A Real-World Evaluation of Cannabidiol-Rich Medical Cannabis for Symptoms

An Assessment of Terpinolene in in silico, in vitro, and in vivo Studies

Plants have been used in medicine for thousands of years, with many active ingredients in various species having been found to possess impressive therapeutic potential. One such class of compounds, terpenes, are expressed throughout the plant Kingdom in countless species. Since their discovery, terpenes are structurally diverse secondary metabolites whose chemical and biological properties have been widely investigated in scientific research. Terpenes are the largest group of natural bioactive compounds. They play an important role in the appearance, scent, and taste, contributing to the defence mechanisms of various plants.

Monoterpenes – the most common of several sub-groups of terpenes – comprise about 90% of the composition of all essential oils. Terpinolene is one such monoterpene that is widely found in aromatic plants – including cannabis. Scientific research has revealed that terpinolene possesses several significant pharmacological activities, including antifungal, antioxidant, and insecticide potential. Furthermore, computational predictions by Bosc and collaborators indicate that terpinolene has significant activity threshold for several biological targets, including Muscarinic acetylcholine receptor M1, Prostanoid EP4 receptor, Serotonin 3a (5-HT3a) receptor, and Proto-oncogene tyrosine-protein kinase ROS (Homo sapiens). 

Terpenes have been acknowledged as effective therapeutic alternatives for the treatment of various conditions in herbal medicine traditions from around the world, including in Mediterranean, Ayurvedic, and Chinese Medicine. Unfortunately, few preclinical studies have investigated these activities through in silico (computational), in vitro, and in vivo studies, and no clinical trials have been conducted to demonstrate the therapeutic potential of terpinolene. 

A recent systematic review (2021) aimed to “report the biological activities of terpinolene demonstrated through preclinical studies… to contribute to further research addressing the therapeutic potential of this monoterpene.”

Methods of the Review

The researchers searched selected databases using the following search terms: terpinolene “AND” activity “OR” properties “OR” therapeutic “OR” treatment. After removing duplicates of articles and applying the inclusion and exclusion criteria, 76 articles were selected for the final analysis. Following the individual verification of full texts, a total of 57 articles were included in the final review. 

Most of the included studies were performed exclusively in vivo (n = 38), followed by studies that were performed in vivo (n = 15), both in vivo and in vitro (n = 2), both in silico and in vitro (n = 1), and exclusively in silico. The existing literature indicated an increasing interest in this subject since the first published study in 1967, with the number of studies rising most significantly since 2009. 

Findings of the Review

Collectively, the articles analysed in this review reported that terpinolene has several pharmacological activities, including anti-cholinesterase, sedative, cytotoxic, cytoprotective, anti-inflammatory, antispasmodic, antiproliferative, antinociceptive, lysozyme ligand and n-nitrosamine inhibitor, and P-glycoprotein. As seen with other terpenes, the anti-oxidant and insecticide potential are the most investigated activities of terpinolene. 

Toxicity and Cytotoxicity

Insecticides are substances that are commonly used to kill insects. They are widely used in farming practices to prevent the infestation of crops; however, many of the insecticides used are synthetic compounds that can cause harm to human health and have toxic effects on various organisms. Therefore, the identification of bioecological alternatives has become an important consideration. Plant-derived products may represent a promising option for the development of insecticides that combat disease vectors without causing environmental damage.

The present review of existing in vitro studies demonstrates that terpinolene has been found to present toxic effects against a variety of organisms, especially against insects, larvae, and mites. Other studies have also demonstrated comparable insecticidal activities of other terpenes, including limonene. For example, a study carried out by Chang et al. (2012), demonstrated the high toxicity of terpinolene, p-cymene, o-cymene, and m-cymene against Blattella Germanica (German Cockroach).

In comparison, the potential of terpinolene using in vivo studies has been poorly investigated. Nevertheless, existing studies found that terpinolene was effective against Tetranychus urticae (Red Spider Mites), in a similar capacity to other monoterpenes limonene and p-cymene. The insecticidal effects of terpinolene were found to be influenced by the time of exposure, method of exposure, and environmental conditions, with results suggesting that “increased exposure in closed environments, for a shorter time and in a way that facilitates exposure to this airborne route tends to optimise the desired toxicity.”

The cytotoxic profile of terpinolene has also been investigated on human cells, in order to determine its in vitro safety and to assess its potential therapeutic potential. Past research suggests that terpinolene could have beneficial roles in cancer; for example, previous research demonstrates that the monoterpene, at concentrations of 50 mg/l, has antiproliferative activity against neuroblastoma (a rare type of paediatric cancer) cells (N2a), which may be related to the inhibition of n-nitrosamine (in silico). However, in another study, terpinolene was seen to cause a marked increase in intracellular production of ROS in cancer cells.

Antioxidant Activity

A total of 11 scientific studies reported the in vitro antioxidant activity of terpinolene. The studies found that high terpinolene concentrations have a protective role against oxidative stimuli by increasing the total antioxidant capacity via induction of Akt1 expression. One study (Lu et al. ) demonstrated that terpinolene concentration-dependently promoted a reduction of total oxidant levels and an increase in the antioxidant substances, which was comparable to the positive control (butylated hydroxytoluene). 

The same study also demonstrated that monoterpene γ-terpinene inhibited lipid peroxidation to the same extent as terpinolene (over 80% inhibition, comparable to the standard antioxidant control). This significant antioxidant potential suggests that terpinolene may be a potential drug candidate for the treatment of pathological processes caused by oxidative stress.

Antimicrobial Activity

The literature assessed in the present review suggests that terpinolene also has antimicrobial activities, such as a parasite, antifungal, antibacterial, virucide, and trypanocide. One study showed that, in addition to being active against nine enteric pathogenic bacteria, terpinolene inhibited the growth of acidophilic bacteria playing important roles in the intestinal flora balance. 

Conclusions

This systematic review lists the various properties of terpinolene in a biological context. Having assessed the existing evidence, the researchers “suggest that the prooxidant and cytotoxic effects of terpenes could be explored in drug development in the context of anticancer and antiparasitic research.” It is also suggested that the antioxidant activity of terpinolene could be useful in preventing cell damage caused by oxidative stress, as well as being potentially useful for neurodegenerative diseases such as Alzheimer’s.

The researchers conclude that the included studies suggest that terpinolene may “have the potential to be used in the development of commercial formulations with repellent effects as well in the composition of insecticides.” Nonetheless, it is also acknowledged that a number of the potential pharmacological effects of terpinolene need to be better investigated. Finally, the authors of this systematic review recommend that future in vivo, in vitro, and clinical studies should explore potential therapeutic applications of terpinolene and its potential benefits to human health.