What are cannabinoids?

The term Cannabinoids originally referred to a class of oxygen containing aromatic compounds which contained 21 carbons that were produced by Cannabis sativa. Now cannabinoids have a broader definition referred to as phytocannabinoids. This includes the original compounds as well as anything that is similar in structure (including synthetic cannabinoids created in labs). The structures of the common cannabinoids are similar to each other: they have a central aromatic ring containing two oxygens ortho to each other (substituted or not), a pentyl group, and a final substituent between the oxygens that varies by species. The only difference in the analogs (THCV, CBDV) is that the pentyl group is replaced with a propyl group, which varies in the way it is made in nature.

How are cannabinoids made in nature?

The method by which molecules are made in plants is called biosynthesis. Cannabinoids begin as cannabigerolic acid, CBGA, which is formed using a prenyltransferase enzyme[1]. An enzyme is nature’s way of accelerating a reaction, in this case the combination of olivetolic acid and geranyl diphosphate (Figure 1). Using prenyltransferase, CBGA undergoes a number of changes; Δ9-THCA synthase converts CBGA to Δ9-THCA, CBDA synthase converts CBGA to CBDA, heat or light causes a decarboxylation of CBGA which gives CBG (Figure 1). There are other transformations (such as the formation of CBCA) which will not be discussed here.

Figure 2. Biosynthesis of cannabinoids CBGA, Δ9-THCA, and CBDA starting from olivetolic acid and geranyl diphosphate. Decarboxylation of CBGA gives CBG (carboxylic acid shown in blue).

Synthase is a specific type of enzyme that has had some research done on how they work. Kuroki et al. obtained a crystal structure of the Δ9-THCA synthase and determined where CBGA interacts via hydrogen bonds to the synthase[2]. The mechanisms appear to slightly change from one paper to another but it appears that the FAD accepts an allylic hydride from CBGA which forms a resonance stabilized carbocation. It is important that the alkene is the E isomer to have the correct configuration for the next cascade step which can react in different ways depending on the synthase. Δ9-THCA synthase uses a substitution type reaction where the phenolate oxygen attacks the tri-substituted alkene which then attacks the second alkene forming the two rings (Figure 2). In the case of the CBDA synthase an elimination reaction where the allylic hydrogen is deprotonated, closing the ring forming CBDA. The FADH2 is regenerated from reducing oxygen to hydrogen peroxide. The acid forms are what nature produces which can be decarboxylated with heat, light, or acid.

Figure 2. Mechanism of the synthase of Δ9-THCA and CBDA.



How do we test for cannabinoids?

  • Homogenize the sample
  • Extract the cannabinoids from the plant material (matrix).
  • Centrifuge, cleanup, and dilute the sample.
  • Inject sample into UHPLC (Thermo Scientific Ultimate 3000, see Fig. X) which separates the cannabinoids and other organic molecules on a column based on their interactions with the stationary phase. They are pushed through a diode array detector, DAD, which looks at parts of the UV-Visible spectrum (where the organic compound adsorb light). Depending on the intensity of the adsorption, we can figure out the concentration for the given cannabinoids. This is called a chromatogram (Figure Y).


Figure X. UHPLC Thermo Scientific Ultimate 3000

Thermo Trace™ Ultimate 3000 HPLC system used to quantify cannabinoids.



Figure Y. Example chromatograph.

Note: We use a UHPLC DAD method to quantify cannabinoids.






*1.5 grams of flower sample is needed for an R&D test (unofficial).
For other requirements see our FAQ page.

[1] M. Fellermeier, M. H. Zenk, FEBS Letters 1998, 427, 283-285.

[2] Y. Shoyama, T. Tamada, K. Kurihara, A. Takeuchi, F. Taura, S. Arai, M. Blaber, Y. Shoyama, S. Morimoto, R. Kuroki, Journal of Molecular Biology 2012, 423, 96-105.



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