| Size | Price | Stock | Qty |
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| 50mg |
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| 500mg | |||
| 1g | |||
| Other Sizes |
| Targets |
DHFR/dihydrofolate reductase
Dihydrofolate reductase (DHFR). 10-Formylfolic acid is described as "the most potent natural inhibitor of dihydrofolate reductase known", but no IC₅₀, Ki, EC₅₀, or DC₅₀ values are provided in this paper [1]. |
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| ln Vitro |
During investigation of folate polyglutamate biosynthesis in rat liver slices utilizing [2-(14)C]folic acid, a folate compound that behaved like a polyglutamate form in the Sephadex G-15 gel filtration system was found to accumulate. Subsequent chromatographic, spectral, chemical, and enzymic studies have indicated that the compound formed in liver slices incubated with [(14)C]folic acid with and without methotrexate was 10-formyl folate. This folate is of interest in that it is the most potent natural inhibitor of dihydrofolate reductase known and may be capable of serving a regulatory function within the cell [1].
In rat liver slice incubations, 10-formylfolic acid is formed from [¹⁴C]folic acid at a rate of approximately 4 nmol/h per gram of tissue (wet weight). Its accumulation occurs both in the presence and absence of methotrexate (1 μM), indicating that its synthesis does not require prior reduction of folate [1]. 10-Formylfolic acid is not a substrate for carboxypeptidase G₁ (from Pseudomonas stutzeri), consistent with its lack of a polyglutamate chain [1]. In a dihydrofolate reductase inhibition assay using purified rat liver DHFR, 10-formylfolic acid inhibited the enzyme in a concentration‑dependent manner. A liver extract containing the compound caused 50% inhibition at an amount corresponding to 80 pmol of 10-formylfolic acid, based on a standard curve with authentic material [1]. |
| Enzyme Assay |
Enzyme inhibition assay for dihydrofolate reductase: The assay mixture (final volume 1.0 mL) contained Tris‑HCl buffer pH 7.0 (100 μmol), KCl (150 μmol), NADPH (0.08 μmol), and rat liver dihydrofolate reductase (16 units). Increasing amounts of liver extract (containing 10-formylfolic acid) were added and incubated for 2 min at 37 °C. The reaction was started by adding 0.02 μmol of dihydrofolate. Appropriate blanks without NADPH were run. The amount of 10-formylfolic acid causing 50% inhibition was calculated by comparison with authentic material [1].
Carboxypeptidase G₁ assay: Carboxypeptidase G₁ from Pseudomonas stutzeri (homogeneous on polyacrylamide gel, specific activity 700 units/mg) was used. The enzyme was assayed as previously described [17]. The purified radioactive folate from liver slices was incubated with the enzyme; lack of substrate activity indicated that the compound was not a polyglutamate [1]. Chemical conversion reactions: 10-Formylfolic acid was reduced with potassium borohydride under anaerobic conditions to the corresponding tetrahydro derivative, which upon addition of HCl converted to 5,10‑methenyltetrahydrofolic acid. Characteristic spectral changes were observed, confirming the presence of a formyl group at the 10‑position [1]. Ultraviolet and fluorescence spectroscopy: Difference spectra of authentic 10‑formylfolic acid and the isolated compound were compared after addition of NaOH to pH 13, showing almost identical changes. Fluorescence was measured with activation at 364 nm; emission maximum at 458 nm (neutral pH) was identical to authentic 10‑formylfolic acid [1]. |
| Cell Assay |
The bioactivity of 10-formyl-7,8-dihydrofolic acid and 10-formyl-folic acid was determined in human leukemia (CCRF-CEM) cells grown in a folate-depleted medium containing methotrexate. Excess 10-formyl-7,8-dihydrofolic acid, (but not 10-formyl folic acid) supported the growth of these cells, but it was less potent than5-formyl-5,6,7,8-tetrahydrofolic acid (a control). 10-formyl-7, 8-dihydrofolic acid (not 10-formyl folic acid) was active as substrate for aminoimidazole carboxamide ribotide transformylase and dihydrofolate reductase. This is the first experimental evidence that 10-formyl-7,8-dihydrofolic acid is a bioactive folate in mammalian cells. These experiments and several other lines of evidence in the literature suggest that 10-formyl-folic acid must be metabolized to bioactive folate by enteric bacteria before it can be utilized by the vertebrate host [2].
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| Animal Protocol |
Animal Protocol: Adult male Sprague‑Dawley rats (approximately 300 g) were killed by decapitation, livers removed rapidly and chilled on ice. Liver slices (1.0 g) were prepared using a Stadie‑Rigg’s hand microtome and incubated in 10 mL Krebs‑Ringer bicarbonate buffer with glucose (5.5 mM) containing 1 μCi of [2‑¹⁴C]folic acid (specific activity 55.3 μCi/μmol). In some experiments, 1 μM methotrexate or unlabeled folic acid (0.1 mM) was added. Incubation was carried out at 37 °C with shaking for 4‑6 h under 95% O₂/5% CO₂. After incubation, slices were washed with ice‑cold saline, dropped into boiling 1% ascorbate (pH 6.0) for 5 min to extract folates, homogenized, and centrifuged. The supernatant was subjected to Sephadex G‑15 gel filtration (2.4×45 cm column, 0.02 M phosphate buffer pH 7.0, 5.5 mL fractions) and DEAE Sephadex A‑25 chromatography (linear gradient 0.1‑0.7 M NaCl in 0.01 M phosphate buffer pH 7.0) to purify the radioactive compound [1].
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| References | |
| Additional Infomation |
According to reports, chili peppers contain 10-formylfolate. Data is available for the year in which it is found.
10-Formylfolic acid (also known as 10-formylfolate) is a naturally occurring folate derivative. It was previously identified as rhizopterin when isolated from Rhizopus nigricans. It supports growth of Streptococcus faecalis and Lactobacillus casei but not Pediococcus cerevisiae. The compound has unusual properties: it behaves like a pteroylpolyglutamate on Sephadex G‑15 (eluting earlier than monoglutamates) but like a monoglutamate on DEAE anion exchange. It fluoresces strongly at 458 nm when excited at 364 nm. Its potent inhibition of dihydrofolate reductase suggests a possible regulatory role in intracellular folate metabolism. The study also raises the possibility that 10-formylfolic acid may be an intermediate in folate polyglutamate formation [1]. |
| Molecular Formula |
C20H19N7O7
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|---|---|
| Exact Mass |
469.135
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| Elemental Analysis |
C, 51.17; H, 4.08; N, 20.89; O, 23.86
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| CAS # |
134-05-4
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| Related CAS # |
10-Formylfolic acid-d4;461426-41-5
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| PubChem CID |
135405023
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| Appearance |
Typically exists as solid at room temperature
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| Density |
1.67g/cm3
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| Index of Refraction |
1.748
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| LogP |
1.114
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| Hydrogen Bond Donor Count |
5
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| Hydrogen Bond Acceptor Count |
10
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| Rotatable Bond Count |
9
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| Heavy Atom Count |
34
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| Complexity |
843
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| Defined Atom Stereocenter Count |
1
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| SMILES |
OC(=O)CCC(C(O)=O)NC(=O)C1=CC=C(C=C1)N(C=O)CC2=NC3=C(N=C2)N=C(N)N=C3=O
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| InChi Key |
UGWUWNVTCLDEOG-ZDUSSCGKSA-N
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| InChi Code |
InChI=1S/C20H19N7O7/c21-20-25-16-15(18(32)26-20)23-11(7-22-16)8-27(9-28)12-3-1-10(2-4-12)17(31)24-13(19(33)34)5-6-14(29)30/h1-4,7,9,13H,5-6,8H2,(H,24,31)(H,29,30)(H,33,34)(H3,21,22,25,26,32)/t13-/m0/s1
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| Chemical Name |
(2S)-2-[[4-[(2-amino-4-oxo-3H-pteridin-6-yl)methyl-formylamino]benzoyl]amino]pentanedioic acid
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| Synonyms |
10-Formylfolic acid; 134-05-4; 10-FORMYLFOLIC ACID (25 MG); N10-Formylfolic Acid; formylfolic acid; 10-Formylpteroylglutamic Acid; 10-Formyl Folic Acid; AI902R79R1;
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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) |
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.) |
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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