Korean J Pain.  2022 Apr;35(2):140-151. 10.3344/kjp.2022.35.2.140.

Antinociceptive, anti-inflammatory, and cytotoxic properties of Origanum vulgare essential oil, rich with β-caryophyllene and β-caryophyllene oxide

• Moghrovyan A
• Parseghyan L
• Sevoyan G
• Darbinyan A
• Sahakyan N
• Gaboyan M
• Karabekian Z
• Voskanyan A

• KMID: 2527765
• DOI: http://doi.org/10.3344/kjp.2022.35.2.140

Abstract

Background
Essential oils are of great interest for their analgesic and anti-inflammatory properties. We aimed to study the content of the essential oil of the Origanum vulgare of the Armenian highlands (OVA) in different periods of vegetation and to investigate its antinociceptive and anti-inflammatory effects in mice (in vivo) and cytotoxic action in cultured cells (in vitro). OVA essential oil was extracted from fresh plant material by hydro-distillation.
Methods
For OVA essential oil contents determination the gas chromatographymass spectrometry method was used. Formalin and hot plate tests and analysis of cell viability using the methyl-thiazolyl-tetrazolium (MTT) assay were used.
Results
The maximal content of β-caryophyllene and β-caryophyllene oxide in OVA essential oil was revealed in the period of blossoming (8.18% and 13.36%, correspondently). In the formalin test, 4% OVA essential oil solution (3.5 mg/mouse) exerts significant antinociceptive and anti-inflammatory effects (P = 0.003). MTT assay shows approximately 60% cytotoxicity in HeLa and Vero cells for 2.0 μL/mL OVA essential oil in media.
Conclusions
The wild oregano herb of Armenian highlands, harvested in the blossoming period, may be considered as a valuable source for developing pain-relieving preparations.

Keyword

Analgesia; Anti-Inflammatory Agents; Beta-Caryophyllene; Caryophyllene Oxide; Cell Survival; Gas Chromatography-Mass Spectrometry; Nociception; Oils; Volatile; Origanum; Pain; Pain Measurement

Figure

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  Fig. 1 GC-MS chromotograph of OVA essential oil in the period of pre-blossoming. 1: (+)-Sabinene, 2: β-Pinene, 3: α-Terpinolene, 4: Eucalyptol, 5: trans-β-Ocimene, 6: 1-Isopropyl-4-methyl-1,4-cyclohexadiene, 7: β-Ocimene, 8: Ethyl amyl ketone, 9: ο-Cymene, 10: trans-β-Terpineol, 11: Neo-allo-ocimene, 12: 3-Octanol, 13: p-Menthan-3-one, 14: Elixene, 15: (-)-β-Bourbonene, 16: β-Linalool, 17: β-Caryophyllene, 18: L-4-terpineol, 19: α-Humulen, 20: D-Germacrene, 21: α-Muurolene, 22: γ-Elemene, 23: 1-Isopropyl-4,7-dimethyl-1,2,3,5,6,8a-hexahydronaphtalene, 24: (R)-(+)-β-Citronellol, 25: β-Caryophyllene epoxide, 26: d-Viridiflorol, 27: Ent-Spathulenol, 28: 1-Methyl-4-(methylethyl)-(E)-2-cyclohexenol, 29: tau-Cadinol, 30: α-Cadinol, 31: 6-Isopropyl-4,8a-dimethyl-1,2,3,5,6,7,8,8a-octahydro-2-naphthalenol, 32: Methyl 1,5,5-trimethyl-6-[(1E)-3-methyl-1,3-butadienyl]-7-oxabicyclo[4.1.0.]hept-2-yl ether, 33: trans-Phytol, 34: Diepicadrene-1-oxide, 35: (3E)-4-(1,5-Dihydroxy-2,6,6-trimethyl-2-cyclohexen-1-yl)-3-buten-2-one, 36: cis-Z-α-Bisabolene epoxide, 37: 3-[(2E)-2-Dodecenyl]dihydro-2,5-furandione, 38: Isoaromadendrene epoxide, 39: 2-Methyl-4-(2,6,6-trimethylcyclohex-1-enyl)but-2-en-1-ol. GC-MS: gas chromatography–mass spectrometry, OVA: Origanum vulgare of the Armenian highlands.

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  Fig. 2 GC-MS chromotograph of OVA essential oil in the period of blossoming. 1: β-Thujene, 2: (+)-Sabinene, 3: β-Pinene, 4: α-Terpinolene, 5: D-Limonene, 6: Eucalyptol, 7: γ-Terpinene, 8: Ethyl amyl ketone, 9: ο-Cymene, 10: 4-methyl-3-(1-methylethylidene)-1-cyclohexene, 11: 2-Hexenal diethyl acetal, trans, 12: 3-Octanol, 13: cis-β-Terpineol, 14: alfa-Copaene, 15: Dihydroedulan II, 16: (-)-β-Bourbonene, 17: β-Linalool, 18: β-Caryophyllene, 19: L-4-terpineol, 20: Alloaromadendrene, 21: Humulen, 22: Germacrene D, 23: I-β-Bisabolene, 24: Elixene, 25: α-trans-Farnesene, 26: β-Caryophyllene epoxide, 27: α-Humulene epoxide II, 28: Ent-Spathulenol, 29: Isoaromadendrene epoxide, 30: Carvacrol/Isothymol, 31: α-Cadinol, 32: Ledene oxide-(II), 33: Tetracyclo[6.3.2.0(2,5).0(1,8)]tridecan-9-ol,4,4-dimethyl-, 34: Dibutyl phthalate, 35: Palmitic acid. GC-MS: gas chromatography–mass spectrometry, OVA: Origanum vulgare of the Armenian highlands.

• 

  Fig. 3 GC-MS chromotograph of OVA essential oil in the period of fruiting. 1: Sabinene, 2: Eucalyptol, 3: trans-β-Ocimene, 4: γ-Terpinene, 5: β-Ocimene, 6: o-Cymene, 7: α-Terpinolene, 8: Ethylamylcarbinol, 9: Matsutake, 10: (-)-β-Bourbonene, 11: β-Linalool, 12: β-Caryophyllene, 13: L-4-terpineol, 14: α-Humulene, 15: α-Terpineol, 16: Germacrene D, 17: I-β-Bisabolene, 18: Elixene, 19: trans-α-Farnesene, 20: (+)-δ-Cadinene, 21: β-Caryophyllene epoxide, 22: Ent-Spathulenol, 23: tau-Cadinol, 24: 6-Isopropenyl-4,8a-dimethyl-3,5,6,7,8,8a-hexahydro-2-(1H)-naphthalenone, 25: α-Cadinol, 26: Tricyclo[5.2.2.0(1,6)]undecan-3-ol, 2-methylene-6,8,8-trimethyl, 27: Tetracyclo[6.3.2.0(2,5).0(1,8)]tridecan-9-ol,4,4-dimethyl, 28: trans-Z-α-Bisabolene epoxide, 29: 15,15`-Bi-1,4,7,10,13-pentaoxacyclohexadecane, 30: 3,6,9,12,15,18,21-Heptaoxatritriacontan-1-ol, 31: 3-Ethyl-5-(2-ethylbutyl)octadecane, 32: Isodecyl octyl phthalate. GC-MS: gas chromatography–mass spectrometry, OVA: Origanum vulgare of the Armenian highlands.

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  Fig. 4 The percentage of OVA essential oil dominant components in different vegetation periods. OVA: Origanum vulgare of the Armenian highlands.

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  Fig. 5 The analgesic effects of Analgin and Diclofenac pretreatment (15 minutes) vs. formalin. The data represent mean ± standard error of mean.

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  Fig. 6 The analgesic properties of various doses of OVA essential oil (OVEO), 15 minutes before formalin injection. The data represent mean ± standard error of mean. OVA: Origanum vulgare of the Armenian highlands.

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  Fig. 7 The analgesic effect of 4% OVA essential oil 15 and 30 minutes before formalin injection. The data represent mean ± standard error of mean. OVA: Origanum vulgare of the Armenian highlands.

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  Fig. 8 Comparing the analgesic effect of 4% OVA essential oil (OVEO) with standard analgesics. The data represent mean ± standard error of mean. OVA: Origanum vulgare of the Armenian highlands.

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  Fig. 9 Significance of obtained data in the second phase of the formalin test. The data represent mean ± standard error of mean (P < 0.05). OVA: Origanum vulgare of the Armenian highlands, OVEO: OVA essential oil, ns: not significant. **P < 0.005, ***P < 0.0005, ****P < 0.0001.

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  Fig. 10 Effect of the OVA essential oil in hot plate test in mice. Control (saline, 0.1 mL, intra peritoneal [IP]) and experimental (0.1 mL, 4% OVEO, IP) groups were injected 15 minutes before the behavioral test. The data represent mean ± standard error of mean. OVA: Origanum vulgare of the Armenian highlands, OVEO: OVA essential oil, ns: not significant.

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  Fig. 11 The growth-inhibitory effect of OVA essential oil on HeLa and Vero cells viability (media with OVA essential oil preliminary addition). OVA: Origanum vulgare of the Armenian highlands. The data represent mean ± standard error of mean.

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  Fig. 12 The cytotoxic effect of OVA essential oil on HeLa and Vero cells viability (media without OVA essential oil addition). OVA: Origanum vulgare of the Armenian highlands. The data represent mean ± standard error of mean.

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  Fig. 13 Significance of cytotoxic effect of OVA essential oil. (A) The essential oil added after cell attachment. (B) The essential oil added before cell attachment (P < 0.05). The data represent mean ± standard error of mean. OVA: Origanum vulgare of the Armenian highlands, ns: not significant. ***P < 0.0005.

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