1,2,4-Trimethoxybenzene
Based on 1 Customer Validation
1,2,4-Trimethoxybenzene is an orally active NLRP3 selective inhibitor. 1,2,4-Trimethoxybenzene can markedly suppress Nigericin (HY-127019) or ATP (HY-B2176)-induced NLRP3 inflammasome activation, thus decreasing caspase-1 activation and IL-1β secretion. 1,2,4-Trimethoxybenzene specifically inhibits the activation of NLRP3 inflammasome without affecting absent in melanoma 2 (AIM2) inflammasome activation. 1,2,4-Trimethoxybenzene inhibits oligomerization of the apoptosis-associated speck-like protein containing a CARD (ASC) and protein-protein interaction between NLRP3 and ASC, thus blocking NLRP3 inflammasome assembly. 1,2,4-Trimethoxybenzene can be used for the study of experimental autoimmune encephalomyelitis (EAE), multiple sclerosis, and type 2 diabetes.
For research use only. We do not sell to patients.
- Purity : 97.0%
- CAS No.: 135-77-3
- Formula: C9H12O3
- Molecular Weight:168.19
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Storage:
Store at room temperature 3 years.
In solvent -80°C, 2 years , -20°C, 1 year
To place orders, for customer services and technical support, please contact: MedChemExpress USA
Tel: 609-228-6898 E-mail: [email protected] [email protected]
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Biological Activity
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Chemical Information
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Solvent & Solubility
- Protocols
- Purity & Documentation
- References
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Help & FAQs
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Biological Activity
Description
IC50 & Target
[1]|
NLRP3 |
Caspase-1 |
IL-1β |
Cellular Effect
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Cell Line
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Type | Value | Description | References |
|---|---|---|---|---|
| HepG2 | IC50 |
>10 μM
Compound: 1
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Cytotoxicity against human HepG2 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
Cytotoxicity against human HepG2 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
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[PMID: 31403289] |
| HL-60 | IC50 |
>10 μM
Compound: 1
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Cytotoxicity against human HL60 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
Cytotoxicity against human HL60 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
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[PMID: 31403289] |
| MCF7 | IC50 |
>10 μM
Compound: 1
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Cytotoxicity against human MCF7 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
Cytotoxicity against human MCF7 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
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[PMID: 31403289] |
| MRC5 | IC50 |
>10 μM
Compound: 1
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Cytotoxicity against human MRC5 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
Cytotoxicity against human MRC5 cells assessed as cell growth inhibition after 72 hrs by alamar blue assay
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[PMID: 31403289] |
In Vitro
1,2,4-Trimethoxybenzene (0.5-1 mM, 1.5 h) significantly inhibits Nigericin (HY-127019) and Lipopolysaccharides (HY-D1056) (LPS)-induced NLRP3 inflammasome activation in immortalized murine bone marrow-derived macrophages (iBMDMs), manifested as reduced caspase-1 (Casp-1) cleavage and IL-1β secretion[1].
1,2,4-Trimethoxybenzene (1 mM, 75-90 min) inhibits LPS and ATP-induced NLRP3 activation in iBMDMs, primary microglia, and Primary macrophages cells[1].
1,2,4-Trimethoxybenzene (75-90 min) inhibits nigericin-induced NLRP3 activation in iBMDMs and primary macrophages, while 1,2,3-TTB had no inhibitory effect, indicating that the inhibitory activity is structure-dependent[1].
1,2,4-Trimethoxybenzene (45 min-2 h) does not inhibit LPS and poly(dA:dT)-induced activation of AIM2 inflammasome in primary peritoneal macrophages and iBMDMs[1].
1,2,4-Trimethoxybenzene (1 h) reduces the formation of ASC specks in iBMDMs and Primary microglia induced by LPS and nigericin: immunofluorescence showed a significant reduction in the number of ASC specks; DSS crosslinking experiments showed an increase in soluble ASCs and a decrease in insoluble ASCs[1].
1,2,4-Trimethoxybenzene (1 h) blocks the protein-protein interaction between NLRP3 and ASC in LPS-induced iBMDMs[1].
1,2,4-Trimethoxybenzene (1 h) inhibits LPS and nigericin-induced NLRP3 oligomerization of iBMDMs and primary macrophages: DSS crosslinking experiments showed reduced formation of NLRP3 monomers, dimers and higher oligomers[1].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.
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Cell Line:Immortalized murine bone marrow-derived macrophages (iBMDMs)
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Concentration:0.5 mM, 1 mM
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Incubation Time:1.5 h
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Result:Caspase-1 (Casp-1) cleavage and IL-1β secretion are reduced.
In Vivo
1,2,4-Trimethoxybenzene (50-100 mg/kg, p.o., once daily for 3 days) enhances the extinction of fear memories and alleviates PTSD-related anxiety and depression-like behaviors in mice by inhibiting the NLRP3 inflammasome[2].
1,2,4-Trimethoxybenzene (50-200 mg/kg, p.o., once daily for 8 weeks) improves T2DM-related cognitive dysfunction by inhibiting NLRP3 inflammasome activation and regulating gut microbiota in type 2 diabetic rats[3].
MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.
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Animal Model:Female C57BL/6 mice (6-8 weeks old) were subcutaneously injected with 150 μg of MOG35-55 peptide, and intraperitoneally injected with 200 ng of pertussis toxin on the day of immunization (day 0) and day 2, to simulate EAE[1].
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Dosage:200 mg/kg
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Administration:P.o., once daily for 17 days
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Result:The incidence of end-stage renal disease (ESRD) decreased, and clinical symptoms were significantly alleviated.
Significantly reduced weight loss indicates a reduction in systemic inflammatory response.
The area of myelin basic protein (MBP) positive in spinal cord sections was significantly larger than in the control group, indicating a reduced degree of demyelination.
The percentage of demyelinated area was significantly lower than in the control group, confirming the protection of myelin structure.
ASC specks were significantly reduced in spinal cord sections, suggesting that NLRP3 inflammasome activation was inhibited.
The expression of NLRP3, ASC, and cleaved caspase-1 proteins was downregulated in spinal cord tissue.
The mRNA expression of pro-inflammatory cytokines IFN-γ and IL-17a and the chemokine CCL-5 was decreased in the spinal cord, while the expression of the anti-inflammatory cytokine IL-4 was upregulated.
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Animal Model:Male C57BL/6 mice (8 weeks old) were intraperitoneally injected with lipopolysaccharide (LPS, 2 mg/kg/d) for 3 consecutive days to induce depressive-like behavior[2].
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Dosage:50 mg/kg, 100 mg/kg
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Administration:P.o., once daily for 3 days
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Result:Sucrose preference was significantly increased (with improved depressive-like behavior), and immobility time in the tail suspension test (TST) and forced swimming test (FST) was significantly reduced.
Activation of the hippocampal NLRP3 inflammasome was inhibited (e.g., reduced expression of cleaved IL-1β, cleaved caspase-1, and ASC spot formation).
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Animal Model:Male Sprague-Dawley rats (8 weeks old, 200 g) were first given a high-sugar, high-fat diet for 5 weeks, and then type 2 diabetes (T2DM) was induced by intraperitoneal injection of streptozotocin (STZ, 30 mg/kg)[3].
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Dosage:50 mg/kg, 100 mg/kg, 200 mg/kg
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Administration:P.o., once daily for 8 weeks
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Result:Significant improvements were observed in blood glucose levels (FBG, OGTT-AUC), insulin resistance index (HOMA-IR), and blood lipids (TC, TG, LDL-C) in rats, along with a decrease in oxidative stress markers (MDA).
Shortened escape latency and increased target quadrant time indicate enhanced learning and memory abilities.
Improved neuronal structure was observed in the hippocampal CA1 region, with ELISA detecting elevated BDNF levels and decreased AChE levels in the hippocampus.
Inhibition of the hippocampal NLRP3 inflammasome pathway (e.g., reduced expression of NLRP3, ASC, caspase-1, GSDMD, and IL-1β).
- Detection of Apoptosis
- TUNEL staining for apoptotic DNA fragmentation
- Annexin V plus membrane-impermeant dye apoptosis staining
- Pull-down
- Immunoprecipitation
- Apoptosis Solutions
- Protocol for Bimolecular Fluorescence Complementation (BiFC) Assay
- Research Protocol for Metabolic Diseases
- Pyroptosis Solutions
- Co-Immunoprecipitation
- Protocol for Yeast Two-Hybrid (Y2H) Assay
Chemical Information
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CAS No. 135-77-3
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Appearance Liquid
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Molecular Weight 168.19
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Formula C9H12O3
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Color Light yellow to yellow
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SMILES
COC(C(OC)=C1)=CC=C1OC
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Shipping
Room temperature in continental US; may vary elsewhere.
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Storage
Store at room temperature 3 years
In solvent -80°C 2 years -20°C 1 year
Solvent & Solubility
In Vitro:
DMSO : 100 mg/mL (594.57 mM; Need ultrasonic; Hygroscopic DMSO has a significant impact on the solubility of product, please use newly opened DMSO)
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year. When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year. When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.
Concentration (start) × Volume (start) = Concentration (final) × Volume (final)
In Vivo:
Select the appropriate dissolution method based on your experimental animal and administration route.
- For the following dissolution methods, please ensure to first prepare a clear stock solution using an In Vitro approach and then sequentially add co-solvents:
- To ensure reliable experimental results, the clarified stock solution can be appropriately stored based on storage conditions. As for the working solution for In Vivo experiments, it is recommended to prepare freshly and use it on the same day.
- The percentages shown for the solvents indicate their volumetric ratio in the final prepared solution. If precipitation or phase separation occurs during preparation, heat and/or sonication can be used to aid dissolution.
Add each solvent one by one: 10% DMSO 40% PEG300 5% Tween-80 45% Saline
Solubility: ≥ 2.5 mg/mL (14.86 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 400 μL PEG300, and mix evenly; then add 50 μL Tween-80 and mix evenly; then add 450 μL Saline to adjust the volume to 1 mL.
Preparation of Saline: Dissolve 0.9 g sodium chloride in ddH₂O and dilute to 100 mL to obtain a clear Saline solution.
Add each solvent one by one: 10% DMSO 90% (20% SBE-β-CD in Saline)
Solubility: ≥ 2.5 mg/mL (14.86 mM); Clear solution
This protocol yields a clear solution of ≥ 2.5 mg/mL (saturation unknown).
Taking 1 mL working solution as an example, add 100 μL DMSO stock solution (25.0 mg/mL) to 900 μL 20% SBE-β-CD in Saline, and mix evenly.
Preparation of 20% SBE-β-CD in Saline (4°C, storage for one week): 2 g SBE-β-CD powder is dissolved in 10 mL Saline, completely dissolve until clear.
In Vivo Dissolution Calculator
Please enter the basic information of animal experiments:
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Recommended: Prepare an additional quantity of animals to account for potential losses during experiments.
Please enter your animal formula composition:
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%DMSO +
Recommended: Keep the proportion of DMSO in working solution below 2% if your animal is weak.
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%+
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+%Tween-80 + +
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%Saline +
The co-solvents required include: DMSO, . All of co-solvents are available by MedChemExpress (MCE). , Tween 80. All of co-solvents are available by MedChemExpress (MCE).
Working solution concentration: 0.22 mg/mL
Method for preparing stock solution: mg drug dissolved in μL DMSO. Stock solution concentration: mg/mL.
1. Take μL DMSO stock solution;
2. Add μL .
μL , mix evenly;
3. Then add μL Tween 80, mix evenly;
4. Then add μL
Please ensure that the stock solution in the first step is dissolved to a clear state, and add co-solvents in sequence. You can use ultrasonic heating (ultrasonic cleaner, recommended frequency 20-40 kHz), vortexing, etc. to assist dissolution.
Purity & Documentation
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Data Sheet (285 KB)
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SDS (394 KB)
- English - EN (394 KB)
- Français - FR (394 KB)
- Deutsch - DE (394 KB)
- Norwegian - NO (394 KB)
- Español - ES (394 KB)
- Swedish - SV (394 KB)
- Italian - IT (394 KB)
- Korean - KR (394 KB)
- Portuguese - PT (394 KB)
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Handling Instructions (2659 KB)
References
[1]. Pan RY, et al. 1,2,4-Trimethoxybenzene selectively inhibits NLRP3 inflammasome activation and attenuates experimental autoimmune encephalomyelitis. Acta Pharmacol Sin. 2021 Nov;42(11):1769-1779. doi: 10.1038/s41401-021-00613-8IF: 8.4 Q1 . Epub 2021 Feb 24. Erratum in: Acta Pharmacol Sin. 2022 Feb;43(2):504. [Content Brief]
[2]. Zhang Y, et al. 1,2,4-Trimethoxybenzene ameliorates depression-like behaviors by inhibiting the activation of NLRP3 inflammasome. Int Immunopharmacol. 2025 Apr 4;151:114361. [Content Brief]
[3]. Zhong P, et al. 1,2,4-Trimethoxybenzene alleviates cognitive dysfunction in type 2 diabetes model rats by inhibiting NLRP3 inflammasome and restoring gut microbiota disorders. Biochem Pharmacol. 2025 Dec;242(Pt 4):117409. [Content Brief]
Complete Stock Solution Preparation Table
Please refer to the solubility information to select the appropriate solvent. Once prepared, please aliquot and store the solution to prevent product inactivation from repeated freeze-thaw cycles.
Storage method and period of stock solution: -80°C, 2 years; -20°C, 1 year. When stored at -80°C, please use it within 2 years. When stored at -20°C, please use it within 1 year.
| Optional Solvent | Concentration Solvent Mass | 1 mg | 5 mg | 10 mg | 25 mg |
|---|---|---|---|---|---|
| DMSO | 1 mM | 5.9457 mL | 29.7283 mL | 59.4566 mL | 148.6414 mL |
| 5 mM | 1.1891 mL | 5.9457 mL | 11.8913 mL | 29.7283 mL | |
| 10 mM | 0.5946 mL | 2.9728 mL | 5.9457 mL | 14.8641 mL | |
| 15 mM | 0.3964 mL | 1.9819 mL | 3.9638 mL | 9.9094 mL | |
| 20 mM | 0.2973 mL | 1.4864 mL | 2.9728 mL | 7.4321 mL | |
| 25 mM | 0.2378 mL | 1.1891 mL | 2.3783 mL | 5.9457 mL | |
| 30 mM | 0.1982 mL | 0.9909 mL | 1.9819 mL | 4.9547 mL | |
| 40 mM | 0.1486 mL | 0.7432 mL | 1.4864 mL | 3.7160 mL | |
| 50 mM | 0.1189 mL | 0.5946 mL | 1.1891 mL | 2.9728 mL | |
| 60 mM | 0.0991 mL | 0.4955 mL | 0.9909 mL | 2.4774 mL | |
| 80 mM | 0.0743 mL | 0.3716 mL | 0.7432 mL | 1.8580 mL | |
| 100 mM | 0.0595 mL | 0.2973 mL | 0.5946 mL | 1.4864 mL |
1,2,4-Trimethoxybenzene Related Classifications
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