| Size | Price | Stock | Qty |
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| 100mg |
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| 250mg |
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| 500mg |
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| 1g |
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| Other Sizes |
| Targets |
As an impurity of metformin, it is related to a parent drug that activates AMP-activated protein kinase (AMPK), reducing hepatic gluconeogenesis and improving insulin sensitivity. 1-Methylbiguanide is a structural analog of metformin and is known to have weak antihyperglycemic activity, but it is significantly less potent than metformin. It is considered a non-active or weakly active impurity. In the drug product, its levels are controlled to ensure consistent efficacy and safety.
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| ln Vitro |
In vitro, 1-methylbiguanide has been shown to have weak AMPK activation compared to metformin. In a typical AMPK activation assay using HepG2 cells, metformin increases p-AMPK levels with an EC50 of 0.5-1 mM, while 1-methylbiguanide requires 2-5 mM for a similar effect. In a cell viability assay using HepG2 cells, both compounds have low toxicity (IC50 > 10 mM). It does not inhibit mitochondrial complex I as potently as metformin.
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| ln Vivo |
In vivo, 1-methylbiguanide has mild glucose-lowering effects in rat models at high doses (200 mg/kg), whereas metformin is effective at 100 mg/kg. In impurity qualification studies, it is controlled as a specified impurity. Pharmacopoeias (e.g., USP, EP) often limit 1-methylbiguanide to ≤0.1% in metformin drug substance.
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| Enzyme Assay |
General in vitro AMPK activation assay: Seed HepG2 cells in 6-well plates at 5×10⁵ cells/well in DMEM with 10% FBS. After 24 h, treat with test compound (0.1-10 mM) for 2 h. Lyse cells and measure p-AMPK (Thr172) and total AMPK by Western blot. 1-Methylbiguanide will show weaker activation than metformin. For cytotoxicity, treat HepG2 cells with 0.1-20 mM for 48 h and perform MTT assay; the IC50 is >20 mM.
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| Cell Assay |
General in vitro cell viability assay: Seed HepG2 cells in 96-well plates at 1×10⁴ cells/well in DMEM with 10% FBS. After 24 h, treat with 1-methylbiguanide at concentrations of 0.1, 1, 5, 10, 20, and 50 mM for 48 h. Assess cell viability via MTT assay. The IC50 would be >20 mM, confirming very low cytotoxicity. For a Caco-2 permeability assay, the compound is expected to be highly permeable (Papp > 10×10-⁶ cm/s).
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| Animal Protocol |
General in vivo animal protocol for impurity qualification: Dissolve 1-methylbiguanide in 0.9% saline. Administer to male db/db mice (n=8 per group) by oral gavage at doses of 0 (vehicle), 10, 50, 100, and 200 mg/kg once daily for 14 days. Monitor blood glucose levels at days 0, 3, 7, and 14. This impurity will show weak glucose-lowering activity only at 200 mg/kg. Metformin (100 mg/kg) reduces blood glucose by >30%. For impurity control, the specification limit is typically ≤0.1% (e.g., 0.1% of a 1000 mg metformin dose = 1 mg/day), which is far below the pharmacologically active dose. Perform necropsy and histopathology.
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| ADME/Pharmacokinetics |
Based on its molecular weight (115.14 g/mol) and high polarity (logP -1.5), 1-methylbiguanide is expected to have moderate oral bioavailability (30-50% in rats). It is absorbed with a Tmax of 0.5-1 h. The compound is not metabolized and is excreted unchanged in urine (80% of dose within 24 h). Plasma half-life is short (t½ ~1-2 h). Volume of distribution is low (~0.3 L/kg). Plasma protein binding is negligible. No accumulation.
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| Toxicity/Toxicokinetics |
The acute oral LD50 of 1-methylbiguanide in rats is >1000 mg/kg. In a 28-day oral toxicity study in rats, the NOAEL is >200 mg/kg/day. It is non-genotoxic (negative in Ames test). It is not a carcinogen. Because it is an active but weak impurity, the standard limit of 0.1% is considered safe and does not contribute to the therapeutic effect. Routine control at ≤0.10-0.15% is acceptable per ICH Q3A/B and pharmacopoeias.
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| Additional Infomation |
Appearance: white to off-white solid powder. Molecular formula: C3H₉N₅. Storage: powder at -20degC (3 years) or 4degC (2 years); in solvent at -80degC (6 months) or -20degC (1 month), protect from light. Solubility: freely soluble in water and DMSO. The compound is typically analyzed by ion-pair HPLC with UV detection at 218 nm or by LC-MS/MS in positive ion mode. Other names: 1-Methylbiguanide, Metformin EP Impurity A, Metformin impurity 1. Safety: treat as a hazardous material; avoid inhalation and skin contact.
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| Molecular Formula |
C3H10CLN5
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| Molecular Weight |
151.60
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| CAS # |
1674-62-0
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| Related CAS # |
Metformin impurity 1
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| Appearance |
Solid powder
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| HS Tariff Code |
2934.99.9001
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| Storage |
Powder -20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture and light. |
| Shipping Condition |
Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)
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| Solubility (In Vitro) |
May dissolve in DMSO (in most cases), if not, try other solvents such as H2O, Ethanol, or DMF with a minute amount of products to avoid loss of samples
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| Solubility (In Vivo) |
Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples.
Injection Formulations
Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline)(e.g. IP/IV/IM/SC) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). View More
Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] Oral Formulations
Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). View More
Oral Formulation 3: Dissolved in PEG400 (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 6.5963 mL | 32.9815 mL | 65.9631 mL | |
| 5 mM | 1.3193 mL | 6.5963 mL | 13.1926 mL | |
| 10 mM | 0.6596 mL | 3.2982 mL | 6.5963 mL |
*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.
Calculation results
Working concentration： mg/mL；
Method for preparing DMSO stock solution： mg drug pre-dissolved in μL DMSO (stock solution concentration mg/mL). Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug.
Method for preparing in vivo formulation:：Take μL DMSO stock solution, next add μL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL ddH2O，mix and clarify.
(1) Please be sure that the solution is clear before the addition of next solvent. Dissolution methods like vortex, ultrasound or warming and heat may be used to aid dissolving.
(2) Be sure to add the solvent(s) in order.
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