| Size | Price | Stock | Qty |
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| 1mg |
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| 5mg |
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| 100mg | |||
| Other Sizes |
| Targets |
Nuclear factor of activated T cells (NFAT) (Inhibits activation) [1]
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| ln Vitro |
Dihydrocucurbitacin B inhibited the proliferation of phytohemagglutinin (PHA)-stimulated human T lymphocytes with an IC₅₀ of 1.48 µM (range 1.0-2.5 µM). At 2.5 µM, it completely inhibited cell proliferation. The compound was not cytotoxic at concentrations up to 10 µM.
Dihydrocucurbitacin B (2.5 µM) arrested the cell cycle of PHA-stimulated T lymphocytes in the G₀/G₁ phase, with minimal progression to the S phase. This effect was similar to dexamethasone (5 µM), while aphidicolin (0.75 µM) arrested the cycle in the S phase. Dihydrocucurbitacin B (2.5-1.0 µM) significantly inhibited the production of interferon-γ (IFN-γ), interleukin-2 (IL-2), and interleukin-10 (IL-10) in PHA-stimulated human T lymphocytes. It also slightly reduced interleukin-4 (IL-4) production, but this was not statistically significant. Reverse transcription-polymerase chain reaction (RT-PCR) analysis showed that Dihydrocucurbitacin B (2.5 µM) attenuated the mRNA expression of IFN-γ, IL-2, IL-4, and IL-10 in stimulated T lymphocytes. Western blot analysis demonstrated that Dihydrocucurbitacin B (2.5 µM) inhibited the expression of key cell cycle cyclins (A₁, B₁, D₂, and E₁) in PHA-stimulated human T lymphocytes. An electrophoretic mobility shift assay (EMSA) showed that Dihydrocucurbitacin B (2.5 µM) inhibited the activation of the transcription factor NFAT in PHA-stimulated T lymphocytes, without affecting calcium influx into the cells. [1] |
| ln Vivo |
In a mouse model of oxazolone-induced delayed-type hypersensitivity (DTH), topical application of Dihydrocucurbitacin B (0.1, 0.3, and 0.5 mg/ear) significantly inhibited ear swelling in a dose-dependent manner.
In a mouse model of dinitrofluorobenzene (DNFB)-induced DTH, topical application of Dihydrocucurbitacin B (0.1, 0.3, and 0.5 mg/ear) also significantly inhibited ear swelling. In a mouse model of sheep red blood cell (SRBC)-induced DTH, intraperitoneal administration of Dihydrocucurbitacin B (10 mg/kg) inhibited paw swelling. Histological analysis of ear tissue from oxazolone- and DNFB-treated mice revealed that Dihydrocucurbitacin B reduced edema, inflammatory cell infiltration (neutrophils, lymphocytes, macrophages), and epidermal changes (papillomatosis, spongiosis) compared to control groups. Ex vivo analysis of inflamed ear tissue from oxazolone- and DNFB-treated mice showed that Dihydrocucurbitacin B reduced the levels of key pro-inflammatory cytokines, including interleukin-1β (IL-1β), interleukin-4 (IL-4), and tumor necrosis factor-α (TNF-α). [1] |
| Cell Assay |
T Lymphocyte Proliferation Assay: Human peripheral blood lymphocytes were isolated and resuspended. Cells were seeded into 96-well plates and stimulated with phytohemagglutinin (PHA). Dihydrocucurbitacin B at various concentrations (1.0 to 2.5 µM) or dexamethasone (5 µM) were added. Plates were incubated for 4 days. Cell proliferation was then quantified using a modified colorimetric MTT assay. Controls included PHA-stimulated cells (100% activity) and unstimulated cells (0% activity).
Cell Cycle Analysis: T lymphocytes were adjusted to a density of 1 × 10⁶ cells/mL and seeded into 24-well plates with or without PHA stimulation. Dihydrocucurbitacin B (2.5 µM), dexamethasone (5 µM), or aphidicolin (0.75 µM) were added. After incubation for specified times, cells were harvested, fixed in ethanol, stained with propidium iodide and ribonuclease A, and analyzed by flow cytometry to determine DNA content and cell cycle phase distribution. Cell Viability and Cytotoxicity Determination: T lymphocytes were cultured and stimulated with PHA for 4 days. Propidium iodide solution was then added, and cell viability was measured by flow cytometry after 30 minutes. Cytokine Production Determination: T lymphocytes were cultured with PHA alone or in combination with various concentrations of Dihydrocucurbitacin B for 4 days. Cell culture supernatants were collected and assayed for IL-2, IL-4, IL-10, and IFN-γ levels using enzyme-linked immunosorbent assay (ELISA) kits. RNA Extraction and RT-PCR for Cytokine mRNA: T cells were treated with or without PHA and cultured with Dihydrocucurbitacin B for 18 hours. Total RNA was extracted using spin columns. RNA concentration and purity were determined spectrophotometrically. Reverse transcription was performed to generate cDNA, which was then used as a template for PCR amplification with specific primers for IFN-γ, IL-2, IL-4, IL-10, and β-actin (as a housekeeping control). PCR products were analyzed by gel electrophoresis and densitometry. [1] |
| Animal Protocol |
Oxazolone- and Dinitrofluorobenzene (DNFB)-Induced DTH in Mice: Female Swiss mice were sensitized by topical application of the allergen (oxazolone or DNFB) on days 1 and 2. On day 6, a challenge dose was applied to the ears. Dihydrocucurbitacin B (0.1, 0.3, or 0.5 mg/ear) dissolved in acetone was topically applied to the ears at 2, 24, 48, and 72 hours after challenge. Ear thickness was measured with a micrometer before treatment and at 24, 48, and 72 hours post-challenge to calculate edema.
Sheep Red Blood Cell (SRBC)-Induced DTH in Mice: Dihydrocucurbitacin B was administered intraperitoneally at a dose of 10 mg/kg. The specific timing of administration relative to SRBC immunization and challenge is not detailed in the provided text. Paw swelling was measured as an indicator of the inflammatory response. [1] |
| Toxicity/Toxicokinetics |
Dihydrocucurbitacin B at concentrations up to 10 µM showed no cytotoxicity to human T lymphocytes, as determined by propidium iodide exclusion method and flow cytometry. [1]
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| References | |
| Additional Infomation |
23,24-Dihydrocucurbitacin B is a 23,24-dihydrocucurbitacin with a lanosterane skeleton that is multiplely substituted with hydroxyl, methyl, and oxo substituents, and an unsaturated bond at position 5; the hydroxyl groups at positions C-25 are acetylated. It is a 23,24-dihydrocucurbitacin, a secondary α-hydroxy ketone, and a tertiary α-hydroxy ketone. Its function is related to that of cucurbitacin B. Dihydrocucurbitacin B has been reported to exist in Begonia nantoensis, Trichosanthes cucumeroides, and other organisms with relevant data. Dihydrocucurbitacin B is a triterpenoid compound isolated from the roots of Cayaponia tayuya. It possesses anti-inflammatory and potential immunomodulatory activities. Its main mechanism of action in inhibiting T cell-mediated delayed-type hypersensitivity (DTH) is the inhibition of the transcription factor NFAT. This inhibition leads to a decrease in the expression of key cell cycle proteins (A₁, B₁, D₂, E₁) and pro-inflammatory cytokines (e.g., IL-2, IFN-γ), thereby arresting the cell cycle at the G₀/G₁ phase and inhibiting the proliferation of activated T lymphocytes. The compound does not appear to act through a glucocorticoid-like mechanism or by directly inhibiting calcium ion influx. [1]
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| Molecular Formula |
C32H48O8
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|---|---|
| Molecular Weight |
560.7187
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| Exact Mass |
560.334
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| CAS # |
13201-14-4
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| PubChem CID |
267250
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| Appearance |
White to off-white solid powder
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| Density |
1.2±0.1 g/cm3
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| Boiling Point |
692.2±55.0 °C at 760 mmHg
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| Flash Point |
215.4±25.0 °C
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| Vapour Pressure |
0.0±4.9 mmHg at 25°C
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| Index of Refraction |
1.559
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| LogP |
2.56
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| Hydrogen Bond Donor Count |
3
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| Hydrogen Bond Acceptor Count |
8
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| Rotatable Bond Count |
7
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| Heavy Atom Count |
40
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| Complexity |
1170
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| Defined Atom Stereocenter Count |
9
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| SMILES |
O([H])[C@]1([H])C([H])([H])[C@@]2(C([H])([H])[H])[C@]3([H])C([H])([H])C([H])=C4C(C([H])([H])[H])(C([H])([H])[H])C([C@]([H])(C([H])([H])[C@@]4([H])[C@]3(C([H])([H])[H])C(C([H])([H])[C@]2(C([H])([H])[H])[C@@]1([H])[C@@](C(C([H])([H])C([H])([H])C(C([H])([H])[H])(C([H])([H])[H])OC(C([H])([H])[H])=O)=O)(C([H])([H])[H])O[H])=O)O[H])=O
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| InChi Key |
QZJJDOYZVRUEDY-NRNCYQGDSA-N
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| InChi Code |
InChI=1S/C32H48O8/c1-17(33)40-27(2,3)13-12-23(36)32(9,39)25-21(35)15-29(6)22-11-10-18-19(14-20(34)26(38)28(18,4)5)31(22,8)24(37)16-30(25,29)7/h10,19-22,25,34-35,39H,11-16H2,1-9H3/t19-,20+,21-,22+,25+,29+,30-,31+,32+/m1/s1
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| Chemical Name |
[(6R)-6-[(2S,8S,9R,10R,13R,14S,16R,17R)-2,16-dihydroxy-4,4,9,13,14-pentamethyl-3,11-dioxo-2,7,8,10,12,15,16,17-octahydro-1H-cyclopenta[a]phenanthren-17-yl]-6-hydroxy-2-methyl-5-oxoheptan-2-yl] acetate
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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 | 1.7834 mL | 8.9171 mL | 17.8342 mL | |
| 5 mM | 0.3567 mL | 1.7834 mL | 3.5668 mL | |
| 10 mM | 0.1783 mL | 0.8917 mL | 1.7834 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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