Cannabis sativa plant has not only cannabinoids as crucial compounds but also the other compounds that play important role as synergistic and/or entourage compound. Cannabis/hemp plant materials and essential oils were analyzed with the help of gas chromatography/mass spectrometry detector for the content of terpenes and terpenoids. The main terpenes/terpenoids and their abundance in the samples were evaluated. Results of this study will be helpful in the next evaluation of these compound in mixture with cannabinoids and their importance in medical treatment.

Since the past 25 years, the interest in cannabis (Cannabis sativa L.) has been growing extremely worldwide. The main reason behind such growing interest is not the treatment of some serious illnesses with cannabis but rather the desire to get rich quickly.

Although medical cannabis in some countries is considered legal, it is not yet a pharmaceutical drug. There are 3 main reasons. (a) Fear and stigma – 80 years of prohibition accompanied with negative propaganda. (b) Lack of standardization – since it is not a single-molecule drug. Cannabis in fact is a plant of over 1,000 chemical constituents, varying by chemotype (chemical phenotype) batch and crop. Sometimes, people incorrectly referred to chemotype as strain, variety, cultivar, or chemovar. Chemotypes are plants of the same genus that are virtually identical in appearance but produce essential oil with different major constituents. Chemotypes are variants within a single botanical species. Today, there are thousands of “strains,” many of which have similar names but cultivated in different climatic regions, and each with unique chemical ingredient profile that activates differently. (c) The cart before the horse – cannabis became legal and approved without standard clinical trials. Now, we have to “back in” to the efficacy to see what type of cannabis works for which medical conditions [1].

Cannabis use as medicine goes back for thousands of years [2], but after the USA started to fight against it since 1937, it disappeared from pharmacopeias all over the world. Fortunately, in 1950, Krejčí [3, 4] from Czechoslovakia discovered antibiotic principle of cannabis and they started to use cannabis clinically at hospitals. Back in 1954, the first scientific conference under “Cannabis as a medicine” was held in Olomouc, Czechoslovakia [5]. Krejčí and Šantavý [6] identified this antibiotic principle compound of cannabis, which is effective against gram-positive microorganisms and some pathogens. They named the compound responsible for this effect cannabidiolic acid [6-8]. It was the first real cannabinoid isolated from C. sativa L. From that time, cannabis grown in Czechoslovakia (mostly the Czechoslovak chemotype Rastislavice) was used for the treatment in the Faculty hospital of Olomouc [9, 10] up to 1990.

The first cannabinoid compound isolated and identified from hemp was cannabinol [11-13]. This compound in fact is an artifact, which originates from Δ9-tetrahydrocannabinol (THC) in the plant resin and after plant harvest, storage, or heating [14]. Decarboxylation product of cannabidiolic acid, cannabidiol (CBD), was fully identified in 1963 [15, 16, 19]. Psychotomimetically active compound of cannabis, THC, was fully characterized in 1964 [17-19]. Today, 177 cannabinoid compounds are known in C. sativa L. [20-23]. Many of them are certainly artifacts originating during harvest, drying, and workup of cannabis plant. After the identification of the above-mentioned cannabinoids, research started on these compounds. Cannabinoid receptors, CB1 [24] and CB2 [25], were discovered in the human body. Full understanding of the whole endocannabinoid system in the human body gave an isolation of natural ligands (today called endocannabinoids) [26-31]. This brought us to understand the medicinal value of cannabis plant, which was used for millennia in treatment [2], and today, it is being legalized for the treatment of different diseases in many countries. Because of illegalization and stigmatization of this plant, there is still a lot of work to be accomplished to acquire full knowledge of the medicinal power of this plant [32].

The first scientific study conducted on cannabis contents goes back to the first half of the 19th century [33]. Bolas and Francis [34] made the first attempt to identify the compounds in the commercial resinous extract of Indian hemp in 1869. They isolated oxy-cannabin, C5H6O2, and acid compound, which crystallize in plates.

It was almost generally accepted that the only active compounds in cannabis are compounds typical for this plant, cannabinoids. Only in the last several years scientists started to speculate about synergic and/or entourage effect of the other cannabis compounds. In the first row today are terpenes/terpenoids, but our plans are also to study flavonoids, flavonoid glycosides, and also polyphenols. It is very likely that the first isolated terpene was just β-caryophyllene.

As workers in Italian hemp fields became gay and giddy from a volatile principle of hemp, Valente searched for the compound that causes it. He employed steam distillation of the fresh leaves of hemp cultivated for fibers to obtain sesquiterpene from essential oil, with the empirical formula C15H24 [35-37]. It was probably β-caryophyllene which is the most common main sesquiterpene in hemp. Vignolo [38, 39] also used distillation in a current of steam to prepare essential oil and subsequently the same sesquiterpene C15H24 as Valente. Wood [40] isolated from charas (because it contains no chlorophyll) a monoterpene C10H16 (probably myrcene that is usually the main monoterpene in hemp) and a sesquiterpene C15H24 (β-caryophyllene?). Simonsen and Todd [41] were the first to name isolated terpene. They extracted p-cymene (C10H16), p-cymenene (C10H12), and humulene (C15H24) from Egyptian hashish.

If the question is what are terpenes or terpenoids in cannabis plant, the answer is terpenes are hydrocarbons and terpenoids are oxygen-containing terpenes. They can be visualized as the result of linking isoprene units “head to tail” to form chains, which can be arranged to form rings (Fig. 1). It is the so-called biogenetic isoprene rule or the C5 rule [42].

Fig. 1.

Linking isoprene units “head to tail” to form terpenes/terpenoids.

Fig. 1.

Linking isoprene units “head to tail” to form terpenes/terpenoids.

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The name terpene was suggested by Kekulé [43] for C10H16 hydrocarbons in 1866. In C. sativa L., plant terpenes/terpenoids can be divided into several groups. Here are examples of different types of terpenes (110 published up-to-date) and terpenoids (121 published up-to-date) in this plant – examples of terpenes/terpenoids from cannabis of this particular group are in given parentheses (Tables 1-4):

Cannabinoids are derived from diterpene structure. The monoterpenes present in cannabis plant have another functional moiety like alcohol (fenchyl alcohol, linalool, and borneol), aldehyde (neral), ketone (carvone), ester (bornyl acetate and linalyl acetate), ether (1,8-cineol), and phenol (thymol and carvacrol). The sesquiterpene molecules include structures like alcohols (farnesol) or ketones (nootkatone). To understand the full effects of terpenes/terpenoids, it is necessary to explain some terms.

Synergy

The term synergy comes from the Attic Greek word συνεργία (synergía, “collaboration”), which is based on the word συνεργός (synergos, “working together”). Explanation is easy. When we have 2 active compounds, they work together better than each one separately, which can be expressed by strict inequality: 1 + 1 > 2.

Entourage Effect

The term entourage effect was introduced in 1988 by Mechoulam and colleagues[44] and was explained as increased activity of an active compound with an inactive one, which can be expressed by strict inequality: 1 + 0 > 1. Standardized cannabis drug preparations, rather than pure cannabinoids, could be generally considered the preferred ones [45, 46]. We believe that all components of the cannabis plant likely exert some therapeutic effect, more than any single compound alone. There is increasing evidence that these compounds work better together than in isolation and that is exactly what is called today “entourage effect.”

The analysis of the sample and the interpretation of the results of the analysis must be evaluated very responsibly. We do not have enough results in this field yet. Chemotype, location of cultivation, conditions of cultivation, season of cultivation, weather and microclimate, stage of plant development, method of processing after harvest, method of storage, storage time before analysis, part of a plant for analysis, and method of sample processing for analysis – all these parameters affect the results of the analysis. So far, changes in the content of cannabinoid substances during the growing season have been studied [47-50] and also dynamics of changes of cannabinoids and terpenes/terpenoids during vegetation period [51, 52].

Terpenes/terpenoids have a wide range of biological and pharmacological activities, for instance, antifungal, antiviral, anticancer, anti-inflammatory, antihyperglycemic, antiparasitic, antioxidant, and antimicrobial. It is not possible to describe all pharmacological effects of terpenes/terpenoids in this paper, but we shall give just some examples for imagination of how important these compounds are in this plant. Monoterpene myrcene is the smallest terpene but the most prevalent terpene found in most varieties of cannabis. Chemotypes high in myrcene will result in a “couch lock” effect (if a sample has over 0.5% myrcene), while chemotypes with low levels of myrcene (<0.5% myrcene) will produce a more energetic high. It is simply the amount of myrcene that is in the sample that dictates how you will be affected. Myrcene is simply the important monoterpene in the plant. Myrcene has antipsychotic, antioxidant, analgesic, anti-inflammatory, sedative, muscle relaxant, and anticancerogenic properties [53-56]. The most important sesquiterpene in the cannabis plant is probably β-caryophyllene. It is a spicy terpene. This compound is the only terpene known to interact with the body’s endocannabinoid system (selectively binds to the CB2 receptor) [57]. Caryophyllene has gastroprotective, analgesic, anticancerogenic, antifungal, antibacterial, antidepressant, anti-inflammatory, antiproliferative, antioxidant, anxiolytic, analgesic, and neuroprotective effects [58, 59]. The presence of β-caryophyllene in many essential oils might contribute strongly to their antiviral ability. β-Caryophyllene displayed a high selectivity index of 140 against herpes simplex virus type 1 in vitro. The selectivity index was determined by the ratio of the cytotoxic concentration of the drug that reduced viable cell number by 50% to antiviral activity, which inhibited plaque numbers by 50% compared with the untreated control [60]. α-Pinene (antibacterial, anti-inflammatory, bronchodilator, antiseptic, and gastroprotective) and β-pinene (antiseptic) are the next important compounds [61]. It is not possible to mention here all the biodynamic terpenes. After all, there may be mentioned, for instance, limonene (antibacterial, gastroprotective, antiproliferative, antifungal, anxiolytic, antidepressant, antimicrobial, antispasmodic, or immunostimulant) [62, 63]. Linalool (sedative, antipsychotic, anticonvulsant, anxiolytic, anesthetic, antidepressant, analgesic, antiepileptic, and antineoplastic) [64], terpineol (antioxidant, antibiotic, and relaxing effect) [65], or caryophyllene oxide (analgesic, anticancer, antifungal, and anti-inflammatory) [66]. Between others are also phellandrene (antifungal and digestive disorders) [67], ocimene (antifungal) [68], camphene (cardiovascular disease) [69], guaiol (antitumor) [70, 71], α-humulene (antibacterial, anti-inflammatory, and antitumor) [72-74], γ-terpinene (analgesic, anti-inflammatory, antimicrobial, and anticancer) [75, 76], β-elemene (antitumor, antineoplastic, and anticancer) [77-80], nerolidol (antiparasitic and antileishmanial) [81, 82], or citral (antifungal, antimicrobial, antiproliferative, cytotoxic, anticancer, and antitumor) [83-90].

Terpenes/terpenoids are largely responsible for the characteristic aroma of cannabis. An essential oil (mostly terpenes or terpenoids), especially when distilled, is not necessarily identical in its chemical composition with the oil that is present in the living plant. Quite often very high-boiling or low-boiling chemicals are simply “lost” due to the nature of the distillation process and due to economic and time constrains. Although most constituents remain intact during distillation, a few undergo chemical changes. Oil also contains substances that are formed from reactive precursors on distillation. The variation in essential oil composition may be due to factors that affect the plant’s environment, such as geographical location, weather conditions, soil type, fertilizer used, the age of the plant, and the time and weather of day or year when it is harvested. Degradation tends to occur on prolonged storage, under poor storage conditions, or when the essential oil is otherwise exposed to air. Atmospheric oxygen can change the chemical composition of an essential oil by reacting with some of its constituents. Oxidation can also affect the efficacy of an essential oil. It can render an essential oil more hazardous [91]. When we use plant material, one must take attention to keep it protected from UV and air oxidation. Otherwise, some terpenes can transform to allergens [92, 93].

The first who pointed out the possible synergic and/or entourage effect of cannabinoids and terpenes is Russo [94]. Only few studies have pharmacology, and further studies have to be accomplished prior to understanding the interactions of cannabinoids and terpenes/terpenoids in humans. Well-arranged review on the terpene biosynthesis in cannabis plant was demonstrated by Kovalchuk et al. [95]. Terpene synthases from C. sativa were characterized [96-98]. Rice and Koziel [99] studied the odorous compounds (as terpenes/terpenoids as the other volatiles). Hazekamp et al. [100] gave a deeper understanding of cannabis effects in laboratory and clinical studies and the usefulness of a terpene/terpenoid approach for chemotaxonomic mapping of cannabis varieties for medicinal use. Gallily et al. [101] investigated the antioxidant and anti-inflammatory properties of 3 different terpene/terpenoid-rich hemp essential oils. They concluded that terpenes/terpenoids may be used to diminute acute inflammation effect, whereas the cannabinoids to inhibit chronic inflammation symptoms. Anti-inflammatory potential of terpenes present in cannabis was studied by Downer [102]. Current evidence of medicinal properties of terpenes was given by Baron [103] and Nuutinen [104] in their reviews. Entourage effect of terpenes/terpenoids and cannabinoids and their pharmacological activity were also studied by Namdar et al. [105] and Ferber et al. [106]. Blasco-Benito et al. [46] extracted fresh cannabis flowers in ethanol, after evaporation, followed by magnetic stirring on hot plate, which achieved cannabinoid decarboxylation. The extract (in mg/g) contains 3.4 THCA, 551.3 THC, 3.7 CBG, and no THCV, CBD, CBDA, cannabinol, and CBC and the 5 main terpenes were 1.9 β-caryophyllene, 0.6 humulene, 0.4 nerolidol, 0.6 linalool, and 0.3 β-pinene. While the ethanolic extract of cannabis flowers has higher antitumor activity than pure THC, this effect was not attributed to any of the 5 most abundant terpenes (THC and the 5 main terpenes in appropriate concentrations did not have higher activity than pure THC). Finlay et al. [107] tried to determine whether terpenes (myrcene, α- and β-pinene, β-caryophyllene, and limonene) in the cannabis plant have detectable receptor-mediated activity or modify the activity of THC, CBD, or the endocannabinoid 2-arachidonoylglycerol at the cannabinoid receptors. This study proves that the putative entourage effect cannot be explained by direct effects at CB1 or CB2. Nuutinen [104] has published a comprehensive paper on the medicinal properties of a number of cannabis mono- and sesquiterpene/terpenoids. These were studied in vitro, on animals, and in clinical trials. Performed studies have shown antimutagenic, antidiabetic, anti-inflammatory, analgesic, antioxidant, antibiotic, anticonvulsive, anticancer, antidepressant, anxiolytic, antitumor, neuroprotective, anti-allergic, and others.

Plant Material and Essential Oils

All 54 chemotype inflorescence samples of cannabis were purchased from Israeli growers. Eight essential oils were of hemp harvested in August/September 2016 in the pre-Alpine region of Slovenia (Upper Savinja Valley), latitude NS 46°20′ 29.525 and longitude E 14°50′ 0.777. These samples of essential oils were prepared by steam distillation of female flowers (upper third of the plant). One sample was from Czech Republic (Bialobrzeskie). Essential oils of cannabis were bought from Cali terpenes (45 samples).

Sample Preparation

A 20 mg of plant material was extracted with hexane, filtered with Filter Fix (25 mm, 0.45 μm Nylon Syringe Filter; SOLUFIX – SIMPLEPURE PTFE, 0.45 μm), and dissolved 20 times with hexane. Essential oil was diluted with hexane to a concentration of 0.1 mg/mL.

Conditions of the Analysis

Instrument: GC/MS (Agilent 7890B GC, Agilent 5977B MSD, PAL 3 [RSI 85]).

Column: Agilent Technologies, Inc., HP-5MS UI, 30 m × 0.25 mm, film 0.25 μm.

Experimental conditions: At first, the column was held at 35°C for 5 min, and afterward, the temperature was raised to 150°C at 5°C/min, then at 15°C/min up to 250°C, hold time 90 min (the inlet temperature was fixed at 250°C; the detector at 280°C; split injection 1:5; initial temperature – 100°C; initial time – 4.0 min), gas – helium (flow rate: 1 mL/min).

Identification: The content compounds were identified by comparison with standards, retention times, retention indices, and the spectral matching of libraries NIST/EPA/NIH Mass Spectral Library 2017, Wiley Registry of Mass Spectral Data 11th Edition, FFNSC3, ©2015, and Adams EO library, Mass Spectral Library, 2205 compounds.

In the text, every time for each mentioned chemotype, the first is a picture of their gas chromatographic/mass spectrometric analysis and after that a table of terpenes/terpenoids.

Comparison of Hemp and Cannabis

Initially, we tried to compare the so-called hemp with the so-called cannabis, which is necessary to explain here. Hemp is a cannabis cultivated for fiber but it is also used for treatment. In fact, it is C. sativa L. with very low concentrations of CBD and THC. Cannabis is called today C. sativa L. mainly used for recreational and/or medicinal use. It has a high concentration of CBD and/or THC. The difference between recreational and medicinal cannabis is in cultivation – medicinal cannabis is cultivated under strict conditions. In fact, many chemotypes developed for recreational use are currently called medicinal cannabis. As we did not have all terpenes standards that exist in this plant, we could not quantify all terpenes and terpenoids; therefore, we used comparison based on relative ratio of the main terpene/terpenoid in the particular chemotype.

Hemp (cannabis for fibers) – chemotype Monoica (listed volatiles that are present at a level more than 5% of the main terpene – monoterpenes are marked in bold) (Fig. 2; Table 5):

Fig. 2.

Gas chromatogram of Monoica hemp chemotype.

Fig. 2.

Gas chromatogram of Monoica hemp chemotype.

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Cannabis (cannabis for treatment) – chemotype Lemon OG Kush (listed volatiles that are present at a level more than 5% of the main terpene – sesquiterpenes/sesquiterpenoids are marked in bold) (Fig. 3; Table 6):

Fig. 3.

Gas chromatogram of Lemon OG Kush cannabis chemotype.

Fig. 3.

Gas chromatogram of Lemon OG Kush cannabis chemotype.

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Results of 108 Inflorescences and Essential Oils Analyses

Fifty-four inflorescences of different cannabis chemotypes (Fuji, Everest, Golan, Tropicana, Kilimanjaro (CannDoc), Marom, Edom, Choco 1, Durban Poison, Jack Herer, Black Diamond, Holy Weed, Pandora’s Box, Tel Aviv, Paris, OG Kush, Bubble Gum, Strawberry 2.0, Lemon Deluxe, Berry Deluxe, Medi Kush, Lemon K2, Blue Haze, Annapurna, Choco 2, Himalaya, Lemon OG Kush, AK-47, Jericho, Chocolope, Doblin, K, Kira Kush, Kilimanjaro (Better), Luli Kush, Maui Waui, White Widow, Topaz, DOV, Alaska, Avidekel, Barak, El-Na, Erez, Or, Tal, Jasmine, Shira, Rafael, Mango, Candy Kush, Local Afgan, Power Plant, and Toffy) + 9 essential oils of different hemp chemotypes (Futura, Felina, Fedora, Ferimon, Monoica, Tiborszallasi, Tisza, Bialobrzeskie from Slovenia, and Bialobrzeskie from Czech Republic) + 45 essential oils of different cannabis chemotypes (Jamaican dream, Girl Scout Cookies, Lemon Cookies, Tangie, Gelato, Chocolate Mint OG, Key Lime Pie, Gorilla Glue, Pink Plant, 24K Gold, Critical Jack, Sweet Tooth, Sugaree, Kashmir Kush, Holy Grail Kush, Chemdawg 4, 3 Kings, Sour Diesel, TNT Kush, Monster, Sensi Star, SFV OG, Veneno, Gipsy Haze, Critical, 707 Truthband, Blackberry Kush, Grapefruit OG, Furious Candy, AK-47, High Level, Cinderella 99, Black Dream, 10K Jack, Cheese, Lavender, Truth, Orange Turbo, Dosidos, Nina Limone, Amnesia, Wifi OG, M.I.B., OG Kush, Mojito) = altogether 108 chemotypes.

a.Fifty-eight different terpenes were found between the 10 main terpenes from each of 108 chemotypes (hemp and cannabis inflorescence and essential oil samples).

Sorting according to the frequency of the main terpene in the 10 main ones at each chemotype (numbers outside parentheses indicate in how many phenotypes was this particular terpene between the 10 main ones, and numbers in parentheses indicate in how many phenotypes was this particular terpene/terpenoid the major one) (Table 7).

b.Forty-four different terpenes were found between the 10 main terpenes of each from 54 chemotypes (cannabis inflorescence samples).

Sorting according to the frequency of the main terpene in the 10 main ones at each chemotype (numbers outside parentheses indicates in how many phenotypes was this particular terpene between the 10 main ones, and numbers in parentheses indicate in how many phenotypes was this particular terpene the major one) (Table 8).

c.Twenty-seven different terpenes were found between the 10 main terpenes of each from 46 chemotypes (cannabis essential oils).

Sorting according to the frequency of the main terpene in the 10 main ones at each chemotype (numbers outside parentheses indicate in how many phenotypes was this particular terpene between the 10 main ones, and numbers in parentheses indicates in how many phenotypes was this particular terpene the major one) (Table 9).

d.Seventeen different terpenes were found between the 10 main terpenes of each from 7 chemotypes (hemp essential oils).

Sorting according to the frequency of the main terpene in the 10 main ones at each chemotype (numbers outside parentheses indicate in how many phenotypes was this particular terpene between the 10 main ones, and numbers in parentheses indicate in how many phenotypes was this particular terpene the major one) (Table 10).

e.Examples of different chemotypes with different main terpenes:

1.α-Pinene-dominant chemotype – Kilimanjaro (Fig. 4; Table 11)

Fig. 4.

Gas chromatogram of Kilimanjaro cannabis chemotype.

Fig. 4.

Gas chromatogram of Kilimanjaro cannabis chemotype.

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2.β-Myrcene dominant chemotype – Durban Poison (Fig. 5; Table 12)

Fig. 5.

Gas chromatogram of Durban Poison cannabis chemotype.

Fig. 5.

Gas chromatogram of Durban Poison cannabis chemotype.

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3.Limonene-dominant chemotype – Sweet Tooth (Fig. 6; Table 13)

Fig. 6.

Gas chromatogram of Sweet Tooth cannabis chemotype.

Fig. 6.

Gas chromatogram of Sweet Tooth cannabis chemotype.

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4.Terpinolene-dominant chemotype – Jack Herer (Fig. 7; Table 14)

Fig. 7.

Gas chromatogram of Jack Herer cannabis chemotype.

Fig. 7.

Gas chromatogram of Jack Herer cannabis chemotype.

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5.Linalool-dominant chemotype – Dosidos (Fig. 8; Table 15)

Fig. 8.

Gas chromatogram of Dosidos cannabis chemotype.

Fig. 8.

Gas chromatogram of Dosidos cannabis chemotype.

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6.β-Caryophyllene-dominant chemotype – Edom (Fig. 9; Table 16)

Fig. 9.

Gas chromatogram of Edom cannabis chemotype.

Fig. 9.

Gas chromatogram of Edom cannabis chemotype.

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7.Selina-3,7(11)-diene-dominant chemotype – Lemon OG Kush (Fig. 10; Table 17)

Fig. 10.

Gas chromatogram of Lemon OG Kush cannabis chemotype.

Fig. 10.

Gas chromatogram of Lemon OG Kush cannabis chemotype.

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8.γ-Selinene-dominant chemotype – Fuji (Fig. 11; Table 18)

Fig. 11.

Gas chromatogram of Fuji cannabis chemotype.

Fig. 11.

Gas chromatogram of Fuji cannabis chemotype.

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9.10-Epi-γ-eudesmol-dominant chemotype – Lemon Deluxe (Fig. 12; Table 19)

Fig. 12.

Gas chromatogram of Lemon Deluxe cannabis chemotype.

Fig. 12.

Gas chromatogram of Lemon Deluxe cannabis chemotype.

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10.β-Eudesmol-dominant chemotype – Alaska (Fig. 13; Table 20)

Fig. 13.

Gas chromatogram of Alaska cannabis chemotype.

Fig. 13.

Gas chromatogram of Alaska cannabis chemotype.

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11.α-Eudesmol-dominant chemotype – El Na (Fig. 14; Table 21)

Fig. 14.

Gas chromatogram of El Na cannabis chemotype.

Fig. 14.

Gas chromatogram of El Na cannabis chemotype.

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12.Bulnesol-dominant chemotype – Berry Deluxe (Fig. 15; Table 22)

Fig. 15.

Gas chromatogram of Berry Deluxe cannabis chemotype.

Fig. 15.

Gas chromatogram of Berry Deluxe cannabis chemotype.

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13.α-Bisabolol-dominant chemotype – Bubble Gum (Fig. 16; Table 23)

Fig. 16.

Gas chromatogram of Bubble Gum cannabis chemotype.

Fig. 16.

Gas chromatogram of Bubble Gum cannabis chemotype.

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f.Comparison of terpene content in fresh and dry samples:

Chemotype Pandora’s Box – fresh (Fig. 17; Table 24);

Fig. 17.

Gas chromatogram of Pandora’s Box cannabis chemotype (fresh sample).

Fig. 17.

Gas chromatogram of Pandora’s Box cannabis chemotype (fresh sample).

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Chemotype Pandora’s Box – dry (Fig. 18; Table 25)

Fig. 18.

Gas chromatogram of Pandora’s Box cannabis chemotype (dry sample).

Fig. 18.

Gas chromatogram of Pandora’s Box cannabis chemotype (dry sample).

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g.All identified volatile compounds in 1 chemotype:

Chemotype Lemon OG Kush (cannabis inflorescence) (Fig. 19; Table 26);

Fig. 19.

Gas chromatogram of Lemon OG Kush cannabis chemotype.

Fig. 19.

Gas chromatogram of Lemon OG Kush cannabis chemotype.

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Chemotype Futura (hemp essential oil) (Fig. 20; Table 27);

Fig. 20.

Gas chromatogram of Futura hemp chemotype.

Fig. 20.

Gas chromatogram of Futura hemp chemotype.

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Chemotype Black Dream (cannabis essential oil) (Fig. 21; Table 28)

Fig. 21.

Gas chromatogram of Black Dream cannabis chemotype.

Fig. 21.

Gas chromatogram of Black Dream cannabis chemotype.

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The purpose of this study was to identify and compare different strains of C. sativa L. with emphasis on terpenes/terpenoids percentages. We tried to identify the most common, the most abundant, and the most interesting compounds in dry flowering tops and etheric oils of hemp and cannabis.

From the chromatograms and tables, one can see that there are many different chemotypes of C. sativa according to their terpenes/terpenoids and that the main volatiles can differ one from the other. This will also influence the industrial, medicinal, and recreational use of this plant. The most important is the content of these volatile compounds in medicine as they will influence the treatment of different diseases.

Comparison of essential oils from hemp and cannabis gave different results. In essential oil of hemp, there were mainly monoterpenes, while in cannabis, the sesquiterpenes/sesquiterpenoids predominate. In any of the 108 chemotypes, the main compounds were β-caryophyllene, β-myrcene, α-pinene, α-humulene, limonene, and β-pinene.

Between all the 108 chemotypes were the ones where the main terpene/terpenoid was β-caryophyllene, β-myrcene, α-pinene, limonene, terpinolene, linalool, selina-3,7(11)-diene, γ-selinene, 10-epi-γ-eudesmol, β-eudesmol, α-eudesmol, bulnesol, or α-bisabolol. In plant material (inflorescence) of cannabis (54 chemotypes), the main compounds were β-caryophyllene, β-myrcene, guaiol, 10-epi-γ-eudesmol, selina-3,7(11)-diene, and α-humulene.

When we evaluated hemp and cannabis inflorescence and essential oil samples for 10 the main terpenes in any from 108 analyzed chemotypes, between them were 58 different terpenes/terpenoids. From these 44 different terpenes/terpenoids were found in 54 chemotypes of cannabis inflorescence samples, 27 different ones in 46 chemotypes of cannabis essential oils, and 17 different ones were found in 8 chemotypes of hemp essential oils. Sometimes we can see that the main terpene/terpenoid does not reach 20% of total ones in the analyzed sample: Kilimanjaro – α-pinene (17.45%) (Fig. 4); Durban Poison – β-myrcene (16.75%) (Fig. 5); Edom – β-caryophyllene (15.74%) (Fig. 9); Lemon OG Kush – selina-3,7(11)-diene (10.03%) (Fig. 10); Fuji – γ-selinene (10.85%) (Fig. 11); Lemon Deluxe – 10-epi-γ-eudesmol (10.37%) (Fig. 12); Alaska – β-eudesmol (8.96%) (Fig. 13); El Na – α-eudesmol (9.66%) (Fig. 14); Berry Deluxe – bulnesol (14.72%) (Fig. 15); and Bubble Gum – α-bisabolol (16.03%) (Fig. 16).

It is also worth to discuss degree of terpenes/terpenoids diversity, since it is very different between various chemotypes. Unfortunately, we did not have access to the hemp inflorescence, so we just present only inflorescences of different cannabis chemotypes (Table 29). When we compare hemp essential oil (Table 30) and cannabis essential oil (Table 31), it became clear that almost in all cannabis essential oil samples, the concentration of the main terpene/terpenoid is higher than that in hemp ones. Moreover, in 8 cases, concentration of the main terpene/terpenoid in cannabis essential oil is higher than 50% of total amount of terpenes/terpenoids. It is interesting that the main terpenes/terpenoids above 20% in the analyzed samples were only β-caryophyllene (39×), α-pinene (8×), β-myrcene (8×), terpinolene (8×), limonene (8×), selina-3,7(11)-diene (2×), and linalool (1×).

Table 29.

Inflorescences of cannabis chemotypes (the main terpene in the sample is above 20% of total)

Inflorescences of cannabis chemotypes (the main terpene in the sample is above 20% of total)
Inflorescences of cannabis chemotypes (the main terpene in the sample is above 20% of total)
Table 30.

Hemp essential oils (the main terpene in the sample is above 20% of total)

Hemp essential oils (the main terpene in the sample is above 20% of total)
Hemp essential oils (the main terpene in the sample is above 20% of total)
Table 31.

Cannabis essential oils (the main terpene/s in the sample is above 20% of total)

Cannabis essential oils (the main terpene/s in the sample is above 20% of total)
Cannabis essential oils (the main terpene/s in the sample is above 20% of total)

β-Caryophyllene and β-myrcene are without any doubt the most common (and often the main ones) terpenes in C. sativa L. plant. Because of their unique properties, these are probably the terpenes that play an important role in medicinal properties of hemp and cannabis.

We would like to inform readers also about the monoterpenes/monoterpenoids-sesquiterpene/sesquiterpenoids ratio in the analyzed samples. Our results show that this ratio does not depend if the sample is hemp, cannabis, or essential oils from them. The content of monoterpenes/monoterpenoids prevailed in most samples, sometimes several times more. Very rarely, the content of sesquiterpenes/sesquiterpenoids was higher, but only slightly more. There is a difference between fresh and dry plant monoterpene content, and between etheric oil and organic solvent extract (it also depends on the type of solvent). Different work-up of different solvent extracts can lead to the loss of monoterpenes. Essential oil and organic solvent extract constituents can be different before and after drying process, and before and after storage. A considerably lower content of monoterpenes can be found in the solvent extract compared with the essential oil from steam distillation. Steam distillation also promotes dehydration of labile alcohols.

The most important is which type of information we need. From biogenetic point of view, the best is headspace gas chromatography of the fresh plant material immediately after its harvest. From pharmacological point of view, the best is to analyze the preparation which is used for medicinal purposes. It is without any doubt that in the course from the living material in the plant to the product we are analyzing, there is a loss of volatile substances, especially monoterpenes.

Many terpenes/terpenoids have a therapeutic power in their mixtures with or without phytocannabinoids, and can be very useful and sometime powerful in treatment of diseases. In the Introduction, we have mentioned several terpenes and their potential medicinal properties. We found more therapeutically interesting structures upon doing our analyses. For instance, 10-epi-γ-eudesmol is highly effective against melanoma and column carcinoma cells proliferation [108]; β-eudesmol is antihepatotoxic, antiangiogenic, and antitumor [109-111]; α-eudesmol induces apoptosis [112]; bulnesol possesses antitussive and expectorant activity [113]; α-bisabolol induces apoptosis of malignant tumor cells, cytotoxicity, and antigenotoxicity [114-118]; guaiol is an anti-inflammatory, antimicrobial, and analgesic terpenoid [70, 71]; and α-humulene acts as an appetite suppressant, antibacterial, and antitumor agent and is an effective anti-inflammatory and analgesic sesquiterpene [73, 119]. Monoterpenoid constituents have antioxidant, anti-inflammatory, and estrogenic effects, and these activities are also relevant to current Alzheimer’s disease therapy. Several monoterpenoid alcohols demonstrated good anti-Parkinson’s disease activity. Many monoterpenoids demonstrated promising neuroprotective activity mediated by various systems [120]. There are some other important terpenes such as selina-3,7(11)-diene or γ-selinene which have never been tested for their pharmacological properties.

We must realize that if we evaluate the relative content of terpenes (%) in the sample, it has nothing to do with their quantitative content in the plant. It is clear from Figure 22 and Table 32 that when we analyze a mixture of standards, where each one has the same concentration (25 ppm in hexane), the area of some peaks is up to 3 times smaller than the area of the largest peak. This is very good for comparing samples and the ratio of the content terpenes/terpenoids, but not for quantifying them.

Table 32.

The relative content of terpenes (%) in the sample analyzed by GC/MS

The relative content of terpenes (%) in the sample analyzed by GC/MS
The relative content of terpenes (%) in the sample analyzed by GC/MS
Fig. 22.

GC/MS chromatogram of 23 terpenes/terpenoids standards (RESTEK Catalog. No. 34095 and 34096). Key: 1, α-pinene; 2, camphene; 3, (−)-β-pinene; 4, β-myrcene; 5, Δ3-carene; 6, α-terpinene; 7, p-cymene; 8, d-limonene; 9, 1,8-cineole; (10, 31% α-ocimene + 11, 69% β-ocimene); 12, γ-terpinene; 13, terpinolene; 14, linalool; 15, (−)-isopulegol; 16, geraniol; 17, β-caryophyllene; 18, α-humulene; (19, 39% cis-nerolidol + 20, 61% trans-nerolidol); 21, (−)-caryophyllene oxide; 22, (−)-guaiol; 23, (−)-α-bisabolol.

Fig. 22.

GC/MS chromatogram of 23 terpenes/terpenoids standards (RESTEK Catalog. No. 34095 and 34096). Key: 1, α-pinene; 2, camphene; 3, (−)-β-pinene; 4, β-myrcene; 5, Δ3-carene; 6, α-terpinene; 7, p-cymene; 8, d-limonene; 9, 1,8-cineole; (10, 31% α-ocimene + 11, 69% β-ocimene); 12, γ-terpinene; 13, terpinolene; 14, linalool; 15, (−)-isopulegol; 16, geraniol; 17, β-caryophyllene; 18, α-humulene; (19, 39% cis-nerolidol + 20, 61% trans-nerolidol); 21, (−)-caryophyllene oxide; 22, (−)-guaiol; 23, (−)-α-bisabolol.

Close modal

Today, we know that in the brain concentrations of anandamide (AEA) are in picomoles and 2-arachidonoylglycerol (2-AG) in nanomoles (what is ng AEA/g brain or µg 2-AG/g brain) [121]. Terpenes/terpenoids are in dried cannabis flower material (which is used for medicinal purposes) usually at concentrations mg/g [122]. As mentioned in Casano et al. [123], “the relative content of terpenoids is strongly inherited while total yield per weight of tissue is more subjected to environmental factors.” Relative content (%) of terpenes/terpenoids is more often used for chemosystematic studies (and it was used also in this publication). What can we deduce from the previous few sentences? Receptors need their ligands in very low concentrations, so it is quite possible that these terpenes/terpenoids are important in the treatment, which are in the plant in very low concentrations and which we are not considering today.

The healing power of cannabis most likely resides in terpenes/terpenoids and phytocannabinoids, of which particular compounds, their amount, and the ratio to each other play the most important role in the treatment of particular diseases. It should be emphasized here that flavonoids, flavonoid glycosides, polyphenol, and the other biodynamic compounds will play this game too.

I would like to express my special appreciation and thanks to Mr. Boris Valte from Slovenia and to Mr. Leopold Svatý from the Czech Republic for hemp essential oils.

The authors have no ethical conflicts to disclose.

The authors have no conflicts of interest to declare.

There were no external funding sources to the study in the preparation of data or the manuscript.

Both authors contributed equally to the manuscript.

1.
Strainprint Technologies Inc., Toronto, Canada
.
2.
Hanuš
LO
.
Pharmacological and therapeutic secrets of plant and brain (endo)cannabinoids
.
Med Res Rev
.
2009
;
29
(
2
):
213
71
.
3.
Krejčí
Z
.
Antibioticky princip Cannabis indica. (The antibiotic effect of Cannabis indica.)
.
[dissertation]
.
Brno, Czechoslovakia
:
Masaryk University
;
1950
. p.
106
.
4.
Krejčí
Z
.
The antibacterial effect of Cannabis indica
.
Lék listy
.
1952
;
7
:
500
.
5.
Konopí jako lék
. (
Cannabis as a medicine.
).
Acta Univ Palacki Olomuc
.
1955
;
6
:
27
114
.
6.
Krejčí
Z
,
Šantavý
F
.
Isolace dalších látek z listí indického konopí Cannabis sativa L. (Isolation of other substances from the leaves of the Indian hemp (Cannabis sativa L., varietas indica)
.
Acta Univ Palacki Olomuc
.
1955
;
6
:
59
66
.
7.
Krejčí
Z
,
Horák
M
,
Šantavý
F
.
Konstituce kyseliny kanabidiolové a kyseliny b.t. 133°C isolovaných Z Cannabis sativa L. (Constitution of the cannabidiolic acid and of an acid of the M.P. 133, isolated from Cannabis sativa L.)
.
Acta Univ Palacki Olomuc
.
1958
;
16
:
9
17
.
8.
Hanuš
L
,
Krejčí
Z
,
Hruban
L
.
Isolation of cannabidiolic acid from Turkish variety of cannabis cultivated for fibre
.
Acta Univ Olomuc Fac Med
.
1975
;
74
:
167
72
.
9.
Kabelík
J
,
Krejčí
Z
,
Šantavý
F
.
Cannabis as a medicament
.
Bull Narc
.
1960
;
12
:
5
23
.
10.
Krejčí
Z
.
Antibakterielní látky v prevenci i terapii infekcí. (The antibacterial substances from cannabis in the treatment and prevention of infections.)
.
[dissertation]
.
Olomouc, Czechoslovakia
:
Palacký University
;
1961
. p.
259
.
11.
Jacob
A
,
Todd
AR
.
Cannabis indica. Part II. Isolation of cannabidiol from Egyptian hashish. Observations on the structure of cannabinol
.
J Chem Soc
.
1940
;
649
53
.
12.
Ghosh
R
,
Todd
AR
,
Wilkinson
S
.
Cannabis indica. Part V. The synthesis of cannabinol
.
J Chem Soc
.
1940
;
1393
6
.
13.
Adams
R
,
Baker
BR
,
Wearn
RB
.
Structure of cannabinol. III. Synthesis of cannabinol, 1-Hydroxy-3-n-amyl-6,6,9-trimethyl-6-dibenzopyran
.
J Amer Chem Soc
.
1940
;
62
:
2204
7
.
14.
Hanuš
L
,
Tesarík
K
,
Krejcí
Z
.
Capillary gas chromatography of natural substances from Cannabis sativa L. I. Cannabinol and cannabinolic acid: artefacts
.
Acta Univ Palacki Olomuc Fac Med
.
1985
;
108
:
29
38
.
15.
Adams
R
,
Wolff
H
,
Cain
CK
,
Clark
JH
.
Structure of cannabidiol. V. Position of the alicyclic double bonds
.
J Amer Chem Soc
.
1940
;
62
:
2215
9
.
16.
Mechoulam
R
,
Shvo
Y
.
Hashish-I
.
Tetrahedron
.
1963
;
19
(
12
):
2073
8
.
17.
Adams
R
,
Pease
DC
,
Cain
CK
,
Baker
BR
,
Clark
JH
,
Wolff
H
,
et al
.
Conversion of cannabidiol to a product with marihuana activity. A type reaction for synthesis of analogous substances. Conversion of cannabidiol to cannabinol
.
J Amer Chem Soc
.
1940
;
62
:
2245
6
.
18.
Gaoni
Y
,
Mechoulam
R
.
Isolation, structure, and partial synthesis of an active constituent of hashish
.
J Am Chem Soc
.
1964
;
86
(
8
):
1646
7
.
19.
Šantavý
F
.
Notes on the structure of cannabidiol compounds
.
Acta Univ Palacki Olomuc Fac Med
.
1964
;
35
:
5
9
.
20.
Hanuš
LO
,
Meyer
SM
,
Muñoz
E
,
Taglialatela-Scafati
O
,
Appendino
G
.
Phytocannabinoids: a unified critical inventory
.
Nat Prod Rep
.
2016
;
33
:
1357
92
.
21.
Berman
P
,
Futoran
K
,
Lewitus
GM
,
Mukha
D
,
Benami
M
,
Shlomi
T
,
et al
.
A new ESI-LC/MS approach for comprehensive metabolic profiling of phytocannabinoids in Cannabis
.
Sci Rep
.
2018
;
8
(
1
):
14280
.
22.
Citti
C
,
Linciano
P
,
Russo
F
,
Luongo
L
,
Iannotta
M
,
Maione
S
,
et al
.
A novel phytocannabinoid isolated from Cannabis sativa L. with an in vivo cannabimimetic activity higher than Δ9-tetrahydrocannabinol: Δ9-tetrahydrocannabiphorol
.
Sci Rep
.
2019
;
9
(
1
):
20335
.
23.
Basas-Jaumandreu
J
,
de las Heras
FXC
.
GC-MS metabolite profile and identification of unusual homologous cannabinoids in high potency Cannabis sativa
.
Planta Med
.
2020
;
86
(
5
):
338
347
.
24.
Devane
WA
,
Dysarz
FA
,
Johnson
MR
,
Melvin
LS
,
Howlett
AC
.
Determination and characterization of a cannabinoid receptor in rat brain
.
Mol Pharmacol
.
1988
;
34
(
5
):
605
13
.
25.
Matsuda
LA
,
Lolait
SJ
,
Brownstein
MJ
,
Young
AC
,
Bonner
TI
.
Structure of a cannabinoid receptor and functional expression of the cloned cDNA
.
Nature
.
1990
;
346
(
6284
):
561
4
.
26.
Devane
WA
,
Hanuš
L
,
Breuer
A
,
Pertwee
RG
,
Stevenson
LA
,
Griffin
G
,
et al
.
Isolation and structure of a brain constituent that binds to the cannabinoid receptor
.
Science
.
1992
;
258
(
5090
):
1946
9
.
27.
Hanuš
LO
.
Discovery and isolation of anandamide and other endocannabinoids, Chapter 12, p. 1828–41
. In:
Lambert
DM
, editor.
Cannabinoids in nature and medicine
.
Zurich
:
Verlag Helvetica Chimica Acta, Wiley-VCH
;
2009
. p.
416
.
28.
Hanuš
LO
.
Discovery and isolation of anandamide and other endocannabinoids
.
Chem Biodivers
.
2007
;
4
(
8
):
1828
41
.
29.
Hanuš
L
,
Gopher
A
,
Almog
S
,
Mechoulam
R
.
Two new unsaturated fatty acid ethanolamides in brain that bind to the cannabinoid receptor
.
J Med Chem
.
1993
;
36
(
20
):
3032
4
.
30.
Mechoulam
R
,
Ben-Shabat
S
,
Hanuš
L
,
Ligumsky
M
,
Kaminski
NE
,
Schatz
AR
,
et al
.
Identification of an endogenous 2-monoglyceride, present in canine gut, that binds to cannabinoid receptors
.
Biochem Pharmacol
.
1995
;
50
(
1
):
83
90
.
31.
Hanuš
L
,
Abu-Lafi
S
,
Fride
E
,
Breuer
A
,
Vogel
Z
,
Shalev
DE
.
2-Arachidonyl glyceryl ether, a novel endogenous agonist of the cannabinoid CB1 receptor
.
Proc Natl Acad Sci U S A
.
2001
;
98
(
7
):
3662
5
.
32.
Bonn-Miller
M
,
Pollack
MO
 Jr
,
Casarett
D
,
Dart
R
,
ElSohly
M
,
Good
L
,
et al
.
Priority considerations for medicinal cannabis-related research
.
Cannabis Cannabinoid Res
.
2019
;
4
(
3
):
139
57
.
33.
Bohlig
JF
.
Cannabis sativa und Urtica dioica, chemisch analysirt
.
Jahrbuch für practische Pharmacie und verwandte Fächer, herausgegeben von der pharmaceutischen Gesellschaft der Pfalz, Verlag von J. J. Tascher, Kaiserslautern
,
1840
. p.
1
58
.
34.
Bolas
T
,
Francis
EEH
.
XXXV.-On the products of the action of nitric acid on the resinous extract of Indian hemp
.
J Chem Soc
.
1869
;
22
:
417
9
.
35.
Valente
L
.
Sull’essenza di canapa. [Essential oil from hemp.]
.
Gazz Chim Ital
.
1880
;
10
:
479
81
.
36.
Valente
L
.
Sull’idrocarburo estratto della canapa. [On the hydrocarbon extract of Indian hemp.]
.
Gazz Chim Ital
.
1881
;
11
:
196
8
.
37.
Valente
L
.
Studi sull’essenza di canapa
.
Atti della Reale Academia dei Lincei
.
1881
;
5
:
126
8
.
38.
Vignolo
G
.
Sull’essenza di Cannabis indica
.
Atti della Reale Accademia dei Lincei
.
1894
;
5
(
3
):
404
7
.
39.
Vignolo
G
.
Sull’essenza di Cannabis indica. (Essence of Cannabis indica.)
.
Gazz Chim Ital
.
1895
;
25
(
i
):
110
4
.
40.
Wood
TB
,
Spivey
WTN
,
Easterfield
TH
.
XL.-Charas. The resin of Indian hemp
.
J Chem Soc Trans
.
1896
;
69
:
539
46
.
41.
Simonsen
JL
,
Todd
AR
.
Cannabis indica. Part X. The essential oil from Egyptian hashish
.
J Chem Soc
.
1942
;
1942
(
1
):
188
91
.
42.
Ružička
L
.
The isoprene rule and the biogenesis of terpenic compounds
.
Experientia
.
1953
;
50
(
4
):
395
405
.
43.
Kekulé
A
.
Lehrbuch der organischen Chemie
.
Erlangen, Germany
:
Ferdinand Enke
;
1866
.
Vol. 2
; p.
464
.
44.
Ben-Shabat
S
,
Fride
E
,
Sheskin
T
,
Tamiri
T
,
Rhee
MH
,
Vogel
Z
,
et al
.
An entourage effect: inactive endogenous fatty acid glycerol esters enhance 2-arachidonoyl-glycerol cannabinoid activity
.
Eur J Pharmacol
.
1998
;
353
(
1
):
23
31
.
45.
Russo
EB
.
The case for the entourage effect and conventional breeding of clinical cannabis: no “strain,” no gain
.
Front Plant Sci
.
2018
;
9
:
1969
.
46.
Blasco-Benito
S
,
Seijo-Vila
M
,
Caro-Villalobos
M
,
Tundidor
I
,
Andradas
C
,
García-Taboada
E
,
et al
.
Appraising the “entourage effect”: antitumor action of a pure cannabinoid versus a botanical drug preparation in preclinical models of breast cancer
.
Biochem Pharmacol
.
2018 Nov
;
157
:
285
93
.
47.
Krejčí
Z
,
Hanuš
L
,
Yoshida
T
,
Braenden
OJ
.
The effect of climatic and ecologic conditions upon the formation and the amount of cannabinoid substances in the cannabis of various provenance
.
Acta Univ Olomuc Fac Med
.
1975
;
74
:
147
60
.
48.
Hanuš
L
,
Krejcí
Z
.
Dynamics of changes in the content of cannabinoid substances during the vegetation period of Cannabis sativa L
.
Acta Univ Palacki Olomuc Fac Med
.
1986
;
114
:
11
29
.
49.
Hanuš
L
,
Subová
D
.
The amount of main cannabinoid substances in hemp, cultivated for industrial fibre production and their changes in the course of one vegetation period
.
Acta Univ Palacki Olomuc Fac Med
.
1989
;
122
:
11
23
.
50.
Hanuš
L
,
Dostálová
M
.
The effect of soil fertilization on the formation and the amount of cannabinoid substances in Cannabis sativa L. in the course of one vegetation period
.
Acta Univ Palacki Olomuc Fac Med
.
1994
;
138
:
11
5
.
51.
Aizpurua-Olaizola
O
,
Soydaner
U
,
Öztürk
E
,
Schibano
D
,
Simsir
Y
,
Navarro
P
,
et al
.
Evolution of the cannabinoid and terpene content during the growth of Cannabis sativa plants from different chemotypes
.
J Nat Prod
.
2016
;
79
(
2
):
324
31
.
52.
Koltai
H
,
Namdar
D
.
Cannabis phytomolecule ‘entourage’: from domestication to medical use
.
Trends Plant Sci
.
Forthcoming
2020
.
53.
Zaklin
R
.
Terpene therapy. Abstracts of Papers, 256th ACS National Meeting & Exposition, Boston, MA, August 19–23, 2018 (2018), CHAS-50
.
54.
Gulluni
N
,
Re
T
,
Loiacono
I
,
Lanzo
G
,
Gori
L
,
Macchi
C
,
et al
.
Cannabis essential oil: a preliminary study for the evaluation of the brain effects
.
Evid Based Complement Alternat Med
.
2018
;
2018
:
1709182
.
55.
Jansen
C
,
Shimoda
LMN
,
Kawakami
JK
,
Ang
L
,
Bacani
AJ
,
Baker
JD
,
et al
.
Myrcene and terpene regulation of TRPV1
.
Channels
.
2019
;
13
(
1
):
344
66
.
56.
Kozioł
A
,
Stryjewska
A
,
Librowski
T
,
Sałat
K
,
Gaweł
M
,
Moniczewski
A
,
et al
.
An overview of the pharmacological properties and potential applications of natural monoterpenes
.
Mini Rev Med Chem
.
2014
;
14
(
14
):
1156
68
.
57.
Gertsch
J
,
Leonti
M
,
Raduner
S
,
Racz
I
,
Jian-Zhong
C
,
Xiang-Qun
X
,
et al
.
Beta-caryophyllene is a dietary cannabinoid
.
Proc Natl Acad Sci U S A
.
2008
;
105
(
26
):
9099
104
.
58.
Sharma
C
,
Al Kaabi
JM
,
Nurulain
SM
,
Goyal
SN
,
Kamal
MA
,
Ojha
S
.
Polypharmacological properties and therapeutic potential of β-caryophyllene: a dietary phytocannabinoid of pharmaceutical promise
.
Curr Pharm Des
.
2016
;
22
(
21
):
3237
–64.[
59.
Francomano
F
,
Caruso
A
,
Barbarossa
A
,
Fazio
A
,
La
Torre
C
,
Ceramella
J
,
et al
.
β-Caryophyllene: a sesquiterpene with countless biological properties
.
Appl Sci
.
2019
;
9
(
24
):
5420
.
60.
Astani
A
,
Reichling
J
,
Schnitzler
P
.
Screening for antiviral activities of isolated compounds from essential oils
.
Evid Based Complement Alternat Med
.
2011
;
2011
:
253643
.
61.
Salehi
B
,
Upadhyay
S
,
Orhan
IE
,
Jugran
AK
,
Jayaweera
SLD
,
Dias
DA
,
et al
.
Therapeutic potential of α- and β-pinene: a miracle gift of nature
.
Biomolecules
.
2019
;
9
:
738
.
62.
Erasto
P
,
Viljoen
AM
.
Limonene: a review: biosynthetic, ecological and pharmacological relevance
.
Nat Prod Commun
.
2008
;
3
:
1193
202
.
63.
Sun
J
.
D-limonene: safety and clinical applications
.
Alt Med Rev
.
2007
;
12
:
259
64
.
64.
Kamatou
GPP
,
Viljoen
AM
.
Linalool: a review of a biologically active compound of commercial importance
.
Nat Prod Commun
.
2008
;
3
:
1183
92
.
65.
Khaleel
C
,
Tabanca
N
,
Buchbauer
G
.
α-Terpineol, a natural monoterpene: a review of its biological properties
.
Open Chem
.
2018
;
16
(
1
):
349
61
.
66.
Fidyt
K
,
Fiedorowicz
A
,
Strządała
L
,
Szumny
A
.
β-Caryophyllene and β-caryophyllene oxide-natural compounds of anticancer and analgesic properties
.
Cancer Med
.
2016
;
5
(
10
):
3007
17
.
67.
Zhang
JH
,
Sun
HL
,
Chen
SY
,
Zeng
L
,
Wang
TT
.
Anti-fungal activity, mechanism studies on α-phellandrene and nonanal against Penicillium cyclopium
.
Bot Stud
.
2017
;
58
(
1
):
13
.
68.
Thakre
AD
,
Mulange
SV
,
Kodgire
SS
,
Zore
GB
,
Karuppayil
SM
.
Effects of cinnamaldehyde, ocimene, camphene, curcumin and farnesene on Candida albicans
.
AiM
.
2016
;
6
(
9
):
627
43
.
69.
Vallianou
I
,
Hadzopoulou-Cladaras
M
.
Camphene, a plant derived monoterpene, exerts its hypolipidemic action by affecting SREBP-1 and MTP expression
.
PLoS One
.
2016
;
11
(
1
):
e0147117
.
70.
Yang
Q
,
Wu
J
,
Luo
Y
,
Huang
N
,
Zhen
N
,
Zhou
Y
,
et al
.
(–)-Guaiol regulates RAD51 stability via autophagy to induce cell apoptosis in non-small cell lung cancer
.
Oncotarget
.
2016
;
7
(
38
):
62585
97
.
71.
Yang
X
,
Zhu
J
,
Wu
J
,
Huang
N
,
Cui
Z
,
Luo
Y
,
et al
.
(–)-Guaiol regulates autophagic cell death depending on mTOR signaling in NSCLC
.
Cancer Biol Ther
.
2018
;
19
(
8
):
706
14
.
72.
Jang
H-I
,
Ki-Jong
R
,
Eom
Y-B
.
Antibacterial and antibiofilm effects of α-humulene against Bacteroides fragilis
.
Can J Microbiol
.
2020
;
66
:
389
399
.
73.
Rogerio
AP
,
Andrade
EL
,
Leite
DF
,
Figueiredo
CP
,
Calixto
JB
.
Preventive and therapeutic anti-inflammatory properties of the sesquiterpene alpha-humulene in experimental airways allergic inflammation
.
Br J Pharmacol
.
2009
;
158
(
4
):
1074
87
.
74.
Legault
J
,
Dahl
W
,
Debiton
E
,
Pichette
A
,
Madelmont
JC
.
Antitumor activity of balsam fir oil: production of reactive oxygen species induced by alpha-humulene as possible mechanism of action
.
Planta Med
.
2003
;
69
(
5
):
402
7
.
75.
Guimarães
AG
,
Quintans
JS
,
Quintans
LJ
 Jr
.
Monoterpenes with analgesic activity: a systematic review
.
Phytother Res
.
2013
;
27
(
1
):
1
15
.
76.
Ramalho
TR
,
Pacheco de Oliveira
MT
,
Lima
AL
,
Bezerra-Santos
CR
,
Piuvezam
MR
.
Erratum for: gamma-terpinene modulates acute inflammatory response in mice
.
Planta Med
.
2015
;
81
(
14
):
E3
54
.
77.
Yao
YQ
,
Ding
X
,
Jia
YC
,
Huang
CX
,
Wang
YZ
,
Xu
YH
.
Anti-tumor effect of beta-elemene in glioblastoma cells depends on p38 MAPK activation
.
Cancer Lett
.
2008
;
264
(
1
):
127
34
.
78.
Li
QQ
,
Wang
G
,
Zhang
M
,
Cuff
CF
,
Huang
L
,
Reed
E
.
Beta-elemene, a novel plant-derived antineoplastic agent, increases cisplatin chemosensitivity of lung tumor cells by triggering apoptosis
.
Oncol Rep
.
2009
;
22
(
1
):
161
70
.
79.
Li
QQ
,
Lee
RX
,
Liang
H
,
Zhong
Y
.
Anticancer activity of β-elemene and its synthetic analogs in human malignant brain tumor cells
.
Anticancer Res
.
2013
;
33
(
1
):
65
76
.
80.
Liu
JS
,
He
SC
,
Zhang
ZL
,
Chen
R
,
Fan
L
,
Qiu
GL
,
et al
.
Anticancer effects of β-elemene in gastric cancer cells and its potential underlying proteins: a proteomic study
.
Oncol Rep
.
2014
;
32
(
6
):
2635
47
.
81.
Silva
MP
,
de Oliveira
RN
,
Mengarda
AC
,
Roquini
DB
,
Allegretti
SM
,
Salvadori
MC
,
et al
.
Antiparasitic activity of nerolidol in a mouse model of schistosomiasis
.
Int J Antimicrob Agents
.
2017
;
50
(
3
):
467
72
.
82.
Alonso
L
,
Fernandes
KS
,
Mendanha
SA
,
Gonçalves
PJ
,
Gomes
RS
,
Dorta
ML
,
et al
.
In vitro antileishmanial and cytotoxic activities of nerolidol are associated with changes in plasma membrane dynamics
.
Biochim Biophys Acta Biomembr
.
2019
;
1861
(
6
):
1049
56
.
83.
Leite
MC
,
Bezerra
AP
,
de Sousa
JP
,
Guerra
FQ
,
Lima
EO
.
Evaluation of antifungal activity and mechanism of action of citral against Candida albicans
.
Evid Based Complement Alternat Med
.
2014
;
2014
:
378280
.
84.
Thomas
ML
,
de Antueno
R
,
Coyle
KM
,
Sultan
M
,
Cruickshank
BM
,
Giacomantonio
MA
,
et al
.
Citral reduces breast tumor growth by inhibiting the cancer stem cell marker ALDH1A3
.
Mol Oncol
.
2016
;
10
(
9
):
1485
96
.
85.
Shi
C
,
Song
K
,
Zhang
X
,
Sun
Y
,
Sui
Y
,
Chen
Y
,
et al
.
Antimicrobial activity and possible mechanism of action of citral against Cronobacter sakazakii
.
PLoS One
.
2016
;
11
(
7
):
e0159006
.
86.
Sheikh
BY
,
Sarker
MMR
,
Kamarudin
MNA
,
Mohan
G
.
Antiproliferative and apoptosis inducing effects of citral via p53 and ROS-induced mitochondrial-mediated apoptosis in human colorectal HCT116 and HT29 cell lines
.
Biomed Pharmacother
.
2017
;
96
:
834
46
.
87.
Sanches
LJ
,
Marinello
PC
,
Panis
C
,
Fagundes
TR
,
Morgado-Díaz
JA
,
de-Freitas-Junior
JC
,
et al
.
Cytotoxicity of citral against melanoma cells: the involvement of oxidative stress generation and cell growth protein reduction
.
Tumour Biol
.
2017
;
39
(
3
):
1010428317695914
.
88.
Zielińska
A
,
Martins-Gomes
C
,
Ferreira
NR
,
Silva
AM
,
Nowak
I
,
Souto
EB
.
Anti-inflammatory and anti-cancer activity of citral: optimization of citral-loaded solid lipid nanoparticles (SLN) using experimental factorial design and LUMiSizer®
.
Int J Pharm
.
2018
;
553
(
1–2
):
428
40
.
89.
Kremer
JL
,
Melo
GP
,
Marinello
PC
,
Bordini
HP
,
Rossaneis
AC
,
Sábio
LR
,
et al
.
Citral prevents UVB-induced skin carcinogenesis in hairless mice
.
J Photochem Photobiol B Biol
.
2019
;
198
:
111565
.
90.
Nigjeh
SE
,
Yeap
SK
,
Nordin
N
,
Rahman
H
,
Rosli
R
.
In vivo anti-tumor effects of citral on 4T1 breast cancer cells via induction of apoptosis and downregulation of aldehyde dehydrogenase activity
.
Molecules
.
2019
;
24
(
18
):
3241
.
91.
Tisserand
R
,
Young
R
.
Essential oil safety: a guide for health care professionals
. 2nd ed.
London
:
Elsevier
;
2014
. p.
780
.
92.
Karlberg
AT
,
Magnusson
K
,
Nilsson
U
.
Air oxidation of d-limonene (the citrus solvent) creates potent allergens
.
Contact Derm
.
1992
;
26
(
5
):
332
40
.
93.
Bråred Christensson
J
,
Karlberg
AT
,
Andersen
KE
,
Bruze
M
,
Johansen
JD
,
Garcia-Bravo
B
,
et al
.
Oxidized limonene and oxidized linalool: concomitant contact allergy to common fragrance terpenes
.
Contact Derm
.
2016
;
74
(
5
):
273
80
.
94.
Russo
EB
.
Taming THC: potential cannabis synergy and phytocannabinoid-terpenoid entourage effects
.
Br J Pharmacol
.
2011
;
163
(
7
):
1344
64
.
95.
Kovalchuk
I
,
Pellino
M
,
Rigault
PO
,
van Velzen
R
,
Ebersbach
J
,
Ashnest
JR
, et al
The genomics of cannabis and its close relatives
.
Ann Rev Plant Biol
.
2020
;
71
:
713
739
.
96.
Allen
KD
,
McKernan
K
,
Pauli
C
,
Roe
J
,
Torres
A
,
Gaudino
R
.
Genomic characterization of the complete terpene synthase gene family from Cannabis sativa
.
PLos One
.
2019
;
14
(
9
):
e0222363
.
97.
Booth
JK
,
Page
JE
,
Bohlmann
J
.
Terpene synthases from Cannabis sativa
.
PLos One
.
2017
;
12
(
3
):
e0173911
.
98.
Gunnewich
N
,
Page
JE
,
Kollner
TG
,
Degenhardt
J
,
Kutchan
TM
.
Functional expression and characterization of trichome-specific (−)-limonene synthase and (+)-α-pinene synthase from Cannabis sativa
.
Nat Prod Commun
.
2007
;
2
(
3
):
223
32
.
99.
Rice
S
,
Koziel
JA
.
Characterizing the smell of marijuana by odor impact of volatile compounds: an application of simultaneous chemical and sensory analysis
.
PLos One
.
2015
;
10
(
12
):
e0144160
.
100.
Hazekamp
A
,
Tejkalová
K
,
Papadimitriou
S
.
Cannabis: from cultivar to chemovar II-A metabolomics approach to cannabis classification
.
Cannabis Cannabinoid Res
.
2016
;
1
(
1
):
202
15
.
101.
Gallily
R
,
Yekhtin
Z
,
Hanuš
LO
.
The anti-inflammatory properties of terpenoids from cannabis
.
Cannabis Cannabinoid Res
.
2018
;
3
(
1
):
282
90
.
102.
Downer
EJ
.
Anti-inflammatory potential of terpenes present in Cannabis sativa L
.
ACS Chem Neurosci
.
2020
;
11
(
5
):
659
62
.
103.
Baron
EP
.
Medicinal properties of cannabinoids, terpenes, and flavonoids in cannabis, and benefits in migraine, headache, and pain: an update on current evidence and cannabis science
.
Headache
.
2018
;
58
(
7
):
1139
86
.
104.
Nuutinen
T
.
Medicinal properties of terpenes found in Cannabis sativa and Humulus lupulus
.
Eur J Med Chem
.
2018
;
157
:
198
228
.
105.
Namdar
D
,
Voet
H
,
Ajjampura
V
,
Nadarajan
S
,
Mayzlish-Gati
E
,
Mazuz
M
,
et al
.
Terpenoids and phytocannabinoids co-produced in Cannabis sativa strains show specific interaction for cell cytotoxic activity
.
Molecules
.
2019
;
24
(
17
):
3031
.
106.
Ferber
SG
,
Namdar
D
,
Hen-Shoval
D
,
Eger
G
,
Koltai
H
,
Shoval
G
,
et al
.
The “entourage effect”: terpenes coupled with cannabinoids for the treatment of mood disorders and anxiety disorders
.
Curr Neuropharmacol
.
2020
;
18
(
2
):
87
96
.
107.
Finlay
DB
,
Sircombe
KJ
,
Nimick
M
,
Jones
C
,
Glass
M
.
Terpenoids from cannabis do not mediate an entourage effect by acting at cannabinoid receptors
.
Front Pharmacol
.
2020
;
11
:
359
.
108.
Venditti
A
,
Frezza
C
,
Sciubba
F
,
Serafini
M
,
Bianco
A
,
Cianfaglione
K
,
et al
.
Volatile components, polar constituents and biological activity of tansy daisy (Tanacetum macrophyllum (Waldst. et Kit.) Schultz Bip.)
.
Ind Crops Prod
.
2018
;
118
:
225
35
.
109.
Kiso
Y
,
Tohkin
M
,
Hikino
H
.
Antihepatotoxic principles of Atractylodes rhizomes
.
J Nat Prod
.
1983 Sep–Oct
;
46
(
5
):
651
4
.
110.
Ma
EL
,
Li
YC
,
Tsuneki
H
,
Xiao
JF
,
Xia
MY
,
Wang
MW
,
et al
.
Beta-eudesmol suppresses tumour growth through inhibition of tumour neovascularisation and tumour cell proliferation
.
J Asian Nat Prod Res
.
2008 Jan–Feb
;
10
(
1–2
):
159
67
.
111.
Tsuneki
H
,
Ma
EL
,
Kobayashi
S
,
Sekizaki
N
,
Maekawa
K
,
Sasaoka
T
,
et al
.
Antiangiogenic activity of beta-eudesmol in vitro and in vivo
.
Eur J Pharmacol
.
2005 Apr 11
;
512
(
2–3
):
105
15
.
112.
Bomfim
DS
,
Ferraz
RP
,
Carvalho
NC
,
Soares
MB
,
Pinheiro
ML
,
Costa
EV
,
et al
.
Eudesmol isomers induce caspase-mediated apoptosis in human hepatocellular carcinoma HepG2 cells
.
Basic Clin Pharmacol Toxicol
.
2013 Nov
;
113
(
5
):
300
6
.
113.
Lu
Y-C
.
Studies on the chemical constituents of the essential oil of Rhododendron tsinghaiense Ching
.
Huaxue Xuebao
.
1980
;
38
(
3
):
241
9
.
114.
Cavalieri
E
,
Mariotto
S
,
Fabrizi
C
,
de Prati
AC
,
Gottardo
R
,
Leone
S
,
et al
.
Alpha-bisabolol, a nontoxic natural compound, strongly induces apoptosis in glioma cells
.
Biochem Biophys Res Commun
.
2004 Mar 12
;
315
(
3
):
589
94
.
115.
Darra
E
,
Lenaz
G
,
Cavalieri
E
,
Fato
R
,
Mariotto
S
,
Bergamini
C
,
et al
.
Alpha-bisabolol: unexpected plant-derived weapon in the struggle against tumour survival?
Ital J Biochem
.
2007 Dec
;
56
(
4
):
323
8
.
116.
Darra
E
,
Abdel-Azeim
S
,
Manara
A
,
Shoji
K
,
Maréchal
JD
,
Mariotto
S
,
et al
.
Insight into the apoptosis-inducing action of alpha-bisabolol towards malignant tumor cells: involvement of lipid rafts and Bid
.
Arch Biochem Biophys
.
2008 Aug 15
;
476
(
2
):
113
23
.
117.
Rigo
A
,
Ferrarini
I
,
Lorenzetto
E
,
Darra
E
,
Liparulo
I
,
Bergamini
C
,
et al
.
BID and the α-bisabolol-triggered cell death program: converging on mitochondria and lysosomes
.
Cell Death Dis
.
2019 Nov 26
;
10
(
12
):
889
.
118.
Anter
J
,
Romero-Jiménez
M
,
Fernández-Bedmar
Z
,
Villatoro-Pulido
M
,
Analla
M
,
Alonso-Moraga
A
,
et al
.
Antigenotoxicity, cytotoxicity, and apoptosis induction by apigenin, bisabolol, and protocatechuic acid
.
J Med Food
.
2011 Mar
;
14
(
3
):
276
83
.
119.
Fernandes
ES
,
Passos
GF
,
Medeiros
R
,
da Cunha
FM
,
Ferreira
J
,
Campos
MM
,
et al
.
Anti-inflammatory effects of compounds alpha-humulene and (−)-trans-caryophyllene isolated from the essential oil of Cordia verbenacea
.
Eur J Pharmacol
.
2007 Aug 27
;
569
(
3
):
228
36
.
120.
Volcho
KP
,
Laev
SS
,
Ashraf
GM
,
Aliev
G
,
Salakhutdinov
NF
.
Application of monoterpenoids and their derivatives for treatment of neurodegenerative disorders
.
Curr Med Chem
.
2018
;
25
(
39
):
5327
46
.
121.
Buczynski
MW
,
Parsons
LH
.
Quantification of brain endocannabinoid levels: methods, interpretations and pitfalls
.
Br J Pharmacol
.
2010
;
160
(
3
):
423
42
.
122.
Fischedick
JT
,
Hazekamp
A
,
Erkelens
T
,
Choi
YH
,
Verpoorte
R
.
Metabolic fingerprinting of Cannabis sativa L., cannabinoids and terpenoids for chemotaxonomic and drug standardization purposes
.
Phytochemistry
.
2010
;
71
(
17–18
):
2058
73
.
123.
Casano
S
,
Grassi
G
,
Martini
V
,
Michelozzi
M
.
Variations in terpene profiles of different strains of cannabis sativa L
.
Acta Hortic
.
2011
;
925
(
925
):
115
21
.
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