Use of selenium-containing complex initiators as inhibitors of deoxynivalenol production in corn

Abstract

The present application relates to the use of selenium-containing complex initiator as a deoxynivalenol synthesis inhibitor in corn, by using selenium-containing complex initiator to coat corn seeds to form corn seeds containing selenium-containing complex initiator; the selenium-containing complex initiator enhances the resistance of corn to fusarium graminearum and inhibits the synthesis of deoxynivalenol by reprogramming seed metabolism and activating jasmonic acid (JA) mediated defense response in seedling stage, simultaneously diverting the phenylpropanoid metabolism of seedling from flavonoid biosynthesis to antifungal phenolic biosynthesis, thereby synchronizing seed vigor with innate immunity, and providing an effective strategy for corn crop protection.

Description

Technical Field

[0001] This invention relates to the field of agricultural biotechnology, and more particularly to the use of a selenium-containing composite initiator as an inhibitor of deoxynivalenol production in maize. Background Technology

[0002] Fusarium graminearum can induce stalk rot and ear rot in maize and produce deoxynivalenol (DON), which contaminates maize. Under suitable temperature and humidity conditions, these fungi can multiply rapidly and produce toxins, severely contaminating maize crops and their processed products. In addition, maize may also produce this toxin if it is not dried sufficiently or stored improperly during storage.

[0003] Several studies have begun to focus on how to inhibit the synthesis of deoxynivalenol, thereby suppressing the growth and toxin production of Fusarium graminearum. For example, CN118923671A discloses a fungal inhibitor composed of resveratrol, which can significantly inhibit the mycelial growth of Fusarium graminearum, disrupt the cell membrane of Fusarium graminearum, inhibit the expression of toxin-producing genes of Fusarium graminearum, and inhibit the production of deoxynivalenol by Fusarium graminearum. CN119498308A discloses the use of the plant hormone methyl jasmonate as an inhibitor of the growth and DON synthesis of Fusarium graminearum, providing a plant-derived inhibitor for the formulation of safe, pollution-free, and green strategies for the control of wheat scab. CN117898307A discloses a biocontrol bacterium, Burkholderia pyrrole, which is used to inhibit the production of deoxynivalenol toxin by Fusarium graminearum.

[0004] Although some progress has been made, the relevant technologies focus more on the growth period after crop seedlings emerge, resulting in limited absorption capacity of crops by the corresponding inhibitors, and the inhibitory effect still needs to be further improved. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a use of a selenium-containing composite initiator as an inhibitor of deoxynivalenol (DON) production in maize. This invention coats maize seeds with the selenium-containing composite initiator, forming maize seeds containing the selenium-containing composite initiator. This can enhance the vitality of maize crops at the seed stage and activate their innate immunity, thereby better inhibiting the production of DON in maize, further improving the overall resistance of maize crops, and making them more conducive to their growth and quality.

[0006] To achieve this objective, the present invention adopts the following technical solution:

[0007] In a first aspect, the present invention provides the use of a selenium-containing composite initiator as an inhibitor of deoxynivalenol production in maize. The use involves coating maize seeds with the selenium-containing composite initiator to form maize seeds containing the selenium-containing composite initiator. By planting the maize seeds containing the selenium-containing composite initiator, the inhibition of deoxynivalenol in maize is achieved.

[0008] The corn seeds containing the selenium complex initiator described in this invention are prepared by a method comprising the following steps:

[0009] (1) A first multi-inlet vortex mixer with four channels is provided. A selenium source is introduced into the first channel of the first multi-inlet vortex mixer as a first liquid flow, a stabilizer is introduced into the second channel as a second liquid flow, water is introduced into the third channel as a third liquid flow, and a reducing agent is introduced into the fourth channel as a fourth liquid flow. The first liquid flow, the second liquid flow, the third liquid flow and the fourth liquid flow collide and mix instantaneously in the first multi-inlet vortex mixer to form a selenium-containing composite suspension.

[0010] (2) Provide a second multi-inlet vortex mixer connected in series with the first multi-inlet vortex mixer in step (1), and introduce the selenium-containing composite suspension obtained in step (1) into the first channel of the second multi-inlet vortex mixer as the first liquid flow, introduce the first ligand into the second channel as the second liquid flow, introduce the second ligand into the third channel as the third liquid flow, and introduce water into the fourth channel as the fourth liquid flow. The first liquid flow, the second liquid flow, the third liquid flow and the fourth liquid flow collide and mix instantaneously in the second multi-inlet vortex mixer to form a selenium-containing coating suspension;

[0011] (3) The disinfected corn seeds are first soaked in the selenium-containing coating suspension obtained in step (2), and then soaked in the cross-linking agent solution. After drying, corn seeds containing selenium complex initiator are formed.

[0012] The "selenium-containing composite initiator" in this invention refers to a preparation process in which selenium is combined with components such as stabilizers and instantaneously collided and mixed in a first multi-inlet vortex mixer and a second multi-inlet vortex mixer connected in series to form a selenium-containing composite. After coating the seeds, it can be used as a maize seed vigor initiator, enabling the maize seeds to exhibit excellent inhibition of the formation of deoxynivalenol in maize during the later stages of planting, thereby effectively inhibiting the production of maize toxins.

[0013] A series of plant physiological results indicate that coating maize seeds with the selenium-containing composite initiator described in this invention can reprogram seed metabolism and activate jasmonic acid (JA)-mediated defense responses during the seedling stage, thereby enhancing maize's resistance to Fusarium graminearum and inhibiting the synthesis of deoxynivalenols. Simultaneously, it shifts seedling phenylpropanoid metabolism from flavonoid biosynthesis to antifungal phenolic biosynthesis, thus establishing a chemical barrier against fungal invasion. This metabolic reprogramming significantly reduces the severity index (DSI) of stem rot and effectively inhibits the biosynthesis of deoxynivalenols.

[0014] Preferably, in step (1) of preparing the corn seeds containing the selenium complex initiator, the selenium source is any one or a combination of at least two of sodium selenite, sodium selenate, or selenium dioxide.

[0015] Preferably, the reducing agent is any one or a combination of at least two of cysteine, hydrazine hydrate, ascorbic acid, sodium sulfite, sodium thiosulfate, or glutathione.

[0016] Preferably, the stabilizer is any one or a combination of at least two of polysaccharides, organic compounds, polymers or peptides, and more preferably a combination of polysaccharides and peptides.

[0017] In this invention, the polysaccharide compound may be, for example, at least one of chitosan, chitosan oligosaccharide, carboxymethyl chitosan, sodium alginate, Codonopsis pilosula polysaccharide, Lycium barbarum polysaccharide, Taraxacum mongolicum polysaccharide, tea polysaccharide, konjac polysaccharide, or Lentinus edodes polysaccharide; the organic compound may be, for example, at least one of melatonin, sodium lauryl sulfate, hexadecyltrimethylammonium bromide, resveratrol, Tween (20, 40, 60, 80), proanthocyanidins, chlorogenic acid, or epigallocatechin gallate; the polymer may be, for example, at least one of polyethylene glycol, polyvinyl alcohol, or polyvinylpyrrolidone; and the polypeptide may be, for example, at least one of soybean peptide, peanut peptide, corn peptide, pea peptide, or rice peptide.

[0018] The stabilizer used in this invention is preferably a combination of polysaccharides and polypeptides. By combining polysaccharides and polypeptides, the selenium-containing composite suspension obtained in step (1) can be made more stable, so that the activity of the selenium composite initiator can be maximized when coating corn seeds, thereby more effectively inhibiting the deoxynivalenol in corn.

[0019] When the stabilizer described in this invention is a combination of polysaccharides and polypeptides, the mass ratio between the polysaccharides and polypeptides can be, for example, (1-5):(1-5), where the first and second "1-5" can be selected from 1, 2, 3, 4, and 5.

[0020] Preferably, the molar ratio of selenium element to reducing agent in the selenium source is 1:(2-6), for example 1:2, 1:3, 1:4, 1:5 or 1:6.

[0021] Preferably, the mass ratio of selenium in the selenium source to the mass of the stabilizer is 1:(5-50), for example 1:5, 1:8, 1:10, 1:12, 1:15, 1:20, 1:25, 1:30, 1:35, 1:40, 1:42, 1:45 or 1:50.

[0022] Preferably, in step (1) of preparing the corn seeds containing the selenium complex initiator, the flow rates of the first and second liquid streams are each independently 4 to 8 mL / min, for example 4 mL / min, 5 mL / min, 6 mL / min, 7 mL / min or 8 mL / min, and the flow rates of the third and fourth liquid streams are each independently 25 to 50 mL / min, for example 25 mL / min, 30 mL / min, 35 mL / min, 40 mL / min or 50 mL / min.

[0023] Preferably, in the selenium-containing composite suspension obtained in step (1), the average particle size of selenium is 20-80 nm, for example 20 nm, 40 nm, 60 nm or 80 nm.

[0024] Preferably, in step (2) of preparing the corn seeds containing the selenium composite initiator, the first ligand is any one or a combination of at least two of carboxymethyl chitosan, sodium alginate, methylcellulose, or cellulose nanocrystals.

[0025] Preferably, the second ligand is any one or a combination of at least two of pectin, xanthan gum, gum arabic, or gelatin.

[0026] In this invention, the mass ratio of the selenium-containing composite suspension obtained in step (1) to the first ligand and the second ligand is not specifically limited, as long as it can achieve good coating of corn seeds. For example, the mass ratio of selenium element to the first ligand and the second ligand in the selenium-containing composite suspension can be 1:(15-30):(15-30), where the first and second "15-30" can be selected from 15, 18, 20, 25 or 30.

[0027] Preferably, in step (2) of preparing the corn seeds containing the selenium complex initiator, the flow rates of the first liquid flow and the second liquid flow are each independently 5-10 mL / min, for example 5 mL / min, 6 mL / min, 8 mL / min, 9 mL / min or 10 mL / min, and the flow rates of the third liquid flow and the fourth liquid flow are each independently 30-60 mL / min, for example 30 mL / min, 40 mL / min, 45 mL / min, 50 mL / min or 60 mL / min.

[0028] Preferably, in step (2) of preparing the corn seeds containing the selenium complex initiator, the concentration of selenium in the selenium-coating suspension is 40-60 mg / L, for example, 40 mg / L, 50 mg / L or 60 mg / L.

[0029] In this invention, by controlling the selenium concentration in the selenium-coated suspension within a suitable range, it can better inhibit the formation of deoxynivalenol in corn, while also maximizing the promotion of corn crop growth.

[0030] Preferably, in step (3) of preparing the corn seeds containing the selenium complex initiator, the soaking time is 2 to 12 hours, for example, 2 hours, 5 hours, 8 hours, 10 hours or 12 hours.

[0031] Preferably, the soaking time is 3 to 6 hours, for example, 3 hours, 4 hours, 5 hours or 6 hours.

[0032] Preferably, the crosslinking agent is any one or a combination of at least two of zinc sulfate, copper sulfate, calcium chloride, or manganese sulfate.

[0033] Preferably, the mass concentration of the crosslinking agent is 1-5%, for example 1%, 2%, 3%, 4% or 5%.

[0034] Compared with the prior art, the present invention has at least the following beneficial effects:

[0035] In this invention, by coating maize seeds with a selenium complex initiator, maize seeds containing the selenium complex initiator are formed. The selenium complex initiator can enhance maize's resistance to Fusarium graminearum and inhibit the synthesis of deoxynivalenol by reprogramming seed metabolism and activating jasmonic acid (JA)-mediated defense responses during the seedling stage. At the same time, it shifts the phenylpropanoid metabolism of seedlings from flavonoid biosynthesis to antifungal phenolic biosynthesis, thereby synchronizing seed vigor with innate immunity. This provides an effective strategy for maize crop protection and achieves effective inhibition of maize toxins. Attached Figure Description

[0036] Figure 1This is a schematic diagram of the process for preparing corn seeds containing a selenium complex initiator in this invention.

[0037] Figure 2 The phenotypic characteristics of maize seedlings corresponding to the control group (CK), reference group (FgCK), Example 1, Examples 3-4, Examples 6-8 and Comparative Example 1 are shown.

[0038] Figure 3 The disease severity index (DSI) of maize seedlings corresponding to the control group (FgCK), Examples 1, Examples 3-4, Examples 6-8 and Comparative Example 1 is shown.

[0039] Figure 4 The jasmonic acid (JA) content in maize seedlings corresponding to the control group (FgCK), Example 1, Examples 3-4, Examples 6-8 and Comparative Example 1 is shown.

[0040] Figure 5 The total flavonoid content in maize seedlings corresponding to the control group (FgCK), Examples 1, 3-4, 6-8 and Comparative Example 1 is shown.

[0041] Figure 6 The total phenol content in maize seedlings corresponding to the reference group (FgCK), Examples 1, 3-4, 6-8 and Comparative Example 1 is shown.

[0042] Figure 7 The correlation and interaction among jasmonic acid, total flavonoids and total phenols in the corn seedlings corresponding to Example 1 are shown. Detailed Implementation

[0043] The technical solution of the present invention will be further described below with reference to the accompanying drawings and specific embodiments. Those skilled in the art should understand that the specific embodiments described are merely illustrative of the present invention and should not be construed as limiting the invention.

[0044] Preparation Example 1

[0045] This preparation example provides a corn seed containing a selenium complex initiator, and the preparation process is as follows: Figure 1 As shown, steps (1) and (2) are carried out in a first multi-inlet vortex mixer and a second multi-inlet vortex mixer connected in series. The preparation process specifically includes the following steps:

[0046] (1) Sodium selenite solution (selenium source) is introduced into the first channel of the first multi-inlet vortex mixer as the first liquid flow, a stabilizer composed of chitosan oligosaccharide and soybean peptide in a mass ratio of 1:2 is introduced into the second channel as the second liquid flow, water is introduced into the third channel as the third liquid flow, and ascorbic acid (reducing agent) is introduced into the fourth channel as the fourth liquid flow.

[0047] The first, second, third, and fourth liquid streams collide and mix instantaneously in a first multi-inlet vortex mixer. The mass ratio of selenium to stabilizer in the sodium selenite solution is 1:30, and the molar ratio of selenium to ascorbic acid in the sodium selenite solution is 1:5. The flow rates of the first and second liquid streams are both 6 mL / min, and the flow rates of the third and fourth liquid streams are both 30 mL / min, forming a selenium-containing composite suspension with an average selenium particle size of 35±5 nm.

[0048] (2) The selenium-containing composite suspension obtained in step (1) is introduced into the first channel of the second multi-inlet vortex mixer, the carboxymethyl chitosan solution (ligand 1) is introduced into the second channel, the pectin solution (ligand 2) is introduced into the third channel, and water is introduced into the fourth channel.

[0049] The first, second, third, and fourth liquid flows collide and mix instantaneously in the second multi-inlet vortex mixer. The mass ratio of selenium to carboxymethyl chitosan and pectin in the selenium-containing composite suspension is 1:20:20. The flow rates of the first and second liquid flows are both 8 mL / min, and the flow rates of the third and fourth liquid flows are both 50 mL / min, forming a selenium-containing coating suspension with a selenium concentration of 50 mg / L.

[0050] (3) After the corn seeds are surface disinfected with 2% H2O2, they are rinsed with distilled water to ensure that the residual disinfectant is completely removed. They are first soaked in the selenium-containing coating suspension obtained in step (2) for 8 hours, and then soaked in calcium chloride with a mass concentration of 2% for 5 hours. After vacuum drying at 35°C for 48 hours, corn seeds containing selenium complex initiator are formed.

[0051] Preparation Example 2

[0052] This preparation example provides a corn seed containing a selenium complex initiator, and the preparation process is as follows: Figure 1 As shown, steps (1) and (2) are carried out in a first multi-inlet vortex mixer and a second multi-inlet vortex mixer connected in series. The preparation process specifically includes the following steps:

[0053] (1) A selenium dioxide solution (selenium source) is introduced into the first channel of the first multi-inlet vortex mixer as the first liquid flow, a stabilizer composed of lentinan and peanut peptide in a mass ratio of 1:1 is introduced into the second channel as the second liquid flow, water is introduced into the third channel as the third liquid flow, and cysteine ​​solution (reducing agent) is introduced into the fourth channel as the fourth liquid flow.

[0054] The first, second, third, and fourth liquid streams collide and mix instantaneously in a first multi-inlet vortex mixer. The mass ratio of selenium to stabilizer in the selenium dioxide solution is 1:10, and the molar ratio of selenium to cysteine ​​in the selenium dioxide solution is 1:3. The flow rates of the first and second liquid streams are both 4 mL / min, and the flow rates of the third and fourth liquid streams are both 40 mL / min, forming a selenium-containing composite suspension with an average selenium particle size of 42±5 nm.

[0055] (2) The selenium-containing composite suspension obtained in step (1) is introduced into the first channel of the second multi-inlet vortex mixer, sodium alginate solution (ligand 1) is introduced into the second channel, gum arabic solution (ligand 2) is introduced into the third channel, and water is introduced into the fourth channel.

[0056] The first, second, third, and fourth liquid flows collide and mix instantaneously in the second multi-inlet vortex mixer. The mass ratio of selenium to sodium alginate and gum arabic in the selenium-containing composite suspension is 1:25:15. The flow rates of the first and second liquid flows are both 6 mL / min, and the flow rates of the third and fourth liquid flows are both 45 mL / min, forming a selenium-containing coating suspension with a selenium concentration of 45 mg / L.

[0057] (3) After the corn seeds are surface disinfected with 2% H2O2, they are rinsed with distilled water to ensure that the residual disinfectant is completely removed. They are first soaked in the selenium-containing coating suspension obtained in step (2) for 10 hours, and then soaked in zinc sulfate with a mass concentration of 3% for 6 hours. After vacuum drying at 40°C for 42 hours, corn seeds containing selenium complex initiator are formed.

[0058] Preparation Example 3

[0059] Compared with Preparation Example 1, the stabilizer combination in step (1) was adjusted to a single stabilizer—chitosan oligosaccharide, and the mass ratio of selenium to chitosan oligosaccharide in the sodium selenite solution was 1:30. Everything else was exactly the same as in Preparation Example 1.

[0060] Preparation Example 4

[0061] Compared with Preparation Example 1, the stabilizer combination in step (1) was adjusted to a single stabilizer—soybean peptide, and the mass ratio of selenium to soybean peptide in the sodium selenite solution was 1:30. Everything else was exactly the same as in Preparation Example 1.

[0062] Preparation Examples 5 to 8

[0063] Compared with Preparation Example 1, the amount of water introduced into the fourth channel in step (2) was changed so that step (2) produced selenium-containing coating suspensions with selenium element concentrations of 5 mg / L (Preparation Example 5), 25 mg / L (Preparation Example 6), 75 mg / L (Preparation Example 7) and 100 mg / L (Preparation Example 8), respectively. Everything else was exactly the same as in Preparation Example 1.

[0064] Preparation Example 9

[0065] Compared with Preparation Example 1, the stabilizer in step (1) was replaced with Tween-20 and soybean peptides in a mass ratio of 1:2, and the mass ratio of selenium to stabilizer in the sodium selenite solution was kept at 1:30. Everything else was exactly the same as in Preparation Example 1.

[0066] Preparation Example 10

[0067] Compared with Preparation Example 1, the stabilizer in step (1) was replaced with a combination of chitosan oligosaccharide and polyethylene glycol in a mass ratio of 1:2, and the mass ratio of selenium to stabilizer in the sodium selenite solution was kept at 1:30. Everything else was exactly the same as Preparation Example 1.

[0068] Comparative Preparation Example 1

[0069] This comparative preparation example provides an inorganic selenium-impregnated corn seed, the preparation method of which includes the following steps:

[0070] (1) After disinfecting the corn seeds with 2% H2O2, rinse them with distilled water to ensure that the residual disinfectant is completely removed;

[0071] (2) The corn seeds disinfected in step (1) are immersed in a sodium selenite solution with a selenium concentration of 50 mg / L. After 15 hours, the seeds are removed and vacuum dried at 35°C for 48 hours to obtain the inorganic selenium-impregnated corn seeds.

[0072] Comparative Preparation Example 2

[0073] This comparative preparation example provides a method for preparing corn seeds coated with a selenium complex using a stirring method, the preparation method comprising the following steps:

[0074] (1) Add sodium selenite solution and a stabilizer composed of chitosan oligosaccharide and soybean peptide in a mass ratio of 1:2 to the first beaker. After stirring for 30 min, add an aqueous solution of ascorbic acid dropwise to the beaker. The mass ratio of selenium in the sodium selenite solution to the stabilizer is 1:30. The molar ratio of selenium in the sodium selenite solution to ascorbic acid is 1:5. The ascorbic acid is added at a rate of 30 mL / min, and a selenium-containing suspension is finally formed.

[0075] (2) Add carboxymethyl chitosan solution and pectin to the second beaker, stir for 30 min, then add the selenium-containing suspension from step (1), and continue stirring for 300 min to form a selenium-containing coating suspension with a selenium element concentration of 50 mg / L, wherein the mass ratio of selenium element to carboxymethyl chitosan and pectin is 1:20:20.

[0076] (3) After the corn seeds are surface disinfected with 2% H2O2, they are rinsed with distilled water to ensure that the residual disinfectant is completely removed. They are first soaked in the selenium-containing coating suspension obtained in step (2) for 8 hours, and then soaked in calcium chloride with a mass concentration of 2% for 5 hours. After vacuum drying at 35°C for 48 hours, corn seeds coated with selenium complex are formed by stirring.

[0077] Test case

[0078] For the corn seeds provided in Preparation Examples 1-10 and Comparative Preparation Examples 1-2, the surface-adhered residual selenium was removed by rinsing with distilled water. Corn seeds without any selenium treatment were used as controls. They were uniformly planted in a controlled environment greenhouse facility with the following growth parameters: light conditions of 14h light / 10h darkness; humidity controlled at 75%; and growth temperature of 24℃.

[0079] 1. Preparation of Fusarium graminearum fungal spore suspension

[0080] Fusarium graminearum strains were inoculated onto PDA plates and cultured in the dark at 25°C for at least 10 days. Several 5×5 mm agar blocks were aseptically cut and placed in sterile CMC medium, and cultured at 200 rpm and 28°C for 3 days. The spore suspension in the CMC medium was filtered through two layers of gauze and then centrifuged at 3800×g for 5 min; the supernatant was discarded, and the concentrated conidia were resuspended in sterile water and washed twice, then diluted to 1.0×10⁻⁶. 6 For a concentration of / ml, add 0.001% Tween-20 to the diluent and set aside.

[0081] 2. Fusarium graminearum inoculation and disease severity index (DSI) calculation

[0082] 2.1 Corn seeds without any selenium treatment, corn seeds from Preparation Examples 1-10, and comparative Preparation Examples 1-2 were cultured for 12 days. Under light conditions of 24°C, 14h light / 10h darkness, a 1mm deep wound was made at the base of the corresponding corn seedling stem using a 1mL syringe. 20µL of water was added to the wound, and 20µL of spore suspension was applied to the wound to form: a control group (FgCK, i.e., inoculated with Fusarium graminearum but without any selenium treatment), Examples 1-10, and comparative Examples 1-2, with a relative humidity of 75%. Three days after inoculation, the phenotype of each corn seedling was observed. At the same time, corn seedlings without any selenium treatment and without inoculation with Fusarium graminearum were cultured in the same manner as a control group (CK).

[0083] Figure 2 The phenotypic characteristics of maize seedlings corresponding to the control group (CK), reference group (FgCK), Example 1, Examples 3-4, Examples 6-8 and Comparative Example 1 are shown.

[0084] pass Figure 2 It can be seen that the control group, which was not inoculated with Fusarium graminearum, showed normal performance; while the reference group, which was inoculated with Fusarium graminearum but not treated with any selenium, showed yellow-brown lesions on its leaves, with most leaves necrotic and the stems severely damaged. Compared with Example 1, the leaves showed severe stress effects and reduced chlorophyll content, while the leaves of Example 1 showed almost no lesions, the stems were harder and thicker, and the degree of rot was lower, showing the best overall performance. Examples 3-4 and Examples 6-8 all showed a certain degree of stem rot, softening, and leaf discoloration, which were slightly worse than Example 1.

[0085] 2.2 The Disease Severity Index (DSI) of each maize seedling was evaluated and statistically analyzed. The calculation of the Disease Severity Index (DSI) was based on the article Sun, Y., Ruan, X., Ma, L., Wang, F., Gao, X. Rapid Screening and Evaluation of Maize Seedling Resistance to Stalk Rot Caused by Fusarium spp. Bio Protoc (2018), 8(10), e2859. DOI:10.21769 / BioProtoc.2859.

[0086] Figure 3 The disease severity index (DSI) of maize seedlings corresponding to the control group (FgCK), Examples 1, Examples 3-4, Examples 6-8 and Comparative Example 1 is shown.

[0087] pass Figure 3 It can be seen that, compared with the control group and Comparative Example 1, the disease severity index of Examples 1, 3-4, and 6-8 in this invention has decreased. This indicates that, compared with no selenium treatment or although selenium treatment was performed, the inorganic selenium impregnation treatment of Comparative Example 1 was used, the method of coating corn seeds with selenium composite initiator used in this invention can more effectively reduce the disease severity index. This also means that this invention better achieves the inhibitory effect on deoxynivalenol.

[0088] pass Figure 3 It can also be seen that the severity index of the disease in Example 1 is the lowest, while that in Examples 3 and 4 is not as good as that in Example 1. This shows that when the stabilizer is adjusted to a single type, it will reduce the inhibitory effect on deoxynivalenol to a certain extent. This also proves that when a combination of polysaccharides and peptides is used as stabilizers, it can better exert the effect of deoxynivalenol production inhibitor.

[0089] The disease severity index of Examples 6-8 is slightly higher than that of Examples 1 and 3-4, indicating that the choice of selenium concentration in this invention has a significant impact on the disease severity index of maize seedlings. When the selenium concentration of Example 6 is 25 mg / L or the selenium concentration of Example 7 is 75 mg / L, the inhibitory effect on Fusarium graminearum fungus is not as good as that of Example 1 with 50 mg / L. When the selenium concentration is further increased, i.e., 100 mg / L is used in Example 8, the disease severity index is lower than that of Examples 6 and 7, but still higher than that of Examples 1 and 3-4. This fully demonstrates that by controlling the selenium concentration within a suitable range (40-60 mg / L), this invention can obtain a more superior inhibitory effect on Fusarium graminearum enol.

[0090] 3. Extraction and analysis of deoxynivalenol

[0091] Accurately weigh 5g of straw sample into a 50mL centrifuge tube, add 20mL of acetonitrile / water / acetic acid solution (70:29:1, v / v / v) to extract deoxynivalenol, vortex for 30min, and then centrifuge. Take 0.5mL of the supernatant and mix it with 0.5mL of water, centrifuge at 12000rpm / min for 10min at 4℃, filter the supernatant through a 0.2μm polytetrafluoroethylene filter membrane, and collect the filtrate in a sample vial. Analyze the deoxynivalenol content using a SHIMADZU 8045 mass spectrometry system.

[0092] The levels of deoxynivalenol (DON) in maize seedlings treated with control group (CK), reference group (FgCK), Examples 1-10 and comparative Examples 1-2 were analyzed and measured. The results are shown in Table 1.

[0093] Table 1

[0094] CK / FgCK 0.032 Example 1 0.015 Example 2 0.016 Example 3 0.018 Example 4 0.019 Example 5 0.022 Example 6 0.022 Example 7 0.023 Example 8 0.021 Example 9 0.023 Example 10 0.022 Comparative Example 1 0.028 Comparative Example 2 0.026

[0095] As shown in Table 1, compared with FgCK and Comparative Examples 1-2, the levels of deoxynivalenol in Examples 1-10 provided by this invention are significantly reduced. This indicates that the selenium composite initiator provided by this invention can significantly alleviate the impact of Fusarium graminearum infection on maize crop growth and effectively reduce the level of deoxynivalenol. This also demonstrates that when Comparative Example 1 uses sodium selenite as a selenium source to directly impregnate maize seeds, its effect on reducing deoxynivalenol content is significantly less than that of this invention; and when Comparative Example 2 only uses stirring preparation instead of the instantaneous nanoprecipitation method, its inhibitory effect on deoxynivalenol is also less than that of this invention.

[0096] The lowest deoxynivalenol content was observed in Examples 1 and 2, indicating the best inhibitory effect on deoxynivalenol. The deoxynivalenol content in Examples 3 and 4 was slightly higher than in Example 1, suggesting that using a single type of stabilizer resulted in a slight decrease in deoxynivalenol inhibition compared to a combination of two stabilizers. Examples 9 and 10 used combinations of polypeptides or polysaccharides with other types of stabilizers. A comparison with Example 1 shows that even though Examples 9 and 10 also used combined stabilizers, replacing the polysaccharides or polypeptides in Example 1 with other types of stabilizers did not significantly reduce the inhibitory effect on deoxynivalenol compared to Example 1. This further confirms that the stabilizer using a combination of polysaccharides and polypeptides has a superior effect in reducing deoxynivalenol levels.

[0097] A comparison of Examples 1 and Examples 5-8 shows that when the selenium concentration is less than the concentration specified in this application (40-60 mg / L) (Examples 5-6), or greater than the concentration specified in this application (Examples 7-8), the inhibitory effect on deoxynivalenol decreases, becoming less effective than in Example 1. Particularly in Example 8, when the selenium concentration is twice that used in this invention, the increased selenium concentration actually makes the inhibitory effect on deoxynivalenol less effective than in Example 1. This fully demonstrates that by controlling the selenium concentration within a suitable range (40-60 mg / L), a superior inhibitory effect on deoxynivalenol can be obtained in this invention.

[0098] 4. Analysis of plant hormone JA, total flavonoids and total phenolic compounds in maize plants

[0099] For metabolite extraction, 100 mg of sample was homogenized in 1 mL of ultrapure water containing 80% methanol and 0.1% formic acid. The homogenate was sonicated for 30 min to improve extraction efficiency, followed by centrifugation at 12,000 rpm for 5 min. To ensure maximum metabolite recovery, the extraction process was repeated twice, and all supernatants were combined. The combined supernatant was dried under nitrogen and then brought to a final volume of 100 μL with 60% ethanol. Before analysis, the sample was purified with 100 mg of C18 material and filtered through a 0.22 μm nylon syringe filter to remove particulate matter.

[0100] JA analysis was performed using ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS / MS) on an Agilent G6465B triple quadrupole system equipped with a reversed-phase C18 column (EclipsePlus C18, 2.1 × 50 mm, 1.8 μm). Total flavonoids and total phenols were determined using the LHT-2-G plant flavonoid assay kit and the TP-2-G plant total phenols assay kit from Suzhou Keming Co., Ltd., respectively.

[0101] Figures 4-6 The following figures show the jasmonic acid (JA) content in maize seedlings corresponding to the control group (FgCK), Examples 1, 3-4, 6-8, and Comparative Example 1. Figure 4 ), total flavonoids ( Figure 5 ) and total phenols ( Figure 6 The content of ).

[0102] pass Figures 4-6 It can be seen that, compared with the reference group and comparative example 1, the corn seedlings of examples 1, 3-4 and 6-8 in this invention have higher contents of jasmonic acid (JA) and total phenols, while the total flavonoid content is lower. This reflects that antibacterial substances are accumulated in the corn seedlings of examples 1, 3-4 and 6-8. This accumulation is mainly due to the retargeting of downstream phenylpropane metabolites after the JA signaling pathway is activated.

[0103] In addition, correlation analyses were performed on the disease severity index (DSI) and DON content with jasmonic acid (JA), total flavonoids, and total phenolic content in the maize seedlings treated in Example 1. Figure 7 As shown.

[0104] Figure 7The correlation and interaction among JA, total flavonoids, and total phenols are shown. The correlation coefficient matrix reveals a highly significant positive correlation (0.79***) between JA and total phenols under Fusarium graminearum infection, while a significant negative correlation (0.72***) is observed between total flavonoids and total phenols. The network diagram on the left further reveals the complexity of the relationship between DON, DSI, and the three: the orange line (total flavonoids) and the green / white line (total phenols) form mutually exclusive connections. This fully verifies that seeds coated with selenium-containing complex initiators enhance Fusarium graminearum resistance and inhibit toxin accumulation by activating JA signaling and antagonizing the balance of flavonoid and phenolic metabolism, promoting the synthesis of antibacterial phenolic substances while inhibiting flavonoid accumulation.

[0105] according to Figure 7 The content shown is combined with the information provided by this invention. Figures 4-6 This also fully demonstrates that the various embodiments provided by the present invention can effectively enhance the resistance of Fusarium graminearum and inhibit toxin accumulation.

[0106] In summary, this invention utilizes a selenium-based complex initiator to coat maize seeds, creating maize seeds containing this initiator. This selenium-based initiator enhances maize's resistance to Fusarium graminearum and inhibits the synthesis of deoxynivalenols by reprogramming seed metabolism and activating jasmonic acid (JA)-mediated defense responses during the seedling stage. Simultaneously, it shifts seedling phenylpropanoid metabolism from flavonoid biosynthesis to antifungal phenolic biosynthesis, thereby synchronizing seed vigor with innate immunity. This provides an effective strategy for maize crop protection and effectively inhibits the formation of maize toxins.

Claims

1. The use of a selenium-containing composite initiator as an inhibitor of deoxynivalenol production in maize, characterized in that, The selenium-containing composite initiator is coated onto corn seeds to form corn seeds containing the selenium-containing composite initiator. The corn seeds containing the selenium complex initiator were prepared using a method comprising the following steps: (1) A first multi-inlet vortex mixer with four channels is provided. A selenium source is introduced into the first channel of the first multi-inlet vortex mixer as a first liquid flow, a stabilizer is introduced into the second channel as a second liquid flow, water is introduced into the third channel as a third liquid flow, and a reducing agent is introduced into the fourth channel as a fourth liquid flow. The first liquid flow, the second liquid flow, the third liquid flow and the fourth liquid flow collide and mix instantaneously in the first multi-inlet vortex mixer to form a selenium-containing composite suspension. (2) Provide a second multi-inlet vortex mixer connected in series with the first multi-inlet vortex mixer in step (1), and introduce the selenium-containing composite suspension obtained in step (1) into the first channel of the second multi-inlet vortex mixer as the first liquid flow, introduce the first ligand into the second channel as the second liquid flow, introduce the second ligand into the third channel as the third liquid flow, and introduce water into the fourth channel as the fourth liquid flow. The first liquid flow, the second liquid flow, the third liquid flow and the fourth liquid flow collide and mix instantaneously in the second multi-inlet vortex mixer to form a selenium-containing coating suspension; (3) The disinfected corn seeds are first soaked in the selenium-containing coating suspension obtained in step (2), and then soaked in the cross-linking agent solution. After drying, corn seeds containing selenium composite initiator are formed. The stabilizer is a combination of polysaccharides and polypeptides; In step (1) of preparing the corn seeds containing the selenium complex initiator, the flow rates of the first liquid flow and the second liquid flow are each independently 4~8 mL / min, and the flow rates of the third liquid flow and the fourth liquid flow are each independently 25~50 mL / min. In step (2) of preparing the corn seeds containing the selenium complex initiator, the first ligand is carboxymethyl chitosan and the second ligand is pectin; or, the first ligand is sodium alginate and the second ligand is gum arabic. In step (2) of preparing the corn seeds containing the selenium composite initiator, the concentration of selenium in the selenium-coating suspension formed is 40~60 mg / L; In step (2) of preparing the corn seeds containing the selenium complex initiator, the flow rates of the first liquid flow and the second liquid flow are each independently 5~10 mL / min, and the flow rates of the third liquid flow and the fourth liquid flow are each independently 30~60 mL / min.

2. The use according to claim 1, characterized in that, In step (1) of preparing the corn seeds containing the selenium complex initiator, the selenium source is any one or a combination of at least two of sodium selenite, sodium selenate, or selenium dioxide.

3. The use according to claim 1, characterized in that, The reducing agent is any one or a combination of at least two of the following: cysteine, hydrazine hydrate, ascorbic acid, sodium sulfite, sodium thiosulfate, or glutathione.

4. The use according to claim 1, characterized in that, The molar ratio of selenium to reducing agent in the selenium source is 1:(2~6).

5. The use according to claim 1, characterized in that, The mass ratio of selenium in the selenium source to the mass of the stabilizer is 1:(5~50).

6. The use according to claim 1, characterized in that, In the selenium-containing composite suspension obtained in step (1), the average particle size of selenium is 20~80nm.

7. The use according to claim 1, characterized in that, In step (3) of preparing the corn seeds containing the selenium complex initiator, the soaking time is 2-12 hours.

8. The use according to claim 1, characterized in that, In step (3) of preparing the corn seeds containing the selenium complex initiator, the soaking time is 3-6 hours.

9. The use according to claim 1, characterized in that, In step (3) of preparing the corn seeds containing the selenium composite initiator, the crosslinking agent is any one or a combination of at least two of zinc sulfate, copper sulfate, calcium chloride or manganese sulfate.

10. The use according to claim 1, characterized in that, The mass concentration of the crosslinking agent is 1-5%.

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