Skatole targets the aryl hydrocarbon receptor (AhR) and p38 MAPK. It acts as a potent endogenous ligand and agonist of AhR in vitro and activates the murine AhR in vivo. Skatole also targets p38 MAPK, inducing apoptosis of intestinal epithelial cells through activating these pathways. Additionally, it has been associated with alkylation of a 57 kDa protein target. Skatole exhibits multiple bioactivities targeting various pathways: it shows inhibitory activity against AmpC Beta-Lactamase (potency 6309.6 nM), acts as a delayed death inhibitor against the malarial parasite plastid (14715.7 nM), inhibits cell surface uPA generation (2818.4 nM), and blocks Ebola virus entry. Its effects on intestinal epithelial cells are mediated through AhR and p38 activation.

Skatole (0-1000 µM, 0-72 h) leads to intestinal epithelial cell apoptosis in a time and dose dependent manner. At 1000 µM, it significantly activates AhR (48 h) and p38 (30 min). At 10 µM for 24 h, skatole significantly reverses the suppression of irisin on Ang II-evoked proliferation, migration, and phenotypic transformation in vascular smooth muscle cells (VSMCs). Skatole exhibits antimicrobial, anti-malarial, anti-cancer, and antiviral properties in various in vitro models. It inhibits cell surface uPA generation (2818.4 nM) and induces DNA re-replication in SW480 colon adenocarcinoma cells. Skatole blocks Ebola virus entry with AC50 values ranging from 0.631 µM to 631.0 nM and shows anti-SARS-CoV-2 activity with IC50 values greater than 20000.0 nM.

In vivo, skatole (3-methylindole) causes pulmonary edema in goats, sheep, rats, and some strains of mice. Response to 3-methylindole varies among species, with mice recovering from 3MI-induced bronchiolar damage with scarring and epithelial metaplasia. Skatole activates the murine AhR in vivo. It is produced by intestinal bacteria and regulates intestinal epithelial cellular functions through activating AhR and p38. Caused no irritation in humans following a 24-hour closed patch test (2% in petrolatum). Lethal doses by intravenous and oral routes produced pulmonary lesions in cattle. Safe when used as a flavoring agent in food according to JECFA.

For in vitro cell-free assays, skatole's interaction with AhR can be studied using radioligand binding or reporter gene assays. p38 MAPK activation can be measured by assessing phosphorylation levels using specific antibodies in Western blot or ELISA-based kinase activity assays. Its inhibitory activity against AmpC Beta-Lactamase can be assessed using chromogenic substrates, measuring the reduction in enzyme activity at varying compound concentrations to determine IC50 values. For Ebola virus entry inhibition, pseudotyped virus entry assays can be employed. For cytotoxicity studies, cell viability can be measured using MTT or similar assays.

For in vitro cellular assays, skatole is typically tested in intestinal epithelial cell lines. Cells are seeded in multi-well plates and treated with skatole at concentrations ranging from 0-1000 µM for 0-72 hours. Apoptosis is assessed using flow cytometry (Annexin V/PI staining) or caspase activity assays. AhR activation is measured after 48 hours of treatment by assessing nuclear translocation or target gene expression. p38 MAPK activation is measured after 30 minutes of treatment by Western blot using phospho-specific antibodies. For VSMC studies, cells are treated with 10 µM skatole for 24 hours to assess proliferation and migration.

For in vivo studies, skatole is typically administered to animal models via oral or intravenous routes. In rodents, it causes pulmonary edema. For AhR activation studies, skatole can be administered to mice, and AhR target gene expression in tissues can be measured. In pulmonary toxicity studies, endpoints include histological examination of lung tissue for edema and epithelial changes. In cattle, lethal doses produce pulmonary lesions. For safety evaluation, skatole can be tested in closed patch tests in humans (2% in petrolatum).

Absorption, Distribution and Excretion
Following intragastric injection of 0.1–0.2 g/kg skatole in cattle or intravenous infusion of 0.06 g/kg skatole, the mean plasma concentration of skatole reached its maximum at 3 hours and 9 hours, respectively. In goats, intravenous infusion of 3-methylindole (3MI) containing (14)C-3MI over 2 hours, with propylene glycol as the solvent, rapidly cleared 3MI from plasma and tissues after infusion, with 81% of the radioactive material excreted in urine within 24 hours. The maximum concentration of unmetabolized 3MI in tissues ranged from 2.6 to 15 μg 3MI/g, with a concentration of 7.5 μg 3MI/g in lung tissue. The proportion of metabolites was highest in lung tissue. The data indicate that 3-methylindole (3MI) does not selectively accumulate in the lungs, and its concentration is lower than that typically associated with direct membrane damage.

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Metabolism/Metabolites
Skatol is produced by bacteria in the gastrointestinal tract (small intestine and rumen) that degrade dietary tryptophan residues…
Adult beef cattle grazing in dry pastures during the summer were transferred to lush pastures to induce acute bovine pulmonary edema and emphysema (ABPE), and to determine whether the concentration of 3-methylindole (3MI) in plasma and rumen fluid was related to… the development of ABPE. In vitro production of 3MI was observed in a culture medium inoculated with rumen fluid, indicating the presence of 3MI-producing microorganisms in the rumen. Clearly, bovine rumen microorganisms convert tryptophan (present in lush pastures) into 3MI, and the absorption of 3MI by animals may lead to the development of ABPE.
Produced by indole-3-acetic acid. Produced in rats and wheat; FRYDMAN RB et al.; FEBS LETTERS 17: 273 (1971). Production of 5-hydroxypyrethrin and 7-hydroxypyrethrin in rats; DALGLIESH CE et al.; BIOCHEM J 70: 13P (1958). Production of 6-hydroxypyrethrin in rabbits; JEPSON JB et al.; BIOCHIM BIOPHYS ACTA 62: 91 (1962). Production of salicylic acid in Pseudomonas; PROCTOR MM; NATURE (LONDON) 181: 1345 (1958). /Excerpt from Table/
Goat jugular vein infusion (14) C-3-methylindole (3MI). The major metabolic pathways of 3-methylindole involve its production, suggesting that a mixed-function oxidase (pyrrole oxygenase) may be the major metabolic system. The minor metabolic pathways involve the oxidation of the methyl carbon of 3-methylindole.
For more metabolic/metabolite (complete) data on 3-methylindole (6 metabolites in total), please visit the HSDB record page.
The known metabolites of 3-methylindole include 3-methylindole-2,3-epoxide and 3-methyleneindoline.
Skatole (CAS 83-34-1) has a molecular formula of C9H9N and a molecular weight of 131.17-131.18. It is a white to light brown solid with a strong fecal odor. Purity: 99.92%. Solubility: DMSO 100 mg/mL (762.37 mM). For in vivo formulation: 10% DMSO >> 40% PEG300 >> 5% Tween-80 >> 45% saline, solubility ≥ 2.5 mg/mL (19.06 mM). Melting point: 95°C; boiling point: 265-266°C. Soluble in hot water, alcohol, benzene, chloroform, ether. Storage: powder -20°C for 3 years, 4°C for 2 years; in solvent -80°C for 6 months, -20°C for 1 month.

Density: 1.11 g/cm3. Acute toxicity: rat oral LD50: 3450 mg/kg; mouse oral LD50 data available. Skatole is considered mildly toxic. Minimum lethal dose (frog, subcutaneous): 1.0 g/kg. It is a mildly toxic compound that causes pulmonary edema in various animal species. No safety concern at current levels of intake when used as a flavouring agent according to JECFA. It can cause irritation; safe when used as a flavoring agent in food.

Skatole regulates intestinal epithelial cellular functions through activating aryl hydrocarbon receptors and p38. Biochem Biophys Res Commun. 2019 Mar 19;510(4):649-655.

Skatole is a methylindole with a methyl substituent at the 3-position. It is produced during the anaerobic metabolism of L-tryptophan in the digestive tract of mammals. It is a metabolite in both mammals and humans. 3-Methylindole has been reported in Tachigali glauca, Tecoma stans, and other organisms with relevant data. See also: ... See more ...

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Mechanism of Action
Nucleophilic thiols, such as glutathione, L-cysteine, and N-acetyl-L-cysteine, can protect microsomal proteins from the alkylation of the active metabolite of 3-methylindole. Bovine lung cytosol components can enhance the protective effect of these thiols. Pretreatment of sheep with diethyl maleate depletes glutathione, thereby exacerbating the pulmonary toxicity of 3-methylindole; while pretreatment with L-cysteine reduces the severity of this effect. These findings are consistent with the hypothesis that the electrophilic metabolite of 3-methylindole is responsible for its pulmonary toxicity and suggest that glutathione and glutathione S-transferases are involved in the detoxification process of this active metabolite. The results of incubation of various indole compounds with goat lung microsomes showed that only 3-methylindole could generate free radicals in an NADPH-dependent microsomal system, which was confirmed by spin trapping experiments. The enzymatic free radical generation of 3-methylindole suggests that the free radical mechanism of microsomal activation may be a specific mechanism for 3-methylindole-induced pulmonary toxicity. This study investigated the bioactivation of 3-methylindole (3MI) in human lung and liver tissues. 3MI is a highly selective pulmonary toxin for goats, and this study aimed to understand human susceptibility to 3MI toxicity. Human lung microsomes were prepared from eight organ transplant donors, and liver microsomes were selected from one of the donors. The turnover rate of 3MI in human lung microsomes was 0.23 ± 0.06 nmol/mg/min, lower than that in human liver microsomes (7.40 nmol/mg/min). These activities were NADPH-dependent and inhibited by the potent cytochrome P450 suicide substrate inhibitor, l-aminobenzotriazole. The covalent binding of 3MI reaction intermediates to human tissues was determined by incubating (14)C-3MI and NADPH with human lung and liver microsomal proteins. While human lung microsomes showed measurable covalent binding activity (2.74 ± 2.57 pmol/mg/min), the intensity of the reaction was only 4% of that in human liver microsomes and was also inhibited by l-aminobenzotriazole. Therefore, the bioactivation of 3MI to covalently bound intermediates is catalyzed by cytochrome P450 in human lung tissue. These activities were compared with those measured in goat tissue. Proteins from goat and human lung and liver microsomal incubation solutions were incubated with radiolabeled 3MI. The radiolabeled proteins were then analyzed by SDS-PAGE and HPLC, respectively, and developed by autoradiography and radiochromatography. The results showed that the 57 kDa protein was the most significant alkylation target associated with 3MI reaction intermediates. These data suggest that humans may be susceptible to 3MI-mediated toxicity, and the specificity of covalent binding and the degree of binding to target proteins may play an important role in the selective susceptibility of organs and species to 3MI pulmonary toxicity.

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Therapeutic Use
Experimental Use: Drug (Veterinary): 5 mg skatole/day intramuscularly for 15 consecutive days in infected guinea pigs weighing 200-300 g showed anti-tuberculosis activity against Mycobacterium tuberculosis.

Skatole is a research-grade compound not approved for therapeutic use. It serves primarily as a pharmacological tool for studying AhR and p38 MAPK signaling, apoptosis, and intestinal epithelial cell function. Its mechanism involves activation of AhR and p38 MAPK pathways leading to apoptosis. Skatole has been used in specific products of the perfume industry and as a flavor additive in ice cream. No clinical trials have been reported. It is an endogenous metabolite produced by intestinal bacteria from tryptophan. Its bioactivities include antimicrobial, anti-malarial, anti-cancer, and antiviral properties.

C9H9N

131.17446

131.073

83-34-1

Skatole-d3;111399-60-1;Skatole-d8;697807-03-7

6736

Off-white to gray solid powder

1.1±0.1 g/cm3

265.1±9.0 °C at 760 mmHg

92-97 °C(lit.)

112.5±11.3 °C

0.0±0.5 mmHg at 25°C

1.655

2.6

1

0

0

10

122

0

ZFRKQXVRDFCRJG-UHFFFAOYSA-N

InChI=1S/C9H9N/c1-7-6-10-9-5-3-2-4-8(7)9/h2-6,10H,1H3

3-methyl-1H-indole

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 : ~100 mg/mL (~762.37 mM)

Solubility in Formulation 1: ≥ 2.5 mg/mL (19.06 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 25.0 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: ≥ 2.5 mg/mL (19.06 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 25.0 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: ≥ 2.5 mg/mL (19.06 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 25.0 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.)