CS264313B2 - Cytokinin, a membrane active agent to increase plant productivity, protein nitrogen content and anion acceptance - Google Patents

Abstract

The composition contains as active substance 0.5 to 90% by weight of an imidazole derivative of the general formula I where R1 represents a hydrogen atom, a chlorine atom or an alkyl group with 1 to 4 carbon atoms substituted by a chlorine atom or a hydroxyl group, or an alkenyl group with 2 to 4 carbon atoms, an amino group, a methylamino group, a hydroxyl group, a mercapto group, a methylmercapto group or an alkoxy group with 1 to 4 carbon atoms, R2 represents an alkyl group with 1 to 4 carbon atoms substituted by a bromine atom and X is an inorganic or organic anion, preferably a chloride, bromide or sulfate anion, and in an amount of 10 to 95% by weight of a liquid and/or solid carrier and 1 to 15% by weight of a surfactant.

Description

The invention relates to a cytokinin, membrane-active agent for increasing plant productivity, protein nitrogen content and anion uptake, which is suitable as a substitute for kinetin and which contains as an active substance 0.5 to 90% by weight of an imidazole derivative of the general formula I ¹ --V^CH

^.ÁííÍh/ i,- ^r ~ C, %

Ι^.Λν> ^Η · ^ΗΧ 'NI r ₂

Jx where represents a hydrogen atom, a chlorine atom, an alkyl group with 1 to 4 carbon atoms optionally substituted by a chlorine atom or a hydroxyl group, an alkenyl group with 2 to 4 carbon atoms, a mercapto group, a hydroxyl group, a methylmercapto group, an alkoxy group with 1 to 4 carbon atoms, an amino group, a methylamino group or a dimethylamino group, further R ₂ represents an alkyl group with 1 to 4 carbon atoms optionally substituted by a bromine atom, and

X is an inorganic or organic anion, preferably a chloride anion, a bromide anion or a sulfate anion, and in an amount of 10 to 95% by weight of a liquid and/or solid carrier and 1 to 15% by weight of surfactants.

Chemicals have long been used in plant cultivation: in addition to artificial fertilizers and pest control products, in recent decades, agents that influence the growth and yield of cultivated plants have been increasingly used: collectively, these agents are referred to as regulators.

These regulators are mostly natural or influence and control various processes of plant growth (Lapja 34, 3, pp. 122 to 126, 1979).

synthetic hormones, which can be used (M. Kólcei, M. Nádasy: Kagyar Kémíkusok

Natural and artificial auxins (indole derivatives) influence the life processes of plants in many ways. Four main types of synthetic auxins are known: indole derivatives, naphthyl derivatives, phenoxy derivatives and benzoic acid derivatives. In practice, methyl ester of naphthyl acetic acid is used as a dusting agent to prevent potato germination, and potassium salt of naphthyl acetic acid is used to root seedlings and prevent premature fruit drop.

The second large group of regulators are gibberellins, which stimulate growth, induce flowering and break dormancy. They are also used in practice.

and

The third group is made up of cytokinins, whose action is very versatile. They stimulate cell division, delay aging processes, maintain application dominance and interrupt dormancy. Of the natural cytoquinones, seatin and lupin are known, but they are not used in practice.

The fourth group includes abscisins, which affect endogenous regulation of aging and, in addition, inhibit growth and promote organ separation. Although more than 70 compounds with a xanthonine structure and abscisin-like action have been synthesized to date, these compounds have not yet been used in practice.

The only gaseous hormone is ethylene, which is produced in large quantities during fruit ripening and which accelerates fruit ripening. In practice, 2-chloroethylphosphonic acid is used for this purpose.

In addition to plant hormones (auxins, gibberellins, cytokinins, growth inhibitors, abscisins, ethylene, etc.), numerous other biologically active substances are used in agriculture. The widespread use of 2-chloroethyltrimethylammonium chloride (CCC) for wheat and barley (US patent number 3,156,544) is worth mentioning.

Endogenous cytokinins (seatin, lupin) and synthetic cytokinins (kinethin, benzyladenine, etc.) increase yield and protein content, but are hardly used in agricultural technology due to their high production costs (F. Skoog, F. M. Strong, 0. 0. fríiller: Science 148, pp. 532 to 533, 1965).

Hungarian patent document number 162 937 describes germination stimulating, bud development promoting and ripening accelerating amide derivatives of 2-chloroethane/thiono/phosphonic acid. Hungarian patent document number 163 510 describes thiourea derivatives which influence fruit separation. Hungarian patent document number 164 190 describes ripening accelerating, senescence retarding, growth inhibiting halogenethanesulfinic acids, for example ichlorothynnylphine acid. According to Hungarian patent document number 164 512, c-ivo prel is treated by sowing with tois/dimethylammonium(-p-chlorobenzylphosphonate derivatives, thereby increasing grain yield and nitrogen uptake.

Hungarian patent documents Nos. 164,567 and 164,866 describe the ripening accelerating action of beta-haloethylsilanes.

Individual pyriiuidine derivatives (for example 2-methylthio-4-ethylamino-5-nitro-5-methylaminopyrimidine) and their salts inhibit germination and inhibit growth in length (Hungarian patent document number 164 885),

Sym-triazinedione derivatives suitable for influencing flowering and sexual reproduction of plants are described in Hungarian patent document number 167 576. Hungarian patent document number 168 916 also describes a composition influencing flowers and generative reproduction which contains allophanimidates.

According to Hungarian patent specification No. 170,761, emergence and growth are accelerated by numerous phthalimide derivatives, for example 3-trichloromethylphthalimide, 1-(3-chlorophthalimido)cyclohexylcarboxamide. Compounds used to increase the sugar content are known from Hungarian patent specification No. 162,520 (2-(aminoalkoxy)-4-phenylthiazole derivatives) and from Hungarian patent specification No. 165,576 (2,4-bis-(N'-methyl-4'-pyridinium-sym-triazine iodide)).

According to Hungarian patent document No. 165,576, it is used to stimulate the vegetative growth of sugar beet in addition to gibberellin, imidazoles and also in addition to fungicides, some herbicides in very small amounts. However, these agents have not been used in practice.

Both endogenous cytokinins (seatin, lupin) and synthetic cytokinins (quinatin, benzyladenine) (F. M. Strong: Topics in microbial chemistry, Wiley, New York 1958, F. Sttog,

F. M. Strong, F. M. Miller: Science 148, pp. 532-533, 1965) are exclusively 6-aminopurine derivatives, the main biological activity of which is the stimulation of the frequency of mitosis. In addition to this action, they have numerous side effects, also useful in agrotechnical and microbiological applications, for example:

a) Does both fresh and dry weight increase when growing callus and mericlone?

bj enrichment of organic latexes in vegetative and generative organs is more favorable?

c) is chlorophyll retention stimulated, thereby increasing its phytochemical effectiveness?

d) is leaf rejuvenation promoted and thus aging suppressed?

e) during phytosynthesis, carbon dioxide is fixed more strongly?

f) does the measurable photolytic efficiency increase with RTO decomposition?

g) the intensity of amino acid and protein synthesis increases, in particular there is a significant absolute and relative increase in the amount of protein nitrogen fractions soluble in 0.5% sodium chloride solution;

h) nucleic acid synthesis is intensified.

Adenine and special derivatives have so far only been prepared in the laboratory in minute quantities. Despite promising experimental results, cytokinins are not yet used in practice due to their high production costs. Among the numerous known growth-retarding substances, not one has a cytokinin effect. The known agents do not have a cytokinin effect, so they are not expected to have a cytokinin effect.

In research into increasing plant growth, productivity and protein content, as well as in experiments to increase yield, it was found that the composition according to the invention, which contains imidazole derivatives of the general formula I in an amount of 0.5 to 90% by weight, liquid and/or solid carriers in an amount of 10 to 95% by weight and 1 to 10% by weight of surfactants, is suitable for influencing physiological processes in plants.

Some imidazole derivatives of the general formula I are known from the literature (Auwers, Mausa; Chem. Ber. 61, 2 414, 1928; Davies Mamalis, Petrow, Sturgeon: J. Pharm. Pharmacoi, 3, 420, 1951; Weidenhagen, Train, Wegner, Nordstrom: Chem. Ber. 75, 1936, 1942; Pozharsky, Simonow: Žur. Obščej. Chlm. 33, 179, 1963); however, the effect of these compounds on plant productivity was also not known until now.

The compositions according to the invention contain, in addition to the carrier and surfactants, compounds of the general formula I and influence physiological processes in plants even in relatively small concentrations (1 to 200 ppm, or 0.5 kg of active substance per hectare).

It has been experimentally proven that the controlling effect of the composition according to the invention on plant productivity, protein synthesis and photosynthesis is stronger than the effect of chemically similar plant growth regulators according to Hungarian patent specification No. 170,761, which only affect plant growth, but not protein synthesis and photosynthesis.

The imidazole derivatives of the general formula I not only have the biological cytokinin activity manifested on cells, but unlike cytokinins, their production and use in agriculture are also economical. The compounds of the general formula I also have membrane activity (i.e. they stimulate the uptake of anions), which is completely absent in endogenous and synthetic cytokinins.

During the research of biological action, it was found that the regulatory effect on nitrogen exchange occurs in almost all tested plants, while the degree of influence on protein synthesis and photosynthesis depends on the type of plant treated.

Well-reproducible methods have been developed for biological testing of the new agent with regard to its action. The effect on protein synthesis is tested by isotope technique using labeled amino acids (glycine- ^1-14 C, glycine- ^2-14 C, methionine [S-methyl ^-14 c]). The stimulation of photosynthesis is also tested by isotope technique, by incorporating labeled carbon dioxide ( ¹⁴ _CO2 ).

The effect on nutrient uptake is measured by determining the incorporation of labeled phosphate ion, while the inhibition of chlorophyll breakdown is measured by extinction using methanol-prepared samples at 665 micrometers.

There is a perfect agreement between the results of laboratory tests and field trials. The composition according to the invention acts on the life processes of plants in this way.

a) the development of the axial link (intermode) is shortened (for example in wine), the number of leaves and the weight of leaf dry matter increase, the nitrogen-carbon ratio acquires a favorable smaller value;

b) by stimulating root growth (root factor), the growth of above-ground shoots is also intensified.

This shortens the vegetative growth period (for example, by 6 weeks in brewing barley) and increases the development of generative organs and harvest;

c) in cultures of perennial, leguminous plants (such as alfalfa) and in cultures of green forage crops (corn for silage, meadows), the uptake of nitrogen and phosphate from the soil increases, the protein content in the leaves increases significantly and the nitrogen-carbon ratio decreases (i.e. improves);

d) when treating corn, soybeans, malting barley, wheat and similar crops, grain yield increases, the total nitrogen content increases by approximately 20%, but the protein content increases to a greater extent;

e) when treating sugar beets, the yield of the tubers increases and their sugar content, measured polarometrically, also increases, so that a greater amount of sugar can be produced per unit area. The genetically determined sugar content in the beet is reached more quickly and thus harvesting can begin earlier;

f) if the vine is treated twice before flowering and then again before ripening, the average weight of the grapes, the weight of 1 000 grapes and the refractometrically detectable sugar content increase;

g) rooting of vines and fruit trees is promoted and the treatment also stimulates the differentiation of the central axis and shoots and budding and ripening, while also influencing the ripening (lignification) of shoots and thus their resistance to frost;

h) the compositions can also be used for growing plant tissue, for differentiation of callus organ tissue; their action is based on increasing the frequency of mitosis;

i) the composition according to the invention, when applied to cultivated plants by spraying, acts as an antidote that compensates for the phytotoxic effect of numerous herbicides.

Treatment with the composition according to the invention is possible before sowing seeds or before planting, but the composition can be used after sowing by spraying on the soil or on the plants themselves in their various stages of development; the method of use depends on which stage of development and which properties are to be influenced. Usually a single treatment is sufficient, but there are also cases, for example in the case of grapevines, when the treatment is repeated.

For treatment, generally minimal doses are required (depending on the composition 0.5 to 20 kg/ha, even 0.5 to 2 kg/ha), but the composition according to the invention, even when used in extremely high doses (20 to 30 kg/ha), has no phytotoxic effect on plants. It is advantageous that the composition according to the invention does not irritate the skin, does not lead to allergies and does not harm the eyes and is practically non-toxic (ΙΌ,-θ 2,500 mg/kg).

. Imidazolium derivatives of the general formula I. can be prepared by methods described in the literature. An appropriately substituted imidazole is reacted in a solvent, for example in acetone, in benzene or in toluene, at a temperature of 50 to 160 ° C with alkyl halides or with alkyl chloroformate. The imidazolium salt formed is filtered off, washed and dried. Depending on the method of its preparation, the compound may contain a little dialkylated product as an impurity. The amount of which is so small that it does not affect the action of the composition according to the invention. If the final product is to be free of dialkylated product, its preparation is carried out in two stages: first, N-alkylimidazole is prepared and from this, in a separate process, an imidazolium salt, for example halide or sulfate, is prepared.

The plant growth regulators according to the invention may be in solid form (dusting agent, granulate). In such a case, kaolin, bentonite, active silicic acid, gypsum, calcium carbonate, talc and similar substances are used as solid carriers and fillers.

Liquid compositions can also be prepared. Of these, compositions with oils are the most preferred. Application forms prepared from them can be applied to plants at any stage of development. Vaseline oil, wood oil, other vegetable oils, as well as non-phytotoxic fractions of mineral oils are suitable as diluents for oily compositions.

Both solid and liquid compositions contain surfactants. Cationic, anionic and nonionic surfactants can be used as surfactants.

Suitable cationic surfactants are quaternary ammonium compounds (cetyltrimethylammonium chloride), as anionic surfactants are salts of dodecylbenzenesulfonic acid, as anionic surfactants are polyoxyethylene alkylphenols, polyoxyethylene sorbitates, polyoxyethylene triglycerides and similar compounds, but mixtures of surfactants and mixtures of anionic and nonionic surfactants can also be used.

The method of preparing the compounds of general formula I is illustrated in Example 1, the preparation of the compositions according to the invention is illustrated in Examples 2 to 5, and the other examples describe the physiological action of the composition according to the invention.

Example

A 2000 ml flask equipped with a stirrer, thermometer, reflux condenser and dropping funnel is charged with 118.2 g (1 mol) of benzimidazole and 800 ml of toluene. 200 ml of isopropyl chloroformate are added with stirring, the temperature being maintained at 20 °C. After the addition is complete, the mixture is heated to 60 °C and stirred at this temperature for two hours. Stirring is continued for two more hours at 90 to 100 °C and then cooled to 15 to 20 °C. The off-white crystals are filtered off, washed and dried at 80 °C.

125.5 g (81.7%) of N-isopropylbenzimidazolium chloride are obtained, which has a melting point of 134-136°C.

Other compounds of formula I can be prepared by analogous methods.

Example 2

A laboratory-grade bead mill is filled with 48 g of technical grade (94%) N-isopropylbenzimidazolium chloride, 4 g of Ultrasil VN-3 as a filler, 20 g of Emulsogen M as an emulsifier and 128 g of technical grade vaseline oil. The mixture is ground after adding 400 g of glass beads with a diameter of 1.5 mm for one hour at a speed of 775 min ¹ . The mixture is then separated from the glass beads using a sieve with a mesh width of 1 mm. The oily preparation containing 20% N-isopropylbenzimidazolium chloride, diluted with water to 1%, forms a stable emulsion without sediment even after 12 hours. .

Example 3

11 g of technical grade (96%) N-isopropylbenzimidazolium chloride, 4 g of Ultrasil VN 3 as filler and 16 g of Emulsogen M as emulsifier and 170 g of vaseline oil are charged into a pearl mill for laboratory purposes. After adding 400 g of glass beads with a diameter of 1.5 mm, the mixture is ground for one hour at a speed of 775 rpm. The glass beads are separated on a sieve with a mesh width of 1 nm.

The product, diluted twenty times with water, forms a stable emulsion.

Example 4

53 g of N-isopropylbenzlmidazolium chloride, 37 g of Zeolex 444, 5 g of powdered sulphite lyes, 3.5 g of Supermittel 1 494 are charged into a powder mixer for laboratory purposes.

1.5 g of Netzer IS and 0.3 g of Tensia defoamer as an antifoaming agent. The substances are mixed and homogenized. Then the mixture is ground in an Ultraplex mill type Alpíne

100 LU.

If a 50% agent is diluted with water to 0.5%, 86 to 90% of the particles are still in suspension after half an hour. The wet residue on the sieve (DIN 100 sieve) is 0.5 to 1.5%.

and

Example 5

84.2 g of N-isopropylbenzimidazolium chloride, 9.5 g of talc, 5 g of powdered sulphite lye are charged into a laboratory powder mixer. 1 g of Netzer IS and 0.3 g of methylene blue. The substances are mixed and homogenized. The mixture is ground in a laboratory mill according to the previous examples. The wet residue on the sieve (DIN 100 sieve) is 2 to 4 mesh.

Example

Effects on tobacco callus growth

The composition according to the invention is diluted with water to a concentration of 10 ppm and mixed into the nutrient medium before incubation. Three-week incubation of tobacco callus is carried out on the treated nutrient medium and also on the untreated control medium. Daily growth, cell growth and mitotic rate are determined by the method of Skoog. The results are shown in Table I

a b u 1 k a I Treatment Daily Number of cells Mitosis rate increment mg g.10 ⁴ % 1. untreated control 16.0 227.7 100 2. N-/n-propyl/benzimidazolium bromide 22.8 327.5 143

It is evident from the table that treatment with the compositions according to the invention increases the daily growth and number of cells and the frequency of mitosis by 43%.

Example7

Effects on tobacco callus growth

The procedure is similar to that of Example 6, but for comparison, nutrient media with 1 ppm quinine are used. The composition according to the invention is used at a concentration of 2 ppm. The results are given in Table IX.

Table II

Number Treatment Callus dry weight mg Speed index growth % 1 chinetin 174.11 97.17 100 2 N-/n-propyl- benzimidazolium- chloride 194.09 116.89 119.0 Appendix 1 to 8 . '

Effect on growth of isolated callus from the root of Datura innoxia milia

The procedure is the same as in the previous examples and nutrient solution is used as a control.

264-313 soil containing kinethin at a concentration of 2 ppm. The results are shown in Table III. The substance of the invention, N-/n-propyl/benzimidazolium bromide, is listed in the table under the letter A.

) u 1 k a III Treatment Weight callus Dry land Value growth Growth rate fresh dry fresh dry g mg % mg/day chinetin 4,969 4 175.06 3.52 15.56 15.56 95.28 chinetin + A 6,577 2 209.13 3.17 20.92 18.78 128.10 untreated 0.396 8 28.08 7.07 0.32 1.65 1.97 A 0.612 0 43.29 7.07 1.04 3.09 6.36

Table IV shows the effect of the composition of the invention on the growth rate in percent.

Table IV

Treatment Control A Chinethin 100 134 Without quinine 100 322 : to 1 and d 9

Inhibition of chlorophyll breakdown in barley, soybeans, broad beans, alfalfa and corn

The composition according to the invention is diluted with water to a concentration of 200 ppm. A leaf segment of the tested plant weighing 200 mg is allowed to float on the surface of the obtained solution for 4 days in the absence of light. Then the chlorophyll is extracted from the leaf segments with methanol.

At the same time, leaf samples are extracted at the initial stage and after 4 days of standing on the surface of distilled water. The chlorophyll concentration in the methanolic extract is determined by measuring the extinction at 665 micrometers. The results are given in Table V.

Table V

Effect of various N-alkylbenzimidazolium bromides on chlorophyll content in alfalfa and corn

Compound of general formula I Alfalfa Maize increase in chlorophyll content

mg % mg % N-methyl derivative 0.83 6.4 0.80 23.0 N-ethyl derivative 0.85 8.9 0.79 21.5 N-bromoethyl derivative 0.64 - 0.71 9.2 N-n-propyl/derivative 0.96 23.0 0.82 26.1 N-/n-butyl/derivative 0.81 3.8 0.76 16.9 untreated 0.79 - 0.65 -

The chlorophyll content in 100 mg of alfalfa and corn in a green state was investigated. The results show that the compositions according to the invention inhibit the breakdown of chlorophyll.

Example 10

Effects on chlorophyll preservation

The effect of the composition according to the invention on the preservation of chlorophyll is compared with the effect of endogenous and synthetic cytokinins. Bean plants of the Pinto species are used as test plants. The effect is determined two weeks after treatment by measuring the extinction at 665 nm.

The values given in the following table show that at a concentration of 200 ppm the compounds according to the invention have a stimulating effect on chlorophyll. The effect is somewhat weaker than the effect of endogenous and synthetic cytokinin. Even at concentrations above 10,000 ppm the compositions according to the invention do not have a phytotoxic effect.

Table VI Treatment Concentration Ppm Extinction Stimulus· % untreated control - 0.671 seatin 20 1,283 91.3 kinethin 30 1,120 67.0 benzyladenine 30 1,229 83.1 A 200 1,081 61.2

Example 11 Stimulation of protein production in sc/ji

The composition according to the invention is diluted with water to a concentration of 200 ppm. 200 mg of soybean leaf segments are allowed to float on the surface of the solution for 24 hours. As controls, segments of the same weight are used, which are allowed to float on the surface of distilled water. The segments are then rinsed with distilled water and placed in a radioactively labeled amino acid (glycine- ^1-14 C; 2f1C1) solution (20 ml) for three hours. The leaf segments are then rinsed with a large amount of water and the protein is isolated from them in the usual way. A stock solution is prepared from the isolated protein and its specific radioactivity is measured by liquid scintillation counting. The values obtained are given in Table VII.

Table VII

Treatment Number

N-/n-propyl/benzimidazolium bromide

Radioactivity yU Ci/100 mg

13.7

Stimulation %

15.2

It is clear that protein formation in soybean leaves treated with the composition of the invention is 15% greater than in untreated control leaves.

Example

Effects on protein synthesis

Pinto bean leaves are left for 18 hours in a solution of natural or synthetic cytokinins and in the composition according to the invention. The effect on protein synthesis is measured by detecting the incorporation of a radioactively labeled amino acid (glycine-2-C). In Table VIII, the radioactivity is in 1000 dpm/200 mg fresh weight.

Table VIII

Treatment

Concentration

PPm

Radioactivity

000 dpm/200 mg in TCA insoluble fraction

Stimulation untreated seatin

3.17

6.18

95.1

AND

) at 1 k and VIII continued Treatment Concentration Radioactivity Stimulation ppm 1,000 dpm/200 mg in TCA insoluble fraction % kinethin 30 5.36 69.4 benzyladenine 30 5.92 87.0 A 200 4.52 42.7

As the table shows, the effect of the composition according to the invention is somewhat weaker than that of cytokinin, but considering that these are not plant hormones occurring in traces, but are readily available compounds, the effect is significant.

Example 13

Pieces of alfalfa, soybean, broad bean and corn leaves weighing 200 mg are allowed to float on the surface of the composition according to the invention, diluted with water to a concentration of 200 ppm, for half an hour.

Then the leaf pieces are introduced into the assimilation chamber of a suitable apparatus. While carefully controlling the illumination by means of a dosing valve, a known amount of radioactive carbon monoxide ( _CO2 ) is introduced from the chamber. After 30 minutes, the leaf samples are frozen in liquid nitrogen, ground in a Potter homogenizer with 50% perchloric acid and processed into a stock solution.

The specific radioactivity of the stock solutions is measured by liquid scintillator.

Untreated leaf samples also exposed to labeled carbon dioxide in the same assimilation chamber are used as controls. The amount taken up by untreated leaves

considered as 100%. In addition, the amount received is i b u 1 k a IX in the table IX states in percentages. Treatment Alphabet , Soybean Bob Corn additionally received amount of CO ₂ % % % % 1. N-methylbenzimidazolium- bromide 125 2. N-ethylbenzimidazolium- bromide 184 3. N-/bromethyl/-benz- imidazolium bromide - 57 4. N-/n-propyl/benz- imidazolium bromide 12 236 - 84 5. N-/n-butyl/benzimidazolium- bromide 41 47

It is clear from the table that the composition according to the invention affects cultivated plants to various but mostly to a large extent.

Example 14

Enhancement of carbon dioxide fixation

The intensity of fixation of photosynthetically bound carbon dioxide was measured on Pinto beans.

The leaf samples are allowed to float for 18 hours in the bioactive solution, fixation is performed by 14 half-hour exposure (at 100 µCi/5,000 ml of _CO2 ).

The following table X shows that seatin (root factor*) has the greatest effect, while synthetic cytokinins are somewhat worse and the composition according to the invention has the same type of effect, but not as strong. However, similar to example 12, it is recalled that according to the invention, it is not a matter of plant hormones occurring in traces, but of readily available compounds.

bulk X Treatment Concentration Radioactivity Growth in the light, in the dark, total ppm dpm/200 mg % untreated - 2.14 0.18 2.32 - seatin 20 5.12 0.46 5.58 140.8 kinethin 30 4.15 0.37 4.52 95.2 benzyladenine 30 4.87 0.41 5.28 128.0 A 200 3.92 0.34 4.26 84.0

Example 15

Stimulation of plant photosynthesis

The composition according to the invention is diluted with water to 500 ppm. Samples of corn leaves weighing 200 mg are treated in this solution for two hours. Then the samples are stored for one hour in RTO at a concentration of 10 µCl/50 ml, then frozen in liquid nitrogen, ground, chromatographically separated by appropriate treatment with glyceraldehyde and their radioactivity is measured. The results are shown in Table XI.

Table XI

Treatment Dry matter dpm/mg untreated control 4.64

A 8.76

Stimulation %

The measurement results show that the composition according to the invention effectively stimulates photolysis.

Example 16

Stimulation of photolytic efficiency

The photolytic activity of endogenous cytokinins (seatin), synthetic cytokinins (chinetin, benzyladenine) and regulators according to the invention is compared. The tests of Pinto bean leaves are left to float in vioactive solutions for 18 hours, the exposure to RTO-photolysis lasts one hour (200 ^uCi/100 ml). The radioactivity is given in dpm/mg of vinous sugar.

Table XIX Treatment Concentration ppm Photolytic efficiency of dextrose dpm/mg Stimulation % untreated - 7.64 - seatin 20 15.94 108 kinethin 30 13.96 82 benzyladenine 30 17.21 125 A 200 14.08 84

Example 17

Stimulation of nutrient uptake

The composition according to the invention is diluted with distilled water to a concentration of 200 ppm. Pieces of leaves weighing 200 mg of wheat, corn and peas are treated with the solution.

Then the leaf pieces are transferred to a labeled phosphate solution with a concentration of 30 ^uCi/50 ml and processed therein for half an hour. Then the leaf pieces are processed in the usual way. The nucleic acid fractions are isolated and dissolved in 6N ammonia. From the solution

a stock solution is prepared and its radioactive phosphate content is measured by liquid scintillation counting. The results are shown in Table XIII. Table XIII Treatment Wheat Corn Peas check 1 000 cpm/g 17.4 31.2 25.3 A 22.8 39.3 32.7 (+31.0 i) (+25.9%) (+29.2 The numbers in parentheses indicate in percentage stimulation in view of untreated control.

It is clear that treatment with the composition according to the invention increases the incorporation of a significant phosphate ion into the nucleic acid fraction by 26 to 30%, from which it can be inferred that there is an increase in nutrient uptake.

Example 18

Stimulation of membrane activity

The composition of the invention also has membrane-active action. This action has not been found with endogenous or synthetic cytokinins. Table XIV shows the stimulation of nitrate uptake for cytokinins and for compositions used according to the invention. Nitrate uptake is manifested by an increase in total nitrogen content and protein nitrogen content.

Experiments carried out on leaves of the Pinto bean plant; evaluation is carried out two weeks after treatment.

Table XIV

Treatment Concentration Dry substance contains nitrogen Protein total protein nitrogen in % of total ppm % % nitrogen untreated - 3.8 2.4 63 seatin 20 3.8 2.8 (+16.6) %) 73 kinethin 30 3.8 2.8 (+16.6) %) 73 benzyladenine 30 3.8 2.8 (+16.6) %) 73 A 200 5.3 (+39.4 %) 4.4 (+83.3) %> 83

The numbers in parentheses indicate the increase expressed as a percentage with respect to the untreated control.

Example 19

Stimulation of the effect on the content of total nitrogen and protein nitrogen in alfalfa

The compositions according to the invention are applied to alfalfa in the open field. The experiments are repeated four times on a plot of 1 hectare each. The composition according to the invention formulated with oil is diluted with water and sprayed once or twice on the plot. The first treatment is carried out when the alfalfa has already emerged and covers the soil. On plots intended for repeated treatment, another dose of the composition according to the invention is applied two weeks after the first treatment. In both cases, a quantity of 2 kg/ha is used.

After the alfalfa was cut, the total nitrogen and protein nitrogen content were determined. The results are shown in Table XV, where the numbers in parentheses indicate the percentage increase relative to the untreated control.

Table XV

Treatment Untreated Compound A

1st treatment

2nd treatment

Total nitrogen content in dry matter %

3.36

3.51 (+4 4)

3.91 (+13%)

Protein nitrogen content in dry matter %

2.25

2.48 (+22%) 2.75 (+22%)

As can be seen from the table, a single treatment increases the nitrogen content in the dry matter by 4%, the protein nitrogen content by 22%. The share of protein nitrogen in the total nitrogen content increases from 67% to 70%, that is, by 5%. With two repeated treatments, the nitrogen content in the dry matter increases by 13%, the protein nitrogen content by 22% and the share of protein nitrogen in the total nitrogen content increases by 5%.

Table XVI

Treatment Green matter yield Crude protein Ratio 100 kg/ha P ₂ Drawbar in dry matter C:N untreated 120 21.3 8.3 A 142 ⁺ (+18.7%) 25.8 ⁺⁺⁺ 10.9 ⁺⁺ 4.30' (+21.5%) (+31.7%) (-21.1

Significance limits:

+ ++ +++ ^SD 5% ^SD 1% ^SD 0.1% The numbers in parentheses indicate the difference from the untreated control in percent.

Example 20

Effects on total nitrogen content and protein nitrogen content in a mixed crop of sdji and sunflower

A mixed crop of soybeans and sunflowers, intended for green fodder, is treated with the composition according to the invention in a field test. The tests are carried out with 4 repetitions on plots of 1 hectare. The composition according to the invention is diluted with water and the spray emulsion is always applied in an amount of 2 kg/ha. The first spraying is carried out at the time when the emerged plants cover the soil. The second treatment is carried out two weeks after the first treatment and the third treatment is carried out two weeks after the second treatment. The green plant matter is harvested on untreated and treated plots and its total nitrogen content and nitrogen content in protein are determined.

Table XVII

Treatment untreated

AND

Nitrogen content in dry matter total protein % %

2.56 2.27

2.80 (+9.7%) 2.75 (+21.1%)

Total/protein nitrogen ratio 100%

65.2

98.9

The numbers in parentheses indicate the percentage increase relative to the untreated control.

The table shows that the treatment increases the total content by almost 10%, the protein nitrogen content by approximately 20%. The share of protein nitrogen in the total nitrogen content increases much more significantly, up to 52%. This improves the feed value of green fodder, which is very important for animal nutrition.

Example 21

Effect of the composition according to the invention on the weight of green matter, on the total nitrogen content and on the protein nitrogen content in silage corn

Experiments are carried out on a plot of 200 m2. The composition according to the invention is diluted with water and the field is treated with it immediately before sowing. The weight of harvested corn for silage from the treated and untreated plots is determined. The composition of the plant matter is also examined. The results are given in Table XVIII.

Table XVIII

Parameter Check Compound A green matter yield (kg) 365 389 dry matter content (%) (dried at air) 20.57 23.48 (+14.1%) crude protein (%) 1.85 2.25 ⁺ (+21.7%) true protein (%) 1.35 1.4 (+4.4%) total carbon content (%) 8.99 10.27 ⁺⁺⁺ total nitrogen content 0.30 0.86 ⁺ C:N ratio 30.86 28.67

The significance limits are the same as in Table XVI. The numbers in parentheses indicate the percentage increase in value with respect to the control. It is clear from the table that the weight of green matter is 10% higher, the dry matter content of the corn for silage is 14% higher and the crude protein content is 36%. The true protein content shows an increase of 4.4%.

The C/N ratio acquires a numerically smaller value, which means a favorable influence and shift of the nitrogen share. The results are summarized in Table XIX. The values, given in brackets, indicate the percentage difference from the untreated control.

Table XIX Treatment Weight of green matter Crude protein ^S 2°5 C/N in dry matter 100 kg/ha % % untreated 310.2 13.2 7.5 31.4 compound A 398.2 ⁺⁺⁺ 17.1 ⁺⁺ . 9.8 ⁺⁺⁺ 23.5 ⁺⁺⁺ (+28.4%) (+29.7%) (30.8%) (-25.4% Compound A is in this case N-/i-propyl/benzimidazolium chloride

The significance limits are the same as in Table XVI.

Example 22

Hybrid corn seed is treated with the composition according to the invention; the composition according to the invention is used as a moistened powder dressing in an amount of 0.5 kg/ha. After the fresh leaf has developed, a spraying is carried out in an amount of 3 kg/ha. The second field treatment is carried out when the flowers appear (1 kg/ha, the third field treatment is carried out at the time of ear maturity (2 kg/ha). Table XX shows the weights converted to corn de-kerned in May, as well as the weight loss of mature corn during storage.

Table XX Treatment Grain yield corn 100 kg/ha Increase in % untreated 132.8 - compound A 149.3 12.4 compound B 208.5 57.1

Compound B means N-/i-propyl/benzimidazolium chloride

The values show that the composition according to the invention increases the yield of corn grain significantly (by 12 to 57%).

Example 23

Effect on soybean yield

The soybean crop is sprayed three times during the growing season at a rate of 2 kg/ha. The first treatment is carried out at the beginning of flowering, the second at the opening of the tip of the flower cluster and the third two weeks after the second spraying. The grain yield is measured. The results are given in Table XXI.

Table XXI

Treatment Grain yield (100 kg/ha) Increase (%) untreated 29.33 compound A 38.67 30.0

The results show that the composition according to the invention significantly increases the yield of soybeans (by 30%).

Example 24

Effects on wheat grain yield and on total nitrogen and protein nitrogen content in grains

Trials are carried out on large plots of wheat of the Libelulla and Mexikoi 226 K varieties. The seed is treated in a similar manner to wet dressing, i.e. a coating is applied to the seed, the amount of active ingredient being 2 kg/ha. Immediately before sowing, the soil is treated with 2 kg/ha of active ingredient. After harvest, the yield, total nitrogen content and protein nitrogen content of the treated wheat and of the untreated wheat are compared. The results are given in Tables XXII and XXIII. Table XXII refers to the Libelulla wheat variety, Table XXIII to the Mexikoi 226 K variety.

Table XXII

Treatment Weight of grains from 100 ears g Stimulation % untreated 215.79 - compound A 357.73 65

It is clear from the table that the yield of wheat grain of the species Libelulla increases by 65% after using the composition according to the invention.

Table XXIII

Treatment Total nitrogen content Protein nitrogen in dry matter untreated compound A %

2.12

2.13 (+1.0 ») %

1.61

1.74 (+8.0 »)

Tests with wheat of the Mexikói 226 K variety show that, in addition to an insignificant increase in the total content, there is a significant increase in the protein nitrogen content.

Example 25

Effects on yield and sugar content in sugar beet

Sugar beet seeds are treated with the composition according to the invention, using 1 kg of the composition according to the invention per hectare. The emerged plants are sprayed twice with a dose of 2 kg/ha each time. The sugar content of the treated plants is higher (16.1% compared to 15%). After harvest, the tuber yield and sugar yield are determined. The results are shown in Table XXIV.

Table XIV

Treatment Yield 200 kg/ha Contents Profit Increase from sugar control % 100 kg/ha % untreated 541 16.00 87.56 - compound A 877 17.05 149.49 70

It is clear that the composition according to the invention significantly increases the sugar yield.

Example 26

Effects on grape yield and sugar content in grapes

The grapevine plants are treated with the composition according to the invention before flowering, then three weeks or six weeks after the first treatment. The composition diluted with water is applied by spraying, using 2 kg/ha.

In parallel, tests are being carried out with commercial plant growth regulators; CCC (chlorocholine chloride), Ethrel (Etafon: 2-chloroethylphosphonic acid) and Nevirol (N-phenylphthaliminic acid) are used. The results given in Table XXV refer to the vine species Pannonia,

Table XXVI shows the results with the species maiden grape from Egr.

Table XXV

Treatment Grape yield 100 kg/ha Sugar content % Sugar yield 100 kg/ha untreated 216 13.0 28.08 compound A 226 15.8 (+21.5 ») 35.7 (+27%) CCC 175 16.0 (+23.0 %) 28.0 Ethrel 194 14.0 (+7.5 «) 27.2 Nevirol 238 15.4 (+18.4 %) 36.52 (+30%)

Values in parentheses express percentage increases relative to the untreated control.

Table XXVI

Treatment Grape yield Sugar content Sugar yield 100 kg/ha % 100 1 kg/ha untreated 151.15 16.7 18.93 compound A 176.92 (+17.0%) 16.7 22.16 (+17.0%) CCC 144.61 17.0 18.44 - Ethrel 164.61 (+8.9 «) 15.7 19', 38 (+2.3 ») Nevirol 169.22 (+11.9%) 16.4 20.81 (+9.9%)

The results show the influence of the vine species on the effect of the composition according to the invention. The composition according to the invention stimulates the grape yield and sugar content in the Pannonia species, while in the case of the girl's grape species the sugar content does not increase, but the grape yield is 17% higher.

Example 27

Effects on the yield of protein nitrogen in pastures

Plots of 4 m2 are marked out on a grassy meadow and sprayed three times with the composition of the invention diluted with water. The treatments are carried out at intervals of two weeks and 2 kg/ha is used for each treatment. The grass is then cut and the green mass, total nitrogen content and protein nitrogen content of the cut grass are determined.

The results are shown in Table XXVII, with the numbers in parentheses indicating the percentage increase compared to the untreated control.

Table XXVII

Treatment Yield kg/4 m ² untreated 5.10 compound A 5.27

Total nitrogen %

4.22

5.36 (+27.0%)

Protein nitrogen %

2.80

3.54 (+26.4%)

Example 28

The composition according to the invention is sprayed once or twice on the common bean (Superalit variety), the effect on the yield (dry beans, i.e. they are not harvested like green vegetables) is shown in Table XXVIII.

Table

XXVIII

Yield increase %

Treatment

Yield 100 kg/ha

untreated 15.00 compound And, one treatment 20. 6. 1979 17.77 18.4 compound And, one treatment 6. 7. 1979 17.03 13.5 compound A/ treatment 20. 6. 1979 treatment 6. 7. 1979 19.52 30.1

Example 29

Some compounds of general formula I are compared with regard to their biological activity. The cytokinin-like activity of the compounds can be characterized by the intensity of photosynthetic fixation of carbon dioxide and the incorporation of radiocarbon-labeled amino acids into the TCA-insoluble protein fraction.

Table XXIX gives values for photosynthetic CO ₂ fixation in maize leaves and glycine-ž-^C incorporation into protein synthesis in beans for benzimidazolium salts with various nitrogen substituents. Radioactivity is given in yiCi/g fresh greens and the mean error of the mean value is not greater than + 7%. The samples examined were left for 18 hours in a solution of the active substance and were exposed radiobiologically for one hour.

Table XXIX

Treatment untreated

C0 ₂ fixation /uCi/g + %

5.72

Incorporation of glycine-2- ¹ ^C juCi/g + %

3.52

Table XXIX continued

Treatment Fixation ^iCi/g ¹⁴ what ₂ + % Incorporation of glycine-2- /iCi/g + % · N-/methyl/benzimidazolium chloride (m.p. 248 to 250 °C) 8.12 42 4.75 35 N-/ethyl/benzimidazolium chloride (m.p. 270 °C) 6.72 18 4.25 21 N-/n-propyl/benzimidazolium bromide (m.p. 160 to 162 °C) 10.52 84 4.96 41 N-/n-butyl/benzimidazolium chloride (mp 227 to 229 °C) Substituent. R ₂ 7.49 31 4.57 30 bromoethyl- 6.46 13 4.36 24 i-propyl-/chloride/ 10.92 92 4.85 35

Example 30

The intensity of photosynthetic fixation of carbon dioxide is determined for various N-/n-propyl/ benzimidazolium bromides (with different substituents in the meaning of the symbol R^). Pinto point plants are used as test plants, the solutions are used at a concentration of 200 ppm and the radioactivity is given in Table XXX in dpm/200 g fresh weight. The results are given in Table XXX.

XXX

Substitute Intensity of fixation carbon dioxide (in position 5/8 or in position 4/7) dpm % increase control 2.32 - methyl 4.26 84.0 ethyl 4.13 78.0 n-propyl 4.22 81.8 isopropyl 4.30 85.3 n-butyl 4.20 81.0 hydroxyethyl 4.10 76.7 chloroethyl 4.00 72.4 allyl 4.15 78.8 chlorine 4.20 81.0 bromine 4.10 76.7 mercapto 4.32 86.2 hydroxy 4.16 79.3 methylmercapto 4.00 72.4 methoxy 4.09 76.2 amino 3.37 45.2 methylamino 3.84 65.0 dimethylamino 3.76 62.0

Claims (1)

SUBJECT OF THE INVENTION Cytokinin, a membrane-active agent for increasing plant productivity, protein nitrogen content and anion uptake, characterized in that it contains, in an amount of 0.5 to 90% by weight, as active substance, an imidazole derivative of the formula I X® '2 where it is hydrogen, chlorine, C1-C4alkyl optionally substituted by chlorine or hydroxyl, C2-C4alkenyl, mercapto, hydroxy, methylmercapto, C1-C4alkoxy , amino, methylamino or dimethylamino, furthermore R1 represents a C1 -C4 alkyl group optionally substituted by bromine, and X represents an inorganic or organic anion, preferably a chloride, bromide or sulphate anion, in an amount of 10 to 95% by weight, and and / or a solid carrier and in an amount of from 1 to 15% by weight of surfactants.