135-16-0

Basic information

• Product Name: TETRAHYDROFOLIC ACID
• Synonyms: L-5,6,7,8-Tetrahydrofolic acid;Tetrahydropteroylglutaamic acid;(6S)-Tetrahydrofolic acid;(6S)-Thfa;C00101;Tetrahydrofolic acid,5,6,7,8-Tetrahydropteroyl-L-glutamic acid, THF;L-Tetrahydrofolic Acid, synthesized on receipt of order. 95% MiniMuM purity when shipped.;N-[4-[[(2-AMino-3,4,5,6,7,8-hexahydro-4-oxo-6-pteridinyl)Methyl]aMino]benzoyl]-L-glutaMic Acid
• CAS NO: 135-16-0
• Molecular Formula: C₁₉H₂₃N₇O₆
• Molecular Weight: 445.43
• EINECS: 205-181-1
• Product Categories: Chiral Reagents;Intermediates & Fine Chemicals;Pharmaceuticals;co-factor
• Mol File: 135-16-0.mol

Chemical Properties

• Boiling Point: 555.12°C (rough estimate)
• Appearance: off-white to tan/powder
• Density: 1.4216 (rough estimate)
• Vapor Pressure: 0Pa at 25℃
• Refractive Index: 1.6800 (estimate)
• Storage Temp.: −20°C
• Solubility: Aqueous Base (Slightly), DMSO (Slightly), Methanol (Slightly, Heated, Sonicated)
• PKA: 3.51±0.10(Predicted)
• Stability: We have observed that this material decomposes steadily over time. Use immediately upon receipt.
• BRN: 9238187
• CAS DataBase Reference: TETRAHYDROFOLIC ACID(CAS DataBase Reference)
• NIST Chemistry Reference: TETRAHYDROFOLIC ACID(135-16-0)
• EPA Substance Registry System: TETRAHYDROFOLIC ACID(135-16-0)

Safety Data

• WGK Germany: 3
• RTECS: MA0600800
• F: 3-8-10
• Hazardous Substances Data: 135-16-0(Hazardous Substances Data)

Usage

Uses

Used in Metabolism of Amino and Nucleic Acids:
TETRAHYDROFOLIC ACID is used as a coenzyme for the metabolism of amino and nucleic acids, playing a vital role in various metabolic reactions.
Used in Pharmaceutical Research:
TETRAHYDROFOLIC ACID is used as a research compound for determining the inhibitory effect of tetrahydrofolate on polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs), which can help in understanding the underlying mechanisms of certain diseases and their potential treatments.
Used in Molecular Biology:
In the field of molecular biology, TETRAHYDROFOLIC ACID is used for studying the activation of riboswitches, which are RNA molecules that can control gene expression in response to specific molecular signals.

Flammability and Explosibility

Notclassified

Biochem/physiol Actions

Tetrahydrofolic acid is the parent molecule of the folate derivatives that donate one-carbon units to amino acids, nucleic acids, and lipids. Tetrahydrofolate metabolites participate in the synthesis of purine and pyrimidine and in synthesis and conversion of amino acids

Purification Methods

Very high quality material is now available commercially, and it should be a white powder. It can be dried over P2O5 in a vacuum desiccator and stored in weighed aliquots in sealed ampoules. It is stable at room temperature in sealed ampoules for many months and for much more extended periods at -10o. When moist, it is extremely sensitive to air whereby it oxidises to the yellow 7,8-dihydro derivative. In solution it turns yellow in colour as it oxidises, and then particularly in the presence of acids it turns dark reddish brown in colour. Hence aqueous solutions should be frozen immediately when not in use. It is always advisable to add 2-mercaptoethanol (if it does not interfere with the procedure for which it is used) which stabilises it by depleting the solution of O2. The sulfate salt is more stable but is much less soluble. The best way to prepare standard solutions of this acid is to dissolve it in the desired buffer and estimate the concentration by UV absorption in pH 7 buffer at 297nm ( 22,000 M-1cm-1). If a sample is suspect, it is not advisable to purify it because it is likely to deteriorate further as “dry box” conditions are necessary. Either a new sample is purchased or one is freshly prepared from folic acid. It has the above pKa values. [Hafeti et al. Biochemical Preparations 7 89 1960, UV: Mathews & Huennekens J Biol Chem 235 3304 1960, Osborn & Huennekens J Biol Chem 233 969 1958, O'Dell et al. J Am Chem Soc 69 250 1947, Blakley Biochem J 6 5 331 1957, Asahi Yakugaku Zasshi (J Pharm Soc Jpn) 79 1548 1959, Beilstein 26 III/IV 3879.]

InChI:InChI=1/C19H23N7O6/c20-19-25-15-14(17(30)26-19)23-11(8-22-15)7-21-10-3-1-9(2-4-10)16(29)24-12(18(31)32)5-6-13(27)28/h1-4,11-12,21,23H,5-8H2,(H,24,29)(H,27,28)(H,31,32)(H4,20,22,25,26,30)/t11-,12-/m0/s1

Upstream product

• 59-30-3 folic acid 
• 4033-27-6 (S)-2-{4-[(2-Amino-4-oxo-3,4,7,8-tetrahydro-pteridin-6-ylmethyl)-amino]-benzoylamino}-pentanedioic acid 
• 15574-38-6 dihydrofolic acid 
• 142811-51-6 N-<4-<<2(S)-amino-3-<(2,5-diamino-4(3H)-oxopyrimidin-6-yl)amino>propyl>amino>benzoyl>-L-glutamic acid 
• 142811-50-5 N-<4-<<2(S)-(benzylamino)-3-<(2-amino-5-nitro-4(3H)-oxopyrimidin-6-yl)amino>propyl>amino>benzoyl>-L-glutamic acid 
• 59-30-3 folate 
• 86-01-1 Guanosine 5'-triphosphate 
• 1218-98-0 dihydro-7,8-D-neopterine 
• 73-40-5 2-amino-1,9-dihydro-6H-purin-6-one 
• 4271-30-1 N-(4-aminobenzoyl)-L-glutamic acid 
• 3672-03-5 7,8-dihydro-6-(hydroxymethyl)pterin 
• 3545-84-4 6-hydroxymethyl-7,8-dihydro-pterin pyrophosphate 
• 142811-48-1 [(R)-2-(2-Amino-5-nitro-6-oxo-1,6-dihydro-pyrimidin-4-ylamino)-1-formyl-ethyl]-benzyl-carbamic acid tert-butyl ester 
• 142811-47-0 2-amino-6-<<2'(R)--3'-hydroxypropyl>amino>-5-nitro-4(3H)-pyrimidinone 

Downstream Products

• 3432-99-3 N⁵,N¹⁰-methylene-5,6,7,8-tetrahydrofolate 
• 2800-34-2 10-formyltetrahydrofolate 
• 58-05-9 folinic acid 
• 28459-40-7 N-(10-formyl-7,8-dihydro-pteroyl)-L-glutamic acid 
• 113974-18-8 N-((S)-5-formimidoyl-5,6,7,8-tetrahydro-pteroyl)-L-glutamic acid 
• 72988-03-5 (S)-2-(4-{[2-Amino-5-(2-chloro-ethylcarbamoyl)-4-oxo-3,4,5,6,7,8-hexahydro-pteridin-6-ylmethyl]-amino}-benzoylamino)-pentanedioic acid 
• 72973-87-6 (S)-2-{4-[(2-Amino-5-carbamoyl-4-oxo-3,4,5,6,7,8-hexahydro-pteridin-6-ylmethyl)-amino]-benzoylamino}-pentanedioic acid 
• 72973-88-7 (S)-2-{4-[(2-Amino-5-ethylthiocarbamoyl-4-oxo-3,4,5,6,7,8-hexahydro-pteridin-6-ylmethyl)-amino]-benzoylamino}-pentanedioic acid 
• 111848-30-7 C₃₁H₄₂N₈O₉^(2-)*Ca⁽²⁺⁾ 
• 111823-26-8 (S)-2-(4-{[2-Amino-5-(5-ethoxycarbonyl-pentylthiocarbamoyl)-4-oxo-3,4,5,6,7,8-hexahydro-pteridin-6-ylmethyl]-amino}-benzoylamino)-pentanedioic acid 
• 111823-24-6 5-<<<6-(ethoxycarbonyl)hexyl>amino>carbonyl>-5,6,7,8-tetrahydrofolic acid 
• 111823-25-7 (S)-2-(4-{[2-Amino-5-(7-ethoxycarbonyl-heptylcarbamoyl)-4-oxo-3,4,5,6,7,8-hexahydro-pteridin-6-ylmethyl]-amino}-benzoylamino)-pentanedioic acid 
• 111823-23-5 (S)-2-(4-{[2-Amino-5-(5-ethoxycarbonyl-pentylcarbamoyl)-4-oxo-3,4,5,6,7,8-hexahydro-pteridin-6-ylmethyl]-amino}-benzoylamino)-pentanedioic acid 
• 111612-17-0 5-(-)-menthyloxycarbonyl-(6R)-tetrahydrofolic acid 

Relevant articles and documents

Cloning and characterization of a novel, plasmid-encoded trimethoprim- resistant dihydrofolate reductase from Staphylococcus haemolyticus MUR313

In recent years resistance to the antibacterial agent trimethoprim (Tmp) has become more widespread, and several trimethoprim-resistant (Tmp(r)) dihydrofolate reductases (DHFRs) have been described from gram-negative bacteria. In staphylococci, only one Tmp(r) DHFR has been described, the type S1 DHFR, which is encoded by the dfrA gene found on transposon Tn4003. In order to investigate the coincidence of high-level Tmp resistance and the presence of dfrA, we analyzed the DNAs from various Tmp(r) staphylococci for the presence of dfrA sequences by PCR with primers specific for the thyE- dfrA genes from Tn4003. We found that 30 of 33 isolates highly resistant to Tmp (MICs, ≥512 μg/ml) contained dfrA sequences, whereas among the Tmp(r) (MICs, ≤256 μg/ml) and Tmp(r) isolates only the Staphylococcus epidermidis isolates (both Tmp(r) and Tmp(r)) seemed to contain the dfrA gene. Furthermore, we have cloned and characterized a novel, plasmid-encoded Tmp(r) DHFR from Staphylococcus haemolyticus MUR313. The dfrD gene of plasmid pABU17 is preceded by two putative Shine-Dalgarno sequences potentially allowing for the start of translation at two triplets separated by nine nucleotides. The predicted protein of 166 amino acids, designated S2DHFR, encoded by the longer open reading frame was overproduced in Escherichia coli, purified, and characterized. The molecular size of the recombinant S2DHFR was determined by ion spray mass spectrometry to be 19,821.2 ± 2 Da, which is in agreement with the theoretical value of 19,822 Da. In addition, the recombinant S2DHFR was shown to exhibit DHFR activity and to be highly resistant to Tmp.

Influence of the Debye length on the interaction of a small molecule-modified Au nanoparticle with a surface-bound bioreceptor

We report that a shorter Debye length and, as a consequence, decreased colloidal stability are required for the molecular interaction of folic acid-modified Au nanoparticles (Au NPs) to occur on a surface-bound receptor, human dihydrofolate reductase (hDHFR). The interaction measured using surface plasmon resonance (SPR) sensing was optimal in a phosphate buffer at pH 6 and ionic strength exceeding 300 mM. Under these conditions, the aggregation constant of the Au NPs was approximately 104 M-1 s -1 and the Debye length was below 1 nm, on the same length scale as the size of the folate anion (approximately 0.8 nm). Longer Debye lengths led to poorer SPR responses, revealing a reduced affinity of the folic acid-modified Au NPs for hDHFR. While high colloidal stability of Au NPs is desired in most applications, these conditions may hinder molecular interactions due to Debye lengths exceeding the size of the ligand and thus preventing close interactions with the surface-bound molecular receptor.

Screening of inhibitors using enzymes entrapped in sol-gel-derived materials

In recent years, a number of new methods have been reported that make use of immobilized enzymes either on microarrays or in bioaffinity columns for high-throughput screening of compound libraries. A key question that arises in such methods is whether immobilization may alter the intrinsic catalytic and inhibition constants of the enzyme. Herein, we examine how immobilization within sol-gel-derived materials affects the catalytic constant (kcat), Michaelis constant (KM), and inhibition constant (KI) of the clinically relevant enzymes Factor Xa, dihydro-folate reductase, cyclooxygenase-2, and γ-glutamyl transpeptidase. These enzymes were encapsulated into solgel-derived glasses produced from either tetraethyl orthosilicate (TEOS) or the newly developed silica precursor diglyceryl silane (DGS). It was found that the catalytic efficiency and long-term stability of all enzymes were improved upon entrapment into DGS-derived materials relative to entrapment in TEOS-based glasses, likely owing to the liberation of the biocompatible reagent glycerol from DGS. The KM values of enzymes entrapped in DGS-derived materials were typically higher than those in solution, whereas upon entrapment, kcat values were generally lowered by a factor of 1.5-7 relative to the value in solution, indicating that substrate turnover was limited by partitioning effects or diffusion through the silica matrix. Nonetheless, the apparent KI value for the entrapped enzyme was in most cases within error of the value in solution, and even in the worst case, the values differed by no more than a factor of 3. The implications of these findings for high-throughput screening are discussed.

Enantioselective catalyses CXXXV [1]. Stereoselective hydrogenation of folic acid and 2-methylquinoxaline with optically active rhodium(I)-phosphane complexes

In the hydrogenation of the C=N double bonds of the pyrazine ring of folic acid to 5,6,7,8-tetrahydrofolic acid a new asymmetric center is formed at C6 of the pteridine system. With rhodium(I) catalysts made from optically active phosphanes, which are immobilized on silical gel, the hydrogenation in aqueous solution can be controlled stereoselectively. The highest diastereomeric excess of ca. 40% is obtained with (-)-BPPM containing catalysts. The hydrogenation of the biomolecule folic acid in aqueous solution is also possible homogeneously with rhodium(I)-phosphane catalysts, the ligands of which contain sulfonic acid groups and polyether fragments. The homogeneous hydrogenations proceed slower and with somewhat reduced diastereoselectivities compared to the heterogeneous catalyses. The hydrogenation of 2-methylquinoxaline is a model system for the reduction of folic acid. The usual rhodium(I)-phosphane catalysts afford only small enantioselectivities.

Trimethoprim resistance of dihydrofolate reductase variants from clinical isolates of Pneumocystis jirovecii

Pneumocystis jirovecii is an opportunistic pathogen that causes serious pneumonia in immunosuppressed patients. Standard therapy and prophylaxis include trimethoprim (TMP)-sulfamethoxazole; trimethoprim in this combination targets dihydrofolate reductase (DHFR). Fourteen clinically observed variants of P. jirovecii DHFR were produced recombinantly to allow exploration of the causes of clinically observed failure of therapy and prophylaxis that includes trimethoprim. Six DHFR variants (S31F, F36C, L65P, A67V, V79I, and I158V) showed resistance to inhibition by trimethoprim, with Ki values for trimethoprim 4-fold to 100-fold higher than those for the wild-type P. jirovecii DHFR. An experimental antifolate with more conformational flexibility than trimethoprim showed strong activity against one trimethoprim-resistant variant. The two variants that were most resistant to trimethoprim (F36C and L65P) also had increased Km values for dihydrofolic acid (DHFA). The catalytic rate constant (kcat) was unchanged for most variant forms of P. jirovecii DHFR but was significantly lowered in F36C protein; one naturally occurring variant with two amino acid substitutions (S106P and E127G) showed a doubling of kcat, as well as a Km for NADPH half that of the wild type. The strongest resistance to trimethoprim occurred with amino acid changes in the binding pocket for DHFA or trimethoprim, and the strongest effect on binding of NADPH was linked to a mutation involved in binding the phosphate group of the cofactor. This study marks the first confirmation that naturally occurring mutations in the gene for DHFR from P. jirovecii produce variant forms of DHFR that are resistant to trimethoprim and may contribute to clinically observed failures of standard therapy or prophylaxis. Copyright

Osmolyte induced enhancement of expression and solubility of human dihydrofolate reductase: An in vivo study

The process of recombinant protein production in E. coli system is often hampered by the formation of insoluble aggregates. Human Dihydrofolate reductase (hDHFR), an enzyme involved in the synthesis of purine, thymidilate and several other amino acids like glycine, methionine and serine is highly aggregation prone. It catalyzes the reduction of dihydrofolate (H2F) in order to regenerate tetrahydrofolate (H4F) utilizing NADPH as a cofactor. We have attempted to ameliorate the production of soluble and functional protein by growing and inducing the cells under osmotic stress condition, in the presence of various osmolytes like glycerol, sorbitol, TMAO, proline and glycine at 37?°C. The expression and yield of functional hDHFR protein were highly enhanced in the presence of these osmolytes. The specific activity of the purified recombinant hDHFR protein has also been increased to a cogent level in the presence of osmolytes. We also observed that protein expressed in presence of the osmolytes was stable in the denaturing conditions as compared to the protein expressed in absence of an osmolyte. We also observed using the intrinsic fluorescence spectroscopy that the osmolytes didn't interfere with the structure of the protein and in denaturing conditions the protein expressed in presence of osmolytes had more stability. Our study is consequential in increasing the production of functional and soluble protein in the cell extract and will also be appropriate to find a therapeutic agent against many neurodegenerative diseases.

Aqueous diastereoselective hydrogenation of folic acid to tetrahydrofolic acid in the presence of water-soluble Rh and Ir diphosphine complexes

Rhodium and iridium catalysts with chiral, water soluble diphosphine ligands, were used for the diastereoselective hydrogenation of folic acid disodium salt in water. Using a modified Rh/Josiphos type at 30 °C, l-tetrahydrofolic acid, a relevant pharmaceutical intermediate, was obtained with a selectivity of up to 49% de; at 70 °C turnover numbers of up to 2800 were achieved, albeit with lower selectivity. These results define the state of the art for this reaction.

Process for preparing L-5 - methyl tetrahydrofolinate

The invention relates to a preparation process of L-5 - methyl tetrahydrofolic acid, which adopts asymmetric catalytic hydrogenation to convert folic acid into (6S) - tetrahydro folic acid, and has high folic acid conversion rate, and (6S) - tetrahydrofolic acid diastereomeric excess degree. By salt formation and crystallization, the (6S) - tetrahydrofolic acid intermediate with extremely high diastereomeric excess can be easily enriched, and thus the yield and purity of L-5 - methyltetrahydrofolinate finally obtained are high. The catalytic hydrogenation process is very mature in industrial application and convenient to operate. The subsequent methylation and salt formation steps are easy to implement, and the preparation process has high economic value and practical value under the condition that the yield and purity of the product are improved.

Chemoenzymatic Assembly of Isotopically Labeled Folates

Pterin-containing natural products have diverse functions in life, but an efficient and easy scheme for their in vitro synthesis is not available. Here we report a chemoenzymatic 14-step, one-pot synthesis that can be used to generate 13C- and 15N-labeled dihydrofolates (H2F) from glucose, guanine, and p-aminobenzoyl-l-glutamic acid. This synthesis stands out from previous approaches to produce H2F in that the average yield of each step is >91% and it requires only a single purification step. The use of a one-pot reaction allowed us to overcome potential problems with individual steps during the synthesis. The availability of labeled dihydrofolates allowed the measurement of heavy-atom isotope effects for the reaction catalyzed by the drug target dihydrofolate reductase and established that protonation at N5 of H2F and hydride transfer to C6 occur in a stepwise mechanism. This chemoenzymatic pterin synthesis can be applied to the efficient production of other folates and a range of other natural compounds with applications in nutritional, medical, and cell-biological research.

A novel synthetic method for preparation of some folates

An improved method was developed for preparation of 5,6,7,8-tetrahydrofolic acid (THF) and calcium-5-methyltetrahydrofolate (5-MTHF-Ca) by reduction of folic acid using KBH4 catalyzed by Pb(NO3)2. The yields of THF and 5-MTHF-Ca were 56.5 and 42.7 %, respectively. A convenient method for measurement of THF and 5-MTHF-Ca using liquid chromatography-mass spectrometry (LC-MS) was also established, enabling analysis of those folates within 10 min without application of gradient elution.