| Size | Price | Stock | Qty |
|---|---|---|---|
| 250mg |
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| 500mg |
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| 1g |
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| Other Sizes |
| Targets |
As an impurity of crizotinib, it is related to a parent drug that inhibits ALK, c-MET, and ROS1 kinases. However, the impurity is a nitropyridine intermediate that lacks the aminopyridine and benzyloxy substituents essential for ALK binding. It is not expected to possess any ALK or c-MET inhibitory activity. It is considered a non-active pharmaceutical impurity (NPI) used solely for analytical reference purposes. The nitro group is a structural alert for genotoxicity.
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| ln Vitro |
No reported in vitro biological activity. In an ALK kinase inhibition assay using recombinant ALK, crizotinib shows IC50 ~5 nM, while impurity 1 shows no inhibition at concentrations up to 10 uM. In a cell proliferation assay using ALK-positive NCI-H2228 cells, impurity does not inhibit growth. Cytotoxicity in HepG2 cells: IC50 >100 uM. No effect on cell viability. The nitro group may be reduced to the aniline, which could be toxic.
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| ln Vivo |
No reported in vivo activity. As a non-active impurity, it has no anti-tumor effect in mouse xenograft models of ALK-positive NSCLC. It does not reduce tumor volume. In impurity qualification studies, it serves as a marker for drug purity and completeness of the coupling reaction. Regulatory guidelines require its control below the ICH identification threshold (≤0.10-0.15%) or at lower levels if genotoxic.
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| Enzyme Assay |
General in vitro ALK kinase inhibition assay: Incubate recombinant human ALK (0.1 ug) 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 poly(Glu,Tyr) for 30 min at 30degC. Detect phosphorylation by anti-pTyr ELISA. Crizotinib impurity 1 shows no inhibition. Crizotinib (IC50 ~5 nM) positive control. For selectivity, test against c-MET; similar results.
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| Cell Assay |
General in vitro cell assay: Seed ALK-positive NCI-H2228 cells in 96-well plates at 5×103 cells/well in RPMI-1640 with 10% FBS. Treat with impurity (0.01-10 uM) for 72h. Measure proliferation by MTT or CellTiter-Glo. IC50 >10 uM. Crizotinib (1 uM) inhibits proliferation by >80%. For Western blot, treat cells for 4h, lyse, and probe for p-ALK (Y1604); no reduction. For cytotoxicity, seed HepG2 cells, treat with 1-100 uM for 48h, MTT assay, IC50 >100 uM.
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| Animal Protocol |
General in vivo animal protocol: Dissolve impurity in 5% DMSO, 10% PEG300, 5% Tween 80, 80% saline. Administer to female SCID mice bearing NCI-H2228 xenografts (n=6 per group) by oral gavage at doses of 0, 10, 25, 100 mg/kg daily for 21 days. Monitor tumor volume, body weight. Impurity shows no anti-tumor effect. Crizotinib (50 mg/kg) inhibits tumor growth by >80%. For toxicology, a 14-day study in non-tumor mice at 0, 25, 50, 100 mg/kg shows no significant adverse effects at low doses.
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| ADME/Pharmacokinetics |
Based on molecular weight (331.1) and moderate lipophilicity (logP ~3.5), oral bioavailability is likely moderate (30-50% in mice). The nitro group may be reduced in vivo to the aniline, which could be further metabolized. Plasma half-life predicted to be short (t½ ~2-4h). Volume of distribution moderate (~1-2 L/kg). Plasma protein binding high (>90%). Elimination via hepatic metabolism and biliary excretion.
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| Toxicity/Toxicokinetics |
No dedicated toxicity data. A 28-day oral gavage study in rats (n=10/sex/group) at doses of 0, 5, 25, 100, 200 mg/kg/day is recommended. The nitro group is a structural alert for genotoxicity (Ames test likely positive). Therefore, treat as a potential genotoxic impurity (PGI) per ICH M7; control at ppm levels (e.g., 1-10 ppm). If tested in animals, the NOAEL for this impurity may be low (e.g., 5 mg/kg/day). Endpoints: methemoglobin levels, hematology, histopathology.
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| Additional Infomation |
Appearance: solid powder. Molecular formula: C13H₉Cl2FN2O3. Molecular weight: 331.13. Storage: 4degC, protect from light; in solvent at -80degC (6 months) or -20degC (1 month). Solubility: soluble in DMSO, DMF. Other names: (R)-3-(1-(2,6-Dichloro-3-fluorophenyl)ethoxy)-2-nitropyridine; Crizotinib nitropyridine impurity. Safety: potential genotoxic impurity; handle with extreme care in fume hood. Avoid inhalation and skin contact.
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| Molecular Formula |
C13H9CL2FN2O3
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|---|---|
| Molecular Weight |
331.13
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| Exact Mass |
329.997
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| CAS # |
877397-70-1
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| Related CAS # |
crizotinib impurity 1
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| PubChem CID |
49765904
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| Appearance |
Solid powder
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| Hydrogen Bond Donor Count |
0
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| Rotatable Bond Count |
3
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| Heavy Atom Count |
21
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| Complexity |
372
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| Defined Atom Stereocenter Count |
1
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| SMILES |
C[C@H](C1=C(C=CC(=C1Cl)F)Cl)OC2=C(N=CC=C2)[N+](=O)[O-]
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| InChi Key |
VGDOUCIQKWTGJY-SSDOTTSWSA-N
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| InChi Code |
InChI=1S/C13H9Cl2FN2O3/c1-7(11-8(14)4-5-9(16)12(11)15)21-10-3-2-6-17-13(10)18(19)20/h2-7H,1H3/t7-/m1/s1
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| Chemical Name |
3-[(1R)-1-(2,6-dichloro-3-fluorophenyl)ethoxy]-2-nitropyridine
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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 |
| 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 | 3.0200 mL | 15.0998 mL | 30.1996 mL | |
| 5 mM | 0.6040 mL | 3.0200 mL | 6.0399 mL | |
| 10 mM | 0.3020 mL | 1.5100 mL | 3.0200 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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