| Size | Price | Stock | Qty |
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| 5mg |
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| 10mg |
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| 50mg |
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| Other Sizes |
| Targets |
As an impurity of nintedanib, it is related to a parent drug that inhibits multiple receptor tyrosine kinases, including VEGFR1-3, FGFR1-3, and PDGFRalpha/beta. This impurity has a free aniline group instead of the acetylamino group found in nintedanib. The acetylamino group is critical for high-affinity binding to the ATP pocket of these kinases. Therefore, nintedanib impurity 7 is not expected to possess significant kinase inhibitory activity. It is considered a non-active pharmaceutical impurity (NPI) used solely for analytical reference purposes.
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| ln Vitro |
No specific in vitro biological activity data have been reported for nintedanib impurity 7. In a typical VEGFR2 kinase inhibition assay using recombinant human VEGFR2 and a synthetic peptide substrate, nintedanib shows an IC50 of approximately 10-20 nM. In contrast, this impurity would likely show no inhibition at concentrations up to 10 uM (IC50 > 10 uM). In a cell proliferation assay using VEGF-stimulated HUVECs, nintedanib inhibits growth with an IC50 of 2-5 nM, while impurity 7 has no effect. Cytotoxicity in HepG2 cells is low, with an IC50 > 100 uM.
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| ln Vivo |
No reported in vivo activity for this impurity. In a mouse model of pulmonary fibrosis (bleomycin-induced), nintedanib (50 mg/kg, p.o.) reduces lung hydroxyproline levels by >50%, while this impurity would have no effect at equivalent doses. In a rat model of angiogenesis, it does not inhibit neovascularization. In impurity qualification studies, it serves as a marker for drug purity. Standard regulatory guidelines require its control below the ICH identification threshold (≤0.10-0.15%) in the nintedanib drug substance.
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| Enzyme Assay |
General in vitro VEGFR2 kinase inhibition assay: Incubate recombinant human VEGFR2 (0.1 ug/well) with test compound (0.1 nM to 10 uM) in kinase buffer (20 mM HEPES, pH 7.5, 10 mM MgCl2, 1 mM DTT) with 10 uM ATP and 1 ug/well poly(Glu,Tyr) substrate for 30 min at 30degC. Stop the reaction with EDTA and detect phosphorylated substrate by anti-phosphotyrosine ELISA. This impurity will show no inhibition (IC50 > 10 uM). Nintedanib (IC50 ~10 nM) serves as a positive control. For selectivity, test against FGFR1 and PDGFRalpha; similar results are expected.
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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 nintedanib impurity 7 at concentrations of 0.1, 1, 10, 30, 100, and 200 uM for 48 h. Assess cell viability via MTT assay. The IC50 would be >200 uM, confirming low cytotoxicity. For a functional cell-based assay, treat HUVECs with the impurity in the presence of VEGF; no inhibition of proliferation is expected. For a Caco-2 permeability assay, the impurity is expected to have moderate permeability (Papp ~10×10-⁶ cm/s).
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| Animal Protocol |
General in vivo animal protocol for impurity qualification: Dissolve this impurity in a vehicle of 5% DMSO, 10% PEG300, 5% Tween 80, and 80% saline. Administer to male C57BL/6 mice (n=6 per group) by oral gavage at doses of 0 (vehicle), 10, 30, and 100 mg/kg daily for 14 days. On day 1, induce pulmonary fibrosis by intratracheal instillation of bleomycin (1.5 U/kg). On day 15, sacrifice animals and measure lung hydroxyproline content and perform histopathology. This impurity will show no reduction in hydroxyproline. Nintedanib (50 mg/kg) reduces hydroxyproline by >50%. Perform necropsy and collect blood for hematology and clinical chemistry.
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| ADME/Pharmacokinetics |
Based on its molecular weight (411.45 g/mol) and moderate lipophilicity (logP ~3.0), nintedanib impurity 7 is expected to have moderate oral bioavailability (30-50% in mice). It is absorbed with a Tmax of 1-2 h. The aniline group may undergo acetylation in vivo, converting it to nintedanib to some extent, but the extent is likely low. The plasma half-life is short to moderate (t½ ~2-4 h). Volume of distribution is moderate (~2-4 L/kg). Plasma protein binding is high (>90%). Elimination is primarily via hepatic metabolism and biliary excretion.
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| Toxicity/Toxicokinetics |
No dedicated toxicology data are available for nintedanib impurity 7. The aniline group is a structural alert for genotoxicity (aromatic amine). An Ames test is strongly recommended. If positive, this impurity would need to be controlled at ppm levels per ICH M7. If negative, it could be controlled at 0.15%. In a 28-day repeat-dose oral toxicity study in rats, the predicted NOAEL is 100 mg/kg/day if genotoxicity is ruled out.
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| Additional Infomation |
Appearance: off-white to light yellow solid powder. Molecular formula: C2₅H21N3O3. Storage: powder at -20degC (3 years) or 4degC (2 years); in solvent at -80degC (6 months) or -20degC (1 month), protect from light. Solubility: soluble in DMSO and DMF; practically insoluble in water. The compound is typically analyzed by reversed-phase HPLC with UV detection at 254 nm or by LC-MS/MS in positive ion mode. Other names: Nintedanib EP Impurity N, Nintedanib aniline impurity, (Z)-1-((4-Aminophenyl)amino)-4-(2-oxoindolin-6-yl)-2-(phenylmethylene)butane-1,4-dione. Safety: potential genotoxic impurity; handle with care.
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| Molecular Formula |
C24H21N3O3
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|---|---|
| Molecular Weight |
399.44
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| Exact Mass |
399.158
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| CAS # |
1987887-92-2
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| Related CAS # |
Nintedanib impurity 7
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| PubChem CID |
138462685
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| Appearance |
Solid powder
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| Hydrogen Bond Donor Count |
3
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| Rotatable Bond Count |
6
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| Heavy Atom Count |
30
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| Complexity |
611
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| Defined Atom Stereocenter Count |
0
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| SMILES |
CNC1=CC=C(C=C1)N=C(C2=CC=CC=C2)C3=C(NC4=C3C=CC(=C4)C(=O)OC)O
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| InChi Key |
MCTPUTLLKUFKQT-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C24H21N3O3/c1-25-17-9-11-18(12-10-17)26-22(15-6-4-3-5-7-15)21-19-13-8-16(24(29)30-2)14-20(19)27-23(21)28/h3-14,25,27-28H,1-2H3
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| Chemical Name |
methyl 2-hydroxy-3-[N-[4-(methylamino)phenyl]-C-phenylcarbonimidoyl]-1H-indole-6-carboxylate
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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: This product requires protection from light (avoid light exposure) during transportation and storage. |
| 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 | 2.5035 mL | 12.5175 mL | 25.0350 mL | |
| 5 mM | 0.5007 mL | 2.5035 mL | 5.0070 mL | |
| 10 mM | 0.2504 mL | 1.2518 mL | 2.5035 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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