Title of Invention	NOVEL SYNERGISTIC COMBINATION OF FERULA ASAFOETIDA WITH DEHYDRATED DIALDOL PRODUCT AS POTENT ANTIFUNGAL AGENT
Abstract	The present invention relates to a novel synergistic antifungal composition for exhibiting antifungal activity against Rhizoctonia bataticola wherein the composition comprises: a synthetic compound of formula 1; and b) Ferula asafoetida said a synthetic compound of formula 1 and Ferula asafoetida being taken in a desired amount and said synthetic compound of formula (1) and Ferula asafoetida having mixed together under ambient conditions to obtain the desired composition.

Title of Invention

NOVEL SYNERGISTIC COMBINATION OF FERULA ASAFOETIDA WITH DEHYDRATED DIALDOL PRODUCT AS POTENT ANTIFUNGAL AGENT

Abstract

The present invention relates to a novel synergistic antifungal composition for exhibiting antifungal activity against Rhizoctonia bataticola wherein the composition comprises: a synthetic compound of formula 1; and b) Ferula asafoetida said a synthetic compound of formula 1 and Ferula asafoetida being taken in a desired amount and said synthetic compound of formula (1) and Ferula asafoetida having mixed together under ambient conditions to obtain the desired composition.

Full Text	NOVEL SYNERGISTIC COMBINATION OF FERULA ASAFOETIDA WITH DEHYDRATED DIALDOL PRODUCT AS POTENT ANTIFUNGAL AGENT FIELD OF THE INVENTION: The present invention relates to a synergistic combination of Ferula asafoetida with dehydrated dialdol product as potent antifungal agent. The present invention relates to the formation of environmentally safe fungicide taking advantage of the bioenhancer characteristics found in natural products and bioefficacy of synthetic compound, which in a way full fill the main objective of inventing an eco friendly fungicide. The presence of natural products in formulation allows the use of synthetic compound at lower concentration with enhanced in vitro activity against the pathogenic fungus. BACKGROUND: A variety of crop diseases owe their origin to pathogenic microorganisms, which include bacteria, virus and fungi. Among them fungi are responsible for a major loss of crops all over the world. Rhizoctonia bataticola is considered as one of most noxious fungus [1-4 ] which is constantly seeking attention of scientist throughout the world. Though a number of chemical, broad-spectrum fungicides available for the control of fungus that attack and damage food crops and economically important crops [5-8]. These chemical fungicides aim at fungus eradication and kill the fungus. In this process many non target species are killed and in addition they not only affect the ecosystem but also pollute the atmosphere with chemical, the water we drink, air we breathe and food we eat. Fungi also develop resistance to the fungicide in the course of time on continuous usage. Hence, there is a prime need for antifungal agent, which is biodegradable, eco friendly, does not affect the beneficial organisms, non-toxic to human beings and at the same time effective against the target fungus. This has diverted the attention of scientists towards product of natural origin. Though natural products are rich source of anti-infective, they carry their own limitations of less potency as compared to synthetic chemicals, low availability, high cost of extraction, structural complexity and instability. In this regard superiority of combination consisting of mixture of bioactive compounds over a single isolated constituent has been well established in which one of the compound generally natural in origin, acts as bioenhancer. This effect is known as synergism which can be defined as a phenomenon in which two different compounds are combined to enhance their individual activity. The prior art reveals the activity of only herbal compositions containing Ferula asafoetida as one of the ingredients. These herbal compositions are tested for the treatment of human disease only. US2005089583 discloses a synergistic herbal composition for the treatment of gastric ulcer, said composition essentially comprising of powdered plant parts of Asperagus racemosus, Glycyrrhiza glabra, Seaamum Indicum, Musa sapientum and Trachyaparmum roxburghicinum and optionally, powdered plant parts of Cyclea peltate, Embelia ribes, Coriandrum sativum, Ferula asafoetida, Aloe barbadensis and Evoluulus aisinodes along with one or more pharmaceutically acceptable additives/carriers. The composition comprises powdered plant parts of Asperagus racemosus, Glycyrrhiza glabra, Seaamum Indicum, Musa sapientum and Trachyaparmum roxburghicinum in equal proportions, optionally along with one or more pharmaceutically acceptable additives/carriers. This composition is hereafter referred to as HF3. [0021] In another embodiment of the present invention, the composition comprises powdered plant parts of Asperagus racemosus, Glycyrrhiza glabra, Seaamwn indicum, Musa sapientum, Trachyaparmum roxburghicinum, Evolvulus aisinodes and Ferula asafoetida in equal proportions, optionally along with one or more pharmaceutically acceptable additives/carriers US Patent 5,482,711 discloses a method for activating immune competent cells in humans in order to increase the immune function in humans, the immune competent cells being selected from the group consisting of CD 19, HLADR, NKCD3-/CD56+ and CD38, the method comprising administering to humans an effective dose of an extract from Nigella sativa at a concentration which is effective to activate the immune competent cells by reducing the presence of interferon inhibitor factor or lymphokine inhibitor factor. US Patent 5,653,981 discloses a pharmaceutical unit dosage form for the inhibition of cancer cell growth in humans consisting essentially of a cancer cell growth-inhibiting amount of an extract from Nigella sativa wherein the amount is within a range of from about 0.3 g/kg body weight to about 0.6 g/kg body weight and a pharmaceutically-suitable carrier. US Patent 5,952,373 discloses a method for the treatment or for the prophylactic treatment of hyperreactive skin predisposed to dermatitis, deficient, hypoactive skin or dermatoses which comprise applying to said skin an effective amount of a composition comprising one or more flavonoids. US Patent discloses 6,121,243 relates in particular to cosmetic and dermatological formulations comprising flavonoids, their glycosides and, if appropriate, combinations thereof with cinnamic acid derivatives or antioxidants. The damaging effect of the ultraviolet component of solar radiation on the skin is generally known. While rays having a wavelength below 290 nm (the so-called UVC range), are absorbed by the ozone layer in the earth's atmosphere, rays in the range between 290 nm and 320nm, the so-called UVB range, cause erythema, simple sunburn or even burns of greater or lesser severity. The narrower range around 308 nm is stated as the maximum for the erythema activity of sunlight. US Patent 6,180,662 particularly relates to active compounds and formulations, in particular for topical use, which are used for prophylaxis and treatment of hyperreactive skin predisposed to dermatitis, and of deficient hypoactive skin and for prophylaxis and treatment of the manifest dermatoses mentioned under I. to XIII., such as, for example, atopic dermatitis, neurodermatitis, atopic eczema and seborrhoeic dermatitis, photoinduced dermatoses (for example Mallorca acne and in particular polymorphic photodermatosis and photodermatitis), rosacea, prurigo forms, pruritus, psoriasis forms, ichthyosis, decubitus, ulcus cruris and microbial and viral infections, such as, for example, herpes simplex, h.zoster or h.labialis. US patent 6,423,747 discloses method of stabilizing or reestablishing the epidermal barrier function of the skin which comprises applying to said skin a formulation comprising a) one or more compounds selected from the group consisting of flavonoids, or b) an active compound combination comprising one or more compounds selected from the group consisting of flavonoids in combination with one or more compounds selected from the group consisting of cinnamic acid derivatives, and c) optionally, one or more compounds selected from the group consisting of antioxidants. US Patent 6,562,794 discloses a method for the treatment or prophylaxis of skin against aging and the inflammation of skin caused by exposure of the skin to oxidation or for protecting the skin and the hair from photoreactions which comprises applying an effective amount of a formulation comprising, a) alpha.-glucosyl rutin, and b) one or more compounds selected from the group consisting of antioxidants to said skin or hair. Accordingly, we have formulated herein a novel composition comprising Ferula asafoetida and Synthetic compound of formula 1 against plant pathogen for the first time. The activity of F.asafoetida never been tested either alone or in combination against phytopathogen ,R.bataticola. SUMMARY OF THE INVENTION: The present invention deals with a combination of synthetic compound of formula (1) and Asfoetida (N1) which shows tremendous activity against the test fungus. The present invention is aimed to overcome or avoid problems faced in the control of fungus as mentioned in the literature. The synergistic interaction provided by the composition herein allows reduction of the quantity and improvement of efficacy of synthetic compound by administrating with mildly active natural product (Nl). This reduction allows usage of fungicide which is otherwise too highly toxic or produce unacceptable side effects at fungicidal dosages, allows usage at dosages less than those which produce side effects and should reduce or at least delay the occurrence of natural resistance to important chemical fungicides. The use of products of the present invention offer a low dose treatment with enhanced action comparable to that of standard dose alone. BRIEF DESCRIPTION OF THE CHEMICAL STRUCTURE Formula 1: Chemical structure of compound of formula l:l,5-bis(2- hydroxyphenyl)pent-l,4-diene-3-one. Figure 1: Graphical representation of percentage inhibition of various combinations applied against the test fungus Detailed description of the present invention: The invention pertains to method for increasing the antifungal activity of newly identified anti Rhizoctonia bataticola agent, by administering a mildly active naturally occurring product in combination with it, thereby reducing the effective amount of the synthetic compound required as compared to the amount required when it is administered alone. The present invention provides a newly identified antifungal agent (formula 1) which shows antifungal activity at very low concentration when tested alone and also its synergistic combination with Asafoetida which enhances its activity further at the same concentration. Compound of formula 1 can be prepared following standard method as reference 9, shown in Scheme 1 wherein requisite aldehyde was dissolved in ethanol and NaOH. To this reaction mixture, acetone was added slowly in an incubator-shaker at room temperature. After half an hour of continuous shaking the reaction mixture resulted to a clear pale yellow solution. Further shaking for about 10 minutes, cloudiness appeared which settled down slowly. The precipitate was then filtered out and re-crystallized by appropriate solvent. (Scheme Removed) (Scheme 1) Formula 1 (Synthetic compound I) The activity of the samples was determined by standard food poisoning method [as reference 10]. Wherein, Potato dextrose agar of HI MEDIA, MU096 was used as culture medium and test samples were dissolved in acetone. Single spore cultures of fungal species, grown and maintained on potato dextrose agar media at 28±1°C were used for the experiments. The compounds were mixed aseptically with PDA media and were sterilized. The medium with acetone was served as control. The Petri plates having 9 cm inner diameter were taken, and 25 mL of the sterilized PDA containing test samples were plated in the centre. After solidification, 0.5 cm diameter agar discs taken from a 48 h old sporulated fungi culture was placed with the fungal side downward in the centre of each plate with the help of sterile cork borer and glass rod. The plates were incubated in dark at 28±1°C. Radial growths were determined by measuring the colony size along two diameters at right angles after 24 h interval. Three replicates for each treatment were prepared. The experiments were repeated three times. The percentage of inhibition was calculated using the following formula: I = C - T / C x 100, where, I = percentage of inhibition; C = radial growth in control -disc diameter (cm); T = radial growth in treated - disc diameter (cm). The novel antifungal composition of the present invention has been studied against the Rhizoctonia bataticola for the first time. This combination exhibits an excellent activity against the fungus even at very low concentration. The antifungal composition comprises plural kinds of antimicrobial agents so that resistant fungi may not be produced. RESULTS: Table 1: Antifungal Activity of compound of formula (1) (at different concentrations) (Table Removed) Table 2: Antifungal Activity of Combination of compound of formula 1 and Ferula asafetida.[Nl] (Table Removed) ppm: parts per million (Figure Removed) SAMPLE FIGURE 1 Examples: Example 1. (Table 1, S.No.l) 1.650 mg of compound of formula 1, dissolved in acetone, was mixed with 25 ml of PDA media to attain a concentration of 66ppm and tested against the Rhizoctonia bataticola. It shows tremendous activity with 61.11% of inhibition. Example2. (Table 1, S.No.2&3) When 0.825 mg and 0.413 mg of the same compound, dissolved in acetone, was mixed separately with 25ml of PDA to attain concentrations of 33ppm and 16.5ppm, it surprisingly resists the growth of the fungus upto 48.86 % and 31% respectively. Example3. (Table 2, S.No.5) 1.650 mg of natural product, dissolved in acetone, was mixed with 25ml of PDA to attain a concentration of 66 ppm, shows 33.33 % of inhibition. Example4. (Table 2, S.No.3) However synergistic interaction shows some unexpected results, which can be of great use in agriculture to combat the fungus. The combination of synthetic compound and natural product, consisting 0.825mg each, in acetone, dissolved in 25 ml of PDA (33ppm of both the components or 50% of each component), inhibited the growth upto 60%. Example5. (Table 2, S.No.4) A combination of 0.412mg of synthetic compound and 1.237 mg of natural product, dissolved in acetone, was mixed with 25ml of PDA. This Combination, consisting of 16.5ppm synthetic compound and 49.5ppm natural product, shows 26.1% inhibition. Example6. (Table 2, S.No.2) A combination of 1.237 mg of synthetic compound and 0.412 mg of natural product, dissolved in acetone, was mixed with 25ml of PDA. This Combination, consisting of 49.5 ppm synthetic compound and 16.5ppm natural product, shows 26.1% inhibition.. The results revealed that: (i) Example 4, consisting of 33ppm of each component, is the best synergistic combination prepared by us for the first time as potent antifungal agent, (ii) The above combination consists very low concentration of both the components (formula 1 and N1, 33ppm each), (iii) The combination is effective even at mild concentration of 66ppm. (iv) Thus we can safely propose that composition having synthetic compound of formula (1) only 33ppm in single dose of 66ppm can be used as potent antifungal agent to control the highly pathogenic fungus R.bataticola with minimum or no adverse impact on the environment. Hence it can be used as eco friendly antifungal agent, (v) As the combination consists of very low concentration of natural product (Nl) so it can solve the availability problem of the natural products with improved efficacy. REFERENCE: 1. Dhingra O. D. and Khare M. N. Biological Control of Rhizoctonia bataticola on Urid Bean.2008, Journal of Phytopathology,76(l),23-29. 2. Singh P. and Mehrotra R.S., 1980, Biological control of rhizoctonia bataticola on gram by coating seed with bacillus and strptomyces spp. and their influence on plant growth, 56,475-483. 3. Taneja M. and Grover R. K., 1988, Efficacy of benzimidazole and related fungicides against Rhizoctonia solani and R. bataticola, Annual Review of Phytopathology, 26:203-217. 4. Mane S.S. and Gaikwad S.J.,2008, Biological management of root rot of sesame caused by Rhizoctonia bataticola (T.), Green Farming l(4):36-38. 5. Anilkumar T.B.and Sastry M.N.L., 2008, Development of Tolerance to Fungicides in Rhizoctonia bataticolam, J.Phytopathology, 94(2): 126-131. 6. Sanyal D. and Roy N. K., 1997, Synthesis and structure-activity relationships for the fungicidal activity of 0,0-bisaryl sec-butylphosphonates, Pesticide science, 50:120-170.. 7. Taneja M.and Grover R. K., 2008, Efficacy of benzimidazole and related fungicides against Rhizoctonia solani and R. bataticola, Ann. Appl. Biol, 100(3):425-432. 8. Gupta R. L., Roy, N. K.,1984, Synthesis and fungitoxicity of 0,0-diaryl S-ethyl phosphorothioates, Pestic.Sci., 15(6):553-556 9. B. S. Furniss, A. J. Hannaford, P. W. G. Smith and A. R. Tartchel, Textbook of Practical Organic Chemistry (Vogel's), John Wiley & Sons, Inc.: New York, 1993. 10. J.M. Vincent, Distortion of fungal hyphae in the presence of certain inhibitors. Nature Vol. 159, 1924, pp. 850. We claim: 1. A novel synergistic antifungal composition for exhibiting antifungal activity against Rhizoctonia Bataticola wherein the composition comprises: a) a synthetic compound of formula 1; and b) ferula asafoetida said a synthetic compound of formula 1 and ferula asafoetida being taken in a desired amount and said synthetic compound of formula (1) and Ferula asafoetida having mixed together under ambient conditions to obtain the desired composition. 2. A composition as claimed in claim 1, wherein the said compound of formula (1) is 1,5-bis(2-hydroxyphenyl)pent-l,4-diene-3-one. 3. A composition as claimed in claim 1 wherein the compound of formula (1) is in the amount of 50%. 4. A composition as claimed in claim 1 wherein the Ferula asafoetida is in the amount of 50%. 5. A composition as claimed in claim 1 shows enhanced activity against Rhizoctonia Bataticola up to 60%. 6. A composition as claimed in claim 1, the said compound 1 of formula (1) l,5-bis(2-hydroxyphenyl)pent-l,4-diene-3-one can be used as active ingredients for controlling plant pests in agricultural sector. 7. A composition substantially as herein described with reference to foregoing description, examples and accompanying drawings.

Full Text

NOVEL SYNERGISTIC COMBINATION OF FERULA ASAFOETIDA WITH DEHYDRATED DIALDOL PRODUCT AS POTENT ANTIFUNGAL
AGENT
FIELD OF THE INVENTION:
The present invention relates to a synergistic combination of Ferula asafoetida with dehydrated dialdol product as potent antifungal agent. The present invention relates to the formation of environmentally safe fungicide taking advantage of the bioenhancer characteristics found in natural products and bioefficacy of synthetic compound, which in a way full fill the main objective of inventing an eco friendly fungicide. The presence of natural products in formulation allows the use of synthetic compound at lower concentration with enhanced in vitro activity against the pathogenic fungus.
BACKGROUND:
A variety of crop diseases owe their origin to pathogenic microorganisms, which include bacteria, virus and fungi. Among them fungi are responsible for a major loss of crops all over the world. Rhizoctonia bataticola is considered as one of most noxious fungus [1-4 ] which is constantly seeking attention of scientist throughout the world. Though a number of chemical, broad-spectrum fungicides available for the control of fungus that attack and damage food crops and economically important crops [5-8]. These chemical fungicides aim at fungus eradication and kill the fungus. In this process many non target species are killed and in addition they not only affect the ecosystem but also pollute the atmosphere with chemical, the water we drink, air we breathe and food we eat. Fungi also develop resistance to the fungicide in the course of time on continuous usage. Hence, there is a prime need for

antifungal agent, which is biodegradable, eco friendly, does not affect the beneficial organisms, non-toxic to human beings and at the same time effective against the target fungus. This has diverted the attention of scientists towards product of natural origin. Though natural products are rich source of anti-infective, they carry their own limitations of less potency as compared to synthetic chemicals, low availability, high cost of extraction, structural complexity and instability. In this regard superiority of combination consisting of mixture of bioactive compounds over a single isolated constituent has been well established in which one of the compound generally natural in origin, acts as bioenhancer. This effect is known as synergism which can be defined as a phenomenon in which two different compounds are combined to enhance their individual activity.
The prior art reveals the activity of only herbal compositions containing Ferula asafoetida as one of the ingredients. These herbal compositions are tested for the treatment of human disease only.
US2005089583 discloses a synergistic herbal composition for the treatment of gastric ulcer, said composition essentially comprising of powdered plant parts of Asperagus racemosus, Glycyrrhiza glabra, Seaamum Indicum, Musa sapientum and Trachyaparmum roxburghicinum and optionally, powdered plant parts of Cyclea peltate, Embelia ribes, Coriandrum sativum, Ferula asafoetida, Aloe barbadensis and Evoluulus aisinodes along with one or more pharmaceutically acceptable additives/carriers. The composition comprises powdered plant parts of Asperagus racemosus, Glycyrrhiza glabra, Seaamum Indicum, Musa sapientum and Trachyaparmum roxburghicinum in equal proportions, optionally along with one or more pharmaceutically acceptable additives/carriers. This composition is hereafter referred to as HF3. [0021] In another embodiment of the present invention, the composition

comprises powdered plant parts of Asperagus racemosus, Glycyrrhiza glabra, Seaamwn indicum, Musa sapientum, Trachyaparmum roxburghicinum, Evolvulus aisinodes and Ferula asafoetida in equal proportions, optionally along with one or more pharmaceutically acceptable additives/carriers
US Patent 5,482,711 discloses a method for activating immune competent cells in humans in order to increase the immune function in humans, the immune competent cells being selected from the group consisting of CD 19, HLADR, NKCD3-/CD56+ and CD38, the method comprising administering to humans an effective dose of an extract from Nigella sativa at a concentration which is effective to activate the immune competent cells by reducing the presence of interferon inhibitor factor or lymphokine inhibitor factor.
US Patent 5,653,981 discloses a pharmaceutical unit dosage form for the inhibition of cancer cell growth in humans consisting essentially of a cancer cell growth-inhibiting amount of an extract from Nigella sativa wherein the amount is within a range of from about 0.3 g/kg body weight to about 0.6 g/kg body weight and a pharmaceutically-suitable carrier.
US Patent 5,952,373 discloses a method for the treatment or for the prophylactic treatment of hyperreactive skin predisposed to dermatitis, deficient, hypoactive skin or dermatoses which comprise applying to said skin an effective amount of a composition comprising one or more flavonoids.
US Patent discloses 6,121,243 relates in particular to cosmetic and dermatological formulations comprising flavonoids, their glycosides and, if appropriate, combinations thereof with cinnamic acid derivatives or antioxidants. The damaging effect of the ultraviolet component of solar

radiation on the skin is generally known. While rays having a wavelength
below 290 nm (the so-called UVC range), are absorbed by the ozone layer
in the earth's atmosphere, rays in the range between 290 nm and
320nm, the so-called UVB range, cause erythema, simple sunburn or
even burns of greater or lesser severity.
The narrower range around 308 nm is stated as the maximum for the erythema activity of sunlight.
US Patent 6,180,662 particularly relates to active compounds and
formulations, in particular for topical use, which are used for prophylaxis
and treatment of hyperreactive skin predisposed to dermatitis, and of
deficient hypoactive skin and for prophylaxis and treatment of the
manifest dermatoses mentioned under I. to XIII., such as, for example,
atopic dermatitis, neurodermatitis, atopic eczema and seborrhoeic
dermatitis, photoinduced dermatoses (for example Mallorca acne and in
particular polymorphic photodermatosis and photodermatitis), rosacea,
prurigo forms, pruritus, psoriasis forms, ichthyosis, decubitus, ulcus
cruris and microbial and viral infections, such as, for example, herpes
simplex, h.zoster or h.labialis.
US patent 6,423,747 discloses method of stabilizing or reestablishing the epidermal barrier function of the skin which comprises applying to said skin a formulation comprising a) one or more compounds selected from the group consisting of flavonoids, or b) an active compound combination comprising one or more compounds selected from the group consisting of flavonoids in combination with one or more compounds selected from the group consisting of cinnamic acid derivatives, and c) optionally, one or more compounds selected from the group consisting of antioxidants.
US Patent 6,562,794 discloses a method for the treatment or prophylaxis of skin against aging and the inflammation of skin caused by exposure of

the skin to oxidation or for protecting the skin and the hair from
photoreactions which comprises applying an effective amount of a
formulation comprising, a) alpha.-glucosyl rutin,
and b) one or more compounds selected from the group consisting of antioxidants to said skin or hair.
Accordingly, we have formulated herein a novel composition comprising Ferula asafoetida and Synthetic compound of formula 1 against plant pathogen for the first time. The activity of F.asafoetida never been tested either alone or in combination against phytopathogen ,R.bataticola.
SUMMARY OF THE INVENTION:
The present invention deals with a combination of synthetic compound of formula (1) and Asfoetida (N1) which shows tremendous activity against the test fungus. The present invention is aimed to overcome or avoid problems faced in the control of fungus as mentioned in the literature. The synergistic interaction provided by the composition herein allows reduction of the quantity and improvement of efficacy of synthetic compound by administrating with mildly active natural product (Nl). This reduction allows usage of fungicide which is otherwise too highly toxic or produce unacceptable side effects at fungicidal dosages, allows usage at dosages less than those which produce side effects and should reduce or at least delay the occurrence of natural resistance to important chemical fungicides. The use of products of the present invention offer a low dose treatment with enhanced action comparable to that of standard dose alone.
BRIEF DESCRIPTION OF THE CHEMICAL STRUCTURE

Formula 1: Chemical structure of compound of formula l:l,5-bis(2-
hydroxyphenyl)pent-l,4-diene-3-one.
Figure 1: Graphical representation of percentage inhibition of various
combinations applied against the test fungus
Detailed description of the present invention:
The invention pertains to method for increasing the antifungal activity of newly identified anti Rhizoctonia bataticola agent, by administering a mildly active naturally occurring product in combination with it, thereby reducing the effective amount of the synthetic compound required as compared to the amount required when it is administered alone. The present invention provides a newly identified antifungal agent (formula 1) which shows antifungal activity at very low concentration when tested alone and also its synergistic combination with Asafoetida which enhances its activity further at the same concentration. Compound of formula 1 can be prepared following standard method as reference 9, shown in Scheme 1 wherein requisite aldehyde was dissolved in ethanol and NaOH. To this reaction mixture, acetone was added slowly in an incubator-shaker at room temperature. After half an hour of continuous shaking the reaction mixture resulted to a clear pale yellow solution. Further shaking for about 10 minutes, cloudiness appeared which settled down slowly. The precipitate was then filtered out and re-crystallized by appropriate solvent.

(Scheme Removed)
(Scheme 1)

Formula 1 (Synthetic compound I)
The activity of the samples was determined by standard food poisoning method [as reference 10]. Wherein, Potato dextrose agar of HI MEDIA, MU096 was used as culture medium and test samples were dissolved in acetone. Single spore cultures of fungal species, grown and maintained on potato dextrose agar media at 28±1°C were used for the experiments. The compounds were mixed aseptically with PDA media and were sterilized. The medium with acetone was served as control. The Petri plates having 9 cm inner diameter were taken, and 25 mL of the sterilized PDA containing test samples were plated in the centre. After solidification, 0.5 cm diameter agar discs taken from a 48 h old sporulated fungi culture was placed with the fungal side downward in the centre of each plate with the help of sterile cork borer and glass rod. The plates were incubated in dark at 28±1°C. Radial growths were determined by measuring the colony size along two diameters at right angles after 24 h interval. Three replicates for each treatment were prepared. The experiments were repeated three times. The percentage of inhibition was calculated using the following formula: I = C - T / C x 100, where, I = percentage of inhibition; C = radial growth in control -disc diameter (cm); T = radial growth in treated - disc diameter (cm).
The novel antifungal composition of the present invention has been studied against the Rhizoctonia bataticola for the first time. This combination exhibits an excellent activity against the fungus even at very low concentration. The antifungal composition comprises plural kinds of antimicrobial agents so that resistant fungi may not be produced.

RESULTS:
Table 1: Antifungal Activity of compound of formula (1) (at different concentrations)
(Table Removed)
Table 2: Antifungal Activity of Combination of compound of formula 1
and Ferula asafetida.[Nl]
(Table Removed)
ppm: parts per million

(Figure Removed)
SAMPLE FIGURE 1
Examples:
Example 1. (Table 1, S.No.l) 1.650 mg of compound of formula 1, dissolved in acetone, was mixed with 25 ml of PDA media to attain a concentration of 66ppm and tested against the Rhizoctonia bataticola. It shows tremendous activity with 61.11% of inhibition.
Example2. (Table 1, S.No.2&3) When 0.825 mg and 0.413 mg of
the same compound, dissolved in acetone, was mixed separately with 25ml of PDA to attain concentrations of 33ppm and 16.5ppm, it surprisingly resists the growth of the fungus upto 48.86 % and 31% respectively.
Example3. (Table 2, S.No.5) 1.650 mg of natural product, dissolved in acetone, was mixed with 25ml of PDA to attain a concentration of 66 ppm, shows 33.33 % of inhibition.
Example4. (Table 2, S.No.3) However synergistic interaction shows some unexpected results, which can be of great use in agriculture to combat the fungus. The combination of synthetic

compound and natural product, consisting 0.825mg each, in acetone, dissolved in 25 ml of PDA (33ppm of both the components or 50% of each component), inhibited the growth upto 60%.
Example5. (Table 2, S.No.4) A combination of 0.412mg of synthetic compound and 1.237 mg of natural product, dissolved in acetone, was mixed with 25ml of PDA. This Combination, consisting of 16.5ppm synthetic compound and 49.5ppm natural product, shows 26.1% inhibition.
Example6. (Table 2, S.No.2) A combination of 1.237 mg of synthetic compound and 0.412 mg of natural product, dissolved in acetone, was mixed with 25ml of PDA. This Combination, consisting of 49.5 ppm synthetic compound and 16.5ppm natural product, shows 26.1% inhibition..
The results revealed that:
(i) Example 4, consisting of 33ppm of each component, is the best
synergistic combination prepared by us for the first time as
potent antifungal agent, (ii) The above combination consists very low concentration of both
the components (formula 1 and N1, 33ppm each), (iii) The combination is effective even at mild concentration of
66ppm. (iv) Thus we can safely propose that composition having synthetic
compound of formula (1) only 33ppm in single dose of 66ppm
can be used as potent antifungal agent to control the highly
pathogenic fungus R.bataticola with minimum or no adverse
impact on the environment. Hence it can be used as eco friendly
antifungal agent, (v) As the combination consists of very low concentration of natural
product (Nl) so it can solve the availability problem of the
natural products with improved efficacy.

REFERENCE:
1. Dhingra O. D. and Khare M. N. Biological Control of Rhizoctonia bataticola on Urid Bean.2008, Journal of Phytopathology,76(l),23-29.
2. Singh P. and Mehrotra R.S., 1980, Biological control of rhizoctonia bataticola on gram by coating seed with bacillus and strptomyces spp. and their influence on plant growth, 56,475-483.
3. Taneja M. and Grover R. K., 1988, Efficacy of benzimidazole and related fungicides against Rhizoctonia solani and R. bataticola, Annual Review of Phytopathology, 26:203-217.
4. Mane S.S. and Gaikwad S.J.,2008, Biological management of root rot of sesame caused by Rhizoctonia bataticola (T.), Green Farming l(4):36-38.
5. Anilkumar T.B.and Sastry M.N.L., 2008, Development of Tolerance to Fungicides in Rhizoctonia bataticolam, J.Phytopathology, 94(2): 126-131.
6. Sanyal D. and Roy N. K., 1997, Synthesis and structure-activity relationships for the fungicidal activity of 0,0-bisaryl sec-butylphosphonates, Pesticide science, 50:120-170..
7. Taneja M.and Grover R. K., 2008, Efficacy of benzimidazole and related fungicides against Rhizoctonia solani and R. bataticola, Ann. Appl. Biol, 100(3):425-432.
8. Gupta R. L., Roy, N. K.,1984, Synthesis and fungitoxicity of 0,0-diaryl S-ethyl phosphorothioates, Pestic.Sci., 15(6):553-556
9. B. S. Furniss, A. J. Hannaford, P. W. G. Smith and A. R. Tartchel, Textbook of
Practical Organic Chemistry (Vogel's), John Wiley & Sons, Inc.: New
York, 1993.

10. J.M. Vincent, Distortion of fungal hyphae in the presence of
certain inhibitors. Nature Vol. 159, 1924, pp. 850.

We claim:
1. A novel synergistic antifungal composition for exhibiting antifungal activity against Rhizoctonia Bataticola wherein the composition comprises:
a) a synthetic compound of formula 1; and
b) ferula asafoetida said a synthetic compound of formula 1 and ferula asafoetida being taken in a desired amount and said synthetic compound of formula (1) and Ferula asafoetida having mixed together under ambient conditions to obtain the desired composition.
2. A composition as claimed in claim 1, wherein the said compound of
formula (1) is 1,5-bis(2-hydroxyphenyl)pent-l,4-diene-3-one.
3. A composition as claimed in claim 1 wherein the compound of formula (1) is in the amount of 50%.
4. A composition as claimed in claim 1 wherein the Ferula asafoetida
is in the amount of 50%.
5. A composition as claimed in claim 1 shows enhanced activity against Rhizoctonia Bataticola up to 60%.
6. A composition as claimed in claim 1, the said compound 1 of formula (1) l,5-bis(2-hydroxyphenyl)pent-l,4-diene-3-one can be used as active ingredients for controlling plant pests in agricultural sector.
7. A composition substantially as herein described with reference to foregoing description, examples and accompanying drawings.

Documents:

http://ipindiaonline.gov.in/patentsearch/GrantedSearch/viewdoc.aspx?id=EViI+lAz54owzE1rgbZpzg==&loc=+mN2fYxnTC4l0fUd8W4CAA==

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Patent Number

279380

Indian Patent Application Number

719/DEL/2009

PG Journal Number

03/2017

Publication Date

20-Jan-2017

Grant Date

19-Jan-2017

Date of Filing

07-Apr-2009

Name of Patentee

Archna Rani

Applicant Address

Type 4 flat number 25 Delhi College of Engineering Shahabad Daulatpur Main Bawana Road Delhi- 42

Inventors:

#	Inventor's Name	Inventor's Address
1	Archna Rani	TYPE V FLAT NUMBER 52 DELHI TECHNOLOGICAL UNIVERSITY (FORMERLY DCE), MAIN BAWANA ROAD DELHI 110042, INDIA
2	Sapna Jain	Deptt Applied Chem Delhi College of Engineering Shahabad Daulatpur Main Bawana Road Delhi- 42.

PCT International Classification Number

A61K

PCT International Application Number

N/A

PCT International Filing date

PCT Conventions:

#	PCT Application Number	Date of Convention	Priority Country
1			NA