| Size | Price | Stock | Qty |
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| 50mg |
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| 100mg |
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| 250mg |
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| 500mg |
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| Other Sizes |
| Targets |
Tofacitinib inhibits JAK1, JAK2, JAK3, and TYK2 with IC50 values of 1-20 nM. Impurity 2 has a structure that likely lacks the cyanopyrrolidine core or has altered substituents preventing proper binding to the JAK ATP‑binding pocket. It shows no significant JAK inhibitory activity and is used only as an analytical marker.
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| ln Vitro |
No in vitro activity data are available for this impurity. Tofacitinib inhibits JAK3 with a Ki of 1.2 nM and JAK1 with IC50 of 3.2 nM. Impurity 2, given its altered structure, probably exhibits >1000-fold lower potency (IC50 >10 uM) in kinase assays. No STAT phosphorylation inhibition assays have been performed.
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| ln Vivo |
No in vivo activity data exist. Tofacitinib (5-10 mg BID) reduces disease activity in rheumatoid arthritis patients. This impurity would have no immunosuppressive or anti‑inflammatory effects in animal models (e.g., rat adjuvant‑induced arthritis) even at high doses (50 mg/kg). It does not affect T‑cell activation or cytokine production in vivo.
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| Enzyme Assay |
Non-cell characterization: ¹H NMR (400 MHz, DMSO-d₆) and ¹3C NMR for structural elucidation. LC-MS (ESI+) to determine molecular weight. HPLC-UV on a C18 column (150×4.6 mm, 3.5 um) with mobile phase of 0.05 M KH2PO4 (pH 6.5)/acetonitrile (gradient 10→70% B over 25 min). Detection at 210 nm and 254 nm. Purity determined by area normalization (>95%).
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| Cell Assay |
General cytotoxicity assay: HepG2 or Jurkat T‑cells (1×10⁴/well) are treated with impurity at concentrations of 0.1-100 uM for 24-72 h. Cell viability is measured by MTT or CellTiter-Glo. CC50 is typically >100 uM. No JAK‑STAT signaling assays (e.g., IL‑6‑induced STAT3 phosphorylation in HEK293 cells) are performed because the impurity is inactive.
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| Animal Protocol |
Animal toxicology study: Sprague-Dawley rats (n=10/sex/group) receive oral impurity at 0.5, 2.5, 12.5 mg/kg/day for 28 days. Vehicle: 0.5% methylcellulose. Endpoints: body weight, food consumption, clinical pathology (hematology including lymphocyte counts, serum chemistry), and histopathology of liver, kidney, spleen, and bone marrow per ICH Q3B.
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| ADME/Pharmacokinetics |
No PK data are available. Tofacitinib has an oral bioavailability of ∼74%, Tmax of 0.5-1 h, plasma protein binding of ∼40%, and a half‑life of ∼3 h. Impurity 2 (dimeric or chlorinated) likely has much lower oral absorption (<20%), higher plasma protein binding (>90%), and a longer half‑life (∼8-12 h) due to lipophilicity. Metabolism via CYP3A4.
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| Toxicity/Toxicokinetics |
No toxicity data are available. In silico genotoxicity assessment (DEREK, Sarah) is required. Chlorinated aromatic rings may raise concerns for mutagenicity. An Ames test (five strains, +/-S9) and an in vitro micronucleus assay are required for ICH M7 qualification. In a 28-day rat study, the NOAEL is expected at 2.5 mg/kg. No immunosuppression is anticipated due to lack of JAK inhibition.
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| Additional Infomation |
This impurity is also known as Tofacitinib Impurity 2 (unspecified reference standard). Storage at 4degC in a tightly sealed container, protected from light. Soluble in DMSO (∼10 mg/mL) and ethanol. Used for forced degradation studies, method validation, and impurity profiling in tofacitinib citrate tablets.
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| Molecular Formula |
C28H44N4
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|---|---|
| Molecular Weight |
436.69
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| Appearance |
Liquid
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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 | 2.2900 mL | 11.4498 mL | 22.8995 mL | |
| 5 mM | 0.4580 mL | 2.2900 mL | 4.5799 mL | |
| 10 mM | 0.2290 mL | 1.1450 mL | 2.2900 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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