WO2022078533A1 - Substituted 1,3-diphenylurea derivatives and 1-phenyl-3-pyridylurea derivatives for plant biotechnology, preparations containing these compounds and use thereof - Google Patents

Substituted 1,3-diphenylurea derivatives and 1-phenyl-3-pyridylurea derivatives for plant biotechnology, preparations containing these compounds and use thereof Download PDF

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WO2022078533A1
WO2022078533A1 PCT/CZ2021/050107 CZ2021050107W WO2022078533A1 WO 2022078533 A1 WO2022078533 A1 WO 2022078533A1 CZ 2021050107 W CZ2021050107 W CZ 2021050107W WO 2022078533 A1 WO2022078533 A1 WO 2022078533A1
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urea
phenyl
group
halogen
general formula
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PCT/CZ2021/050107
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French (fr)
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Jaroslav NISLER
Miroslav Strnad
Lukas Spichal
Stefaan Werbrouck
Nino MURVANIDZE
Alena KADLECOVA
Danuse TARKOWSKA
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Ustav Experimentalni Botaniky Av Cr, V.V.I.
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D213/72—Nitrogen atoms
    • C07D213/75—Amino or imino radicals, acylated by carboxylic or carbonic acids, or by sulfur or nitrogen analogues thereof, e.g. carbamates
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N37/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids
    • A01N37/18—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing the group —CO—N<, e.g. carboxylic acid amides or imides; Thio analogues thereof
    • A01N37/28—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom having three bonds to hetero atoms with at the most two bonds to halogen, e.g. carboxylic acids containing the group —CO—N<, e.g. carboxylic acid amides or imides; Thio analogues thereof containing the group; Thio analogues thereof
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N47/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid
    • A01N47/08—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid the carbon atom having one or more single bonds to nitrogen atoms
    • A01N47/28—Ureas or thioureas containing the groups >N—CO—N< or >N—CS—N<
    • A01N47/30—Derivatives containing the group >N—CO—N aryl or >N—CS—N—aryl
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N47/00—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid
    • A01N47/08—Biocides, pest repellants or attractants, or plant growth regulators containing organic compounds containing a carbon atom not being member of a ring and having no bond to a carbon or hydrogen atom, e.g. derivatives of carbonic acid the carbon atom having one or more single bonds to nitrogen atoms
    • A01N47/28—Ureas or thioureas containing the groups >N—CO—N< or >N—CS—N<
    • A01N47/36—Ureas or thioureas containing the groups >N—CO—N< or >N—CS—N< containing the group >N—CO—N< directly attached to at least one heterocyclic ring; Thio analogues thereof
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
    • A01P21/00—Plant growth regulators
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01P—BIOCIDAL, PEST REPELLANT, PEST ATTRACTANT OR PLANT GROWTH REGULATORY ACTIVITY OF CHEMICAL COMPOUNDS OR PREPARATIONS
    • A01P5/00—Nematocides
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C275/00—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups
    • C07C275/28—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C275/00—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups
    • C07C275/28—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton
    • C07C275/30—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton being further substituted by halogen atoms, or by nitro or nitroso groups
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C275/00—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups
    • C07C275/28—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton
    • C07C275/32—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton being further substituted by singly-bound oxygen atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C275/00—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups
    • C07C275/28—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton
    • C07C275/32—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton being further substituted by singly-bound oxygen atoms
    • C07C275/34—Derivatives of urea, i.e. compounds containing any of the groups, the nitrogen atoms not being part of nitro or nitroso groups having nitrogen atoms of urea groups bound to carbon atoms of six-membered aromatic rings of a carbon skeleton being further substituted by singly-bound oxygen atoms having nitrogen atoms of urea groups and singly-bound oxygen atoms bound to carbon atoms of the same non-condensed six-membered aromatic ring

Definitions

  • the invention relates to substituted l-phenyl-3-yl-urea derivatives, their use in plant biotechnology and preparations containing these derivatives.
  • Plant biotechnology can be defined as a set of techniques that, using tissue cultures and/or genetic engineering, result in the production of plants that exhibit new or improved desirable properties.
  • Plant tissue cultures are used for the rapid propagation of plant material in vitro. They are used to propagate a large number of plant species that cannot (or it is expensive) be grown from seeds. As examples may serve the commercially available bananas. New plants are obtained from different parts of the mother plant (from its callus, organs, from different types of tissues, etc.). Should new plants develop (regenerate), it is necessary to add plant hormones that induce regeneration to the growth medium where new plants are regenerated. Key hormones for organogenesis and morphogenesis are auxins and cytokinins, which stimulates the growth of plant roots and stems.
  • the present invention relates to substituted l-phenyl-3- yl-urea derivatives, which have a positive synergistic effect on plant regeneration in tissue cultures.
  • the object of the present invention are l-phenyl-3-yl-urea derivatives of the general formula (I), wherein
  • R1 is selected from the group consisting of halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methylsulfanyl group may optionally be independently substituted with halogen;
  • R2 is selected from the group consisting of hydrogen, halogen or methyl
  • R3 is selected from the group consisting of hydrogen, halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methyl sulfanyl group may optionally be independently substituted with halogen;
  • R4 is selected from the group consisting of hydrogen, halogen or methyl; with the proviso that when Y is nitrogen, then R1 is halogen or methoxy group; and with the proviso that the following compounds are excluded from l-phenyl-3-yl-urea derivatives of the general formula (I): l-(3,5-dichlorophenyl)-3-(2,6-dichlorpyridin-4-yl)urea, l-(3,5-difluorophenyl)-3-(3-(trifluormethylsulfanyl)phenyl)urea, l-(3,5-dichlorophenyl)-3-(3-(trifluormethylsulfanyl)phenyl)urea, l-(3,5-difluorophenyl)-3-(3-methoxyphenyl)urea, l-(3-bromophenyl)-3-(3,5-dichlorophenyl)urea,
  • halogen is selected from the group comprising fluorine, chlorine, bromine and iodine atom; methyl group denotes -CH3; methoxy group denotes -OCH3; methylsulfanyl group denotes -SCH3; trifluormethoxy group denotes -OCF3; trifluoromethylsulfany group denotes -SCF3.
  • Optional substitution of the hydrogen atoms of the methoxy group and the methylsulfanyl group by halogen atoms means the substitution of one, two or three hydrogen atoms by one or more halogen atoms (F, Cl, Br, I).
  • the halogen atoms may be the same or different.
  • the resulting substituent R1 and/or R3, derived from methoxy or methylsulphanyl group may thus be for example:
  • the present invention further includes the compounds of general formula (I) in the form of salts with alkali metals, ammonium or amines, as well as in the form of addition salts with acids.
  • R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
  • R2 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
  • R3 is selected from the group consisting of hydrogen, fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
  • R4 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
  • Y is carbon atom
  • R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methyl sulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
  • Y is nitrogen atom
  • R1 is selected from the group consisting of fluor, chlor, brom or methoxy group.
  • Y is nitrogen atom
  • R1 is selected from the group consisting of F, Cl, Br and -OCH3
  • R2 is selected from the group consisting of H, F, Cl and Br.
  • Y is carbon atom
  • R1 is selected from the group consisting of F, Cl, Br, -OCH3, -OCF3, -SCH3 and -SCF3
  • R2 is selected from the group consisting of H, F, Cl and Br.
  • At least one of R3 and R4 is not hydrogen, preferably R3 and R4 are not hydrogen atoms.
  • R2 is not hydrogen atom.
  • Y is nitrogen and the l-phenyl-3-yl-urea derivative of formula (I) is selected from the group consisting of the following, wherein:
  • R1 is F and R2 is H: l-(3-fluorophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-chlorophenyl)-3-(2- fluoropyridin-4-yl)urea, l-(3-bromophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3,5-difluorophenyl)- 3-(2-fluoropyridin-4-yl)urea, l-(3,5-dichlorophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3,5- dibromophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-chloro-5-fluorophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-bromo-5-chlorophen
  • R1 and R2 are F: l-(3-fluorophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3-chlorophenyl)-3- (2,6-difluoropyridin-4-yl)urea, l-(3-bromophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3,5- difluorophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3,5-dichlorophenyl)-3-(2,6-difluoropyridin-
  • R1 is Cl and R2 is H: l-(3,5-dichlorophenyl)-3-(2-chloropyridin-4-yl)urea, l-(3,5- dibromophenyl)-3-(2-chloropyridin-4-yl)urea, l-(3-chloro-5-fluorophenyl)-3-(2-chloropyridin-4- yl)urea, l-(3-bromo-5-chlorophenyl)-3-(2-chloropyridin-4-yl)urea, l-(2-chloropyridin-4-yl)-3-(3- methoxyphenyl)urea, l-(2-chloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea, l-(2- chloropyridin-4-yl)-3-(3-(methylsulfanyl)phenyl)urea, l-
  • R 1 and R2 are Cl: l-(3-chlorophenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3-bromophenyl)-3- (2,6-dichloropyridin-4-yl)urea, l-(3,5-dibromophenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3- chloro-5-fluorophenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3-bromo-5-chlorophenyl)-3-(2,6- dichloropyridin-4-yl)urea, l-(3-chloro-5-methylphenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(2,6- dichloropyridin-4-yl)-3-(3-methoxypheny
  • R1 is Br and R2 is H: l-(2-bromopyridin-4-yl)-3-(3-fluorophenyl)urea, l-(2-bromopyridin-4-yl)- 3-(3-chlorophenyl)urea, l-(3-bromophenyl)-3-(2-bromopyridin-4-yl)urea, l-(2-bromopyridin-4- y 1) -3 -(3 ,5 -difluorophenyl)urea, 1 -(2-bromopyridin-4-yl)-3-(3 ,5-dichlorophenyl)urea, 1 -(3 ,5 - dibromophenyl)-3-(2-bromopyridin-4-yl)urea, l-(2-bromopyridin-4-yl)-3-(3-chloro-5- fluorophenyl)urea, l-(3
  • Y is carbon atom and the l-phenyl-3-yl-urea derivative of formula (I) is selected from the group consisting of the following, wherein:
  • R1 is SCF3 and R2 is H: l-(3-fluorophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3- chlorophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3-bromophenyl)-3-(3-trifluoromethyl- sulfanylphenyl)urea, l-(3,5-dibromophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3- chloro-5-fluorophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3-bromo-5-chlorophenyl)-3- (3-trifluoromethylsulfanylphenyl)urea, l-(3-chloro-5-methylphenyl)-3-(3
  • R1 is SCH3 and R2 is H: l-(3-fluorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3- chlorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3-bromophenyl)-3-(3- methylsulfanylphenyl)urea, l-(3,5-difluorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3,5-dibromophenyl)-3-(3- methylsulfanylphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3- bromo-5-ch
  • R1 is OCF3 and R2 is H: l-(3-fluorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3-bromophenyl)-3-(3- trifluoromethoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3- trifluoromethoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-trifluoromethoxyphenyl)urea, 1- (3-bromo-5-chlorophenyl
  • R1 is OCF3 and R2 is F: l-(3-fluorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3-bromophenyl)-3-(3-fluoro-5- trifluoromethoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)- moco-vina, l-(3,5-dichlorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3,5- dibromophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3-chloro-5-fluoropheny
  • R1 is OCF 3 and R2 is Cl: l-(3-chloro-5-trifluoromethoxyphenyl)-3-(3-fluorophenyl)urea, l-(3- chlorophenyl)-3-(3-chloro-5-trifluoromethoxyphenyl)urea, l-(3-bromophenyl)-3-(3-chloro-5-tri- fluoromethoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-chloro-5-trifluoromethoxyphenyl)urea, l-(3,5-dichlorophenyl)-3-(3-chloro-5-trifluoromethoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3- chloro-5-trifluoromethoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3
  • R1 is OCF 3 and R2 is Br: l-(3-bromo-5-trifluoromethoxyphenyl)-3-(3-fluorophenyl)urea, l-(3- bromo-5-trifluoromethoxyphenyl)-3-(3-chlorophenyl)urea, l-(3-bromophenyl)-3-(3-bromo-5- trifluoromethoxyphenyl)urea, l-(3-bromo-5-trifluoromethoxyphenyl)-3-(3,5- difluorophenyl)urea, l-(3-bromo-5-trifluoromethoxyphenyl)-3-(3,5-dichlorophenyl)urea, l-(3,5- dibromophenyl)-3-(3-bromo-5-trifluoromethoxyphenyl)urea, l-(3-bromo-5- trifluoromethoxyphenyl
  • R1 is OCH3 and R2 is H: l-(3,5-dichlorophenyl)-3-(3-methoxyphenyl)urea, l-(3,5- dibromophenyl)-3-(3-methoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3- methoxyphenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3-methoxyphenyl)urea,
  • R1 is OCH3 and R2 is F: l-(3-fluorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3-bromophenyl)-3-(3-fluoro-5- methoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3-fluoro-5- methoxyphenyl)urea, 1 -(3 -chloro-5-fluorophenyl)-3 -(3 -fluoro-5-methoxyphenyl)
  • R1 is OCH3 and R2 is Cl: l-(3-chloro-5-methoxyphenyl)-3-(3-fluorophenyl)urea, l-(3- chlorophenyl)-3-(3-chloro-5-methoxyphenyl)urea, l-(3-bromophenyl)-3-(3-chloro-5- methoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-chloro-5-methoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-chloro-5-methoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3-chloro-5- methoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-chloro-5-methoxyphenyl)urea,
  • R1 is OCH3 and R2 is Br: l-(3-fluorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3-bromo-5-methoxyphenyl)-3-(3- bromophenyl)urea, l-(3,5-difluorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3-bromo-5- methoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-bromo-5-methoxyphenyl)urea,
  • R1 is Br and R2 is H: l-(3-bromophenyl)-3-(3,5-difluorophenyl)urea, l-(3-bromophenyl)-3-(3,5- dibromophenyl)urea, l-(3-bromophenyl)-3-(3-chloro-5-fluorophenyl)urea, l-(3-bromophenyl)-3- (3 -bromo-5-chlorophenyl)urea,
  • R1 is Br and R2 is F: l-(3-bromo-5-fluorophenyl)-3-(3-fluorophenyl)urea, l-(3-bromo-5- fluorophenyl)-3-(3-chlorophenyl)urea, l-(3-bromo-5-fluorophenyl)-3-(3-bromophenyl)urea, 1- (3-bromo-5-fluorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-bromo-5-fluorophenyl)-3-(3,5- dichlorophenyl)urea, l-(3-bromo-5-fluorophenyl)-3-(3,5-dibromophenyl)urea, l-(3-bromo-5- fluorophenyl)-3-(3-chloro-5-fluorophenyl)urea, l-(
  • R1 is Br and R2 is Cl: l-(3-bromo-5-chlorophenyl)-3-(3-fluorophenyl)urea, l-(3-bromo-5- chlorophenyl)-3-(3-chlorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3-bromophenyl)urea, 1- (3-bromo-5-chlorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3,5- dichlorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3,5-dibromophenyl)urea, l-(3-bromo-5- chlorophenyl)-3 -(3 -chloro-5-fluorophenyl)urea, 1 ,3
  • R1 and R2 are Br: l-(3,5-dibromophenyl)-3-(3-fluorophenyl)urea, l-(3,5-dibromophenyl)-3-(3- dichlorophenyl)urea, l-(3,5-dibromophenyl)-3-(3-bromophenyl)urea, l-(3,5-dibromophenyl)-3- (3,5-difluorophenyl)urea, l-(3,5-dibromophenyl)-3-(3,5-dichlorophenyl)urea, 1 ,3-bis-(3,5- dibromophenyl)urea, l-(3,5-dibromophenyl)-3-(3-chloro-5-fluorophenyl)urea, l-(3,5- dibromophenyl) 3 -(3 -bromo-5-ch
  • R1 is Cl and R2 is H: l-(3-chlorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-chlorophenyl)-3-(3,5- dichlorophenyl)urea, l-(3-chlorophenyl)-3-(3,5-dibromophenyl)urea, l-(3-chlorophenyl)-3-(3- chloro-5-fluorophenyl)urea, l-(3-chlorophenyl)-3-(3-bromo-5-chlorophenyl)urea,
  • R1 is Cl and R2 is F: l-(3-chloro-5-fluorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-chloro-5- fluorophenyl)-3-(3,5-dichlorophenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3,5- dibromophenyl)urea, l-3-bis-(3-chloro-5-fluorophenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3- bromo-5-chlorophenyl)urea,
  • R 1 and R2 are Cl: l-(3,5-dichlorophenyl)-3-(3,5-difluorophenyl)urea, l-(3,5-dichlorophenyl)-3- (3,5 -dibromophenyl)urea, 1 -(3 ,5-dichlorophenyl)-3 -(3 -chloro-5-fluorophenyl)urea, 1 -(3 ,5 - dichlorophenyl) 3 -(3 -bromo-5-chlorophenyl)urea, R1 is F and R2 is H: l-(3,5-difluorophenyl)-3-(3-fluorophenyl)urea, l-(3,5-dichlorophenyl)-3- (3 -fluorophenyl)urea, 1 -(3 ,5-dibromophenyl)-3 -(3 -fluoropheny
  • R1 is selected from the group consisting of halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methylsulfanyl group may optionally be independently substituted with halogen;
  • R2 is selected from the group consisting of hydrogen, halogen or methyl
  • R3 is selected from the group consisting of hydrogen, halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methyl sulfanyl group may optionally be independently substituted with halogen;
  • R4 is selected from the group consisting of hydrogen, halogen or methyl; with the proviso that when Y is nitrogen, then R1 is halogen or methoxy group; in plant biotechnologies, preferably to increase the regeneration and viability of new plants in tissue cultures, especially in the production of crops.
  • R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
  • R2 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
  • R3 is selected from the group consisting of hydrogen, fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
  • R4 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
  • Y is carbon atom
  • R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
  • Y is nitrogen atom
  • R1 is selected from the group consisting of fluor, chlor, brom or methoxy group.
  • Y is nitrogen atom
  • R1 is selected from the group consisting of F, Cl, Br and - OCH3
  • R2 is selected from the group consisting of H, F, Cl and Br.
  • At least one of R3 and R4 is not hydrogen, preferably R3 and R4 are not hydrogen atoms.
  • R2 is not hydrogen atom.
  • the compound of the general formula (la) is selected from the group comprising: l-(2,6-dichloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea (13), l-(2-bromopyridin-4-yl)- 3-(3-((trifluoromethyl)sulfanyl)phenyl)-urea (16), l-(3,5-dichlorophenyl)-3-(3-
  • the compounds of the present invention of the general formula (I) and (la) show a positive synergistic effect on plant regeneration in tissue cultures in the presence of ’-isopcntcnyladcninc (iP). This effect increases the plant regeneration index and thus reduces their regeneration (production) costs. It has further been observed that the compounds of the present invention of the general formula (I) and (la) show a positive synergistic effect with the cytokinin /rans-zeatin (tZ) on the delay of chlorophyll breakdown in excised wheat leaves.
  • the compounds of the present invention of the general formula (I) and (la) inhibit reproduction and vitality of Caenorhabditis elegans in concentrations lower than 10 pM. All these compounds are very toxic for this soil free-living nematode, thus could be used as anthelmintic agent.
  • Another object of the present invention is thus the use of the compounds of the general formula (I) and (la) or of the preparations containing these compounds in agriculture as anthelmintic agents, preferably against Caenorhabditis elegans.
  • the compounds of the present invention of the general formula (I) and (la) can be applied to whole plants, plant organs, or to plant cells and plant tissues.
  • the compounds of the general formula (I) and (la) are suitable for use in plant tissue cultures, where they prevent the degradation of cytokinins and thus increase the viability of new plants, regeneration of buds and shoots from explants, callus growth, reduce hyperhydricity of plants grown in vitro and show very strong micropropagation and anti- senescence activity.
  • the present invention further includes antisenescent and/or antistress preparations for plants, plant organs and plant cells, comprising at least one compound of the general formula (I) or (la) and at least one auxiliary substance (excipients).
  • the auxiliary substances can be in solid or liquid state.
  • Excipient refers to solvents, solid carriers, surface active agents (surfactants), emulsifiers, dispersants, wetting agents, stabilizers, antifoaming agents, preservatives, viscosity agents, adhesives, and also fertilizers or other active ingredients.
  • Suitable solvents are, for example, aromatic hydrocarbons having from 8 to 12 carbon atoms (e.g. xylene or naphthalene derivatives), phthalates (for example dibutyl phthalate, dioctyl phthalate), aliphatic hydrocarbons (e.g. cyclohexane, paraffin), alcohols, glycols and their ethers and esters (e.g. ethanol, ethylene glycol, 2- methoxyethanol, 2-ethoxyethanol), ketones (e.g. cyclohexanone, N-methyl-2-pyrrolidone, DMSO, DMF), vegetable oils, water.
  • aromatic hydrocarbons having from 8 to 12 carbon atoms
  • phthalates for example dibutyl phthalate, dioctyl phthalate
  • aliphatic hydrocarbons e.g. cyclohexane, paraffin
  • alcohols e.g. cyclohexane, paraffin
  • alcohols e.
  • Solid carriers are typically calcite, talc, kaolin, montmorillonite or attapulgite, silicic acid, polymers, crushed limestone.
  • Suitable surfactants are nonionic, cationic or anionic surfactants (alkali metal salts, alkaline earth metal salts or ammonium salts of higher fatty acids having a carbon number of 10 to 20, especially sodium or potassium salts of oleic or stearic acid, sulfonated fatty acids, sulfonated benzimidazole derivatives, alkyl sulfonates, phosphoric acid esters).
  • Nonionic surfactants are typically derivatives of polyglycol ether and aliphatic or cycloaliphatic alcohols or saturated or unsaturated fatty acids and alkylphenols, water-soluble adducts of polyethylene oxide with polyethylene glycol.
  • Stabilizers are, for example, vegetable oils or epoxidized vegetable oils (epoxidized palm oil, rapeseed or olive oil).
  • the antifoaming agent is, for example, silicone oil.
  • anionic, non-ionic and cationic surfactants examples include WO 97/34485.
  • the invention further relates to a method for inhibiting stress and/or senescence in plants, plant organs and/or plant cells, which comprises applying at least one compound with cytokinin arrangement, from group comprosing: czs-zeatin, /rans-zeatin, ’-isopcntcnyladcnin, ’-bcnzyladcnin, kinetin, meto-topolin, their nukleosides and/or nukleotides and at least one compound of the general formula (I) or (la) to a plant, plant organ and/or plant cell.
  • cytokinin from group comprosing: czs-zeatin, /rans-zeatin, ’-isopcntcnyladcnin, ’-bcnzyladcnin, kinetin, meto-topolin, their nukleosides and/or nukleotides and at least one compound of the general formula (I) or
  • the compounds of the general formula (I) and (la) are used in unmodified form or, preferably, together with excipients conventionally employed in the art of preparations. To this end they are conveniently formulated as concentrates of active compounds as well as suspensions and dispersions, preferentially isotonic water solutions, suspensions and dispersion, diluted emulsions, soluble powders, dusts, granulates, creams, gels, oil suspensions and also encapsulations, e.g. in polymeric substances.
  • the methods of application such as spraying, atomizing, dusting, scattering, coating or pouring, are chosen in accordance with the intended objectives and the prevailing circumstances.
  • the preparations may be sterilized and/or contain further excipients of neutral nature such as preservatives, stabilizers, wetting agents or emulgators, solubilizing agents, as well as fertilizers, micronutrient donors or other formulations for obtaining special effects.
  • excipients of neutral nature such as preservatives, stabilizers, wetting agents or emulgators, solubilizing agents, as well as fertilizers, micronutrient donors or other formulations for obtaining special effects.
  • the compounds of the general formula (I) and (la) of the present invention can be prepared by a method in which an aniline (or pyridine-4-amine) bearing R1 and/or R2 is converted to an isocyanate by a conventional method using diphosgene (Kurita K. et al, (1976) J. Org. Chem. 41, 2070-71). The resulting isocyanate then reacts with an aniline bearing R3 and/or R4 to give the desired product of general formula (I) or (la).
  • compositions comprising the compounds of general formula I or (la) (active ingredients) and, where appropriate, one or more solid or liquid excipients, are prepared in a manner known per se e.g. by mixing and/or grinding the active ingredients with excipients, e.g. solvents or solid carriers.
  • excipients e.g. solvents or solid carriers.
  • surface-active compounds surfactants may also be used in the preparations.
  • suitable surface-active compounds are non-ionic, cationic and/or anionic surfactants and surfactant mixtures having good emulsifying, dispersing and wetting properties.
  • suitable anionic, non-ionic and cationic surfactants are listed, for example, in WO 97/34485.
  • compositions containg compounds of the general formula (I) or (la) according to the invention are the surfactants conventionally used in formulation technology, which are described, inter alia, in "McCutcheon's Detergents and Emulsifiers Annual” MC Publishing Corp., Ridgewood New Jersey, 1981; Stache, H., “Tensid-Taschenbuch", Carl Hanser Verlag, Kunststoff, 1981; and M. and J. Ash, "Encyclopedia of Surfactants", Vol.1-3, Chemical Publishing Co., New York, 1980-81.
  • the formulation of the preparation contains from 0.01 to 99.09 % (w/w), in particular from 0.1 to 95 % (w/w), of the active ingredient corresponding to the compound or mixture of compounds of the general formula (I) or (la), and from 0.01 to 99.09 % (w/w) of a mixture of additives or other carriers, depending on the methods of application, preferably from 5 to 99.9 % (w/w) of a mixture of additives or other carriers; and optionally may further comprise from 0.1 to 40 % (w/w) of a wetting agent, preferably from 0.5 to 30 % (w/w) of a wetting agent, more preferably from 1 to 20 % (w/w) of a wetting agent.
  • compositions may also comprise further ingredients, such as stabilisers, e.g. vegetable oils or epoxidised vegetable oils (epoxidised palm oil, coconut iol, rapeseed oil or soybean oil), antifoams, e.g. silicone oil, preservatives, stabilizers, wetting agents or emulsifiers, viscosity factors, binders, tackifiers, and also fertilisers or other active ingredients.
  • Dusts active ingredient mixture: 0.1 to 10 %, solid carrier: 99.9 to 90 %,
  • Suspension concentrates active ingredient mixture: 0.5 to 75 %, water: 94 to 24 %, surfactant: 1 to 40 %,
  • Wettable powders active ingredient mixture: 0.5 to 90 %, surfactant: 0.5 to 20 %, solid carrier: 5 to 95 %,
  • Granules active ingredient mixture: 0.1 to 30 %, solid carrier: 99.9 to 70 %.
  • compositions may also comprise further ingredients, such as stabilisers, e.g. vegetable oils or epoxidised vegetable oils (epoxidised coconut oil, rapeseed oil or soybean oil), anti-foams, e.g. silicone oil, preservatives, viscosity regulators, binders, tackifiers, and also fertilisers or other active ingredients.
  • stabilisers e.g. vegetable oils or epoxidised vegetable oils (epoxidised coconut oil, rapeseed oil or soybean oil)
  • anti-foams e.g. silicone oil
  • preservatives e.g. silicone oil
  • viscosity regulators binders
  • tackifiers e.g., tackifiers, and also fertilisers or other active ingredients.
  • the method of preparation of the compounds of the general formula (I) or (la) is based on the reaction of 1 equivalent of substituted isocyanatobenzene bearing R3 and R4 with 1 equivalent of substituted aniline (or pyridin-4-amine) bearing R1 and R2 according to Scheme 1 (step B).
  • the reaction is carried out in an inert solvent such as THF or DCM.
  • isocyanates - l-chloro-3-isocyanato-5-(trifluoromethoxy)benzene, l-bromo-3-isocyanato-5- (trifluoromethoxy)benzene, l-chloro-3-isocyanato-5-methoxybenzene, l-bromo-3-isocyanato-5- methoxybenzene were prepared by reacting a corresponding substituted aniline with diphosgene in THF (Kurita K. et al, (1976) J. Org. Chem. 41, 2070-71) according to Scheme 1 (step A).
  • Figure 1 shows: the effect of compound 24 (0.01 pM) on the regeneration of new plants from tobacco explant in the presence of 0.01 pM iP.
  • Figure 2 shows: the excessive formation of degenerate chrysanthemum stems after application of 1 pM TDZ ( Figure 2 A) and the increased formation of newly regenerated and viable chrysanthemum plants after application of 1 pM iP and 1 pM 24 (l-(3,5-dichlorophenyl)-3-(3- (trifluoromethoxy)phenyl)urea) ( Figure 2B).
  • Example 1 Preparation of l-bromo-3-isocyanato-5-(trifluoromethoxy)benzene 3-Bromo-5-(trifluoromethoxy)anilin (1 ekvivalent) was dissolved in dry THF and mixed with trietylamine (2 ekvivalents). This mixture was slowly added dropwise to a solution of diphosgene (0.6 equivalent) in THF at -20 °C. Subsequently, the temperature of the reaction mixture was gradually brought to room temperature or higher (up to 60 °C). After cooling to 4 °C, the precipitated triethylamine hydrochloride was filtered off and the filtrate was evaporated to dryness.
  • Example 2 l-(2-Chloropyridin-4-yl)-3-(3-methoxyphenyl)urea (7) l-Isocyanato-3 -methoxybenzene (75 mg, 0.5 mmol) was dissolved in DCM (5 mF) and 2- chloropyridin-4-amine (64 mg, 0,5 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The white crystalline product crystallized from DCM, and after cooling the reaction mixture to 4 °C, the product was isolated by filtration and dried in a desiccator (isolated yield - 115 mg, 82 %). EM 277.
  • Example 5 l-(2,6-Dichloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea (14) l-Isocyanato-3-(trifluoromethoxy)benzene (101 mg, 0.5 mmol) was dissolved in DCM (5 mL) and 2,6-dichloropyridin-4-amin (81 mg, 0,5 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The white crystalline product crystallized from DCM, and after cooling the reaction mixture to 4 °C, the product was isolated by filtration and dried in a desiccator (isolated yield -144 mg, 79%). EM 365.
  • Example 6 l-(3-chloro-5-fluorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (23) 3-Chloro-5-fluoroaniline (36 mg, 0.25 mmol) and l-isocyanato-3-(trifluoromethoxy)benzene (50 mg, 0.25 mmol) were mixed in DCM and the reaction mixture was stirred at r.t. for 1 h. Then, triethylamine was added (20 .L) until basic pH occurred and reaction mixture was stirred at r.t. overnight. DCM was evaporated and water was added to the white powder.
  • Example 7 l-(3,5-Dichlorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (24) 3,5-Dichlorophenylisocyanate (40 mg, 0,21 mmol) was dissolved in THF (2 mL) and 3- trifluoromethoxyanilin (38 mg, 0,21 mmol) was added. The reaction mixture was stirred at room temperature for 2 h, then THF was evaporated and white crystalline solid was filtered from DCM and dried in a desiccator (isolated yield - 50 mg, 64%). EM 364.
  • Example 11 l-(3,5-Dibromophenyl)-3-(3,5-dichlorophenyl)urea (37) 3,5-Dichlorophenylisocyanate (56 mg, 0,3 mmol) was dissolved in acetone and (3,5- dibromo)aniline (75 mg, 0,3 mmol) was added. The reaction mixture was stirred at room temperature overnight. The oven crystalized from acetone in a fridge. White crystalline solid was filtered off and dried in a desiccator (isolated yield - 85 mg, 65%). EM 438.
  • the number(s) in the abreviation indicates the position(s) of the substituent on the phenyl (pyridyl) ring, the dash separates the two aromatic cycles of the urea.
  • the abbreviations do not include the words phenyl and urea. Abbreviations begin with a nucleus that bears R1 and R2.
  • Example 13 Synergistic effect of compounds of general formula (I) and (la) with cytokinin iP on increasing the regeneration index of tobacco plants.
  • Table 2 Effect of compounds on the regeneration index (number of new plants per explant) of tobacco and the amount of retained chlorophyll in wheat leaves (in %).
  • the values of the regeneration index for compounds 1-40 are the result of the synergistic effect of these compounds in 0.1 pM concentration and of cytokinin iP at the same concentration.
  • the regeneration index value is the average of 15 explants ( ⁇ standard error).
  • the chlorophyll content value (see Example 12) is the average of 5 measurements of chlorophyll content in 5 leaves ( ⁇ standard error).
  • Example 14 Anti-senescence synergistic effect of compounds of general formula (I) and (la) with /rans-zeatin in a senescence test on leaf segments of wheat cultivated in the dark. Seeds of winter wheat Triticum aestivum cv. Hereward was rinsed under running water for 4 hours and then seeded in vermiculite saturated with Knop's nutrient solution (Ca(NO3)2, MgSCU, KNO3, KH2PO4, all ⁇ 1%). The pots were placed in an air-conditioned growth chamber with a 16/8 hour light period (light intensity 50 pmol.m ⁇ .s' 1 ) and a temperature of 15 °C.
  • the seedlings had developed the first flag leaf and the second leaf began to grow.
  • the top sections of the first leaves were removed so that their weight was 25 mg.
  • the basal ends of these leaf segments were placed in the wells of microtiter polystyrene plates containing 150 pL of a solution of a test derivative of formula (I) or (la) (1 leaf segment per well) or a combination thereof with trans- zeatin.
  • the plates were placed in a plastic box with filter paper, which was saturated with water for maximum humidity. After 96 hours of incubation in the dark at 25 °C, the leaf segments were removed and the chlorophyll was extracted in 5 mL of 80 % ethanol by heating at 80 °C for 1 hour.
  • Trans-zeatin applied alone at 1 micromolar concentration retained 13 % chlorophyll.
  • the compounds of formula (I) and (la) applied alone at 1 micromolar concentration did not show a significant positive effect on the chlorophyll retention in the wheat leaves (retained 0 to 5 % over the negative control).
  • Trans-zeatin applied alone at 10 micromolar concentration was able to retain 28 % chlorophyll compared to the negative control.
  • the compounds of formula (I) and (la) applied alone at a concentration of 10 micromolar were able to retain between 5 and 16 % of chlorophyll, thus showing a weak positive effect on the delay of senescence in the segments of wheat leaves caused by darkness. Results are summarised in Table 2.
  • Example 15 Comparison of the influence of selected growth regulators and their combined application with iP on chrysanthemum regeneration
  • TDZ thidiazuron; l-phenyl-3-(l,2,3-thiadiazol-5-yl)urea
  • Example 16 The effect of tested compounds on vitality of Caenorhabditis elegans
  • CGC Caenorhabditis genetic center
  • M9 buffer for 1 L: 6 g Na2HPO4, 3 g KH2PO4, 5 g NaCl, 0.25 g MgSO4 x 7H2O
  • bleaching solution for 10 mL: 8 mL water, 1,5
  • LI larvae were diluted with S -complete medium to a final concentration of 200 - 300 worms per ml, fed with 3 mg/ml bacteria and distributed onto 96-well plates. Larvae were treated with the test compounds (in five different concentartions) or DMSO (negative control) in equal volume. Anthelmintic drug ivermectin was used as positive control. Populations were left to grow at 20 °C for 4 days. After this period, healthy worms would reach adulthood and start reproducing, thus producing eggs. The eggs produce enzyme chitinase to evolve in LI larvae stadium.
  • the non- fluorescent (Anorogenic) chitinase substrate 4-methylumbelliferyl -D-N,N',N"- triacetylchitotrioside (20 pM) was added to the wells to detect the Auoroscent product of the chitinase reaction - 4-methylumbelliferon.
  • the plates were incubated at 37 °C for Ih. The reaction was then stopped by the addition of alkaline buffer (I M glycine/1 M NaOH, pH 10.6), and the intensity of the Auorescence of 4-methylumbelliferon was measured on a plate reader. Prior to the assay, the plates were visually checked under a microscope, and abnormalities, such as toxicity or developmental impairments, were noted. The toxic effect of the tested compounds were performed in a wider concentration range to obtain the /C50 valeus. Those were calculated using Graphpad Prism. Experiments were repeated at least three times.
  • the active ingredient in the formulations may 1 any compound of formula (I) or mixture of two or more compounds of general formula (I).
  • Emulsifiable concentrates a) b) c) d) active ingredient mixture 5 % 10 % 25 % 50 % calcium dodecylbenzenesulfonate 6 % 8 % 6 % 8 % castor oil poly glycol ether 4 % - 4 % 4 %
  • the solutions are suitable for use in the form of microdrops.
  • Wettable powders a) b) c) d) active ingredient mixture 5 % 25 % 50 % 80 % sodium ligno sulfonate 4 % 3 % sodium lauryl sulfate 2 % 3 % - 4 % sodium diisobutylnaphthalenesulfonate 6 % 5 % 6 % octylphenol polyglycol ether 1 % 2 % - (7-8 mol of ethylene oxide) highly dispersed silicic acid 1 % 3 % 5 % 10 % kaolin 87 % 61 % 37 %
  • the active ingredient is mixed thoroughly with the adjuvants and the mixture is thoroughly ground in a suitable mill, affording wettable powders which can be diluted with water to give suspensions of any desired concentration.
  • Coated granules a) b) c) active ingredient mixture 0.1 % 5 % 15 % highly dispersed silicic acid 0.9 % 2 % 2 % inorganic carrier 99.0 % 93 % 83 %
  • the active ingredient is dissolved in methylene chloroide and applied to the carrier by spraying, and the solvent is then evaporated off in vacuo.
  • Coated granules a) b) c) active ingredient mixture 0.1 % 5 % 15 % polyethylene glycol MW 200 1.0 % 2 % 3 % highly dispersed silicic acid 0.9 % 1 % 2 % inorganic carrier 98.0 % 92 % 80 % (AE 0.1-1 mm) e.g. CaCCh or SiCh
  • active ingredient mixture 0.1 % 5 % 15 % polyethylene glycol MW 200 1.0 % 2 % 3 % highly dispersed silicic acid 0.9 % 1 % 2 % inorganic carrier 98.0 % 92 % 80 % (AE 0.1-1 mm) e.g. CaCCh or SiCh
  • the finely ground active ingredient is uniformly applied, in a mixer, to the carrier moistened with polyethylene glycol. Non-dusty coated granules are obtained in this manner.
  • the active ingredient is mixed and ground with the adjuvants, and the mixture is moistened with water.
  • the mixture is extruded and then dried in a stream of air.
  • Ready-to-use dusts are obtained by mixing the active ingredient with the carriers and grinding the mixture in a suitable mill.
  • Suspension concentrates a) b) c) d) active ingredient mixture 3 % 10 % 25 % 50 % ethylene glycol 5 % 5 % 5 % 5 % nonylphenol polyglycol ether 1 % 2 % (15 mol of ethylene oxide) sodium ligno sulfonate 3 % 3 % 4 % 5 % c arboxy methy Icellulo se 1 % 1 % 1 % 1 % 1 % 37 % aqueous formaldehyde 0.2 % 0.2 % 0.2 % 0.2 % solution silicone oil emulsion 0.8 % 0.8 % 0.8 % 0.8 % % water 86 % 78 % 64 % 38 %
  • the finely ground active ingredient is intimately mixed with the adjutants, giving a suspension concentrate from which suspensions of any desired concentration can be obtained by dilution with water.

Abstract

The present invention relates to substituted 1-phenyl-3-yl-urea derivatives of the general formula (I), and preparations containing these derivatives, (I) wherein Y is nitrogen or carbon; R1 is halogen, methoxy group, methylsulfanyl group, optionally the hydrogens of methoxy and methylsulfanyl group may be substituted by halogen, R2 and R4 are independently hydrogen, fluor, chlorine, bromine or methyl and R3 is hydrogen or R1, with the proviso that when Y is nitrogen, then R1 is halogen or methoxy group. The invention further relates to the use of 1-phenyl-3-yl-urea derivatives of the general formula (I) in plant biotechnologies for increasing the regeneration and viability of new plants, in particular in the production of useful plants, and to preparations containing these derivatives.

Description

SUBSTITUTED 1 ,3-DIPHENYLUREA DERIVATIVES AND 1-PHENYL-3-PYRIDYLUREA DERIVATIVES FOR PLANT BIOTECHNOLOGY, PREPARATIONS CONTAINING THESE COMPOUNDS AND USE THEREOF
Technical Field
The invention relates to substituted l-phenyl-3-yl-urea derivatives, their use in plant biotechnology and preparations containing these derivatives.
Background Art
Plant biotechnology can be defined as a set of techniques that, using tissue cultures and/or genetic engineering, result in the production of plants that exhibit new or improved desirable properties. Plant tissue cultures are used for the rapid propagation of plant material in vitro. They are used to propagate a large number of plant species that cannot (or it is expensive) be grown from seeds. As examples may serve the commercially available bananas. New plants are obtained from different parts of the mother plant (from its callus, organs, from different types of tissues, etc.). Should new plants develop (regenerate), it is necessary to add plant hormones that induce regeneration to the growth medium where new plants are regenerated. Key hormones for organogenesis and morphogenesis are auxins and cytokinins, which stimulates the growth of plant roots and stems. Despite the wide range of commercially available auxins and cytokinins, some plant species still fail to regenerate or their regeneration index (number of regenerated plants from one part of the mother plant) is very low. There is thus a constant need of new types of compounds to increas the plant regeneration index.
Disclosure of the Invention
It is the aim of the present invention to provide novel compounds from the group of phytohormones which would increase plant regeneration. The present invention relates to substituted l-phenyl-3- yl-urea derivatives, which have a positive synergistic effect on plant regeneration in tissue cultures.
The object of the present invention are l-phenyl-3-yl-urea derivatives of the general formula (I),
Figure imgf000003_0001
wherein
Y is nitrogen atom (=N-) or carbon atom (=C(H)-);
R1 is selected from the group consisting of halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methylsulfanyl group may optionally be independently substituted with halogen;
R2 is selected from the group consisting of hydrogen, halogen or methyl;
R3 is selected from the group consisting of hydrogen, halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methyl sulfanyl group may optionally be independently substituted with halogen;
R4 is selected from the group consisting of hydrogen, halogen or methyl; with the proviso that when Y is nitrogen, then R1 is halogen or methoxy group; and with the proviso that the following compounds are excluded from l-phenyl-3-yl-urea derivatives of the general formula (I): l-(3,5-dichlorophenyl)-3-(2,6-dichlorpyridin-4-yl)urea, l-(3,5-difluorophenyl)-3-(3-(trifluormethylsulfanyl)phenyl)urea, l-(3,5-dichlorophenyl)-3-(3-(trifluormethylsulfanyl)phenyl)urea, l-(3,5-difluorophenyl)-3-(3-methoxyphenyl)urea, l-(3-bromophenyl)-3-(3,5-dichlorophenyl)urea,
1.3-bis-(3,5-dichlorophenyl)urea,
1.3-bis-(3,5-difluorophenyl)urea,
1.3-bis(3-(trifluoromethoxy)phenyl)urea,
1 - (3 -bromophenyl) -3 - (pheny l)urea, l-(3-fluorophenyl)-3-(phenyl)urea, l-(3-methoxyphenyl)-3-(phenyl)urea, l-(3-fluorophenyl)-3-(3-methylphenyl)urea, l-(3-methylphenyl)-3-(2-chloropyridin-4-yl)urea, l-(3-chlorophenyl)-3-(2-chloropyridin-4-yl)urea, l-(phenyl)-3-(2-chloropyridin-4-yl)urea, l-(phenyl)-3-(2-methoxypyridin-4-yl)urea, l-(phenyl)-3-(2-fluoropyridin-4-yl)urea, l-(phenyl)-3-(2-bromopyridin-4-yl)urea, l-(phenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3-fluorophenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(phenyl)-3-(2,6-dibromopyridin-4-yl)urea, l-(phenyl)-3-(2-chloro-6-methoxypyridin-4-yl)urea, l-(3-fluorophenyl)-3-(2-chloropyridin-4-yl)urea, l-(3,5-difluorophenyl)-3-(2-chloropyridin-4-yl)urea.
The generic substituent groups have meanings as defined herein below: halogen is selected from the group comprising fluorine, chlorine, bromine and iodine atom; methyl group denotes -CH3; methoxy group denotes -OCH3; methylsulfanyl group denotes -SCH3; trifluormethoxy group denotes -OCF3; trifluoromethylsulfany group denotes -SCF3.
Optional substitution of the hydrogen atoms of the methoxy group and the methylsulfanyl group by halogen atoms means the substitution of one, two or three hydrogen atoms by one or more halogen atoms (F, Cl, Br, I). The halogen atoms may be the same or different.
The resulting substituent R1 and/or R3, derived from methoxy or methylsulphanyl group may thus be for example:
-OCCh; -OCBn; -OCI3;
-OCFCh; -OCFBn; -OCFI2; -OCCIF2; -OCCIBn; -OCCII2; -OCBrF2; -OCBrCh; -OCBrh; -
OCIF2; -OCICI2; -OCIBn;
-OCFCIBr; -OCFC1I; -OCCIBrI;
-OCHF2; -OCHCh; -OCHBn; -OCHI2;
-OCHFC1; -OCHFBr; -OCHFI; -OCHClBr; -OCHC1I; -OCHBrI;
-OCH2CI; -OCFbBr; -OCH2I; -OCH2F; -SCCh; -SCBr3; -SCI3;
-SCFCI2; -SCFBn; -SCFI2; -SCCIF2; -SCClBr2; -SCCII2; -SCBrF2; -SCBrCh; -SCBrI2; -SCIF2; -
SCICI2; -SCIBn;
-SCFCIBr; -SCFC1I; -SCCIBrI;
-SCHF2; -SCHCI2; -SCHBn; -SCHI2;
-SCHFC1; -SCHFBr; -SCHFI; -SCHClBr; -SCHC1I; -SCHBrI;
-SCH2CI; -SCFbBr; -SCH2I; -SCH2F.
The present invention further includes the compounds of general formula (I) in the form of salts with alkali metals, ammonium or amines, as well as in the form of addition salts with acids.
In one preferred embodiment R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
In one preferred embodiment R2 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
In one preferred embodiment R3 is selected from the group consisting of hydrogen, fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
In one preferred embodiment R4 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
In one preferred embodiment Y is carbon atom, R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methyl sulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
In one embodiment Y is nitrogen atom, R1 is selected from the group consisting of fluor, chlor, brom or methoxy group.
In one preferred embodiment Y is nitrogen atom, R1 is selected from the group consisting of F, Cl, Br and -OCH3; and R2 is selected from the group consisting of H, F, Cl and Br. In one preferred embodiment Y is carbon atom; R1 is selected from the group consisting of F, Cl, Br, -OCH3, -OCF3, -SCH3 and -SCF3; and R2 is selected from the group consisting of H, F, Cl and Br.
In one embodiment, at least one of R3 and R4 is not hydrogen, preferably R3 and R4 are not hydrogen atoms.
In one embodiment R2 is not hydrogen atom.
In one embodiment, Y is nitrogen and the l-phenyl-3-yl-urea derivative of formula (I) is selected from the group consisting of the following, wherein:
R1 is F and R2 is H: l-(3-fluorophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-chlorophenyl)-3-(2- fluoropyridin-4-yl)urea, l-(3-bromophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3,5-difluorophenyl)- 3-(2-fluoropyridin-4-yl)urea, l-(3,5-dichlorophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3,5- dibromophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-chloro-5-fluorophenyl)-3-(2-fluoropyridin-4- yl)urea, l-(3-bromo-5-chlorophenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-chloro-5-methylphenyl)-
3-(2-fluoropyridin-4-yl)urea, l-(2-fluoropyridin-4-yl)-3-(3-methoxyphenyl)urea, l-(2- fluoropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea, l-(2-fluoropyridin-4-yl)-3-(3-
(methylsulfanyl)phenyl)urea, l-(2-fluropyridin-4-yl)-3-(3-
((trifluoromethyl)sulfanyl)phenyl)urea, l-(3-chloro-5-methoxyphenyl)-3-(2-fluoropyridin-4- yl)urea, l-(3-bromo-5-methoxyphenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-chloro-5- (trifluoromethoxy)phenyl)-3-(2-fluoropyridin-4-yl)urea, l-(3-bromo-5-
(trifluoromethoxy)phenyl)-3-(2-fluoropyridin-4-yl)urea,
R1 and R2 are F: l-(3-fluorophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3-chlorophenyl)-3- (2,6-difluoropyridin-4-yl)urea, l-(3-bromophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3,5- difluorophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3,5-dichlorophenyl)-3-(2,6-difluoropyridin-
4-yl)urea, l-(3,5-dibromophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3-chloro-5-fluorophenyl)- 3-(2,6-difluoropyridin-4-yl)urea, l-(3-bromo-5-chlorophenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3-chloro-5-methylphenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(2,6-difluoropyridin-4-yl)-3-(3- methoxyphenyl)urea, l-(2,6-difluoropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea, l-(2,6- difluoropyridin-4-yl)-3-(3-(methylsulfanyl)phenyl)urea, l-(2,6-difluropyridin-4-yl)-3-(3- ((trifluoromethyl)sulfanyl)phenyl)urea, l-(3-chloro-5-methoxyphenyl)-3-(2,6-difluoropyridin-4- yl)urea, l-(3-bromo-5-methoxyphenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3-chloro-5- ( trifluoromethoxy )phenyl)-3-(2,6-difluoropyridin-4-yl)urea, l-(3-bromo-5-
( trifluoromethoxy )phenyl)-3-(2,6-difluoropyridin-4-yl)urea,
R1 is Cl and R2 is H: l-(3,5-dichlorophenyl)-3-(2-chloropyridin-4-yl)urea, l-(3,5- dibromophenyl)-3-(2-chloropyridin-4-yl)urea, l-(3-chloro-5-fluorophenyl)-3-(2-chloropyridin-4- yl)urea, l-(3-bromo-5-chlorophenyl)-3-(2-chloropyridin-4-yl)urea, l-(2-chloropyridin-4-yl)-3-(3- methoxyphenyl)urea, l-(2-chloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea, l-(2- chloropyridin-4-yl)-3-(3-(methylsulfanyl)phenyl)urea, l-(2-chloropyridin-4-yl)-3-(3-
((trifluoromethyl)sulfanyl)phenyl)urea, l-(3-chloro-5-methoxyphenyl)-3-(2-chloropyridin-4- yl)urea, l-(3-bromo-5-methoxyphenyl)-3-(2-chloropyridin-4-yl)urea, l-(3-chloro-5- ( trifluoromethoxy )phenyl)-3-(2-chloropyridin-4-yl)urea, l-(3-bromo-5-
( trifluoromethoxy )phenyl)-3-(2-chloropyridin-4-yl)urea,
R 1 and R2 are Cl: l-(3-chlorophenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3-bromophenyl)-3- (2,6-dichloropyridin-4-yl)urea, l-(3,5-dibromophenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3- chloro-5-fluorophenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3-bromo-5-chlorophenyl)-3-(2,6- dichloropyridin-4-yl)urea, l-(3-chloro-5-methylphenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(2,6- dichloropyridin-4-yl)-3-(3-methoxyphenyl)urea, l-(2,6-dichloropyridin-4-yl)-3-(3-
( trifluoromethoxy )phenyl)urea, l-(2,6-dichloropyridin-4-yl)-3-(3-(methylsulfanyl)phenyl)urea, l-(2,6-dichloropyridin-4-yl)-3-(3-((trifluoromethyl)sulfanyl)phenyl)urea, l-(3-chloro-5- methoxyphenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3-bromo-5-methoxyphenyl)-3-(2,6- dichloropyridin-4-yl)urea, l-(3-chloro-5-(trifluoromethoxy)phenyl)-3-(2,6-dichloropyridin-4- yl)urea, l-(3-bromo-5-(trifluoromethoxy)phenyl)-3-(2,6-dichloropyridin-4-yl)urea,
R1 is Br and R2 is H: l-(2-bromopyridin-4-yl)-3-(3-fluorophenyl)urea, l-(2-bromopyridin-4-yl)- 3-(3-chlorophenyl)urea, l-(3-bromophenyl)-3-(2-bromopyridin-4-yl)urea, l-(2-bromopyridin-4- y 1) -3 -(3 ,5 -difluorophenyl)urea, 1 -(2-bromopyridin-4-yl)-3-(3 ,5-dichlorophenyl)urea, 1 -(3 ,5 - dibromophenyl)-3-(2-bromopyridin-4-yl)urea, l-(2-bromopyridin-4-yl)-3-(3-chloro-5- fluorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(2-bromopyridin-4-yl)urea, l-(2- bromopyridin-4-yl)-3-(3-methoxyphenyl)urea, l-(2-bromopyridin-4-yl)-3-(3-
( trifluoromethoxy )phenyl)urea, l-(2-bromopyridin-4-yl)-3-(3-(methylsulfanyl)phenyl)urea, l-(2- bromopyridin-4-yl)-3-(3-((trifluoromethyl)sulfanyl)phenyl)urea, l-(3-chloro-5-methoxyphenyl)- 3-(2-bromopyridin-4-yl)urea, l-(3-chloro-5-(trifluoromethoxy)phenyl)-3-(2-bromopyridin-4- yl)urea, R1 is OCH3 and R2 is H: l-(3-fluorophenyl)-3-(2-methoxypyridin-4-yl)urea, l-(3- chlorophenyl)-3-(2-methoxypyridin-4-yl)urea, l-(3-bromophenyl)-3-(2-methoxypyridin-4- yl)urea, l-(3,5-difluorophenyl)-3-(2-methoxypyridin-4-yl)urea, l-(3,5-dichlorophenyl)-3-(2- methoxypyridin-4-yl)urea, l-(3,5-dibromophenyl)-3-(2-methoxypyridin-4-yl)urea, l-(3-chloro-5- fluorophenyl)-3-(2-methoxypyridin-4-yl)urea, l-(3-bromo-5-chlorophenyl)-3-(2- methoxypyridin-4-yl)urea.
In one embodiment, Y is carbon atom and the l-phenyl-3-yl-urea derivative of formula (I) is selected from the group consisting of the following, wherein:
R1 is SCF3 and R2 is H: l-(3-fluorophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3- chlorophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3-bromophenyl)-3-(3-trifluoromethyl- sulfanylphenyl)urea, l-(3,5-dibromophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3- chloro-5-fluorophenyl)-3-(3-trifluoromethylsulfanylphenyl)urea, l-(3-bromo-5-chlorophenyl)-3- (3-trifluoromethylsulfanylphenyl)urea, l-(3-chloro-5-methylphenyl)-3-(3- trifluoromethylsulfanylphenyl)urea,
R1 is SCH3 and R2 is H: l-(3-fluorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3- chlorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3-bromophenyl)-3-(3- methylsulfanylphenyl)urea, l-(3,5-difluorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3,5-dibromophenyl)-3-(3- methylsulfanylphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3- bromo-5-chlorophenyl)-3-(3-methylsulfanylphenyl)urea, l-(3-chloro-5-methylphenyl)-3-(3- methylsulfanylphenyl)urea,
R1 is OCF3 and R2 is H: l-(3-fluorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3-bromophenyl)-3-(3- trifluoromethoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-trifluoromethoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3- trifluoromethoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-trifluoromethoxyphenyl)urea, 1- (3-bromo-5-chlorophenyl)-3-(3-trifluoromethoxyphenyl)urea,
R1 is OCF3 and R2 is F: l-(3-fluorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3-bromophenyl)-3-(3-fluoro-5- trifluoromethoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)- moco-vina, l-(3,5-dichlorophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3,5- dibromophenyl)-3-(3-fluoro-5-trifluoromethoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3- fluoro-5-trifluoro-methoxyphenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3-fluoro-5- trifluoromethoxyphenyl)urea,
R1 is OCF3 and R2 is Cl: l-(3-chloro-5-trifluoromethoxyphenyl)-3-(3-fluorophenyl)urea, l-(3- chlorophenyl)-3-(3-chloro-5-trifluoromethoxyphenyl)urea, l-(3-bromophenyl)-3-(3-chloro-5-tri- fluoromethoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-chloro-5-trifluoromethoxyphenyl)urea, l-(3,5-dichlorophenyl)-3-(3-chloro-5-trifluoromethoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3- chloro-5-trifluoromethoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-chloro-5-trifluorometh- oxyphenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3-chloro-5-trifluoromethoxyphenyl)urea,
R1 is OCF3 and R2 is Br: l-(3-bromo-5-trifluoromethoxyphenyl)-3-(3-fluorophenyl)urea, l-(3- bromo-5-trifluoromethoxyphenyl)-3-(3-chlorophenyl)urea, l-(3-bromophenyl)-3-(3-bromo-5- trifluoromethoxyphenyl)urea, l-(3-bromo-5-trifluoromethoxyphenyl)-3-(3,5- difluorophenyl)urea, l-(3-bromo-5-trifluoromethoxyphenyl)-3-(3,5-dichlorophenyl)urea, l-(3,5- dibromophenyl)-3-(3-bromo-5-trifluoromethoxyphenyl)urea, l-(3-bromo-5- trifluoromethoxyphenyl)-3-(3-chloro-5-fluorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3- bromo-5-trifluoromethoxyphenyl)urea,
R1 is OCH3 and R2 is H: l-(3,5-dichlorophenyl)-3-(3-methoxyphenyl)urea, l-(3,5- dibromophenyl)-3-(3-methoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3- methoxyphenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3-methoxyphenyl)urea,
R1 is OCH3 and R2 is F: l-(3-fluorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3-bromophenyl)-3-(3-fluoro-5- methoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3-fluoro-5- methoxyphenyl)urea, 1 -(3 -chloro-5-fluorophenyl)-3 -(3 -fluoro-5-methoxyphenyl)urea, 1 -(3- bromo-5-chlorophenyl)-3-(3-fluoro-5-methoxyphenyl)urea,
R1 is OCH3 and R2 is Cl: l-(3-chloro-5-methoxyphenyl)-3-(3-fluorophenyl)urea, l-(3- chlorophenyl)-3-(3-chloro-5-methoxyphenyl)urea, l-(3-bromophenyl)-3-(3-chloro-5- methoxyphenyl)urea, l-(3,5-difluorophenyl)-3-(3-chloro-5-methoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-chloro-5-methoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3-chloro-5- methoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-chloro-5-methoxyphenyl)urea, l-(3- bromo-5-chlorophenyl)-3 -(3 -chloro-5-methoxyphenyl)urea,
R1 is OCH3 and R2 is Br: l-(3-fluorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3- chlorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3-bromo-5-methoxyphenyl)-3-(3- bromophenyl)urea, l-(3,5-difluorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3,5- dichlorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3,5-dibromophenyl)-3-(3-bromo-5- methoxyphenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3-bromo-5-methoxyphenyl)urea, l-(3- bromo-5-chlorophenyl)-3-(3-bromo-5-methoxyphenyl)urea,
R1 is Br and R2 is H: l-(3-bromophenyl)-3-(3,5-difluorophenyl)urea, l-(3-bromophenyl)-3-(3,5- dibromophenyl)urea, l-(3-bromophenyl)-3-(3-chloro-5-fluorophenyl)urea, l-(3-bromophenyl)-3- (3 -bromo-5-chlorophenyl)urea,
R1 is Br and R2 is F: l-(3-bromo-5-fluorophenyl)-3-(3-fluorophenyl)urea, l-(3-bromo-5- fluorophenyl)-3-(3-chlorophenyl)urea, l-(3-bromo-5-fluorophenyl)-3-(3-bromophenyl)urea, 1- (3-bromo-5-fluorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-bromo-5-fluorophenyl)-3-(3,5- dichlorophenyl)urea, l-(3-bromo-5-fluorophenyl)-3-(3,5-dibromophenyl)urea, l-(3-bromo-5- fluorophenyl)-3-(3-chloro-5-fluorophenyl)urea, l-(3-bromo-5-fluorophenyl)-3-(3-bromo-5- chlorophenyl)urea,
R1 is Br and R2 is Cl: l-(3-bromo-5-chlorophenyl)-3-(3-fluorophenyl)urea, l-(3-bromo-5- chlorophenyl)-3-(3-chlorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3-bromophenyl)urea, 1- (3-bromo-5-chlorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3,5- dichlorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3,5-dibromophenyl)urea, l-(3-bromo-5- chlorophenyl)-3 -(3 -chloro-5-fluorophenyl)urea, 1 ,3 -bis-(3 -bromo-5-chlorophenyl)urea,
R1 and R2 are Br: l-(3,5-dibromophenyl)-3-(3-fluorophenyl)urea, l-(3,5-dibromophenyl)-3-(3- dichlorophenyl)urea, l-(3,5-dibromophenyl)-3-(3-bromophenyl)urea, l-(3,5-dibromophenyl)-3- (3,5-difluorophenyl)urea, l-(3,5-dibromophenyl)-3-(3,5-dichlorophenyl)urea, 1 ,3-bis-(3,5- dibromophenyl)urea, l-(3,5-dibromophenyl)-3-(3-chloro-5-fluorophenyl)urea, l-(3,5- dibromophenyl) 3 -(3 -bromo-5-chlorophenyl)urea,
R1 is Cl and R2 is H: l-(3-chlorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-chlorophenyl)-3-(3,5- dichlorophenyl)urea, l-(3-chlorophenyl)-3-(3,5-dibromophenyl)urea, l-(3-chlorophenyl)-3-(3- chloro-5-fluorophenyl)urea, l-(3-chlorophenyl)-3-(3-bromo-5-chlorophenyl)urea,
R1 is Cl and R2 is F: l-(3-chloro-5-fluorophenyl)-3-(3,5-difluorophenyl)urea, l-(3-chloro-5- fluorophenyl)-3-(3,5-dichlorophenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3,5- dibromophenyl)urea, l-3-bis-(3-chloro-5-fluorophenyl)urea, l-(3-chloro-5-fluorophenyl)-3-(3- bromo-5-chlorophenyl)urea,
R 1 and R2 are Cl: l-(3,5-dichlorophenyl)-3-(3,5-difluorophenyl)urea, l-(3,5-dichlorophenyl)-3- (3,5 -dibromophenyl)urea, 1 -(3 ,5-dichlorophenyl)-3 -(3 -chloro-5-fluorophenyl)urea, 1 -(3 ,5 - dichlorophenyl) 3 -(3 -bromo-5-chlorophenyl)urea, R1 is F and R2 is H: l-(3,5-difluorophenyl)-3-(3-fluorophenyl)urea, l-(3,5-dichlorophenyl)-3- (3 -fluorophenyl)urea, 1 -(3 ,5-dibromophenyl)-3 -(3 -fluorophenyl)urea, 1 -(3 -chloro-5- fluorophenyl)-3-(3-fluorophenyl)urea, l-(3-bromo-5-chlorophenyl)-3-(3-fluorophenyl)urea, R 1 and R2 are F - l-(3-bromo-5-chlorophenyl)-3-(3,5-difluorophenyl)urea.
Further aspect of the invention is the use of a compound of general formula (la)
Figure imgf000011_0001
wherein
Y is nitrogen atom (=N-) or carbon atom (=C(H)-);
R1 is selected from the group consisting of halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methylsulfanyl group may optionally be independently substituted with halogen;
R2 is selected from the group consisting of hydrogen, halogen or methyl;
R3 is selected from the group consisting of hydrogen, halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methyl sulfanyl group may optionally be independently substituted with halogen;
R4 is selected from the group consisting of hydrogen, halogen or methyl; with the proviso that when Y is nitrogen, then R1 is halogen or methoxy group; in plant biotechnologies, preferably to increase the regeneration and viability of new plants in tissue cultures, especially in the production of crops.
In one preferred embodiment of the use of the compound of the general formula (la) in plant biotechnology, R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group. In one preferred embodiment of the use of the compound of the general formula (la) in plant biotechnology, R2 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
In one preferred embodiment of the use of the compound of the general formula (la) in plant biotechnology, R3 is selected from the group consisting of hydrogen, fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
In one preferred embodiment of the use of the compound of the general formula (la) in plant biotechnology, R4 is selected from the group consisting of hydrogen, fluor, chlor, brom and methyl.
In one embodiment of the use of the compound of the general formula (la) in plant biotechnology,
Y is carbon atom, R1 is selected from the group consisting of fluor, chlor, brom, methoxy group, methylsulfanyl group, trifluoromethoxy group or trifluoromethylsulfanyl group.
In one embodiment of the use of the compound of the general formula (la) in plant biotechnology,
Y is nitrogen atom, R1 is selected from the group consisting of fluor, chlor, brom or methoxy group.
In one preferred embodiment of the use of the compound of the general formula (la) in plant biotechnology, Y is nitrogen atom, R1 is selected from the group consisting of F, Cl, Br and - OCH3; and R2 is selected from the group consisting of H, F, Cl and Br.
In one preferred embodiment of the use of the compound of the general formula (la) in plant biotechnology, Y is carbon atom; R1 is selected from the group consisting of F, Cl, Br, -OCH3, - OCF3, -SCH3 and -SCF3; and R2 is selected from the group consisting of H, F, Cl and Br.
In one embodiment, at least one of R3 and R4 is not hydrogen, preferably R3 and R4 are not hydrogen atoms.
In one embodiment R2 is not hydrogen atom. In one preferred embodiment of the use of the compound of the general formula (la) in plant biotechnology, the compound of the general formula (la) is selected from the group comprising: l-(2,6-dichloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea (13), l-(2-bromopyridin-4-yl)- 3-(3-((trifluoromethyl)sulfanyl)phenyl)-urea (16), l-(3,5-dichlorophenyl)-3-(3-
((trifluoromethyl)sulfanyl)phenyl)urea (19), l-(3-chloro-5-fluorophenyl)-3-(3-
(trifluoromethoxy)phenyl)urea (23), l-(3,5-difluorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (24), l-(3,5-dichlorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (25), l-(3,5-dichlorophenyl)-3- (3-fluoro-5-(trifluoromethoxy)phenyl)urea (26), l-(3-chloro-5-(trifluoromethoxy)phenyl)-3-(3- chlorophenyl)urea (27).
It has been observed that the compounds of the present invention of the general formula (I) and (la) show a positive synergistic effect on plant regeneration in tissue cultures in the presence of ’-isopcntcnyladcninc (iP). This effect increases the plant regeneration index and thus reduces their regeneration (production) costs. It has further been observed that the compounds of the present invention of the general formula (I) and (la) show a positive synergistic effect with the cytokinin /rans-zeatin (tZ) on the delay of chlorophyll breakdown in excised wheat leaves.
It has further been observed that the compounds of the present invention of the general formula (I) and (la) inhibit reproduction and vitality of Caenorhabditis elegans in concentrations lower than 10 pM. All these compounds are very toxic for this soil free-living nematode, thus could be used as anthelmintic agent. Another object of the present invention is thus the use of the compounds of the general formula (I) and (la) or of the preparations containing these compounds in agriculture as anthelmintic agents, preferably against Caenorhabditis elegans.
The compounds of the present invention of the general formula (I) and (la) can be applied to whole plants, plant organs, or to plant cells and plant tissues. The compounds of the general formula (I) and (la) are suitable for use in plant tissue cultures, where they prevent the degradation of cytokinins and thus increase the viability of new plants, regeneration of buds and shoots from explants, callus growth, reduce hyperhydricity of plants grown in vitro and show very strong micropropagation and anti- senescence activity. The present invention further includes antisenescent and/or antistress preparations for plants, plant organs and plant cells, comprising at least one compound of the general formula (I) or (la) and at least one auxiliary substance (excipients). The auxiliary substances can be in solid or liquid state. Excipient refers to solvents, solid carriers, surface active agents (surfactants), emulsifiers, dispersants, wetting agents, stabilizers, antifoaming agents, preservatives, viscosity agents, adhesives, and also fertilizers or other active ingredients.
Suitable solvents are, for example, aromatic hydrocarbons having from 8 to 12 carbon atoms (e.g. xylene or naphthalene derivatives), phthalates (for example dibutyl phthalate, dioctyl phthalate), aliphatic hydrocarbons (e.g. cyclohexane, paraffin), alcohols, glycols and their ethers and esters (e.g. ethanol, ethylene glycol, 2- methoxyethanol, 2-ethoxyethanol), ketones (e.g. cyclohexanone, N-methyl-2-pyrrolidone, DMSO, DMF), vegetable oils, water.
Solid carriers are typically calcite, talc, kaolin, montmorillonite or attapulgite, silicic acid, polymers, crushed limestone.
Suitable surfactants are nonionic, cationic or anionic surfactants (alkali metal salts, alkaline earth metal salts or ammonium salts of higher fatty acids having a carbon number of 10 to 20, especially sodium or potassium salts of oleic or stearic acid, sulfonated fatty acids, sulfonated benzimidazole derivatives, alkyl sulfonates, phosphoric acid esters). Nonionic surfactants are typically derivatives of polyglycol ether and aliphatic or cycloaliphatic alcohols or saturated or unsaturated fatty acids and alkylphenols, water-soluble adducts of polyethylene oxide with polyethylene glycol.
Stabilizers are, for example, vegetable oils or epoxidized vegetable oils (epoxidized palm oil, rapeseed or olive oil).
The antifoaming agent is, for example, silicone oil.
Examples of suitable anionic, non-ionic and cationic surfactants are listed, for example, in WO 97/34485.
The invention further relates to a method for inhibiting stress and/or senescence in plants, plant organs and/or plant cells, which comprises applying at least one compound with cytokinin aktivity, from group comprosing: czs-zeatin, /rans-zeatin, ’-isopcntcnyladcnin, ’-bcnzyladcnin, kinetin, meto-topolin, their nukleosides and/or nukleotides and at least one compound of the general formula (I) or (la) to a plant, plant organ and/or plant cell.
The compounds of the general formula (I) and (la) are used in unmodified form or, preferably, together with excipients conventionally employed in the art of preparations. To this end they are conveniently formulated as concentrates of active compounds as well as suspensions and dispersions, preferentially isotonic water solutions, suspensions and dispersion, diluted emulsions, soluble powders, dusts, granulates, creams, gels, oil suspensions and also encapsulations, e.g. in polymeric substances. As with the type of the preparation, the methods of application, such as spraying, atomizing, dusting, scattering, coating or pouring, are chosen in accordance with the intended objectives and the prevailing circumstances. The preparations may be sterilized and/or contain further excipients of neutral nature such as preservatives, stabilizers, wetting agents or emulgators, solubilizing agents, as well as fertilizers, micronutrient donors or other formulations for obtaining special effects.
The compounds of the general formula (I) and (la) of the present invention can be prepared by a method in which an aniline (or pyridine-4-amine) bearing R1 and/or R2 is converted to an isocyanate by a conventional method using diphosgene (Kurita K. et al, (1976) J. Org. Chem. 41, 2070-71). The resulting isocyanate then reacts with an aniline bearing R3 and/or R4 to give the desired product of general formula (I) or (la).
Preparations
The preparations comprising the compounds of general formula I or (la) (active ingredients) and, where appropriate, one or more solid or liquid excipients, are prepared in a manner known per se e.g. by mixing and/or grinding the active ingredients with excipients, e.g. solvents or solid carriers. In addition, surface-active compounds (surfactants) may also be used in the preparations.
Depending on the nature of the compound of general formula (I) or (la) to be formulated, suitable surface-active compounds are non-ionic, cationic and/or anionic surfactants and surfactant mixtures having good emulsifying, dispersing and wetting properties. Examples of suitable anionic, non-ionic and cationic surfactants are listed, for example, in WO 97/34485. Also suitable in the preparation of the compositions containg compounds of the general formula (I) or (la) according to the invention are the surfactants conventionally used in formulation technology, which are described, inter alia, in "McCutcheon's Detergents and Emulsifiers Annual" MC Publishing Corp., Ridgewood New Jersey, 1981; Stache, H., "Tensid-Taschenbuch", Carl Hanser Verlag, Munich, 1981; and M. and J. Ash, "Encyclopedia of Surfactants", Vol.1-3, Chemical Publishing Co., New York, 1980-81.
The formulation of the preparation contains from 0.01 to 99.09 % (w/w), in particular from 0.1 to 95 % (w/w), of the active ingredient corresponding to the compound or mixture of compounds of the general formula (I) or (la), and from 0.01 to 99.09 % (w/w) of a mixture of additives or other carriers, depending on the methods of application, preferably from 5 to 99.9 % (w/w) of a mixture of additives or other carriers; and optionally may further comprise from 0.1 to 40 % (w/w) of a wetting agent, preferably from 0.5 to 30 % (w/w) of a wetting agent, more preferably from 1 to 20 % (w/w) of a wetting agent.
Whereas commercial products are usually formulated as concentrates, the end user will normally employ dilute formulations. The compositions may also comprise further ingredients, such as stabilisers, e.g. vegetable oils or epoxidised vegetable oils (epoxidised palm oil, coconut iol, rapeseed oil or soybean oil), antifoams, e.g. silicone oil, preservatives, stabilizers, wetting agents or emulsifiers, viscosity factors, binders, tackifiers, and also fertilisers or other active ingredients. Preferred formulations have the following compositions: (% = percent by weight) Emulsifiable concentrates: active ingredient mixture: 0.01 to 90 %, surfactant: 1 to 30 %, liquid carrier: 5 to 94 %
Dusts: active ingredient mixture: 0.1 to 10 %, solid carrier: 99.9 to 90 %,
Suspension concentrates: active ingredient mixture: 0.5 to 75 %, water: 94 to 24 %, surfactant: 1 to 40 %,
Wettable powders: active ingredient mixture: 0.5 to 90 %, surfactant: 0.5 to 20 %, solid carrier: 5 to 95 %,
Granules: active ingredient mixture: 0.1 to 30 %, solid carrier: 99.9 to 70 %.
The compositions may also comprise further ingredients, such as stabilisers, e.g. vegetable oils or epoxidised vegetable oils (epoxidised coconut oil, rapeseed oil or soybean oil), anti-foams, e.g. silicone oil, preservatives, viscosity regulators, binders, tackifiers, and also fertilisers or other active ingredients. For the use of the compounds of general formula (I) or (la), or of compositions comprising them, in the protection of crop plants against the damaging effects of growth regulators, various methods and techniques come into consideration.
The method of preparation of the compounds of the general formula (I) or (la) is based on the reaction of 1 equivalent of substituted isocyanatobenzene bearing R3 and R4 with 1 equivalent of substituted aniline (or pyridin-4-amine) bearing R1 and R2 according to Scheme 1 (step B). The reaction is carried out in an inert solvent such as THF or DCM. Commercially unavailable isocyanates - l-chloro-3-isocyanato-5-(trifluoromethoxy)benzene, l-bromo-3-isocyanato-5- (trifluoromethoxy)benzene, l-chloro-3-isocyanato-5-methoxybenzene, l-bromo-3-isocyanato-5- methoxybenzene were prepared by reacting a corresponding substituted aniline with diphosgene in THF (Kurita K. et al, (1976) J. Org. Chem. 41, 2070-71) according to Scheme 1 (step A).
Figure imgf000017_0001
Scheme 1.
Brief description of drawings
Figure 1 shows: the effect of compound 24 (0.01 pM) on the regeneration of new plants from tobacco explant in the presence of 0.01 pM iP.
Figure 2 shows: the excessive formation of degenerate chrysanthemum stems after application of 1 pM TDZ (Figure 2 A) and the increased formation of newly regenerated and viable chrysanthemum plants after application of 1 pM iP and 1 pM 24 (l-(3,5-dichlorophenyl)-3-(3- (trifluoromethoxy)phenyl)urea) (Figure 2B). Examples
Example 1. Preparation of l-bromo-3-isocyanato-5-(trifluoromethoxy)benzene 3-Bromo-5-(trifluoromethoxy)anilin (1 ekvivalent) was dissolved in dry THF and mixed with trietylamine (2 ekvivalents). This mixture was slowly added dropwise to a solution of diphosgene (0.6 equivalent) in THF at -20 °C. Subsequently, the temperature of the reaction mixture was gradually brought to room temperature or higher (up to 60 °C). After cooling to 4 °C, the precipitated triethylamine hydrochloride was filtered off and the filtrate was evaporated to dryness. Crude isocyanate - l-bromo-3-isocyanato-5-(trifluoromethoxy)benzene remained in the reaction vessel and was used for the next synthesis. Other isocyanates used in the description of the present invention were prepared according to this example.
Example 2. l-(2-Chloropyridin-4-yl)-3-(3-methoxyphenyl)urea (7) l-Isocyanato-3 -methoxybenzene (75 mg, 0.5 mmol) was dissolved in DCM (5 mF) and 2- chloropyridin-4-amine (64 mg, 0,5 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The white crystalline product crystallized from DCM, and after cooling the reaction mixture to 4 °C, the product was isolated by filtration and dried in a desiccator (isolated yield - 115 mg, 82 %). EM 277. 'H NMR (500 MHz, DMSO- d): 6 3.69 (3H, s, OCH3), 6.56 (1H, dd, Ji=8.5 Hz, 72=2.0 HZ, ArH), 6.91 (1H, dd, 77=8.0 Hz, 72=1.0 Hz, ArH), 7.12 (1H, t, 7=2.5 Hz, ArH), 7.16 (1H, t, 7=8.0 Hz, ArH), 7.27 (1H, dd, 77=6.0 Hz, 72=2.5 Hz, ArH), 7.60 (1H, ds, 7=1.5 Hz, ArH), 8.13 (1H, d, 7=6.0 Hz, ArH), 8.95 (1H, s, NH), 9.30 (1H, s, NH). MS-ESF: 278.1 [M+H]+ (100), 280.0 [M+H]+ (55).
Example 3. l-(2-Chloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea (8)
1-Isocyanato-3-(trifluoromethoxy)benzene (101 mg, 0.5 mmol) was dissolved in DCM (5 mL) and
2-chloropyridin-4-amin (64 mg, 0,5 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The white crystalline product crystallized from DCM, and after cooling the reaction mixture to 4 °C, the product was isolated by filtration and dried in a desiccator (isolated yield - 132 mg, 80 %). EM 331. 'H NMR (500 MHz, DMSO-76): 6 6.97 (1H, d, 7=8.25 Hz, ArH), 7.29-7.33 (2H, m, ArH), 7.40 (1H, dt, 7r=8.25 Hz, 7a=2.75 Hz, ArH), 7.61-7.65 (2H, m, ArH), 8.16 (1H, dd, 7 =5.81 Hz, 72=2.45 Hz, ArH), 9.29 (1H, s, NH), 9.43 (1H, s, NH). MS-ESF: 353.8, 355.8 [M+Na]+ (100, 35). MS-ESI : 329.9, 331.9 [M-H]’ (100, 40). Example 4. l-(2,6-Dichloropyridin-4-yl)-3-(3-methoxyphenyl)urea (13) l-Isocyanato-3 -methoxybenzene (75 mg, 0,5 mmol) was dissolved in DCM (5 mL) and 2,6- dichloropyridin-4-amine (81 mg, 0,5 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The white crystalline product crystallized from DCM, and after cooling the reaction mixture to 4 °C, the product was isolated by filtration and dried in a desiccator (isolated yield - 118 mg, 76 %). EM 311. 'H NMR (500 MHz, DMSO- d): 6 3.69 (3H, s, OCH3), 6.58 (1H, dd, J i=l..5 Hz, J2=2.5 HZ, ArH), 6.92 (1H, dd, Ji=1.5 Hz, 72=1.0 Hz, ArH), 7.12 (1H, t, 7=2.0 Hz, ArH), 7.17 (1H, t, 7=8.0 Hz, ArH), 7.51 (2H, s, ArH), 9.11 (1H, s, NH), 9.49 (1H, s, NH). MS- ESI : 310.1, 311.8 [M-H]' (100, 70).
Example 5. l-(2,6-Dichloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea (14) l-Isocyanato-3-(trifluoromethoxy)benzene (101 mg, 0.5 mmol) was dissolved in DCM (5 mL) and 2,6-dichloropyridin-4-amin (81 mg, 0,5 mmol) was added. The reaction mixture was stirred at room temperature for 2 h. The white crystalline product crystallized from DCM, and after cooling the reaction mixture to 4 °C, the product was isolated by filtration and dried in a desiccator (isolated yield -144 mg, 79%). EM 365. 'H NMR (500 MHz, DMSO-76): 8 6.98 (1H, d, 7=8.0 Hz, ArH), 7.31 (1H, d, 7=8.0 Hz, ArH), 7.40 (1H, t, 7=9.0 Hz, ArH), 7.52 (2H, s, ArH), 7.61 (1H, s, ArH), 9.45 (1H, s, NH), 9.63 (1H, s, NH). MS-ESF: 387.8, 389.8 [M+Na]+ (100, 70). MS-ESI' : 363.9, 365.8 [M-H]' (100, 70).
Example 6. l-(3-chloro-5-fluorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (23) 3-Chloro-5-fluoroaniline (36 mg, 0.25 mmol) and l-isocyanato-3-(trifluoromethoxy)benzene (50 mg, 0.25 mmol) were mixed in DCM and the reaction mixture was stirred at r.t. for 1 h. Then, triethylamine was added (20 .L) until basic pH occurred and reaction mixture was stirred at r.t. overnight. DCM was evaporated and water was added to the white powder. White crystals were filtered off and dried; yield: 60 mg (70%); EM 348; 'H NMR (500 MHz, DMSO-<7s): 6 6.94 (1H, d(br), 7=7.1 Hz, ArH), 6.97 (1H, td, 7d=8.3 Hz, Jt=2.2 Hz, ArH), 7.24-7.32 (2H, m, ArH), 7.34- 7.40 (2H, t+m, 7f=8.3 Hz, ArH), 7.62 (1H, s(br), ArH), 9.13 (1H, s(br), NH), 9.17 (1H, s(br), NH); MS-ESF: 349.1, 350.0 [M+H]+ (100, 40); MS-ESI': 347.0, 348.3 [M-H]' (100, 30).
Example 7. l-(3,5-Dichlorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (24) 3,5-Dichlorophenylisocyanate (40 mg, 0,21 mmol) was dissolved in THF (2 mL) and 3- trifluoromethoxyanilin (38 mg, 0,21 mmol) was added. The reaction mixture was stirred at room temperature for 2 h, then THF was evaporated and white crystalline solid was filtered from DCM and dried in a desiccator (isolated yield - 50 mg, 64%). EM 364. 'H NMR (500 MHz, DMSO-tfc): 8 6.94 (1H, d, J=8.6 Hz, ArH), 7.16 (1H, t, J=1.7 Hz, ArH), 7.29 (1H, d, J=8.0 Hz, ArH), 7.37 (1H, t, J=8.0 Hz, ArH), 7.50 (2H, d, J=1.7 Hz, ArH), 7.63 (1H, s(br), ArH), 9.11 (1H, s (br), NH), 9.18 (1H, s (br), NH). MS-ESF CV 20 V, m/z (rel. int.): 365.0 and 367.0 [M+H] + (100 and 70). MS- ESI' CV 20 V, m/z (rel. int.): 363.0 and 365.0 [M-H]' (100 and 70).
Example 8. l-(3,5-difluorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (25)
3.5-Difluoroaniline (32 mg, 0.25 mmol) and l-isocyanato-3-(trifluoromethoxy)benzene (50 mg, 0.25 mmol) were mixed in DCM and the reaction mixture was stirred at r.t. for 1 h. Then, triethylamine was added (20 .L) until basic pH occurred and reaction mixture was stirred at r.t. overnight. DCM was evaporated and water was added to the white powder. White crystals were filtered off and dried; yield: 60 mg (72%); EM 332; 'H NM R (500 MHz, DMSO- d): 6 6.77 (1H, tt, Ji=9.3 Hz, 72=2.3 Hz, ArH), 6.93 (1H, d(br), 7=8.0 Hz, ArH), 7.15 (2H, dd, 7;=10.0 Hz, 72=2.2 Hz, ArH), 7.27 (1H, d(br), 7=8.0 Hz, ArH), 7.37 (1H, t, 7=8.0 Hz, ArH), 7.62 (1H, s(br), ArH), 9.15 (2H, s(br), 2xNH); MS-ESF: 333.2 [M+H]+ (100); MS-ESI': 331.0[M-H]' (100).
Example 9. l-(3-Chloro-5-(trifluoromethoxy)phenyl)-3-(3,5-dichlorophenyl)urea (28)
3.5-Dichlorophenylisocyanate (40 mg, 0,21 mmol) was dissolved in THF and (3-chloro-5- trifluoromethoxy)aniline (45 mg, 0,21 mmol) was added. The reaction mixture was stirred at room temperature for 2 h, then THF was evaporated and chloroform (1 mL) was added. White crystalline solid was filtered off and dried in a desiccator (isolated yield - 65 mg, 76%). EM 398. 'H NMR (500 MHz, DMSO-76): 6 7.12 (1H, s(br), ArH), 7.18 (1H, t, J=1.7 Hz, ArH), 7.50 (3H, d+?, Jd=l .7 Hz, 3xArH), 7.54 (1H, t, J=1.7 Hz, ArH), 9.25 (1H, s, NH), 9.36 (1H, s, NH). MS-ESI' CV 20 V, m/z (rel. int.): 397.0, 399.0 and 400.8 [M-H]' (100, 80 and 30), 795.0, 796.8 and 799.2 [2M-H]' (50, 100 and 50).
Example 10. l-(3-Bromo-5-(trifluoromethoxy)phenyl)-3-(3,5-dichlorophenyl)urea (29)
3,5-Dichlorophenylisocyanate (37 mg, 0,2 mmol) was dissolved in acetone and (3-bromo-5- trifluoromethoxy)aniline (51 mg, 0,2 mmol) was added. The reaction mixture was stirred at room temperature overnight, then acetone was evaporated and chloroform added. White crystalline solid was filtered off and dried in a desiccator (isolated yield - 70 mg, 80%). EM 443. 'H NMR (500 MHz, DMSO-76): 6 7.18 (1H, t, J=1.7 Hz, ArH), 7.22 (1H, s (br), ArH), 7.51 (2H, d, J=1.7 Hz, ArH), 7.53 (1H, s (br), ArH), 7.67 (1H, t, J=1.7 Hz, ArH), 9.24 (1H, s, NH), 9.34 (1H, s, NH). MS-ESI+: 443.0, 445.0, 446.9 [M+H]+ (60, 100, 50).
Example 11. l-(3,5-Dibromophenyl)-3-(3,5-dichlorophenyl)urea (37) 3,5-Dichlorophenylisocyanate (56 mg, 0,3 mmol) was dissolved in acetone and (3,5- dibromo)aniline (75 mg, 0,3 mmol) was added. The reaction mixture was stirred at room temperature overnight. The produkt crystalized from acetone in a fridge. White crystalline solid was filtered off and dried in a desiccator (isolated yield - 85 mg, 65%). EM 438. 'H NMR (500 MHz, DMSO- d): 8 7.17 (1H, t, J=1.7 Hz, ArH), 7.39 (1H, t, J=1.7 Hz, ArH), 7.50 (2H, d, J=1.7 Hz, ArH), 7.67 (2H, d, J=1.7 Hz, ArH), 9.23 (1H, s (br), NH), 9.26 (1H, s (br), NH). MS-ESF: 436.9, 438.9, 440.9, 443.0 [M+H]+ (40, 100, 90, 40).
Example 12. l,3-Bis-(3,5-dichlorophenyl)urea (39)
3,5-Dichlorophenylisocyanate (188 mg, 1 mmol) was dissolved in THF and 3,5-dichloroanilin (162 mg, 1 mmol) was added. The reaction mixture was stirred at room temperature for 2 h, then THF was evaporated and DCM (1 mL) was added. White crystalline solid was filtered off and dried in a desiccator (isolated yield - 310 mg, 88%). EM 348. 'H NMR (500 MHz, DMSO-tTs): 6 7.17 (2H, t, J=1.7 Hz, ArH), 7.50 (4H, d, J=1.7 Hz, ArH), 9.24 (2H, s, NH). MS-ESF CV 20 V, m/z (rel. int.): 348.8 [M+H] + (80), 350.8 [M+H] + (100), 352.8 [M+H] + (60). MS-ESI" CV 20 V, m/z (rel. int.): 346.8 [M-H]’(60), 348.7 [M-H]’(100), 350.7 [M-H]’(50), 696.8 [2M-H]’(50), 698.8 [2M-H]" (60), 700.8 [2M-H]" (50).
Following the procedures of Examples 1-12, additional compounds were prepared. These are listed in Table 1.
Table 1. Overview of substituents in positions R1-R4 for compounds synthesized and their MS- ESI values [M+H]+ identified. If the substituent is not indicated (-), hydrogen (H) is in the position. Explanations: CP-chloropyridyl, P - pyridyl, D - di, F - fluor, Cl (C) - chlorine, Br - bromine, Me - methyl, MeO - methoxy group, MeS - methylsulfanyl group, TFM - trifluoromethoxy group, TFS - trifluoromethylsulfanyl group. The number(s) in the abreviation indicates the position(s) of the substituent on the phenyl (pyridyl) ring, the dash separates the two aromatic cycles of the urea. The abbreviations do not include the words phenyl and urea. Abbreviations begin with a nucleus that bears R1 and R2.
Figure imgf000022_0001
Figure imgf000023_0001
Example 13. Synergistic effect of compounds of general formula (I) and (la) with cytokinin iP on increasing the regeneration index of tobacco plants.
The tobacco leaves were divided into small round disks (d = 0.5 cm) and placed in petri dishes with 10 ml of Murashige and Skoog medium, (sucrose 30 g/L, agar 7 g/L, pH 5.8). This medium further contained 0.1 .M ’-isopcntcnyladcninc (iP) or a combination of 0.1 .M iP and 0.1 .M of compounds 1-40. Each combination was performed in 3 replicates with 15 explants per combination. Plant regeneration was assessed after 3 weeks. The results are shown in Table 2. It can be seen from the results that all compounds 1-40 of the present invention increase the regeneration index of tobacco plants treated with 0.1 .M iP. While 0.1 .M iP alone allowed to the regeneration of about 5 plants from one disc (explant), compounds 1-40 were able to increase this regeneration to approximately 7-12 plants. The best substances thus doubled the number of regenerated plants from one explant. In the extreme case, 0.01 .M compound 25 positively affected the regeneration of new plants from the tobacco explant in the presence of 0.01 .M iP (Figure 1). Compounds 1-40 applied alone (without iP) have no effect on the regeneration of new plants from the tobacco leaf (regeneration index is 0), however in combination with iP they act surprisingly synergistically and increase the efficiency of the iP, which is commonly used.
Table 2. Effect of compounds on the regeneration index (number of new plants per explant) of tobacco and the amount of retained chlorophyll in wheat leaves (in %). The values of the regeneration index for compounds 1-40 are the result of the synergistic effect of these compounds in 0.1 pM concentration and of cytokinin iP at the same concentration. The regeneration index value is the average of 15 explants (± standard error). The chlorophyll content value (see Example 12) is the average of 5 measurements of chlorophyll content in 5 leaves (± standard error). Comp. - compound; nt. - not tested; x - irrelevant.
Figure imgf000024_0001
Figure imgf000025_0001
Example 14. Anti-senescence synergistic effect of compounds of general formula (I) and (la) with /rans-zeatin in a senescence test on leaf segments of wheat cultivated in the dark. Seeds of winter wheat Triticum aestivum cv. Hereward was rinsed under running water for 4 hours and then seeded in vermiculite saturated with Knop's nutrient solution (Ca(NO3)2, MgSCU, KNO3, KH2PO4, all <1%). The pots were placed in an air-conditioned growth chamber with a 16/8 hour light period (light intensity 50 pmol.m^.s'1) and a temperature of 15 °C. After 7 days, the seedlings had developed the first flag leaf and the second leaf began to grow. The top sections of the first leaves were removed so that their weight was 25 mg. The basal ends of these leaf segments were placed in the wells of microtiter polystyrene plates containing 150 pL of a solution of a test derivative of formula (I) or (la) (1 leaf segment per well) or a combination thereof with trans- zeatin. The plates were placed in a plastic box with filter paper, which was saturated with water for maximum humidity. After 96 hours of incubation in the dark at 25 °C, the leaf segments were removed and the chlorophyll was extracted in 5 mL of 80 % ethanol by heating at 80 °C for 1 hour. After extraction, the sample volume was restored to 5 mL by adding 80 % ethanol. The absorbance of the extracts was measured at 665 nm. As a green control, freshly cut leaves were used, from which chlorophyll was extracted in the same way (this value represents 100 %). Leaves incubated in deionized water only for 96 hours in the dark served as a negative control (chlorophyll content in these leaves was set to be 0 %). The calculated values are the average of 5 replicates and the whole experiment was repeated at least 2 times. The effect of the compounds of formula (I) and (la) and /rans-zeatin was first tested at 1 and 10 micromolar concentrations.
Trans-zeatin applied alone at 1 micromolar concentration retained 13 % chlorophyll. The compounds of formula (I) and (la) applied alone at 1 micromolar concentration, did not show a significant positive effect on the chlorophyll retention in the wheat leaves (retained 0 to 5 % over the negative control). Trans-zeatin applied alone at 10 micromolar concentration was able to retain 28 % chlorophyll compared to the negative control. The compounds of formula (I) and (la) applied alone at a concentration of 10 micromolar were able to retain between 5 and 16 % of chlorophyll, thus showing a weak positive effect on the delay of senescence in the segments of wheat leaves caused by darkness. Results are summarised in Table 2.
Furthermore, the effect of the combination of /rans-zeatin at 10 micromolar concentration and the individual compounds of formula (I) and (la) at 1 micromolar concentration (at this concentration the substances were almost not active themselves) and 10 micromolar concentration on chlorophyll retention in excised wheat leaves incubated in the dark was tested. All newly prepared and tested compounds of formula (I) and (la) at 1 and 10 micromolar concentration showed a clear synergistic effect with /rans-zeatin at 10 micromolar concentration (see Table 2).
Example 15. Comparison of the influence of selected growth regulators and their combined application with iP on chrysanthemum regeneration
Tested growth regulators:
TDZ (thidiazuron; l-phenyl-3-(l,2,3-thiadiazol-5-yl)urea)
DPU (1,3 -diphenylurea)
Compound No. 24 of the present invention (l-(3,5-dichlorophenyl)-3-(3-(trifluoromethoxy) phenyl)urea)
Compound No. 29 of the present invention (l-(3-bromo-5-(trifluoromethoxy)phenyl)-3-(3,5- dichlorophenyl)urea) Compound No. 37 of the present invention (l-(3,5-dibromophenyl)-3-(3,5-dichlorophenyl) urea)
Method:
Chrysanthemum leaves were divided into small disks (d = 0.5 cm) and placed in petri dishes with 10 mL of MS medium (medium contained sucrose 30 g/L, plant agar 7 g/L, pH 5.8). This medium further contained the plant growth regulators tested or their combination with an iP. Compound concentrations and results are listed in Table 3. Each combination was performed in 3 replicates with 5 explants per combination. Plant regeneration was assessed after 3 weeks. Table 3: Effect of tested compounds on the regeneration index (number of new viable plants per explant) of chrysanthemum. The regeneration index value is the average of 15 explants (± standard error).
Figure imgf000027_0001
iP - N6-(2-isopentenyl)adenine; TDZ - thidiazuron, DPU - diphenylurea; * - there is an excessive formation of new plants (stems), which are degenerate and incapable of life. 1
The results show that the application of 1 pM iP or 0.1 pM TDZ leads to the regeneration of chrysanthemum. The 10 pM iP application does not lead to a significant increase in the number of regenerated plants. Application of a higher concentration of TDZ (1 pM) leads to increased formation of new plants, but they are degenerate and incapable of life (Figure 2A). The same phenomenon occurs after co-application of 1 pM iP with 1 pM TDZ. In contrast, DPU application alone has no effect on plant regeneration and in combination with 1 pM iP shows no synergistic effect. Compounds 24, 29 and 37 of the present invention do not affect the regeneration of chrysanthemum plants when applied alone. Surprisingly, however, when applied in combination with a 1 pM iP, a very strong synergistic effect is observed - approximately twice the number of newly regenerated viable plants (Figure 2B). The regeneration index of these compounds is more than double compared to the regeneration index of TDZ or DPU.
It is clear from the above that only the combination of iP with substances 24, 29 or 37 led to an increase in the number of newly regenerated and viable chrysanthemum plants. The example shows that the synergy between iP and the growth regulator is present only when compounds of the present invention are used. Neither TDZ nor DPU show any synergy.
Example 16. The effect of tested compounds on vitality of Caenorhabditis elegans
C. elegans cultivation
The Wild-type N2 (Bristol) Caenorhabditis elegans strain and bacteria Escherichia coli OP50 used in this experiment was obtained from the Caenorhabditis genetic center (CGC). Worms were cultivated in 20 °C on plates containing Nematode Growth Medium agar (for 1 L: 3.0g NaCl, 2.5g Pepton, 17g Agar; add water to 975 mb; after autoclaving add 0.5 mL of IM CaCh, 1 mL of 5mg/mL cholesterol in ethanol, 1 mL of IM MgSO4, 25 mL KH2PO4 pH = 6) seeded with Escherichia coli OP50. Bacteria were grown overnight in 37 °C in Lysogeny broth medium (for 1 L: 10 g tryptone, 5 g yeast extract, 10 g NaCl). Worms were age-synchronised before further use.
C. elegans synchronization
Worms were age-synchronized by bleaching procedure: Plates containing mostly pregnant adults were washed with 2 mL of M9 buffer (for 1 L: 6 g Na2HPO4, 3 g KH2PO4, 5 g NaCl, 0.25 g MgSO4 x 7H2O) and liquid was transferred into Eppendorf tubes, which were shortly spined on a microcentrifuge. Buffer was removed and replaced with bleaching solution (for 10 mL: 8 mL water, 1,5 mL 14% NaClO, 0,5 mL 10M NaOH). Tubes were placed on a shaker set to maximum rpm (1500) until all worms were dissolved (approximately 10 minutes), leaving only resistant chitin-containing eggs. Eggs were washed 3 times with M9 buffer and twice with S complete medium [for 1 L: 977 mL S-basal (5.9 g NaCl, 50 mL mL KH2PO4 pH = 6), 10 mL IM potassium citrate pH 6 (for 1 L: 268.8 g tripotassium citrate, 26.3 g citric acid monohydrate), 10 mL trace metals solution (for 1 L: 1.86 g Na2EDTA, 0.69 g FeSCU x 7H2O, 0.20 g MnCh x 4H2O, 0.29 g ZnSO4 x 7H2O, 0.016 g CuSO4), 3 mL IM CaCh, 3 mL IM MgSCL, 1 mL of 5mg/mL cholesterol in ethanol)]. Eggs were left to hatch overnight in 20 °C on a shaker (150 rpm) to LI larvae stadium.
Toxicity evaluation in Caenorhabditis elegans
LI larvae were diluted with S -complete medium to a final concentration of 200 - 300 worms per ml, fed with 3 mg/ml bacteria and distributed onto 96-well plates. Larvae were treated with the test compounds (in five different concentartions) or DMSO (negative control) in equal volume. Anthelmintic drug ivermectin was used as positive control. Populations were left to grow at 20 °C for 4 days. After this period, healthy worms would reach adulthood and start reproducing, thus producing eggs. The eggs produce enzyme chitinase to evolve in LI larvae stadium. The non- fluorescent (Anorogenic) chitinase substrate 4-methylumbelliferyl -D-N,N',N"- triacetylchitotrioside (20 pM) was added to the wells to detect the Auoroscent product of the chitinase reaction - 4-methylumbelliferon. The plates were incubated at 37 °C for Ih. The reaction was then stopped by the addition of alkaline buffer (I M glycine/1 M NaOH, pH 10.6), and the intensity of the Auorescence of 4-methylumbelliferon was measured on a plate reader. Prior to the assay, the plates were visually checked under a microscope, and abnormalities, such as toxicity or developmental impairments, were noted. The toxic effect of the tested compounds were performed in a wider concentration range to obtain the /C50 valeus. Those were calculated using Graphpad Prism. Experiments were repeated at least three times.
From the results obtained (Table 4) it is clear that all of the tested compounds inhibit reproduction and vitality of C. elegans in concentrations lower than 10 pM. All these compounds can be categorised as very toxic for this soil free-living nematode, thus could be used as anthelmintic agent. Table 4. The /C7o values of the tested compounds against C. elegans. compound IC50 (pM)
DMSO (0.1 %) Not toxic ivermectin < 0.1
4 1.8 ± 0.13
8 0.60 ± 0.26
13 0.34 ± 0.01
23 2.1 ± 1.7
24 1.1 ± 0.1
25 3.6 ± 2.3
28 0.68 ± 0.20
29 0.85 ± 0.27
37 8.9 ± 4.4
39 8.6 ± 1.9
Example 17. Formulations
The active ingredient in the formulations may 1 any compound of formula (I) or mixture of two or more compounds of general formula (I).
Al. Emulsifiable concentrates a) b) c) d) active ingredient mixture 5 % 10 % 25 % 50 % calcium dodecylbenzenesulfonate 6 % 8 % 6 % 8 % castor oil poly glycol ether 4 % - 4 % 4 %
(36 mol of ethylene oxide) octylphenol polyglycol ether 2 % - 2 % (7-8 mol of ethylene oxide) cyclohexanone 10 % 20 % arom. hydrocarbon mixture C9-C12 83 % 82 % 53 % 18 % Emulsions of any desired concentration can be tained from such concentrates by dilution with water.
A2. Solutions a) b) c) d) active ingredient mixture 5 % 10 % 50 % 90 % l-methoxy-3-(3-methoxypropoxy)propane 20 % 20 % polyethylene glycol MW 400 20 % 10 % - N-methyl-2-pyrrolidone - - 30 % 10 % arom. hydrocarbon mixture C9-C12 75 % 60 %
The solutions are suitable for use in the form of microdrops.
A3. Wettable powders a) b) c) d) active ingredient mixture 5 % 25 % 50 % 80 % sodium ligno sulfonate 4 % 3 % sodium lauryl sulfate 2 % 3 % - 4 % sodium diisobutylnaphthalenesulfonate 6 % 5 % 6 % octylphenol polyglycol ether 1 % 2 % - (7-8 mol of ethylene oxide) highly dispersed silicic acid 1 % 3 % 5 % 10 % kaolin 87 % 61 % 37 %
The active ingredient is mixed thoroughly with the adjuvants and the mixture is thoroughly ground in a suitable mill, affording wettable powders which can be diluted with water to give suspensions of any desired concentration.
A4. Coated granules a) b) c) active ingredient mixture 0.1 % 5 % 15 % highly dispersed silicic acid 0.9 % 2 % 2 % inorganic carrier 99.0 % 93 % 83 %
(AE 0.1 -1 mm) e.g. CaCCh or SiCh
The active ingredient is dissolved in methylene chloroide and applied to the carrier by spraying, and the solvent is then evaporated off in vacuo.
A5. Coated granules a) b) c) active ingredient mixture 0.1 % 5 % 15 % polyethylene glycol MW 200 1.0 % 2 % 3 % highly dispersed silicic acid 0.9 % 1 % 2 % inorganic carrier 98.0 % 92 % 80 % (AE 0.1-1 mm) e.g. CaCCh or SiCh The finely ground active ingredient is uniformly applied, in a mixer, to the carrier moistened with polyethylene glycol. Non-dusty coated granules are obtained in this manner.
A6. Extruder granules a) b) c) d) active ingredient mixture 0.1 % 3 % 5 % 15 % sodium lignosulfonate 1.5 % 2 % 3 % 4 % carboxymethylcellulose 1.4 % 2 % 2 % 2 % kaolin 97 % 93 % 90 % 79 %
The active ingredient is mixed and ground with the adjuvants, and the mixture is moistened with water. The mixture is extruded and then dried in a stream of air.
A7. Dusts a) b) c) active ingredient mixture 0.1 % 1 % 5 % talcum 39.9 % 49 % 35 % kaolin 60 % 50 % 60 %
Ready-to-use dusts are obtained by mixing the active ingredient with the carriers and grinding the mixture in a suitable mill.
A8. Suspension concentrates a) b) c) d) active ingredient mixture 3 % 10 % 25 % 50 % ethylene glycol 5 % 5 % 5 % 5 % nonylphenol polyglycol ether 1 % 2 % (15 mol of ethylene oxide) sodium ligno sulfonate 3 % 3 % 4 % 5 % c arboxy methy Icellulo se 1 % 1 % 1 % 1 % 37 % aqueous formaldehyde 0.2 % 0.2 % 0.2 % 0.2 % solution silicone oil emulsion 0.8 % 0.8 % 0.8 % 0.8 % water 86 % 78 % 64 % 38 %
The finely ground active ingredient is intimately mixed with the adjutants, giving a suspension concentrate from which suspensions of any desired concentration can be obtained by dilution with water.

Claims

32 CLAIMS
1. Derivatives of l-phenyl-3-yl-urea of the general formula (I),
Figure imgf000033_0001
wherein
Y is nitrogen or carbon atom;
R1 is selected from the group consisting of halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methylsulfanyl group may be independently substituted with halogen;
R2 is selected from the group consisting of hydrogen, halogen or methyl;
R3 is selected from the group consisting of hydrogen, halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methyl sulfanyl group may be independently substituted with halogen;
R4 is selected from the group consisting of hydrogen, halogen or methyl; with the proviso that when Y is nitrogen, then R1 is halogen or methoxy group; and with the proviso that the following compounds are excluded from the scope of claim 1: l-(3,5- dichlorophenyl)-3-(2,6-dichlorpyridin-4-yl)urea, l-(3,5-difluorophenyl)-3-(3-
(trifluormethylsulfanyl)phenyl)urea, l-(3,5-dichlorophenyl)-3-(3-
(trifluormethylsulfanyl)phenyl)urea, l-(3,5-difluorophenyl)-3-(3-methoxyphenyl)urea, l-(3- bromophenyl)-3-(3,5-dichlorophenyl)urea, l,3-bis-(3,5-dichlorophenyl)urea, 1 ,3-bis-(3,5- difluorophenyl)urea, l,3-bis(3-(trifluoromethoxy)phenyl)urea, l-(3-bromophenyl)-3- (phenyl)urea, l-(3-fluorophenyl)-3-(phenyl)urea, l-(3-methoxyphenyl)-3-(phenyl)urea, l-(3- fluorophenyl)-3-(3-methylphenyl)urea, l-(3-methylphenyl)-3-(2-chloropyridin-4-yl)urea, l-(3- chlorophenyl)-3-(2-chloropyridin-4-yl)urea, l-(phenyl)-3-(2-chloropyridin-4-yl)urea, l-(phenyl)- 3-(2-methoxypyridin-4-yl)urea, l-(phenyl)-3-(2-fluoropyridin-4-yl)urea, l-(phenyl)-3-(2- 33 bromopyridin-4-yl)urea, l-(phenyl)-3-(2,6-dichloropyridin-4-yl)urea, l-(3-fluorophenyl)-3-(2,6- dichloropyridin-4-yl)urea, l-(phenyl)-3-(2,6-dibromopyridin-4-yl)urea, l-(phenyl)-3-(2-chloro-6- methoxypyridin-4-yl)urea, 1 -(3 -fluorophenyl)-3 -(2-chloropyridin-4-yl)urea, 1 -(3 ,5 - difluorophenyl)-3-(2-chloropyridin-4-yl)urea.
2. Derivatives of l-phenyl-3-yl-urea of the general formula (I) according to claim 1, wherein R1 is selected from the group consisting of fluor, chlorine, bromine, methoxy group, methylsulfanyl group, trifluoromethoxy group and trifluormethylsulfanyl group.
3. Derivatives of l-phenyl-3-yl-urea of the general formula (I) according to claim 1 or 2, wherein R2 is selected from the group consisting of hydrogen, fluor, chlorine, bromine and methyl.
4. Derivatives of l-phenyl-3-yl-urea of the general formula (I) according to claim 1, 2 or 3, wherein R3 is selected from the group consisting of hydrogen, fluor, chlorine, bromine, methoxy group, methylsulfanyl group, trifluoromethoxy group and trifluormethylsulfanyl group.
5. Derivatives of l-phenyl-3-yl-urea of the general formula (I) according to any one of the preceding claims 1 to 4, wherein R4 is selected from the group consisting of hydrogen, fluor, chlorine, bromine and methyl.
6. Use of l-phenyl-3-yl-urea derivatives of the general formula (la)
Figure imgf000034_0001
in plant biotechnologies; wherein
Y is nitrogen or carbon atom; R1 is selected from the group consisting of halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methylsulfanyl group may be independently substituted with halogen;
R2 is selected from the group consisting of hydrogen, halogen or methyl;
R3 is selected from the group consisting of hydrogen, halogen, methoxy group, methylsulfanyl group, wherein the hydrogen atoms of the methoxy group and the methyl sulfanyl group may be independently substituted with halogen;
R4 is selected from the group consisting of hydrogen, halogen or methyl; with the proviso that when Y is nitrogen, then R1 is halogen or methoxy group.
7. Use according to claim 6 of the l-phenyl-3-yl-urea derivatives of the general formula (la), wherein l-phenyl-3-yl-urea derivatives are selected from the group consisting of l-(2,6- dichloropyridin-4-yl)-3-(3-(trifluoromethoxy)phenyl)urea (13), l-(2-brompyridin-4-yl)-3-(3- ((trifluormethyl)sulfanyl)phenyl)urea (16), l-(3,5-dichlorphenyl)-3-(3-
((trifluoromethyl)sulfanyl)phenyl)urea (19), l-(3-chloro-5-fluorophenyl)-3-(3-
(trifluoromethoxy)phenyl)urea (23), l-(3,5-difluorophenyl)-3-(3-(trifluoromethoxy)phenyl)urea (24), l-(3,5-dichloorphenyl)-3-(3-(trifluoromethoxy)phenyl)urea (25), l-(3,5-dichlorophenyl)-3- (3-fluoro-5-(trifluoromethoxy)phenyl)urea (26), l-(3-chloro-5-(trifluoromethoxy)phenyl)-3-(3- chlorophenyl)urea (27).
8. Use according to claim 6 or 7 of the l-phenyl-3-yl-urea derivatives of the general formula (la) for regeneration of plant tissue cultures.
9. Use of the l-phenyl-3-yl-urea derivatives of the general formula (la) for inhibiting senescence in whole plants, plant organs, tissues and/or cells.
10. Use of the l-phenyl-3-yl-urea derivatives of the general formula (la) to stimulate seedling growth and plant growth and development.
11. Use of the l-phenyl-3-yl-urea derivatives of the general formula (la) in preparations intended for the cloning of plant cells, tissues, organs and plants.
12. A preparation for treatment of plants, characterized in that it contains at least one l-phenyl-3- yl-urea derivative of the general formula (I) or (la) as defined in any one of the preceding claims 1 to 6 or a salt thereof with an alkali metal, ammonia or an amine in the form of a racemate or an optically active isomer, or an acid addition salt thereof; and at least one excipient selected from the group comprising solvents, solid carriers, surfactants, emulsifiers, dispersants, wetting agents, wetting agents, stabilizers, antifoaming agents, preservatives, viscosities, binders, adhesives, and fertilizers.
13. Use of the compounds of the general formula (I) and (la) or of the preparations according to claim 12 in agriculture as anthelmintic agents, preferably against Caenorhabditis elegans.
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