| Size | Price | Stock | Qty |
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| 25mg |
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| 50mg |
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| 100mg |
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| Other Sizes |
| ln Vitro |
- Graveoline reduced A375 cell viability in a dose-dependent manner with an IC50 value of 22.23 ± 0.79 μg/mL after 48 h treatment. In contrast, it displayed minimal and negligible cytotoxic effect on normal peripheral blood mononuclear cells (PBMC). [1]
- Graveoline induced autophagy as evidenced by: detection of acidic vesicular organelles (AVOs) using acridine orange staining (orange-red fluorescence); conversion of LC3-I (18 kDa) to LC3-II (16 kDa) by immunoblot; increased punctate pattern of LC3 by confocal microscopy; up-regulation of Atg5 and Atg7; down-regulation of Bcl-2 and mTOR; increased expression of Beclin-1 by immunoblot and confocal microscopy. [1] - In the presence of 3-methyladenine (3-MA, autophagy inhibitor), Beclin-1 expression was down-regulated, AVOs formation was reduced, and cell viability increased to 61.86% (compared to 50% with graveoline alone) at the IC50 dose, indicating autophagy led to cell death. 3-MA alone increased viability to 91.17%. [1] - Graveoline induced apoptosis as shown by Annexin V-FITC/PI dual stain flow cytometry, with progressive increment in Annexin V+ cell population. [1] - Blockage of autophagy by 3-MA failed to stop apoptosis induction; 3-MA showed an increase in early apoptosis in the presence of graveoline compared to graveoline alone. Conversely, caspase inhibitor z-DEVD-fmk successfully inhibited apoptosis but slightly increased autophagosome formation, and slightly increased cell viability compared to graveoline alone, suggesting autophagic cell death occurred independent of apoptosis. [1] - Graveoline elevated reactive oxygen species (ROS) level at early hours (between 4th and 6th hour) as measured by H2DCFDA staining and flow cytometry. [1] - In the presence of ascorbic acid (ROS scavenger), graveoline failed to elevate ROS level and failed to induce significant cell death (cell viability increased to 88.62% at IC50 dose), indicating cell death was ROS-mediated. [1] - Inhibition of autophagy by 3-MA did not affect ROS generation induced by graveoline, suggesting ROS generation was upstream of autophagy induction. [1] |
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| Cell Assay |
- Cell viability assay (MTT): A375 cells and PBMC were exposed to different concentrations (0–35 μg/mL) of graveoline for 48 h. MTT assay was performed to determine cell viability. The IC50 value for A375 cells was 22.23 ± 0.79 μg/mL. [1]
- Detection and quantification of AVOs with acridine orange: After 48 h exposure to graveoline (22.23 ± 0.79 μg/mL), cells were collected, fixed with 70% chilled ethanol, stained with 5 mM acridine orange, and observed under fluorescence microscope. Quantitative estimation of fluorescence changes was carried out by flow cytometry. [1] - Immunoblot analysis: Cells were treated with graveoline (22.23 ± 0.79 μg/mL) for different time points (0–48 h). Antibodies used: anti-LC3, anti-Beclin-1, anti-Atg5, anti-Atg7, anti-mTOR, anti-Bcl-2, and anti-GAPDH (loading control). Protein expression was detected by immunoblot. [1] - Confocal microscopy for Beclin-1 and LC3: Treated cells on coverslips were fixed, permeabilized, blocked with 2% BSA + 0.5% Triton X100 in PBS, incubated with primary antibodies (Beclin-1 and LC3) for 2 h at 37°C, then with FITC-tagged secondary antibody, stained with DAPI (10 μg/mL), and mounted. Images captured by confocal microscopy. [1] - Annexin V-FITC/PI dual stain assay: Approximately 10^5 cells were treated with graveoline (22.23 ± 0.79 μg/mL for 48 h) in the presence or absence of z-DEVD-fmk or 3-MA. Cells were stained with FITC-tagged Annexin V (10 μM) and PI (5 μM). Fluorescence measured through FL-1 filter (530 nm) and FL-2 filter (585 nm); 10^4 events acquired. Data analyzed by flow cytometry. [1] - Intracellular ROS generation analysis: Cells treated with graveoline (22.23 ± 0.79 μg/mL) for different time points (0–48 h) in the presence or absence of ascorbic acid (300 μM) or 3-MA were incubated with 10 μM H2DCFDA. Qualitative analysis under fluorescence microscope; quantitative estimation by flow cytometry with excitation 480 nm and emission 530 nm using FL-1H filter. [1] |
| Toxicity/Toxicokinetics |
Graveoline displayed minimal and negligible cytotoxic effect on normal human peripheral blood mononuclear cells (PBMC) at concentrations up to 35 μg/mL for 48 h, as determined by MTT assay. [1]
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| References |
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| Additional Infomation |
Graveoline is a member of the quinoline family of compounds. It has been reported to be found in Oenothera biloba, Griffith's clover, and other organisms for which relevant data exist.
- Graveoline is an alkaloid isolated from ethanolic extract of Ruta graveolens. Its structural formula and mass spectra (m/z: 280.14 [M+1]+) are presented. [1] - Graveoline induced both apoptotic and autophagic cell death in skin melanoma A375 cells, and autophagic cell death was independent of apoptosis. This property is desirable for anti-cancer drug design, particularly because cancer cells can become resistant to apoptosis but can still be killed by autophagy. [1] - The molecular mechanism involves ROS generation that is upstream of autophagy induction; ROS elevation is essential for graveoline-induced cell death. [1] |
| Molecular Formula |
C17H13NO3
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|---|---|
| Molecular Weight |
279.2900
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| Exact Mass |
279.09
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| CAS # |
485-61-0
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| Related CAS # |
Graveoline; 485-61-0
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| PubChem CID |
353825
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| Appearance |
White to off-white solid
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| Density |
1.327g/cm3
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| Boiling Point |
436.7ºC at 760mmHg
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| Melting Point |
204-205 °C
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| Flash Point |
217.9ºC
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| Vapour Pressure |
7.91E-08mmHg at 25°C
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| Index of Refraction |
1.652
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| Source |
Originated from plants: Rutaceae Ruta graveolens Linn.
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| LogP |
2.934
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| Hydrogen Bond Donor Count |
0
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| Hydrogen Bond Acceptor Count |
4
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| Rotatable Bond Count |
1
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| Heavy Atom Count |
21
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| Complexity |
461
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| Defined Atom Stereocenter Count |
0
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| SMILES |
O1C([H])([H])OC2C([H])=C([H])C(=C([H])C1=2)C1=C([H])C(C2=C([H])C([H])=C([H])C([H])=C2N1C([H])([H])[H])=O
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| InChi Key |
COBBNRKBTCBWQP-UHFFFAOYSA-N
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| InChi Code |
InChI=1S/C17H13NO3/c1-18-13-5-3-2-4-12(13)15(19)9-14(18)11-6-7-16-17(8-11)21-10-20-16/h2-9H,10H2,1H3
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| Chemical Name |
2-(1,3-benzodioxol-5-yl)-1-methylquinolin-4-one
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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) |
DMSO: ~20 mg/mL (~71.6 mM; with ultrasonication and warming by heating to 60°C)
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|---|---|
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 1 mg/mL (3.58 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween-80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), Clear solution.
For example, if 1 mL of working solution is to be prepared, you can add 100 μL of DMSO stock solution (10.0 mg/mL) to 400 μL of PEG300 and mix well; then add 50 μL of Tween-80 and mix well; finally add 450 μL of physiological saline and adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution. Solubility in Formulation 2: ≥ 1 mg/mL (3.58 mM) (saturation unknown) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), Clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of DMSO stock solution (10.0 mg/mL) to 900 μL of 20% SBE-β-CD saline and mix well. Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Solubility in Formulation 3: ≥ 1 mg/mL (3.58 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (add these co-solvents sequentially from left to right, and one by one), Clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of DMSO stock solution (10.0 mg/mL) to 900 μL of corn oil and mix well. (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 3.5805 mL | 17.9025 mL | 35.8051 mL | |
| 5 mM | 0.7161 mL | 3.5805 mL | 7.1610 mL | |
| 10 mM | 0.3581 mL | 1.7903 mL | 3.5805 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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