Compositions and methods for controlling nematodes

Abstract

Compositions and methods for controlling nematodes are described herein, e.g., nematodes that infest plants or animals. The compounds include oxazoles, oxadiazoles and thiadiazoles.

Claims

1. A method for control of plant parasitic nematodes, the method comprising administering to a plant, a seed or soil a composition comprising an effective amount of a compound of Formula III or a salt thereof, ##STR00193## wherein, A is phenyl or pyrazyl, each of which can be optionally independently substituted with one or more substituents selected from the group consisting of halogen, CF.sub.3, CH.sub.3, OCF.sub.3, OCH.sub.3, CN and C(H)O; and C is thienyl, furanyl, oxazolyl, or isoxazolyl, each of which can be optionally independently substituted with one or more substituents selected from the group consisting of fluorine, chlorine, CH.sub.3 and OCF.sub.3.

2. The method of claim 1 wherein A is phenyl.

3. The method of claim 1 wherein C is thienyl.

4. The method of claim 1 wherein C is furanyl.

5. The method of claim 1 wherein the composition comprises a compound of Formula IIIa or a salt thereof, ##STR00194## wherein, R.sub.1 and R.sub.5 are independently selected from the group consisting of hydrogen, CH.sub.3, F, Cl, Br, CF.sub.3, and OCF.sub.3; R.sub.2 and R.sub.4 are independently selected from the group consisting of hydrogen, F, Cl, Br, and CF.sub.3; R.sub.3 is selected from the group consisting of hydrogen, CH.sub.3, CF.sub.3, F, Cl, Br, OCF.sub.3, OCH.sub.3, CN, and C(H)O; R.sub.7 and R.sub.8 are independently selected the group consisting of from hydrogen and fluorine; R.sub.9 is selected from the group consisting of hydrogen, F, Cl, CH.sub.3, and OCF.sub.3; and E is O or S.

6. The method of claim 5 wherein E is S.

7. The method of claim 5 wherein E is O.

8. The method of claim 1 wherein the composition comprises a compound of Formula IIIb or a salt thereof, ##STR00195## wherein, R.sub.1 and R.sub.5 are independently selected from the group consisting of hydrogen, CH.sub.3, F, Cl, Br, CF.sub.3, and OCF.sub.3; R.sub.2 and R.sub.4 are independently selected from the group consisting of hydrogen, F, Cl, Br, and CF.sub.3; R.sub.3 is selected from the group consisting of hydrogen, CH.sub.3, CF.sub.3, F, Cl, Br, OCF.sub.3, OCH.sub.3, CN, and C(H)O; R.sub.8 is selected from the group consisting of hydrogen and fluorine; R.sub.6 and R.sub.9 are independently selected from the group consisting of hydrogen, F, Cl, CH.sub.3, and OCF.sub.3; and E is O or S.

9. The method of claim 8 wherein E is S.

10. The method of claim 8 wherein E is O.

11. The method of claim 1 wherein the composition comprises 2-(4-chlorophenyl)-5-(thiophen-2-yl)-1,3,4-oxadiazole.

12. The method of claim 1 wherein the composition comprises a surfactant.

13. The method of claim 1 wherein the composition comprises a co-solvent.

14. The method of claim 1 wherein the composition further comprises one or more of an insecticide, a fungicide, an herbicide, or another pesticide.

15. The method of claim 14 wherein the insecticide, fungicide, herbicide or the pesticide is selected from the group consisting of avermectin, ivermectin, milbemycin, imidacloprid, aldicarb, oxamyl, fenamiphos, fosthiazate, metam sodium, etridiazole, penta-chloro-nitrobenzene (PCNB), flutolanil, metalaxyl, mefenoxam, fosetyl-al, silthiofam, fludioxonil, myclobutanil, azoxystrobin, chlorothalonil, propiconazole, tebuconazole, pyraclostrobin, trifloxysulfuron, glyphosate, and halosulfuron.

16. The method of claim 1 wherein the plant parasitic nemoatode is selected from the group consisting of: M incognita, H. glycines, B. longicaudatus, H. contortus, A. suum, and B. malayi, or wherein the plant parasitic nematode is a nematode of one of the following genera: Pratylenchus, Heterodera, Globodera, Meloidogyne, Rotylenchulus, Hoplolaimus, Belonolaimus, Longidorus, Paratrichodorus, Ditylenchus, Xiphinema, Helicotylenchus, Radopholus, Hirschmanniella, Tylenchorhynchus, and Trichodorus.

17. The method of claim 1 wherein the composition is applied to a seed.

Description

BRIEF DESCRIPTION OF DRAWINGS

(1) FIG. 1: Root galling seen in plants with no chemical applications (Fall trial).

(2) FIG. 2: Typical root galling seen in plants treated with 2 kg/ha 4776 (Fall trial).

(3) FIG. 3: Typical root galling in plants treated with 2 kg/ha 4559 (Fall trial).

(4) FIG. 4: Typical root galling in plants treated with 2 kg/ha of the commercial nematicide oxamyl (Fall trial).

(5) FIG. 5: Root galling seen in plants with no chemical applications (Summer trial).

(6) FIG. 6: Typical root galling seen in plants treated with 4 kg/ha 5823 (Summer trial).

(7) FIG. 7: Typical root galling in plants treated with 4 kg/ha 5938 (Summer trial).

DESCRIPTION OF THE PREFERRED EMBODIMENTS

(8) Described herein are certain compounds, some of which are oxazole, oxadiazole and thiadiazole analogs with potent broad spectrum nematicidal activity.

(9) The nematicidal compounds may be supplied to plants exogenously, through sprays for example. These compounds may also be applied as a seed coat. The compounds can be applied to plants or the environment of plants needing nematode control, or to animals or the food of animals needing nematode parasite control. The compositions may be applied by, for example drench or drip techniques. With drip applications compounds can be applied directly to the base of the plants or the soil immediately adjacent to the plants. The composition may be applied through existing drip irrigation systems. This procedure is particularly applicable for cotton, strawberries, tomatoes, potatoes, vegetables and ornamental plants. Alternatively, a drench application can be used where a sufficient quantity of nematicidal composition is applied such that it drains to the root area of the plants. The drench technique can be used for a variety of crops and turf grasses. The drench technique can also be used for animals. Preferably, the nematicidal compositions would be administered orally to promote activity against internal parasitic nematodes. Nematicidal compositions may also be administered in some cases by injection of the host animal or by topical applications.

(10) The concentration of the nematicidal composition should be sufficient to control the parasite without causing significant phytotoxicity to the desired plant or undue toxicity to the animal host. The compounds disclosed in this invention have a good therapeutic window.

(11) We have surprisingly found that certain oxazole, oxadiazole and thiadiazole analogs (e.g., 5-(4-chloro-2-fluorophenyl)-2-(thiophen-2-yl)oxazole, 3-(4-chloro-2-methylphenyl)-5-(furan-2-yl)-1,2,4-oxadiazole, 3-(2,4-dichlorophenyl)-5-(furan-2-yl)-1,2,4-thiadiazole) have nematicidal potencies comparable with organophosphate and carbamate standards yet display excellent selectivity for nematodes over plants and animals. Thus, these analogs will provide useful compounds for nematode parasite control.

(12) The nematicidal agents described herein can be applied in conjunction with another pesticidal agents. The second agent may, for example, be applied simultaneously or sequentially. Such pesticidal agents can include for example, avermectins for animal applications.

(13) The aforementioned nematicidal compositions can be used to treat diseases or infestations caused by nematodes of the following non-limiting, exemplary genera: Anguina, Ditylenchus, Tylenchorhynchus, Pratylenchus, Radopholus, Hirschmanniella, Nacobbus, Hoplolaimus, Scutellonema, Rotylenchus, Helicotylenchus, Rotylenchulus, Belonolaimus, Heterodera, other cyst nematodes, Meloidogyne, Criconemoides, Hemicycliophora, Paratylenchus, Tylenchulus, Aphelenchoides, Bursaphelenchus, Rhadinaphelenchus, Longidorus, Xiphinema, Trichodorus, and Paratrichodorus, Dirofiliaria, Onchocerca, Brugia, Acanthocheilonema, Aelurostrongylus, Anchlostoma, Angiostrongylus, Ascaris, Bunostomum, Capillaria, Chabertia, Cooperia, Crenosoma, Dictyocaulus, Dioctophyme, Dipetalonema, Dracunculus, Enterobius, Filaroides, Haemonchus, Lagochilascaris, Loa, Manseonella, Muellerius, Necator, Nematodirus, Oesophagostomum, Ostertagia, Parafilaria, Parascaris, Physaloptera, Protostrongylus, Setaria, Spirocerca, Stephanogilaria, Strongyloides, Strongylus, Thelazia, Toxascaris, Toxocara, Trichinella, Trichostrongylus, Trichuris, Uncinaria, and Wuchereria. Particularly preferred are nematodes including Dirofilaria, Onchocerca, Brugia, Acanthocheilonema, Dipetalonema, Loa, Mansonella, Parafilaria, Setaria, Stephanofilaria, and Wucheria, Pratylenchus, Heterodera, Meloidogyne, Paratylenchus. Species that are particularly preferred are: Ancylostoma caninum, Haemonchus contortus, Trichinella spiralis, Trichurs muris, Dirofilaria immitis, Dirofilaria tenuis, Dirofilaria repens, Dirofilari ursi, Ascaris suum, Toxocara canis, Toxocara cati, Strongyloides ratti, Parastrongyloides trichosuri, Heterodera glycines, Globodera pallida, Meloidogyne javanica, Meloidogyne incognita, and Meloidogyne arenaria, Radopholus similis, Longidorus elongatus, Meloidogyne hapla, and Pratylenchus penetrans.

(14) The following examples are, therefore, to be construed as merely illustrative, and not limitative of the remainder of the disclosure in any way whatsoever. All of the publications cited herein are hereby incorporated by reference in their entirety.

EXAMPLES

Example 1

M. incognita Testing of Several Nematicidal Compounds in a Miniaturized Greenhouse Assay

(15) Overview:

(16) The test compound is dissolved in an acetone solution and added to water. A sprouted cucumber seedling is placed into a vial with dry sand and the water-chemical solution is added immediately. Twenty four hours later Meloidogyne incognita eggs are added to the vials and 10 to 12 days later the roots are evaluated for nematode galling.

(17) Procedure:

(18) Cucumber seeds are sprouted for 3 days in moist paper towels. Acceptable sprouts should be 3 to 4 cm long with several lateral roots just emerging. Stock solutions of chemistry are prepared in a mixture of acetone and Triton X100 (412 mg in 500 mL) to a final concentration of 5 mg/mL. The chemical stock solution is then added to 10 mL deionized water plus 0.015% Triton X100 and mixed thoroughly. This is enough to test each condition in triplicate. Ten mL dry sand is added to each vial. At this time the solubility of the chemistry is visually determined and recorded as either ppt (large precipitates) or cloudy (fine precipitates). Seedlings are planted by tilting the vial and laying the seedling in the correct orientation so that the cotyledons are just above the sand and then tilting back to cover the radicles with sand. 3.3 ml water/chemical mix is added to each vial and the vials placed in racks under fluorescent light banks. The vials are inoculated two days after planting by adding 500 vermiform M. incognita eggs to each vial in 50 uL of deionized or spring water. The vials are then kept under the fluorescent lamps at ambient room temperature and watered as needed with 1 mL deionized water, usually twice during duration of test. Harvest of the cucumber plants is done 10 to 12 days after inoculation by washing sand off the roots. A root gall rating and visual phytotoxicity rating is assigned using the following scales: Gall rating scale (Gall: % root mass galled): 0=0-5%; 1=6-20%; 2=21-50%; and 3=51-100%. The average of the triplicate gall rating is then calculated: green=0.00-0.33 (no galls); yellow=0.67-1.33 (mild galling); orange=1.67-2.33 (moderate galling); red=2.67-3.00 (severe galling). Visual phytotoxicity scale is also assigned (Vis. tox; visual reduction in root mass compared to the control): rs1=mild stunting; rs2=moderate stunting; rs3=severe stunting.

(19) TABLE-US-00001 TABLE 1A Potent nematicidal oxadiazole and oxazole 2-thiophene and 2-furan analogs showing examples of substitutions compatible with high activity 8 ppm gall Name Analog ratings 1822 0 1846 0 4417 0.33 4559 0 4775 0 0 4776 0 4948 0 4971 0.67 5006 0 5012 0.67 5082 1.67 5090 1.67 5132 1.33 5181 0.33 5212 0 1 5213 0.33 5292 0.67 5297 0.33 5456 0.67 5467 0 5468 1 5475 1.33 5478 0 5479 0 5499 0 0 5523 0 5527 0.67 5556 0.33 5586 0.67 5587 0 5618 1.33 5622 0 5623 0 5625 0.33 5663 0 0 5666 1.33 5671 0.67 5672 0 Oxamyl 0.67 (1 ppm)

(20) A variety of single substitutions on or in the six membered aromatic ring (e.g., pyridine or pyrazine in place of phenyl) of the phenyl-2-furan and phenyl-2-thiophene oxadiazoles and oxazoles are compatible with high nematicidal activity. Examples of preferred single substitutions include halogens, CH.sub.3, CF.sub.3, OCF.sub.3 and OCH.sub.3 especially in the para position (4-position) of the phenyl ring. The phenyl ring can also be multiply substituted in a way compatible with high nematicidal efficacy. Ring numbering system is shown below.

(21) ##STR00064##

(22) TABLE-US-00002 TABLE 1B Examples of nematicidal thiadiazole, oxadiazole and oxazole analogs with potency comparable to commercial standards Name Analog 1 ppm gall ratings* 4776 1.sup.a, 1.sup.b, 0.33.sup.c, 0.33.sup.d 1822 0.33.sup.a, 0.67.sup.b, 0.33.sup.c, 0.sup.d 4559 1.sup.a 5499 1.sup.a 1846 1.33.sup.a, 0.67.sup.b 5467 0 1.67.sup.a, 1.33.sup.b 5479 1.sup.a, 0.67.sup.b 5523 1.sup.a, 1.33.sup.b 5527 1.67.sup.a, 1.sup.b 5823 1.67.sup.a, 0.33.sup.b, 0.33.sup.e 5825 0.sup.a, 0.33.sup.b 5383 1.33.sup.a 5864 1.sup.a 5882 0.67.sup.a 5969 1.sup.e 5915 0 0.33.sup.e 5970 1.sup.e 5938 0.67.sup.e 5960 0.33.sup.e Oxamyl 0.67.sup.a, 1.sup.b, 1.33.sup.c, 1.33.sup.d, 1.sup.e Fenamiphos 0.sup.c, 0.sup.d, 0.sup.e *Data with the same letters are taken from the same test.

(23) Several phenyl-2-furan and phenyl 2-thiophene oxadiazoles, oxazoles and thiadiazoles have nematicidal potency equivalent to the commercial carbamate nematicide oxamyl and the commercial organophosphate nematicide fenamiphos. Oxamyl and fenamiphos are highly toxic compounds classified as toxicity Class I chemicals by the US Environmental Protection Agency. Also noteworthy is the fact that some multiply substituted analogs are especially nematicidal.

(24) TABLE-US-00003 TABLE 1C Nematicidal activity 3-furan and 3-thiophene analogs 1 ppm gall Name Analog rating* 5885 1.sup.a 5867 1.sup.a 5869 1.sup.a 5886 1.33.sup.b 5887 1.sup.b 1822 0.sup.a, 0.33.sup.b 4776 0 1.sup.a, 0.33.sup.b, 1.sup.c 5882 0.67.sup.c 5876 1.67.sup.c Oxamyl 1.33.sup.a, 1.sup.b, 0.67.sup.c *Data with the same letters are taken from the same test.

(25) Strong nematicidal activity is not limited to 2-furan and 2-thiophene analogs and is also seen with 3-furan and 3-thiophene. Additionally certain substitutions on the 5-membered thiophene or furan rings appear to be well tolerated.

(26) TABLE-US-00004 TABLE 1D Comparison of nematicidal oxazoles and oxadiazoles with nematicidal pyrazoles and thiazoles 8 ppm 1 ppm gall gall Name Analog rating* rating* 5725 1.33.sup.a 3.sup.a 5735 0.sup.a 2.sup.a 5738 0.sup.a 1.33.sup.a 5741 0.sup.a 1.sup.a 4776 0.sup.a 0.sup.a 1822 0.sup.a 1.33.sup.a 5663 0.sup.b 1.67.sup.b 1787 00 1.67.sup.b 3.sup.b 5645 01 0.sup.b 2.sup.b Oxamyl 1.33.sup.a, 1.sup.b *Data with the same letters are taken from the same test.

(27) Oxazoles and oxadiazole analogs of the current invention show significant enhancement in nematicidal potency over comparable nematicidal pyrazoles or nematicidal thiazoles.

Example 2

General Greenhouse Testing Protocols

(28) Soybean Planting and Growth:

(29) Soybeans seeds are planted in 100% sand in two inch square plastic pots. Chemical treatment is done when the soybeans show the first trifoliate beginning to emerge about 10 to 12 days after planting. At least four hours after chemical application the nematode soybean cyst nematode (SCN) eggs are applied and 28 days after the egg inoculation the test is harvested.

(30) Cucumber Planting and Growth

(31) Cucumber seeds are planted in a sandy soil mixture in two inch square plastic pots. When the cotyledons are fully opened and just as the first leaf begins to emerge, usually 7 days after planting, chemistry for the 7 day treatment is applied. One week later the chemistry for the 0 day treatment is applied. Separate plants are used for each application. The plants are generally in the 1-2 leaf stage now. At least four hours after the chemistry application the pots are inoculated with root knot nematode (RKN) eggs. Plants are rated for galling 14 days after the egg inoculation.

(32) Chemical Formulation and Application

(33) One milligram of chemistry per four pots is equal to one kilogram per hectare of chemical. A standard test uses four replications. For rates above 2 kg/ha, the desired amount of chemical is weighed into a 30 ml vial (example: 8 kg/ha rate=8 mg chemical in 30 ml vial). The chemical is dissolved in 2 ml of appropriate solvent, generally acetone. For rates below 2 kg/ha, 2 milligrams of chemistry is weighed into the vial and dissolved in 2 ml of the solvent. The appropriate amount of chemical concentrate is then pipetted into a separate 30 ml vial and solvent is added to bring the volume to 2 ml (example 0.5 kg/ha=0.5 ml of concentrate+1.5 ml solvent). Each dissolved concentrate is then brought to a total of 20 milliliters using 0.05% Triton X 100 surfactant solution.

(34) Chemical and Nematode Application

(35) Pots to be treated are moist but not saturated. To each of four pots, five milliliters of the appropriate chemical solution is pipetted to the media surface making sure to avoid contact with the base of the plant. Immediately following chemical application, using a mist nozzle, the pot surface is wetted sufficiently to saturate the pot watering in the chemistry. The chemical application is done in the morning.

(36) Nematode eggs, either SCN or RKN, are added to distilled water to create a concentration of 1000 vermiform eggs per liter of water. At least four hours after chemical treatment the eggs are applied to the treated pots plus non-treated check plants. A small hole about 1 cm deep is punched into the pot surface. One milliliter of the nematode egg slurry is pipetted into the hole. Immediately afterwards the hole is gently covered. Watering of the test plants is then restricted to only water as needed to prevent wilt for a period of 24 hours. After the 24 hour restricted watering, normal sub-irrigation watering is done for the duration of the test.

(37) TABLE-US-00005 TABLE 2A SCN greenhouse sand assay on soybean plants Name Analog 2 kg* 1 kg* 0.5 kg* 0.25 kg* 0.1 kg* 1822 02 100.sup.a 4559 03 98.sup.a 4776 04 99.sup.a 89.sup.c 78.sup.c 5181 05 100.sup.a 5292 06 92.sup.a 4417 07 94.sup.b 4775 08 95.sup.b 5823 09 69.sup.d 38.sup.d 5915 0 74.sup.d 44.sup.d 5938 89.sup.d 60.sup.d 5939 88.sup.d 64.sup.d Fenamiphos 98.sup.a 98.sup.b 94.sup.c 26.sup.d 5.sup.d *Rate in kg/ha. Data shows percent control (i.e., cyst number reduction) relative to the control blank treatment. Data with the same letters are taken from the same test.

(38) The oxazoles, oxadiazoles and thiadiazoles of this invention are highly efficacious nematicides against soybean cyst nematode with potencies comparable to fenamiphos demonstrating that this area of chemistry has broad nematicidal spectrum.

(39) TABLE-US-00006 TABLE 2B RKN greenhouse soil assay on cucumber plants 0 day kg/ha rate* 7 day kg/ha rate* Name Analog 1 0.25 0.1 0.05 1 0.25 0.1 0.05 5823 95.sup.a 98.sup.c 85.sup.a 91.sup.c 53.sup.a 38.sup.c 5825 94.sup.b 89.sup.a 84.sup.b 50.sup.a 53.sup.a 97.sup.b 5860 85.sup.a 47.sup.a 86.sup.a 1822 89.sup.a 81.sup.b 60.sup.a 64.sup.b 47.sup.a 7.sup.a 85.sup.a 75.sup.b 4776 99.sup.b 5960 76.sup.c 75.sup.c 75.sup.c 5961 81.sup.c 88.sup.c 73.sup.c *Data shows percent control (i.e., galling reduction) relative to the control blank treatment. Data with the same letters are taken from the same test.

(40) Certain oxazoles, oxadiazoles and thiadiazoles are highly efficacious nematicides in bioactive soil with potencies comparable to fenamiphos and activities that are resistant to biotic or abiotic degradation over a timeframe of least one week.

(41) TABLE-US-00007 TABLE 2C RKN greenhouse soil assay on cucumber plants showing comparison of two different formulations. Name Analogs Acetone 1 mg/kg* Radex 1 mg/kg* 1822 0 94 98 5825 96 96 1846 88 86 5523 86 86 5527 91 80 5479 91 96 5467 73 88 Fenam 98 99 *Data shows percent control (i.e., galling reduction) relative to the appropriate control blank treatment. The Acetone formulation is the standard 10% acetone in 0.05% Triton X 100 formulation described above. The Radix formulation was prepared by adding 10 mg of each compound to 150 mg of a mixture of 12% Triton X 100, 11% Agsolex 8, 33% Agsolex 1 and 44% Steposol SC (all by weight). Final was 6.25% active ingredient by weight.

(42) The nematicidal activity of this area of chemistry is not compromised on moving from a typical screening formulation with high amounts of acetone to an emulsifiable concentrate format typical used in commercial applications.

Example 3

Belonalaimus Longicaudatus (Sting Nematode) Testing Protocols

(43) Populations of sting (Belonolaimus longicaudatus) nematodes are maintained on St. Augustine turf grass on soil in 15-cm pots. At test initiation the turf is removed from the pots and the soil containing nematode eggs, juveniles, and adults is subdivided into pots each containing a volume of 125 cm.sup.3 The compounds to be tested are dissolved in 3 ml of acetone using 3, 6, or 15 mg to achieve equivalent surface area application rates of 2, 4, or 10 kg/ha, respectively. The 3 ml acetone stock solution is added to 30 ml of water, and 5 ml of that solution is used to drench each of 6 replicate test pots prepared as described above. The treated pots containing nematodes are incubated in the laboratory at ambient temperature of approximately 25 C. After 3 days the soil from each pot is washed onto a modified Baermann apparatus comprised of a screen supporting a layer of filter paper on which the soil sample is placed and set in a dish of water. The samples are then incubated at 25 C. for 24 hours to allow the live nematodes to migrate through the paper and screen and into a water reservoir to be collected for counting with a light microscope. Nematodes that have been killed or immobilized by the test compounds are not able to migrate into the reservoir.

(44) TABLE-US-00008 TABLE 3 Efficacy against the sting nematode in a bench top soil assay Name Analog 2 kg/ha 4 kg/ha 10 kg/ha Other 4417 24 13 7 4559 39 47 33 4775 15 7 4 4776 0 16 19 20 Positive.sup.# 20 Negative.sup.# 65 Water 62 *Number of nematodes recovered from treated soil after 3 days incubation with the compound .sup.#11.1 kg fenamiphos used as positive control, acetone formulation blank used to dissolve compounds in the negative control.

(45) Certain oxazoles and oxadiazoles are highly efficacious nematicides against the sting nematode which is a serious pest on turf grass. These analogs have potencies comparable to fenamiphos demonstrating that this area of chemistry has broad nematicidal spectrum.

Example 4

C. elegans Testing Protocols

(46) Various compounds were tested for nematicidal activity against C. elegans using contact assays in wells. The assays were performed as described below. The test compounds were solubilized in DMSO at 10 mg/ml to create 100 stock solutions. A dilution series was created by diluting the stock solution with DMSO. For each well assay 4 ul of the appropriate dilution is added to a well of a test plate.

(47) A 400 ul aliquot of bacterial stock (in M9 buffer with ampicillin and nystatin) are added to each well of the test plate. Worms are added and the test plate placed on a rotary shaker and held at 20 C. Worms are examined and scored at 4 hrs, 24 hrs, 48 hrs and 72 hours.

(48) L1 worms and L4 worms were used in the assay. L1 worms are prepared by plating eggs on a plate without a bacterial feeding layer. The eggs hatch and arrest at the L1 stage. This L1 stage population is then used to create a stock for the experiments. To create an L4 stage stock a small number of worms are taken from an overgrown and starved plate of worms and seeded on a plate with a bacterial feeder layer. A 25 ul aliquot of worms is added to each well in the assay.

(49) TABLE-US-00009 TABLE 4 Three day C. elegans well assay of nematicidal oxadiazole and oxazole analogs L1 L1 L1 L4 L4 L4 Name Analog 1 D* 2 D* 3 D* 1 D* 2 D* 3 D* 5820 0.4 0.4 0.4 no (25F1) (6.3F1) 5821 0.4 0.4 0.4 no (0.4F1) (0.4F1) 5822 1.6 0.4 0.4 no 1.6 (1.6F1) 5823 0.4 0.4 0.4 1.6 0.4 (0.4F1) 5824 1.6 0.4 0.4 no no (1.6F1) 5825 0.4 0.4 0.4 1.6 1.6 (1.6F1) 5826 6.3 1.6 1.6 6.3 6.3 (6.3F1) 5827 6.3 1.6 1.6 25 6.3 (6.3F1) 5828 1.6 1.6 1.6 no no no 5845 0 no 1.6 0.4 no 25 (25F1) 5846 1.6 0.4 0.4 1.6 1.6 (1.6F1) 5847 no 0.4 0.4 no 1.6 (1.6F1) 5848 1.6 0.4 0.4 1.6 1.6 (1.6F1) 5849 6.3 0.4 1.6 no (6.3F1) (6.3F1) 5850 1.6 0.4 0.4 1.6 1.6 (1.6F1) 5860 1.6 0.4 0.4 1.6 1.6 (1.6F1) 5861 0.4 0.4 0.4 1.6 1.6 (1.6F1) 5905 0.4 0.4 0.4 ND ND ND 5906 0.4 0.4 0.4 ND ND ND 5938 0 1.6 1.6 1.6 ND ND ND 5939 0.4 0.4 0.4 ND ND ND 5915 0.4 0.4 0.4 ND ND ND *EC50 in parts per million of compound after one day, two days or three days of exposure for L1 larvae or L4 larvae. L4 data in parentheses refer to effects on the second generation larvae. ND: Experiment not done.

(50) The free living nematode C. elegans is highly diverged genetically from the tylenchid parasites such as soybean cyst nematode and root knot nematode. Therefore the nematicidal activity of these oxazoles, oxadiazoles and thiadiazoles against C. elegans L1 larvae and L4 larvae further confirms that this is chemistry is broadly active against various nematode species and stages.

Example 5

Mouse Acute Toxicity Testing

(51) Acute oral toxicity testing was performed in mice in accordance with test method P203.UDP, as administered by Eurofins/Product Safety Laboratories (Dayton, N.J.). CD-1/Swiss derived albino mice were obtained and group housed in suspended solid bottom caging. The mice were fed rodent chow and filtered tap water was supplied ad libitum. Following acclimation to the laboratory setting, a group of animals was fasted overnight by removing food from the cages. After the fasting period, three female mice were selected based on vitality and initial body weights. The individual compound doses were calculated from these body weights.

(52) The test substance was prepared as a 1% (50 mg/kg) or 5% (500 mg/kg) weight to weight (w/w) mixture in a 0.5% w/w solution of carboxymethylcellulose (CMC) in distilled water. A tissue homogenizer was used to create a homogeneous mixture. A dose of 50 or 500 mg/kg was administered to three healthy mice per dose level by oral intubation using a ball-tipped gavage needle attached to a syringe. After administration, the animals were returned to their cages, and feed was replaced immediately after dosing.

(53) The animals were observed for mortality, signs of gross toxicity and behavioral changes during the first several hours post dosing and at least once daily for up to 14 days. Body weights were recorded prior to initiation and on Days 7 and 14 or a soon as possible after death.

(54) Results were obtained for the following compounds:

(55) 1822:

(56) ##STR00153##

(57) At a dose of 50 mg/kg all animals survived, gained body weight, and appeared active and healthy. There were no signs of gross toxicity, adverse pharmacologic effects, or abnormal behavior. At a dose of 500 mg/kg all animals died within three days of test substance administration.

(58) 4417:

(59) ##STR00154##

(60) At a dose of 500 mg/kg two animals appeared active and healthy and gained body weight over the 14-day observation period. One animal died within four days of substance administration.

(61) 4775:

(62) ##STR00155##

(63) At a dose of 500 mg/kg all animals survived, gained body weight, and appeared active and healthy. There were no signs of gross toxicity, adverse pharmacologic effects, or abnormal behavior.

(64) 4776:

(65) ##STR00156##

(66) At a dose of 500 mg/kg two animals died within three days of substance administration. One animal appeared active and healthy during the entire study and gained weight over the 14-day observation period

(67) 5960:

(68) ##STR00157##

(69) At a dose of 500 mg/kg all animals survived, gained body weight, and appeared active and healthy. There were no signs of gross toxicity, adverse pharmacologic effects, or abnormal behavior.

(70) Based on these mouse studies the oral toxicity of 1822 appears to be between 50 mg/kg and 500 mg/kg, that of 4776 mg/kg to be slightly lower than 500 mg/kg, that of 4417 to be slightly higher than 500 mg/kg and that of 4775 and 5960 to be greater than 500 mg/kg. In comparison, the oral LD50 for aldicarb, oxamyl and fenamiphos in mice are 300 ug/kg, 2.3 mg/kg and 22.7 mg/kg respectively.

(71) Consequently, although the oxazole and oxadiazole chemistry of this invention has broad spectrum nematicidal activity these compounds nonetheless show considerable improvement in safety over the commercial organophosphate and carbamate standards and over abamectin (oral mouse LD50 13.6 mg/kg) the active ingredient the nematicidal seed treatment Avicta.

Example 6

Advanced Greenhouse Testing Protocols

(72) Pre-plant Incorporated Test (PPI)

(73) The PPI test examines the effect of pre-incorporation of compounds in soil and longer aging to simulate in furrow methods of nematicide application in the field. The PPI test exposes compounds to a higher volume of soil and drying which can result in more severe soil binding. Compounds are also aged for longer periods which can lead to more extensive biotic and abiotic degradation further limiting activity.

(74) The chemically treated soil (sandy soil mix) for all treatment days (e.g., 7 days, 14 days, 21 days) treatments is potted into their appropriate pots. On the same day the 7 day treatment pots are seeded. One week later eggs are applied and 14 days after egg application the test is harvested. The 14 day treatments are planted 7 days after the first planting. The 14 day planting and 7 day inoculation happen on the same day. One week later the 14 day treatments are inoculated with eggs. These are harvested 14 days after the inoculation. The 21 day treatments are planted 14 days after the first planting. The 14 day inoculation and 21 day planting are done on the same day. One week later the 21 day plants are inoculated with eggs. The 7 day treatment is harvested the same day as the 21 day inoculation. Fourteen days after inoculation the 21 day plants are harvested.

(75) TABLE-US-00010 Treatment Planting Inoculation Harvest 7 day day 0 day 7 day 21 14 day day 7 day 14 day 28 21 day day 14 day 21 day 35

(76) For each compound a stock is prepared using 4 mg material in 4 ml of acetone. The soil is mixed by placing 80 ml of field soil and 320 ml of sand in a plastic bag and mixing well. The formulation for treatment is done by adding 2.13 ml (8 kg/ha rate), 1.06 ml (4 kg/ha rate) or 0.53 ml (2 kg/ha rate) to a vial and raising it with 10 ml in 0.05%100. Soil is then treated by adding the entire 10 ml to the 400 ml of mix in the bag. The treated soil is immediately mixed well in the sealed bag to distribute the compound evenly. Approximately 95 ml is used to fill each 2-inch square pot up to the top with some soil compression and flattening. For each compound and for the control treatments 4 pots are filled. All pots are watered until moist but with no run-out through the bottom.

(77) The PPI test simulates 8, 4 and 2 kg/ha rates incorporated 15 cm deep in the field and is equivalent to the 2, 1 and 0.5 kg/ha drench application rates in the standard 2-inch pot cucumber greenhouse assay.

(78) TABLE-US-00011 TABLE 6A Seven day pre-plant incorporated greenhouse studies of root knot nematode on cucumber plants 8 ka/ha 4 kg/ha Name Analog rate* rate* 1822 99 99 5213 98 85 Fenamiphos 100 96 *Data shows percent control (i.e., galling reduction) relative to the control blank treatment.

(79) TABLE-US-00012 TABLE 6B Fourteen day pre-plant incorporated greenhouse studies of root knot nematode on cucumber plants Name Analog 8 ka/ha rate* 4 kg/ha rate* 2 kg/ha rate* 1822 0 100.sup.a 97.sup.a 67.sup.a 5467 100.sup.a 76.sup.a 71.sup.a 5479 100.sup.a 89.sup.a 71.sup.a 5523 99.sup.a 87.sup.a 59.sup.a 5527 96.sup.a 90.sup.a 57.sup.a 5823 100.sup.a 100.sup.b 98.sup.a 94.sup.b 85.sup.a 5825 96.sup.a 98.sup.a 69.sup.a 5915 99.sup.b 70.sup.b 5938 100.sup.b 90.sup.b Fenamiphos 100.sup.a 99.sup.a 88.sup.a 100.sup.b 100.sup.b *Data shows percent control (i.e., galling reduction) relative to the control blank treatment. Data with the same letters are taken from the same test.

(80) TABLE-US-00013 TABLE 6C Twenty one day pre-plant incorporated greenhouse studies of root knot nematode on cucumber plants 8 ka/ha 4 kg/ha Name Analog rate* rate* 1822 95 82 4776 0 80 50 Fenamiphos 99 84 *Data shows percent control (i.e., galling reduction) relative to the control blank treatment.

Example 7

Fall Nematicidal Field Testing

(81) Site Establishment

(82) The test site was located at 3511 Highway F in New Melle, Mo., Saint Charles county. The soil was a native prairie/pasture covering silty clay loam soil. Holes were dug using a Bobcat 763 skid loader with a 12-inch auger to a depth of 18 inches. Total volume of each hole was about 1.2 cubic feet. Six cubic yards of topsoil and 9 tons of river sand were purchased from Dardenne Farms Topsoil. Mixing to a ratio of 4 volumes sand to 1 volume soil was accomplished using a trailer-mounted 9 cubic foot concrete mixer. Holes were filled and then re-filled 5 days later after settling. The mixture was 92.5% sand, 2.5% silt, and 5% clay. Organic matter was 0.2% and pH was 6.8.

(83) Plots were planted with squash seeds and had a uniform stand of squash seedlings (2 per plot, 10 cm apart) with the first true leaf emerging just prior to treatment.

(84) Treatment and Inoculation

(85) Treatments were arranged in a block design with the blocks laid out perpendicular to the primary slope and parallel to secondary slope. There are 7 inoculated controls and 5 non-inoculated controls and the distributions of disease severity appeared independent of location.

(86) TABLE-US-00014 Randomized complete block design N blk 1 blk 2 blk 3 blk 4 blk 5 blk 6 1 6 8 3 7 9 5 10 5 1 9 9 2 7 9 4 9 8 4 5 7 2 2 7 6 2 6 10 8 6 3 1 3 10 6 5 8 9 4 8 1 4 9 3 10 5 4 3 7 4 1 7 3 2 10 8 2 6 5 1

(87) Application rates are expressed as kg of active material per hectare, and the mg per plot is based on the surface area of the bored and filled holes (0.000008559 Ha). The DC compounds were formulated immediately before application as follows: 1) the amount required to treat all six replicates was dissolved in 300 ml of acetone, 2) for each plot 50 ml of that solution was added to a graduated cylinder with 2 ml of 12.5% Triton X100 and the volume was raised to 500 ml with tap water. The resultant mix is the same as used in the standard greenhouse assays (10% acetone, 0.05%100). The oxamyl treatments were prepared from Vydate 2 L formulated the same way. The 500 ml was placed in a watering can and the entire volume was evenly sprinkled over the surface of the plot. No runoff occurred and pooling, if any, was short lived. The final drench volume was 0.58 ml/cm.sup.2, compared to 0.2 ml/cm.sup.2 used in our greenhouse, however the microplots are much deeper so the drench volume applied per soil volume treated is roughly the same.

(88) TABLE-US-00015 TABLE 7A Compound treatment list Field Name Analog Rate* Amount* 4417 2 17 4776 2 17 4559 2 17 Oxamyl 5 43 2 17 NT NI *Field rate in kg ai per hectare and amount of compound added in mg ai per plot. NT = non treated (i.e., inoculated with nematodes but not treated with chemicals) NI = non inoculated (i.e., not treated with chemicals or inoculated with nematodes)

(89) Meloidogyne incognita eggs were harvested over a two week period and stored at approximately 5 C. until needed. A stock of 5.6 million vermiform eggs was adjusted to 9000/ml in 620 ml. One day after treatment, two holes were made in each plot about 7 cm apart and equidistant from the squash plants. Five ml of egg suspension was pipetted into each hole, which was then sealed and the plot lightly watered. A total of 90,000 vermiform eggs were added to each plot.

(90) Early Observations

(91) Two days after treatment slight phytotoxicity was seen with the 4417 2 kg/ha treatment. The hypocotyls of affected seedlings were water-soaked at the soil line. Leaf diameter of the first true leaves measured five days after treatment (5 DAT) also showed a slight reduction for the 4417 treatment. None of the compounds appeared to affect the onset of bloom.

(92) TABLE-US-00016 TABLE 7B Root Ratings Name Analog TW28 RG28 RW28 RG43 RW43 4417 97.5 cde 25.5 abc 5.3 ab 38.2 bc 16.4 a 4776 250.8 abc 19.0 bc 7.5 ab 26.8 cd 13.0 ab 4559 150.8 cd 34.2 ab 6.4 ab 24.0 cde 13.2 ab Oxamyl5 232.1 abcd 10.5 c 6.6 ab 17.3 def 12.7 b Oxamyl2 136.8 d 45.0 a 5.6 b 42.3 ab 14.3 ab NT 322.4 a 38.6 a 7.9 a 54.7 a 15.5 ab NI 263.1 ab 0.4 c 6.9 ab 0.0 g 14.4 ab *Means with a letter in common are not significantly different at P = 0.1 using Students T test. TW28 = top weight at 28 days after treatment RG28 = % root galls at 28 days after treatment TW28 = root weight at 28 days after treatment RG43 = % root galls at 43 days after treatment TW43 = root weight at 43 days after treatment NT = non treated (i.e., inoculated with nematodes but not treated with chemicals) NI = non inoculated (i.e., not treated with chemicals or inoculated with nematodes) Oxamyl5 and Oxamyl2 are oxamyl at 5 kg ai/ha and 2 kg ai/ha respectively

(93) The first root evaluation was at 28 DAT. The tops were cut off and weighed immediately in the field, and the roots were carefully dug out so as not to disturb the remaining plant.

(94) The earlier phytotoxicity seen with 4417 is mirrored in a reduction in top weights at 28 DAT. However root weights 28 DAT were not affected and roots weights recorded from the second harvest (43 DAT) revealed no effect from any of the treatments.

(95) Root gall damage was estimated at 28 DAT and 43 DAT using a percent binning scale of 0, 1, 5, 10, 25, 33, 50, 66, 75, 90, and 100% representing the % of root mass significantly impacted by galls. At both sampling times all three compounds provided control of root galling that was numerically superior to oxamyl at an equivalent rate. 4776 was statistically better than oxamyl at both the 28 day and the 43 day time point whereas 4559 was significantly better than oxamyl at the 43 day time point.

(96) In summary all three compounds provide equivalent or superior nematode control to oxamyl under field conditions. Thus these nematicidal analogs are superior to many of the newer more selective nematicide candidates which lack field efficacy at reasonable use rates and are lack sufficient longevity to be of commercial interest.

Example 8

Summer Nematicidal Field Evaluation of Pre-plant Incorporated (PPI) Compounds for Control of Meloidogyne incognita on Squash

(97) Test plots of 33 cm diameter holes were bored 41 cm deep into clay soil and filled with a mixture of 80% sand and 20% silt loam soil. Technical compound for each treatment was dissolved in 50 ml acetone containing 250 ul of Triton X-100 surfactant. This solution was added to 450 ml water and poured onto 95 liters of sand/soil mixture in a rotating drum mixer. While continuing to rotate the mixing drum 66 grams of chopped, galled, tomato roots was added and thoroughly distributed. The treated soil was sufficient to fill the top 15 cm of each of the 6 replicate plots, thus simulating a PPI treatment. The plots were then watered lightly and a mixture of M. incognita eggs and larvae were injected 5 cm deep at 5 points within the plot (100 k eggs/larvae in 10 ml per plot). Three-week old squash (cv. Liberator III) with 1 fully expanded true leaf was planted 4 days after soil treatment, one per plot.

(98) TABLE-US-00017 feeder root 0-3 vigor 0-3 vigor root wgt (g) top wgt (lbs) total fruit gall % (3 = ave) 16DAP 21DAP 31DAP 31DAP (lbs) 31DAP 31DAP 5523 4 kg 3.0 3.0 26.3 1.31 1.24 26 3.0 5823 4 kg 3.0 3.0 22.6 1.45 1.44 3 2.7 5891 4 kg 3.0 2.8 27.5 1.43 1.22 28 3.0 5938 4 kg 2.5 2.7 24.1 1.60 1.22 9 2.7 5960 4 kg 3.0 3.0 32.6 1.58 1.61 24 3.3 fosthiazate 2 kg 3.0 3.0 26.4 2.01 1.25 5 2.3 oxamyl 4 kg 2.7 2.5 37.0 1.16 1.09 85 3.0 Blank 1.5 1.2 23.4 0.30 0.38 90 2.7

(99) Chopped gall inoculum combined with eggs/juveniles provided high pressure and rapid development of symptoms. PPI applications of DC5823 and DC5938 provided excellent control at 4 kg/ha. DC5523, DC5891, and DC5960 also provided significant control at 4 kg/ha.

Example 9

Seed Treatment Test of Root Knot Nematode on Cucumber Plants and Soybean Cyst Nematode on Soybean Plants

(100) For a given concentration the chemical is dissolved in 500 ul of acetone and one gram of cucumber seed (RKN test) or soybean seed (SCN test) is added (e.g., 20 mg active ingredient in 500 ul acetone plus 1 gram of seed). The seed solutions are agitated until all seeds were thoroughly covered with the chemical solution. The acetone is then allowed to evaporate by air drying the seeds. The seeds are planted in 2-inch pots containing sandy soil and then the pots are inoculated with 1000 Meloidogyne incognita (RKN) or 1000 Heterodera glycines (SCN) eggs per pot three days after planting. Plants are rated for galling 14 days after egg inoculation for RKN or 28 days after egg inoculation for SCN.

(101) TABLE-US-00018 TABLE 9A Seed treatment activity against root knot nematode using cucumber seeds 20 mg ai/gram Name Analog seed* 1822 76 4775 77 4776 58 Abamectin.sup.# 84 *Data shows percent control (i.e., galling reduction) relative to the control blank treatment. .sup.#Abamectin positive control at 10 mg ai/gram seed.

(102) TABLE-US-00019 TABLE 9B Seed treatment activity against soybean cyst nematode using soybean seeds 1.5 0.375 Name Analog mg* mg* 5527 0 71.sup.a 43.sup.a 5479 88.sup.a 83.sup.b 67.sup.a 69.sup.b 1822 70.sup.a 58.sup.a 5847 80.sup.b 66.sup.b 5878 77.sup.b 43.sup.b 5953 77.sup.b 44.sup.b Oxamyl 71.sup.b 4.sup.b Thiodi- 23.sup.a 6.sup.a carb Aba- 24.sup.a 14.sup.a mectin *Data shows percent cyst reduction relative to control blank treatment. Rates are mg ai/gram seed. Data with the same letters are taken from the same test.

(103) Oxadiazole, thiadiazole and oxazole analogs are versatile nematicides showing activity as seed treatments in addition to drench applications and soil pre-incorporation methods.

Example 10

The Claimed Structures do not Induce an Apoptosis Marker in Mammalian Cells and do not Kill Nematodes by Causing Apoptosis

(104) Previous studies have shown that induction of the pro-apoptotic caspase-3 protease through the cleavage of specific fluorogenic substrates is a reliable method of measuring the induction of apoptosis, and certain chloro and bromo substituted thiophene and furan oxadiazoles were identified after high-throughput screening for caspase-3 induction in mammalian cells (Zhang H Z, Kasibhatla S, Kuemmerle J, Kemnitzer W, Ollis-Mason K, Qiu L, Crogan-Grundy C, Tseng B, Drewe J, Cai S X. Discovery and structure-activity relationship of 3-aryl-5-aryl-1,2,4-oxadiazoles as a new series of apoptosis inducers and potential anticancer agents. J Med Chem. 2005 48(16):5215-23).

(105) To evaluate whether the compound classes of this invention are able to induce apoptosis, caspase-3 activity was determined after compound exposure in rat hepatoma derived H4IIE cells using a caspase substrate (DEVD, Asp-Glu-Val-Asp) labeled with a fluorescent molecule, 7-Amino-4-methylcoumarin (AMC). Caspase 3 cleaves the tetrapeptide between D and AMC, thus releasing the fluorogenic green AMC. Following the test article exposure to cells in 96-well plates, medium was aspirated from plates and PBS added to each well. Plates were stored at 80 C. to lyse cells and store samples until further analysis. On the day of analysis, plates were removed from freezer and thawed. Caspase buffer with fluorescent substrate was added to each well and incubated at room temperature for one hour. AMC release was measured in a spectrofluorometer at an excitation wavelength of 360 nm and an emission wavelength of 460 nm. Values are expressed as relative fluorescent units (RFU). In contrast to paclitaxel, camptothecin, and staurosporine, which were reportedly capable of inducing apoptosis in a variety of cell lines at or below doses of 1 M doses, no induction of caspase-3 is observed for DC1822, DC5823, DC5915, and DC5938 at concentrations up to 300 M in this system.

(106) To confirm that these compounds do not affect nematodes by induction of apoptosis, Caenorhabditis elegans mutants defective in the apoptotic pathway, ced-3(n717) and ced-4(N1162) mutants (Ellis H M, Horvitz H R. Genetic control of programmed cell death in the nematode C. elegans. 1986 Cell 44:817-829), were evaluated for susceptibility to 10 g/ml DC5823 on NGM agar plates. No observable phenotypic difference in susceptibility between the wild-type C. elegans strain (N2 Bristol) and the ced-3 and ced-4 mutants were observed, including time to mortality.

(107) These data indicate that the claimed structures do not affect apoptosis in either mammalian cells or nematodes.

Example 11

Description of Synthesis of the Compounds of the Formula I to VII

(108) The compounds of this invention of the Formulas I to VII may be prepared using methods known to those skilled in the art. Specifically, the compounds of this invention with Formulae Ia and Ib can be prepared as illustrated by the exemplary reaction in Scheme 1. The alpha aminoketones 3 are prepared from the acetophenones 1 in a two-step procedure that involves bromination with 4(-dimethylamino)pyridine tribromide and subsequent amination of the bromide intermediate 2 with sodium diformylamide. The aminoketone 3 is then reacted with an appropriate acyl chloride 4 to yield the acylaminoketone 5. A cyclization of the linear precursor 5 to the 2,5-disubstituted-1,3-oxazole analog 6 is accomplished with phosporousoxychloride in DMF in good yields.

(109) ##STR00186##

(110) Specifically, the compounds of this invention with Formulae IIa and IIb can be prepared as illustrated by the exemplary reaction in Scheme 2. The alpha aminoketone 2 is prepared from the bromide precursor 1 by amination with sodium diformylamide and then reacted with acyl chloride 3 to yield the acylaminoketone 4. A cyclization of the linear precursor 4 to the 2,5-disubstituted-1,3-oxazole analog 5 is accomplished with phosporousoxychloride in DMF in good yields.

(111) ##STR00187##

(112) Specifically, the compounds of this invention with Formulae Ma and III b can be prepared as illustrated by the exemplary reaction in Scheme 3. The benzohydrazide 1 is reacted with the acyl chloride 2 in chloroform in the presence of trietylamine (TEA) at ambient temperature to give acyl benzohydrazide 3. A cyclization of the diacylhydrazine 3 to the 2,5-disubstituted-1,3,4-oxadizaole compound 4 is accomplished with phosporouschloride (POCl3) in DMF.

(113) ##STR00188##

(114) Specifically, the compounds of this invention with Formulae IVa and IVb can be prepared as illustrated by the exemplary reaction in Scheme 4. The benzonitrle 1 is converted to the corresponding hydroxyimidate 2 when reacted with hydroxylamine hydrochloride in the presence of DIEA in methanol at room temperature overnight. Then the benzohydroxyimidate 2 is acylated with an appropriate furan or thiophene carbonyl chloride (R2-COY) in the presence of pyridine, followed with DCC dehydration to give the 3,5-disubstituted-1,2,4-oxadiazole product.

(115) ##STR00189##

(116) Specifically, the compounds of this invention with Formulae Va and Vb can be prepared as illustrated by the exemplary reaction in Scheme 5.

(117) ##STR00190##

(118) First, the appropriate analog of furan or thiophene nitrile 1 is converted to the corresponding hydroxyimidate 2 by reacting with hydroxylamine in methanol in the presence of DIEA. Then, the intermediate 2 is reacted with the appropriately substituted benzoyl chloride 3 in pyridine-dioxnae to give the desired 3,5-disubstituted-1,2,4-oxadiazole product 4.

(119) Specifically, the compounds of this invention with Formulae VIa and VIb can be prepared as illustrated by the exemplary reaction in Scheme 6. The synthesis starts with the reaction of an appropriate benzamide substrate 1 with chlorocarbonylsulfenyl chloride to yield the oxathiazolone compound 2. In the next step the oxathiazoline intermediate 2 is reacted with an appropriate furan or thiophene nitrile in toluene under microwave conditions to give the desired 3,5-disubstituted-1,2,4-thiadiazole product 3.

(120) ##STR00191##

(121) Specifically, the compounds of this invention with Formulae VIIa and VIIb can be prepared as illustrated by the exemplary reaction in Scheme 7. An appropriate furan or thiophene carboxamide substrate 1 is converted to the oxathiazolone intermediate by reacting with chlorocarbonylsulfenyl chloride. Then, the oxathiazoline intermediate 2 is reacted with an appropriate benzonitrile compound in toluene under microwave conditions to give the desired 3,5-disubstituted-1,2,4-thiadiazole product 4.

(122) ##STR00192##

Formula Ia Example

5-(4-chloro-2-fluorophenyl)-2-(thiophen-2-yl)oxazole

(123) A mixture of 4-chloro-2-fluoroacetophenone (17.5 g, 100 mmol), 4-(dimethylamino)pyridine tribromide (40.0 g, 110 mmol) and acetic acid (100 mL) was stirred at room temperature for 24 h. Water (150 mL) was added and after stirring for 30 min the precipitated solid was collected by filtration, washed with water, and dried in vacuo to give the desired bromide intermediate as a white solid (24 g, 95%).

(124) To a solution of the bromide compound (24 g, 90 mmol) in acetonitrile (300 mL) was added sodium diformylamide (9.0 g, 95 mmol). The mixture was heated to reflux for 2 h and cooled to r.t overnight. The mixture was filtered to remove NaBr. The filtrate was concentrated to give diformylamide intermediate as a brown oil, 23.6 g. EtOH (300 mL) and 30% HCl (90 mL) were added and the mixture was stirred at 50 C. for 5 h and cooled to room temperature overnight, during which time the product crystallized out. The solid was collected by filtration, washed with dichloromethane, and dried to constant weight to give the desired aminoketone hydrochloride as white solid (6.3 g, 31%). that was sued as is in the next step.

(125) The synthesis of acylamino ketone was performed as described in the literature (J. Med. Chem. 1986, 29, 333-341). A suspension of 2-amino-1-(4-chloro-2-fluorophenyl)ethanone hydrochloride (6.3 g, 28 mmol) in water (50 mL) and EtOAc (100 mL) was cooled in an ice-bath. NaHCO.sub.3 (11.9 g, 140 mmol) was added in portions, followed by 2-thiophene carbonyl chloride (4.25 g, 29 mmol). The mixture was stirred at room temperature for 16 h. Water (50 mL) was added and the mixture was extracted with EtOAc (250 mL). The organic layers were combined, washed with brine, dried (MgSO.sub.4), filtered, and concentrated in vacuo to give acylamino ketone 5 as yellow solid (7.7 g, 92%). The organic layers were combined, dried (MgSO.sub.4), and concentrated in vacuo to give crude product, 7.8 g, which was purified by crystallization from EtOH (25 mL). Yield 5.0 g (69%) of yellow solid.

(126) Molecular Formula: C.sub.13H.sub.7ClFNOS; MW 279.72.

(127) HPLC-ESMS: t.sub.R=6.04 min; m/z: 279.9 (M+H); HPLC purity 98.0% (216 nm); 99% (250 nm)

(128) .sup.1H-NMR (300 MHz, CDCl.sub.3): 7.74-7.85 (m, 2H), 7.52-7.56 (m, 1H), 7.46-7.51 (m, 1H), 7.21-7.31 (m, 2H), 7.14-7.20 (m, 1H)

Formula IIa Example

2-(4-chloro-2-fluorophenyl)-5-(thiophen-2-yl) oxazole

(129) A mixture of 2-(2-bromoacetyl)thiophene (2.05 g, 10 mmol), sodium diformyl amide (1.05 g, 11 mmol) and acetonitrile (20 mL) was heated to reflux for 4 h. The mixture was cooled to r.t. and filtered to remove NaBr. The filtrate was concentrated in vacuo to give a brown oil, 2.0 g. EtOH 930 mL) was added followed by concentrated HCl (30%, 10 mL). The mixture was stirred at r.t. overnight. Concentration in vacuo gave a sticky solid, 2.1 g. The resulted aminoketone hydrochloride was contaminated by some NH.sub.4Cl (based on H1-NMR spectra) and used as is in the next step.

(130) A mixture of the crude amine.HCl in EtOAc (40 mL) and water (20 mL) was vigorously stirred and cooled in ice-water bath. NaHCO.sub.3 (8.3 g, 100 mmol) was added, followed by 4-chloro-2-fluorobenzoyl chloride (1.9 g, 10 mmol). The mixture was stirred at r.t. overnight. The layers were separated. The water layer was extracted with EtOAc (50 mL). The combined organic layers were washed with water, dried (MgSO.sub.4) and concentrated to a brown solid, 2.0 g. The resulted crude product was a mixture of the desired acylaminoketone and 4-chloro-2-fluorobenzamide (formed by reaction of ammonium chloride present in the starting aminoketo compound with the acyl chloride).

(131) The acylaminoketone intermediate was dissolved n DMF (25 mL). and then POCl.sub.3 (2.3 g, 15 mmol) was added and the mixture was stirred at r.t. for 2.5 days. Ice-water was added and the mixture was extracted with EtOAc (350 mL). The organic layer was washed with water (330 ml), dried (MgSO.sub.4) and concentrated to a brown solid/oil, 1.7 g. A column chromatography (Hep/EtOAc 2/1) gave 1.0 g of a solid which was still not pure. Crystallization from MeOH (5 mL) gave pure (0.6 g, 22%) 2-(4-chloro-2-fluorophenyl)-5-(thiophen-2-yl)oxazole with HPLC purity >99.0% (215 and 254 nm).

(132) Molecular Formula: C.sub.13H.sub.7ClFNOS, MW 279.72; LC-MS: t.sub.R=9.46 min m/z: 279.9 (M+H).

(133) .sup.1H-NMR (300 MHz, CDCl.sub.3): 7.98-8.08 (m, 1H), 7.22-7.42 (m, 5H), 7.08-7.14 (m, 1H)

Formula IIIa Example

2-(4-Chloro-phenyl)-5-thiophen-2-yl-[1,3,4]oxadiazole

(134) To 250 mL round bottom flask was added 2.0 g (11.7 mmol, 1 eq) of 4-chlorobenzhydrazide (1) in 100 mL of amelene stabilized chloroform, followed by addition of 4 mL (29.25 mmol, 2.5 eq) of TEA. Then, 1.4 mL (12.87 mmol, 1.1 eq) of 2-thiophenecarbonyl chloride (2) was added drop-wise and the mixture was stirred at ambient temperature for 1 h. Reaction progress was monitored by LCMS on a twelve minute gradient. The formed white precipitate was filtered, washed with chloroform and then dried on the high vacuum for two hours. The resulting material was confirmed to be the desired diacylhydrazide and was used in the next step without further purification. The crude diacyl-hydrazide was dissolved in 60 mL of POCl.sub.3 under heating. The resulting mixture was then heated under reflux in oil bath (100-110 C.) for 5-7 h. The reaction progress was monitored by LCMS on a twelve minute gradient. Once the cyclization reaction was completed as determined by LCMS, POCl.sub.3 was carefully evaporated in vacuum and the reaction was then neutralized with a 1 N solution of ammonium hydroxide. The product was extracted with ethyl acetate (300 mL) from saturated solution of NaHCO.sub.3 (200 mL), washed with a brine (2200 mL), then dried over sodium sulfate, filtered and evaporated to dryness. The product was purified by flash column chromatography (hexane.fwdarw.12% ethyl acetate/hexane), and then recrystallized from mixture of hexane/ethyl acetate (5:1) to give 1.3 g of the desired compound 2-(4-Chloro-phenyl)-5-thiophen-2-yl-[1,3,4]oxadiazole (42%) as a white solid.

(135) Chemical Formula: C.sub.12H.sub.7ClN.sub.2OS; MW 262.71; ESMS: m/z 263 (M+H);

(136) .sup.1H-NMR (250 MHz, D.sub.6-DMSO): 8.08-8.12 (m, 2H), 7.96-7.99 (m, 2H), 7.69-7.72 (m, 2H), 7.32-7.35 (m, 1H)

Formula IVa Example

3-(4-Chloro-2-methyl-phenyl)-5-furan-2-yl-[1,2,4-]oxadiazole

(137) In a 500 mL round-bottom flask, 4-chloro-2-methylbenzonitrile (10 g, 66 mmol) was dissolved in 200 mL of methanol. To the mixture was added hydroxylammonium chloride (4.56 g, 66 mmol) followed by DIEA (diisopropylethylamine) (23 mL, 132 mmol). The mixture was heated at reflux for overnight. The solvents were removed. The residue was dissolved in 200 mL of CHCl.sub.3. To the mixture was added 2-furoyl chloride (10.5 ml, 66 mmol) followed by DIEA (23 mL, 132 mmol). After reaction completion, the mixture was extracted with chloroform and water. The organic layer was separated, washed with brine, dried over Na.sub.2SO.sub.4, filtered and evaporated to dryness. The residue was dissolved in 200 mL of dioxanes. To the mixture was added 1 eq of DIC (N, N-diisopropylcarbodiimide) followed by 1 eq of DIEA. The mixture was then heated at reflux overnight. After reaction completion, the mixture was cooled down. The solvents were removed in vacuo. The residue was then extracted with ethyl acetate and water. The organic layer was separated, washed with brine, dried over Na.sub.2SO.sub.4, filtered and evaporated to dryness. The crude was purified by flash chromatography on silica gel in a 0-20% ethyl acetate/hexanes gradient to afford 4.96 g of the desired compound 3-(4-Chloro-2-methyl-phenyl)-5-furan-2-yl-[1,2,4-]oxadiazole as a white powder in an overall yield of 28.8%.

(138) Molecular Formula: C.sub.13H.sub.9ClN.sub.2O.sub.2; MW 260.04; HPLC purity 99.9% (254 nm); LC-ESMS: t.sub.R=7.55 min; m/z 261.1 (M+1);

(139) .sup.1H-NMR (250 MHz, D.sub.6-DMSO): 8.18-8.19 (m, 1H), 7.98-8.01 (d, J=8.3, 1H), 7.64-7.65 (m, 1H), 7.52-7.56 (m, 1H), 7.46-7.50 (m, 1H), 6.87-6.89 (m, 1H), 2.59 (s, 3H)

Formula IVa Example

3-(4-Bromo-2-methyl-phenyl)-5-furan-2-yl-[1,2,4]-oxadiazole

(140) In a 500 mL round-bottom flask, 4-bromo-2-methylbenzonitrile (5 g, 25 mmol) was dissolved in 200 mL of methanol. To the mixture was added hydroxylammonium chloride (1.72 g, 25 mmol) followed by DIEA (diisopropylethylamine) (8.7 mL, 50 mmol). The mixture was heated at reflux for overnight. The solvents were removed. The residue was dissolved in 200 mL of CHCl.sub.3. To the mixture was added 2-furoyl chloride (3.97 ml, 25 mmol) followed by DIEA (8.7 mL, 50 mmol). After reaction completion, the mixture was extracted with chloroform and water. The organic layer was separated, washed with brine, dried over Na.sub.2SO.sub.4, filtered and evaporated to dryness. The residue was dissolved in 200 mL of dioxanes. To the mixture was added 1 eq of DIC (N, N-diisopropylcarbodiimide) followed by 1 eq of DIEA. The mixture was then heated at reflux overnight. After reaction completion, the mixture was cooled down. The solvents were removed in vacuo. The residue was then extracted with ethyl acetate and water. The organic layer was separated, washed with brine, dried over Na.sub.2SO.sub.4, filtered and evaporated to dryness. The crude was purified by flash chromatography on silica gel in a 0-20% ethyl acetate/hexanes gradient to afford 2.23 g of the desired compound 3-(4-Bromo-2-methyl-phenyl)-5-furan-2-yl-[1,2,4]-oxadiazole as a white powder in an overall yield of 36%.

(141) Chemical Formula: C.sub.13H.sub.9BrN.sub.2O.sub.2; MW: 305.13; HPLC Purity >99.0%; (254 nm) ESMS: t.sub.R=7.81 min; m/z 305.1 (M+.sup.1);

(142) .sup.1H-NMR (250 MHz, D.sub.6-DMSO): 8.18-8.19 (m, 1 H), 7.92 (d, J=8.3, 1H), 7.58-7.70 (m, 3H), 6.86-6.90 (m, 1H), 2.59 (s, 3H)

Formula Va Example

5-(4-chloro-2-methylphenyl)-3-(furan-2-yl)-1,2,4-oxadiazole

(143) To a solution of 2-furonitrile (1.9 g, 20 mmol) in MeOH (50 mL) was added hydroxylamine hydrochloride (1.4 g, 20 mmol) and triethylamine (2.1 g, 20 mmol). The mixture was heated to reflux overnight. After cooling to room temperature the mixture was concentrated in vacuo. The residue was stirred with EtOAc (50 mL). The solid was filtered off and the filtrate was concentrated to a thick oil, 2.5 g (99%). The H-NMR spectra was in accordance with the desired hydroxyamidine compound which was contaminated with Et.sub.3N.HCl. The crude product resulted in this reaction was used without the purification in the next step.

(144) To a suspension of 4-chloro-2-methylbenzoic acid (3.4 g, 20 mmol) in dichloromethane (50 mL) was added one drop of DMF followed by oxalylchloride (3.2 g, 25 mmol). The mixture was stirred overnight during which time all solid dissolved. The mixture was concentrated in vacuo and stripped with dichloromethane to remove excess oxalylchloride. The residual acid chloride was taken in dioxane/pyridine (10/1, 55 mL) and hydroxyamidine compound (2.5 g, 20 mmol) was added. The mixture was heated to reflux for 3 h. After cooling to room temperature, water was added (100 mL) and the resulting solid was collected by filtration and dried to give 6.2 g of crude product. Recrystallizaton from MeOH (40 mL) gave pure 5-(4-chloro-2-methylphenyl)-3-(furan-2-yl)-1,2,4-oxadiazole 2.6 g (yield 47%).

(145) Molecular Formula: C.sub.13H.sub.9ClN.sub.2O2; MW 260.04; HPLC purity: >99.9% (216 nm); 99.9% (324 nm); LC-ESMS: t.sub.R=9.46 min; m/z 261.1 (M+1);

(146) .sup.1H-NMR (300 MHz, CDCl.sub.3): 8.10 (dd, J=8.1, 1H), 7.63-7.66 (m, 1H), 7.32-7.42 (m, 2H), 7.18-7.22 (d d, J=2.7, 0.9, 1H), 6.58-6.62 (m, 1H), 2.89 (s, 3H)

Formula VIa Example

(2,4-dichlorophenyl)-5-(furan-2-yl)-1,2,4-thiadiazole

(147) A mixture of 2,4-dichlorobenzamide (25 g, 131.5 mmol) and chlorocarbonylsulfenylchloride (19 g, 145 mmol) in toluene (150 mL) was heated to reflux for 4 h (HCl-gas formation was observed with pH paper). After cooling to r.t. the mixture was concentrated in vacuo to give the desired oxathiazolone compound as an off-white solid (32.4 g, 99%). that was used in the next without purification. In a 20 mL vial a mixture of oxathiazolone 8a (2 g, 8 mmol) and 2-furonitrile (10 g, 107 mmol) was heated in the microwave at 190 C. for 20 min. The reaction was performed 10 times and the combined mixture was distilled (Kugerrohr) at 100 C./20 mbar to remove excess 2-furonitrile (the recovered 2-furonitrile was used again). The mixture was further distilled at 150 C./10 mbar to remove the byproduct nitrile 10 (yellow solid, 6.5 g, 47%). The residue of the distillation (circa 10 g) was taken in dichloromethane (50 mL), filtered and the filtrate concentrated to a brown solid, 8 g. Recrystallization by dissolution in hot MeOH (50 mL) and addition of water (10 mL) gave pure (2,4-dichlorophenyl)-5-(furan-2-yl)-1,2,4-thiadiazole as brown solid, 4.7 g, in a 20% yield.

(148) Chemical Formula: C.sub.12H.sub.6Cl.sub.2N.sub.2OS; MW: 297.16; HPLC-ESMS: t.sub.R=6.5; m/z: 296.96; 298.95 (M+1); HPLC purity >99% (221 nm), >99% (263 nm), >99.0% (306 nm)

(149) .sup.1H-NMR (300 MHz, CDCl.sub.3): 7.90 (dd, J=8.4, 1H), 7.57-7.58 (m, 1H), 7.29 (dd, J=8.4, 1.8) 7,48, d, J=1.8, 1H), 7.15-7.20 (m, 1H), 6.55-6.59 (m, 1H)

Formula VIa Example

3-(4-chloro-2-methylphenyl)-5-(furan-2-yl)-1,2,4-thiadiazole

(150) A magnetically stirred mixture of acid 4-chloro-2-methylbenzoic acid (50 g, 0.29 mol), dichloromethane (200 mL), and 0.5 mL DMF was cooled in an ice-bath. The cooler was connected to a gas absorption trap. Oxalyl chloride (44.5 g, 0.35 mmol) was added dropwise in 1 h. The mixture was stirred at r.t. overnight during which time all solid dissolved. The solution was concentrated in vacuo and stripped with dichloromethane to remove excess oxalyl chloride. The residue was taken in THF (200 mL) and mechanically stirred in an ice-water bath. Aqueous 25% ammonia (100 mL) was added in 15 min, which resulted in the formation of a precipitate. The THF was removed with the rotavap and extra water (100 mL) was added. The suspension was stirred at r.t. overnight. The solid was collected by filtration and dried in vacuo to give 2-methyl-4-chlorobenzamide (43.7 g, yield 89%) that was used without purification in the next step.

(151) A mechanically stirred mixture of 2-methyl-4-chlorobenzamide (31.35 g, 185 mmol), toluene (400 mL), and chlorocarbonylsulfenylchloride (25 g, 190 mmol) was heated to reflux for 3 h. After cooling to room temperature the mixture was concentrated in vacuo to give a yellow solid 40 g (95%). H-NMR showed that this was a mixture of the desired oxathiazolone compound and nitrie by-product and starting amide in a ratio 85:10:5. This mixture was used in the next step without further purification.

(152) The crude oxathiazolone compound (2.0 g, 8.8 mmol) and 2-furonitrile (16 g, 170 mmol) were mixed and heated for 20 min at 190 C. in the microwave. Ten batches were combined and Kugelrohr distilled at 100 C./30 mbar to recover excess 2-furonitrile (used again in next microwave reactions). The residue was further distilled at 150 C./20 mbar to remove the nitrile by-product. The residue, 5.5 g was combined with the residue of another ten microwave reactions (4.5 g) and purified by column chromatography. The resulting 4.5 g (85% pure by HPLC) was recrystallized from MeOH (50 mL) to give pure 3-(4-chloro-2-methylphenyl)-5-(furan-2-yl)-1,2,4-thiadiazole as light brown solid, 3.6 g (7.5% yield).

(153) Chemical Formula: C.sub.13H.sub.9ClN.sub.2OS; MW: 278.7; HPLC-ESMS: t.sub.R=6.36 min and m/z 277.0 (M+1); HPLC purity: >95% (220 nm) 95% (270 nm).

(154) .sup.1-H-NMR (300 MHz, CDCl.sub.3): 8.06, (dd, J=7.8, 1H), 7.62-7.63 (m, 1H), 7.22-7.31 (m, 3H), 6.61-6.63 (m, 1H), 2.66 (s, 3H)

Formula VIa Example

3-(4-chlorophenyl)-5-(furan-2-yl)-1,2,4-thiadiazole

(155) A mechanically stirred mixture of 4-chlorobenzamide (20.23 g, 130 mmol), toluene (150 mL), and chlorocarbonylsulfenylchloride (19 g, 145 mmol) was heated to reflux for 3 h. After cooling to r.t. the mixture was concentrated in vacuo to give a yellow solid foam, 27.65 g (100%). H-NMR showed that this was almost pure oxathiazolone compound that was used as is in the next step. The oxathiazolone compound (1.71 g, 8 mmol) and 2-furonitrile (15 g, 160 mmol) were mixed and heated for 20 min at 190 C. in the microwave. Ten batches were combined and Kugelrohr distilled at 100 C./30 mbar to recover excess 2-furonitrile (used again in next microwave reactions). The residue was further distilled at 150 C./20 mbar to remove the nitrile byproduct. The residue, 5 g, was recrystallized from MeOH to give 3.5 g of solid. This was combined with the residue of another 5 microwave reactions (2.6 g) and purified by column chromatography. The resulting 4.4 g (90% pure by HPLC) was recrystallized from Heptane/EtOAc=7/1 (50 mL) to give pure 3-(4-chlorophenyl)-5-(furan-2-yl)-1,2,4-thiadiazole as light brown solid, 3.35 g (10% yield).

(156) Chemical Formula: C.sub.12H.sub.7ClN.sub.2OS; Molecular Weight: 262.71; HPLC-ESMS: t.sub.R=6.06 min; m/z: 263.00, 264.99 (M+1)

(157) .sup.1H-NMR (300 MHz, CDCl.sub.3): 8.24-8.33 (m, 2H), 7.63-7.65 (m, 1H), 7.42-7.50 (m, 2H), 7.23-7.28 (m, 1H), 6.62-64 (m, 1H)

Formula VIIa Example

5-(2-chloro-4-methylphenyl)-3-(furan-2-yl)-1,2,4-thiadiazole

(158) A magnetically stirred mixture of 2-furoylamide (prepared from 2-furoylchloride and aqueous ammonia, 1.13 g, 10 mmol) and chlorocarbonylsulfenylchloride (2.0 g, 15 mol) in toluene (20 mL) was heated to reflux for 4 h. After cooling to room temperature the mixture was concentrated to give 1.7 g of the desired oxathiazolone as a yellow solid (almost in a quantitative yield) that was used in the next step without further purification.

(159) A mixture of the oxathiazolone compound (170 mg, 1 mmol) and 4-chloro-2-methylbenzonitrile (3.03 g, 20 mmol) was heated in the microwave at 190 C. for 20 min. A second reaction was performed and the mixtures were combined. Excess of the nitrile by-product (furonitrile) were removed in vacuo (120 C., 0.3 mbar). The residual brown solid (100 mg) was taken in hot MeOH (10 mL) and decanted from insolable material (presumably sulphur). The MeOH solution was left at room temperature overnight. The precipitated solid was collected and dried to give compound 5-(2-chloro-4-methylphenyl)-3-(furan-2-yl)-1,2,4-thiadiazole as brown solid, 40 mg (7%). NMR conform structure.

(160) Chemical Formula: C.sub.13H.sub.9ClN.sub.2OS; MW: 278.7; HPLC-ESMS: t.sub.R=6.36 min and m/z 277.01 (M+1); HPLC purity: 93.5 (216 nm) 91% (324 nm); .sup.1H-NMR (300 MHz, CDCl.sub.3): 7.87 (dd, J=8.1, 1H), 7.51-7.60 (m, 1H), 7.24-7.32 (m, 2H), 7.15-7.20 (m, 1H), 6.50-6.56 (m, 1H), 2.58 (s, 3H)