5-methoxyflavone, a flavonoid analog, is a novel DNA polymerase-ß inhibitor and neuroprotective agent against ß-amyloid toxicity. 5-Methoxyflavone was confirmed to be an inhibitor of DNA pol-β activity using cultured primary neurons, a valuable model for examining the neuroprotective effects of putative DNA pol-β inhibitors because treatment with Aβ causes DNA replication and cell death in these neurons. 5-methoxyflavone decreased the number of S-phase neurons and the subsequent apoptotic death induced by Aβ, which is consistent with the inhibition of DNA pol-β. 5-Methoxyflavone is the first flavonoid compound that, as opposed to having broad antioxidant and anti-inflammatory effects, is able to stop neurodegeneration through a specific molecular mechanism.

Physicochemical Properties

Molecular Formula	C16H12O3
Molecular Weight	252.26
Exact Mass	252.079
CAS #	42079-78-7
Related CAS #	42079-78-7
PubChem CID	94525
Appearance	Yellow solid powder
Density	1.24g/cm3
Boiling Point	422.5ºC at 760mmHg
Melting Point	131-133°C
Flash Point	201.1ºC
Index of Refraction	1.548
LogP	3.468
Hydrogen Bond Donor Count	0
Hydrogen Bond Acceptor Count	3
Rotatable Bond Count	2
Heavy Atom Count	19
Complexity	368
Defined Atom Stereocenter Count	0
SMILES	O1C(=C([H])C(C2C(=C([H])C([H])=C([H])C1=2)OC([H])([H])[H])=O)C1C([H])=C([H])C([H])=C([H])C=1[H]
InChi Key	XRQSPUXANRGDAV-UHFFFAOYSA-N
InChi Code	InChI=1S/C16H12O3/c1-18-13-8-5-9-14-16(13)12(17)10-15(19-14)11-6-3-2-4-7-11/h2-10H,1H3
Chemical Name	5-methoxy-2-phenylchromen-4-one
Synonyms	5-Methoxyflavone
HS Tariff Code	2934.99.9001
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)

Biological Activity

Targets	DNA polymerase β
ln Vitro	In vitro activity: 5-Methoxyflavone (compound 1) has been identified as a potential compound that can inhibit DNA pol-β in multiple instances and prevent the initiation of the cell cycle and subsequent neuronal apoptosis in primary neuronal cultures challenged with Aβ. On 92TAg cells, 5-methoxyflavone (10–30 μM) can greatly increase the toxicity of MMS. 5-Polymerase activity on a gapped substrate is significantly reduced by 1- or 10-methoxyflavone[1]. 5-Methoxyflavone (5-MF, 0-100 μg/mL) causes the proapoptotic protein BAX to increase and the antiapoptotic proteins cFLIP and Mcl-1 to decrease over time. 5-MF induces TRAIL-R1 (DR4) and TRAIL-R2 (DR5) in a manner that is dependent on time[2].
ln Vivo	5-Methoxyflavone (100, 150 mg/kg, i.p) dramatically reduces the amount of time it takes to lose the righting reflex. 5. A notable and dose-dependent decrease in spontaneous locomotor activity is shown by methylflavone (50, 100, and 150 mg/kg, i.p.). 5. The rearing response is decreased by methylflavone (50, 100 mg/kg, i.p.). 5-The grooming response was totally eliminated by methylflavone (100, 125, and 150 mg/kg, i.p. ), just like in animals treated with diazepam[3].
Enzyme Assay	Human DNA Pol-β Assay [1] DNA pol-β inhibition was evaluated with a specific DNA pol-β assay kit. The assay was carried out strictly following the manufacturer’s instructions. Briefly, a reaction mixture containing a gapped DNA template, dNTPs (both supplied with the kit), and DNA pol-β was incubated for 30 min at RT with or without the compound to be tested. The formation of repaired duplexes, which, unlike gapped DNA, selectively incorporate a fluorescent dye in the presence of Reagent U (both supplied by the kit) was determined by measuring at 535 nm with the excitation wavelength at 485 nm in a fluorimeter. OA (50 μM) was used as a positive control for inhibition. Activity was evaluated as fluorescent signal to no-enzyme-background ratio.
Cell Assay	The MTT assay is used to assess the cell viability after treating MOLT-4, U937, or PBMC cells with DMF, TMF, PMF, 5-MF, or 2'-MF at different concentrations for 24 hours.
Animal Protocol
References	[1]. Identification of 5-Methoxyflavone as a Novel DNA Polymerase-Beta Inhibitor and Neuroprotective Agent against Beta-Amyloid Toxicity. J Nat Prod. 2015 Nov 25;78(11):2704-11. [2]. Sedative-hypnotic like effect of 5-methoxyflavone in mice and investigation on possible mechanisms by in vivo and in silico methods. Biomed Pharmacother. 2018 Dec;108:85-94. [3]. Methoxyflavone derivatives modulate the effect of TRAIL-induced apoptosis in human leukemic cell lines. J Hematol Oncol. 2011 Dec 21;4:52.
Additional Infomation	5-Methoxyflavone is an ether and a member of flavonoids.

Solubility Data

Solubility (In Vitro)

DMSO: 50~125 mg/mL (198.2~495.5 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.08 mg/mL (8.25 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 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 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 to the above solution and mix evenly; then add 450 μL normal saline to 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: ≥ 2.08 mg/mL (8.25 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 20.8 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. 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: ≥ 2.08 mg/mL (8.25 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 20.8 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	3.9642 mL	19.8208 mL	39.6416 mL
	5 mM	0.7928 mL	3.9642 mL	7.9283 mL
	10 mM	0.3964 mL	1.9821 mL	3.9642 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.