TECHNICAL FIELD

The present disclosure relates to, process of preparing diyne compounds, and particularly, to the preparation of 14-(furan-2-yl)tetradeca-11,13-diynoic acid (14 FTDA), and salts thereof.

BACKGROUND

Diyne compounds have been found to inhibit the function of the Olel protein in a wide variety of fungal pathogens and are thus capable of inhibiting fungal growth in humans and animals. This potent antifungal activity due to the inhibition of Olel protein has been found to be a superior alternative to the generally used fungicides. There are four broad classes of fungicidal drug generally used. These are the polyenes that binds with sterols in fungal cell membrane, principally ergosterol; imidazoles and triazoles which inhibit cytochrome P450 14a-demethylase; allylamines that inhibit the enzyme squaleneepoxidase; and echnocandins that inhibit the synthesis of glucan in the cell wall. These drugs however, have serious side effects.

WO 2001/025197 discloses enediyne as antifungal compounds. Another disclosure, WO 2011/006061 discloses antifungal Diyne compounds as inhibitor of the function of the Olel protein.

However, there is a need to explore alternatives for the preparation of the diyne compounds to facilitate higher yield or cost effectiveness by for example, minimum usage of reagents to make it feasible to bring it into higher scale synthesis.

SUMMARY

The present disclosure relates to a process for the preparation of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA), and salts thereof comprising: coupling of a halo-ethynyl furan with an acetylene acid having terminal alkyne, in the presence of a metal catalyst and a base.

In an aspect of the present subject matter, the process comprises of a coupling reaction of a halo-ethynyl compound with an acetylene acid having terminal alkyne, to obtain 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA). In one aspect the halo ethynyl compound is bromo-ethynyl furan and the acetylene acid is do-dec-11-ynoic acid. In another aspect, the present disclosure relates to process of preparation of salts of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA).

These and other features, aspects, and advantages of the present subject matter will become better understood with reference to the following description. This summary is provided to introduce a selection of concepts in a simplified form. This summary is not intended to identify key features or essential features of the disclosure, nor is it intended to be used to limit the scope of the subject matter.

BRIEF DESCRIPTION OF THE DRAWINGS

The above and other features, aspects, and advantages of the subject matter will be better understood with regard to the following description, and accompanying drawings where:

Figure 1 shows normal hyphal growth as compared with changes in the plane of hyphal growth and abnormal thickening of the hyphae due to the antifungal effect of the potassium salt of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA).

DETAIL DESCIPTION:

The present disclosure provides a process for the preparation of the antifungal compound, 14-(furan-2-yl)tetradeca-l 1,13-diynoic acid (or 14-FTDA).

In accordance with the present disclosure, it provides a process for the preparation of 14-(furan-2-yl)tetradeca-l 1,13-diynoic acid (14-FTDA), and salts thereof comprising: coupling of a halo-ethynyl furan with an acetylene acid having terminal alkyne, in the presence of a metal catalyst and a base, to obtain 14-(ftiran-2-yl) tetradeca-11,13-diynoic acid (14-FTDA).

The halo-ethynyl furan, in accordance with the present disclosure, is selected from fluro- ethynyl furan, chloro-ethynyl furan, bromo-ethynyl furan, or iodo-ethynyl furan. In one preferred embodiment, the halo-ethynyl furan is bromo-ethynly furan.

In another embodiment, acetylene acid is do-dec-11-ynoic acid. The coupling of the halo ethynyl compound directly with an acetylenic acid compound renders the process economically more viable, which facilitates in bringing the process to higher scale synthesis.

The metal catalyst used in the present disclosure is a copper catalyst; and the base used in the present disclosure is hydroxylamine hydrochloride.

The halo-ethynyl furan described in the present disclosure is prepared by a process comprising: reacting furfural with an triphenylphosphine (TPP), and carbon tetra halide, in the presence of chlorinated solvent to obtain 2-(2,2-dihalovinyl)furan; and reacting 2-(2,2-dihalovinyl)furan with an alkali hydroxide, and benzyltriethyl ammonium chloride, in the presence of chlorinated solvent to obtain a halo-ethynyl furan.

In yet another embodiment, the chlorinated solvent is selected from chloroform, methylene dichloride, ethylene dichloride, or mixture thereof and the carbon tetra halide used in the present disclosure is carbon tetra bromide, carbon tetra chloride, carbon tetra iodide or carbon tetra fluoride.

The halo-ethynyl furan described in the present disclosure can also prepared by reacting 2-ethynylfuran with an alkali hydroxide and halogen. The alkali hydroxide used in the present disclosure is selected from sodium hydroxide (NaOH) or potassium hydroxide (KOH) and the halogen used in the present disclosure is selected from bromine (Br2), chlorine (Cl2), iodine (I2) or fluorine (F2).

Another aspect of the present disclosure provides a process for the preparation of 14-(furan-2-yl)tetradeca-11,13-diynoic acid (14-FTDA), and salts thereof comprising: reacting furfural with triphenylphosphine (TPP), and carbon tetra bromide, in the presence of methylene dichloride to obtain 2-(2,2-dibromovinyl)furan; reacting 2-(2,2-dibromovinyl)furan with potassium hydroxide, and benzyltriethyl ammonium chloride, in the presence of methylene dichloride to obtain 2-(bromoethylnyl)furan; and coupling 2-(bromoethylnyl)furan with do-dec-11-ynoic acid, in the presence of copper chloride and hydroxylamine hydrochloride, to obtain 14-(furan-2-yl) tetradeca-11, 13-diynoic acid (14-FTDA).

Yet another aspect of the present disclosure, it provides a process for the preparation of 14-(furan-2-yl)tetradeca-11,13-diynoic acid (14-FTDA), and salts thereof comprising: reacting 2-ethynylfuran with sodium hydroxide and bromine to obtain 2-(bromoethylnyl)furan; and coupling 2-(bromoethylnyl)furan with do-dec-11-ynoic acid, in the presence of copper chloride and hydroxylamine hydrochloride, to obtain 14-(furan-2-yl) tetradeca-11, 13-diynoic acid (14-FTDA).

Still another aspect of the present disclosure, it provides a process for the preparation of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA), and salts thereof comprising: coupling of a halo-ethynyl furan with an acetylene acid having terminal alkyne, in the presence of a metal catalyst and a base, to obtain 14-(furan-2-yl) tetradeca-11,13- diynoic acid (14-FTDA); and reacting 14-(furan-2-yl)tetadeca-l 1,13-diynoic acid (14 FTDA) with alkali hydroxide to obtain salt of 14-(furan-2-yl)tetadeca-l 1,13-diynoic acid (14 FTDA).

The alkali hydroxide used in the present disclosure is selected from potassium hydroxide or sodium hydroxide.

In another embodiment, the disclosure provides potassium salt of the compound 14-(furan-2-yl)tetradeca-l 1,13-diynoic acid (14-FTDA), which is potassium 14-(furan-2-yl) tetradeca-11,13-diynoate of formula I-A.

In another embodiment, the disclosure provides sodium salt of the compound 14-(furan-2-yl)tetradeca-l 1,13-diynoic acid (14-FTDA), which is sodium 14-(furan-2-yl) tetradeca-11,13-diynoate of formula I-B.

The compounds 14-(furan-2-yl)tetradeca-l 1,13-diynoic acid (14-FTDA) and its salts, being highly soluble and suitable for formulation, provide highly effective components for formulations as fungicides for a variety of fungal pathogens in humans and animals.

In yet another embodiment, the disclosure also provides process for preparation of salts, derivative and analogs of the above compound, 14-(furan-2-yl)tetradeca-l 1,13-diynoic acid (14-FTDA). Such salts include alkali metal for example, potassium and sodium.

The salts are prepared by reacting 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) with aqueous potassium hydroxide or sodium hydroxide to get respective salts, namely, potassium 14-(furan-2-yl) tetradeca-11,13-diynoate and sodium 14-(furan-2-yl) tetradeca-11,13 -diynoate.

In one of the embodiment, 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) is prepared by the process described below:

To a suspension of Cu(I)Cl in an alcohol (methanol), 70% alkylamine (ethylamine) is added. The reaction mixture is cooled and hydroxyl amine hydrochloride is added. A solution of compound dodec-11-ynoic acid dissolved in an alcohol such as CI -4 alcohol such as methanol), is added slowly till the reaction mixture becomes brown suspension. The reaction mixture is stirred and allowed to attain room temperature between 20°C - 30°C. The reaction mixture is cooled again to about 0°C to 5°C and a solution of Bromoethynyl furanin alcohol (methanol) is added. The reaction mixture is allowed to attain room temperature 20 -30°C with stirring for 6 - 16 hours preferably overnight.

The above reaction mixture is concentrated to remove alcohol (methanol) and acidified preferably with 5M H2SO4 solution and diluted with ethyl acetate and then filtered through celitebed. The layers are separated; the aqueous layer is extracted with ethyl acetate. The combined organic layer was washed with brine, dried over anhydrous. Sodium sulphate filtered and concentrated. The crude mixture is purified by Column chromatography over Silica-gel using EA-Hexanes as eluent. The filtrate is concentrated and further recrystallized to get a compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA).

In another embodiment, the compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) is prepared in accordance with the present disclosure as:

Alkyl amine is added to a suspension of Cu(I)Cl in an alcohol. The reaction mixture is cooled and required amount of hydroxylamine hydrochloride is added in order to make the solution as colorless. To this solution, alcoholic solution of dodec-11-ynoic acid is added slowly. The reaction mixture is stirred at temperature ranging from 0 to 10°C and allowed to attain room temperature say 20 - 30°C. The above solution is cooled to 0-5°C and 2-bromo ethynyl furan in alcohol is added slowly. The reaction mixture is allowed to attain room temperature and it is stirred for 4-16 hours. Excess alcohol is removed and acidified with inorganic acid solution and diluted with ethyl acetate. Organic and aqueous layers are separated. The aqueous layer is extracted with ethyl acetate. The combined organic layer is washed with brine, dried over anhydrous sodium sulphate and concentrated. The crude mixture is purified through silica bed and re-crystallized with organic solvent to get 14-(furan-2-yl) tetradeca-11, 13-diynoic acid (14-FTDA) as off-white solid.

The disclosure provides preparation of the compound bromoethyl furan, used in the preparation of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) in accordance with the present disclosure, as described below.

To solution of carbon tetra bromide in dichloromethane, aromatic aldehyde is added. To the above, organophosphorus compound such as triphenylphosphine (TPP) is added. The reaction is carried out for a period of 6 - 16 hours maintaining the temperature between 0 to 10°C. Further, required quantity of petroleum ether is added and semi-solid mass is decanted. This is further extracted with organic solvent such as petroleum ether, diethyl ether, and hexane to obtain a aromatic dibromo vinyl compound.

To a 60% aq. KOH (5L) at 0°C, a solution of Benzyltriethyl ammonium chloride in a chlorinated solvent such as methylene dichloride (DCM) is added. Then a solution of Aromatic dibromo vinyl compound in chlorinated solvent such as methylene dichloride (DCM) is added maintaining the temperature at about -5 to +5°C. The reaction mixture is stirred and diluted with organic solvent such as methylene dichloride (DCM). The layers were separated and the aqueous phase is extracted. The combined organic layer is dried over anhydrous sodium sulphate and passed through a bed of silica to obtain a bromoethynyl furan.

In yet another embodiment, the disclosure provides preparation of the compound bromoethyl furan, in accordance with the present invention, as-2-ethynylfuran in dioxane is reacted with bromine in aqueous sodium hydroxide solution. The reaction mass is separated by adding ice cold water and extracted with diethyl ether followed by removal of solvent to obtain a brominated ethynyl furan compound.

In accordance with the present disclosure, chlorinated solvents can be any solvent selected from chlorofprm, methylene dichloride, ethylene dichloride and a mixture thereof.

The organophosphorus compound is triphenylphosphine (TPP). The alcohol is selected from CM such as methanol, ethanol, propanol and butanol. The alkylamine is selected from alkyl amine, a secondary alkyl amine, and a tertiary alkyl amine such as methylamine (mono, di and tri), ethylamine (mono and di), isopropyl amice (mono-n-propyl amime, di-n-propyl amine and tri-n-propyl amine), butylamine, cyclohexyl amine, ethylenediamine, ethanolamine, propylamine, etc.

The previously described versions of the subject matter and its equivalent thereof have many advantages, including directly coupling reaction ofacetylenic acid i.e., Dodec-11-ynoic acid with halo-ethynyl compound, without obtaining the acid in ester form. Hence the process also avoids further hydrolysis of ester form of the 14-(furan-2-yl) tetradeca-11,13-diynoic acid (H-FTDA).In addition, the optimization of fine coupling reaction directly on acid moiety i.e. Dodec-11-ynoic acid with bromo-2-ethynyl furan makes the process simpler and more viable for upgrading to higher scale production.

The compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salts thereof of the present disclosure are useful as fungicides against any one or more fungal pathogen selected from the group consisting of Candida spp. (for example C. albicans, C. krusei, C. glabrata, C. tropicalis, C. parapsilosis, C. guilliermondii, C. haemulonii, C. lusitaniae, C. lipolytica, C. norvegensis, C. viswanathii, C. kefyr or C. dubliniensis), Aspergillus spp. (for example A. fumigatus, A. flavus, A. niger or A. terreus,) Histoplasma capsulatum, Coccidioides immitis, Coccidioides posadasii, Cryptococcus spp. (for example C. neoformans (for example var. neoformans or var. gattii), C. bidus, C. laurentii, or C. fusarium), Zygomycetes (such as Rhizopus oryzae, R. micropsorus, R. pusillus, Cunninghamelle bertholletiae, Saksenaea vasiformis, Mucor circinelloides, M. ramosissimus, Absidia corymbifera, Apophysomyces elegans, Cokeromyces recurvatus or Syncephalastrum racemosum), Malassezia spp. (for example M. furfur or M. globosa), Hyalohyphomycetes (for example Fusarium solani or Scedosporium spp., such as S. prolificans or S. apiospermum), Dermatophytes (for example Trichophyton spp. (for example T. mentagrophytes, T. rubrum or T. tonsurans), Epidermophyton floccosum, Microsporum spp (for example M. cookei, M. canis, M. vanbreuseghemii, M. gallinae or M. gypseum) or Trichosporon terrestre), Blastomyces dermatitidis, Sporothrix schenkii, Chromomycotic fungi (for example Fonsecaea pedrosoi, F. compacta, Cladophylophora carrionii or Phialophora verrucosajand Madurella spp. (for example M. mycetomatis or M. griseum), Pneumocystis jirovecii, Pneumocystis carinii, Ascomycota Botiytis cinerea; Magnaporthe grisea; Anamorph: Pyricularia oryzae Colletotrichum gleoesporioides- Chilli strain; Colletotrichum gleoesporioides- mango strain; Fusarium verticillioides; Fusarium oxysporum; Alternaria solani; Uncinula necator Syn Erysiphe necator; Macrophomina phaseolina; Syn. Sclerotium bataticola and Rizoctonia bataticola; Botryodiplodia theobromae; Basidiomycota Sclerotium rolfsii; Rhizoctonia solani; Puccinia arachidis; Oomycota Pythium aphanidermatum; and Plasmopara viticola Syn. Personopora viticola.

The compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salts thereof of the present disclosure provides a fungicidal formulation for use in an agricultural setting or of enhancing the fungicidal activity of a formulation for use in an agricultural setting, comprising adding one or more of the inventive fungicides to a formulation.

In preferred embodiments, the formulation is used to combat a fungal pathogen in a plant, a grass or a field.

The compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salts thereof of the present disclosure is used for the preparation of a treatment for an agricultural condition which can be improved or prevented by treatment of the agricultural condition with an Olel protein inhibitor.

The present disclosure also provides a kit for an agricultural fungicide comprising one or more of the inventive compounds or fungicides of structure 14-(furan-2-yl) tetradeca-11,13 -diynoic acid (14-FTDA) or its salts thereof FORMULATIONS

The inventive compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) disclosed herein should be understood to include any pharmaceutical^ acceptable salts encompassing either salts with inorganic acids or organic acids like hydrohalogenic acids, e.g. hydrochloric or hydrobromic acid; sulfuric acid, phosphoric acid, nitric acid, citric acid, formic acid, acetic acid, maleic acid, tartaric acid, methanesulfonic acid, p-toluenesulfonic acid and the like or in case the compound is acidic in nature with an organic base such as, for example, triethylamine, triethanolamine, tert-burylamine, or an inorganic base like an alkali or earth alkali base, e.g. sodium hydroxide, potassium hydroxide, calcium hydroxide etc.

Because of their ability to inhibit a wide variety of fungal pathogens occurring in humans and/or animals, occurring both systemically and topically, the compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salts thereof can be used for the treatment of diseases which are associated with an infection by such type of pathogens. They are valuable antifungal treatments.

The compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salts thereof can be administered orally, rectally, parenterally e.g. by intravenous, intramuscular, subcutaneous, intrathecal or transdermal administration or sublingually or as ophthalmic preparation or administered as aerosol. Examples of applications are capsules, tablets, orally administered suspensions or solutions, intravenous solutions, suppositories, injections, eye-drops, ointments or aerosols/nebulizers.

Preferred applications are oral or i.v. systemic formulations and ointment, pellet, liquid or liquid suspension topical formulations. The dosage used depends upon the type of the specific active ingredient, the age and the requirements of the patient and the kind of application. The preparations with the inventive compounds can contain inert or as well pharmacodynamically active excipients. Tablets or granules, for example, could contain a number of binding agents, filling excipients, carrier substances or diluents.

These pharmaceutical compositions may contain the compounds of the invention as well as their pharmaceutically acceptable salts in combination with inorganic and/or organic excipients which are usual in the pharmaceutical industry like lactose, maize or derivatives thereof, talcum, stearic acid or salts of these materials.

For gelatine capsules vegetable oils, waxes, lipids, liquid or half-liquid polyols etc. may be used. For the preparation of solutions and syrups e.g. water, polyols, saccharose, glucose etc. are used. Injectables are prepared by using e.g. water, polyols, alcohols, glycerin, vegetable oils, lecithin, liposomes etc. Suppositories may be prepared by using natural or hydrogenated oils, waxes, fatty acids (fats), liquid or half-liquid polyols etc.

The compositions may contain in addition preservatives, stabilisation improving substances, viscosity improving or regulating substances, solubility improving substances, sweeteners, dyes, taste improving compounds, salts to change the osmotic pressure, buffer, antioxidants etc.

Figure 1 shows (upper pictures, controls) normal hyphal growth as compared with changes in the plane of hyphal growth and abnormal thickening of the hyphae due to the antifungal effect of the potassium salt of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) (two lower pictures).

Mechanism of inhibition of Olel protein

An Olel protein inhibitor is inherently fungicidal because the Olel protein is an essential protein to the fungal organism. In the biosynthesis of lipids, the Olel protein converts stearic acid to oleic acid. Oleic acid is an essential component of lipids and thus essential to the fungal organism. Without oleic acid the organism fails to survive due to collapse of the nuclear membrane. Agricultural fungal infection

The Olel protein inhibitors of the present disclosure provide potent broad spectrum antifungal agents for a wide variety of agricultural purposes. The inventive Olel protein inhibitors are suitable and efficacious for treating a fungal infection in the agricultural setting, including reducing the risk of a fungal infection, and in particular may be used for methods of treating an infection in a plant, or a grass, by contacting a plant with an Olel protein inhibitor according to the invention. Plants include trees, crops, grasses, and flowering plants.

Another aspect of the present disclosure relates to a pesticide composition comprising compound 14-(furan-2-yl)tetradeca-11,13-diynoic acid (14-FTDA) or its derivatives, especially its salts, such as compound sodium salt, or its analogs, providing effective and potent Olel protein inhibitors, in defeating or lessening agricultural fungal pathogens, providing effective and potent compounds for use in the agricultural setting.

Further, the present disclosure contemplates methods for preventing or controlling fungal infections in plants, parts of plants, seeds, or at their locus of growth.

Efficacy in Agricultural species

The compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salts and analogs and derivatives have been tested on various agricultural setting fungal pathogens namely Ascomycota Botiytis cinerea; Magnaporthe grisea anamorph: Pyricularia oryzae; Colletotrichum gleoesporioides- Chilli strain; Colletotrichum gleoesporioides- mango strain; Fusarium verticillioid.es; Fusarium oxysporum; Alternaria solani; Uncinula necator Syn Erysiphe necator; Macrophomina phaseolinaSyn. Sclerotium bataticola and Rizoctonia bataticola; Botryodiplodia theobromae; Basidiomycota Sclerotium rolfsii; Rhizoctonia solani; Puccinia arachidis; Oomycota Pythium aphanidermatum; Plasmopara viticola Syn. The criteria for evaluating the compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salt thereof was based on (i) yield losses and disease severity caused on crops and other plants (for example, ornamental and amenity grasses); (ii) host infected by fungi; (iii) difficulty in providing control measures with existing fungicides; representation across major classes of pathogenic fungi; and representation across major groups of fungal diseases viz., rust, rot (root and fruit), leaf spots, mildews and wilts.

Conidia/spores are the major source of spreading diseases, and if the sporulation is affected, disease spread in the farming field is contained. Thus, inhibiting sporulation is an indirect way of conducting disease control. Moreover, if sporulation is affected, the emergence of disease resistance is minimised, because the genetic changes which make the pathogen to adopt for the fungicide will not be carried to the next generation. The asexual fruit body of the plant pathogenic fungus Colletotrichum gloeosporioid.es is called acervulus. Acervuli, visible to the naked eye and salmon colour, are produced in concentric circles. When the potassium salt of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) was loaded on a sterile paper disc in the path of pathogen growth, the growth of the mycelia is arrested. Although the pathogen continues growth somewhat, there is no sporulation observed in mycelia grown around the region where the disc is loaded with test compound. The mycelia grown in the region diffused with the test compound was weak and did not differentiate into conidiophores - no sporulation observed. Malformation and inhibition takes place in the spore germination in M. grisea.

Spore germination in the M. grisea control starts with small beak-like germination, which extends into long germ tube and culminates into an appressorium. The appressorium will be densely melanized to withstand the high turgor pressure created during penetration of infection peg through the plant cell wall. The germ tube starts from one or two terminal cells of a three-celled spore, and the process is completed in 8-hrs. When the spores were incubated in sterile water containing different concentrations of test compound (potassium salt of compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA)), more than 50% of spores did not germinate, and where small beak-like germination started in germinated spores: in some more spores, though the appressorium is formed, they were not melanised enough to withstand the pressure; breakage in the germ tube near the formation of appressoria was observed; some appressoria burst due to turgor pressure; and in some spores germ tubes formed from the middle cell, instead of from the terminal cells.

Resistant fungus infections

Fungal infections may be resistant to treatment for many reasons, resistant to treatment with a particular antifungal agent, or because of acquired resistance. The compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) or its salts thereof are used for treating infection by a fungus resistant to one or more alternative treatment and that acts via: a) inhibiting ergosterol biosynthesis;

b) binding to ergosterol;
c) inhibiting 1, 3-0-glucan synthase;
d) inhibiting epoxidase;
e) inhibiting Leucyl-tRNA synthetase; and/or
f) inhibition of elongation factor 2.

Particularly, such resistant antifungal treatments may be benzoic acid, ciclopirox, tolnaftate, undecylenic acid, flucytosine, griseofulvin, haloprogin and sodium bicarbonate or may be polyenes, azoles, allylamines or echinocandins. Polyene antifungal agents have multiple conjugated double bonds, and typically, also comprise a heavily hydroxylated region, exemplified by Natamycin, Rimocidin, Filipin, Nystatin, Amphotericin B or Candicin. Azole antifungal agents may for example be imidazole or triazole or thiazole antifungal agents. Imidazole antifungal agents may for example include miconazole, ketoconazole, clotromazole, econazole, bifonazole, butoconazole, fenticonazole, isoconazole, oxiconazole, seraconazole, sulconazole or tioconazole. Triazole antifungal agents may for example include fluconazole, itraconazole, isavuconazole, ravuconazole, posaconazole, voriconazole, terconazole, and abafungin. Allylamine antifungals include Terbinafine, Amorolfine, Naftifine or Butenafme. Non-limiting examples of echinocandins include Anidulafungin, Caspofungin or Micafungin.

The compound 14-(furan-2-yl)tetadeca-11,13-diynoic acid (14 FTDA) and salts thereof can be prepared as described above in detail; is outlined in the reaction schemes as described hereinafter, and the description thereof.

Scheme 1: General route in accordance with the present disclosure for the preparation of 14-(furan-2-yl)tetadeca-l 1,13-diynoic acid (14 FTDA);

14-(furan-2-yl)tetadeca-11,13-diynoic acid (14-FTDA) is prepared by reacting with halo-ethynyl furan and acetylenic acid in the presence of copper chloride and a base. Scheme 2: Route for the preparation of halo-ethynyl furan (HI):


Furfural is reacted with triphenylphosphine (TPP), and carbon tetra halide, in the presence of chlorinated solvent to obtain 2-(2,2-dihalovinyl)furan (II); and

2-(2,2-dihalovinyl)furan (II) is reacted with an alkali hydroxide, and benzyltriethyl ammonium chloride, in the presence of chlorinated solvent to obtain a halo-ethynyl furan (HI). Scheme 3: Route for the preparation of halo-ethynyl furan (III):

2-ethynylfuran is reacted with an alkali hydroxide and halogen to obtain a halo-ethynyl furan (III).

Examples

The disclosure will now be illustrated with working examples, which is intended to illustrate the working of disclosure and not intended to take restrictively to imply any limitations on the scope of the present disclosure.

Example 1: Preparation of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA): Step 1: Preparation of 2-(2,2-dibromovinyl) furan (compound 2)

To a stirred solution of CBr4(51.82g; 0.156 mol) in methylene dichloride (DCM) (300 ml), furfural (lOg, 0.104 mol) was added at room temperature. The reaction mixture was cooled to 5 to 10°C. A solution of triphenylphosphine (TPP) (81.83g; 0.132 mol) in methylene dichloride (DCM) (100 ml) was added over a period of 3 hours maintaining the temperature at about 0°C-5°C. The above reaction is exothermic. The reaction mixture was stirred for 6 to 8 hours maintaining the temperature between 10-15°C. TLC was checked.

Required quantity of petroleum ether was added in the reaction mixture. The reaction mass was taken out from the reactor. The clear solution was decanted from the precipitate (semisolid mass) and filtered through a bed of celite. The clear solution was evaporated under vacuum to gummy residue. It was further extracted with petroleum ether and concentrated to obtain 2-(2,2-dibromovinyl) furan (compound 2) as brown oily liquid (yield 60%) Step 2: Preparation of 2-(Bromoethynyl) furan (compound 3)

To a solution of benzyl tri ethyl ammonium chloride (2gm, 0.0089mol) in methylene dichloride (DCM), a solution of 2-(2,2-dibromovinyl) furan (9g, 0.035 mol) in methylene dichloride (DCM) was added slowly maintaining the temperature 0°C. Then to this a 60% solution of potassium hydroxide (KOH) (35ml) was added slowly. The reaction mixture was stirred at 0°C for lh. The reaction mixture was then diluted with methylene dichloride (DCM) and stirred for lOmin. The layers were separated and the aqueous phase was extracted with methylene dichloride (DCM). The combined organic layer was dried over anhydrous sodium sulphate and passed through a bed of silica. It was concentrated and stored as such under cold condition. (Yield: 96%) Step 3: Preparation of 14-(furan-2-yl) tetradeca-11, 13-diynoic acid (14-FTDA)

To a suspension of Cu(I)Cl (3.56g, 0.035 mol) in methanol, 70% ethyl amine (15.49g, 0.344mol) was added. The reaction mixture became blue in color. It was cooled under ice-salt bath and hydroxyl amine hydrochloride (3.52g, 0.051 mol) was added in portion till the reaction became colorless. A solution of dodec-11-ynoic acid (5gm, 0.025 mol) dissolved in methanol was added slowly, reaction mixture becomes brown suspension. Reaction mixture was stirred at 0°C for 10 min and then ice bath was removed and allowed to come at RT. Again it was cooled to 0°C and a solution of 2-bromo ethynyl furan (6.07g, 0.035 mol) in methanol was added slowly. The reaction mixture was allowed slowly to warm up to RT and stirred for overnight. Then methanol was removed from the reaction mixture and acidified it with 5M H2SO4 solution and was diluted with ethyl acetate. It was filtered through celite bed. The layers were separated. The aqueous layer was extracted with ethyl acetate. The combined organic layer was washed with brine, dried over anhyd. sodium sulphate filtered and concentrated. The crude mixture was purified through a silica bed and further recrystallized from pet ether to get off-white solid (yield 45%) Example 2: Preparation of 14-(fiiran-2-yl) tetradeca-11,13-diynoic acid (14-FTDA):

Step 1: Preparation of 2-BromoEthynyl Furan:

Bromine (9.46g, 59.2 mmol) was added to a solution of ION aqueous NaOH (13.6ml) under stirring at 0-5°C. 2-ethynylfuran(5gm,54.34 mmol) as a solution in 1,4 dioxane (25ml) was added drop wise to the above reaction mass over a period of 30 minutes. Reaction mixture was continued under stirring for another 30 min without further cooling and poured 100ml of ice cold water in to the reaction mixture. The product was extracted with diethyl ether and dried over magnesium sulphate followed by the removal of solvent. The product was obtained as yellow oil. Step 2: Preparation of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA):

70% ethyl amine (15.49g, 0.344mol) was added to a suspension of Cu(I)Cl (3.56g, 0.035 mol) in methanol. The reaction mixture changes in to blue in color. It was cooled under ice-salt bath and hydroxyl amine hydrochloride (3.52g, 0.051 mol) was added in portion till the reaction became colorless. A solution of dodec-11-ynoic acid (5gm, 0.025 mol) dissolved in methanol was added slowly, reaction mixture becomes brown suspension. Reaction mixture was stirred at 0°C for 10 min and then ice bath was removed and allowed to come at RT. Again it was cooled to 0°C and a solution of 2-bromo ethynyl furan (6.07g, 0.035 mol) in methanol was added slowly. The reaction mixture was allowed slowly to warm up to RT and stirred for overnight. Then methanol was removed from the reaction mixture and acidified it with 5M H2SO4 solution and was diluted with ethyl acetate. It was filtered through celite bed. The layers were separated. The aqueous layer was extracted with ethyl acetate. The combined organic layer was washed with brine, dried over anhyd. sodium sulphate filtered and concentrated. The crude mixture was purified through a silica bed and further recrystallized from pet ether to get off-white solid (yield 45%).

Example 3: Antifungal efficacy in-vitro

The in-vitro efficacy of potassium salt of 14-(furan-2-yl) tetradeca-11, 13-diynoic acid (14-FTDA) was investigated on 8 Candida reference strains. The efficacy was determined according by broth micro dilution and the MIC given as ng/ml (ref table 1) Table 1: Susceptibility of reference Candida strains to compound Potassium salt of 14-(furan-2-yl) tetradeca-11, 13-diynoic acid (14-FTDA).

Example 4:

Agriculture Assays:

In vitro mycelial growth inhibition assays - poisoned plate:

Growth of fungi was carried out in potato dextrose agar media at 40-450C3 and test compound was added at different concentrations and at pH 5.8 for compound 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA) and at pH 7.0 for its potassium salt, with a water control instead of test compound. Radial mycelial growth was measured at regular intervals 4, 8, 12 and 16 days for slow growing fungi and at 3, 6 and 8 days for fast growing fungi. When the mycelia reached the end of the plate, measurements were stopped. Morphological changes in the hyphal growth and sporalation patterns were also observed. For dose response studies a range of concentrations and a range of inhibition obtained falling below and above 50% inhibition (see Table 2 below) Spore germination studies for plant fungal pathogens- hanging drop method:

Spore/conidial suspension of 5-10x103 spores per ml (for larger spores) and 5-10 x 105 for smaller spores, was tested against different concentrations of the potassium salt of compound 14FDTA. Spore germination was carried out in sterile distilled water, under a moist chamber, in cavity slides. After overnight incubation the spores were observed. Where applicable, the solvent dimethyl sulfoxide was added to both test and control. Spore suspensions were prepared from good sporulating field isolates and grown in appropriate media, generally potato dextrose agar, or for Magnaporthe grisea oatmeal agar. For specific sporulating structure, spore suspensions were prepared devoid of mycelial bits. Spore number was adjusted using haemocytometer. Final spore concentration of spores was 5-10 X 103 spores per ml. Photographic recordings were made of perfect/good germination; recording of any malformation such as disintegration, shrinking of germ tubes or spores. Inhibition was calculated by comparison with the control germination (inhibition = [(% of spore germination in control with DMSO - % of spore germination in treated with compound)/( [(% of spore germination in control with DMSO)]. Leaf disk assay:

Leaf disk assay was carried out by the cavity well plate method for powdery mildew disease of grapes. Leaf disks of 14 mm in diameter were cut with a cork borer from the healthy leaves (second and third from the tip) of grapevine plants, and were dipped in lOO uJ of each test compound at different concentrations for two minutes, as in treatment details shown below. The control leaf disks were dipped only in sterile water for two minutes. The compound treated leaf disks were placed, abaxial side up, in TC-24 well plates containing water agar medium. The disks were inoculated by placing 20|xl of inoculums (1-5 x 106 spores/ml) on the centre of the disk. After inoculation, the cavity well plates were incubated at 20°C for 10 days. After incubation, the powdery mildew lesions on the leaf disks were rated to 0-9 scale, in which, 0 was no visible symptoms and 9 represented more than 50% leaf area with mildew growth/lesion. Percent disease index (PDI) was calculated as follows:

PDI= Sum of individual ratings x 100
Total no. of leaf disk maximum disease grade observed

After the observation, the conidia were washed from the leaf disks in known volume of a fixative solution of ethanol-formaldehyde-acetic acid (90:5:5, v/v/v) and counted with a hemocytometer.

Table 2. Mycelial growth inhibition

Changes in the hyphal tips of Botrytis cinerea using potassium salt of 14-(furan-2-yl) tetradeca-11,13-diynoic acid, were investigated to measure the effect of the Olel inhibitor upon growth and thriving. The compound was found to be toxic to the fungus Botrytis cinerea as described in the following. Apical dominance is an important criterion for growth of hyphae of fungi: at the apical tip of the mycelia branching is not seen near the growing tip. Apical dominance is maintained but branching of hyphae will start at sub-apical point, a distance away from the growth point. Under abnormal conditions of stress, apical dominance is lost, extensive branching begins, resulting in the growth of the fungal mycelia being arrested. When the test compound is incorporated in the media in which the fungi is present, hyphal tip splitting and branching is seen with loss of apical dominance and polarity. Figure 1 show (in the two lower pictures) the changes in the plane of hyphal growth and abnormal thickening of the hyphae when the mycelia are inoculated in the plates with potassium salt of 14-(furan-2-yl) tetradeca-11,13-diynoic acid, (2ug/ml).

Hyphae from the inoculated disc start growing against gravity and are thicker than the normal hyphae seen in the control (the two upper pictures). Abnormal thickening of the hyphae shows the stress created by the presence of the Olel inhibitor.

Although the subject matter has been described in considerable detail with reference to certain preferred embodiments thereof, other embodiments are possible. As such, the spirit and scope of the appended claims should not be limited to the description of the preferred embodiment contained therein.

I/we claim;

1. A process for the preparation of 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA), and salts thereof comprising:

coupling of a halo-ethynyl furan with an acetylene acid having terminal alkyne, in the presence of a metal catalyst and a base, to obtain 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA).

2. The process as claimed in claim 1, wherein the halo-ethynyl furan is selected from fluro-ethynyl furan, chloro-ethynyl furan, bromo-ethynyl furan, or iodo-ethynyl furan.

3. The process as claimed in claim 1, wherein the acetylene acid is do-dec- 11-ynoic acid.

4. The process as claimed in claim 1, wherein the metal catalyst is a copper catalyst.

5. The process as claimed in claim 1, wherein the base is hydroxylamine hydrochloride.

6. The process as claimed in claim 1, wherein halo-ethynyl furan is prepared by a process comprising:

reacting furfural with triphenylphosphine, and carbon tetra bromide, in the presence of chlorinated solvent to obtain 2-(2,2-dibromovinyl)furan; and

reacting 2-(2,2-dibromovinyl)furan with an alkali hydroxide, and benzyltriethyl ammonium chloride, in the presence of chlorinated solvent to obtain a halo-ethynyl furan.

7. The process as claimed in claim 6, the chlorinated solvent is selected from chloroform, methylene dichloride, ethylene dichloride, or mixture thereof.

8. The process as claimed in claim 1, wherein the halo-ethynyl furan is prepared by reacting 2-ethynylfuran with an alkali hydroxide and halogen.

9. The process as claimed in claim 6 or 8, wherein the alkali hydroxide is selected from sodium hydroxide (NaOH) or potassium hydroxide (KOH).

10. The process as claimed in claim 8, wherein the halogen is selected from bromine (Br2>, chlorine (CI2), iodine (I2) or fluorine (F2).

11. The process as claimed in claim 1, comprising:

reacting furfural with triphenylphosphine (TPP), and carbon tetra bromide, in the presence of methylene dichloride to obtain 2-(2,2-dibromovinyl)furan;

reacting 2-(2,2-dibromovinyl)furan with an potassium hydroxide, and benzyltriethyl ammonium chloride, in the presence of methylene dichloride to obtain 2-(bromoethylnyl)furan; and

coupling 2-(bromoethylnyl)furan with do-dec-11-ynoic acid, in the presence of copper chloride and hydroxylamine hydrochloride, to obtain 14-(furan-2-yl) tetradeca-11,13-diynoic acid (14-FTDA).

12. The process as claimed in claim 1, comprising:

reacting 2-ethynylfuran with an sodium hydroxide and bromine to obtain 2-(bromoethylnyl)furan; and

coupling 2-(bromoethylnyl)furan with do-dec-11-ynoic acid, in the presence of copper chloride and hydroxylamine hydrochloride, to obtain 14-(furan-2-yl) tetradeca-11, 13-diynoic acid (14-FTDA).

13. The process as claimed in claim 1, further comprising:

reacting 14-(furan-2-yl)tetadeca-11,13-diynoic acid (14 FTDA) with alkali hydroxide to obtain salt of 14-(furan-2-yl)tetadeca-l 1,13-diynoic acid (14 FTDA).

14. The process as claimed in claim 13, wherein the alkali hydroxide is selected from potassium hydroxide or sodium hydroxide.