Menthol targets the transient receptor potential melastatin 8 (TRPM8) channel, a cold temperature-sensing ion channel. TRPM8 is activated by cold temperatures and cooling agents such as menthol. By modulating TRPM8, menthol exerts analgesic and anti-irritation mechanisms. Menthol also inhibits neuronal membrane Ca²⁺ currents, which contributes to its cooling sensation and local anesthetic effects. Additionally, menthol has been shown to improve oral nicotine rejection in mice. Its analgesic properties are utilized in medicinal preparations for the relief of acute and inflammatory pain, such as sports injuries and arthritis.

In vitro, Menthol exhibits antibacterial, antifungal, antipruritic, anticancer, and analgesic properties. The compound stimulates cold receptors by inhibiting neuronal membrane Ca²⁺ currents, producing a cooling sensation. As a TRPM8 modulator, menthol regulates the cold temperature-sensing ion channel to exert analgesic and anti-irritation mechanisms. The compound also functions as an effective fumigant. Its in vitro activities are concentration-dependent, with efficacy observed across a range of concentrations depending on the assay and target organism. Menthol is widely used as a flavoring agent and in topical medicinal preparations.

In vivo, Menthol produces a cooling sensation through stimulation of cold receptors. It has been shown to improve oral nicotine rejection in mice. The compound's analgesic properties are utilized in medicinal preparations for the relief of acute and inflammatory pain, such as sports injuries and arthritis. In smokers, elevated levels of the major DL-Menthol metabolite, menthol glucuronide, have been observed in urine. Notably, a significant negative correlation exists between urinary menthol glucuronide levels and NNAL-glucuronide (the major detoxification metabolite of the tobacco carcinogen NNK), suggesting that menthol may inhibit the clearance of this important tobacco carcinogen.

In vitro enzyme/receptor binding (non-cellular) assays for Menthol typically involve measuring its interaction with the TRPM8 channel. Radioligand binding assays using membrane preparations from cells expressing TRPM8 can measure displacement of labeled ligands by menthol. Alternatively, calcium flux assays using purified TRPM8 protein or membrane preparations can assess menthol-induced channel activation in a cell-free system. The compound's ability to inhibit neuronal membrane Ca²⁺ currents can be studied using lipid bilayer reconstitution or patch-clamp electrophysiology on isolated membrane patches. These assays help characterize menthol's binding affinity and mechanism of action on the TRPM8 channel.

In vitro cellular experiments with Menthol are performed using various cell lines expressing TRPM8 or neuronal cells. Cells are cultured in appropriate media and treated with varying concentrations of menthol (typically 10-1000 μM) for varying durations. TRPM8 activation is assessed by measuring intracellular calcium levels using fluorescent calcium indicators such as Fura-2 or Fluo-4. Neuronal cell membrane potential and ion currents are measured using patch-clamp electrophysiology. Cell viability and cytotoxicity are assessed using MTT or LDH release assays. For antimicrobial studies, bacterial or fungal cultures are treated with menthol, and growth inhibition is measured by optical density or colony counting. Cells are maintained at 37°C in 5% CO₂ with appropriate media supplements.

In vivo animal studies with Menthol have been conducted in various models. In mice, menthol has been shown to improve oral nicotine rejection. In smokers, urinary levels of menthol glucuronide have been studied, revealing a significant negative correlation with NNAL-glucuronide levels, suggesting that menthol may inhibit the clearance of the tobacco carcinogen NNK. For analgesic studies, rodent models of pain (e.g., formalin test, hot plate test) are used to evaluate menthol's pain-relieving effects. The compound is typically administered topically or orally. Efficacy is assessed by measuring pain responses, inflammation markers, and behavioral outcomes. Menthol's analgesic properties are well-established for relief of acute and inflammatory pain.

Absorption, Distribution and Excretion
In rabbits, the percentage of levomenthol excreted after binding with glucuronic acid depends on the dose; the higher the dose, the lower the degree of binding. /L-menthol/ Not all glucuronides are excreted via renal tubular secretion… High molecular weight conjugates, such as androsterone glucuronides… are excreted only by glomerular filtration, while low molecular weight menthol conjugates… are excreted via renal tubules in addition to glomerular filtration… /L-menthol/ Many structurally diverse substances are known to be excreted into bile; these include menthol glucuronides… /L-menthol/ Topical application can result in absorption. /DL-menthol/ For more complete data on the absorption, distribution, and excretion of menthol (7 types), please visit the HSDB records page.

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Metabolism/Metabolites In the United States, all cigarette flavorings except menthol are banned. The cooling effect of menthol may promote the absorption of tobacco toxins. We investigated the levels of tobacco exposure biomarkers in U.S. smokers, including those who smoked menthol and non-menthol cigarettes. We studied 4,603 white, African American, and Mexican American individuals aged 20 and older who participated in the National Health and Nutrition Examination Survey (NHANES) from 1999 to 2010 and had data on cigarette type and serum cotinine, cadmium, and lead levels. In this study, total 4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanol (NNAL) was measured in 1,607 participants with available measurement data. Of these, 3,210 (74.3%) smoked non-menthol cigarettes, and 1,393 (25.7%) smoked menthol cigarettes. The geometric mean concentrations of cotinine in the serum of non-menthol cigarette smokers and menthol cigarette smokers were 163.1 ng/mL and 175.9 ng/mL, respectively; the concentrations of cadmium in the serum were 0.95 μg/L and 1.02 μg/L, respectively; the concentrations of lead in the serum were 1.87 μg/dL and 1.75 μg/dL, respectively; and the concentrations of nephrine-neutralizing alcohol (NNAL) in the urine were 0.27 ng/mL and 0.23 ng/mL, respectively. After multivariate adjustment, the concentration ratios [95% confidence interval (CI)] of cotinine, cadmium, lead, and NNAL in the blood of menthol cigarette smokers and non-menthol cigarette smokers were 1.03 (0.95–1.11), 1.10 (1.04–1.16), 0.95 (0.90–1.01), and 0.81 (0.65–1.01), respectively. In a representative sample of adult smokers in the United States, current menthol cigarette use was associated with elevated blood levels of cadmium (a known carcinogen and highly toxic metal), but not with other biomarkers. These findings provide information about potential differences in exposure to toxic components between menthol cigarette smokers and non-menthol cigarette smokers. /L-Menthol/
Researchers recently noted that in Black smokers, nicotine and carcinogen exposure, measured by biomarkers such as cotinine and (4-(methylnitrosoamino)-1-(3-pyridyl)-1-butanol) (NNAL), was not associated with daily smoking volume (CPD). Researchers also noted no difference in nicotine exposure between menthol cigarette smokers and non-menthol cigarette smokers. In this study, we examined NNAL exposure in U.S. smokers by race, daily smoking volume, and menthol cigarette use. We analyzed urinary NNAL concentrations from over 1,500 daily smokers who participated in the National Health and Nutrition Examination Survey between 2007 and 2010. For comparison, we also analyzed serum cotinine concentrations in these smokers. We used linear regression analysis to estimate the mean concentration of the biomarker by daily smoking volume (CPD) and racial/ethnic group, and tested the association between biomarker concentrations and menthol cigarette use, while controlling for other demographic and smoking characteristics. Biomarker concentrations increased with increasing daily smoking volume in white, black, and Hispanic smokers, but NNAL concentrations tended to be stable at lower daily smoking volumes compared to other smokers. The mean NNAL concentrations in menthol cigarette smokers were lower than those in non-menthol cigarette smokers across all smokers (β = -0.165, p = .032) and across all white smokers (β = -0.207, p = .048). From the National Health Survey data, we found that nicotine and carcinogen exposure generally increased with increasing daily smoking volume, but NNAL exposure patterns differed across racial/ethnic groups at high daily smoking volumes. We also found that NNAL exposure differed between menthol smokers and non-menthol smokers among all smokers and among white smokers. Corynebacterium RWM1 strain was able to grow using (-)-menthol, (-)-menthone, and other acyclic monoterpenes as the sole carbon source. Growth on menthol was very slow, with a doubling time exceeding 24 hours, and growth on (-)-menthone was also not rapid (doubling time 12 hours). Growth was inhibited at concentrations exceeding 0.025% for both carbon sources. Cultures grown on (-)-menthone transiently accumulated 3,7-dimethyl-6-hydroxyoctanoate during growth, while cells grown on (-)-menthol oxidized (-)-menthol, (-)-menthone, 3,7-dimethyl-6-octanolide, and 3,7-dimethyl-6-hydroxyoctanoate. Although menthol oxidase or menthol dehydrogenase were not detected in cell extracts cultured with (-)-menthol or (-)-menthone, an inducible NADPH-coupled monooxygenase active to (-)-menthone was readily detected. In the crude cell extract, only 3,7-dimethyl-6-hydroxyoctanoate was detected as a reaction product. The oxidation of the lactone 3,7-dimethyl-6-octanoate was confirmed when (-)-menthone monooxygenase was separated from the inducible 3,7-dimethyl-6-octanolide hydrolase by hydroxyapatite chromatography. /L-Menthol/
L-Menthol was rapidly but incompletely glucuronidated. Except for one subject who had pre-administered cimetidine (1 g/day for 1 week, an oxidative drug metabolism inhibitor), all other subjects showed increased production of levmenthyl glucuronide; increased levmenthyl glucuronide production was also observed in all subjects who had pre-administered the drug-metabolizing enzyme inducer phenobarbital (60 mg once nightly for 10 days). /Levomenthyl/
For more complete data on the metabolism/metabolites of menthol (16 in total), please visit the HSDB record page.
The known human metabolites of new menthol include (2S,3S,4S,5R,6R)-3,4,5-trihydroxy-6-(5-methyl-2-propyl-2-ylcyclohexyl)oxaoxane-2-carboxylic acid.
Pharmacokinetic properties of Menthol indicate that it is well-absorbed following oral or topical administration. The compound has molecular formula C₁₀H₂₀O and molecular weight 156.26 g/mol. It is soluble in DMSO at 50 mg/mL (319.98 mM). Storage: powder at -20°C for 3 years; in solvent at -80°C for 1 year. Menthol is metabolized primarily by glucuronidation to menthol glucuronide, which is excreted in urine. The compound's density is 0.904 at 59°F. As a small lipophilic molecule (molecular weight 156.26), it is expected to have excellent tissue distribution and blood-brain barrier penetration. The half-life is relatively short due to rapid metabolism and clearance.

Toxicity Summary
Identification and Uses: Menthol is produced in crystalline or granular form. DL-menthol is used as a flavoring agent, disinfectant, and cooling agent in confectionery, liqueurs, chewing gum, toothpaste, cosmetics, cold ointments, and human medicines. L-menthol is widely used in cigarettes, cosmetics, toothpaste, chewing gum, confectionery, and pharmaceuticals. D-menthol is for research use only. Human Exposure and Toxicity: Maximum dose tests were conducted on 25 volunteers. The substance was tested at an 8% concentration in petroleum jelly without causing sensitization. Ingestion of high doses of menthol may cause abdominal pain, convulsions, nausea, vomiting, dizziness, ataxia, drowsiness, and coma. Menthol may cause anaphylactic reactions (e.g., contact dermatitis, flushing, and headache) in some individuals. In rare cases, all in children under 1 year of age, application of menthol to the nostrils or nasal cavity can cause reflex apnea. In a representative sample of US adults, menthol cigarette smoking was associated with increased all-cause mortality, cardiovascular disease mortality, and cancer mortality, with no difference compared to non-menthol cigarette smoking. In systematic reviews, menthol cigarette smoking was negatively associated with cancer risk compared to non-menthol cigarette smoking, but there is evidence that it increases the risk of cardiovascular disease. Epidemiological studies have shown no difference in nicotine addiction between smokers who smoke menthol cigarettes and those who smoke non-menthol cigarettes in the US. Animal studies: All menthol isomers studied, when applied directly to the skin undiluted, were moderately irritating to the skin. Menthol isomers were slightly irritating to the eyes. In laboratory animals, acute toxicity of menthol administered orally, by injection, and by the skin was low. Hepatic and renal changes were observed in some animals, primarily involving oral administration. Inhalation of menthol may cause respiratory damage. There is no conclusive evidence of carcinogenicity in rats and mice. In gavage studies on various animals (rats, mice, rabbits, and hamsters), L-menthol showed no embryotoxicity or fetal toxicity, nor teratogenicity, at non-maternally toxic doses. The menthol isomers were considered non-genotoxic in in vitro bacterial and mammalian assays. In vivo studies showed that L-menthol and D/L-menthol were not mutagenic in dominant lethality assays, cytogenetic assays, or mouse bone marrow micronucleus assays.

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Interactions
This study used two equations and a bilayer in vitro skin model to investigate the effect of combined use of L-menthol ((-)-menthol) and ethanol on the transdermal absorption of the model drug. Using a nonlinear least squares method, six coefficients were determined using the two equations and experimentally measured full-thickness skin permeability coefficient and full-thickness skin/carrier concentration ratio. The addition of menthol to water and 40% ethanol increased the drug diffusion coefficient in the lipid and pore channels of the stratum corneum. Adding ethanol to water and 5% menthol improves drug solubility in the carrier, reduces skin polarity, and enhances the role of pore channels in full-thickness skin penetration. The conclusion is that levonorgestrel and ethanol have a synergistic effect on in vitro transdermal drug absorption. /levonorgestrel/
Most drugs have very limited transocular penetration; therefore, finding safe and effective penetration enhancers is crucial in current ophthalmic research. This article uses a novel approach combining traditional Chinese and Western medicine to improve the corneal permeability of the water-lipid-insoluble targeted drug baicalin in vitro. Rabbits were divided into three groups. The first group was given borneol (0.05%, 0.1%), menthol (0.1%, 0.2%), or labrasol (1%, 2%) alone; the second group was given a combination of labrasol and borneol or menthol; the third group was the control group. Compared with the control group, the use of borneol, menthol, or labrasol alone significantly improved the in vitro permeability of baicalin. Furthermore, the combination of labrasol and menthol or borneol significantly enhanced its penetration effect. Among various combined penetration enhancers, the combination of 0.1% borneol and 2% larasol showed the best efficacy, with an apparent permeability approximately 16.35 times that of the control group. Furthermore, calculations of corneal hydration levels and the Draize test indicated that these penetration enhancers had good safety in rabbit corneas. This study confirms that the combined use of borneol or menthol (both derived from traditional Chinese medicine) with Larasol can improve the corneal permeability of both water-insoluble and lipid-insoluble drugs. /DL-Menthol/
Inflammation and oxidative stress are associated with a variety of pathological processes, including the development of skin tumors. Skin cancer is the most common type of cancer, leading to a considerably high morbidity and mortality rate, yet progress in its treatment remains slow. Therefore, chemoprevention and other strategies are being considered. Menthol has shown high anticancer activity against a variety of human cancers, but its effects on skin cancer have not yet been evaluated. This study investigated the chemopreventive effects of menthol on skin inflammation, oxidative stress, and carcinogenesis induced by 9,10-dimethylbenzo[a]anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA) in female ICR mice. Pretreatment with different doses of menthol significantly inhibited tumor formation and growth, and significantly reduced tumor incidence and volume. Furthermore, menthol inhibited TPA-induced skin proliferation and inflammation, and significantly suppressed cyclooxygenase-2 (COX-2) expression. Menthol pretreatment also inhibited reactive oxygen species (ROS) generation and affected the activity of various antioxidant enzymes in the skin. Menthol administration downregulated the expression of NF-κB, Erk, and p38. Therefore, inflammation and oxidative stress jointly played a key role in the chemopreventive effect of menthol on the development of skin tumors in mice. /DL-Menthol/
This study aimed to investigate the anti-apoptotic, antioxidant, and anti-inflammatory properties of menthol on ethanol-induced gastric ulcers in rats. Wistar rats were orally administered solvent, carbenolide (100 mg/kg), or menthol (50 mg/kg), followed by ethanol-induced gastric ulceration. After sacrifice, gastric tissue samples were prepared for histological sectioning and biochemical analysis. Immunohistochemical analysis was performed on heat shock protein-70 (HSP-70) and apoptosis protein Bax, which possess cytoprotective and anti-apoptotic effects. Neutrophils were manually counted. Myeloperoxidase (MPO) activity was measured. To determine antioxidant function levels, enzyme-linked immunosorbent assay (ELISA) was used to detect the levels of glutathione (GSH), glutathione peroxidase (GSH-Px), glutathione reductase (GR), and superoxide dismutase (SOD). Simultaneously, ELISA kits were used to detect the levels of pro-inflammatory cytokines tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), as well as the anti-inflammatory cytokine interleukin-10 (IL-10). Compared with the solvent control group, the gastric mucosal protection rate of the menthol-treated group reached 92%. The immunomarking area of HSP-70 increased while the immunomarking area of Bax protein decreased in the menthol-treated group. Menthol treatment reduced the activities of myeloperoxidase (MPO) and SOD, and increased the protein levels of GSH, GSH-Px, and GR. The levels of TNF-α and IL-6 also decreased, while the level of IL-10 increased. In summary, oral menthol exhibits gastric protective effects through anti-apoptotic, antioxidant, and anti-inflammatory mechanisms. /DL-Menthol/
For more complete data on menthol interactions (7 items in total), please visit the HSDB record page.

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Non-human toxicity values
Oral LD50 in rats: 2900 mg/kg /DL-menthol/
Oral LD50 in rats: 3180 mg/kg /DL-menthol/
Intramuscular LD50 in rats: 10,000 mg/kg /DL-menthol/
Oral LD50 in cats: 1500-1600 mg/kg /DL-menthol/
For more complete (16 values) data on non-human toxicity of menthol, please visit the HSDB record page.

Toxicological information for Menthol indicates that it is generally recognized as safe (GRAS) for use in food and pharmaceutical products at appropriate concentrations. The compound appears as a white solid and is derived from natural sources. At high concentrations, menthol may cause skin or mucosal irritation. It should be handled with appropriate safety precautions including use of personal protective equipment (gloves, safety goggles, lab coat) and working in a well-ventilated area. The compound's pungent odor may cause respiratory irritation in sensitive individuals. Menthol is for research use and not for human therapeutic use without proper formulation and regulatory approval. Purity: 99.88%. Storage: powder at -20°C for 3 years; in solvent at -80°C for 1 year.

[1]. Menthol: a natural analgesic compound. Neuroscience Letters. 2002 Apr.

[2]. Menthol decreases oral nicotine aversion in C57BL/6 mice through a TRPM8-dependent mechanism. Tob Control. 2016 Nov;25(Suppl 2):ii50-ii54.

[3]. Menthol attenuates respiratory irritation and elevates blood cotinine in cigarette smoke exposed mice. PLoS One. 2015 Feb 13;10(2):e0117128.

Therapeutic Uses
Antipruritic Agent
Exploring Therapies: Menthol is primarily undergoing human trials due to its pharmacological properties, such as enhancing lung capacity and airway volume.
Menthol is a natural product of the peppermint plant (Lamiaceae), a monoterpene widely used as a natural ingredient in cosmetics, a flavoring agent, and an intermediate in the production of other compounds. Various extracts of peppermint contain menthol as the main active ingredient, and it has been used for centuries as a traditional medicine to treat a variety of ailments, including infections, insomnia, and irritable bowel syndrome, as well as as an anthelmintic. /Traditional Medicine//
Veterinary Uses: Its vapor has been used clinically to treat respiratory diseases in horses, pigs, and poultry. …When administered parenterally, it is used in irritant expectorant mixtures…
For more complete data on the therapeutic uses of menthol (6 in total), please visit the HSDB records page.

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Drug Warning
Studies have found that L-menthol has a superior "cooling" effect compared to other isomers; L-menthol also has a superior smell and taste, while some isomers produce a sharp, irritating, and unpleasant sensation. /L-menthol/
Neonatal glucose-6-phosphate dehydrogenase deficiency can cause severe jaundice after taking menthol because newborns cannot bind menthol.
There have been reports of allergic reactions to the use of menthol products (including cigarettes). Individual cases of laryngospasm have been reported in infants using menthol nasal drops, and a few cases of neurological or digestive disorders have been associated with excessive inhalation or oral administration of menthol.
Menthol…can cause allergic reactions in some people (such as contact dermatitis, flushing, and headache). Applying menthol-containing ointments inside an infant's nostrils to treat cold symptoms can cause the infant to faint immediately.
Veterinarians: Overdose can lead to seizures and eventually even death.
Menthol (CAS 1490-04-6) is a naturally occurring cyclic monoterpene alcohol obtained from peppermint or mint oils. The compound has molecular formula C₁₀H₂₀O and molecular weight 156.26 g/mol. Menthol has flavoring and local anesthetic properties and produces a cooling sensation by stimulating cold receptors through inhibition of neuronal membrane Ca²⁺ currents. The compound is a TRPM8 modulator and exhibits antibacterial, antifungal, antipruritic, anticancer, and analgesic properties. The naturally predominant stereoisomer is (-)-Menthol (levomenthol, L-menthol). Also known as DL-Menthol and 薄荷脑. Purity: 99.88%. Storage: powder at -20°C for 3 years; in solvent at -80°C for 1 year.

C10H20O

156.2652

156.151

1490-04-6

Menthol-d4;1217765-02-0

1254

White to off-white solid powder

0.9±0.1 g/cm3

215.4±8.0 °C at 760 mmHg

34-36 ℃(lit.)

93.3±0.0 °C

0.0±0.9 mmHg at 25°C

1.457

3.2

1

1

1

11

120

0

NOOLISFMXDJSKH-UHFFFAOYSA-N

InChI=1S/C10H20O/c1-7(2)9-5-4-8(3)6-10(9)11/h7-11H,4-6H2,1-3H3

5-methyl-2-propan-2-ylcyclohexan-1-ol

2934.99.9001

Powder      -20°C    3 years

                     4°C     2 years

In solvent   -80°C    6 months

                  -20°C    1 month

Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

DMSO : ~250 mg/mL (~1599.80 mM)

Solubility in Formulation 1: ≥ 2.08 mg/mL (13.31 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL.
Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.

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Solubility in Formulation 2: ≥ 2.08 mg/mL (13.31 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution.

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Solubility in Formulation 3: ≥ 2.08 mg/mL (13.31 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.

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 (Please use freshly prepared in vivo formulations for optimal results.)