RU2844593C1 - Biopreparation based on chitosan asparaginate for soil treatment - Google Patents

Abstract

FIELD: chemistry; agriculture.

SUBSTANCE: invention relates to chemistry and agriculture, specifically to use of a biopreparation based on chitosan asparaginate for soil treatment, where said biopreparation is an aqueous dispersion of chitosan asparaginate particles with size of 1200-1600 nm, obtained from an aqueous solution of chitosan with a molecular weight of 200 kDa in L- or D-aspartic acid, additionally containing silicon tetraglycerolate solution in 3-mol excess of glycerol, with following ratio of components: chitosan with molecular weight 200 kDa – 0.08-0.34 g/dl; L- or D-aspartic acid – 0.07-0.27 g/dl; silicon tetraglycerolate solution in 3-mol excess of glycerol – 0.08-0.32 g/dl; water – balance.

EFFECT: invention provides an increase in the content of nitrogen, carbon and hydrogen in the soil while maintaining the level of soil acidity, a decrease in the degree of structuring of the treated soil due to a decrease in its bulk density, increase of productivity of herbaceous crops of family leguminous, cereals, cruciferae, cabbages at cultivation on the treated soil and watering.

3 cl, 6 dwg, 4 tbl, 14 ex

Description

Field of technology to which the invention relates

The invention relates to agriculture and can be used in plant growing, forestry and fruit farming to improve the fertility and agrochemical properties of soils, as well as to increase the yield of agricultural crops.

State of the art

The widespread use of chemical fertilizers, fungicides, insecticides, herbicides, pesticides and synthetic plant protection products causes a serious imbalance in the environment and has an adverse effect on both natural ecosystems, in particular the soil ecosystem, and human health. In this regard, the use of safe biopreparations (bioagrochemicals) in agriculture will help solve the problems of environmental pollution and human health, improve soil fertility, and increase the yield and quality of agricultural crops.

Among the materials that have demonstrated high efficiency in seed priming, stimulating plant growth functions, protecting agricultural crops from pests, diseases, etc., we can highlight preparations based on the aminopolysaccharide chitosan (see Russian Federation Patent No. 2316963 under IPC class A01N 59/00, A01N 25/04, A01N 37/02, A61K 31/722, published on 20.02.2008; see Russian Federation Patent No. 2257711 under IPC class A01N 25/32, published on 10.08.2005). Chitosan is also of interest as a soil additive as a nutrient that increases the efficiency of chemical fertilizers and does not affect the beneficial soil microbiota, which can be considered as a “green” approach to agrobiotechnology (see the articles by Zimin Yu.A., Sroslov G.A., Postnov M.V. Application of chitosan-based biopreparations in agriculture // Natural systems and resources. 2018. Vol. 8. No. 3. P. 22-28; Riseh R.S., Hassanisaadi M., Vatankhah M., Babaki S.A., Barka E.A. Chitosan as a potential natural compound to manage plant diseases // Int. J. Biol. Macromol. 2022. 220. P. 998-1009). Chitosan nanomaterials used as a carrier for fertilizers act as plant immune system enhancers due to slow, controlled and targeted delivery of nutrients to plants (see the article Riseh R.S., Vazvani M.G., Kennedy J.F. The application of chitosan as a carrier for fertilizer: A review // Int. J. Biol. Macromol. 2023 V. 252. ID 126483). In addition, the use of nanotechnology to produce agronanochemicals can increase the stability and effectiveness of preparations, as well as reduce the concentration of the active substance.

It is known to obtain and use an agrochemical composition based on polydisperse chitosan (see Russian Patent No. 2675485 under IPC class A01N 43/16, published on 19.12. 2018). The composition contains from 0.001 to 70% of polydisperse chitosan and one or more non-phytotoxic acids or their salts, wherein the acid is selected from hydrochloric, sulfuric, phosphoric, lactic, formic, acetic, pyruvic, oxalic, malonic, succinic, adipic, citric, glutamic, benzoic, salicylic, 2-(indolyl-3)acetic, 4-(indolyl-3)butyric, sorbic, humic acids, methionine. To obtain the composition, chitosan with a molecular weight of 10 to 300 kDa and a degree of deacetylation of 65 to 98 mol.% is gradually introduced into depolymerization conditions under the action of mineral or organic acids, oxidizers, nitrosating agents or enzymes at a temperature of 50-140°C for 30-300 min. The invention makes it possible to increase the yield of agricultural crops, improve plant survival under stress conditions, and reduce the pesticide load during plant cultivation.

The disadvantages of the proposed agrochemical composition are that gradient depolymerization of the polymer at a high temperature, in some cases exceeding 100°C, is used to obtain polydisperse chitosan. In preferred variants, aggressive mineral acids (hydrochloric, sulfuric, phosphoric, nitrous), organic and inorganic peroxides, halogens are used for gradient depolymerization, which can significantly reduce both the environmental friendliness of the finished product and its biological activity. The use of the agrochemical composition to increase soil fertility in the method was not reported.

The organo-mineral fertilizer-ameliorant “Nara-2” is known for improving and healing soils (see Russian Federation Patent No. 2115641 under IPC class C05F 7/00, C05G 1/00, C09K 17/40, published on 20.07.1998). The fertilizer-ameliorant contains a mixture of peat and sapropel, zeolite tuff with a particle size of 1-5 mm, lime flour and chitin or its derivatives or chitosan or its derivatives in the following ratio of components, wt.%: mixture of peat and sapropel - 55.0-84.5, zeolite tuff - 10-30, lime flour - 5-10, chitin or its derivatives or chitosan or its derivatives - 0.5-5.0. The proposed agent has increased selective sorption activity for heavy metals and radionuclides, exhibits a high meliorating effect and helps to increase the fertility of soils.

The disadvantage of the ameliorant fertilizer is the alkaline environment of the soil due to the presence of limestone flour in its composition, as a result of which chitosan will not be able to pass into salt form and will be in an inactive, water-insoluble form.

The closest to the claimed one is a plant growth biostimulator made of chitosan aspartate (see Russian Patent No. 2782614 under class IPC A01N25/00, published on October 31, 2022), containing chitosan with a molecular weight of 200 kDa - 0.01-0.001 g/l, L -aspartic acid - 0.01-0.001 g/l, water - the rest. The plant growth biostimulator made of chitosan aspartate is obtained by the reaction of salt formation of chitosan with aspartic acid in an aqueous medium. Chitosan with an average viscosity molecular weight of 200 kDa and a deacetylation degree of 82±2 mol.% produced by ZAO Bioprogress, Russian Federation is used; L -aspartic acid obtained by biocatalytic synthesis using the E.coli strain VKPM 7188, produced by Bioamid CJSC, Russian Federation; distilled water. Chitosan powder with a molecular weight of 200 kDa and a deacetylation degree of 82 mol% and L -aspartic acid powder, while maintaining the molar ratio [chitosan]: [acid] = 0.8-1.3, are dispersed in 1 liter of distilled water at 40-50 °C using a magnetic stirrer for 2-3 hours until completely dissolved. The finished chitosan aspartate solution is dried by spray or freeze drying to obtain an air-dry powder with a humidity of no more than 8-10%. To obtain a biostimulant, air-dried chitosan aspartate powder is dissolved in water at a rate of 0.10-0.75 g of powder per 1 l of water for seed treatment or 0.01-0.1 g per 10 l of water for watering vegetative plants. The biostimulant increases the germination of test seeds (wheat, oats, carrots, arugula, beans, pumpkin), and also accelerates the growth and development of test plants (lettuce, spinach), which leads to an increase in their green mass and, accordingly, yield compared to the control (germination and watering using water).

The disadvantages of the biostimulant in the proposed form are the unspecified effect on the agrochemical and agrophysical properties of soils. In addition, the chitosan aspartate solution is unstable in terms of aggregation and sedimentation: opalescence is observed 5 days after solution preparation, and phase separation with precipitation is observed 7 days after solution preparation (see the articles Lugovitskaya TN, Shipovskaya AB, Shmakov SL, Shipenok XM Formation, structure, properties of chitosan aspartate and metastable state of its solutions for obtaining nanoparticles // Carbohydrate Polymers. 2022. Vol. 277. ID: 118773; Shipenok KM, Lugovitskaya TN, Shipovskaya AB Structure formation during the synthesis of chitosan L- and D -asparaginate nano-particles // Russian Journal of Physical Chemistry A. 2024. Vol. 98. No. 7. P. 1584-1591).

Disclosure of invention

The technical problem of the claimed invention consists in modifying soil with a biopreparation made from chitosan aspartate to improve the agrochemical and agrophysical properties of the soil while increasing the yield of herbaceous crops.

The technical result is an increase in the content of nitrogen, carbon and hydrogen in the soil modified with chitosan aspartate, while maintaining the soil acidity level, a decrease in the degree of structuring of the modified soil due to a decrease in its bulk density, an increase in the yield of herbaceous crops of the legume, cereal, cruciferous and brassica families when grown on the modified soil and watered with water.

Nitrogen is involved in the synthesis of plant proteins and molecules that transmit genetic information, promotes the growth of stems, leaves, flowers and fruits, and also develops the root system, which, accordingly, increases the absorption of nutrients by the plant. Activates metabolic processes in the plant's body: from chlorophyll photosynthesis to the absorption of vitamins. To increase the concentration of inorganic nitrogen in the soil, mineral nitrogen fertilizers are added, and organic nitrogen - amino acid fertilizers.

Carbon is part of the soil organic matter - a natural source of supplying plants with mineral nutrients, controls the processes of nitrogen fixation, denitrification, mineralization and immobilization of nitrogen, serves as a source of energy and nutrition for microorganisms, and is sensitive to the effects of fertilizers.

Hydrogen regulates the action of plant hormones such as auxin, cytokine, and also reduces nitrates to ammonia with subsequent conversion into useful organic compounds.

The degree of structuring determines the basic agrotechnical properties of the soil: looseness and porosity, which determine the moisture permeability, air permeability, density and moisture capacity of the soil; flowability, which determines the speed of horizontal movement of soil and groundwater near or directly in the root-inhabiting layer of plant communities.

To solve the stated problem and achieve the stated result, the biopreparation based on chitosan aspartate for soil treatment is an aqueous dispersion of chitosan aspartate particles with a size of 1200-1600 nm, obtained from an aqueous solution of chitosan with a molecular weight of 200 kDa in L- or D -aspartic acid, additionally containing a solution of silicon tetraglycerolate in a 3-molar excess of glycerol, with the following ratio of components:

chitosan with a molecular weight of 200 kDa 0.08-0.34 g/dl L- or D -aspartic acid 0.07-0.27 g/dl silicon tetraglycerolate in a 3-molar excess of glycerol 0.08-0.32 g/dl water the rest

When aspartic acid is contained in the D -form, the particle size of chitosan aspartate in the biopreparation is 1200-1400 nm, and when aspartic acid is contained in the L -form, it is 1400-1600 nm.

Thus, a biopreparation based on chitosan aspartate was obtained in the form of an aqueous dispersion of L- or D -chitosan aspartate particles with the preservation of the salt form of chitosan, which is responsible for the manifestation of high effects of biological activity of this biopolymer.

Implementation of the invention

The invention is explained by illustrations, which show:

Photo 1 shows a photo of Tríticum aestívum wheat shoots after 9 days of growth and vegetation in a specific soil of an urban ecosystem of an anthropogenically modified biome (urbostratozem), treated before planting test seeds with distilled water (Example 4 - (1)), aqueous dispersions of L -chitosan aspartate (Example 5 - (2)) or D -chitosan aspartate (Example 6 - (4)) with C _CTZ = 0.25 g/dl, C _{L - AspA} = 0.25 g/dl or C _{D - AspA} = 0.25 g/dl ([CTZ]:[AspA]=1.3), C _Si = 0.32 g/dl.

Photo 2 shows a photo of Pisum sativum pea shoots after 8 days of growth and vegetation in the “Good Helper Universal” soil mixture, treated before planting the test seeds with distilled water (Example 12 - (1)), aqueous dispersions of L -chitosan aspartate (Example 13 - (4)) or D -chitosan aspartate (Example 14 - (4)) with C _CTZ = 0.08 g/dl, C _{L - AspA} = 0.07 g/dl or C _{D - AspA} = 0.07 g/dl ([CTZ]:[AspA]=1.0), C _Si = 0.08 g/dl.

The biopreparation chitosan aspartate is obtained by the reaction of salt formation of chitosan with L- or D -aspartic acid in an aqueous medium, accompanied by counterionic condensation of the chitosan polycation with the counterion of the acid residue and spontaneous formation of hollow particles of L- or D -chitosan aspartate while maintaining the salt (charged) form of the biopolymer and, accordingly, its biological activity.

To obtain a biopreparation based on L- or D -aspartate of chitosan, the following are used: powdered chitosan (CP) produced by ZAO Bioprogress (Shchelkovo, Russian Federation) with an average-viscosity molecular weight of 200 kDa, a degree of deacetylation of 82 mol.% and a humidity of 8±1 wt.%; L -aspartic acid ( L -AspA) produced by ZAO Bioamid (Saratov, Russian Federation); D -aspartic acid ( D -AspA) purchased from ZAO Vekton (St. Petersburg, Russian Federation); silicon tetraglycerolate in a 3-molar excess of glycerol (Si(OGly) ₄ ), synthesized in laboratory conditions at the I. Ya. Postovsky Institute of Organic Synthesis, Ural Branch of the Russian Academy of Sciences (Yekaterinburg, Russian Federation); distilled water.

The process of dissolving chitosan in L- or D -AspA and obtaining an aqueous dispersion of L- or D -aspartate of chitosan is carried out on a setup consisting of a Labdevices HMS-100D (China) flask heater with a reflux condenser and a thermometer for temperature control, filter and cooling cells. For gravimetric measurements, an Ohaus Adventurer AR 1530 analytical scale is used, weighing accuracy ±0.002 g.

Pour 50 ml of distilled water into a 100 ml flask and heat to 50°C, then add the calculated amount of CTZ and stir for 20 min at 400 rpm to swell the polysaccharide particles. Then add the calculated amount of L- or D -AspA and 50 ml of water, stir under the same conditions (temperature, stirrer speed) for 2-3 hours until the CTZ is completely dissolved and filter through a Schott-160 filter. In a 100 ml flask, 25 ml of filtered aqueous solution of CTZ in L -AspA or CTZ in D -AspA are collected, the calculated amount of glycerol solution Si(OGly) ₄ , which is controlled gravimetrically, is added using an automatic pipette-dispenser, stirred under the same conditions (temperature, stirrer rotation speed) for 6 hours and cooled to room temperature in a heat-resistant glass with insulation at a cooling rate of 15 deg./hour.

Silicon tetraglycerolate is a biologically active gelling agent and a bioavailable source of the microelement Si, which improves the health and stress resistance of plants, helps plants fight fungal diseases and other infections, and increases crop yield and quality. In an aqueous environment, it undergoes hydrolysis and condensation to form silicon polyolate (a polysiloxane spatial network).

As a result of the sol-gel synthesis of Si(OGly) ₄ , an aqueous dispersion of chitosan L- or D -aspartate (CTZ⋅ L -AspA + _H2O or CTZ⋅ D -AspA + _H2O ) is formed, consisting of hollow particles coated with a polysiloxane shell, 1400-1600 nm in size for chitosan L -aspartate or 1200-1400 nm for chitosan D -aspartate. The aqueous dispersion of CTZ⋅ L -AspA or CTZ⋅ D -AspA is aggregationally and sedimentationally stable for up to 300 days of storage under room atmosphere conditions.

An aqueous dispersion of chitosan L- or D -aspartate is prepared at a chitosan concentration of C _CTZ = 0.08-0.34 g/dl, a concentration of L -aspartic acid C _{L - AspA} = 0.07-0.27 g/dl or D -aspartic acid C _{D - AspA} = 0.07-0.27 g/dl, maintaining a molar ratio of [CTZ]:[AspA] = 1.0-1.3, and a concentration of silicon tetraglycerolate in a 3-molar excess of glycerol C _Si = 0.08-0.32 g/dl.

To evaluate the effectiveness of the influence of the biopreparation L- or D -aspartate chitosan on the agrochemical and agrophysical properties of the soil, growth-stimulating activity (germination, height and weight of shoots) in relation to test plants of herbaceous crops of the legume, cereal, cruciferous, and cabbage families, an agrotechnical method of cultivating agricultural crops is used, based on an assessment of the effect of modifying the soil with the studied preparations on the growth and vegetation of test plants.

Two types of soil are used: specific soil of the urban ecosystem of the anthropogenically modified biome (urbostratozem) and the soil “Good Helper Universal”.

To modify the soil, an aqueous dispersion of HTZ L -AspA or HTZ D -AspA is added to the soil mixture pre-sifted through a sieve with a pore diameter of 0.1 mm at a rate of 300 ml of the modifier dispersion per 300 g of soil. The mixture is thoroughly mixed until a homogeneous state is obtained and left in a room atmosphere for 5 days until the soil mixture is completely dry. The soil mixture treated with water according to the method described above is used as a control. Samples of the control soil and the soil treated with the modifier are analyzed for functional and elemental composition using IR spectroscopy, elemental analysis, pH-metry and the gravimetric method for determining the bulk density of bulk objects.

IR spectra of soil samples are recorded on a B-Rad FTS 175 FTIR spectrometer in the range of 400-5000 cm ^-1 with a spectral resolution of 0.5 cm ^-1 and an absolute error of ±0.1 cm ^-1 .

Elemental analysis of soil samples for nitrogen, carbon and hydrogen content is carried out on a CHN EA 3000 elemental analyzer (EuroVector Instruments, Italy) with an accuracy of ±0.3% (absolute error limits).

Determination of the pH of the aqueous extract of soil samples is carried out in accordance with GOST 26583-85.

The bulk density of soil samples is determined by the gravimetric method. The soil sample is poured through a funnel into a pre-weighed 50 ml cylinder and weighed on an analytical balance. The bulk density (kg/ ^m3 ) was calculated using the ratio m / V , where m is the mass of the soil, kg; V is the volume of the soil, ^m3 .

Bulk density allows us to give a generalized description of such physical properties of soil as looseness, porosity, density, flowability, water permeability, moisture capacity, and air exchange.

Growth-promoting activity is established in vivo in relation to herbaceous dicotyledonous test plants using the example of Brussels sprouts Brassica oleracea var. Gemmifera , radish Raphanus raphanistrum subsp. Sativus and pea Pisum sativum , as well as in relation to monocotyledonous test plants using the example of wheat Tríticum aestívum .

Seeds of 50 test plants are planted in special plastic cups filled with 20 g of moistened original (control) and modified soil, and grown at 22±2°C for 8-9 days. Watering is carried out with water at room temperature on the day of planting, then as the soil dries to a depth of 1.0-1.5 cm. The growth-stimulating effect is assessed by the change in seed germination, height and weight of vegetative shoots of test plants in the modified soil compared to the original, as well as by the volume of irrigation water spent during the growing season.

Group of examples of evaluation of functional and elemental composition of soil. Physicochemical characteristics of the original soil and modified with water dispersion of L- and D -aspartate chitosan are given in Table 1.

Example 1. The original soil of an urban ecosystem of an anthropogenically modified biome is watered at a rate of 300 ml of water per 300 g of soil, mixed until smooth, dried in room atmosphere for 5 days until completely dry, the chemical and elemental composition is analyzed using IR spectroscopy and elemental analysis, the hydrogen index of the aqueous extract and bulk density are determined. The main absorption bands in the IR spectrum of the original soil, ν/cm: 3696, 3620 (free OH); 3414 (OH); 2922, 2852 (C-H); 1639 (C=O); 1031 (C-O); 800-480 - "fingerprint" region. The content of elements in the original soil, %: nitrogen - 0.75, carbon - 12.41, hydrogen - 1.52. The pH of the aqueous extract is 6.5, the bulk density is 2.1×10 ³ kg/m ³ .

Example 2 is performed similarly to Example 1. The difference is that the original soil (urbostratozem) is treated with an aqueous dispersion of chitosan L -aspartate with a chitosan concentration of C _CTZ = 0.25 g / dl, a concentration of L -aspartic acid C _{L - AspA} = 0.25 g / dl ([CTZ]: [AspA] = 1.3) and a concentration of silicon tetraglycerolate in a 3-molar excess of glycerol C _Si = 0.32 g / dl. The main absorption bands in the IR spectrum of the modified soil, ν / cm: 3696, 3620 (free OH); 3409 (OH, decreased intensity compared to the control according to Example 1); 2920, 2851 (C-H); 1638 (C = O); 1032 (C-O); 780-390 - "fingerprint" area. The content of elements in the modified soil, %: nitrogen - 0.83, carbon - 14.28, hydrogen - 2.27. The pH of the aqueous extract is 6.4, the bulk density is 1.9×10 ³ kg/m ³ .

Example 3 is performed similarly to Example 1. The difference is that the original soil (urbostratozem) is treated with an aqueous dispersion of chitosan D -aspartate with a chitosan concentration of C _CTZ = 0.25 g / dl, a concentration of D -aspartic acid C _{D - AspA} = 0.25 g / dl ([CTZ]: [AspA] = 1.3) and a concentration of silicon tetraglycerolate in a 3-molar excess of glycerol C _Si = 0.32 g / dl. The main absorption bands in the IR spectrum of the modified soil, ν / cm: 3696, 3620 (free OH); 3429 (OH, decreased intensity compared to the control according to Example 1); 2920, 2850 (C-H); 1636 (C = O); 1031 (C-O); 800-390 - "fingerprint" area. The content of elements in the modified soil, %: nitrogen - 0.81, carbon - 14.20, hydrogen - 2.26. The pH of the aqueous extract was 6.4, the bulk density was 1.8×10 ³ kg/m ³ .

Comparative analysis of the physicochemical characteristics of the original soil and the soil modified with L- or D -aspartate of chitosan according to examples 1-3 shows that the functional composition and acidity of the soil colloid do not change during modification. At the same time, during modification of the soil, the content of organic nitrogen (by 8.0-10.7%), carbon and hydrogen increases, and spontaneous (i.e. without the use of mechanical destruction and the introduction of disintegrants, such as sand) soil loosening (decrease in the degree of structuring) occurs, which is expressed by a decrease in the values of bulk density and is confirmed by IR spectroscopy (a decrease in the intensity of the absorption band in the IR spectra in the frequency range of 3900-3200 cm ^-1 indicates a decrease in the number of hydrogen bonds).

A group of examples of assessing the growth and vegetation of test plants in a specific soil of an urban ecosystem of an anthropogenically modified biome (urbostratozem): the original and modified aqueous dispersion of L- or D -aspartate chitosan.

The characteristics of the used biopreparation are given in Table 2. The evaluation of germination, height and weight of shoots, as well as a comparative analysis of the volume of irrigation water spent when growing test plants in the original urbostratozem and modified with an aqueous dispersion of L- or D -aspartate of chitosan are given in Table 3.

Example 4. Wheat seedsTriticum aestivumplanted in specific soil of the urban ecosystem of anthropogenically modified biome (urbostratozem), processed according to example 1, water until the soil is moistened to a depth of 1.0-1.5 cm, then similarly every 2 days of vegetation and growth of the test plant (1). The germination of test seeds on the 4th day was 76%. The height and weight of shoots after 8-9 days of growth of the test plant was 8.8 mm and 0.14 kg. The total volume of irrigation water for 8-9 days of vegetation of the test plant was 40 ml. Wheat shootsTriticum aestivumafter 9 days of growth and vegetation in the original urbostratozem are shown in photo 1, example 4 - (1).

Example 5 is performed similarly to example 4. The difference is that the soil mixture is treated according to example 2 with an aqueous dispersion of chitosan L -aspartate with C _CTZ = 0.25 g/dl, C _{L -AspA} = 0.25 g/dl, [CTZ]:[AspA] = 1.3 and C _S = 0.32 g/dl (2) or with C _CTZ = 0.13 g/dl, C _{L -AspA} = 0.13 g/dl, [CTZ]:[AspA] = 1.3 and C _Si = 0.16 g/dl (3). The germination of test seeds on the 4th day for samples (2) and (3) was 86 and 82%, respectively, which is 13.2 and 7.9% higher than the control. The height of the test plant shoots after 8-9 days of growth for samples (2) and (3) was 13.5 and 9.9 mm, which exceeded the green mass increase by 53.4 and 12.5% compared to the control, respectively, the shoot weight was 0.17 and 0.16 kg, which increased the yield by 21.4 and 14.3% compared to the control. The total volume of irrigation water for 8-9 days of test plant vegetation was 35 ml. Tríticum aestívum wheat shoots after 9 days of growth and vegetation in urbostratozem modified with an aqueous dispersion of chitosan L -aspartate with C _CTZ = 0.25 g / dl and C _{L - AspA} = 0.25 g / dl are shown in Photo 1, example 5 - (2).

Example 6 is performed similarly to example 4. The difference is that the soil mixture is treated according to example 3 with an aqueous dispersion of chitosan D -aspartate with C _CTZ = 0.25 g/dl, C _{D -AspA} = 0.25 g/dl, [CTZ]:[AspA] = 1.3 and C _Si = 0.32 g/dl (4) or C _CTZ = 0.13 g/dl, C _{D -AspA} = 0.13 g/dl, [CTZ]:[AspA] = 1.3 and C _Si = 0.16 g/dl (5). The germination of test seeds on the 4th day for samples (4) and (5) was 90 and 96%, respectively, which is 18.4 and 26.3% higher than the control. The height of the test plant shoots after 8-9 days of growth for samples (4) and (5) was 16.0 and 11.4 mm, which exceeded the green mass increase by 81.8 and 29.6% compared to the control, the shoot weight was 0.23 and 0.17 kg, which increased the yield by 64.3 and 21.4% compared to the control. The total volume of irrigation water for 8-9 days of test plant vegetation was 32 ml. Tríticum aestívum wheat shoots after 9 days of growth and vegetation in urbostratozem modified with an aqueous dispersion of chitosan D -aspartate _{with CCTZ} = 0.25 g / dl and C _{L - AspA} = 0.25 g / dl are shown in Photo 1, example 6 - (4).

Example 7 is similar to example 4. The difference is that the seeds of Brussels sprouts Brassica oleracea var. Gemmifera (1) are planted. The germination of test seeds on the 4th day was 50%. The height and weight of shoots after 8 days of growth of the test plant were 7.0 mm and 0.056 kg. The total volume of irrigation water for 8 days of vegetation of the test plant was 50 ml.

Example 8 is performed similarly to Example 7. The difference is that the soil mixture modified according to Example 5 is used. The germination of test seeds on the 4th day for samples (2) and (3) was 53 and 60%, respectively, which is 6.0 and 20.0% higher than the control. The height of the test plant shoots after 8 days of growth for samples (2) and (3) was 8.5 and 8.3 mm, which was 21.4 and 18.6% higher than the increase in green mass compared to the control, the shoot weight was 0.061 and 0.072 kg, which increased the yield by 8.9 and 28.6% compared to the control. The total volume of irrigation water for 8 days of test plant vegetation was 40 ml.

Example 9 is performed similarly to example 4. The difference is that the seeds of the common radish Raphanus raphanistrum subsp. Sativus (1) are planted. The germination of the test seeds on the 4th day was 80%. The height and weight of the shoots after 9 days of growth of the test plant were 9.0 mm and 0.16 kg. The total volume of irrigation water for 9 days of vegetation of the test plant was 50 ml.

Example 10 is performed similarly to example 9. The difference is that the soil mixture modified according to example 5 with C _HTZ = 0.25 g / dl, C _{L -AspA} = 0.25 g / dl and C _Si = 0.32 g / dl (2) is used. The germination of test seeds on the 4th day was 100%, which is 25.0% higher than the control. The height of the shoots of the test plant after 9 days of growth was 10.6 mm, which was 17.8% higher than the increase in green mass compared to the control, the shoot weight was 0.18 kg, which increased the yield by 12.5% compared to the control. The total volume of irrigation water for 9 days of vegetation of the test plant was 50 ml.

Example 11 is performed similarly to example 9. The difference is that the soil mixture modified according to example 6 with _CHTZ = 0.25 g / dl, C _{D -AspA} = 0.25 g / dl and C _Si = 0.32 g / dl (4) is used. The germination of test seeds on the 4th day was 100%, which is 25.0% higher than the control. The height of the shoots of the test plant after 9 days of growth was 11.3 mm, which was 25.6% higher than the increase in green mass compared to the control, the shoot weight was 0.19 kg, which increased the yield by 18.8% compared to the control. The total volume of irrigation water for 9 days of vegetation of the test plant was 50 ml.

A group of examples of growth and vegetation assessment of test plants in the soil mixture "Dobryi Pomoshchnik Universal": the original and modified with an aqueous dispersion of L- or D -aspartate chitosan. The characteristics of the used biopreparation are given in Table 2. The assessment of germination, height and weight of shoots, as well as a comparative analysis of the volume of irrigation water spent when growing test plants in the original and modified with an aqueous dispersion of L- or D -aspartate chitosan soil mixture "Dobryi Pomoshchnik Universal" are given in Table 4.

Example 12. is performed similarly to example 4. The difference is that the soil mixture "Good Helper Universal" is used, processed according to example 1, and the seeds of Pisum sativum peas are planted (1). The germination of test seeds on the 8th day was 68%. The height and weight of shoots after 8 days of growth of the test plant were 67 mm and 0.21 kg. The total volume of irrigation water for 8 days of vegetation of the test plant was 100 ml. Shoots of Pisum sativum peas after 8 days of growth and vegetation in the original soil mixture are shown in photo 2, example 12 - (1).

Example 13 is performed similarly to example 12. The difference is that the soil used is treated according to example 2 with an aqueous dispersion _{of chitosan L-aspartate with CCTZ = 0.34 g/dl, C L -AspA = 0.27 g/dl, [CTZ]:[AspA]=1.0 and C Si = 0.32 g/dl (2) or CCTZ} = 0.17 _g / _{dl ,} C _L -AspA = _{0.13 g} /dl, [CTZ] _: [AspA]=1.0 and C _Si = 0.16 g/dl (3) or CCTZ = 0.08 g/dl, C _{L -AspA} = 0.07 g/dl, [CTZ]:[AspA]=1.0 and C _Si = 0.08 g/dl (4). The germination of test seeds on the 8th day for samples (2), (3) and (4) was 75, 75 and 79%, respectively, which is 10.3, 10.3 and 16.2% higher than the control. The height of the test plant shoots after 8 days of growth for samples (2), (3) and (4) was 74, 88 and 80 mm, which was 11.4, 31.3 and 19.4% higher than the increase in green mass compared to the control, the shoot weight was 0.27, 0.28 and 0.28 kg, which increased the yield by 28.6, 33.3 and 33.3% compared to the control. The total volume of irrigation water for 8 days of test plant vegetation was 80 ml. Shoots of Pisum sativum peas after 8 days of growth and vegetation in the “Universal Good Helper” soil modified with an aqueous dispersion of L -chitosan aspartate with C _CTZ = 0.08 g/dl and C _{L - AspA} = 0.07 g/dl are shown in photo 2, example 13 - (4).

Example 14 is performed similarly to example 12. The difference is that the soil used is treated according to example 3 with an aqueous dispersion _{of chitosan D-aspartate with CCTZ = 0.34 g/dl, C D -AspA = 0.27 g/dl, [CTZ]:[AspA]=1.0 and C Si = 0.32 g/dl (2) or CCTZ} = 0.17 _g / _{dl ,} C _D -AspA = _{0.13 g} /dl, [CTZ]:[AspA]=1.0 and C _Si = 0.16 g/ _{dl (3) or CCTZ} = 0.08 g/dl, C _{D -AspA} = 0.07 g/dl, [CTZ]:[AspA]=1.0 and C _Si = 0.08 g/dl (4). The germination of test seeds on the 8th day for samples (2), (3) and (4) was 78, 75 and 81%, respectively, which is 14.7, 10.3 and 19.1% higher than the control. The height of the test plant shoots after 8 days of growth for samples (2), (3) and (4) was 74, 91 and 75 mm, which was 11.4, 35.8 and 12.0% higher than the increase in green mass compared to the control, the shoot weight was 0.30, 0.28 and 0.27 kg, which increased the yield by 42.9, 33.3 and 28.6% compared to the control. The total volume of irrigation water for 8 days of test plant vegetation was 80 ml. Shoots of Pisum sativum peas after 8 days of growth and vegetation in the “Good Helper Universal” soil modified with an aqueous dispersion of D -chitosan aspartate with C _CTZ = 0.08 g/dl and C _{L - AspA} = 0.07 g/dl are shown in photo 2, example 14 - (4).

A comparative in vivo analysis using two types of soil (urbostratozem, fertile soil) in examples 4-14 shows that dispersions of L- or D -aspartate chitosan, when introduced into a soil colloid, have a highly effective growth-stimulating effect on dicotyledonous and monocotyledonous test plants of herbaceous crops of the legume, cereal, cruciferous, and brassica families.

When growing wheat Tríticum aestívum , Brussels sprouts Brassica oleracea var. Gemmifera and radish Raphanus raphanistrum subsp. Sativus in urbostratozem treated with an aqueous dispersion of L- or D -aspartate of chitosan with a polymer and acid concentration of 0.13-0.25 g/dl, a Si(OGly) ₄ concentration of 0.16-0.32 g/dl compared to unmodified soil (control) showed an increase in test seed germination by 8-26% (wheat), 6-20% (cabbage) and 25% (radish), green mass by 12-82% (wheat), 18-21% (cabbage) and 17-26% (radish), yield by 14-64% (wheat), 9-29% (cabbage) and 12-19% (radish), as well as a decrease in the volume of irrigation water by an average of 12% (wheat) and 20% (cabbage), see examples 4-11.

When growing Pisum sativum peas in the “Good Helper Universal” soil treated with an aqueous dispersion of L- or D -chitosan aspartate with a polymer concentration of 0.08-0.34 g/dl, an acid concentration of 0.07-0.27 g/dl and a Si(OGly) ₄ concentration of 0.08-0.32 g/dl, compared to the control, an increase in the germination of test seeds by 10-19%, green mass by 11-36%, yield by 28-43%, as well as a decrease in the volume of irrigation water by an average of 20% are observed, see examples 12-14.

It has been experimentally established that when using the ratio of components (chitosan, L- or D -aspartic acid, Si(OGly) ₄ ), taken in greater or lesser quantities, declared in examples 1-14 intervals, the technical result of the invention is not achieved. With a lower content of C _HTZ , C _{L -AspA} or C _{D -AspA} , Si(OGly) ₄ , the germination of test seeds, green mass and yield of test plants differ little from their germination, increase in green mass and yield in the original unmodified soil, with a higher content, the acidity of the soil increases, which has a negative effect on soil fertility and plant life.

Thus, in the invention, when assessing the effect of modifying the soil with an aqueous dispersion of L- or D -aspartate of chitosan, a significant increase in soil fertility is shown, in particular, an improvement in the agrochemical, agrophysical and agrobiological properties of the soil, expressed in an increase in the content of nitrogen, carbon and hydrogen with an unchanged acidity of the soil colloid, a decrease in the degree of structuring and bulk density of the soil, as well as an increase in the yield of crops of herbaceous plants of the legume, cereal, cruciferous and cabbage families.

The biopreparation based on an aqueous dispersion of L- or D -aspartate chitosan is easy to use, does not require specific conditions for production and is highly stable during storage. The components used to obtain the biopreparation (chitosan, L- or D -aspartic acid) belong to the class of biologically active substances and do not harm the environment, animals or humans.

Table 1

Physicochemical characteristics of soil of urban ecosystem of anthropogenically modified biome: original and modified with aqueous dispersion of nanostructured L- or D -aspartate chitosan

Example No. Modifier Concentration of modifier components, g/dl Molnoe
ratio
[HTZ]:[AspA] Content of elements, % pH Bulk density (×10 ^-3 ), kg/m ³ HTZ AspA Si(OGly) ₄ Nitrogen Carbon Hydrogen 1 _H2O (control) - - - - 0.75 12.41 1.52 6.5 2.1 2 HTZ· L -AspA
+
_H2O 0.25 0.25 0.32 1.3 0.83 14.28 2.27 6.4 1.9 3 HTZ· D -AspA +
_H2O 0.25 0.25 0.32 1.3 0.81 14.20 2.26 6.4 1.8

Table 3

The effect of modification of specific soil of urban ecosystem of anthropogenically altered biome (urbostratozem) with aqueous dispersion of nanostructured L- or D -aspartate chitosan on growth and vegetation of test plants of wheat Tríticum aestívum , Brussels sprouts Brassica oleracea var. Gemmifera , radish Raphanus raphanistrum subsp. Sativus and on the volume of irrigation water consumed

Example No. Soil modifier Germination
on the 4th day of vegetation, % Increase in germination compared to control, % Vegetation time, days Total volume of _H2O for irrigation during the growing season, ml Growth-promoting activity Height of shoots, mm Increase in green mass compared to control, % Weight of shoots, kg Increase in yield compared to control, % Wheat Triticum aestivum 4 (1) _H2O (control) 76 - 8-9 40 8.8±4.7 - 0.14±0.01 - 5 (2) HTZ· L -AspA + H ₂ O 86 13.2 9 35 13.5±1.5 53.4±5.9 0.17±0.06 21.4±7.5 (3) 82 7.9 8 - 9.9±4.7 12.5±5.9 0.16±0.03 14.3±2.7 6 (4) HTZ· D -AspA + H ₂ O 90 18.4 9 32 16.0±1.0 81.8±5.1 0.23±0.03 64.3±8.4 (5) 96 26.3 8 - 11.4±3.6 29.6±9.3 0.17±0.04 21.4±5.0 Brussels sprouts Brassica oleracea var. Gemmifera 7 (1) _H2O (control) 50 - 8 50 7.0±3.3 - 0.056±0.002 - 8 (2) HTZ· L -AspA + H ₂ O 53 6.0 8 40 8.5±2.7 21.4±6.8 0.061±0.007 8.9±1.0 (3) 60 20.0 8 40 8.3±2.9 18.6±6.5 0.072±0.004 28.6±1.6 Radish Raphanus raphanistrum subsp. Sativus 9 (1) _H2O (control) 80 - 9 50 9.0±1.5 - 0.16±0.04 - 10 (2) HTZ· L -AspA + H ₂ O 100 25.0 9 50 10.6±1.3 17.8±2.2 0.18±0.06 12.5±4.2 11 (4) HTZ· D -AspA + H ₂ O 100 25.0 9 50 11.3±2.0 25.6±4.5 0.19±0.02 18.8±2.0

Table 4

The effect of modification of the soil "Good Helper Universal" with an aqueous dispersion of nanostructured L- or D -aspartate chitosan on the growth and vegetation of the test plant pea Pisum sativum and the volume of irrigation water consumed

Example No. Soil modifier Total volume of _H2O for irrigation for 8 days of vegetation, ml Vegetation time, days Germination rate, % Increase in germination compared to control, % Growth-promoting activity Height of shoots, mm Increase in green mass compared to control, % Weight of shoots, kg Increase in yield compared to control, % Pea Pisum sativum 12 (1) _H2O (control) 100 4 36±5 - 13±3 - 0.21±0.07 - 6 61±6 37±8 8 68±3 67±8 13 (2) HTZ· L -AspA + H ₂ O 80 4 22±7 10.3 10±2 11.4 0.27±0.07 28.6 6 56±5 44±2 8 75±3 74±6 (3) 80 4 48±8 10.3 17±5 31.3 0.28±0.02 33.3 6 68±4 54±8 8 75±5 88±9 (4) 80 4 43±2 16.2 12±2 19.4 0.28±0.07 33.3 6 72±2 31±3 8 79±1 80±5 14 (2) HTZ· D -AspA + H ₂ O 80 4 30±8 14.7 13±3 11.4 0.30±0.07 42.9 6 71±9 44±8 8 78±7 74±3 (3) 80 4 36±9 10.3 17±7 35.8 0.28±0.03 33.3 6 67±9 55±5 8 75±7 91±8 (4) 80 4 35±2 19.1 10±1 12.0 0.27±0.02 28.6 6 72±5 43±5 8 81±4 75±4

Claims (5)

1. Use of a biopreparation based on chitosan aspartate, which is an aqueous dispersion of chitosan aspartate particles with a size of 1200-1600 nm, obtained from an aqueous solution of chitosan with a molecular weight of 200 kDa in L- or D -aspartic acid, additionally containing a solution of silicon tetraglycerolate in a 3-molar excess of glycerol, with the following ratio of components: chitosan with a molecular weight of 200 kDa 0.08-0.34 g/dl L- or D -aspartic acid 0.07-0.27 g/dl silicon tetraglycerolate in a 3-molar excess of glycerol 0.08-0.32 g/dl water the rest, as a biological preparation for soil treatment. 2. The use according to item 1, characterized in that the particle size of chitosan aspartate is 1200-1400 nm when using aspartic acid in the D -form. 3. The use according to item 1, characterized in that the particle size of chitosan aspartate is 1400-1600 nm when using aspartic acid in the L -form.