of Chemistry and Polymer Science.
Her methods combine comprehensive two-dimensional liquid chromatography with high-resolution mass spectrometry.
Liquid chromatography separates the many different chemicals in a mixture so that researchers can study them individually. In two-dimensional liquid chromatography, compounds are separated in two different ways, giving scientists much greater resolving power when working with samples containing many closely related chemicals.
High-resolution mass spectrometry then helps identify those compounds by measuring their molecular masses with extremely high precision.
Together, the techniques allowed the researchers to examine the phenolic chemistry of Cannabis in far greater detail.
“We were looking for a new application for the methods that I developed, after successfully testing them on rooibos tea, grapes and wine. I then decided to apply the methods to Cannabis because I knew it was a complex sample, and that Cannabis phenolics have not been well characterized,” she explains.
Separating Rare Compounds From Abundant Ones
According to Prof. André de Villiers, Muller’s study leader and main author on the paper, the separation achieved using the technique was key to spotting the rare compounds.
He said he was blown away by the chromatographic results Muller obtained.
“The excellent performance of two-dimensional liquid chromatography allowed separation of the flavoalkaloids from the much more abundant flavonoids, which is why we were able to detect these rare compounds for the first time in Cannabis.”
De Villiers leads the analytical chemistry research group in SU’s Department of Chemistry and Polymer Science.
The result highlights an important challenge in plant chemistry. Rare compounds can easily be obscured by other chemicals that occur in much larger quantities. Better separation methods can therefore reveal substances that were present all along but were difficult to distinguish using less powerful techniques.
Looking Beyond THC and Other Cannabinoids
Much of the scientific attention surrounding Cannabis has historically focused on cannabinoids, the group of compounds that includes substances responsible for many of the plant’s well known pharmacological and mood effects.
De Villiers argues that the new findings show how much remains to be learned about the plant’s non-cannabinoid chemistry.
The leaves in particular may deserve closer scrutiny. Plant material that is currently regarded as waste could contain phenolic compounds with properties that make them interesting for future biomedical research.
“Our analysis again highlights the medicinal potential of Cannabis plant material, currently regarded as waste. Cannabis exhibits a rich and unique non-cannabinoid phenolic profile, which could be relevant from a biomedical research perspective,” he concludes.
