Newly identified phytocannabinoids have four- and seven-carbon side alkyl chains

Almost 150 structurally and physiologically distinct cannabinoid compounds are unique to Cannabis. These compounds collectively known as phytocannabinoids are produced by an enzymatic reaction between a resorcinol and an isoprenoid group. The precursor for the most ubiquitous naturally occurring phytocannabinoids is olivetolic acid, which undergoes a condensation reaction leading to the formation of cannabigerolic acid (CBGA) (Figure 1). Specific synthases convert CBGA to cannabichromenic acid (CBCA) or cannabidiolic acid (CBDA), which can be converted to tetrahydrocannabinolic acid (THCA). These carboxylated forms are converted to a corresponding decarboxylated form when exposed to heat. The best-known decarboxylated cannabinoids are cannabidiol (CBD) and Δ⁹-THC (the principal psychoactive constituent of Cannabis).

Figure 1. Synthesis and degradation of olivetol series phytocannabinoids.

Alkyl Chain Length Differences

The alkyl side chain on the resorcinol moiety of olivetol cannabinoids is five carbon atoms long. However, other phytocannabinoids such as those derived from varinolic acid contain three carbon atoms on their side chain, and those derived from orcinolic acid contain one carbon atom. You can refer to Cayman’s Phyocannabinoid Guide to see the biosynthesis and degradation pathways of phytocannabinoids derived from the olivetol, varinol, and orcinol series in detail. A recently identified butyl phytocannabinoid series represented by cannabidibutol (CBDB) and Δ⁹-tetrahydrocannabutol (Δ⁹-THCB) has a four-carbon alkyl chain (Figure 2). Since evidence of a corresponding biosynthesis enzyme has not been found, these butyl phytocannabinoids are thought to derive from microbial ω-oxidation and decarboxylation of their corresponding five-carbon homologs. Phytocannabinoids with an alkyl side chain longer than five carbon atoms were not thought to occur naturally until cannabidiphorol (CBDP) and Δ⁹-tetrahydrocannabiphorol (Δ⁹-THCP), seven-carbon homologs of CBD and Δ⁹-THC, were identified in a medicinal Cannabis variety, the Italian FM2 (Figure 2). These phytocannabinoids are named for their resorcinol moiety 5-heptyl-benzene-1,3-diol, which is commonly referred to as sphaerophorol.

Figure 2. Newly identified butyl and phorol phytocannabinoids are analogs of CBD and Δ⁹-THC with differing alkyl side chain lengths.

Side Alkyl Chain Influences Biological Activity

The length of the alkyl side chain is incredibly important for the interaction of Δ⁹-THC with the human cannabinoid receptors CB₁ and CB₂ as it directly correlates with receptor binding affinity. Structure-activity relationship studies of Δ⁹-THC and Δ⁸-THC homologs demonstrate a minimum of three carbons is necessary to bind the CB₁ receptor.¹ Maximum activity has been registered with an eight-carbon side chain. Indeed, synthetic analogs of Δ⁹-THC with a side chain longer than five carbons have shown cannabimimetic properties several times more potent than Δ⁹-THC itself. For example, JWH 091 a seven-carbon chain homolog was shown to have nearly double the receptor affinity from that of Δ⁹-THC or Δ⁸-THC.¹ The optimal binding affinity for CB₁ lies between five and eight carbons. Any higher number of carbon atoms is associated with a decrease in activity.

Interestingly, the binding activity of Δ⁹-THCB at human CB₁ receptor in vitro was reported to be three-times higher than Δ⁹-THC (K_is = 15 and 40 nM, respectively) and five-times higher than Δ⁹-THCV (K_i = 75.4 nM) with no significant differences in CB₂ receptor binding affinity.² The binding activity of Δ⁹-THCP against human CB₁ receptor (K_i = 1.2 nM) was over 30-times more potent than Δ⁹-THC and comparable to that of the potent full CB₁ agonist (±)-CP 55,940 (K_i = 0.9 nM).³ In the cannabinoid tetrad pharmacological test, Δ⁹-THCP induced hypomotility, analgesia, catalepsy, and decreased rectal temperature indicating a THC-like cannabimimetic activity.³ The presence of these new phytocannabinoids could account for the pharmacological properties of some Cannabis varieties difficult to explain only by the presence of Δ⁹-THC. Because Δ⁹-THCP has higher binding affinity for the CB₁ receptor and greater cannabimimetic activity than Δ⁹-THC itself, attention may shift to this newly discovered phytocannabinoid to explore its pharmacological potential to treat a wide range of disorders.

Identifying Impurities

While the binding affinity of CBDB and CBDP to CB₁ and CB₂ receptors has not been reported, the binding affinity of CBD is known to be poor. If the change in alkyl chain length affects the binding affinity to CB₁ and CB₂ as it does for Δ⁹-THCP, then CBDP could potentially be a more potent form of CBD. Knowledge of this becomes particularly important to the production of pharmaceutical-grade CBD as any impurities present could alter its efficacy. Indeed, cannabidivarin (CBDV), the three-carbon alkyl side chain homolog of CBD, and CBDB have both been identified as major impurities in CBD produced by hemp extraction.⁴ The amount of these two impurities in the final product could be relatively high, up to 1% CBDV and 0.5% CBDB (w/w), which would necessitate chromatographic purification to generate CBD that is suitable for pharmaceutical use. Cayman offers analytical standards for CBDV, CBDB, and CBDP that can be applied to suitable analytical methods to quantify these compounds.

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References

1. Bow, E.W. and Rimoldi. J.M. The structure-function relationships of classical cannabinoids: CB1/CB2 modulation. Perspect. Medicin. Chem.8, 17-39 (2016).

2. Linciano, P., Citti, C., Luongo, L., et al. Isolation of a high-affinity cannabinoid for the human CB1 receptor from a medicinal Cannabis sativa variety: Δ⁹-Tetrahydrocannabutol, the butyl homologue of Δ⁹-tetrahydrocannabinol. J. Nat. Prod.83(1), 88-98 (2020).

3. Citti, C., Linciano, P., Russo, F., et al. A novel phytocannabinoid isolated from Cannabis sativa L. with an in vivo cannabimimetic activity higher than Δ⁹-tetrahydrocannabinol: Δ⁹-Tetrahydrocannabiphorol. Sci. Rep. 9:20335 (2019).

4. Citti, C., Linciano, P., Forni, F., et al. Analysis of impurities of cannabidiol from hemp. Isolation, characterization and synthesis of cannabidibutol, the novel cannabidiol butyl analog. J. Pharm. Biomed. Anal. 175:112752 (2019).