Terpenes and Terpenoids: New Study Confirms Entourage Effect

Identifying specific terpene:cannabinoid combinations could be a new means of therapy improvement, finds a new study published in Nature. The first animal model study on terpenes cannabimimetic confirms the entourage effect of terpenes and terpenoids when combined with cannabinoids. It also finds that these effects depend on CB1 and A2a receptors, as well as independent mechanisms.

Terpenes and terpenoids

Terpenes are basic hydrocarbons constituents of essential oils found in many plants, while terpenoids are oxygen-containing terpenes. Both groups of compounds are responsible for the aroma of cannabis. In the cannabis plant, cannabinoids and terpenoids have a common parent compound – geranyl pyrophosphate. Apart from that, the research has shown that they are quite potent, affecting animal and human behavior even when inhaled from the ambient air. For example, a study on mice has shown that the effects on mice inhaling terpenoids for one hour are comparable with the effects of the human intake of THC for pain or symptoms of multiple sclerosis.

In the cannabis plant, terpenoid production increases with light exposure but decreases with soil fertility. The glasshouse experience has demonstrated higher yields if plants experience relative nitrogen lack just before harvest, favoring floral over foliar growth. Other influencing factors can be chemotype, location of cultivation, conditions of cultivation, the season of cultivation, weather, microclimate, stage of plant development, method of processing after harvest, method of storage, storage time before analysis, part of a plant for analysis, as well as the method of sample processing.

A study conducted by Lumír Ondřej Hanuš and Yotam Hod and published in Karger in 2020 investigated 54 inflorescences of different cannabis chemotypes, nine essential oils of different hemp chemotypes, and 45 essential oils of different cannabis chemotypes. Fifty-eight different terpenes were found between the 10 main terpenes from each of 108 chemotypes in hemp and cannabis inflorescence and essential oil samples. The study aimed to identify and compare different strains of Cannabis sativa L. with emphasis on terpenes/terpenoids percentages, and these are the chemotypes established to have different terpenes dominance:

α-Pinene-dominant chemotype – Kilimanjaro
β-Myrcene dominant chemotype – Durban Poison
Limonene-dominant chemotype – Sweet Tooth
Terpinolene-dominant chemotype – Jack Herer
Linalool-dominant chemotype – Dosidos
β-Caryophyllene-dominant chemotype – Edom
Selina-3,7(11)-diene-dominant chemotype – Lemon OG Kush
γ-Selinene-dominant chemotype – Fuji
10-Epi-γ-eudesmol-dominant chemotype – Lemon Deluxe
β-Eudesmol-dominant chemotype – Alaska
α-Eudesmol-dominant chemotype – El Na
Bulnesol-dominant chemotype – Berry Deluxe
α-Bisabolol-dominant chemotype – Bubble Gum

Terpenes/Terpenoids therapeutic properties

Terpenes and terpenoids have a wide range of biological and pharmacological activities. They have antifungal, antiviral, anticancer, anti-inflammatory, antihyperglycemic, antiparasitic, antioxidant, and antimicrobial properties. Here is a list of the most important terpenes found in the Cannabis sativa plant with their therapeutic properties.

Myrcene is the smallest but the most prevalent terpene found in most varieties of cannabis. When used in a combination with THC, plant chemotypes high in myrcene will result in a “couch-lock” effect, while chemotypes with low levels of myrcene (<0.5% myrcene) will produce a more energetic high. Myrcene has antipsychotic, antioxidant, analgesic, anti-inflammatory, sedative, muscle relaxant, and anticancer properties.

The most important terpene in the cannabis plant is probably β-caryophyllene. It is a spicy terpene with gastroprotective, analgesic, anticancer, antifungal, antibacterial, antidepressant, anti-inflammatory, antiproliferative, antioxidant, anxiolytic, analgesic, and neuroprotective effects. The presence of β-caryophyllene in many essential oils might contribute strongly to their antiviral ability. It also displayed a high selectivity index against herpes simplex virus type 1.

The next important compounds are α-pinene with antibacterial, anti-inflammatory, bronchodilator, antiseptic, and gastroprotective activity, and β-pinene, efficient as an antiseptic.

Limonene, most commonly associated with citrus fruits, is also present in Cannabis sativa. It has antibacterial, gastroprotective, antiproliferative, antifungal, anxiolytic, antidepressant, antimicrobial, antispasmodic, or immunostimulant activity.

Linalool, common to lavender, can be used as a sedative, antipsychotic, anticonvulsant, anxiolytic, anesthetic, antidepressant, analgesic, antiepileptic, and antineoplastic.

Terpineol has antioxidant, antibiotic, and relaxing effects, and caryophyllene oxide can be used as an analgesic, anticancer, antifungal, and anti-inflammatory compound.

There is also phellandrene (antifungal and digestive disorders), ocimene (antifungal), camphene (cardiovascular disease), guaiol (anti-inflammatory, antimicrobial, and analgesic), α-humulene (appetite suppressant, antibacterial, anti-inflammatory, and antitumor), γ-terpinene (analgesic, anti-inflammatory, antimicrobial, and anticancer), β-elemene (antitumor, antineoplastic, and anticancer), nerolidol (antiparasitic and antileishmanial), and citral (antifungal, antimicrobial, antiproliferative, cytotoxic, anticancer, and antitumor).

Furthermore, 10-epi-γ-eudesmol is highly effective against melanoma and column carcinoma cells proliferation; β-eudesmol is antihepatotoxic, antiangiogenic, and antitumor; α-eudesmol induces apoptosis; bulnesol possesses antitussive and expectorant activity; α-bisabolol induces apoptosis of malignant tumor cells, cytotoxicity, and antigenotoxicity.

Entourage effect

What puzzles the scientists who focus on phytocannabinoid properties are terpenes’ cannabimimetic activities. It seems that the terpenes/terpenoids found in the Cannabis sativa plant can produce an ‘entourage effect‘ – they can modulate cannabinoids in a way that results in unique and improved outcomes. The entourage effect was first described in 1998 by Shimon Ben-Shabat and Raphael Mechoulam with an idea that cannabinoids within the cannabis plant work together, or possess synergy, and affect the body in a mechanism similar to the body’s endocannabinoid system.

One of the scientists who contributed the most to this theory was Ethan Russo, whose highly cited review published in the British Journal of Pharmacology in 2011 provides an excellent insight into the subject and introduces possibilities for phytocannabinoid-terpenoid synergy. For example, limonene and linalool could be promising anti-acne agents, CBG with α-pinene could produce the safest possible new antiseptic agent while α-pinene improves mental clarity and could be used as an antidote for THC intoxication.

The latest research confirms the entourage effect theory, proving that terpenes and terpenoids can boost activity in cannabinoids.

The new study

The newest study from the University of Arizona published in April in Nature investigated if terpenes and terpenoids when combined with cannabinoids can produce an additive effect. It observed α-humulene, β-pinene, linalool, geraniol, and β-caryophyllene. These were selected because of their quantities in the Cannabis sativa plant, with α-humulene, β-pinene, linalool, and β-caryophyllene being found in higher amounts. In a serial of experiments, the scientists tested the activity of those compounds in the cannabinoid tetrad of behaviors mediated by the CB1 receptor (antinociception, hypolocomotion, catalepsy, and hypothermia) at several doses (50–200 mg/kg). β -caryophyllene, already identified as a selective CB2 agonist, was administered at 100 mg/kg.

The findings showed that the tested terpenes generally although selectively increase the behavioral effects of the cannabinoid, supporting the potential modulation of cannabinoids by terpenes.

This was also the first study to identify the CB1 and A2a receptors as terpene targets and describe the role of these receptors in producing terpene cannabimimetic effects in vivo. The results show that behavioral effects depend on a mix of CB1-dependent, A2a-dependent, and independent mechanisms.

According to the results, cannabis terpenes can have significant pharmacological effects and could be used to selectively modulate the impact of cannabis or cannabinoid therapy. These findings show that terpenes could be used to enhance the analgesic properties of such a therapy, without worsening the side effects. However, this must still be confirmed in new trials using relevant phytocannabinoid instead of the synthetic cannabinoid used in this study. Identifying specific terpene: cannabinoid combinations with a maximized therapeutic index for a particular disease state could provide a new means to improve human therapy with these drugs, the scientists conclude.

Sex differences with linalool

For all experiments in this study, both male and female mice were used, and in nearly every case, there was no difference in results. However, specific sex differences occurred with linalool. When linalool was combined with WIN55,212-2 in the test of pain response; males showed greater additive effects that occurred earlier in the time, while females showed a delay in response and no potentiation over linalool alone. When observing hypolocomotion, the scientists also found the difference suggesting this behavior is mediated by CB1 in males and A2a in females. These observations suggest that other terpene sex differences could be found, although the mechanism for this difference is unknown.