Size | Price | Stock | Qty |
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10mg |
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50mg |
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100mg |
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Other Sizes |
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Purity: ≥98%
Targets |
NOX; TRPA1 (EC50: 1 to 3 μM)
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ln Vitro |
NADPH oxidase (NOX) and TRPA1 activator, diphenyl iodide chloride has an EC50 of 1 to 3 μM. Applying diphenyl iodide chloride at concentrations ranging from 0.03 to 10 μM allowed for the successful induction of Ca2+ responses in HEK-TRPA1 cells. Diphenyl iodide chloride, however, was unable to cause Ca2+ responses in control HEK cells, not even at the comparatively high dose of 10 μM. causes the reaction Ca2+[1]. Lipopolysaccharide (LPS) induction was observed upon the addition of diphenyl iodide chloride to the co-culture. Diphenyl ammonium treatment dramatically decreased LPS-induced O2-production by 2.0-fold, bringing it down to within 27% of the control [2].
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ln Vivo |
Licking or biting behavior is induced by injecting 2 mM diphenyl iodonium chloride into the fibrous claw's sole [1]. Diphenylene iodide chloride treatment effectively decreased LPS-induced O4-promoted cell loss either immediately after injection or 24 hours later. White matter fiber disorganization caused by lipopolysaccharide injection was much reduced when diphenyl iodonium was administered either right away or 24 hours later. Nevertheless, LPS-induced white matter damage does not seem to be repaired by diphenyleneiodonium (DPI) treatment 48 hours after LPS injection. The buildup of gp91phox and p67phox in membrane modules can be considerably decreased by DPI therapy given either immediately after the injection of LPS or 24 hours later [2].
The contribution of microglial activation to preoligodendroglial (preOL) damage in the central nervous system (CNS) is considered to be one of the principal causes of periventricular leukomalacia (PVL) pathogenesis. The present study explores the effect of diphenyleneiodonium (DPI) , a NADPH oxidase (NOX) inhibitor, on protection of preOLs from bacterial lipopolysaccharide (LPS)-induced microglial toxicity in vivo and in vitro. In vitro, preOLs co-cultured with microglia exhibited increased preOL apoptosis, accompanied by overproduction of superoxide anion (O(2)(-)) and the formation of peroxynitrite (ONOO(-)) after LPS exposure. LPS also significantly up-regulated accumulation of activated microglial NOX subunits p67-phox and gp91-phox in the plasma membrane. diphenyleneiodonium (DPI) (10μm) was found to significantly attenuate up-regulation of this NOX activity. In vivo, DPI was administered (1mg/kg/day) by subcutaneous injection for 3 days to two-day-old neonatal Sprague-Dawley rats subjected to intracerebral injection of LPS. Treatment with DPI within 24h of LPS injection significantly ameliorated white matter injury, decreasing preOL loss, O(2)(-) generation, and ONOO(-) formation, and inhibiting p67-phox, gp91-phox synthesis and p67phox membrane translocation in microglia. These results indicated that LPS-induced preOL apoptosis may have been mediated by microglia-derived ONOO(-). DPI prevented this LPS-induced brain injury, most likely by inhibiting ONOO(-) formation via NOX, thereby preventing preOL loss and immature white matter injury [2]. |
Cell Assay |
Glial cell cultures were established in two stages; initial mixed culture of neuroglia, followed by isolation and purification of glial cells. Primary, mixed glial cell cultures were first prepared from the cerebral cortex of 2-day-old Sprague–Dawley rats, as described previously with modifications. Briefly, cells isolated from the cerebral cortex were plated at a density of 106 cells/cm2 in 75 cm2 culture flasks containing DMEM/F12 (Hyclone, USA) with 10% fetal bovine serum (FBS; Gibco, USA), feeding every other day for 7–9 days.
Primary preOLs and microglia were then isolated from the primary mixed glial cultures by shaking and differential detachment, as described. Microglia, isolated by shaking mixed glial culture flasks for 2 h at 200 rpm, were maintained in DMEM/F12 (1:1) with 10% FBS for 2–3 days. Primary microglia were found to be >95% pure by immunofluorescence staining (data not shown) for the microglial marker, isolectin B4. Mixed glial cultures were then subjected to further shaking at 220 rpm overnight to separate preOLs from the astrocyte layer (Li et al., 2005). The resulting culture supernatant cell suspension was then plated onto uncoated petri dishes for 1 h to remove residual adherent microglia and astrocytes. Isolated preOLs were maintained in serum-free, preOL-conditioned medium (DMEM/F12, 0.1% bovine serum albumin, 50 μg/ml human apo-transferrin, 50 μg/ml insulin, 30 nM sodium selenite, 10 nM D-biotin, 10 nM hydrocortisone, 10 ng/ml PDGF, 10 ng/ml bFGF) for 3–5 days. Biotinylated monoclonal antibodies specific for oligodendrocyte markers O4 and O1 were used to distinguish O4+O1− late OL precursors from other developing stage-specific OLs immunocytochemically. Primary preOLs were found to be >99% O4+O1− (data not shown). Purified microglia and preOLs were then co-cultured using a Transwell culture system (Corning, USA). Co-cultured cells were divided into three groups: control, LPS-activated, and LPS plus DPI. Microglia were cultured in Transwells over established preOL layers and exposed to either LPS (100 ng/ml;Escherichia coli O111:B4, Sigma), LPS+DPI (10 μmol/l; Sigma, USA) or untreated. The medium supernatants and cellular protein fractions from the co-cultures were then collected for further analysis after 48 h incubation [2]. |
Animal Protocol |
Two-day-old Sprague–Dawley rat pups of both sexes were used for intracerebral injections in a neonatal PVL injury model, as previously described. Briefly, pups were anesthesized with isoflurane (1.5%) and placed under a stereotaxic apparatus with an adapter for neonatal rats. A scalp incision was made in the skull surface to expose the bregma. Intracerebral injection was then performed at a location 1.0 mm posterior, 1.0 mm left of the bregma, and 2.0 mm deep into the skull surface. An LPS suspension in sterile saline (1 mg/kg) was injected into the left hemisphere of the rat brain at a rate of 0.5 μl/min, as previously determined. After injection, the needle was kept in position for an additional 2 min and then retrieved slowly to prevent leakage. The wound was sutured and the pups placed on a thermal blanket (34–35 °C) for recovery. Intracerebral injection was survived by 80% of the animals.
All rat pups were randomly divided into control, LPS, and LPS+DPI groups, and the LPS+DPI group was further divided into DPI-0 h, DPI-24 h and DPI-48 h subgroups. Rat pups injected with sterile PBS were utilized for the control group. For the DPI group, rat pups were subcutaneously injected with DPI (single dose of 1 mg/kg/day), beginning either 0 h, 24 h or 48 h after LPS injection and continuing for a total of 3d, as previously described. All animal studies were approved by the Ethics Committee for Laboratory Animals, Xinhua Hospital Affiliate to Shanghai Jiao Tong University School of Medicine. Five days after LPS injection, all pups were anesthetized with sodium pentobarbital (100 mg/kg) and sacrificed by transcardiac perfusion with cold normal saline, followed by 4% paraformaldehyde for brain section preparation, or by decapitation for brain tissue collection used in immunoblot assays [2]. The ddy mice (6 to 7 wk of age) are individually placed in transparent cages for 30 min before experiments. An intraplantar injection of 10 μL Diphenyleneiodonium chloride (2 mM, solvent: Kolliphor EL with 20% DMSO) is then injected into the right hindpaw with or without intraperitoneal administration with HC030031 (300 mg/kg at 0.5 h prior to injection of Diphenyleneiodonium chloride; solvent: saline with 0.5% methyl cellulose). The time spent licking or biting the injected paw is recorded for 45 min after injection [2]. |
References |
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Additional Infomation |
Dibenziodolium chloride is an organic chloride salt having dibenziodolium as the counterion. It has a role as an EC 1.6.3.1. [NAD(P)H oxidase (H2O2-forming)] inhibitor and a G-protein-coupled receptor agonist. It contains a dibenziodolium.
Transient receptor potential ankyrin 1 (TRPA1) is a Ca(2+)-permeable nonselective cation channel expressed in neuronal and nonneuronal cells and plays an important role in acute and inflammatory pain. Here, we show that an NADPH oxidase (NOX) inhibitor, diphenyleneiodonium (DPI), functions as a TRPA1 activator in human embryonic kidney cells expressing human TRPA1 (HEK-TRPA1) and in human fibroblast-like synoviocytes. Application of DPI at 0.03-10 μM induced a Ca(2+) response in HEK-TRPA1 cells in a concentration-dependent manner. The Ca(2+) response was effectively blocked by a selective TRPA1 antagonist, HC-030031 (HC). In contrast, DPI had no effect on HEK cells expressing TRPV1-V4 or TRPM8. Four other NOX inhibitors, apocynin (APO), VAS2870 (VAS), plumbagin, and 2-acetylphenothiazine, also induced a Ca(2+) response in HEK-TRPA1 cells, which was inhibited by pretreatment with HC. In the presence of 5 mM glutathione, the Ca(2+) response to DPI was effectively reduced. Moreover, mutation of cysteine 621 in TRPA1 substantially inhibited the DPI-induced Ca(2+) response, while it did not inhibit the APO- and VAS-induced responses. The channel activity was induced by DPI in excised membrane patches with both outside-out and inside-out configurations. Internal application of neomycin significantly inhibited the DPI-induced inward currents. In inflammatory synoviocytes with TRPA1, DPI evoked a Ca(2+) response that was sensitive to HC. In mice, intraplantar injection of DPI caused a pain-related response which was inhibited by preadministration with HC. Taken together, our findings demonstrate that DPI and other NOX inhibitors activate human TRPA1 without mediating NOX.[1] |
Molecular Formula |
C12H8CLI
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Molecular Weight |
314.55
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Exact Mass |
313.936
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Elemental Analysis |
C, 45.82; H, 2.56; Cl, 11.27; I, 40.34
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CAS # |
4673-26-1
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Related CAS # |
244-54-2; 4673-26-1 (Cl)
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PubChem CID |
2733504
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Appearance |
White to off-white solid
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Melting Point |
312-322ºC
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Hydrogen Bond Donor Count |
0
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Hydrogen Bond Acceptor Count |
1
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Rotatable Bond Count |
0
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Heavy Atom Count |
14
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Complexity |
170
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Defined Atom Stereocenter Count |
0
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SMILES |
C12=CC=CC=C1C3=CC=CC=C3[I+]2.[Cl-]
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InChi Key |
FCFZKAVCDNTYID-UHFFFAOYSA-M
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InChi Code |
InChI=1S/C12H8I.ClH/c1-3-7-11-9(5-1)10-6-2-4-8-12(10)13-11;/h1-8H;1H/q+1;/p-1
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Chemical Name |
8-iodoniatricyclo[7.4.0.02,7]trideca-1(13),2,4,6,9,11-hexaene;chloride; Dibenziodolium chloride
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Synonyms |
DPI; Dibenziodolium chloride; iphenyleneiodonium chloride; 4673-26-1; Dibenziodolium chloride; DPI; UNII-7M9D81YZ2N; 7M9D81YZ2N; CHEBI:77967; MFCD00214165; Diphenyleneiodonium Chloride
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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, avoid exposure to moisture. |
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) |
DMSO : ~6 mg/mL (~19.07 mM)
H2O : < 0.1 mg/mL |
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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 | 3.1791 mL | 15.8957 mL | 31.7914 mL | |
5 mM | 0.6358 mL | 3.1791 mL | 6.3583 mL | |
10 mM | 0.3179 mL | 1.5896 mL | 3.1791 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.