EP2953935A2

EP2953935A2 - Method for producing pyrazoles, novel pyrazoles and applications thereof

Info

Publication number: EP2953935A2
Application number: EP14709725.7A
Authority: EP
Inventors: Frédéric TARAN; Manon Chaumontet; Sergii KOLODYCH; Evelia RASOLOFONJATOVO ANDOVOLA
Original assignee: Commissariat a lEnergie Atomique et aux Energies Alternatives CEA
Current assignee: Commissariat a lEnergie Atomique et aux Energies Alternatives CEA
Priority date: 2013-02-11
Filing date: 2014-02-07
Publication date: 2015-12-16
Also published as: WO2014122407A3; US20150368295A1; FR3001967B1; US9802980B2; FR3001967A1; WO2014122407A2

Abstract

A method for preparing a pyrazole of formula (I) in which R,R' and R'' have different meanings, characterised in that it involves reacting a sydnone of formula (II) in which R and R' have the meanings already indicated, with an alkyne of formula (III) in which R'' has the meaning already indicated, in the presence of copper, to obtain a pyrazole compound of formula (I) that is then isolated and salified if desired.

Description

Process for producing pyrazoles, new pyrazoles and their

applications

The present invention relates to a method of manufacturing pyrazoles, new pyrazoles impossible to manufacture to date, and their applications.

There are many methods of forming pyrazoles. These can be summarized in two main solutions as described in the recent review by Antonio Simon-Fuentes, Chem. Rev. 2011, 111, 6984-7034.

A first solution consists in using the cyclo-condensation of 1,3-dielectrophiles, such as beta-diketones, with hydrazines. This solution has been known for many years and is probably the most widely used today for the synthesis of pyrazoles. However, this method has a number of limitations. In particular, it can not be applied to all the substrates, in particular the hydrazine substrates having nucleophilic groups such as amines can not be used. Similarly, dielectrophilic substrates having other electrophilic centers, such as aldehydes, ketones or Michael acceptors are to be avoided. In addition, the reaction conditions most often require the use of acids and / or high temperatures.

The second solution is to prepare the pyrazoles by 1,3-dipolar cycloaddition from alkynes and dipoles such as diazoalkanes, nitrilimines or sydnones. The reaction with the diazo alkanes and the nitrilimines requires strong base conditions and the use of organic solvents which is a significant disadvantage when fragile substrates and / or solely soluble in aqueous medium must be employed. In addition, these reactions generate two regioisomers that need to be separated.

The reaction with sydnones is described in the literature according to two solutions.

A first solution is to prepare the pyrazoles thermally. Thermal sydnone / alkyne cycloaddition has been known for many years and has been described in particular by Kirsi Harju, Johanna Vesterinen and Jari Yli-Kauhaluoma, Org. Lett. 2009, 10, 2219-2221 and by Duncan L. Browne, Matthew D. Helm, Andrew Plant, and Joseph PA Harrity, Angew. Chem. Int. Ed. 2007, 46, 8656-8658. The temperatures required for the reaction are typically greater than 120 ° C.

Another solution, described for example by Wu Chunrui, Yuesi Fang, Richard C. Larock and Feng Shi, Org. Lett. 2010, 10, 2234-2237 or by Yuesi Fang, Wu Chunrui, Richard C. Larock and Feng Shi, J. Org. Chem. 2011, 76, 8840-8851, consists of preparing the pyrazoles from very reactive alkynes. The sydnone / alkyne cycloaddition can then be carried out at room temperature when aromatic compounds called arynes having an extremely reactive triple bond are used. Arynes are not stable and must be prepared in situ from appropriate precursors.

Each of these solutions has significant disadvantages.

The disadvantages of the first solution are related to the drastic conditions of preparation. High temperatures, typically of the order of 150 ° C, are incompatible with many fragile substrates. The thermal reaction does not tolerate the presence of many reaction functions. More regioisomeric products are formed, which reduces the synthesis yields and requires tedious purifications.

The second solution uses very particular alkynes. It is therefore inherently limited in terms of the diversity of substrates that can be used. Only pyrazoles called 2H-lndazoles can be synthesized by this reaction.

It would therefore be desirable to have a method that allows the manufacture of a variety of pyrazoles, including fragile or reactive groups. It would also be desirable for this process to be as general as possible and compatible with complex molecules and biological media.

On the other hand, to increase the reaction yield, it would be desirable for this process to be regio- and chemo-selective.

Chunrui Wu also describes in TETRAHEDRON LETTERS, vol. 52, No. 29, July 201 1 Palladium-catalyzed oxidative coupling between mono-substituted sydnones and alkynes in toluene at 75 ° C. In certain reactions, a cupric salt combined with a silver-based compound is used to regenerate palladium II. In attempting to prepare acetylenic compounds, the author has by chance obtained a pyridyl-substituted pyrazole.

After extensive research, the inventors have developed a method of manufacturing pyrazoles, including the preparation of new pyrazoles impossible to manufacture to date.

These pyrazoles correspond to formula I

and have virtually no limitation as to the nature of the substituents R and R "which can be of a very large variety, ranging from simple alkyl to proteins as will be seen hereinafter in the examples.

This is why the present application relates to a process for preparing a pyrazole of formula I

in which

R represents a hydrogen atom, a halogen atom, a thio or thioalkyl group, a hydroxyl or alkoxy group, an amino or aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is unsubstituted; or substituted one or more times with F, Cl, Br, I, CN, OH, NH ₂ , NH-C 1 -C 8 alkyl, N- (C 1 -C 8 alkyl) ₂ , SH, an aryl radical containing 6 to 10 carbon atoms. carbon which is unsubstituted or substituted one or more times with an alkyl or alkoxy radical containing 1 to 8 carbon atoms, a heterocycle, an amino acid, a polypeptide, a protein, a nucleic acid, a DNA molecule, a polysaccharide , a polymer, a nanoparticle, a dendrimer.

R 'represents a hydrogen atom, a halogen atom, a thio or -thioalkyl group, a hydroxyl or alkoxy group, an amino or -aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is not substituted or substituted one or more times with F, Cl, Br, I, CN, OH, NH ₂ , NH-C 1 -C 8 alkyl, N- (C 1 -C 8 alkyl) ₂ , SH, an aryl radical containing 6 to 10 atoms of carbon which is unsubstituted or substituted one or more times with an alkyl or alkoxy radical containing 1 to 8 carbon atoms, a heterocycle,

R "represents a hydrogen atom, a halogen atom, a thio group, a hydroxyl group, an amino group, a hydrocarbon group or a hydrocarbon group containing from 1 to 100 heteroatoms, R" particularly representing an atom of hydrogen, a halogen atom, a thio or thioalkyl group, a hydroxy or alkoxy group, an amino or -aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is unsubstituted or substituted one or more times with Substituted or unsubstituted F, Cl, Br, I, CN, OH, O-heterocycle, NH ₂ , -NH-C 1 -C 8 alkyl, -N- (C 1 -C 8 alkyl) ₂ , -NH-R '", or - Alkyl C1 -C3-NH-R '"where R'" represents a sulfonated group, SH, an aryl radical containing 6 to 10 carbon atoms which is unsubstituted or substituted one or more times with an alkyl or aikoxy radical containing 1 to 8 carbon atoms, a heterocycle, an amino acid, a polypeptide, a protein, a nucleic acid, a DNA molecule, a polysaccharide, a polymer, a nanoparticle, a dendrimer,

characterized in that a sydnone of formula II is reacted

in which R and R 'have the meanings already indicated, with an alkyne of formula III

wherein R "has the meaning already indicated, in the presence of copper used as a metal catalyst, to obtain a pyrazole compound of formula I which is isolated and salified if desired.

The halogen atom may be a bromine or iodine atom, and preferably a chlorine atom.

The alkyl radical is straight chain or branched chain and may contain 1 to 8 carbon atoms and preferably 1 to 5 carbon atoms. Suitable alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, sec-butyl and tert-butyl. It is the same for the alkyl radical of the alkylthio group, alkoxy or alkylamino.

The term aryl refers to an aromatic carbocyclic radical containing 6 to 10 carbon atoms unless otherwise indicated. Suitable aryl groups include phenyl, naphthyl and biphenyl. The substituted aryl groups include the aryl groups described above which are substituted one or more times, preferably once with halogen, alkyl, hydroxy, alkoxy, nitro, methylenedioxy, ethylenedioxy, amino, alkylamino, dialkylamino, hydroxyalkyl, hydroxyalkoxy, carboxy.

The heterocycle may be a saturated, partially saturated and fully unsaturated heterocyclic group having one, two or three rings and a total number of 5 to 10 ring atoms wherein at least one of the ring atoms is an N, O atom. or S. Preferably, the heterocyclic group contains from 1 to 3 ring heteroatoms selected from N, O and S. Suitable saturated and partially saturated heterocyclic rings include tetrahydrofuranyl, tetrahydrothienyl, tetrahydropyranyl, dihydropyranyl, pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl, oxoazolinyl, isoxazolinyl, and the like. Suitable heteroaryl groups include furyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, pyridyl, pyrimidinyl, benzopyranyl, indolyl, quinolinyl, isoquinolinyl, naphthyridinyl, and the like. Other examples of suitable heterocyclic rings are 2-quinolinyl, 1,3-benzodioxyl, 2-thienyl, 2-benzofuranyl, 2-benzothiophenyl, 3-thienyl, 2,3-benzenediyl, 5-dihydro-5-benzofuranyl, 4-indoyl, 4-pyridyl, 3-quinolinyl, 4-quinolinyl, 1,4-benzodioxan-6-yl, 3-indoyl, 2-pyrrolyl, benzopyran, 6-yl, a 5-indolyl, a 1,5-benzoxepin-8-yl, a 3-pyridyl, a 6-coumarinyl, a 5-benzofuranyl, a 2-isoimidazol-4-yl, a 3-pyrazolyl, a 3-carbazolyl, 2-thiazolyl, 2-oxazolyl, 1-imidazolyl and 2-imidazolyl.

The heterocycle may be substituted in one or more substitutable sites with, for example, halogen, aryl, C1-C5 alkyl, hydroxy, C1-C5 alkoxy, cyano, trifluoromethyl, nitro, an oxo, a amino, a C1-C5 alkyl amino or a C1-C5 amino dialkyl or even another heterocycle which itself may also be substituted.

The radicals which are substituted one or more times preferably contain 1 to 3 substituents.

A polypeptide may for example be angotensin II, substance P, neurokine A, calcitonin, oxytocin.

A protein may be for example an antibody, an enzyme, hemoglobin, transferrin, streptavidin.

A nucleic acid may be for example a molecule consisting of DNA or RNA.

A polysaccharide may be for example chitosan, cellulose, alginates, dextran.

A polymer may be for example polyethylene glycol (PEG), polystyrene, polypropylene, polyethylene, polyacrylamide.

A nanoparticle may be selected for example parm i quantum dots, carbon nanotubes, metal nanoparticles (Au, Fe ₃ O _4, Cr ₂ O _3, Al ₂ O _3, BaFei ₂ Oi _9).

A dendrimer may be chosen for example from PAMAM dendrimers and Cyclotriphosphazene-PMMH.

When R '"represents a sulfonated group, it is preferably one of the sulfonated groups illustrated by the examples.

In the present application and in what it says, the expression "hydrocarbon group" designates a linear, branched, cyclic or polycyclic hydrocarbon group, these cycles being able to be fused or not, containing from 1 to 100 heteroatoms, these groups being able to be unsubstituted or substituted by one or more atoms or groups such as those mentioned for the radicals R 'and R ".

It should be noted that the sydnones of formula II are dipoles existing in several tautomeric forms. Only one of the forms is represented above.

Under preferred conditions of implementation of the method described above, R represents an optionally substituted aryl or heteroaryl radical, in particular an optionally substituted aryl radical, particularly an optionally substituted phenyl radical, these different radicals and substituent having in particular the meanings indicated above.

In other preferred conditions of implementation of the method described above, R 'represents a hydrogen atom.

In still other preferred conditions of implementation of the method described above, R "represents a heteroaryl radical or an aryl radical or an alkyl radical containing 1 to 8 carbon atoms, preferably one of the last two .

Under particularly preferred conditions, R and R ', R' and R ", R and R", and R, R 'and R "have the preferred values indicated above.

The catalyst metal of the reaction is copper. Copper forms that can be used in the process are, for example, and preferably either copper (I) salts such as Cul, CuOTf, CuCl, CuOAc, etc. and preferably Cul, or copper (II) salts such as CuSO ₄ , CuOTf ₂ , CuOAc ₂ , ... and preferably CuSO ₄ reduced in situ to Cu (1) salts by a reducing agent such as ascorbic acid, sodium bisulfite, zinc or metallic copper, and preferably ascorbic acid.

The copper salts can either be used as such or preferably in complexed form with ligands, preferably nitrogenous, such as the phenanthroline compounds such as IV below, triazoles such as V or VII below, pyridines such as IX and X below, of imidazoles such as VI II below, benzimidazoles such as VI below and phosphines, preferably nitrogen ligands such as those mentioned above, particularly phenanthrolines. These nitrogen ligands allow an increase in the catalytic activity of the copper but also the regioselectivity of the reaction, in particular in the case where R 'is different from H.

The above copper salts are preferably reduced in situ to salts having a lower oxidation state.

The solvents which can be used for the implementation of the present process can be organic solvents, water or organo-aqueous mixtures. It should be noted that quite remarkably, an optionally complex physiological fluid such as plasma, urine, cell lysates or cell culture media can be used as a solvent medium in the present invention, without disturbing the reaction.

The present process is insensitive to the pH of the editorial medium. However, it is preferred to operate in a neutral medium, or particularly in an alkaline medium. For this purpose, for example, an organic base such as triethylamine or triethanolamine can be used.

The reaction of a sydnone of formula II with an alkyne of formula III is preferably carried out between 10 ° C and 120 ° C, preferably between 15 ° C and 100 ° C, particularly between 15 ° C and 70 ° C. For the preparation of pyrazole compounds of formula I which are sensitive or fragile such as a polypeptide, a protein, a nucleic acid, a DNA molecule, a polysaccharide, a nanoparticle or a dendrimer, the procedure is preferably carried out between 15 ° and 50 °. C, preferably between 20 and 40 ° C. When the pyrazole compound of formula I is not particularly sensitive or fragile, it is preferably carried out between 45 and 72 ° C, preferably between 55 and 65 ° C.

The methods that are the subject of the present invention possess very interesting properties. They notably enjoy remarkable reaction yields, generally greater than 95%. They benefit from total regioselectivity, ie only one regioisomer is formed during the reaction.

To prepare the pyrazoles of formula I, starting reactants having functional groups can be reacted without altering them.

The reaction is applicable regardless of the structure of the reagents defined above implemented.

The reaction is carried out under mild reaction conditions, compatible in particular with fragile molecules or biomolecules such as proteins, enzymes, DNA molecules, etc. These properties are illustrated below in the experimental part.

They justify the use of the methods described above, in the preparation of original compounds of pyrazole, hitherto inaccessible.

Therefore, the present application also relates to pyrazole compounds of formula I described above, wherein R 'and R "have the meaning and preferred values already indicated and R represents a polypeptide such as, for example, angiotensin II, substance P, neurokine A, calcitonin, oxytocin, a protein such as an antibody, an enzyme, hemoglobin, transferrin, streptavidin, a nucleic acid such as for example a molecule consisting of DNA or RNA, a polysaccharide such as chitosan, cellulose, alginates, dextran, a polymer such as polyethylene glycol (PEG), polystyrene, polypropylene, polyethylene, polyacrylamide, a nanoparticle as per examples of quantum dots, carbon nanotubes, metal nanoparticles (Au, Fe3O ₄ , Cr ₂ O ₃ , Al ₂ O 3,

a dendrimer such as PAMAM dendrimers, Cyclotriphosphazene-PMMH.

R "in this case preferably represents a radical having a fluorophore function, for example cyanines Cy3 and Cy5, Alexa Fluor derivatives, coumarin, fluorescein and rhodamine derivatives such as tetramethylrhodamine (TAMRA), a radiotracer such as for example the DOTA-Gd, DOTA-G a and DOTA- ⁶⁴ Cu complexes or a molecule containing an atom ¹⁸ F or ¹²⁵ l, an active principle of human therapy (medicament) such as taxol, taxotere, navel bi ne, vinblastine, a molecule capable of interacting with a protein or DNA, for example biotin, tetraacetylfucose, RGD peptides, anthracyclines.

The present application also relates to pyrazole compounds of formula I described above, in which R and R 'have the meaning and the preferred values already indicated and R "represents a polypeptide such as, for example, angiotensin II, P, neurokin A, calcitonin, oxytocin, a protein such as an antibody, an enzyme, hemoglobin, transferrin, streptavidin, a nucleic acid such as a molecule consisting of DNA or RNA, a polysaccharide such as chitosan, cellulose, alginates, dextran, a polymer such as polyethylene glycol (PEG), polystyrene, polypropylene, polyethylene, polyacrylamide , a nanoparticle such as quantum dots, carbon nanotubes, metal nanoparticles (Au, Fe3O ₄ , Cr ₂ O ₃ , Al ₂ O 3, a dendrimer such as PAMAM dendrimers, cyclotriphosphazene-PMMH.

In this case, R preferably represents a radical having a fluorophore function, for example cyanines C y 3 and C y 5, Alexa Fluor derivatives, coumarin, fluorescein and rhodamine derivatives such as tetramethylrhodamine (TAMRA). , a radiotracer such as for example the complexes of DOTA-Gd, DOTA-Ga and DOTA- ⁶⁴ Cu or a molecule comprising an ¹⁸ F or ¹²⁵ I atom, an active principle of human therapy (drug) such as for example taxol, taxotere, navel bine, vinblastine; a molecule capable of interacting with a protein or DNA such as, for example, biotin, tetraacetylfucose, RGD peptides, anthracyclines.

The present invention is not intended, either with regard to the aforementioned preparation process or with regard to new pyrazole compounds of formula I above, Chunrui Wu 1 - (4-chlorophenyl) -4-pyridyl-2 pyrazole.

The novel pyrazole compounds of formula I described above which are the subject of the present invention possess very interesting properties. They are endowed notably with remarkable properties such as: i) a high detection sensitivity in the case of pyrazoles having a group R or R "= fluorophore or radiotracer, / ^' / ^' ) the ability to effectively bind a biomolecule of interest in the case of pyrazoles having a group R or R "= active principle or a molecule capable of interacting with a protein or DNA.

These properties are illustrated below in the experimental part. They justify the use of the novel pyrazole compounds described above, in many applications such as protein detection. Therefore, the present application also relates to the use of the novel pyrazole compounds described above as: f) biological tracers for medical imaging or diagnosis (Thazoles containing an ¹⁸ F atom and an antibody) for example), / ^{^'/')} functional polymers for the capture and detection of biomolecules of interest, / ^{^'/'} / ^') molecules for the detection and analysis of DNA, RNA and proteins following the incorporation of fluorescent groups via the process of the invention, (V) nano-objects with therapeutic activity (in the case of dimerimers or polyfunctionalized nanoparticles, via the process of the invention, by an active principle), v) radiopharmaceutical compounds (thazoles comprising an antibody specific for tumor cells and a radioelement for example)

The preferred conditions of implementation of the pyrazole compounds described above also apply to the other objects of the invention referred to above, in particular to their applications and to their manufacturing process.

The following examples illustrate the present application.

Examples 1 to 6 Pyrazole compounds of formula I wherein R is phenyl, R 'is hydrogen and R "is benzyl are prepared according to the reaction below,

according to the procedures of the invention set forth below:

Procedure A

200 μl of a freshly prepared aqueous solution of copper sulfate pentahydrate (5 mg, 0.02 mmol) (0.2 eq) in disodium bathophenanthrolinedisulfonic acid trihydrate (1.18 mg, 0.5%) were added. , 02 mmol) and triethanolamine (15 mg, 0.1 mmol), to a solution of sydnone (0.1 mmol), alkyne (0.1 mmol), sodium ascorbate (39.6 mg 0.2 mmol) and triethanolamine (15 mg, 0.1 mmol) in 1.8 ml of water. The resulting mixture was stirred at 60 ° C. for 16 h, then cooled with 2 ml of a 0.05 M solution of HEDTA and extracted with 3 × 5 ml of methylene chloride. The combined organic phases were dried over anhydrous sodium sulphate and concentrated to obtain the expected product, further purification by flash chromatography. Procedure B

A solution of sydnone (0.5 mmol), alkyne (0.5 mmol), triethylamine (0.5 mmol) and 1, 10-phenanthroline (0.1 mmol) was poured into 10 ml of dimethylformamide. anhydrous in a flask containing copper iodide (0.1 mmol) under Argon. The resulting mixture was stirred at 60 ° C under argon for 16h and then concentrated. The residue was dissolved in 10 mL of methylene chloride and washed with 10 mL of 0.05 M HEDTA solution. The organic phase was dried over anhydrous sodium sulphate and concentrated to give the expected product, further purification by flash chromatography.

Procedure C Copper sulphate pentahydrate (5 mg, 0.02 mmol) (0.2 eq) was added to a solution of sydnone (0.1 mmol), alkyne (0.1 mmol), ascorbate of sodium (39.6 mg, 0.2 mmol), triethylamine (0.1 mmol) and 1, 10-phenanthroline (0.02 mmol) in 2 ml of a tert-butanol / water mixture (1: 1). The resulting mixture was stirred for 16h at 60 ° C and then cooled with 2 ml of a 0.05 M solution of HEDTA and extracted with 3 x 5 ml of methylene chloride. The combined organic phases were dried over anhydrous sodium sulphate and concentrated to obtain the expected product, further purification by flash chromatography.

Procedure C

200 μl of a freshly prepared aqueous solution of copper sulfate pentahydrate (5 mg, 0.02 mmol) (0.2 eq.) In disodium bathophenanthroline disulfonic acid trihydrate (11.8 mg. , 02 mmol) and triethanolamine (15 mg, 0.1 mmol), to a solution of sydnone (0.1 mmol), of alkyne (0.1 mmol), of sodium ascorbate (39.6 mg 0.2 mmol) in 1.8 ml of tert-butanol / water (55:45). The resulting mixture was stirred for 16 h at 60 ° C and then cooled with 2 mL of a 0.05 M solution of HEDTA and extracted with 3 x 5 mL of methylene chloride. The combined organic phases were dried over anhydrous sodium sulfate and concentrated to give the expected product, further purified by flash chromatography.

Procedure D

The procedure was as in procedure C, but carrying out the reaction at room temperature.

Procedure E

The procedure was as in procedure B, but without triethylamine. Procedure F

The procedure was as in procedure C, but without copper ligand (without 1,10-phenanthroline) and at 100 ° C instead of 60 ° C. Procedure G

The procedure was as in procedure C, but using the 3,10-diphenyldiimidazo [1,2-a: 2 ', 1'-c] quinoxaline ligand (Villa) in place of 1,10-phenanthroline.

3,10-diphenyldiimidazo [1,2-a: 2 ', 1'-c] quinoxaline

The results obtained are reported in Table 1 below. Table 1

Thus, one has an excellent yield, in particular by using the protocols A, B, C and C ', and in addition the regioselectivity of the reaction is total in favor of the compound 3a, this in very varied mediums, aqueous, organic, or mixed.

Comparative Example 1

By way of comparison, the same reagents have also been used to carry out their reaction thermally according to the prior art. under the following conditions: Solvent = DMF - Reflux (153 ° C) - reaction time = 16h.

A yield of only 15% was obtained, and a mixture of isomers 3a and 3a 'in a ratio of 3a / 3a' = 25/75. 1-Phenethyl-1-phenyl-1H-pyrazole

The title product of Example 7 was prepared from 3-phenylsydnone and 4-phenyl-1-butyne using procedure C, with purification by flash chromatography (ethyl acetate (EtOAc) / cyclohexane =

20/80). The product was obtained in the form of a yellow oil with a yield of 96%.

By proceeding under the conditions of procedure B, the product was obtained with a yield of 90%.

1 H NMR (400 MHz, CDCl ₃ , δ ppm): 7.67 (s, 1H), 7.65 (s, 2H), 7.56 (s, 1H),

7.46 (t, J = 8.0 Hz, 2H), 7.20 - 7.37 (m, 6H), 2.93 - 3.00 (m, 2H), 2.86 - 2 93

(m, 2H).

¹³ C NMR (100 MHz, CDCl ₃ , δ ppm): 141, 6, 141, 0, 140.3, 129.5, 128.6, 128.5, 126.2, 125.0, 123.1, 18.9, 37.2, 26.3.

Example 9: (1-Phenyl-1H-pyrazol-4-yl) methyl benzoate

The title product of Example 9 was prepared from 3-phenylsydnone and prop-2-yn-1-yl benzoate using procedure C, with purification by flash chromatography (EtOAc / cyclohexane = 20/80). ). The product was obtained as a yellow oil with a yield of 93%. ¹ H NMR (400 MHz, CDCl ₃ , δ ppm): 8.05 (m, 3H), 7.82 (s, 1H), 7.68 (d, J = 7.7 Hz, 2H) , 7.56 (t, J = 7.5 Hz, 1H), 7.49-7.38 (m, 4H), 7.30 (t, J = 7.5 Hz, 1H), , 34 (s, 2H). ¹³ C NMR (100 MHz, CDCl ₃ , δ ppm): 166.7, 141, 7, 140.1, 133.2, 130.2, 129.8, 129.6, 128.5, 127.7, 126.9, 19.4, 18.6, 57.8.

Example 10 2- (-Phenyl-1H-pyrazol-4-yl) propan-2-ol

The title product of Example 10 was prepared from 3-phenylsydnone and 2-methyl-3-butyn-2-ol using procedure C, with purification by flash chromatography (EtOAc / cyclohexane = 10/90 ). The product was obtained as a white solid with a yield of 83%.

1H NMR (400 MHz, Methanol-d ₄ , δ ppm): 8.1 2 (s, 1H), 7.75-7.65 (m, 3H), 7.47 (t, J = 8, 1 Hz, 2H), 7.31 (t, J = 7.3Hz, 1H), 1.59 (s, 6H).

1 ³ C NMR (100 MHz, Methanol-d ₄ , δ ppm): 141, 6, 1 39.7, 1 35.0, 130.7, 1 27.8, 125.7, 120.4, 68, 7, 31, 9.

The title product of Example 11 was prepared from 3-phenylsydnone and phenylacetylene using procedure C, with purification by flash chromatography (EtOAc / cyclohexane = 20/80). The product was obtained as a white solid with a yield of 80%.

¹ H NMR (400 MHz, CDCl ₃ , δ ppm): 8.19 (s, 1H), 8.04 (s, 1H), 7.77 (d, J = 8.1 Hz, 2H) , 7.59 (d, J = 7.7 Hz, 2H), 7.51 (t, J = 7.8 Hz, 2H), 7.44 (t, J = 7.7 Hz, 2H). ), 7.37 - 7.27 (m, 2H).

¹³ C NMR (100 MHz, CDCl ₃ , δ ppm): 140.2, 138.9, 132.2, 129.6, 129.1, 127.0, 126.7, 125.8, 125.0, 123.5, 1 19.2.

Example 12 4-heptyl-1-phenyl-1H-pyrazole The title product of Example 12 was prepared from 3-phenylsydnone and 1-nonyne using procedure C, with purification by flash chromatography (EtOAc / cyclohexane = 20/80). The product was obtained as a yellow oil with a yield of 61%.

1H NMR (400 MHz, CDCl ₃ , δ ppm): 7.70 (s, 1H), 7.66 (d, J = 7.7 Hz, 2H), 7.55 (s, 1H), 7.43 (t, J = 7.9 Hz, 2H), 7.25 (t, J = 7.3 Hz, 1H), 2.52 (t, J = 7.7 Hz, 2H) , 1.61 (quin, J = 7.4 Hz, 2H), 1.38-1.8 (m, 8H), 0.89 (t, J = 6.8 Hz, 3H). ¹³ C NMR (100 MHz, CDCI ₃ , δ ppm): 141, 1, 140.4, 129.5, 126.1, 124.8, 124.2, 1 18.9, 32.0, 30.9 , 29.4, 29.3, 24.3, 22.8, 14.2.

Example 1 - (4-Phenethyl-1H-pyrazol-1-yl) benzoic acid

The title product of Example 13 was prepared from 3- (4-carboxyphenyl) sydnone and 4-phenyl-1-butyne using procedure C. The product was obtained as a white solid with a yield of 99%. ¹ H NMR (400 MHz, DMSO-d _6, δ ppm): 12.98 (br s, 1H..), 8.40 (s, 1H), 8.04 (d, J = 8.6 Hz, 2H), 7.90 (d, J = 8.6Hz, 2H), 7.63 (s, 1H), 7.32-7.21 (m, 4H), 7.17; (t, J = 6.8Hz, 1H), 2.90 (t, J = 7.7Hz, 2H), 2.79 (t, J = 7.7Hz, 2H).

1 ³ C NMR (100 MHz, DMSO-d ₆ , δ ppm): 166.7, 42.8, 141, 8, 141, 4, 130.9, 128.3, 128.3, 127.7, 126.0, 125.9, 123.6, 17.4, 36.0, 25.6.

Examples 14, 15, 16: 4- (4- (2-hydroxypropan-2-yl) -1H-pyrazol-1-yl) benzoic acid

The title product of Example 14 was prepared from 3- (4-carboxyphenyl) sydnone and 2-methyl-3-butyn-2-ol using procedure A, operating at 37 ° C. product was obtained as a white solid with a yield of 93%. By proceeding under the conditions of procedure A but using as a solvent human blood plasma, the product was obtained in 88% yield.

By proceeding under the conditions of procedure C, the product was obtained with a yield of 91%.

¹ H NMR (400 MHz, Methanol-d ₄ , δ ppm): 8.27 (s, 1H), 8.14 (d, J = 8.8 Hz, 2H), 7.87 (d, J) = 8.8 Hz, 2H), 7.77 (s, 1H), 1.61 (s, 6H)

¹³ C NMR (100 MHz, Methanol-d ₄ , δ ppm): 169.2, 144.8, 140.7, 135.7, 1 32.5, 125.6, 1 19.4, 68.7, 31, 9

Example 17: (2R, 3R, 4S, 5R, 6R) -2- (acetoxymethyl) triacetate

The title product of Example 17 was prepared from 3-

(phenyl) sydnone and 2-propynyl-tetra-O-acetyl-D-glucopyranoside using procedure C. The product was obtained as a white solid in 96% yield.

¹ H NMR (400 MHz, CDCl ₃ , δ ppm): 7.89 (s, 1H), 7.68-7.62 (m, 3H), 7.44 (t, J = 7.9 Hz) , 2H), 7.28 (t, J = 7.2 Hz, 1H), 5.19 (t, J = 9.5 Hz, 1H), 5.10 (t, J = 9.5 Hz, 1H), 5.03 (t, J = 9.1Hz, 1H), 4.83 (d, J = 12.4Hz, 1H), 4.66-4.59 (m, 2H), 4.27 (dd, J = 4.8, 12.2 Hz, 1H), 4.18 (dd, J = 2.3, 12.2 Hz, 1H), 3.76 - 3.66 (m, 1H), 2.09 (s, 3H), 2.01 (s, 3H), 1.99 (s, 3H), 1.98 (s, 3H).

1 ³ C NMR (100 MHz, CDCI ₃ , δ ppm): 170.8, 170.3, 169.5, 169.4, 140.9, 140.0, 129.5, 126.8, 126.6 , 1 19.2, 1 19.0, 99.2, 72.9, 71, 9, 71, 3, 69.2, 68.5, 62.1, 62.0, 20.8, 20.7 , 20.6. Example 18 4- (4-methoxyphenyl) -1-phenyl-1H-pyrazole

The title product of Example 18 was prepared from 3- (phenyl) sydnone and 1-ethynyl-4-methoxybenzene using procedure C, with purification by flash chromatography (EtOAc / cyclohexane = 20/80). . The product was obtained as a white solid with a yield of 64%.

¹ H NMR (400 MHz, CDCl ₃ , δ ppm): 8.09 (s, 1H), 7.94 (s, 1H), 7.73 (d, J = 8.2 Hz, 2H) , 7.48 (t, J = 8.2 Hz, 4H), 7.30 (t, J = 7.3 Hz, 1H), 6.95 (d, J = 8.6 Hz, 2H). ), 3.85 (s, 3H).

¹³ C NMR (100 MHz, CDCl ₃ , δ ppm): 158.8, 140.2, 138.7, 129.6, 127.1, 126.7, 124.8, 124.8, 123.0, 19.2, 14.6, 55.5.

The title product of Example 19 was prepared from 3- (phenyl) sydnone and 2-ethynylpyridine using procedure C. The product was obtained as a white solid in 95% yield .

1 H NMR (400 MHz, CDCl ₃ , δ ppm): 8.62 (d, J = 4.8 Hz, 1H), 8.55 (s, 1H), 8.20 (s, 1H), 7.78 (d, J = 7.9 Hz, 2H), 7.72 (dt, J = 1, 6, 7.7 Hz, 1H), 7.57 (d, J = 8.1 Hz). , 1H), 7.50 (t, J = 8.0 Hz, 2H), 7.33 (t, J = 7.3 Hz, 1H), 7.20 - 7.12 (m, 1 H). ¹³ C NMR (100 MHz, CDCl ₃ , δ ppm): 151, 5, 149.7, 140.0, 139.3, 136.9, 129.6, 126.9, 125.4, 125.2, 121, 6, 19.9, 19.3. Example 20: 1-Phenyl-4- (thiophen-3-yl) -1H-pyrazole

The title product of Example 20 was prepared from 3- (phenyl) sydnone and 2-ethynylthiophene using procedure C. The product was obtained as a white solid in 99% yield. %. ¹ H NMR (400 MHz, CDCl ₃ , δ ppm): 8.1 (s, 1H), 7.95 (s, 1H), 7.75 (d, J = 8.0 Hz, 2H) ), 7.49 (t, J = 8.0 Hz, 2H), 7.42 to 7.38 (m, 1H), 7.38 to 7.27 (m, 3H) ¹³ C NMR ( 100 MHz, CDCI ₃ , δ ppm): 140.1, 139.2, 132.9, 129.6, 126.6, 126.4, 126.2, 123.4, 120.4, 1 19.1 , 15.1.

Example 21 4- (4 - ((5- (dimethylamino) naphthalene-1 -

The title product of Example 21 was prepared from 3- (4-carboxyphenyl) sydone and 5- (dimethylamino) -N- (prop-2-yn-1-yl) naphthalene-1-sulfonamide. using procedure C, operating at 37 ° C. and with purification by flash chromatography (EtOAc / cyclohexane / AcOH = 49/49/2). The product was obtained in the form of a yellow solid with a yield of 59%.

¹ H NMR (400 MHz, Methanol-d ₄ , δ ppm): 8.40 (d, J = 8.4 Hz, 1H), 8.28 (d, J = 8.6 Hz, 1H), 8.20 (d, J = 7.4Hz, 1H), 8.03 (d, J = 8.8Hz, 2H), 7.59-7.42 (m, 5H), 7, (S, 1H), 7.17 (d, J = 7.5 Hz, 1H), 4.10 (s, 2H), 2.73 (s, 6H). ¹³ C NMR (100 MHz, Methanol-d ₄ , δ ppm): 169.2, 153.3, 144.1, 142.3, 137.5, 132.3, 1 31, 4, 131, 1, 131 , 1, 130.7, 129.9, 129.2, 127.7, 1 24.3, 121, 9, 120.5, 1 19.2, 1 16.4, 45.8, 38.0. Example 22: 2- (6- (Diethylamino) -3- (diethylimino) -3H-xanthen-9-yl) -5- (N - ((1-phenyl-1-pyrazol-4-yl) methyl) sulfamoyl) benzenesulfonate

The title product of Example 22 was prepared from 3- (phenyl) sydnone and 2- (6- (diethylamino) -3- (diethyliminio) -3H-xanthen-9-yl) -5- ( N- (prop-2-yn-1-yl) sulfamoyl) benzenesulfonate using procedure C and isolation by filtration from the reaction medium. The product was obtained as a violet solid with a yield of 92%.

¹ H NMR (400 MHz, DMSO-d _6, δ ppm): 8.47 (d, J = 1, 6 Hz, 1H), 8.38 (br s, 1H..), 8.32 ( s, 1H), 7.91 (dd, J = 1.6, 8.0 Hz, 1H), 7.76 (d, J = 7.9 Hz, 2H), 7.51 (s, 1H), 7.45 (t, J = 7.9 Hz, 2H), 7.39 (d, J = 7.9 Hz, 1H), 7.28 (t, J = 7.3 Hz). , 1H), 6.98 - 6.86 (m, 6H), 4.12 (d, J = 2.8 Hz, 2H), 3.69 - 3.56 (m, 8H), 1, 20 (t, J = 7.0 Hz, 12H).

MS (ESI) m / z: 714.3 [M + H] ⁺ .

Example 23: -phenyl-1H-pyrazol-4-yl) methyl) amine

Copper sulfate pentahydrate (0.15 mmol) was added to a solution of 3-phenylsydnone (0.9 mmol), tripropargylamine (0.1 mmol), sodium ascorbate (1.5 mmol), triethylamine (0.9 mmol) and 1.10 g. phenanthroline (0.15 mmol) in 18 ml of a tert-butanol / water mixture (1: 1).

The resultant mixture was stirred in a closed flask at 60 ° C. for 16 h. The second portion of sodium ascorbate (1.5 mmol), 1.10-phenanthroline (0.15 mmol) was then added to the mixture. and copper sulfate pentahydrate (0.15 mmol) and the new mixture was stirred at 60 ° C. for a further 24 hours.

After cooling to room temperature, the mixture was diluted with 20 ml of 0.05 M HEDTA solution and extracted with 3x30 ml of methylene chloride. The combined organic phases were dried over anhydrous sodium sulphate and concentrated to obtain 127.3 mg of the expected product in the form of a white solid, 0.26 mmol, 85% yield, further purified by flash chromatography (CH ₂ Cl ₂ / EtOAc = 80/20 to 0/100).

1 H NMR (400 MHz, CDCl ₃ , δ ppm): 7.91 (s, 3H), 7.73 (s, 3H), 7.71 (d, J = 8.0 Hz, 6H), 7.47 (t, J = 7.9 Hz, 6H), 7.30 (t, J = 8.2 Hz, 3H), 3.67 (s, 6H).

1 ³ C NMR (100 MHz, CDCl ₃ , δ ppm): 141, 7, 140.2, 129.6, 126.5, 126.4, 120.3, 1 19.0, 47.1. Example 24 Labeling of a Peptide with a Fluorophore: Acid

2- (2- (2- (4- (4 - ((5- (dimethylamino) naphthalene-1-sulfonamido) methyl) -1H-

To a solution of 3- (4 - ((2 - ((1 - ((carboxymethyl) amino) -3- (1H-indol-3-yl) -1-oxopropan-2-yl) amino) -2- oxoethyl) carbamoyl) phenyl) -1,2,3-oxadiazol-3-ium-5-olate (6 mg, 12 μιτιοΙ), N-dansyl-N-propargylamine (4.5 mg, 16 μιτιοΙ), sodium ascorbate ( 7 mg, 36 μιτιοΙ) and triethanolamine (1.8 mg, 12 μιτιοΙ) in 1 ml of water was added 200 μΙ of a freshly prepared aqueous solution of copper sulfate pentahydrate (0.9 mg, 3.6 μιτιοΙ), disodium salt trihydrate bathophenanthrolinedisulfonic acid (2.1 mg, 3. 6 μ ιτι ο Ι) and triethanolamine (1 .8 mg, 12 μιτιοΙ). The resulting mixture was stirred at 37 ° C for 16h. Liquid chromatography / mass spectrometry analysis of the resulting mixture showed complete conversion of the starting material. The mixture was concentrated and purified by high performance liquid chromatography on a C18 XBridge 19 x 150 mm column, 5 μιτι to give the expected product labeled with dansyl.

1H NMR (400 MHz, Methanol-d ₄ , δ ppm): 8.43 (d, J = 8.4 Hz, 1H), 8.30 (d, J = 8.6 Hz, 1H), 8 , 22 (d, J = 6.6 Hz, 1H), 7.86 (d, J = 8.8 Hz, 2H), 7.63-7.55 (m, 3H), 7.54 7.46 (m, 3H), 7.31 (d, J = 8.1 Hz, 1H), 7.22 (s, 1H), 7.20 (d, J = 7.5 Hz). , 1H), 7.16 (s, 1H), 7.07 (t, J = 7.1 Hz, 1H), 7.00 (t, J = 7.5 Hz, 1H), 4 , 80 (dd, J = 5.4, 7.8 Hz, 1H), 4.12 (s, 2H), 4.06 (d, J = 16.5 Hz, 1H), 4.01 - 3.83 (m, 3H), 3.44 - 3.38 (m, 1H), 3.25 - 3.12 (m, 1H), 2.76 (s, 6H)

MS (ESI) m / z: 751.4 [M + H] ⁺ . Example 25: Marking a protein with a fluoro hore

The procedure is as above but using as starting sydnone the sydnone coupled to the bovine serum albumin prepared hereinafter. It was operated at room temperature for 16 hours under the following conditions:

Sydnone coupled with bovine serum albumin: 1 mg / mL - 0.18 μιτιοΙ, 1 eq.

N-dansyl-N-propargylamine: 1, 8 μιτιοΙ, 10 eq.

CuSO4: 1, 8 μηηοΙ, 10 eq.

Bathophenanthroline: 1, 8 μιτιοΙ, 10 eq.

Sodium ascorbate: 18 μιτιοΙ, 100 eq.

After revelation of the proteins with Coomassie blue and measurement of the fluorescence of dansyl A _ex = 320 nm, A _em = 320 nm, it was found that an average number of dansyl residues of 1.6 was recorded on serum albumin. bovine.

Thus, the method of the invention can be implemented on starting sydnones carrying biological molecules.

Example 26: (1H-phenyl-1H-pyrazol-4-yl) methyl 2- (4 - ((5-chloroethyl)

3-fluoropyridin-2-yl) oxy) phenoxy) propanoate

The title product of Example 26 was prepared from 3- (phenyl) sydnone and Clodinafop-propargyl using procedure C and purification by flash chromatography (EtOAc / cyclohexane = 30/70). The product was obtained as a white solid with a yield of 62%. ¹ H NMR (400 MHz, Methanol-d ₄ , δ ppm): 8.15 (s, 1H), 7.79 (d, J = 2.2 Hz, 1H), 7.75 (dd, J) = 2.2, 9.5 Hz, 1H), 7.70 (s, 1H), 7.66 (d, J = 7.9 Hz, 2H), 7.44 (t, J = 7); , 9 Hz, 2H), 7.30 (t, J = 7.3 Hz, 1H), 6.99 (d, J = 9.0 Hz, 2H), 6.86 (d, J = 9.0 Hz, 2H), 5.19 (s, 2H), 4.85 (q, J = 6.8Hz, 1H), 1.58 (d, J = 6.8Hz, 3H). H). ¹³ C NMR (100 MHz, CDCI ₃ , δ ppm): 173.8, 156.4, 152.7, 148.6, 148.5, 142.7, 141, 3, 141, 2, 1, 30.7 , 129.6, 1 28.1, 1 26.5, 1 26.3, 1 23.4, 120.5, 1 1 9.8, 1 1 7.2, 74.3, 59.0, 18 9.

MS (ESI) m / z: 468.0 [M ⁽³⁵ Cl) + H] ^+, 469.8 [M ⁽³⁷ Cl) + H] ⁺

Example 27: 1 - ((2R, 4S, 5R) -4-hydroxy-5- (hydroxymethyl)

The title product of Example 27 was prepared from 3- (phenyl) sydnone and 5-ethynyl-2'-deoxyuridine using procedure C and isolation by filtration from the reaction medium. The product was obtained as a white solid with a yield of 84%.

¹ H NMR (400 MHz, DMSO-d6, δ ppm): 8.68 (s, 1H), 8.48 (s, 1H), 8.05 (s, 1H),

7.81 (d, J = 7.9 Hz, 2H), 7.50 (t, J = 7.9 Hz, 2H), 7.32 (t, J = 7.9 Hz, 1H) , 6.23

(t, J = 6.4Hz, 1H), 4.33 (brs, 1H), 3.84 (d, J = 3.0Hz, 1H), 3.73 (dd, J) = 3.0,

11.9 Hz, 1H), 3.66 (dd, J = 3.0, 11.9 Hz, 1H), 2.32-2.13 (m, 2H).

¹³ C NM R (100 MHz, DMSO-d ₆ , δ ppm): 162.1, 150.0, 139.9, 138.9, 135.8,

130.0, 126.7, 124.9, 18.7, 16.6, 106.2, 87.8, 85.0, 70.2, 61, 1, 31.0.

MS (ESI) m / z: 371.0 [M + H] ⁺ , 741.3 [2M + H] ⁺ .

Example 28 Ethyl 1-phenyl-1H-pyrazole-4-carboxylate The title product of Example 28 was prepared from 3-phenylsydnone and ethyl propionate using protocol F. The product was obtained in 84% yield after purification by chromatography (EtOAc / cyclohexane = 10/90).

¹ H NMR (400 MHz, CDCl ₃ , δ ppm): 8.39 (s, 1H), 8.08 (s, 1H), 7.68 (d, J = 8.0 Hz, 2H), 7.45 (t, J = 8.0 Hz, 2H), 7.32 (t, J = 7.3 Hz, 1H), 4.32 (q, J = 7.1 Hz, 2H), 1 , 36 (t, J = 7.1 Hz, 3H).

¹³ C NMR (100 MHz, CDCl ₃ , δ ppm): 162.9, 142.2, 139.4, 130.0, 129.6, 127.6, 19.6, 17.0, 60, 5, 14.5.

IR (NaCl, cm- ¹ ): 3124, 3063, 2984, 2901, 1709, 1597, 1560, 1505, 1463, 1413, 1390, 1258, 1150, 1027, 996, 951, 887, 825, 770, 757, 688, 654.

MS (ESI) m / z: 217.1 [M + H] ⁺ .

HRMS (ESI) m / z: Calc, for C12H13N2O2: 217.0977; found: 217.0971.

Example 29: 5-Bromo-4-phenethyl-1-phenyl-1H-pyrazole.

The title product of Example 29 was prepared from 3-phenyl-4-bromosydnone and 4-phenyl-1-butyne using Protocol G. The product was obtained as a yellow solid with a yield of 74% after purification by chromatography (Heptane / Ethyl acetate: 95/5).

1H NMR (Chloroform -d, 400MHz): d = 7.38-7.55 (m, 6H), 7.28-7.34 (m, 2H), 7.19-7.25 (m, 3H), 2.89 - 2.95 (m, 2H), 2.77 - 2.83 (m, 2H) ppm;

¹³ C NMR (Chloroform -d, 101 MHz): d = 141, 4, 140.9, 139.4, 129.0, 1 28.6, 128.6, 128.3, 126.3, 125.6 , 122.3, 12.6, 36.3, 26.7 ppm;

IR (NaCl, cm- ¹ ): 3062, 3027, 2923, 2856, 1598, 1556, 1499, 1455, 1407, 1391, 1241, 1087, 1068, 961, 91, 851, 836, 760, 694;

LCMS (ESI) Ci ₇ Hi ₅ BrN ₂ (M ^[79 Br] H +) 326.9, (M ^[81 Br] + H) 328.8. Example 30: 5-Bromo-1- (4-methoxyphenyl) -4-phenethyl-1H-pyrazole

The title product of Example 30 was prepared from 3- (p-methoxyphenyl) -4-bromosydnone and 4-phenyl-1-butyne using Protocol G. The product was obtained under form of a yellow solid with a yield of 70% after purification by chromatography (Heptane / Ethyl acetate: 95/5).

¹ H NMR (Chloroform -d, 400MHz): d = 7.49 (s, 1H), 7.39 to 7.44 (m, 2H), 7.28 to 7.34 (m, 2H) , 7.19 - 7.25 (m, 3H), 6.95 - 7.01 (m, 2H), 3.86 (s, 3H), 2.88 - 2.95 (m, 2 H), 2.75 - 2.82 (m, 2H) ppm;

¹³ C NMR (Chloroform -d, 101 MHz): d = 159.4, 141, 3, 140.3, 132.4, 128.6 (2C), 128.4 (2C), 127.0 (2C) 126.1, 121, 7, 14.0 (2C), 12.9, 55.6, 36.1, 26.7 ppm;

LCMS (ESI) Ci ₈ Hi ₇ BrN ₂ O (M [ ^'y Br] + H) 357.3, (Mf Br] + H) 359.4.

The starting sydnones of formula II can be prepared as follows: Step A: preparation of N-nitrosyl amino acids

A solution of 6.90 g of NaNO ₂ in 100 ml of water is added dropwise over 40 minutes to a vigorous suspension of N-aryl amino acid (0.1 mol) in 10% strength. aqueous hydrochloric acid (100 mL). The resulting mixture was stirred at room temperature under nitrogen for 14H. The expected product was collected by filtration, washed with a little methanol and dried to produce the expected N-nitrosylated compound.

Thus, 4 - ((carboxymethyl) (nitroso) amino) benzoic acid was prepared as a white solid with a yield of 72%.

1 H NMR (400 MHz, DMSO-d ₆ , δ ppm): 8.08 (d, J = 8.6 Hz, 2H), 7.76 (d, J = 8.6 Hz, 2H), 4 , 81 (s, 2H). ¹³ C NMR (100 MHz, DMSO-d ₆ , δ ppm): 167.0, 16.6, 144.5, 130.7, 129.3, 19.0, 46.2.

2- (Nitroso (phenyl) amino) acetic acid was also prepared as a white solid in 89% yield.

1H NMR (400 MHz, DMSO-d ₆ , δ ppm): 7.62 (d, J = 7.9 Hz, 2H), 7.54 (t, J = 7.9 Hz, 2H), 7 , 43 (t, J = 7.3 Hz, 1H), 4.76 (s, 2H).

¹³ C NMR (100 MHz, DMSO-d ₆ , δ ppm): 167.2, 141, 3, 129.5, 127.6, 19.9, 46.9.

Stage B: Preparation of svdnones of Formula II

A mixture of 0.1 mol of N-nitrosylated amino acid and 100 ml of acetic anhydride was mixed for 3 hours at 100 °. The solution obtained was concentrated on a rotary evaporator. The residue was triturated with 100 ml of water.

The precipitate was then recovered by filtration and recrystallized from methanol to obtain the expected pure product.

Thus, 3- (4-carboxyphenyl) -1,2,3-oxadiazol-3-ium-

5-olate as a white solid with a yield of 52%.

¹ H NMR (400 MHz, DMF-d _7, δ ppm): 8.32 (d, J = 8.5 Hz, 2H), 8.19 (d, J = 8.5

Hz, 2H), 7.88 (s, 1H)

¹³ C NMR (100 MHz, DMF-d ₇ , δ ppm): 169.7, 167.1, 138.9, 135.7, 132.3, 123.1, 96.1.

3-Phenyl-1,2,3-oxadiazol-3-ium-5-olate was also prepared as a white solid in 55% yield.

1H NMR (400 MHz, DMSO-d ₆ , δ ppm): 7.94 (dd, J = 1.5, 8.1 Hz, 2H), 7.78 (s, 1H), 7.77- 7.67 (m, 3H)

1 ³ C NMR (100 MHz, DMSO-d _6, δ ppm): 168.5, 132.4, 130.2, 126.6, 121, 6, 94.9.

Preparation of a tripeptide-bearing svdnone of formula II: 3- (4 - ((2 - ((1 - ((carboxymethyl) amino) -3- (1H-indol-3-yl) -1-oxopropanol 2-yl) amino) -2-oxoethyl) carbamoyl) phenyl) -1,2,3-oxadiazol-3-ium-5-olate.

N, N'-Dicyclohexylcarbodiinnide (13 mg, 0.063 mmol) was added to a solution of 3- (4-carboxyphenyl) sydnone (13 mg, 0.063 mmol) and N-Hydroxysuccininnide (8.7 mg, 0.076 mmol). in anhydrous dimethylformamide (1 ml) at 0 ° C. The resulting solution was stirred at room temperature for 24 h and then filtered. Then, the tripeptide H-Gly-Trp-Gly-OH (20 mg, 0.063 mmol) was added to the filtrate and the solution was stirred at room temperature for a further 16 hours. The mixture was then concentrated and the residue was concentrated. purified by high performance semi-preparative liquid chromatography on Phenomenex Synergi Fusion-RP column 21.2 x 150 mm, 5 μιτι, to produce 13 mg of the expected product in the form of a white solid with a yield of 41%.

¹ H NMR (400 MHz, Methanol-d ₄ , δ ppm): 8.05 (d, J = 8.8 Hz, 2H), 7.98 (d, J = 8.8 Hz, 2H), 7.58 (d, J = 8.8 Hz, 1H), 7.48 (s, 1H), 7.28 (d, J = 8.1 Hz, 1H), 7.14 (s, 1H), 7.04 (t, J = 7.3 Hz, 1H), 6.97 (t, J = 7.2 Hz, 1H), 4.79 (dd, J = 5.5, 7.7 Hz, 1H), 4.07 (d, J = 16.5 Hz, 1H), 3.97 (d, J = 16.5 Hz, 1H), 3.94 - 3.82 (m, 2H), 3.36 (dd, J = 5.2, 14.7 Hz, 1H), 3.17 (dd, J = 8.1, 14.8 Hz, 1H).

MS (ESI) m / z: 507.1 [M + H] ⁺ . Preparation of a svdnone of formula II carrying a protein: bovine serum albumin (BSA)

The procedure was as above, but using the bovine serum albumin instead of the tripeptide to obtain the expected product, in which the bovine serum albumin carries 12 sydnones.

Claims

1. A process for preparing a pyrazole of formula I

in which

R represents a hydrogen atom, a halogen atom, a thio or -thioalkyl group, a hydroxy or aikoxy group, an amino or aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is unsubstituted; or substituted one or more times with F, Cl, Br, I, CN, OH, NH ₂ , NH-C 1 -C 8 alkyl, N- (C 1 -C 8 alkyl) ₂ , SH, an aryl radical containing 6 to 10 carbon atoms. carbon which is unsubstituted or substituted one or more times with an alkyl or alkoxy radical containing 1 to 8 carbon atoms, a heterocycle, an amino acid, a polypeptide, a protein, a nucleic acid, a DNA molecule, a polysaccharide , a polymer, a nanoparticle, a dendrimer.

R 'represents a hydrogen atom, a halogen atom, a thio or -thioalkyl group, a hydroxy or aikoxy group, an amino or -aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is not substituted or substituted one or more times with F, Cl, Br, I, CN, OH, NH ₂ , NH-C 1 -C 8 alkyl, N- (C 1 -C 8 alkyl) ₂ , SH, an aryl radical containing 6 to 10 atoms of carbon which is unsubstituted or substituted one or more times with an alkyl or alkoxy radical containing 1 to 8 carbon atoms, a heterocycle,

R "represents a hydrogen atom, a halogen atom, a thio group, a hydroxyl group, an amino group, a hydrocarbon group or a hydrocarbon group containing from 1 to 100 heteroatoms, R" particularly representing an atom of hydrogen, a halogen atom, a thio or thioalkyl group, a hydroxy or alkoxy group, an amino or -aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is unsubstituted or substituted one or more times with Substituted or unsubstituted F, Cl, Br, I, CN, OH, O-heterocycle, NH ₂ , -NH-C 1 -C 8 alkyl, -N- (C 1 -C 8 alkyl) ₂ , -NH-R '", or - Alkyl C1 -C3-NH-R '"where R'" represents a sulfonated group, SH, an aryl radical containing 6 to 10 carbon atoms which is unsubstituted or substituted one or more times with an alkyl or alkoxy radical containing 1 to 8 carbon atoms, a heterocycle, an amino acid, a polypeptide, a protein, a nucleic acid, a DNA molecule, a polysaccharide, a polymer, a nanoparticle, a dendrimer,

characterized in that a sydnone of formula II is made

In which R "has the meaning already indicated, in the presence of copper used as a catalyst metal, to obtain a pyrazole compound of formula I which is isolated and salified if desired.

2. A process according to claim 1, characterized in that R represents an unsubstituted or substituted aryl or heteroaryl radical.

3. A process according to claim 1 or 2, characterized in that R 'represents a hydrogen atom.

4. A process according to one of claims 1 to 3, characterized in that R "represents a heteroaryl radical or an aryl radical or an alkyl radical containing from 1 to 8 carbon atoms.

5. A method according to one of claims 1 to 4, characterized in that the catalyst metal is used in the form of a copper salt complexed with a nitrogen ligand.

6. A method according to one of claims 1 to 4, characterized in that the catalyst metal is used in the form of a copper salt complexed with a nitrogen ligand selected from phenanthroline compounds, triazoles, pyridines, imidazoles, and benzimidazoles.

7. A pyrazole compound of formula I in which

R "represents a hydrogen atom, a halogen atom, a thio group, a hydroxyl group, an amino group, a hydrocarbon group or a hydrocarbon group containing from 1 to 100 heteroatoms, R" particularly representing an atom of hydrogen, a halogen atom, a thio or thioalkyl group, a hydroxy or alkoxy group, an amino or -aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is unsubstituted or substituted one or more times with Substituted or unsubstituted F, Cl, Br, I, CN, OH, O-heterocycle, NH ₂ , -NH-C 1 -C 8 alkyl, -N- (C 1 -C 8 alkyl) ₂ , -NH-R '", or - Alkyl C1 -C3-NH-R '"where R'" represents a sulfonated group, SH, an aryl radical containing 6 to 10 carbon atoms which is unsubstituted or substituted one or more times with an alkyl or aikoxy radical containing 1 to 8 carbon atoms, a heterocycle, an amino acid, a polypeptide, a protein, a nucleic acid, a e molecule of DNA, a polysaccharide, a polymer, a nanoparticle, a dendrimer, and

R represents a polypeptide, a protein, a nucleic acid, DNA, a polysaccharide, a polymer, a nanoparticle or a dendrimer.

8. A pyrazole compound according to claim 7 characterized in that R "has the function of a fluorophore, a radiotracer, an active principle of human therapy (drug), a molecule capable of interacting with a protein or DNA.

9. A prazole compound of formula I

in which R represents a hydrogen atom, a halogen atom, a thio or -thioalkyl group, a hydroxy or aikoxy group, an amino or aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is unsubstituted; or substituted one or more times with F, Cl, Br, I, CN, OH, NH ₂ , NH-C 1 -C 8 alkyl, N- (C 1 -C 8 alkyl) ₂ , SH, an aryl radical containing 6 to 10 carbon atoms. carbon which is unsubstituted or substituted one or more times with an alkyl or alkoxy radical containing 1 to 8 carbon atoms, a heterocycle, an amino acid, a polypeptide, a protein, a nucleic acid, a DNA molecule, a polysaccharide , a polymer, a nanoparticle, a dendrimer.

R 'represents a hydrogen atom, a halogen atom, a thio or -thioalkyl group, a hydroxy or aikoxy group, an amino or -aminoalkyl group, an alkyl radical containing from 1 to 8 carbon atoms which is not substituted or substituted one or more times with F, Cl, Br, I, CN, OH, NH ₂ , NH-C 1 -C 8 alkyl, N- (C 1 -C 8 alkyl) ₂ , SH, an aryl radical containing 6 to 10 atoms carbon which is unsubstituted or substituted one or more times with an alkyl or alkoxy radical containing 1 to 8 carbon atoms, a heterocycle, and

- R "represents a polypeptide, a protein, a nucleic acid, a DNA molecule, a polysaccharide, a polymer, a nanoparticle, a dendrimer.

10. A pyrazole compound according to claim 9 characterized in that R has the function of a fluorophore, a radiotracer, an active principle of human therapy (drug), a molecule capable of interacting with a protein or DNA.

1 1. Use of a novel pyrazole compound as defined in any one of claims 7 to 10 or prepared by the method as defined in one of claims 1 to 6, in the detection of proteins.