Antibacterial agent containing polar deep sea antibacterial peptide, fertilizer and preparation method of antibacterial agent

Abstract

The invention relates to an antibacterial agent containing polar deep sea antibacterial peptide, a fertilizer and a preparation method thereof, the antibacterial agent is characterized by comprising antibacterial peptide and ethanolamine, and the antibacterial peptide is a metabolite of marine bacillus; the antibacterial agent has a broad-spectrum antibacterial property and a good antibacterial effect, and has a relatively good effect when being applied to agriculture.

Description

Technical Field

[0001] The present invention relates to an antibacterial agent containing polar deep-sea antibacterial peptides, a fertilizer and a preparation method thereof. Background Art

[0002] Antimicrobial peptides have broad-spectrum antibacterial, antiviral and antifungal activities. They mainly form electrostatic attraction with negatively charged bacterial membranes through cationic residues, cover the surface of bacterial membranes, and penetrate hydrophobic residues into the phospholipid bilayer. Relying on the amphiphilic properties of antimicrobial peptides, they disintegrate cell membrane components, causing the bacterial contents to flow out and die. Unlike antibiotics that target intracellular targets, the antibacterial mechanism of antimicrobial peptides that physically breaks the membrane makes it difficult for bacteria to develop drug resistance.

[0003] Deep-sea antimicrobial peptides are a class of potent bioactive compounds with broad-spectrum antibacterial, antiviral, and antifungal activities. They are key components of the immune defenses of marine organisms, adapted to the harsher marine environment and possessing high stability. However, their antimicrobial properties need to be improved. However, their disadvantage is that their peptide chains are too long, resulting in a high molecular weight. This makes them incompatible with other antimicrobial substances. Even if these are combined, their antimicrobial properties may be reduced, failing to achieve the desired antimicrobial properties.

[0004] The polar deep-sea antimicrobial peptide used in this application is a polar deep-sea antimicrobial peptide obtained by cultivating Bacillus isolated from marine sediment samples obtained during the sixth Arctic scientific expedition. It has good antibacterial properties and great research potential.

[0005] Currently, there are no products or methods that further enhance the antimicrobial properties of polar deep-sea antimicrobial peptides. There are also no solutions to the problem of polar deep-sea antimicrobial peptides being too long, which affects their compatibility and results in poor results when combined with other ingredients. There are also no products or methods that combine polar deep-sea antimicrobial peptides with fertilizers with minimal impact on their antimicrobial properties. Summary of the Invention

[0006] The present invention provides an antibacterial agent and fertilizer containing polar deep-sea antimicrobial peptides and a preparation method thereof, and solves the following technical problems: 1) further improving the antibacterial properties of polar deep-sea antimicrobial peptides; 2) improving the compatibility of polar deep-sea antimicrobial peptides and improving their antimicrobial properties; and 3) combining polar deep-sea antimicrobial peptides with fertilizers to affect their antimicrobial properties.

[0007] In order to solve the above technical problems, the present invention adopts the following technical solutions: An antimicrobial agent containing polar deep-sea antimicrobial peptides, comprising antimicrobial peptides and ethanolamine, wherein the antimicrobial peptides are metabolites of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY; The ethanolamine is one or more of monoethanolamine, diethanolamine and triethanolamine; The mass ratio of antimicrobial peptide to ethanolamine is 100:0.3-1.

[0008] It also includes chitosan oligosaccharide; the mass ratio of antimicrobial peptide to chitosan oligosaccharide is 100:0-20; The ethanolamine is one or both of diethanolamine and triethanolamine.

[0009] The pH is 4.0-6.5.

[0010] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding antimicrobial peptide and ethanolamine into water, dissolving them, and stirring them to obtain an antimicrobial agent aqueous solution of polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0011] The method for preparing an antimicrobial agent containing polar deep-sea antimicrobial peptides is characterized by following the steps: adding antimicrobial peptide and ethanolamine into water, dissolving them, and stirring to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0012] Fertilizers containing polar deep-sea antimicrobial peptides, including polar deep-sea antimicrobial peptide antimicrobial agents and urea; and do not contain ionic fertilizer raw materials; The ionic fertilizer raw materials include one or more of potassium nitrate, potassium dihydrogen phosphate, potassium chloride, potassium sulfate, ammonium chloride, ammonium sulfate and ammonium dihydrogen phosphate.

[0013] The invention also includes auxiliary materials; the auxiliary materials are one or more of bentonite, talc, attapulgite and humic acid.

[0014] A method for preparing a fertilizer containing polar deep-sea antimicrobial peptides, comprising adding an antimicrobial agent of polar deep-sea antimicrobial peptides to urea and mixing the mixture to obtain a fertilizer containing polar deep-sea antimicrobial peptides; or The antimicrobial agent of the polar deep-sea antimicrobial peptide is added to urea and mixed evenly, and then granulated to obtain a fertilizer containing the polar deep-sea antimicrobial peptide; or, Add urea to the antibacterial agent aqueous solution of polar deep-sea antimicrobial peptide and dissolve it to obtain a fertilizer containing polar deep-sea antimicrobial peptide; or, Adding the antibacterial agent of urea polar deep-sea antimicrobial peptide into water and dissolving it to obtain a fertilizer containing polar deep-sea antimicrobial peptide; or, The antibacterial agent of the polar deep-sea antimicrobial peptide is added to the urine slurry, and the urea containing the polar deep-sea antimicrobial peptide is obtained by spraying and granulating. The mass ratio of the antimicrobial agent, auxiliary materials and urea of ​​the polar deep-sea antimicrobial peptide is 1-950:0-500:50-999.

[0015] It also includes drying, cooling and screening.

[0016] The invention has the following beneficial technical effects: 1. This application adds ethanolamine. After ethanolamine has a small molecular weight and electrostatically coordinates with the antimicrobial peptide, the product has no special phenomenon. The amino group of ethanolamine and the carboxyl group of the antimicrobial peptide electrostatically coordinate to form a stable complex, which can enhance the adsorption of the antimicrobial peptide and have a better antibacterial and bactericidal effect; diethanolamine or triethanolamine has a better effect.

[0017] 2. This application limits the amount of chitosan oligosaccharide added. If too much chitosan oligosaccharide is added, the concentration of chitosan oligosaccharide in the solution will be high, and the amino groups on the chitosan oligosaccharide will compete with the amino groups of ethanolamine, thereby affecting the effect of the product.

[0018] 3. This application defines the pH in order to keep the antimicrobial peptide positively charged without affecting the efficacy of the product.

[0019] 4. The present application prepares the antimicrobial peptide and ethanolamine into an antimicrobial peptide-ethanolamine solution first, thereby improving the compatibility of the antimicrobial peptide without affecting the antimicrobial effect.

[0020] 5. The fertilizer containing polar deep-sea antimicrobial peptides in this application does not contain ionic fertilizer raw materials. This is because the fertilizer standard requires that the single nutrient content of nitrogen, phosphorus, and potassium be ≥4%. When the concentration of nitrogen, phosphorus, and potassium in the ionic fertilizer is ≥4%, it will affect the effectiveness of the antimicrobial peptides. However, when only urea is used, it will not affect the effectiveness of the antimicrobial peptides.

[0021] 6. This application combines antimicrobial peptides, ethanolamine and chitosan oligosaccharides to further enhance the antimicrobial effect of antimicrobial peptides. DETAILED DESCRIPTION

[0022] In order to better understand the above technical solution, the technical solution of the present invention is described in detail below with reference to preferred examples. Example 1

[0023] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide and monoethanolamine in a mass ratio of 100:0.5, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0024] The pH is 6.2.

[0025] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and monoethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0026] The fertilizer containing polar deep-sea antimicrobial peptide is composed of the antimicrobial agent of polar deep-sea antimicrobial peptide and urea.

[0027] The preparation method of the fertilizer containing the polar deep-sea antimicrobial peptide comprises the following steps: adding 4 parts of an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide into 996 parts of urea slurry, mixing the mixture, and spraying and granulating the mixture to obtain urea containing the polar deep-sea antimicrobial peptide.

[0028] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 2

[0029] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide and diethanolamine in a mass ratio of 100:0.5, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0030] The pH is 6.2.

[0031] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and diethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0032] The fertilizer containing polar deep-sea antimicrobial peptide is composed of the antimicrobial agent of polar deep-sea antimicrobial peptide and urea.

[0033] The preparation method of the fertilizer containing the polar deep-sea antimicrobial peptide comprises the following steps: adding 4 parts of an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide into 996 parts of urea slurry, mixing the mixture, and spraying and granulating the mixture to obtain urea containing the polar deep-sea antimicrobial peptide.

[0034] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 3

[0035] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide and triethanolamine in a mass ratio of 100:0.5, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0036] The pH is 6.2.

[0037] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding antimicrobial peptide and triethanolamine into water to dissolve, stirring, and adjusting pH to obtain polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0038] The fertilizer containing polar deep-sea antimicrobial peptide is composed of the antimicrobial agent of polar deep-sea antimicrobial peptide and urea.

[0039] The preparation method of the fertilizer containing the polar deep-sea antimicrobial peptide comprises the following steps: adding 4 parts of an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide into 996 parts of urea slurry, mixing the mixture, and spraying and granulating the mixture to obtain urea containing the polar deep-sea antimicrobial peptide.

[0040] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 4

[0041] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide, monoethanolamine, and chitosan oligosaccharide in a mass ratio of 100:0.5:15, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0042] The pH is 6.2.

[0043] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and monoethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0044] The fertilizer containing polar deep-sea antimicrobial peptide is composed of the antimicrobial agent of polar deep-sea antimicrobial peptide and urea.

[0045] The preparation method of the fertilizer containing the polar deep-sea antimicrobial peptide comprises the following steps: adding 4 parts of an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide into 996 parts of urea slurry, mixing the mixture, and spraying and granulating the mixture to obtain urea containing the polar deep-sea antimicrobial peptide.

[0046] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 5

[0047] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide, diethanolamine, and chitosan oligosaccharide in a mass ratio of 100:0.5:15, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0048] The pH is 6.2.

[0049] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and diethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0050] The fertilizer containing polar deep-sea antimicrobial peptide is composed of the antimicrobial agent of polar deep-sea antimicrobial peptide and urea.

[0051] The preparation method of the fertilizer containing the polar deep-sea antimicrobial peptide comprises the following steps: adding 4 parts of an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide into 996 parts of urea slurry, mixing the mixture, and spraying and granulating the mixture to obtain urea containing the polar deep-sea antimicrobial peptide.

[0052] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 6

[0053] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide, triethanolamine, and chitosan oligosaccharide in a mass ratio of 100:0.5:15, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0054] The pH is 6.2.

[0055] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and triethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0056] The fertilizer containing polar deep-sea antimicrobial peptide is composed of the antimicrobial agent of polar deep-sea antimicrobial peptide and urea.

[0057] The preparation method of the fertilizer containing the polar deep-sea antimicrobial peptide comprises the following steps: adding 4 parts of an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide into 996 parts of urea slurry, mixing the mixture, and spraying and granulating the mixture to obtain urea containing the polar deep-sea antimicrobial peptide.

[0058] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 7

[0059] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide, triethanolamine, and chitosan oligosaccharide in a mass ratio of 100:0.5:15, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0060] The pH is 6.2.

[0061] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and triethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0062] The fertilizer containing polar deep-sea antimicrobial peptide is composed of polar deep-sea antimicrobial peptide antimicrobial agent powder and urea in a mass ratio of 80:20.

[0063] The invention discloses a method for preparing a fertilizer containing polar deep-sea antimicrobial peptides. The method comprises the following steps: uniformly mixing urea and antimicrobial agent powder of the polar deep-sea antimicrobial peptides to obtain a water-soluble fertilizer containing the polar deep-sea antimicrobial peptides. Example 8

[0064] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide and diethanolamine in a mass ratio of 100:0.8, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0065] The pH is 5.7.

[0066] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and diethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0067] A fertilizer containing polar deep-sea antimicrobial peptide, which is composed of an antimicrobial agent of polar deep-sea antimicrobial peptide, urea, humic acid and bentonite; The invention discloses a method for preparing a fertilizer containing polar deep-sea antimicrobial peptides. The method comprises the following steps: uniformly mixing 650 parts of urea, 100 parts of humic acid and 250 parts of bentonite; feeding the mixture into a granulation drum via a conveyor belt; spraying 3 parts of an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptides into the granulation drum at an inlet end thereof to form granules together with the urea, humic acid and bentonite; and drying, cooling and screening the mixture to obtain humic acid urea.

[0068] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 9

[0069] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide, triethanolamine, and chitosan oligosaccharide in a mass ratio of 100:0.3:10, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0070] The pH is 5.2.

[0071] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: adding the antimicrobial peptide and triethanolamine into water to dissolve, stirring, and adjusting the pH to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0072] The fertilizer containing polar deep-sea antimicrobial peptide is composed of polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution and urea in a mass ratio of 920:80.

[0073] The invention discloses a preparation method of a fertilizer containing polar deep-sea antimicrobial peptides. Urea is added into an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptides for dissolution, thereby obtaining a water-soluble fertilizer containing the polar deep-sea antimicrobial peptides.

[0074] The solid content of the polar deep-sea antimicrobial peptide antimicrobial aqueous solution is 20%. Example 10

[0075] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide, monoethanolamine, and chitosan oligosaccharide in a mass ratio of 100:0.9:12, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0076] The pH is 6.1.

[0077] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: Adding monoethanolamine to the antimicrobial peptide solution, mixing evenly, stirring, and adjusting the pH to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0078] The fertilizer containing polar deep-sea antimicrobial peptide is composed of polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution and urea in a mass ratio of 80:20.

[0079] The invention discloses a preparation method of a fertilizer containing polar deep-sea antimicrobial peptides. Urea is added into an antimicrobial agent aqueous solution of the polar deep-sea antimicrobial peptides for dissolution, thereby obtaining a water-soluble fertilizer containing the polar deep-sea antimicrobial peptides.

[0080] The liquid-solid content of antimicrobial peptide is 20%. Example 11

[0081] An antimicrobial agent containing a polar deep-sea antimicrobial peptide, comprising an antimicrobial peptide, monoethanolamine, and chitosan oligosaccharide in a mass ratio of 100:0.9:12, wherein the antimicrobial peptide is a metabolite of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY.

[0082] The pH is 6.1.

[0083] The preparation method of the antimicrobial agent containing polar deep-sea antimicrobial peptide is carried out according to the following steps: Adding monoethanolamine to the antimicrobial peptide solution, mixing evenly, stirring, and adjusting the pH to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

[0084] The urea containing polar deep-sea antimicrobial peptide consists of the antimicrobial agent of polar deep-sea antimicrobial peptide and urea.

[0085] The preparation method of urea containing polar deep-sea antimicrobial peptide comprises the following steps: adding 6 parts of polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution into urine slurry and mixing the mixture; and spraying and granulating the mixture to obtain urea containing polar deep-sea antimicrobial peptide.

[0086] The liquid-solid content of antimicrobial peptide is 20%.

[0087] Except for the different treatment methods, the other conditions in the above embodiments are the same.

[0088] The beneficial effects of the present invention are further illustrated below in conjunction with test data: Experiment 1 1.1 Experimental location: Gansu Smart Agriculture Engineering Technology Research Center.

[0089] 1.2 Experimental testing: Observe the state of the solution; Minimum inhibitory concentration (μg / mL) against Staphylococcus aureus, Rhizoctonia solani, and Fusarium graminearum.

[0090] 1.3 Test materials: Comparative 1 (except that monoethanolamine was not added, all other materials were the same as those in Example 1), Comparison 2 (chitosan oligosaccharide), Comparison 3 (except that monoethanolamine was not added, all others were consistent with Example 4), Comparison 4 (except that the antimicrobial peptide, monoethanolamine and chitosan oligosaccharide were composed in a mass ratio of 100:0.5:25, all others were consistent with Example 4) and the antibacterial powder of polar deep-sea antimicrobial peptide prepared in Examples 1 to 6.

[0091] 1.4 Detection method: Dilute the third generation bacterial suspension of the three bacteria to 1×10 6 cfu / mL, and each treatment was diluted with PBS to different concentrations. The bacterial suspension and 100 μL of each treatment dilution were mixed in a 96-well polystyrene microplate (each concentration was replicated three times). After incubating the 96-well plate at 37°C for 24 hours, the minimum inhibitory concentration (MIC) was determined by visually observing the turbidity in the wells.

[0092] Except for the different treatments in this experiment, the other treatments were the same.

[0093] 2 Results and Analysis Observe the state of the solution; the minimum inhibitory concentration (μg / mL) for Staphylococcus aureus (recorded as Staphylococcus aureus), Rhizoctonia solani (recorded as Rhizoctonia solani), and Fusarium graminearum (recorded as Fusarium graminearum) is shown in Table 1

[0094] Note: The phenomenon in this test is the phenomenon after mixing and standing for 2 hours. Comparison 3 and Comparison 4 have precipitation, and the upper clear liquid is absorbed.

[0095] From the comparison of the data of Comparative 1, Example 1, Example 2 and Example 3, it can be seen that the addition of ethanolamine can further enhance the antibacterial effect of the antimicrobial peptide.

[0096] From the comparison of the data of Comparison 3, Comparison 4 and Example 4, it can be seen that when ethanolamine is not added, the compatibility of antimicrobial peptides and chitosan oligosaccharides is poor, and the antibacterial activity is reduced after compatibility; the concentration of chitosan oligosaccharides also affects the antibacterial activity of antimicrobial peptides. It may be that the increased concentration of chitosan oligosaccharides competes with ethanolamine and electrostatically coordinates with the antimicrobial peptide, thereby producing large molecules and forming precipitation, which in turn affects the use effect.

[0097] From the comparison of the data of Examples 1, 2, and 3 with those of Examples 4, 5, and 6, it can be seen that the effects of using diethanolamine and triethanolamine are better than those of monoethanolamine.

[0098] Experiment 2 Fertilizer standards require that the minimum single nutrient content be ≥4% or 40g / L.

[0099] 1.1 Experimental location: Gansu Smart Agriculture Engineering Technology Research Center.

[0100] 1.2 Experimental testing: Observe the state of the solution; Minimum inhibitory concentration (μg / mL) against Staphylococcus aureus, Rhizoctonia solani, and Fusarium graminearum.

[0101] 1.3 Test materials: blank (the antibacterial powder of polar deep-sea antimicrobial peptide prepared in Example 6), comparative 5 (the urea in Example 7 was replaced with industrial-grade potassium sulfate, and the rest were consistent with Example 7), comparative 6 (the urea in Example 7 was replaced with industrial-grade ammonium dihydrogen phosphate, and the rest were consistent with Example 7), comparative 7 (the 7 parts of urea in Example 7 were replaced with 7 parts of industrial-grade potassium sulfate, and the rest were unchanged, and the rest were consistent with Example 7), comparative 8 (the 6 parts of urea in Example 7 were replaced with 6 parts of industrial-grade ammonium dihydrogen phosphate, and the rest were unchanged, and the rest were consistent with Example 7) and the water-soluble fertilizer prepared in Example 7.

[0102] Note: Comparison 6 and Comparison 7 meet the fertilizer standard requirement of a single macronutrient content of not less than 4%. The content of P2O5 in Comparison 6 is 4%; the content of K2O in Comparison 7 is 4%. The content of K2O in industrial-grade potassium sulfate is 54%; the content of P2O5 in ammonium dihydrogen phosphate is 61%.

[0103] 1.4 Detection method: Dilute the third generation bacterial suspension of the three bacteria to 1×10 6cfu / mL, and each treatment was diluted with PBS to different concentrations. The bacterial suspension and 100 μL of each treatment dilution were mixed in a 96-well polystyrene microplate (each concentration was replicated three times). After incubating the 96-well plate at 37°C for 24 hours, the minimum inhibitory concentration (MIC) was determined by visually observing the turbidity in the wells.

[0104] The concentration of antimicrobial peptides was used as the standard for each treatment dilution.

[0105] Except for the different treatments in this experiment, the other treatments were the same.

[0106]

[0107] Note: The blank uses the data of Example 6 in Table 1, because the antibacterial powders of polar deep-sea antimicrobial peptides in Example 6 and Example 7 are the same.

[0108] As can be seen from the solutions in Table 2 for the blank, Comparative 5, Comparative 6, Comparative 7, Comparative 8, and Example 7, no precipitation was produced after dissolving the antimicrobial powder of the polar deep-sea antimicrobial peptide in water-soluble fertilizer, indicating that the antimicrobial agent of the polar deep-sea antimicrobial peptide and fertilizer are compatible. The antimicrobial activity of the polar deep-sea antimicrobial peptide antimicrobial agent was minimally affected when combined with urea, resulting in a superior antimicrobial effect. However, when ionic fertilizer raw materials were added, the antimicrobial activity was significantly weakened, even when meeting the minimum requirements of fertilizer standards. Furthermore, the antimicrobial activity decreased with increasing amounts of ionic fertilizer raw materials. Therefore, to maintain the antimicrobial activity of the polar deep-sea antimicrobial peptide antimicrobial agent, it should not be combined with ionic fertilizer raw materials.

[0109] Discussion: The reason why ionic fertilizer raw materials affect the antibacterial activity of polar deep-sea antimicrobial peptides may be salt shielding. The shielding effect of potassium sulfate is higher than that of ammonium dihydrogen phosphate, which may be related to the fact that potassium sulfate is a strong acid and strong base salt, while ammonium dihydrogen phosphate is a weak acid and weak base salt.

Claims

1. An antimicrobial agent containing polar deep-sea antimicrobial peptides, characterized in that: The invention comprises antimicrobial peptides and ethanolamine, wherein the antimicrobial peptides are metabolites of marine Bacillus; The antimicrobial peptide consists of 93 amino acids, and the sequence is as follows: MKLKKVLTGSALSLALLVSAAPAFAANPDGTHSEQTSVASSDFKANIYTKTETNDTGIFINSYTDADGITWYLKKITKNNNGTWTAYYEGRKY; The ethanolamine is one or more of monoethanolamine, diethanolamine and triethanolamine; The mass ratio of antimicrobial peptide to ethanolamine is 100:0.3-1.

2. The antimicrobial agent containing polar deep-sea antimicrobial peptide according to claim 1, characterized in that: It also includes chitosan oligosaccharide; the mass ratio of antimicrobial peptide to chitosan oligosaccharide is 100:0-20; The ethanolamine is one or both of diethanolamine and triethanolamine.

3. The antimicrobial agent containing the polar deep-sea antimicrobial peptide according to claim 1 or 2, characterized in that: The pH is 4.0-6.

5.

4. The method for preparing an antimicrobial agent containing a polar deep-sea antimicrobial peptide according to claim 1, wherein: Follow the steps below, adding antimicrobial peptide and ethanolamine into water, dissolving them, and stirring them to obtain an antimicrobial agent aqueous solution of polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

5. The method for preparing the antimicrobial agent containing polar deep-sea antimicrobial peptide according to claim 1, characterized in that: Follow the steps below, adding antimicrobial peptide and ethanolamine into water, dissolving them, and stirring to obtain an antimicrobial peptide-ethanolamine solution; Adding chitosan oligosaccharide to the antimicrobial peptide-ethanolamine solution and dissolving it to obtain the polar deep-sea antimicrobial peptide antimicrobial agent aqueous solution; The antibacterial agent of the polar deep-sea antimicrobial peptide is spray-dried to obtain the antibacterial agent powder of the polar deep-sea antimicrobial peptide; The antibacterial agent of the polar deep-sea antimicrobial peptide comprises an antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and an antibacterial agent powder of the polar deep-sea antimicrobial peptide.

6. The fertilizer containing polar deep-sea antimicrobial peptide according to any one of claims 1 to 5, characterized in that: Including antimicrobial agents and urea from polar deep-sea antimicrobial peptides; and does not contain ionic fertilizer raw materials; The ionic fertilizer raw materials include one or more of potassium nitrate, potassium dihydrogen phosphate, potassium chloride, potassium sulfate, ammonium chloride, ammonium sulfate and ammonium dihydrogen phosphate.

7. The fertilizer containing polar deep-sea antimicrobial peptide according to claim 6, characterized in that: The invention also includes auxiliary materials; the auxiliary materials are one or more of bentonite, talc, attapulgite and humic acid.

8. The method for preparing a fertilizer containing polar deep-sea antimicrobial peptide according to claim 6 or 7, characterized in that: Adding the antibacterial agent of the polar deep-sea antimicrobial peptide to urea and mixing them evenly to obtain a fertilizer containing the polar deep-sea antimicrobial peptide; or, The antibacterial agent of the polar deep-sea antimicrobial peptide is added to urea and mixed evenly, and then granulated to obtain a fertilizer containing the polar deep-sea antimicrobial peptide; or, Adding urea to the antibacterial agent aqueous solution of the polar deep-sea antimicrobial peptide and dissolving it to obtain a fertilizer containing the polar deep-sea antimicrobial peptide; or, The antibacterial agent of urea polar deep-sea antimicrobial peptide is added into water and dissolved to obtain a fertilizer containing polar deep-sea antimicrobial peptide; or, The antibacterial agent of the polar deep-sea antimicrobial peptide is added to the urine slurry, and the urea containing the polar deep-sea antimicrobial peptide is obtained by spraying and granulating. The mass ratio of the antimicrobial agent, auxiliary materials and urea of ​​the polar deep-sea antimicrobial peptide is 1-950:0-500:50-999.

9. The method for preparing a fertilizer containing polar deep-sea antimicrobial peptides according to claim 8, wherein: It also includes drying, cooling and screening.

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