AU735367B2 - A novel method to protect plants from fungal infection - Google Patents

Description

Our Ref: 725630 P/00/011 Regulation 3:2

AUSTRALIA

Patents Act 1990

ORIGINAL

COMPLETE SPECIFICATION STANDARD PATENT Applicant(s): Agrogene, Ltd.

27 Hartzfeld Street 55600 Kiryat Ono

ISRAEL

DAVIES COLLISON CAVE Patent Trade Mark Attorneys Level 10, 10 Barrack Street SYDNEY NSW 2000 Address for Service: Invention Title: A novel method to protect plants from fungal infection The following statement is a full description of this invention, including the best method of performing it known to me:- 5020 A-NOVEL ETO TOP TECT PLANTS FROM FUNGAL INFECTION This application is a divisional of "parent" No. 700579 (formerly No. 15119/95).

The invention of this divisional application relates to the novel non-fungicidal compounds: -N-benzoyl-3-aminobutyric acid 1-methyl-1-butyl ester, -N-benzoyl-3aniinobutyric acid 1-methyl-5-pentyl ester, N-benzoyl-3-aminobutyric acid n-heptyl ester, -N-benzoyl-3-aminobutyric acid n-hexyl ester, -N-benzoyl-3-aminobutyric acid 1-methyl-1-heptyl ester, -N-benzoyl-3-aminobutyric acid 1-niethyl-l-hexyl ester, -N-benzoyl-3-aminobutyric acid 1-methyl-1-octyl ester, -N-benzoyl- '3aminobutyric acid 1-methyl-1-pentyl ester, -N-benzoyl-3-aminobutyric acid I- :methyl-l-pentyl ester, -N-2-phenylethyl-3-aminobutyric acid n-octyl ester, -N-2- *.a.phenylethyl-3 -aminobutryric acid Il-methyl-lI-pentyl ester and -N-chloromethyl-3 aminobutyric acid. When applied to a crop and its locus the compounds of the :*invention induce local and systemic resistance. of the crop against funrgal. diseases.

The ensuing description is substantially identical to the description of "parent" No.

700579. The parent description has been fully readopted to facilitate identification of the parent/d 'ivisional relationship. It is reiterated that the invention of this *divisional application is as defined above, and in the ensuing claims.

The present invezi-tion concerns a novel method to protect *..plants fromu pathogeniv- at tack. The present invention mo-re partiCUlarly concerns a method of applying selected **non-furnicidAl compounds and composi tions to a crop and its locus to.immunjze, vis-induce local and svst e I C resistance of the crop agaiins! fungal (iseas-ts Wherein such action is referred to in this application as "induced plant defence" (hereinafter IPD. Such compounds lack any anti-fungal activity when applied directiy to the fungus but when applied to a crop they enhance its, own immunization canacity via altering its metabolism.

The present invention also concerns novel non-fungicidal compounds.

BACKGROUND OF THE INVENTION IPD (also known as SAR. Systemic Aquired Resistance) in a crop results from altered metabolism of plant tissue and, is manifested by various defense mechanisms including the accumulation in the crop of soluble proteins referred to as pathogenesis-related (PR) proteins. Some.PR-proteins have been shown to be hydrolytic enzymes such as chitinases and B-1, 3 gluconases, while others are shown to be peroxidases.

Also accumulated are a group of these proteins having a molecular weight or about 10 to 20 kDaltons referred to as P14 proteins, which are now known to be anti-fungal.

All or part of these proteins are believed to participate in the defense system of a crop. Various isonicotinoyl-pyridinyl-hydrazine-derivatives such as dichloroisonicotinic acid (INA), and benzothiadiazole compounds such as Ciba-Geigy CGA 245704) have been described in the patent literature as immunizing healthy plants against fungal diseases (European Patent Publication Numbers 268 775, 0 288 976; and 313 512). The use of threo-DL-0-methylaspartic acid •9 and of DL-B-aminobutyric acid for the control of root rot of peas caused by Aphanomyces euteiches Drechs, has also been described (Papavizas, Plant Disease Reporter, 4.8, 537-541 (1964), Papavizas, Plant Disease Reporter, 51, 125-129. (1967), The use of D-alanine, D-and DL leucine and DL-a-aminobutyric acid at 0.03 M was described to reduce scab in apple caused by Venturia inaequalis (Kuc et al., 49:313-315,1959).

Van AndetI. showed (Ti jdschur Pl antenziek ten. '64 307-3217, 1958 that DL-ser ine D-ser ine (and to a lesser eN tent L-serine phen I ser intic DL-t hreon ine hutt no t DL- a-ami nobtt c ac id nor DL-l a minobtvr i !-cid0 61etiaV 1d as chnothree a 1il he funzl.is ciadosnorium cucumrerinum on c-ucumber (Ibid. page 318).

Oort and Van Andel (1960, Mededel.

Landsbo-rowhager school Dpzoekin,,ssta. Staat Gent 981-992) showed that DL-B-aminobutyric acid applied to leaves of tomato protected those leaves against P h vt op Itt ho T inf es ta ns -(page 987).

**Various derivzatives of DL-B-arninobutvric acid and B-aminocrotonic acid have been descri bed in the patent literature as funsicides agains t Phvt ophthora infes tans in toma to an,:l P Iastronar-a yit icol a in g rapes (Ge rman Patent NO.1,120,802).

Recently, Cohen, Nidernian, Mosinger and Fluhr (Plant Phys iol .1994 104 :58-66 repor ted thatr PR proteins are :involved in IPD in tomatoes.

Sys ternic acqui re res is tance i s very of ten speci f ic to a crop and a di sease For example INA and CGA 24-1704 c-an immunize tobacco against the fungus Peronospora tabacina but not potato nor tomato against P. infestans (Y.Cohen, unpublished). Also known in the li terature is that various isomers of a compound may show di ff erent abilities to induce systemic resis tance. For example, DL-13-aminobutyric acid induces res istance against Fusarium wilt in tomato whereas a-aminoisobutyric acid induces resistance against powdery mildew in whea t (Kalix et al, in Modern Fungicides and Antifungal Compounds, eds. Lyr, Russel and Sissler Intercept 1995).

The literature also teaches that even enantiomers of the same molecule greatly differ in their IPD ability. Thus Cohen showed (Physio Molecular Plant Pathol. 44:273-2-88, 1994) that (R)-B-aminobutvric acid can immunize tobacco against P.tabacina whereas (S-)-1-aminobutyric cannot.

Oort and Van Andel (Ibid) concluded: We do not understand why one of two related amino acids has an evident effect and the Other has not, and wh' one influences a give-n plant pathogen combination but has noor hardly any influence on another" (Ibid page 987).

-3 The method of the present invention is not obvious in view of the prior art cited, for the following reasons: 1. Oort and Van Andel (Mededel Landbourhoogeschool Opzockkinssta. Staat Gent 25:981-992, 1960) showed that BABA (DL-B-aminobutyric acid) applied exclusively to the leaves of tomato 2 days before *inoculation reduced infection with P. infestans (Ibid page 987 line The present invention shows that BABA protects plants against disease when applied to either the leaves or to the roots, or even injected to the stem, and may be applied either before or after inoculation. Also data of OOrt and Van Andel are merly qualitative as no BABA concentration nor percent protection are given.

2. USA 3,899,585 (Misato et .al) 12 August 1975 teaches in Table 1 that test compound NO.13 (which is the closest to the present invention) 2-aminobutvric acid lauryl ester hydrochloride applied to rice leaves reduces rice blast disease. The present invention teaches that 2-aminobutyric acid was totally ineffective in reducing diseases in various crops (see Cohen Phytopathology 83: 55-59,1994, Cohen, Physiol.

Molecul. Plant Pathol. 44: 273-288-1994). Also it teaches that test compound NO 13 should be applied preventively rather than curatively, unlike the present invention which allows application of the test compound curatively.

Interestingly enough test compound No 13 is probably inactive in cucumber against downy mildew as it is missing from Example 4 (Table 4) unlike the present invention showing that BABA is active against this disease.

3. USA 3,991,208 (Dudzinzki et al) 09 November 1976 teaches that a tertiary amine group attached to a 2-carbon of a long chain alkyl, and carboxyethyl group substituent of the nitrogen atom are surface active compounds that are antibacterial (against Gram positive bacteria). Such compounds are not amino acids, have no effect on fungi and do not control fungal plant diseases either directly or indirectly and therefore have no relevance to the present invention.

4 4. USA 481,219 (Watkinson) 06 November 1984 teaches that timber decay due to fungal attack by mainly Basidiomycetes may be prevented by composition containing a nitrogenous compcund and a sacharide compound. The nitrogenous compound selected from a group consisting DL-methionine sulfcxide, lysine HC1, and aminoisobutyric acid. The latter compound is (CH3)2 C(NH2) COOH (Column 2 line 28) which is 2-amino-iso butyric acid. According to the present invention this compound is not suitable for inducing systemic resistance against fungal plant pathogens, but BABA does, not to say that the present invention deals with protection of alive green plants and not of a dead woody tissue like timber. Moreover, according to Watkinson the composition must contain a sugar as an energy source for the fungus, whereas the Spresent invention does not.

USA 5,096,700 (Seibel et al) 17 March 1992 teaches that halogenated amino acid derivatives are useful antibacterial agents in humans. This prior art is indeed not relevant to the present invention because first it deals with bacterial human diseases and not in fungal diseases of crop plants; second it involves halogenated (at least one halogen atom bound to the carbon backbone at position 2) amino acids, which is not required for BABA to induce IPD; 6. GB, 1,048,507, (Harinack et al) 16 November 1965 teaches that glycine derivatives are effective systemic fungicides in crop plants although they are not effective against fungus spore germination in in-vitro tests (page 1 line This prior art depart from the present invention due to the fact that glycine is a 2-amino acid (a-amino acid) in which the NH2 group is bonded to carbon 2, while the present invention deals with 3-amino acids (8-amino acids) in which the NH2 group is bound to carbon 3. According to the present invention only 3-aminobutyric acids but not 2-aminobutyric acids, have systemic IPD effect.

-il lir~ 5 The present invention deals with compounds that protect crop plants against fungal attack via immunization, namely altering plant metabolism so as it can resist fungal colonization in its tissues. It was shown by Cohen et al (Plant Physiology 104: 59-66 1994) that BABA enhances the accumulation of PR-proteins in tomato. This accumalation was correlated with resistance to P.infestans. However, this is probably not the case in other crops such as curcurbits and tobacco in which BABA also induces IPD 9 response. In curcubits BABA induces the accumulation of callose and lignin in the infected sites which probably stop the fungus whereas in tobacco the IPD mechanism remains obscure (Cohen, Physiol. Molecule Plant. Pathol 44: 273-288, 1994) S OBJECTIVES OF THE INVENTION It is the objective of the present invention to provide a novel method to induce IPD. It is a further objective induce IPD in selected crops. It is a further objective of the present invention to provide novel compounds which maybe used to induce IPD.

SUMMARY OF THE INVENTION We have found a novel method for protecting a crop as hereinafter-defined in need of protection against fungal diseases caused by fungi by inducing local and systemic resistance in said crop in need of such resistance comprising applying to the crop or its locus a composition containing an effective amount of a compound of formula R R 7 ^0

N

R

R

K

5 RI

R

2 Rl and R 2 are independently hydrogen. Ci-a alkyl, phenyl, and.phenyl CI-4 alkyl

R

3 is hydrogen; Ci-23 alkyl; carboxy C 1 -4 alkyl; phenyl

C

1 -4 alkyl; wherein the phenyl moiety is unsubstituted or monosubstituted by halogen; or C2- 2 3 alkoxy-carbonyl Ci-4alkyl;

R

4 and Rs are independently hydrogen or Ci-9 alkyl; p:jwPOOCSNEH'%S96B37.SPE 13M1 1M9 -6- R6 and R7 are independently hydrogen; Cl-8 alkyl; cyclohexyl; hydroxy-ethyl C2-8 alkanoyl; phenyl C1-4 alkyl, benzoyl wherein the phenyl moiety is unsubstituted or monosubstituted by halogen; C2-8 alkoxy-carbonyl; CONHR8 wherein R8 is hydrogen, CI-8 alkyl, phenyl, phenyl C1-4 alkyl; phenyl C2-4 alkyloxycarbonyl; X is0, Sor NH and salts thereof, and the crop is selected from the group consisting of corn, cucumber, melon, broccoli, cauliflower, kohirabi, potatoes, cabbage, sunflower, tobacco, grapes, cotton, maize, sorghum, pearl millet, rice, lettuce, hop, avocado, citrus, soybean, onion and tomato, the composition being present *9 in an amount sufficient to induce local systemic resistance in the crop to control the fungal 10 disease with the proviso that the crop is not tomato when the fungal disease is Fusariumn Qxysponlm f. sp. lycopersici.

In accordance with a further aspect of the invention, there is provided the novel non- .9.-fungicidal compounds: -N-benzoyl-3-aminbUtyric acid 1-methyl-1-butyl ester, -Nbenzoyl-3-aminobutyric acid 1-mnethyl-5-pentyl ester, N-benzoyl-3-aminobutyric acid n-heptyl ester, -N-benzoyl-3-aminobutyric acid n-hexyl ester, -N-benzoyl-3aminobutyric acid 1lmethyl-1-heptyl, ester, -N-benzoyl-3-an-dnobutyric acid 1-methyl-' 1-hexyl ester, -N-benzoyl-3-aminobutyric acid 1.-methyl-1-octyl ester, -N-benzoyl-3- 20 Iaminobutyric acid 1 -methyl-1-pentyl ester, N-benzoyl-3-aniinobutyric acid 1-methyl- 1-pentyl ester, -N-2-phenylethyl-3-aininobutyric acid n-octyl ester, -N-2-phenylethyl- 3-ammnobutyric acid 1-methyl-i -pentyl ester,, -N.-choromethyl-3-aminobutyric acid and -3-aminobutyric acid sodium salt.

DETAILED DESCRIPTION OF THE INVENTION Alkyl as used herein refers to straight chains, branched and cyclic forms and preferably contain one to four carbon atoms.

RI and R2 are preferably independently hydrogen, methyl or phenyl, more preferably R1 is hydrogen or methyl and R2 is hydrogen.

-6a- R3 is preferably hydrogen.

R4 and R5 are preferably indpendently hydrogen or C1-3 alkyl, more preferably R4 is hydrogen or methyl and R5 is hydrogen or C1-3 alkyl, more preferably R4 is hydrogen or methyl and is hydrogen.

R6 and R7 are preferably independently hydrogen, C1-6 alkyl, benzyl optionally substituted by halogen, more preferably R6 is hydrogen or methyl and R7 is hydrogen.

X is preferably oxygen.

Preferred compounds of the invention are the P-amino-butyric acids and gthe P-aminovaleric acids; and most preferred is R-p-aminobutyric acid. The structures of various aminobutyric acids are shown in Scheme 1.

o* *o 7 Scheme 1. Am-inobutyric acids ~s~~COOH H{ NH 2 -00"Y COOH

NH

2 (AABA) L-2-Amino-l-butyric acid DL-2-Amino-n-butyric acid H H H 2 (S)-3-Amino-fl-butyriC acid *HA CO 4-Amino-n-butyric acid

(GABA)

H H2 0.

COOH

(R)-3-Amino-n-butyric acid

CH

3

COCH

CH

KNH

2 2-Amino-iso-butyric acid

COOH

(BABA)

DL-3-Amino-n-butyric acid

~COOH

CH

3 DL-3-Amino-iso-butyric acid 8 Salt forms of the compound of formula contemplated in this application include acid addition salts such as those obtained by the addition of HC1,CF3 C02H, toluene sulfonic acid, methane sulfonic acid and (CO 2

H)

2 Y NR6R7H I XR 3 R R1 R2 wherein Y is the residue of the acid; alkali metal salts such as those obtained by treatment with NaOH, KOH or LiOH NR R7H <4 X M

.R

1 2 5 wherein M is an alkali metal such as Na, K or Li; and acid addition/amine salts such as those obtained by treatment with HCI and an amine such as diethylamine, propyl amine, benzylamine.

0 0 Cl NR 6 R H R

XNH

2 Rab

R

5 R R 2 5 1 2 wherein Ra and Rb are substituents.

Preferred crops in which the method of the present invention is applicable are cucumbers, melon, broccoli, cauliflower, kohlrabi, potatoes, sunflower, tobacco, grapes, cotton, maize, sorghum, cabbage, pearl millet, rice, onion and hop. Most preferred are sunflower, grapes, cucumber, melon, broccoli, kohlrabi, cauliflower, potatoes, tobacco and maize.

9 Production methods The novel compounds encompassed by the present application are structurally related to known compounds and can be easily prepared by either derivatising the known compounds or by modifying the procedures for preparing the known compound, as required. These procedures will be apparent to those skilled in the art The following procedures are illustrative.

Compounds of the formula (la) (Ia) R R 6 7 0

R

4

OR

3

R

1 wherein R 1 and R 4 7 are as previously defined and R3 represents hydrogen or Ci-a alkyl can be obtained from a compound of the formula (IIa).

OR R3 (IIa)

OR

R R 1 5 1 To prepare compounds of formula (la) where R 6 is H and

R

7 is as previously defined, the compound of formula (IIa) is reacted with NR 7

H

2 wherein R 7 is as previously defined. Reactions of this type are described in the literature, by A. Zilkha and J. Rivlin, J.Org.

Chem. 1957, 23, 94.

To prepare compounds of formula (la) where Re and R7 are as previously defined but excluding hydrogen, the compound of formula (IIa) is reacted with NR 6

R

7 Li, wherein R 6 and R 7 are as previously defined but excluding hydrogen. Reactions of this type are described in the literature, by Davies et al., Tetrahedron: Asymmetry, Vol.2, No.3, pp. 183-186 (1991).

Compounds of the formula (IIa) are either known or obtainable from known compounds according to standard procedures.

10 As can be appreciated, in such cases where R 4 and Rs do not represent-the same substituent, the carbon atom to which they attach is chiral. Procedures for preparing each enantiomer form are either specifically described in the literature, in EP 0 144 980 or in Davies, supra, or can be prepared according to analogous procedures.

The present method was found to be effective against a variety of diseases. Examples are late blight, downy mildew, blue mold, leaf spots, fusarium wilt, trunk rot, fruit brown rot, damping off, white rust, black shunk and Phytophthoras root rots.

The compounds of this invention will typically be applied to crops or their locus before or after the onset or after the initial signs of fungal attack and may be applied to the foliar surfaces of the crop. The :eee amount of the active ingredient to be employed will be sufficient to induce the systemic resistance of the crop to control the fungi and will vary depending on such factors as the species of fungi to be controlled, the type of treatment (for example, spraying dusting, seed treatment, soil drench), the condition of the crop, and the particular active ingredient used.

As an application to the crop or its locus, the compounds will be applied to the crops with a dosage rate of from 0.1 to 5 kg/ha, preferably from 0.2 to 2kg/ha. with application being repeated as necessary, typically at intervals of every one to three weeks.

*e Depending on circumstances, the compounds of this invention may be used in association with other pesticides, fungicides, insecticides, acaricides, herbicides, or plant growth regulating agents in order to enhance their activity or to widen their spectrum of activity.

11 The compounds of this invention are conveniently employed as fungicidal compositions in association with agriculturally acceptable carriers or diluents. Such compositions also form part of the present invention.

They may contain, aside from a compound of formula (I) as active agent, other active agents, such as fungicides.

They may be employed in either solid or liquid application forms in the form of a wettable powder, an emulsion concentrate, a water dispersible suspension concentrate ("flowable"), a dusting powder, a granulate, a delayed release form incorporating conventional carriers, diluents and/or adjuvants. Such compositions may be produced in conventional manner, e.g. by mixing the active ingredient with a carrier and other formulating ingredients.

Particular formulations to be applied in spraying forms such as water dispersible concentrates or wettable powders may contain surfactant such as wetting and dispersing agents, the condensation product of formaldehyde with naphthalene sulphonate, an alkyl-aryl-sulphonate, a lignin sulphonate, a fatty alkyl *,.*sulphate an ethoxylated alkylphenol and an ethoxylated fatty alcohol.

In general, the formulations include from 0.01 to 90% by weight of active agent, said active agent consisting either of at least one .compound of formula or mixture thereof with other active agents, such as fungicides. Concentrate forms of compositions generally contain between about 2 and 80%, preferably between about 5 and 70% by weight of active agent. Application forms of formulation may, for example, contain from 0.01% to 20% by weight, preferably from 0.01% to 5% by weight, of active agent.

-12- Formulation Example I: Wettable powder parts by weight of a compound of formula are ground with 2 parts of lauryl sulphate, 3 parts sodium lignin the sulphonate and 45 parts of finely divided kaolininite until the mean particle size is below microns. The resulting wettable powder so obtained is diluted with water before use to a concentration of between 0.01% to 5% active ingredient. The resulting spray liquor may be applied by foliar spray as well as by root drench application.

Formulation Example II: emulsion concentrate parts by weight of a compound of formula I, 65 parts of xylene, 10 parts of the mixed reaction product of an alkylphenol with xyleneoxide and calcium-dodecyl-benzene sulphonate are thoroughly mixed until a homogeneous solution is obtained. The resulting emulsion concentrate is diluted with water before use.

Formulation Example III: Granulate (for soil treatments) Onto 94.5 parts by weight of quartz sand in a tumbler mixer is sprayed 0.5 parts by weight of a binder (non-ionic tenside) and is thoroughly mixed. 5 parts by weight of compound of the formula in powdered form S" are then added and thoroughly mixed to obtain a granulate formulation with a particle size in the range of from about 0.3 to about 0.7 mm. The granulate may be applied by incorporation into the soil adjacent the plants to be tested.

13 Formulation Example IV: Seed or Tuber Dressing parts by weight of compound of the formula parts of dialkylphenoxy-poly-(ethylenoxy) ethanol, parts of fine silica,, 44 parts of fine kaolin, parts of a colorant crystal violet) and 0.5 parts of xantham gum are mixed and ground in a contraplex mill at approximately 10,000 rpm to an average particle size of below 20 microns.

The resulting formulation is applied to the seeds or tubers as an aqueous suspension in an apparatus suitable for that purpose. Where the compound of the formula

(I)

is liquid, it is first absorbed on the carriers, if desired with the air of a small amount of a volatile solvent such as acetone. The resulting powder is first allowed to dry if a solvent is used, then the other ingredients are added and the rest of the procedure is carried out.

Formulation Example V: Soil Drench Drip Irrigation 2 parts by weight of compound of the formula are dissolved in 1,000 parts of water. The resulting formulation is applied to plants by drip irrigation As previously mentioned, the compounds of formula

(I)

are effective in activating or enhancing a crop s defense system against fungal diseases caused by fungi.

SSuch activity can be demonstrated in using the general procedures of the following tests: Test A: IPD in potato plants against Phytophthora Sinfestans Potato plants (cultivar Bintie) are grown from tubers in pots filled with sandy loam, peat and erlite mixed in equal volumes, in the greenhouse -14 (20-220C). The plants are ready for testing when they have 6 or 7 compound leaves.

Tests are carried out with the metalaxyl-resistant isolate MR1 of Phvtophthora infestans. as well as the MS2, MS3, MR2 and MR3 isolates (Kadish and Cohen, Phytopathology, 78: 912-9155 1988). The fungus is grown on potato tuber slices at 150C in the dark. Fresh sporangia are harvested at 6 days after inoculating the slices into double distilled water (40C) and their concentration adjusted to 10,000 sporangia/ml before used for challenge inoculations.

The compounds of this application are dissolved in water and sprayed on either the lower or upper leaf surfaces of the potato plants with a fine atomizer (about 10 ml per plant). The plants are left on the bench until the droplets dry and then are placed in a growth cabinet calibrated to 200C and 14 hours of light per 10 day.

Challenge inoculation with P.infes tans is carried out at time intervals ranging from 30 minutes to 12 days after treatment with the compounds, by spraying a sporangial suspension on the upper leaf surfaces (about 15 ml per :*plant). In one experiment, inoculum droplets (10 containing about 100 sporangia) are placed on the leaf surfaces, two droplets per leaflet, one on each side of the main vein. In another experiment the compounds are applied curatively, 24 hours after inoculation with P.infestans.

Inoculated plants are kept at 100% RH in the dark for 24 hours at 180C and then returned to a growth cabinet maintained at 200 C with 12 hours light per day. Disease severity is monitored 4-8 days after inoculation by visually estimating the proportion or a leaf area covered by blight lesions.

r oeoo Test B: IPD in tobacco plants against Peronospora tabacina Tobacco plants (cultivars Ky-14 or Ky-16) are grown from seed in pots in the greenhouse. When reaching the 10-leaf stage or older the compounds of this application are injected into the stem of the plant. At 1-3 days before injection or at 1-10 days after injection plants are challenge-inoculated with conidia of the fungus Peronospora tabacina Adam which causes the blue mold disease. Conidia are harvested from previously infected tobacco plants. Inoculation is done with 10,000-100,000 conidia/ml, with approximately 50ml per plant. The procedure described above for inoculation, maintaining and scoring the disease are also applicable here.

While the invention will now be described in connection with certain preferred embodiments in the following examples, it will be understood that it is not intended to limit the invention to these particular embodiments.

On the contrary, it is intended to cover all alter- S natives, modifications and equivalents as may be included within the scope of the invention as defined by the appended claims. Thus, the following examples, which include preferred embodiments, will serve to illustrate the practice of this invention, it being understood that S the particulars shown are by way of example and for Spurposes of illustrative discussion of preferred embodiments of the present invention only and are presented in the cause of providing what is believed to be the most useful and readily understood description of procedures, as well as of the principles and conceptual aspects of the invention.

16

EXAMPLES

EXAMPLE 1: N- (2-hydroxyethyl) -6 -aminobutyric acid A solution of 86g of crotonic acid (1 mole) and ethanolamine (1 mole) in pyridine (200ml) is refluxed for 2-3 hours and subsequently cooled. The resulting product is filtered and recrystallized to yield the title compound having m.p. 178-1800 C (compoundl.1, Table 1).

Following analogous procedure, the compounds 1.2-1.7, 1.10, 1.11, and 1.13-1.15 set forth in Table 1 are obtained.

EXAMPLE 2: 3-aminohexanoic acid A mixture of 2 hexenoic acid (7.0g, 0.06 mol) and concentrated aqueous ammonium hydroxide (70ml) is heated go* for 24 hours in an autoclave at 1500C. The cooled mixture is treated with carbon black and filtered. After evaporation of the solvent the crude product is e recrystallized from ethanol to give the title compound m.p. 2030C (compound 1.21, Table 1).

Following an analogous procedure, the compounds 1.8, 1.9, 1.12 and 1.18-1.20 of Table 1 are obtained.

EXAMPLE 3: N-benzoyl-3-aminobutyric acid To a cooled solution of 3 aminobutyric acid (13g) in 2M NaOH (130ml) is added benzoyl chloride (19.7g) over the course of two hours. The mixture is allowed to warm to room temperature. Washing with diethyl ether, acidifying of the aqueous phase with 20% HCI, extraction with diethyl ether, drying over MgSO4, evaporation of the solvent and recrystallization in ether/hexane gives the title compound m.p. 150-1520C (compound 1.16, Table 1).

i7- EXAMPLE DL.-N-benzol-3-amin-obtr ic acid n-oct-l-ester To 3.0 r DL-N-benzoyl-3-anfobutyric acid in 30 ml Dichioro methane, was added 3.0 g PCl5 in smnall portions, keeping, the mixture temperature at 50C. The mixtiure was allowed to warm to 22-200C and stirred during 5 hr. Petrol ether 60-80 (150 ml) was added, the chloride was filtered and dried, yielding 2.3 g p roduct To the chloride in 20rnl Dichioroethane was added 4.1 g. n-octanol in 10 ml Dichioroethane, during 10 minutes at 200C. The mixture 0.00 was heated to 650C during 6 hr. After distillation, 20 ml oo~oDichioroethane was added and washed with MX1 ml sodium oo::oobicarbonate dried over M2504. Evaporation of the solvent gave 2.0 gof the titled compound having a molecular weight of 319.

0 0 *000 18 Fol lowing an analogous procedure, compouind 1. 17 of Table 1 is obtained.

TABLE I.

COMPOUNDS PREPARED OF THE FO-RMUjLA: R 4 HR 6

O

R R Cpd R 1

R

4

R

5 R6 mlp.(UC) 1.1 H CT-b H hydroxvethyl 178-180 1.2 H CH 3 H isopropyl 167-169 1.3 H C-H 3 H benzyl 178-180 1.4 H CH 3 H cyclohexyl 161-163 H C1H 3 H n-hexyl 151-153 1.6 H Gil 3 H p-chlorobenzyl 152-154 00 1.7 H CH 3 H benzyl 180-182 1.8 H ethyl H H 128-130 1.9 H CH 3 Gil 3 H 216 1.10 CH 3 H H benzyl 148 o. 1.11 H CH 3 H phenyl-ethyl 164 1.20 H phenyl H H 220-221 1.13 H CH- 3 H n-octyl 150 1.14 H Gil 3 H n-decyl 148 1.15 H ethyl H benzyl 157-160 1.16 H Gil 3 H benzoyl 150-152 1.17 H Gil 3 H benzyloxycarbonyll28-130 1.18 H ethyl H H 178-180 1.19 H Gil 3 H H 209-210* 0 1.20 H CH 3 H Gil 3 86-87** 1.21 H propyl H H 203 enantiomer Amonohydr ate 19- EXAMPLE 4: N-benzvloxvcarbonvl 3 aminobutvric acid (4-chlorophenvl)-l- ethylamide Z-protected B aminobutyric acid (0.02ml), (4-chlorophenyl)-l- ethylamine and 1.1 equivalents of DCC (dicyclohexyl carbonimide) are stirred in ethyl acetate at room temperature for 16 hours. The precipitate is filtered, the filtrate evaporated and chromatographed on silica gel (hexane/ethyl acetate 1:1) to give the title compound as a mixture of diastereomers m.p. 168-178 0

C.

EXAMPLE 5: B-aminobutyric acid hydrochloride 5.15 g B-aminobutyric acid (50 mol) are dissolved in 650ml methanol. After addition of 5.5ml concentrated

HCI

the solution is evaporated. The residue is triturated in *diethyl ether, decanted and dried. A colorless oil is isolated. Microanalysis: C,34.4; H, 7.2; N, 10.0; CI, 25.4.

EXAMPLE 6: B-aminobutvric acid sodium salt 2.06g B-aminobutyric acid (20mol) are dissolved in 100ml of a mixture of water:methanol One equivalent NaOH in 10 ml water is added. The solution is evaporated and the resulting amorphous solid is dried.

Microanalysis: C, 37.4; H, 6.7; N, 10.9.

EXAMPLE 7: 8-aminobutvric acid diethvlammonium chloride 1.4 g 8-aminobutyric acid (10 mol) are dissolved in 100 ml of methanol. Diethylamine (0.9 g, 12.3 mol) is added and the residue is evaporated. The oily residue is washed with ether, decanted and dried to afford an amorphous material. H-NMR (CD 3 OD,200MHz) 1.29 9H, 3 CH 3 2.25-2.45 2H, CH 2 3.14 4H. CH 2

CH

3 3.34-3.58 1H,CH) EXAMPLE 8: Protection of tomato plants against fusarium wilt Tomato plants were grown in sterile soil in the greenhouse. When they reached the 4-leaf stage they were treated with compound of the formula solution by a soil drench. Four days later the plants were uprooted, washed with water and their root system immersed for two minutes in conidial suspension (107 conidialml) of the fungus Fusarium oxvsporum f.sp. lvcopersici. Plants were then transplanted (without washing) in pots filled with sterile soil.Twelve days later all challenged control plants wilted of the disease whereas none of the challenged treated plants wilted. Growth of the latter plants was similar to that of control, unchallengeduninoculated plants. Results are shown in Table 2.

EXAMPLE 9: Following the method of Example 8, a similar experiment S" was run using lower concentration of BABA. The results are shown in Table 3.

21 TABLE 2 Protection of tomato plants (cv. Rehovot 13) against fusarium wilt caused by Fusarium oxysporum f.sp.

Ivcopersici by aminobutyric acids (soil drench) Compound None

AABA

BABA

GABA

Percent of plants Healthy Wilted 0 34 100 7 100 66 0 93 Plants were soil-drenched with 2000 ppm of the compound and inoculated 4 days later; rating was taken 12 days after inoculation.

TABLE 3 Protection of tomato plants (cv. Rehovot 13) against fusarium wilt caused by Fusarium oxysporum f.sp.

Iycopersici by lower concentrations of BABA (soil drench) Concentration 0 (ppm) Percent Wilted Plants 0 100 250 53 500 43 1000 0 FXAMPLE Folltowing .rh, ahove d.,?scribed mtH)ods, the -f fect of ami nobut'r atccis,. on downy mi 1 dew in sufflower was s tudied. The resuilt F appear i n TablIe which shows the mar1-keod a c t ivity of FABA in percent protec t ioal.

EXAMPLE I I: Following the above described methods, the effect of 3minobutyric acids on PlasmoDara viticola in grape plants were studied. The results which appear in Table shonw the good protection gi ven by 1..ABA.

EXAMPLE 12: Followintz the above described mthlods. the effect of .~.aiinobutyric acids on downev mi ldew in cucumber and mielon plants were studied. The results, which appear in Table 6, show the good protection given by BABA.

EXAMPLES 13 Following the above described methods, the effect of R-BABA and S-BABA against Peronosipora parasitica and Alternaria brassicicola in broccoli, kohlrabi, and cauliflower were studied; and the results, which appear in Tables 7 9, respectively, show the good protection given by R-BABA.

23 TABLE 4: The effect of aminobutyric acids on systemic downy mildew of sunflowers (cv. caused by Plasmopara halstedii Compound

AABA

BABA

GABA

Method of Application Conc.

mg /1 Percent Protection 100 0 spray spray spray 2000 2000 2000 mg /Plant

AABA

BABA

GABA

AABA

BABA

GABA

soil drench soil drench soil drench root uptake root uptake root uptake 0 100 0 0 100 0 In all experiments inoculation with the fungus was done either 1 day before treatment (curative) or 2 days after treatment.

a 24 TABLE The effect of aminobutyric acids on downy mildew caused by Plasmopara viticola in grape plants (cv. Sauvignon blanc or Cabernet sauvignon) Conc.

ppm per plant 100 200 500 1000 Application mode soil drench spraying) intact plants in pots %protection with AABA BABA GABA 10 0 90 0 50 floating 5 95 0 100 leaf discs 10 100 Disease rating was taken 9 days after inoculation. BABA has also curative effect as follows: When applied to inoculated leaf discs at 0, 1, 2 and 3 days after inoculation percent protection was 100, 86, 50 and percent, respectively.

TABLE 6: The e ff ec t of aminobu tyr ic ac ids on downy mil1dew caused by Pseudoper-onospora cubensi s in cucumber and melon -plants (foliar spray-) Or leaf discs floating) Plant cul tivar cucumber (Dlii a) Conc.

P-PM

250 500 1000 2000 250 500 1000 2000 Percent protection by whole plants AABA BABA GABA mel1on (Gal ia) 4 4* 4 *4 4.

4 4 .4 .4 leaf discs cucumber (Dlii a) 94 97 100 *100 melon 6 0 19 0 (Ananas) 12 0 25 0 0 70 0 37 85 Disease rating was taken 7 days after inoculation.

4444 4444 4 -26- TABLE 7: (Broccoli) Protection of (cv. Shugon) against Peronospora parasitica and Alternaria brassicicola with amino butyric acids applied as a' foliar spray.

Compound

R-BABA.

Conc. ppm 125 250 500 1000 2000 percent protection P. parasi t ica A. bras si c icol1a 33 50 95 100 100 0.0.

S-B ABA 500 1000 2000 not tested) S. S

SS

S*SS

S

S

-2'f'7- TABLE 8: (Kohlrabi) Protect ion of (cv. Whi-te Wien) against Peronospora parasitica and Alternaria brassicicola with amino butyric acids applied as a foliar spray.

Compound Conc .ppm percent protection P. paras iti ca A. bras sici col a

R-BABA

0 *OOb *0 0 *0*e 0* 00 0 0

S

0 0* *0 S 45 0 555.

SO

bO 0 125 250 500 1000 2000 500 1000 2000 100 100 0 not tested) 5-BABA

S.

055S

S

*000

S

0000 2.8 TABLE 9: Protect ion of Caul if lower (cv. Nuri t) against Peronospora parasitica and Al ternaria brassicicola with amino butyric acids applied as a foliar spray.

Compound Conc ppm percent protection P. par a sit ica A. bras sic icolIa

R-BABA

S-BABA

125 250 500 1000 2000 500 1000 2000 33 50 95 100 100 0 0 0 0 not tested) 29 EXAMPLE 16: Following the above described methods, the effect of a formulated DL BABA in potatoes was studied. The results are shown in Table EXAMPLE 17: Following the above described methods, the markedly good effect of DL BABA against late blight in potatoes (Bintie) in growth chambers was studied. The results are shown in Table 11 EXAMPLE 18: Following the method of Example 16 but running field trials in both Alpha and Bintje cultivars, the results are shown in Table 12.

EXAMPLES 19-20: Resistance to Peronospora tabacina induced in tobacco plants was studied as outlined by Y. Cohen (Physiological and Molecular Plant Pathology (1994) 44:273-88) where the active ingredients were applied as a stem injection or foliar spray. Results are shown in Table 13.

EXAMPLE 21: Following the method of Examples 19-20, the effect of a soil drench with DL- BABA (3mg per plant) on blue mold development in tobacco cv. Ky 16 showed an 80 percent control of the disease some 20 days after challenge inoculations.

30 TABLE EFFECT OF 25% WP FORMULATED DL-BABA.IN

POTATOES

ppm DL-BABA Percent Protection against Phvtophthora infect ions Cofltro I 31 62 125 250 500 I ,000 2,000 31.- TABLE 11 Effect of aminohutyric acids against late blight in potato crops in growth chamber Compound Days after inocula tion Percent blighted leaf area Cont rol1

GABA

DL-AABA

DL- BABA 32 TABLE 12: Percentage control of late blight epidemics induced by Phytophthora infestans (isolate MR1) in potato crops treated with BABA (25 WP) in three independent field experiments.

Experiment and cultivar Autumn Alpha Winter Bintje Spring Alpha dose Kg a.i.

per ha Interval 7 between sprays, days 10 14 0 0.2 0.4 0.8 0 24.6 52.6 55.1 r 0 0.2 0.4 0.8 0 0.57 1.15 2.30 0 0.57 1.15 2.30 0 55.0 57.4 62.6 0 38.0 77.7 75.0 0 35.2 64.5 76.0 0 42.1 52.5 62.6 0 38.4 47.1 58.0 Spring Bintje 33 TABLE 13: Resistanice to. Peronospora t abacina in tobac,-co plants .amincbutyric acids.

Compound Folia Sprav Stem Iniection c (Oc Protect ion) Disease Severity (mean SD) Water 0 2. 0 Oa DL- XXBA 10 t. 3 0 .21b DL-BkBXk 78 0. 7 0. 2c -BABA 99 0. 07 0.09d GABA Oa SAa 80 1.5 0b INAb 61 0. 5 Oc a Sodium salicylate b 2,6 dichioro-iso-licotifliC acid c The letters refer to statistics.

34 EXAMPLE 22: Sunflower plants were protected against downy mildew caused by Plasmopora halstedii by treating the seeds with BABA. Thus, the seeds were soaked for 24 hours in solution containing 10mg BABA per ml, and then sown in pots in the greenhouse.Two weeks later the developed plants were inoculated with Plasmopora halstedii The progress of the disease was assessed after seven more days with the following results: While the control had 100 percent of the plants remained infected; the treated plants had only a 2 percent rate of infection.

EXAMPLE 23: Maize (Line 3376) seeds were allowed to sprout in water for 5 days. They were then dipped in a BABA solution for S" one day. The sprouted seeds were washed and placed in contact with Fusarium moniliforme for one day and then planted in pots. After two weeks the progress of the disease was as follows: BABA (ppm) 0 125 250 500 2,000 Percent plants 70 80 40 0 0 infected.

AiU l ii.

35 EXAMPLE:24 The activity of N-benzoyl-3-aminobutyric acid n-octyl ester against late blight (Phytophthora infestan on potato in growth chamber was studied. Six week old plants in pots were sprayed with the compounds and inoculated two days later. Disease records taken 7 days post-inoculation are listed in Table 14.

TABLE 14 Activity of N-benzoyl-3-aminobutyric acid n-octyl ester against late blight (Phvtophthora infestans) on potato in growth chambers.

Concentration ppm protection 500 77 1000 89 2000 96 a a 36 EXAMPLE The effect of BABA against grey mold in tomatoes and cucumbers was studied. Young tomatoes (cultivar Baby) plants (4-leaf stage) and young cucumbers (cultivar Dlila) plants (1-leaf stage) were sprayed with BABA and inociilated two days later with spores suspension of Botrvtis cinera.

The plants were kept under a moist Perspex cover in a Irowth chamber for four days and then moni tored for infection. The results are listed in Table go 9 TABLE The effect of BABA against grey mould, incited by Botrvtis cinerea, in tomato and cucumbers.

BABA conc Percent plants infected (ppm) Tomato Cucumber 0 100 100 62 20 125 0 200 0 0 500 0 0 1000 0 0 2000 0 0 37 EXAMPLE 26 -To asessthe e acy of BA-BA against leaf blast disease (Pyricularia oryzae) in rice plants.

-To assess the phytoto'5ucity of the product in rice planrts (if any).

b) Mvaterial and Method: -Location: Plant Protection Center-South Vietnam.

Long Dinh village, Chau Thanh distrit Tien Gang provine 0Se-Crop: R-ice plants. Variety: GMv 296.

-Experiniental desigh:

CRD.

-Treatment: 6 treatments as follows: Tramn -Doag *-AB *p AB 500*p 31 *BA

A*OOP

1-BABA 200 ppm 25 EC 625 ppm *0A- 16n-O Untreated 38 c) Test method: +Each treatment included5 clay pots (30 cm in diameter and 28 cm in height).

+A 14 day old rice seedling was transplanted in each clay pot.

+Water was always kept enough in the clay pots for the rice plants growth.

+3g of urea was added to each pot at 3 days after transplanting.

+Inoculation was .'one when rice plants were at 30 day old.

Blast diseased rice leaves were collected from the fields, cit into small pieces and kept moist (temperature of 250 C and 90% humidity) in three days for fungal growth and sporulation then, broadcasted on to the rice plants.

+After inoculation, the clay pots were placed under shade conditior for infection.

S Application timing: One spray at 7 days before inoculation of leaf blast pathogen for treatment 1 to 8 (23 day old), and one spray when the disease appears for treatment 9 (37 day old).

Assessment: Tssess the disease incidence and severity. All leaves per hill were taken scoring at before and every week after occurrence of the disease.

Disease incidence aid severity were computed as follows: Number of leaves infected DI X100 Total of leaves observed 9n9+Sn8+7n7+6n6+5n5+ 4 n 4 +3n3+2n2+nl X100 39 Table 16: leaf blast disease incidence.

TIr-eatm~ent Dosagye Incidence Disease(% (ppm) 1-BABA 2-BABA 3B ABA 4-BABA

EC

6-Untreated 100 500 1,000 625 7DAI* 9.2ab 8. 4b 10. 6ab 9.3 ab 1 1. lab 14DMI 21DAI 46.9b 5 1.9b 4 5.l1b 46.3b 47.l1b 48.9b 42. 8b 41.5b 40.5b 43.lb 28DMI 47.9b 46.6b 43.Ob 49.3b 44. 9b 81 .4a 46.4b 46.3 b 44.4b 44. 6b 42.5b 79. 6a 11.7 8.8 ab 65.2a.

8-1.8a, Cy 19.0 28.6 19.5 36.0 Note.

DAI* Day after infection Table 17: Leaf blast seventy Treatment Dosage Severity Disease 7DMI* 14DMI 21DMI 28DM I1-BAkBA 100 5.3a 15. b 18.4b 19.6b 16.1b 2-BABA 500 6.1a 12,7b 20.4b 20.2b 14.4b BABA 1,000 4.1a l11Ob 18. 3b 18.0b 1-5.2b 4-BABA 2,000 5.2a IL.9b 18.6b 18. lb 15.1b 5-Proniconazole25 625 5.l1a 15.9b 15.Ob 15. lb 15.4b

EC

6-10-untreated 4.8a.

12.0 19. 8a 26.3a 25.Oa 37.2 a Cv 3 4. 91 19.5 40.0 19.1 34.9 19.5 Table 18: Plant height (cm) and number of tillers per hill Treatment I1-BABA 2-BABA 3-BABA Dosage 7DMi 14DAI 21DAM (ppm) 100

N*

3 .n 46.7ns

N

12.2

P

55.8 N P 14. 58.1

N

14. 6ns 28DMI

P

68.1 a 65.4 a (9.5 a 500 3.6 45.8 12.4 55.2 13. 60.6 1,000 3.6 45.4 11.1 58.4 .14. 63. 8 1 13.2 13.7 4-MVCW 21-21 S -Propiconazole 25 EC 6-untreated

CV(%)

2,000 3.7 45.6 11.4 55.3 14.

625 -3.3 45.3 11.7 55.1 12.5 3.3 45.7 10.6. 50.6 -11.

8- 8.6 10.3 63.2 I2 2 66.2 a 61.8 12.3 69.4 a 58.0 11.3 53.4 b 16.9 6.3 Note:- Number of tillers per bI.

Plant height (cm) Table 19: Phytotoxicity Treatment Dosage(ppm) Phytotoxixcit IDAA 3DA.A 7DAA 1-BABA 2-BABA 3-BABA 4-BABA 25

EC

100 500 1,000 2,000 625 14DAA

I

I

I

I

Note: DANA Day after application.

41 (d)-Result and Discussion: 7 days after inoculation, rice leaves were infected by Pyricularia oryzea. Their disease incidence and severty (DI DS) were relatively uniform in all treatments. After that they increased, but the increases were lower than that of untreated treatments. (there was significantly different of disease seventy between trated treatments and untreated one by Duncant's test).

It means that BABA and Propiconazole at the tested dosages as mentioned above could prohibit the blast disease expansion on rice; However, there was no significant difference in the efficacy against blast disease between treated treatments.

There was different in height, but there was not different in number of tillers per hill between treated treatments and untreated one.

No phytotoxiccity was recorded after BABA application.

(e)--Conclusion: BABA with four tested dosages of 100 ppm-5 0 0 ppm-l,000ppm- 2 ,000 ppm, and Propiconazole 25 EC at the dosage of 625 were effective to control the development of blast disease on rice plnats. With BABA better at lower doseges.

BABA did not cause crop injury to rice plants.

In conclusion, BABA is as good as and even better than fungicide propiconazole against Pyriculana oruzae in rice plants, working by inducing local and systemic raisteace 42 nl: Number of leaves at score 1 .n3: Number of leaves at score 3; Number of leaves at score 5; n7: -do 7; n9: -do- 9.

N: Number of leaves observed.

Leaf blast scale: following standard (IRIR 2 d Edition, July 1980) Score 0 1 2.

4 Number of leaves at score 2.

Number of leaves at score 4.

-do- 6.

-do- 8.

r evaluation system for rice Description No lesions Small brown specks of pinhead size.

Larger brown specks Small, roundish to slightly elongated, necrotic gray sports about l-2mm in diameter with a brown margin.

Typical blast lesions, elliptical, 1-2 cm long, usually confined to the area of the 2 main veins, infecting less than 2% of the leaf area.

Typical blast lesions, infecting less than of leaf area.

Typical blast lesions, infecting 10-25% leaf area.

Typical blast lesions, infecting 26-50% leaf area.

Typical blast lesions, infecting 51-75% leaf area and many leaves dead.

All leaved dead.

r r +Plant phytotoxicity: Phytotoxicity was visually evaluated by EWRS's classification scale 1-9 no effect, 9: complete kill) at 1,3,7 and 14 days after treatment.

Count number of tillers, and measure the plant height every week.

43 EXAMPLE 27 The Preparation of n-benzoyl-3-aminobutyric acid- -methyl-1-butylester The mixture of 2.0 g n-benzoyl-3-aminobutryic acid, 1.5 g 2-pentanol, 30 ml toluene and 0.004 g H 2

SO

4 (conc.) was heated under reflux during 6.5 hr. The water formed in the remission was separated by azeotropic distillation. The cooled solution was washed with water (3X15ml), and with 2% sodium bicarbonate (3X5ml). After the evaporation of the solvent, and distillation of the excess alcohol at 30 0 C (0.4 mm Hg) oily product was obtained. The structure was confirmed by Mass spectroscopy and NMR spectroscopic methods. By proceeding as described above, using the suitable mass spectroscopy and starting materials, compounds 2 to 8 were prepared.

EXAMPLE 28 The Preparation of N-benzyl-3-aminobutyric acid-1-methyl-l-pentyl ester To 30 g crotonic acid dissolved in 120 g Ethyl alcohol, was added 37.2 g benzylamine at 10-15"C during 25 min. The mixture was heated at 75-77°C during 5 hr. The reaction was cooled to room temperature and n-benzyl-3-aminobutyric acid was filtered to afford 43.8g.

To 4.0 g of the benzyl product in 50 ml Toluene was added thionyl chloride (2.4 g) at room temperature during 15 minutes. The mixture was heated at 80C during 1 hr.

After cooling, 7.2 g 2-hexanol was added at room temperature during 15 min. The Sreaction was heated at 80*C during 4 hr. The mixture was washed with 2X50 ml water and 3X50 ml sodium bicarbonate After evaporation and distillation of the excess alcohol 4.4 g.compound was obtained. By further purification on a silica gel column using a mixture of Dichloroethane:Ethyl Acetate (90:10) as eluant, 3.2 g of the ester was obtained.

The structure was proved by NMR and Mass Spectroscopy. By proceeding as described above, using the suitable starting materials, compoundsl0 and 11 were prepared.

-44 The Preparation of ft-Aininobutyric acid hydrochloride g 0-aminobutyric acid was dissolved in 20 mld water. After the addition of 8.0 g ml 32, the solution was evaporated, anid the compound,ws dried.

EXAMPLE_ The Preparation of O-Amino, butyric acid sodium salt 6.0 g j3-arinobutyric acid was dissolve in 20 ml water. 2.3 g. NaOH dissolved in 12 ml water was added. The solution was evaporated, and the solid was dried.

EXAMPLE 31 The activity -of some BABA derivatives against powdery ruldew caused by Erysiphe Graminis in wheat, 8 days after inoculation was studied. The results are shown in Table 20, which is to be compared with the case without any treatment which showed only a efficacy of control.

Table 20: Actt of some BABA derivatives against ]powdery mildew caused b Erysiphe Graminis in Wheat 8 days after inoculatio Dose: 1000 ppm CH3C(H)NHR6CH2C(O)0R3 9 Compound No. IR 6 Percent Eficacy of Control

C

6 HsCH 2

C

6

HSCH

2

CH

2 C6H 5

CH

2

CH

2

CH

2

CI

H

CH(CH3)(CHA)CH 3

(CH

2 )7CH3

CH(CH

3 XCH2) 3

CH

3

H

Na 45 EXAMPLE 32 Follow the methods described in Examples 16 and 24, the activity of some BABA derivatives against late blight caused by Ptytophotra Infestans in potato was studied.

The results are shown in Table 2 1, which is to be compared with the case without any treatment which showed only a 12% efficacy of control.

Table 21: Actt of some BABA derivatives aninst late'bfigbt caused b PhytoRhitor Infestans in Potato. 6 day after inoculation Dose: 2000 ppm and 1000 ppm CE13C(H)NHR 6

CH

2 C(O)0R 3 a.

S

a a.

Compound No. Control Efficacy Dose yom

C

6 HsCO

C

6 HsCO

C

6 HsCO

C

6 HsCO

C

6 HsCO

-C

6 HsCO

C

6

H

5

CO

C

6

H

5

CO

CG~jsCH 2

CH

2 C6H 5

CH

2

CH

2

CH

2

CI

A

CH(CH

3

XCH

2 2

CH

3

CH(CH

3

XCH

2 2 CH3

(C-H

2 )6CH 3 (Cfl 2 )sCH 3

CH(CH

3

)(CH

2

)CH

3

ICH(CH)(CH

24

CH

3

CH(CH

3

XCH

23

CH

3 2000 2000 2000 2000 2000 2000 2000 2000 1000 1000 1000 1000

S

-46- EXAMPLE 33 Foilowig. the above described methods the activity of some BABA derivatives against late blight caused by Phytophtora Infestans in tomato was studied. The results are shown in Table 22, which is to be compared with the case without any treatment which showed only a 5% efficacy of control..

Table 22: Activity of some BABA derivatives UAainst late bliat causedby Phtophtora Infestans in Tomato. 4 days after inoculation Dose: 1000 ppm

CH

3

C(HI)NEOR

4 CH2C(O)0R 3 Compound No., R6 Control Efficacy

C

6 HsCH 2 CH2 C6j{CH 2

CH

2 CH2CI

H

(CH)7CH 3

CH(CH

3

)(CH

2 3 CH3

H

Na

Claims (8)

1. -N-benzoyl-3-axninobutyric acid 1-methyl-l-butyl ester.

2. -N-benzoyl-3-aminobutyric acid l-methyl-5-pentyl ester.

3. a. zoy5-.J-uniutyric acid n-heptyl ester.

4. -N-benzoyl-3-aminobutyric acid n-hexyl ester. -N-benzoyl-3-aminobutyric acid 1-methyl-1-heptyl ester. 15

6. -N-benzoyl-3-aminobutyric acid l-methyl-1-hexyl ester.

7. -e zyA3a...t rc cd 1 m t y -l otletr A: -N-benzoyl-3-axninobutyric acid 1-methyl-1-octyl ester. One.

8. -N-benzoy1-3-aminobutyric acid 1 -methyl-i -pentyl ester.

19. -N-b-penyletl3-aminobutyic acid 1 -methyl- -pentyl ester. 12. -N-choromethyl-3..aminobutyric acid. DATED this 30th day of April, 2001 1AGROGENE, LD By its Patent Attorneys kA <~DAVIES COLLISON CAVE