Table 3.

Summary of the pharmacological properties of Thymus vulgaris L.

Pharmacological Activity	Type of Study	Models used	Plant part/material	Type of extract/compound	Doses used	Controls	Possible/reported mechanisms	Results	References
Antibacterial activity	in vitro	Escherichia coli, Klebsiella pneumoniae, Yersinia enterocolitica, Staphylococcus aureus, Listeria monocytogenes, Enterococcus faecalis	Aerial parts	Ethanol	100 μl of 6.25–0.025% serial dilutions	Solvent ethanol	Not defined	MIC was observed as >6.25, 3.12,>6.25, 0.2, 0.39, 0.78 mm respectively at 100 μl concentration (p < 0.05). These bacteria showed mixed response towards antibiotics.	[53]
	in vivo	Methicillin-resistant Staphylococcus aureus (MRSA) in mice	Not defined	Methanol	200 mg/ml per kg of body weight	Positive control-infected, negative control-normal mice, Antibiotics	Not defined	For the bacteria isolated from throat and lungs, MIC was found to be 2.93 and 3.83 CFU (log10)/ml, respectively (p < 0.05).	[54]
	in vitro	Salmonella typhirium	Aerial parts	Essential oil	100 μl of 1/20–1/200 v/v serial dilutions	Amoxicillin, cefotaxime	Not defined	MIC of 25.5 mm at 100 μl concentration, in accordance with amoxicillin (23.0 mm) and cefotaxime (15.0 mm) (p < 0.05).	[55]
	in vitro	Salmonella Enteritidis Biofilm	Dried plant	Essential oil	5–0.0024 μl/ml.	Growth control (broth + microbe), negative control (broth + propylene glycol + microbe), sterility control (broth + test oil), positive control (broth + streptomycin + microbe)	Possible inhibition of bacterial adsorption and biofilm matrix formation	Biofilm inhibition at MIC/MBC 0.156/0.315 μl/ml by oil, thymol, and carvacrol. Oil reduced the metabolic activity by 9.6–70.5%, (p < 0.05).	[13]
	in vitro	Staphylococcus aureus (MDR)	Not defined	Essential oil	10 μl of 2.87–11.5 μg/ml	Cefotaxime	Not defined	A significant inhibition with 35–40 mm inhibition zone at 2.8–11.5 at μg/ml was observed for MDR variants, whereas cefotaxime showed MIC at 32 μg/mL concentration (p < 0.05)	[11]
	in vitro	Pseudomonas aeruginosa	Leaves and branches	Essential oil	100 μl oil of different concentrations	Not defined	Not defined	Minimum bactericidal concentration (MBC) was found to be 8% at 100 μl concentration	[56]
	in vitro	Actinobacillus pleuropneumoniae, Streptococcus suis, Actinobacillussuis, Haemophilus parasuis, Pasteurella multocida, and Bordetella bronchiseptica	Not defined	Essential oil	100 μl of 1.25 to 0.01% (v/v)	Media + microbes + PBS	Not defined	MIC values ranging from 0.039% to 0.078% at dilutions 0.01–1.25% v/v (p < 0.05)	[57]
Antioxidant activity	in vivo	Antioxidant enzyme levels in rabbits	Not defined	Aqueous extract	50 mg/kg of body weight	Water	Not defined	Levels of antioxidant enzymes catalase, glutathione, glutathione-S-transferase, and superoxide dismutase increased by 14.12%, 27.69%, 98.75% and 78.29%, respectively (p < 0.05)	[58]
	in vivo	Antioxidant enzyme levels in rats	Dried leaves	Aqueous extract	500 mg/kg body weight	Paracetamol (200 mg/kg)	Not defined	Alanine aminotransferase, aspartate aminotransferase, and alkaline phosphatase content increased by 2 units/mL. In combination with paracetamol, the enzyme levels increased by 15–20 units/ml (p < 0.01)	[59]
	in vitro	Radical scavenging activity using DPPH	Leaf and stem	Aqueous extract	0.0125–3.0 mg/ml	Not defined	Polysaccharide biding with BSA brings out the radical scavenging	Radical free scavenging activity of 92.0% at the concentration of 1.5 mg/mL, in accordance with butylated hydroxyanisole (BHA-95.7%) and butylated hydroxytoluene (BHT-96.6%).	[52]
	in vitro	FRAP, ABTS, and FIC	Not defined	Essential oil	0.23–30 mg/ml	Not defined	Not defined	FRAP, FIC, ABTS assays showed IC50 values 12.69, 13.29, and 6.46 mg/ml respectively (p < 0.05)	[60]
	in vitro	Primary and secondary lipid oxidation products in oil in water (O/W) emulsions through DPPH	Dry waste plant	Ethanolic extract	100 μl of different concentrations	Not defined	Not defined	IC50 of 93 μg/ml compared to BHT (89 μg/mL) (p > 0.05)	[61]
Antifungal activity	in vitro	Sclerotinia sclerotiorum, Botrytis cinerea, Phytophthora parasitica, Pythium aphanidermatum, Fusarium oxysporum, Alternaria brassicae, Trichoderma aggressivumf.sp. europaeum, Cladobotryum mycophilum	Not defined	Essential oil	5,10,15,20,30% (v/v)	Media + Tween 20	Not defined	Mycelial growth inhibition values were found to be ranging between 13.9 to 41.4 mm at 5% concentration with ED50 values ranging from 9.3-18.0% for all the species (p < 0.05)	[14]
	in vitro	Clinical isolates of Candida albicans and C. glabrata species	Not defined	Essential oil	0.005–2.5% (v/v)	Amphotericin B	Possible ergosterol binding	MIC and MFC were in the range of 0.04–22.9 mg/ml for all the isolates. Thyme oil reduced the fungal growth in the initial hours (4–8). It inhibited the growth with sorbitol at lower MIC (0.08 mg/ml) (p < 0.05)	[15]
	in vivo	C. albicans in a Caenorhabditis elegans nematode model	Not defined	Thymol	32, 64, and 128 mg/l	Kanamycin (45 μg/ml), ampicillin (100 μg/mL), and streptomycin (100 μg/ml)	Enhancing pmk-1and sec-1 gene expressions, which in turn enhance p38 MAPK signalling pathway	Complete inhibition of fungi and biofilm at 64 mg/l and 128 mg/ml, compared to control used (p < 0.05). Growth reduction at 12 h, compared to control (36 h). Thymol enhances the expressions of pmk-1and sec-1 genes, in turn p38 MAPK signalling pathway	[62]
	in vitro	Cryptococcus neoformans	Not defined	Essential oil	0.07–10 mg/ml	FLC (0.06–128 μg/mL), ITC (0.0078–2 μg/ml), VRC (0.0078–32 μg/mL)	Possible membrane deterioration by thymol, Possible ergosterol binding	MIC and MFC were found to be in 0.56–1.12 mg/ml in accordance with controls. Thymol showed better activity 0.02–0.08 mg/ml (p < 0.05)	[63]
	in vitro	Aspergillus flavus	Not defined	Essential oil vapour and liquid phases	0, 1, 5, 10, and 20 μg/ml	Aflatoxin B	Downregulating of fungal development genes brlA, abaA, wetA and aflatoxin biosynthesis genes aflR, aflD, and aflK	Vapor and liquid phases reduced growth at 20 and 400 μg/ml, respectively. Thyme oil 10 μg/mL of reduced production of afltoxin by 97.0 and 56.4% through vapour and liquid phases, respectively. (p < 0.05).	[16]
Anti-inflammatory activity	in vitro	NO radical scavenging in murine macrophage cell line J774A.1	Flowering tops	Aqueous extract	8.5, 16, 50.4, 84 μg/ml	Dexamethasone	Possible cellular mechanisms of suppression of iNOS induction by flavonoids	Significant scavenging of NO radicals with 80.3% of the activity at 16 μg/ml concentration was observed in accordance with control (p < 0.05)	[19]
	in vitro	5-lipoxygenease (5-LOX) production, lipopolysaccharide (LPS) induced TNF-α, IL-1β, and IL-8 secretions using THP-1 cells	Dried plant	Essential oil	30 μl of different concentrations	α-bisabolol	Not defined	5-LOX got inhibited at 0.005 μg/ml (IC50) of thyme oil, compared to α-bisabolol (0.049 μg/mL). TNF-α, IL-1β, and IL-8 got inhibited at 0.01 μg/ml.	[20]
	in vivo	Mice with carrageenan-induced paw edema	Aerial parts and dried leaves	Essential oil	100, 200 and 400 mg/kg	Tween 80 and diclofenac	Not defined	Paw thickness was found reducing at a dose of 400 mg/kg. Results were in accordance with both the controls Tween 80 and diclofenac (p < 0.001). Toxic level of thyme oil was found (4500 mg/kg), where sedation was observed at 5000 mg/kg.	[43]
	in vivo	Mice with carrageenan-induced pleurisy	Leaves	Essential oil	250, 500 and 750 mg/kg	Croton oil	Carvacrol may act by inhibiting cytokines and leukotrienes, and these mediators are likely not involved in the mechanism of action of thymol	All the concentrations reduced inflammatory exudates as well as migrated leucocytes in ear edema. Individual assessment showed thymol (34.2%) and carvacrol (47.3%) are attributable for the anti-inflammatory activity (p < 0.05)	[64]
Anti-cancerous activity	in vitro	MCF7 (breast adenocarcinoma), HCT15 (colon carcinoma), HeLa (cervical carcinoma), HepG2 (hepatocellular carcinoma), and NCI-H460 (non-small cell lung cancer) cell lines	Dried aerial parts	Essential oil	10–100 μg/ml	Ellipticine (0.24–65.2 μg/ml)	Possible involvement of thymol in the stimulation of active proliferation of pulp fibroblasts	T. vulgaris L. oil showed inhibition of growth at 76.02–180.40 μg/ml concentration (GI50). It did not show any effect on non-tumour liver PLP cells, even at a high concentration of 400 μg/ml (p < 0.05)	[65]
	in vitro	THP-1 leukemia cell line	Not defined	Essential oil	10–500 μg/ml	DMSO	Not defined	At a concentration of 100 μg/ml and >200 μg/ml, thyme oil prevented the proliferation of THP-1 leukemia cells	[66]
	in vitro	H460 lung cancer cell line	Not defined	Hydroalcoholic extract	0.04–0.6%	Glyceraldehyde 3-phosphate dehydrogenase	Possible interference in pro-inflammatory cytokines	H460 lung cancer cell line was found to be sensitive at 0.11% of hydroalcoholic extract (p < 0.05) and downregulated NF-κB p65 and NF-κB p52 proteins along with the reduction of IL-1β and IL-8 gene expression in LPS model	[21]
	in vivo	Mammary carcinoma rat and 4T1 mouse models	Dried plant	Thyme powder	50 mg/kg body weight	Untreated models	Possible interference with pro-inflammatory cytokines, Possible upregulation of caspase genes at epigenetic level	Thyme powder reduced the volume of 4T1 tumours by 85% at 1% concentration. In rat model, the same concentration decreased the tumour frequency by 53% (p < 0.05). Upregulation of caspase-2 and caspase-3 enzymes, along with bcl-2 and Bax proteins	[67]
	in vitro	HL-60 acute promyelotic leukemia cell line, human peripheral blood mononuclear cell (PBMC)	Not defined	Thymol	5, 25, 50, 75 and 100 μM for 24 h	Camptothecin (5 μM)	Apoptosis induced by thymol in HL-60 cells was associated with ROS production, increase in mitochondrial H2O2 production, decrease in Bcl-2 protein, increase in Bax protein levels, enhancing apoptosis inducing factor (AIF) in mitochondria and caspase activation	Thymol showed no cytotoxic effect on human peripheral blood mononuclear cell (PBMC) at 5 and 25μM concentrations. However, extensive cytotoxicity was observed at >50 μM, after 24 h	[68]
	in vitro	Synthesized silver nanoparticles against T47D human breast cancer cells	Dried leaves	Silver nanoparticles and ethanol extract	12.5–200 μg/ml	Untreated cells	Nanoparticles could trig- ger translocation of phosphatidylserine (PS) from the inner membrane indicating apoptosis pathway rather than necrosis	T47D cells showed high sensitivity towards nanoparticles (90%) compared to the extract (75%). T47D cells treated with nanoparticles showed 18.40% early and 0.69% late apoptosis with varying IC50 concentrations (12.5–100 μg/mL). Same was observed in case of plant extract, where 15.67% early and 1.70% late apoptosis was found (p < 0.05)	[47]
Antiviral activity	in vitro	Influenza virus	Not defined	Essential oil vapour and liquid phases	3.12–100 μl/ml	Canova oil	Possible interaction with hemagglutinin (HA)	Liquid phase at 3.1 μl/ml concentration completely inhibited the viral growth, which was better than that of control used (canola oil). Significant inhibition of HA was observed. Also, 50% of the culture was reduced depicted as TC50 14.34 μl/ml (p < 0.05)	[33]
	in vitro	Herpes simplex virus (HSV) on RC-37 (African green monkey kidney cells)	Not defined	Essential oil	10–750 μg/ml	Untreated cells	Not defined	Cytotoxicity ranged between 20 μg/ml for citral and 1250 μg/ml for 1,8-cineole. IC50 values for1,8-cineole was 1200 μg/ml. Thyme oil proved to reduce the viral load by >96%, whereas all monoterpenes by >80% (p < 0.05)	[69]
	in vitro	HIV-1 in HeLa HL3T1 cell line	Not defined	Essential oil	7.5–240 μg/ml	Neomycin, cisplatin	Possible alteration in the structure of Tat/TAR-RNA complex	EMSA showed a notable inhibitory potential of oil (3–6 μg/ml), compared to the control in case of Tat/TAR-RNA complex inhibition. Reduction activity test against Tat-induced HIV-1 LTR transcription resulted in RT50 = 0,83 μg/ml, a notable inhibitory potential which reduced viral transcription to 52% (p < 0.05)	[70]
	in vitro	HIV-1 subtype A in PBMC cell line	Dried plant	Methanol extract	10, 100, 200, 800 and 1600 μg/ml	DMSO, Zidovudine	Not defined	The cytotoxicity value (CC50) on PBMC was found to be 200 μg/ml. Antiviral assay revealed EC50 value of >500 μg/ml. Mean fluorescent intensity (MFI) of the CD4+ expressions were found to be 22.72 in PBMC (p < 0.05)	[71]
Antidiabetic activity	in vitro	Inhibition of α-glucosidase and α-amylase enzymes	Not defined	Aqueous, methanol and ethanol extracts	4, 8, 15, and 20 μg/ml	Acarbose	Not defined	Methanol extract resulted in maximum inhibition of α-glucosidase (IC50 4.35, 22.04, 30.77, 43.13), though less compared to Acarbose (IC50 16.11, 44.6, 53.03, 63.70). Similarly, α-amylase got reduced maximally by the same extract (IC50 6.39, 11.47, 17.01, 22.93), less compared to Acarbose (IC50 12.37, 25.16, 36.08, 44.97)	[72]
Anxiolytic activity	in vivo	Elevated plus-maze (EPM) rat model	Dried plant	Aqueous extract	50 mg/kg, 100 mg/kg, and 200 mg/kg	Saline fed groups	Possible relation with antioxidant activity of phytochemicals	The aqueous extract exhibited a significant increase in rat movement into the open arms at 100 mg/kg (p < 0.05) and 200 mg/kg (p < 0.01).	[74]
UV-protective activity	in vitro	Human skin cells	Not defined	Aqueous extract, thymol	1.82 μg/ml extract and 1 μg/ml thymol	Normal cells without UV treatment, but with extract treatment	Reduction of ROS induced DNA damage, Possible involvement of polyphenols in protectivity	Aqueous extract of thyme leaf (1.82 μg/ml) and thymol (1 μg/ml) reduced the release lactic acid dehydrogenase (LDH), in cultured skin cells treated with UV rays. Cell proliferation was observed in thyme pre-treated skin cells in accordance with control, along with the reduction in DNA damage (p < 0.01)	[75]
Anthelminthic activity	in vitro	Eimeria spp. oocysts from Turkey fowls	Not defined	Essential oil	0, 1, 2, 4, 8, 10, 20, 40, 80, and 800 mg/ml	Ammonia and diclazuril	Not defined	Thyme oil showed significant anti-helminthic activity against 4 species of Eimeria spp. at IC50 53.42 mg/ml for 5×104 oocysts (p < 0.05)	[40]
Anti-anti-alzheimer's activity	in vivo	Acetylcholine esterase and nicotinic acetylcholine receptor in C. elegans nematode model	Aerial parts and leaves	Essential oil	10, 20, 40, 60, 80, and 100 ppm	10% DMSO	Upregulation of unc-17, unc-50, and cho-1 genes by ρ-Cymene	Enhancement of the nicotinic acetylcholine receptor activity, upregulation of unc-17, unc-50, and cho-1 genes at 40 and 60 ppm ρ-Cymene was attributed for gene upregulation activity along with downregulating ace-1 and ace-2 at 20 and 100 ppm (p < 0.05). Thymol and γ-terpinene enhanced synaptic acetylcholine levels in combination (40 ppm)	[76]
Anti-osteoporotic activity	in vivo	Rat model with low calcium intake	Dried leaves	Leaf powder	5% w/w	Standard diet + normal calcium (Ca 0.5% w/w), standard diet + low calcium (Ca 0.1% w/w), Thyme powder (5% w/w) + low calcium (Ca 0.1% w/w)	Possible promotion of calcium resorption in the gut	Significant increase in the bone mass (2.93 g/kg), length 32.8 mm), and density (0.13 g/cm2), compared to low calcium diet control (2.46 g/kg, 32.2 mm, and 0.09 g/cm2, respectively) (p < 0.05)	[77]
Anti-pulpotomy activity	in vivo	Formocresolpulpotomy in humans	Not defined	Ethanolic extract	Suitable consistency	Formocresol	Not defined	Thyme ethanolic extract along with zinc oxide reduced pain and tenderness, Enhanced bone and root resorption. Clinical and radiographic evaluations showed 94.4% and 88.2% success, respectively with no statistical significance compared to the control, formocresol 88.2% (p > 0.05)	[78]