A seed coating material with a biopolymer based controlled released system

Abstract

The invention is related to a seed coating material with a biopolymer-based controlled released system, which comprises of three layers and a coating method of this coating material to seed.

Description

[0001] DESCRIPTION

[0002] A SEED COATING MATERIAL WITH A BIOPOLYMER BASED CONTROLLED RELEASED SYSTEM TECHNICAL FIELD

[0003] The invention is related to a seed coating material with a biopolymer-based controlled released system, which comprises of three layers and a coating method of this coating material to seed.

[0004] BACKGROUND

[0005] Seed is one of the most important and most valuable inputs of agricultural production. Today, most of the agricultural production (90%) for food production purposes is performed by using seeds (Gbkgbl and Duman, 2018). For this reason, increasing seed quality affects agricultural production directly. There are different criteria that determine the seed quality, like; pest and disease-free, high germination rate, and homozygous germination. However, seed quality decreases because of both the structure of the seed and environmental factors (Gray, 1989). In order to obtain high-quality seed, both the seed production process should be well managed, and the correct seed storage conditions should be obtained later on. However, management of pathogens, climate change and soil structure is not possible. For this reason, seed coating technologies are important applications, which are developed to increase seed quality purposes.

[0006] Seed coating applications are performed for different purposes, such as increasing seed germination and vigor, facilitating planting, making misshapen and small seeds sowable by machine, protecting seeds against pests and diseases, etc (Gbkgbl and Duman, 2018). In 2022, 2.4 billion dollars were spent globally for seed coating, and it is estimated that this cost will increase up to 3.6 billion dollars in 2027 (Seed Coating Market, 2022). The development of coating materials which is cheaper, more efficient, and less harmful to the environment in such a large market has great importance.

[0007] Although different techniques have been developed for seed coating, the common purpose of all techniques is a good coating, homogeneity and sticking of coating material to the seed. Seed coating is performed by two approaches: film coating and pelleting. The seeds are coated by a thin layer of powder chemicals during film coating. During the coating process, adhesive liquids (water or polymer) are used for the adhesion of chemicals to the seed surface. Most of the chemicals, used during coating process, are pesticides that aim to protect seed from pathogens. In some cases, nutrients or plant growth regulators are used as coating material to increase seed germination.

[0008] Another seed coating approach is pelleting. In pelleting technique, it is aimed to homogenization of seeds and make seeds suitable for sowing by mechines by changing the seed shape, increasing the weight. In pelleting technique, seeds are coated layer by layer and each layer contains different chemicals for different purposes. The first layer in pelleting technique is the layer that protects seed from chemicals. Later on, different materials such as pesticides, fungicides, nutrients and growth regulators are used respectively (Gbkgbl and Duman, 2018).

[0009] As it mentioned above, there are different techniques and materilas that are used for seed coating purpose. The technologies and materials used for this purpose is developed day by day. One of the most important problem of seed coating is not being proper for sustainable agriculture due to using synthetic chemicals. For this reason, the effect of chemicals used in agricultural applications on humans and the environment is very important. The first aim of seed coating is protecting of seed from possible pests. Because, seed or soil born pathogens are most important factors that decrease seed germination and plant development. Not performing a healthy seed germination process directly affects crop production and yield (Kaufman, 1991 ; Pedrini et. al., 2017). For this reason, choosing right coating technique and material is very important. Besides, amount and environmental effect of materials should be considered during coating process (Taylor and Harman 1990; Pedrini et. al., 2017).

[0010] Various chemicals are used during seed coating. Although the content of these chemicals varies depending on the purpose of coating, they are generally similar. During seed coating, various chemicals like fungicides and pesticides to protect the seeds from different pathogens and nutrition to support seed germination are used. However, the harm to the environment and humans by these chemicals is known. Besides, direct application of chemicals and their penetration into the soil even when they are not needed harms sustainable agriculture. For this reason, a search have been made to develop of different coating materials. The fact that the materials to be used in seed coating do not harm the environment, humans and plants, or that this harm is at a much lower level than the synthetic chemicals used, makes them more preferred (Nuyttens et al., 2013; Accilleni et al., 2019; Pedrini et al., 2020; Zhang et al., 2020). Additionally, it will be a huge step in terms of sustainable agriculture that prevents the release of coating materials directly into the soil.

[0011] Synthetic chemicals, natural products like plant extracts and essential oils, biological agents and minerals are used in seed coating (Afzal vd., 2020). The main ingredients of seed coatings are active compounds. These compounds aim to increase seed germination and plant development as well as protect seeds from biotic and abiotic stress factors starting from the moment of planting. Chemicals like fungicides, pesticides, insecticides, nematicides and bactericides are products used in seed coating to neutralize pathogens to which the seed may be exposed (Rocha et al., 2019). Biostimulants and nutrition are preferred to increase seed germination and plant development (Pedrini et. al., 2017).

[0012] In order to adhere the active compounds to seed surface various liquids are used. The most important of these is water, which is good solvent. However, adjuvants are preferred, due to the solubility of chemicals in water is low (Afzal vd., 2020). Besides, surfactants are also used to adhere chemicals to seed surface. Surfactants increases seed germination apart form adhere (Madesen et. al., 2017). Additionally, to determine the coating level of seeds and label the coated seeds different colorants are used, which consist of 60% of coating material.

[0013] Other materials used in seed coatings are solid materials. This products are applied to seed surface as fine powder (Taylor and Harman, 1990; Chen and Turnblad, 2002). During this method water is used to adhere the materials onto seed surface. However, disadvantage of solid materials is making difficult to seed germination. In order to make easy to seed germination additional materials such as limestone, gypsum, silica, zeolite are used during coating process (Sooter et. al., 1978; Kangsopa et. al., 2018).

[0014] In order to contribute to sustainable agriculture, non-toxic or less harmless materials to the environment and humans have been preferred instead of synthetic chemicals. In the study, conducted by Ben-Jabeur et al. (2022), thyme oil was used for coating wheat seeds, and it was stated that thyme oil increased the activity of enzymes like amylase, involved in seed germination and had a positive effect on seed germination and plant development. In another study, various fungicides and fungicides mixtures, dry powder obtained from some plants (Bauhinia purpurea, Caesalpinia gilliesii, Cassia fistula, Cassia senna, Chrysanthemum frutescens, Euonymus japonicus ve Thespesia populnea var. acutiloba) were used for sugar beet seed coating to determine the impact aginst fungal pathogen Sclerotium rolfsii. In the study, it was observed that the most effective substance against S. rolfsii was Thiram+Cabonix, but other plant extracts, especially Thespesia populnea var. acutiloba, were also effective (Derbalah et. al., 2012). In other study that used soybean powder for seed coating purpose, it was stated that seed germination was decreased but, plant development was increased (Amirkhani et. al., 2016). For determination of the effect of cloves and star anise plants against Callosobruchus maculatus (F.) and C. chinensis L. insects, mung bean seeds were coated by film coating technique with mentioned paint extracts. As a result, it was emphasized that 1 % of star anise extract had effect on insects, mentioned above, without decreasing seed germination and could be used as an alternative to synthetic chemicals (Pumnunan et. al., 2021 ). In another study, essential oils of geranium, rose, lemon and mint plants were extracted and used for the coating of bean seeds. It was stated that the essential oils used for seed coating decreased the harmful effects of Fusarium oxysporum and Rhizoctonia solani fungi in beans and could be an alternative to chemical fungicides (El-Mougy et. al., 2007).

[0015] There is no control mechanism during the release of traditional agricultural chemicals. Therefore, the release of these chemicals into the environment and polluting soil and water resources is inevitable. In addition, these chemicals not only harm the environment, but there are also situations where they are washed from the soil by irrigation systems or evaporate before the plant can use them. Thus, these chemicals are intended to be used in agricultural production cannot be uptaken by plants effectively. On the other side, using of the mentioned chemicals in agriculture is inevitable. The role of pesticides and fungicides in controlling of biotic stress is highly important during controlling of agricultural pathogens and an important part of agricultural production. For this reason, it is not possible to eliminate of these chemicals from agricultural production under today’s conditions. However, it is possible to using less amount of chemicals more efficiently by developing technology. In this way, it is possible to use fertilizers and pesticides, which are indispensable to agricultural production, in a more controlled and safe manner. For this reason, controlled release systems have been used in agriculture. Various materials are used for controlled release systems. These materials are used as polymeric, metallic, or clay based and in different forms in general.

[0016] There are different advantages of controlled release systems: • In controlled release systems, only one application is sufficient, which provides an advantage in the quantity of chemicals and application frequency,

[0017] • It is possible to obstruct leaf scorch, loss of chemicals, toxic applications, and dermal irritations by controlled release systems,

[0018] • Since the release is controlled, it does not deteriorate the soil quality.

[0019] Apart from all these advantages, there are some disadvantages of controlled release systems:

[0020] • There is costly production process, so there is a marketing shortage,

[0021] • Some materials, which are used in coating, are not biodegradable and have harmful properties,

[0022] • Storage is harder; they can be affected by humidity and climate conditions easily,

[0023] • The needed quantity of chemical cannot be obtained and same amount of chemical is released (Azeem, 2014).

[0024] The release of chemicals contained in controleed release systems is performed through different mechanisms. This technology is still on progress and there are different methods in literature. There are basically three different release systems: diffusion or swelling, degradation of polymer coating, and fracture or dissolution (Trinh and Kushaari, 2016).

[0025] There are fertilizers and pesticides developed by controlled release systems in literature. In a study conducted by Xu et al. (2022), copper ions and NPK fertilizer were released by pH change and the presence of pathogens after encapsulated in biodegradable nanoparticles. Besides, slow or immediate release of active compounds were obtained by the changing in the concentration of polymer materials (Xu et. al., 2022). Sodium alginate (Na-Ag) was mixed with perlite and clay separately and nanocapsules, which contained KNO3, were formed and and by controlled release of KNO3, faster germination of parsley seeds and improved plant growth were achieved (Durmu§, 2019).

[0026] Another material that is a possible alternative for controlled fertilizer release is lignin. As a result of modified coatings produced by lignin and various chemicals, controlled release of especially urea was performed and the positive effect on plant development was observed (Chen et. al., 2020). An alternative fertilizer was produced by providing a controlled release of granular fertilizers coated with starch and polyvinyl alcohol. In the study, there was no plant experiment, but the controlled release of fertilizer was performed (Han et al., 2009). Urea was coated using castor oil-based polyurethane coating material and controlled release of urea was achieved. In the experiment conducted on corn plants, it was stated that controlled release of urea contributed to plant growth (Zhao et al., 2020).

[0027] In traditional seed coating techniques, seeds are coated with necessary chemicals and these chemicals are released around the seed by sowing the seeds into soil. There is no control mechanism in this release system. However, it is possible to use the controlled release system, which has been used in pharmacy for many years and has become popular in agriculture recently, in seed coating. Thus, controlled and slow release of coating materials can be performed. In this way, not only will the effectiveness period of these materials be extended, but their release will also be possible if they are necessary, thanks to controlled release.

[0028] Thiram, which is used as fungicides in traditional seed coating, was used for coating of soybean seeds by polyethylene glycol. After coating, Thiram release was achieved in a controlled manner and seed germination increased (Kaushik et. al., 2013). Rice seeds were encapsulated through electrospinning technique by ethyl cellulose based Thiram. As a result of the study, while there was no increase in the seed germination, it was stated that coating significantly decreased examined fungal diseases (Castaneda et. al., 2014). In another study, tomato and lettuce seeds were coated with nanofibers containing CuO nanoparticles using cellulose acetate and gelatin and their effectiveness on Fusarium species was examined. As a result of the study, seed germination and plant development were increased with and without pathogens by obtaining slow release of CuO compared to traditional film coating (Xu et. al., 2020).

[0029] AIM OF THE INVENTION

[0030] The aim of the invention is to develop seed coating material which provides controlled release. Another aim of the study is to use essential oils and nanoparticles, which are harmless of less harmful in coating material instead of harmful chemicals.

[0031] Another aim of the study is to develop biodegradable seed coating material that provides controlled release. For this purpose, in the first step soil born fungus were isolated, which causes damping off disease in sugar beet plants. Later on, the antifungal effect on this fungus of various medicinal plants (marjoram, lavender, sage) and silver nanoparticles (AgNP), which antifungal effect was stated in the literature.

[0032] The matrix of the coating material consists of polycaprolactone (PCL), cellulose acetate (CA) and chitosan (CH) polymers. Coating material degrades by pH change during germination and enzymes secreted from pathogens due to natural characteristics of polymers. Thus, release of active compounds that is encapsulated in coating material will be performed during germination and presence of pathogens. In this case, no pollution is caused in the soil.

[0033] LIST OF FIGURES

[0034] Figure 1. Schematic presentation of coating material

[0035] Correspondence of numberings given in the figure:

[0036] 1 . Bottom layer

[0037] 2. Middle layer

[0038] 3. Upper layer

[0039] DETAILED DESCRIPTION OF INVENTION

[0040] Polycaprolactone (PCL), cellulose acetate (CA) and chitosan (CH) polymers were used for coating. Polycaprolactone is a biodegradable polymer in the presence of amylase enzyme. Chitosan degrades under high pH conditions, when dissolves in low pH conditions. Thus, degradation of coating material through presence of pathogens (due to amylase enzyme secreted by pathogens) and decrease in pH during germination occurred and essential oils and AgNPs were released. The aim of choosing CA was to prevent early degradation of coating material due to its hydrophobic structure. The solvents of the polymers are also selected from solvents that are safe and suitable for sustainable agriculture. For this purpose, polymers were dissolved in formic acid / acetic acid / water 40 / 40 / 20 (v / v / v) solvents.

[0041] The seed coating subject to the invention consists of three layers. The first layer (first shell layer) consisted only of polymer matrix to prevent the interaction of essential oils and AgNP to seeds.

[0042] The second layer, the middle layer, is named as core layer. In this layer there is essential oils and AgNPs. Thus, release of these component occurs at a high rate. The release in this layer will be long term. The third layer, named as second shell layer, has an immediate release content to ensure first and rapid response. The essential oil and AgNPs content in this layer is half of the second layer.

[0043] Within the scope of this invention, coating is performed by three different polymer concentrations (Figure 1 ). By the changing in concentrations, the degradation of coating materials changes from slow to immediate.

[0044] For immediate release in the bottom layer (1 ) CH / CA / Zein / Starch / PCL- 40 / 30 / 10 / 10 / 10 (3% w / v), middle layer (2) CH / CA / PCL - 25 / 25 / 50 (2% w / v) + AgNP / EO / Thiram (50%) and upper layer (3) CH / CA / Zein / Starch / PCL - 35 / 30 / 15 / 15 / 5 (3% w / v) + AgNP / EO / Thiram (50%) structure was formed.

[0045] For middle release in the bottom layer (1 ) CH / CA / Zein / Starch / PCL- 40 / 30 / 10 / 10 / 10 (3% w / v), middle layer (2) CH / CA / PCL - 25 / 25 / 50 (2% w / v) + AgNP / EO / Thiram (75%) and upper layer (3) CH / CA / Zein / Starch / PCL - 40 / 30 / 10 / 10 / 10 (3% w / v) + AgNP / EO / Thiram (25%) structure was formed.

[0046] For slow release in the bottom layer (1 ) CH / CA / Zein / Starch / PCL-40 / 30 / 10 / 10 / 10 (3% w / v), middle layer (2) CH / CA / PCL - 25 / 25 / 50 (2% w / v) + AgNP / EO / Thiram (100%) and upper layer (3) CH / CA / Zein / Starch / PCL - 40 / 30 / 10 / 10 / 10 (3% w / v) structure was formed.

[0047] Layers were prepared sequentially. After preparing the first layer, the second layer was added onto the first layer, and finally, the third layer was prepared. Preparation steps of layers are explained below.

[0048] To preparation of shell layers (Middle (2) and upper layers (3)) firstly starch and PCL were dissolved in formic acid for 10 hours. Later, it was added to acetic acid and water in the solution and stirred for 2 hours. CH was added in the solution and mixed for 10 more hours. Finally, solution for shell layer was prepared by addition of CA, zein and active compounds and mixed for 2 hours.

[0049] In order to preparation the core layer (Bottom layer (1 )) PCL was stirred in acetic acid for 4 hours at room temperature. Later, water was added and stirred 8 more hours. Finally, CH, CA and active compounds were added and mixed for 4 hours. Thus, the precursor solution was prepared for core layer. Content of layers is presented on Table 1 . The prepared solutions were sterilized by filter sterilization. Table 1. Content of coating materials

[0050] First Layer

[0051] Solvent (mL) Biopolymer (%3 w / v) (mg) Active compound

[0052] AA FA H20 CA CH Zein Starch PCL

[0053] 1. CM 4 4 2 120 90 30 30 15 0

[0054] 2. CM 4 4 2 120 90 30 30 15 0

[0055] 3. CM 4 4 2 120 90 30 30 15 0

[0056] Second Layer

[0057] Solvent (mL) Biopolymer (%2 w / v) (mg) Active compound

[0058] AA FA H2O CA CH PCL At least one of

[0059] _ 9ssential oils,

[0060] 1. CM 4 4 2 50 50 100 AgNP or

[0061] 2. CM 4 4 2 50 50 100 ™ram

[0062] 3. CM 4 4 2 50 50 100

[0063] Third Layer

[0064] Solvent (mL) Biopolymer (%3 w / v) (mg) Active compound

[0065] AA FA H2O CA CH Zein Starch PCL At least one of

[0066] _ 9ssential oils,

[0067] 1. CM 4 4 2 105 90 45 45 15AgNpor

[0068] 2. CM 4 4 2 120 90 30 30 15Thlram

[0069] 3. CM 4 4 2 120 90 30 30 15

[0070] CM: Coating material, AA: Acetic acid, FA: Formic acid, CA: Cellulose acetate, CH: chitosan, PCL: Polycaprolactone

[0071] The other step was coating of seeds. For this purpose, the seeds were dipped in the solutions and dried for 24 hours. Thus, the solvents were removed, and polymerization was performed. Later, the seeds were dipped in the second solution and dried for 24 hours. Thus, the solvents were removed, and polymerization was performed. This step for the second layer was performed 3 times. In short, the second layer consisted of three layers. Finally, the seeds were coated by dipping of seeds in the third solution and then dried. The coating material contained at least one of the following essential oils, AgNPs and Thiram as active compounds.

Claims

CLAIMS1. A seed coating material with a biopolymer-based controlled release system, characterized by comprising a three-layered structure further comprising a bottom layer (1 ) CH / CA / Zein / Starch / PCL - 40 / 30 / 10 / 10 / 10 (3% w / v), a middle layer (2) CH / CA / PCL - 25 / 25 / 50 (2% w / v) + at least one of AgNP / EO / Thiram (50%) and an upper layer (3) CH / CA / Zein / Starch / PCL- 35 / 30 / 15 / 15 / 5 (3% w / v ) + at least one (50%) of AgNP / EO / Thiram.

2. A seed coating material with a biopolymer-based controlled release system, characterized by comprising a three-layered structure further comprising a bottom layer (1 ) CH / CA / Zein / Starch / PCL - 40 / 30 / 10 / 10 / 10 (3% w / v), a middle layer (2) CH / CA / PCL - 25 / 25 / 50 (2% w / v) + at least one of AgNP / EO / Thiram (75%) and an upper layer (3) CH / CA / Zein / Starch / PCL- 40 / 30 / 10 / 10 / 10 (3% w / v ) + at least one (50%) of AgNP / EO / Thiram.

3. A seed coating material with a biopolymer-based controlled release system, characterized by comprising a three-layered structure further comprising a bottom layer (1 ) CH / CA / Zein / Starch / PCL - 40 / 30 / 10 / 10 / 10 (3% w / v), a middle layer (2) CH / CA / PCL - 25 / 25 / 50 (2% w / v) + at least one of AgNP / EO / Thiram (100%) and an upper layer (3) CH / CA / Zein / Starch / PCL- 40 / 30 / 10 / 10 / 10 (3% w / v ) and characterized by containing a three-layered structure.

4. A seed coating method of seed coating materials according to any claims from Claims 1-3, characterized by comprising the steps below;- To prepare the shell layers (middle (2) and upper layers (3)), first dissolve starch and PCL in formic acid for 10 hours at room temperature,- Then, acetic acid and water are added to the solution and mixed for another 2 hours,- Adddition of CH to the prepared solution and stirring for another 10 hours,- Preparation the solution required for the shell layer by adding SA, zein and active ingredients and mixing for another 2 hours,- For preparation the core layer (Second layer (2)), dissolving PCL in formic acid and acetic acid at room temperature for 4 hours,- Then, add water and mixing for another 8 hours,- Finally, addition of CH, CA and active compounds and mix for another 4 hours.- Sterilization the prepared solutions with a filter,- Dipping the seeds into the first prepared layer and removing the solution by waiting for 24 hours,- After the first layer dries, dipping it into the polymer solution prepared for the second layer 3 times in succession and removing the solvent for 24 hours after each dipping process,- It is characterized by the stages of drying after dipping into the polymer solutions that prepared for the third layer.

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