CN116041115A - Water-soluble seaweed fertilizer containing bacillus californicus and polyglutamic acid as well as production method and application thereof - Google Patents

Water-soluble seaweed fertilizer containing bacillus californicus and polyglutamic acid as well as production method and application thereof Download PDF

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CN116041115A
CN116041115A CN202211259111.5A CN202211259111A CN116041115A CN 116041115 A CN116041115 A CN 116041115A CN 202211259111 A CN202211259111 A CN 202211259111A CN 116041115 A CN116041115 A CN 116041115A
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bacillus
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californicus
lyhd
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孙志强
李文华
黄晓亚
常城
于海军
郑素莲
李佼倩
赵云福
邹雨平
曹守花
田大伟
王蕾
林程琳
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Yantai Luyun Biotechnology Co ltd
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    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05C—NITROGENOUS FERTILISERS
    • C05C11/00—Other nitrogenous fertilisers
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05F—ORGANIC FERTILISERS NOT COVERED BY SUBCLASSES C05B, C05C, e.g. FERTILISERS FROM WASTE OR REFUSE
    • C05F17/00—Preparation of fertilisers characterised by biological or biochemical treatment steps, e.g. composting or fermentation
    • C05F17/20—Preparation of fertilisers characterised by biological or biochemical treatment steps, e.g. composting or fermentation using specific microorganisms or substances, e.g. enzymes, for activating or stimulating the treatment
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
    • C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
    • C05G3/60—Biocides or preservatives, e.g. disinfectants, pesticides or herbicides; Pest repellants or attractants
    • C—CHEMISTRY; METALLURGY
    • C05—FERTILISERS; MANUFACTURE THEREOF
    • C05G—MIXTURES OF FERTILISERS COVERED INDIVIDUALLY BY DIFFERENT SUBCLASSES OF CLASS C05; MIXTURES OF ONE OR MORE FERTILISERS WITH MATERIALS NOT HAVING A SPECIFIC FERTILISING ACTIVITY, e.g. PESTICIDES, SOIL-CONDITIONERS, WETTING AGENTS; FERTILISERS CHARACTERISED BY THEIR FORM
    • C05G3/00—Mixtures of one or more fertilisers with additives not having a specially fertilising activity
    • C05G3/80—Soil conditioners
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/07—Bacillus
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02W—CLIMATE CHANGE MITIGATION TECHNOLOGIES RELATED TO WASTEWATER TREATMENT OR WASTE MANAGEMENT
    • Y02W30/00—Technologies for solid waste management
    • Y02W30/40—Bio-organic fraction processing; Production of fertilisers from the organic fraction of waste or refuse

Abstract

The invention relates to a water-soluble fertilizer containing bacillus californicus (Bacillus cabrialesii) and polyglutamic acid, and a preparation method and application thereof, wherein the water-soluble seaweed fertilizer comprises bacillus californicus LYHD-001 microbial inoculum; the preservation number of the bacillus californicus LYHD-001 strain is CCTCCNO: M2021209, the preservation date is 2021, 3 and 8 days, the preservation address is China, the university of Wuhan and the university of Wuhan, and the classification name is Bacillus cabrialesii LYHD-001. The method for preparing the bacillus californicus LYHD-001 microbial inoculum and the water-soluble seaweed fertilizer containing the bacillus californicus agent and polyglutamic acid by using the bacillus californicus LYHD-001 has the advantages of short fermentation time, stable and controllable process flow, small and easily-absorbed product molecular weight and obvious fertilizer efficiency, and is suitable for large-scale production.

Description

Water-soluble seaweed fertilizer containing bacillus californicus and polyglutamic acid as well as production method and application thereof
Technical Field
The invention relates to a water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid, and a production method and application thereof, and belongs to the technical field of biological bacterial fertilizers.
Background
Seaweed is an algae plant with strong stress resistance and fast growth, and widely exists in various coastal areas. Algae have been used mainly as food for people, and in recent years, excessive use of chemical fertilizers causes problems such as soil hardening, pollution of nitrate in vegetables, pollution of groundwater and the like, which seriously threatens the health of human beings. Seaweed fertilizer is a natural fertilizer, and is widely paid attention to by the advantages of safety, effectiveness, degradability and the like.
Compared with chemical fertilizers, the seaweed fertilizer can effectively promote the growth of crops due to the auxin, auxin analogues and the like contained in brown seaweed such as kelp and the like. Wherein, a large amount of abscisic acid is found in the extract of the Ascophyllum nodosum, and the abscisic acid is substances which can promote the storage proteins, sugar and the like of fruits, thereby improving the yield; brown algae contain various betaines and betaine compounds. Betaine can be used as a compatible solute in plants, so that osmotic stress caused by salinity and drought can be relieved, and the stress resistance of the plants can be enhanced; alginic acid extracted from kelp can improve soil aggregate structure, so as to be beneficial to the maintenance of soil moisture, and the good soil ventilation property can stimulate the activity of crop root systems and the growth of rhizosphere microorganisms; simultaneously, the kelp also has the function of reducing the nitrate content in fruits and vegetables, and can improve the content of vitamins, soluble sugar, carotene, carotenoid and the like in the vegetables.
However, the most critical step in the seaweed fertilizer production process is the digestion of algae, which aims at breaking up cells to dissolve out intracellular contents, and degrading macromolecular substances in cells into soluble micromolecular substances which are easy to be absorbed and utilized. The preparation process and the seaweed wall breaking method of the seaweed fertilizer reported at home and abroad mainly comprise a mechanical method, a chemical method and a biological method.
The mechanical method and the chemical method have the problems of low cell disruption rate, low extraction efficiency, strict conditions, high cost, pollution, denaturation and inactivation of bioactive substances and the like, and are not suitable for large-scale production; the biological method mainly comprises a biological enzyme method and a microbial fermentation method, wherein the enzyme method mainly utilizes enzyme to degrade crude fibers in algae and release contents. At present, the seaweed bio-organic fertilizer is widely prepared by utilizing compound enzyme to carry out enzymolysis on kelp. However, the technical bottleneck for restricting the development of the seaweed fertilizer is that the enzyme for directly degrading the alginic acid metal salt with more than two valences in brown seaweed into the water-soluble brown seaweed oligosaccharide with small molecules is lacking, and the effects of common cellulase, protease and the like are not ideal, so that a microbial fermentation method for producing alginic acid lyase by using marine microorganisms is generated. The alginic acid lyase has wide sources and can be separated from marine microorganisms. However, the problems of overlong fermentation production time, poor controllability of the fermentation process, undefined fermentation end point and the like still exist at present, the quality of the product is good and uneven, the requirement of large-scale production can not be met, and high-quality enzyme-producing microorganism strains and optimization of the production process flow are urgently needed.
At present, the soil in China has complex and various problems, and the outside air temperature is very variable. This results in that in the planting fertilisation, it is often difficult to achieve the intended purpose using only a single fertiliser or microbial fertilizer. The soil is cultivated in successive years, so that the nutrition loss is serious, a large amount of fertilizers harden the soil, and the microbial fertilizer cannot adapt to severe soil environment to cause massive death because of insufficient organic matters after entering the soil, so that the microbial agent cannot exert the effect even if a small amount of fertilizers survive because of being incapable of forming population effect.
Disclosure of Invention
The invention aims at the problems and provides a water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid and a production method thereof; the bacillus californica is bacillus californica LYHD-001, the preservation number of the strain is CCTCC NO: M2021209, the preservation date is 2021, 3 and 8 days, the preservation address is China, wuhan university and Wuhan university, and the classification name is Bacillus cabrialesii LYHD-001. Firstly, the bacillus californicus separated from the marine sediment is utilized to thoroughly degrade brown algae such as kelp and phyllophyllum nodosum, so that nutritional ingredients in the seaweed are fully released, and polyglutamic acid is used for assisting in adjusting and improving soil environment, so that water locking, fertilizer conservation and fixation of bacillus californicus of soil are facilitated, and stress resistance and yield of planted crops are improved; the invention further provides a method for preparing the bacillus californicus agent and the water-soluble seaweed fertilizer containing the bacillus californicus agent and polyglutamic acid by utilizing the bacillus californicus, and the method is optimized, short in fermentation time, stable and controllable in process flow, small in product molecular weight, easy to absorb, obvious in fertilizer efficiency and suitable for large-scale production. The technical scheme of the invention is as follows:
a water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid comprises bacillus caldarius LYHD-001 microbial agent; the preservation number of the bacillus californicus LYHD-001 strain is CCTCC NO: M2021209, the preservation date is 2021, 3 and 8 days, the preservation address is China, the university of Wuhan and the university of Wuhan, and the classification name is Bacillus cabrialesii LYHD-001.
Furthermore, the water-soluble seaweed fertilizer also comprises polyglutamic acid organic matters.
The invention also discloses a preparation method of the water-soluble seaweed fertilizer, which comprises the following steps:
uniformly mixing 20-30 parts of polyglutamic acid, 20-30 parts of bacillus californicus LYHD-001 seaweed fermentation broth, 10-20 parts of amino acid, 5-10 parts of trace elements and 5-10 parts of fulvic acid;
the mass ratio of the microelements is 2:2:1, a mixture of ferrous sulfate, zinc sulfate and boric acid.
Further, the preparation method of the bacillus californicus LYHD-001 microbial agent comprises the following steps:
(1) Preparing seaweed mix: uniformly mixing the dried kelp and the pickled algae according to the mass ratio of 8:2, cleaning with tap water, and cutting into uniform seaweed mixture small pieces; preferably, the area of the small piece of sheet is 3×3mm;
(2) Preparing a product A: loading seaweed mixture small slices and water into a fermentation tank according to a mass ratio of 1-10:100, adding fermentation nutrients at the same time, and sterilizing at 100-150 ℃ for 15-35 min to obtain a product A;
the fermentation nutrient substances are prepared according to the following mass ratio: 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 15 parts of sodium chloride, 0.3 part of magnesium sulfate, 0.4 part of monopotassium phosphate, 0.01 part of ferrous sulfate, 0.1 part of yeast powder, 1 part of tween 80 and 1000 parts of distilled water, wherein the sterilization temperature is 121 ℃ and the sterilization time is 30 minutes;
(3) Preparing seed liquid: inoculating bacillus californicus LYHD-001 into a liquid culture medium for culturing for 20-30 hours; the volume ratio between the bacillus californicus LYHD-001 and the culture medium is 1-10:100;
the bacillus californicus LYHD-001 has a preservation number of CCTCCNO: M2021209, a preservation date of 2021, 3 months and 8 days, a preservation address of China, wuhan university and Wuhan university, and a classification name of Bacillus cabrialesii LYHD-001;
the pH value of the liquid culture medium ranges from 7.0 to 7.2, and the liquid culture medium is prepared according to the following mass ratio:
0.1 part of sodium alginate, 5 parts of ammonium sulfate, 10 parts of sodium chloride, 200 parts of seaweed mixture, 0.1 part of yeast extract, 50 parts of Vickers salt solution and 1000 parts of distilled water;
(4) Preparing a product B, namely adding seed liquid into the product A to obtain the product B, wherein the mass ratio of the seed liquid to the product A is 1-10:100;
(5) Preparing a product C, namely fermenting the product B for 2-6 days under the conditions of ventilation and stirring at the temperature of 20-40 ℃ and the pH of 5-9 to obtain the product C;
(6) Preparing a product D, namely adjusting the pH value of the product C to 5.0-9.0, heating to 30-40 ℃, adding alginic acid with the mass fraction of 1% -2%, and preserving for 3-72 h to obtain the product D;
(7) Preparing a product E, namely adjusting the temperature of the product D to 30-50 ℃, adjusting the pH value to 4-9, and filtering to obtain the product E;
(8) The bacillus californicus LYHD-001 microbial agent is prepared by adjusting the temperature of the product E to 30-50 ℃, adding polyglutamic acid, amino acid, fulvic acid and microelements, uniformly mixing, and adjusting the pH value to 4-9.
Preferably, in the step (2), the mass ratio of the seaweed small pieces to the water is 4:100.
Preferably, when the seed solution is prepared in the step (3), the bacillus calmette guerin LYHD-001 is inoculated into a liquid culture medium and cultured for 30 hours to obtain the seed solution, wherein the volume ratio of the bacillus calmette guerin to the culture medium is 8:100.
Preferably, the mass ratio of the seed liquid to the product A in the step (4) is 8:100.
Preferably, in the step (5), the temperature is 33 ℃, the pH is 7.0-7.2, and the fermentation time is 48 hours; in the step (6), the pH is 7.0, the temperature is 40 ℃, and the mixture is preserved for 48 hours after 1% alginic acid is added; the temperature in the step (7) is 35 ℃, the pH is 6.5, the temperature in the step (8) is 40 ℃, and the pH is 6.5.
The invention also comprises application of the water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid in soil regulation.
Compared with the prior art, the invention has the following advantages:
1. the bacillus californicus LYHD-001 obtained from the marine sediment by the invention is used for fermenting kelp and phyllophyllum nodosum, the fermentation process flow is stable and controllable, the fertilizer efficiency is obvious, and the method is suitable for large-scale production.
2. The fermentation enzymolysis product of bacillus calmette guerin obtained by the invention is filtered, the soluble nutrient substances are reserved, the carrier is a fully soluble organic matter, the obtained product is a fully water-soluble fertilizer, the plant is easy to absorb and utilize, meanwhile, the fermentation enzymolysis condition is mild, the activity of various plant hormones and other nutrient substances in seaweed is not damaged, and the bacillus calmette guerin can be used as an important signal molecule, so that the nutrient accumulation of crops can be promoted, and the quality and the yield of agricultural products can be effectively improved.
3. The bacillus californicus obtained by the invention not only has the capability of producing seaweed lyase, but also has a certain antibacterial effect, has a certain biocontrol effect on soil-borne diseases, and can improve the root system growth environment of plants and enhance the fertilizer efficiency.
Drawings
FIG. 1 is a gram stain and front panel colony pattern of a microscopic observation of LYHD-001 of the strain of the invention;
FIG. 2 is a schematic representation of a phylogenetic tree of the strain LYHD-001 constructed based on the 16SrDNA sequence.
Detailed Description
The invention will be further described with reference to specific embodiments, and advantages and features of the invention will become apparent from the description. The embodiments are merely exemplary and do not limit the scope of the invention in any way. It will be understood by those skilled in the art that various changes and substitutions of details and forms of the technical solution of the present invention may be made without departing from the spirit and scope of the present invention, but these changes and substitutions fall within the scope of the present invention.
Bacillus californicus LYHD-001 and identification
Bacillus californicus LYHD-001, the strain has a preservation number of CCTCCNO: M2021209, a preservation date of 2021, 3 months and 8 days, and a preservation address of China, university of Wuhan and Wuhan, and classified name of Bacillus cabrialesii LYHD-001.
After LYHD-001 strain grows on the common beef extract culture medium plate for 48h, the colony is irregular, light yellow, the whole colony is opaque, and the surface is wrinkled. The strain is observed to be rod-shaped by microscopy, and spores can be generated at the tail end or near the tail end, and gram positive can be generated. The growth temperature is 20-45 ℃, the PH is 6-8, the NaCl concentration is 0-20%, the optimal growth temperature of the strain is 33 ℃, and the optimal growth pH is 7.0.
Can hydrolyze esculin and gelatin, namely beta-glucosidase and protease are detected positive, and mannose and mannitol can be assimilated; carbon source substances such as trehalose, glycerol, galactose, fructose, glucose, mannose, maltose, sucrose and the like can be used; sorbitol, starch, lactose cannot be utilized. The results are shown in tables 1 and 2.
TABLE 1 physiological and biochemical characteristics of LYHD-001 strain-enzyme activity and carbon source assimilation
Figure SMS_1
Figure SMS_2
TABLE 2 physiological and biochemical characteristics of LYHD-001 strain-acid production Using carbon source
Figure SMS_3
Figure SMS_4
The 16SrDNA sequence of Bacillus calmette-guerin LYHD-001 is SEQ ID NO.1, and the phylogenetic tree is shown in FIG. 2. Based on its physiological and biochemical characteristics, the 16SrDNA sequence and phylogenetic tree information, the strain was identified as Bacillus californicus Bacillus cabrialesii.
Example 1A Water-soluble seaweed fertilizer containing Bacillus Carlsbergensis and polyglutamic acid and a method for producing the same
(1) Preparing seaweed mix pieces of flakes: uniformly mixing the dried kelp and the pickled algae according to the mass ratio of 8:2, cleaning with tap water, and cutting into uniform seaweed mixture small pieces with the mass ratio of 3 multiplied by 3mm;
(2) Preparing a product A: loading seaweed mixture small slices and water into a fermentation tank according to a mass ratio of 1:100, adding fermentation nutrients at the same time, and sterilizing at 121 ℃ for 20min to obtain a product A;
the fermentation nutrient substances are prepared according to the following mass ratio: 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 15 parts of sodium chloride, 0.3 part of magnesium sulfate, 0.4 part of monopotassium phosphate, 0.01 part of ferrous sulfate, 0.1 part of yeast powder, 1 part of tween 80 and 1000 parts of distilled water, wherein the sterilization temperature is 121 ℃ and the sterilization time is 30 minutes;
(3) Preparing seed liquid: inoculating bacillus californicus LYHD-001 into the liquid culture medium for culturing for 20 hours; the bacillus californicus is LYHD-001, and the preservation number is CCTCC NO: M2021209;
the pH value of the liquid culture medium ranges from 7.0 to 7.2, and the liquid culture medium is prepared according to the following mass ratio:
0.1 part of sodium alginate, 5 parts of ammonium sulfate, 10 parts of sodium chloride, 200 parts of seaweed, 0.1 part of yeast extract, 50 parts of Vickers salt solution and 1000 parts of distilled water; the volume ratio between the bacillus californicus LYHD-001 and the liquid culture medium is 10:100;
(4) Preparing a product B, namely adding seed liquid into the product A, wherein the mass ratio of the seed liquid to the product A is 2:100, and obtaining the product B;
(5) Preparing a product C, namely fermenting the product B for 4 days under the conditions of 30 ℃ of temperature, pH 7.0-7.2 and aeration stirring to obtain the product C;
(6) Preparing a product D, namely adjusting the pH value of the product C to 7.0, heating to 35 ℃, adding alginic acid with the mass fraction of 1%, and preserving for 72 hours to obtain the product D;
(7) Preparing a product E, namely adjusting the temperature of the product D to 35 ℃, adjusting the pH value to 6.5, and filtering to obtain the product E;
(8) Preparing a water-soluble seaweed fertilizer, namely regulating the temperature of a product E to 40 ℃, and mixing the product E with polyglutamic acid, amino acid, fulvic acid and microelements according to the mass ratio of 30:30:20:10:5:5, wherein the microelements are as follows: 2:1, a mixture of ferrous sulfate, zinc sulfate and boric acid; after being uniformly mixed, the pH value is adjusted to 6.5, and the water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid is obtained.
Example 2A Water-soluble seaweed fertilizer containing Bacillus Carlsbergensis and polyglutamic acid and a method for producing the same
(1) Preparing seaweed mix pieces of flakes: uniformly mixing the dried kelp and the pickled algae according to the mass ratio of 7:3, cleaning with tap water, and cutting into uniform seaweed mixture small pieces with the mass ratio of 4 multiplied by 4 mm;
(2) Preparing a product A, namely filling small seaweed mixture slices and water into a fermentation tank according to a mass ratio of 2:100, adding fermentation nutrients at the same time, and sterilizing for 30min at 115 ℃ to obtain the product A;
the fermentation nutrient is prepared from 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 15 parts of sodium chloride, 0.3 part of magnesium sulfate, 0.4 part of monopotassium phosphate, 0.01 part of ferrous sulfate, 0.1 part of yeast powder, 1 part of tween 80 and 1000 parts of distilled water according to the mass ratio, wherein the sterilization temperature is 121 ℃ and the sterilization time is 30 minutes;
(3) Preparing seed liquid by inoculating Bacillus californicus LYHD-001 into liquid culture medium, and culturing for 24 hr; the bacillus californicus is LYHD-001, and the preservation number is CCTCC NO: M2021209; the pH 7.0-7.2 of the liquid culture medium is prepared from the following mixture of 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 10 parts of sodium chloride, 200 parts of seaweed, 0.1 part of yeast extract, 50 parts of Vickers salt solution and 1000 parts of distilled water according to the mass ratio; the volume ratio between the bacillus californicus LYHD-001 and the liquid culture medium is 10:100;
(4) Preparing a product B, namely adding seed liquid into the product A, wherein the mass ratio of the seed liquid to the product A is 4:100, and obtaining the product B;
(5) Preparing a product C, namely fermenting the product B for 3 days under the conditions of 31 ℃ and pH 7.0-7.2 and aeration stirring to obtain the product C;
(6) Preparing a product D, namely adjusting the pH value of the product C to 7.0, heating to 35 ℃, adding 1% alginic acid, and preserving for 72 hours to obtain the product D;
(7) Preparing a product E, namely adjusting the temperature of the product D to 35 ℃, adjusting the pH value to 6.5, and filtering to obtain the product E;
(8) Preparing a water-soluble seaweed fertilizer, namely mixing the product E with polyglutamic acid, amino acid, fulvic acid and microelements according to the mass ratio of 25:30:20:10:10:5 after the temperature of the product E reaches 40 ℃, wherein the microelements are as follows: 2:1, uniformly mixing the mixture of ferrous sulfate, zinc sulfate and boric acid, and regulating the pH value to 6.5 to obtain the water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid.
Example 3A Water-soluble seaweed fertilizer containing Bacillus Carlsbergensis and polyglutamic acid and a method for producing the same
(1) Preparing seaweed mixture small pieces, namely uniformly mixing the dried kelp and the seaweed in a mass ratio of 8:2, washing with tap water, and then cutting into uniform seaweed mixture small pieces with the mass ratio of 3X 3mm;
(2) Preparing a product A, namely filling small seaweed mixture slices and water into a fermentation tank according to a mass ratio of 4:100, adding fermentation nutrients at the same time, and sterilizing for 30min at 121 ℃ to obtain the product A;
the fermentation nutrient is prepared from 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 15 parts of sodium chloride, 0.3 part of magnesium sulfate, 0.4 part of monopotassium phosphate, 0.01 part of ferrous sulfate, 0.1 part of yeast powder, 1 part of tween 80 and 1000 parts of distilled water according to the mass ratio, wherein the sterilization temperature is 121 ℃ and the sterilization time is 30 minutes;
(3) Preparing seed liquid by inoculating Bacillus californicus LYHD-001 into liquid culture medium, and culturing for 30 hr; the bacillus californicus is LYHD-001, and the preservation number is CCTCC NO: M2021209; the pH 7.0-7.2 of the liquid culture medium is prepared from the following mixture of 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 10 parts of sodium chloride, 200 parts of seaweed, 0.1 part of yeast extract, 50 parts of Vickers salt solution and 1000 parts of distilled water according to the mass ratio; the volume ratio between the bacillus californicus LYHD-001 and the liquid culture medium is 10:100;
(4) Preparing a product B, namely adding seed liquid into the product A, wherein the mass ratio of the seed liquid to the product A is 8:100, and obtaining the product B;
(5) Preparing a product C, namely fermenting the product B for 2 days under the conditions of air ventilation and stirring at the temperature of 33 ℃ and pH of 7.0-7.2 to obtain the product C;
(6) Preparing a product D, namely adjusting the pH value of the product C to 7.0, heating to 35 ℃, adding 1% alginic acid, and preserving for 72 hours to obtain the product D;
(7) Preparing a product E, namely adjusting the temperature of the product D to 35 ℃, adjusting the pH value to 6.5, and filtering to obtain the product E;
(8) Preparing a water-soluble seaweed fertilizer, namely mixing the product E with polyglutamic acid, amino acid, fulvic acid and microelements according to the proportion of 30:20:20:10:10:10, wherein the mass ratio of the microelements is 2:2:1, uniformly mixing the mixture of ferrous sulfate, zinc sulfate and boric acid, and regulating the pH value to 6.5 to obtain the water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid.
Example 4A Water-soluble seaweed fertilizer containing Bacillus Carlsbergensis and polyglutamic acid and a method for producing the same
(1) Preparing seaweed mixture small pieces, namely uniformly mixing the dried kelp and the seaweed in a mass ratio of 7:3, washing with tap water, and then cutting into uniform seaweed mixture small pieces with the mass ratio of 4X 4 mm;
(2) Preparing a product A, namely filling small seaweed mixture slices and water into a fermentation tank according to a mass ratio of 6:100, adding fermentation nutrients at the same time, and sterilizing at 121 ℃ for 35min to obtain the product A;
the fermentation nutrient is prepared from 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 15 parts of sodium chloride, 0.3 part of magnesium sulfate, 0.4 part of monopotassium phosphate, 0.01 part of ferrous sulfate, 0.1 part of yeast powder, 1 part of tween 80 and 1000 parts of distilled water according to the mass ratio, wherein the sterilization temperature is 121 ℃ and the sterilization time is 30 minutes;
(3) Preparing seed liquid by inoculating Bacillus californicus LYHD-001 into liquid culture medium, and culturing for 40 hr; the bacillus californicus is LYHD-001, and the preservation number is CCTCC NO: M2021209;
the pH 7.0-7.2 of the liquid culture medium is prepared from the following mixture of 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 10 parts of sodium chloride, 200 parts of seaweed, 0.1 part of yeast extract, 50 parts of Vickers salt solution and 1000 parts of distilled water according to the mass ratio. The volume ratio between the bacillus caldarius and the liquid culture medium is 10:100;
(4) Preparing a product B, namely adding seed liquid into the product A, wherein the mass ratio of the seed liquid to the product A is 10:100, and obtaining the product B;
(5) Preparing a product C, namely fermenting the product B for 2 days under the conditions of 35 ℃ of temperature, pH 7.0-7.2 and aeration stirring to obtain the product C;
(6) Preparing a product D, namely adjusting the pH value of the product C to 7.0, heating to 35 ℃, adding 1% alginic acid, and preserving for 48 hours to obtain the product D;
(7) Preparing a product E, namely adjusting the temperature of the product D to 35 ℃, adjusting the pH value to 6.5, and filtering to obtain the product E;
(8) Preparing a water-soluble seaweed fertilizer, namely mixing the product E with polyglutamic acid, amino acid, fulvic acid and microelements according to the mass ratio of 20:30:25:10:10:5 after the temperature of the product E reaches 40 ℃, wherein the microelements are as follows: 2:1, uniformly mixing the mixture of ferrous sulfate, zinc sulfate and boric acid, and regulating the pH value to 6.5 to obtain the water-soluble seaweed fertilizer containing bacillus caldarius and polyglutamic acid.
Comparative example 1:
(1) Preparing seaweed mix pieces of flakes: uniformly mixing the dried kelp and the pickled algae according to the mass ratio of 7:3, cleaning with tap water, and cutting into uniform seaweed mixture small pieces with the mass ratio of 4 multiplied by 4 mm;
(2) Preparing a product A, namely filling small seaweed mixture slices and water into a fermentation tank according to a mass ratio of 2:100, adding fermentation nutrients at the same time, and sterilizing for 30min at 115 ℃ to obtain the product A;
the fermentation nutrient is prepared from 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 15 parts of sodium chloride, 0.3 part of magnesium sulfate, 0.4 part of monopotassium phosphate, 0.01 part of ferrous sulfate, 0.1 part of yeast powder, 1 part of tween 80 and 1000 parts of distilled water according to the mass ratio, wherein the sterilization temperature is 121 ℃ and the sterilization time is 30 minutes;
(3) Preparing seed liquid by inoculating Bacillus californicus LYHD-001 into liquid culture medium, and culturing for 24 hr; the bacillus californicus is LYHD-001, and the preservation number is CCTCC NO: M2021209; the seaweed type white seaweed compound is prepared from the following mixture of 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 10 parts of sodium chloride, 200 parts of seaweed, 0.1 part of yeast extract, 50 parts of Vickers salt solution and 1000 parts of distilled water according to the mass ratio; the volume ratio between the bacillus californicus LYHD-001 and the liquid culture medium is 10:100;
(4) Preparing a product B, namely adding seed liquid into the product A, wherein the mass ratio of the seed liquid to the product A is 4:100, and obtaining the product B;
(5) Preparing a product C, namely fermenting the product B at normal temperature under the condition of ventilation and stirring for 3 days to obtain the product C;
(6) Preparing a product D, namely adding 1% of alginic acid, and preserving for 72 hours at normal temperature to obtain the product D;
(7) Preparing a product E, namely filtering the product D to obtain the product E;
(8) Comparative example 1 was prepared by mixing product E with polyglutamic acid, amino acid, fulvic acid and trace elements in a ratio of 25:30:20:10:10:5 uniformly.
Comparative example 1 the temperature and pH in the fermentation process were not adjusted on the basis of example 2, the original temperature of the environment and the pH of the material itself were maintained, other conditions and material ratios were the same as in example 2, and the procedure was the same as in example 2.
Comparative example 2
The seaweed extract, the polyglutamic acid, the amino acid powder, the fulvic acid and the trace elements are uniformly mixed in an amount of 3 parts, 3 parts and 2 parts to obtain a comparative example 2. The seaweed extract is prepared by extracting soluble organic components from brown seaweed by alkaline hydrolysis.
Test example 1: and detecting various indexes of the product according to the agricultural industry standard NY/T798-2015 of the people's republic of China and the chemical industry standard HG/T5050-2016 of the people's republic of China, wherein the detection results are shown in Table 3.
TABLE 3 quality test results for examples and comparative examples of the present invention
Figure SMS_5
The effective live bacteria in the soil are detected by the bacillus caldarius adopted by the invention.
The results show that the indexes of the microbial fertilizer produced by the invention are obviously higher than the national standard, the content of heavy metal in the fertilizer is obviously reduced through fermentation treatment, the mixed bacterial rate is obviously reduced, the effective viable count in the product can be greatly improved by adopting the fermentation technology, and the microbial fertilizer has obvious advantages in similar products.
Test example 2: the related fertilizers of the embodiment and the comparative example are applied to cucumber planting plots, the variety is Qiandeyi, the planting density is 3000 plants/mu, and the plot area is 30m 2 The dosage per mu is 10 kg/time, and the whole growth period is 5 times. The planting period is 1 year, and specific performance indexes of soil properties are shown in table 4.
TABLE 4 various performance indexes after soil improvement
Figure SMS_6
Figure SMS_7
The effective live bacteria in the soil are detected by the bacillus caldarius adopted by the invention.
The results show that the strain adopted by the invention survives and grows in the soil, the soil performance is obviously improved, the pH of the soil is 6.2-6.4, the organic matters in the soil are 14.22-14.43g/kg, and the content of the soil N, P, K is improved to a certain extent compared with the blank.
The effect of applying the products of the examples and comparative examples of the invention on cucumber crops is compared with that of not applying seaweed bacterial fertilizer products, and the fertilizer efficiency results are shown in Table 5.
Table 5 results of fertilizer efficiency comparisons
Figure SMS_8
The result shows that under the condition of equivalent fertilization amount, the product of the invention can obviously improve the yield of crops by 8.67-10.03%.
TABLE 6 cucumber quality comparison results
Figure SMS_9
The result shows that under the condition of equivalent fertilization amount, the product of the invention obviously improves the quality of crops and improves the commodity rate by 14.35-18.22%.
TABLE 7 occurrence of cucumber wilt in the growing period
Figure SMS_10
The result shows that the product of the invention can obviously reduce the incidence of crop wilt under the condition of equivalent fertilization amount.

Claims (10)

1. The water-soluble seaweed fertilizer containing bacillus californicus and polyglutamic acid is characterized by comprising bacillus californicus LYHD-001 microbial inoculum; the preservation number of the bacillus californicus LYHD-001 strain is CCTCC NO: M2021209, the preservation date is 2021, 3 and 8 days, the preservation address is China, the university of Wuhan and the university of Wuhan, and the classification name is Bacillus cabrialesii LYHD-001.
2. The water-soluble seaweed fertilizer of claim 1, further comprising polyglutamic acid organic matter.
3. The method for preparing the water-soluble seaweed fertilizer as claimed in claim 2, comprising the following steps:
uniformly mixing 20-30 parts of polyglutamic acid, 20-30 parts of bacillus californicus LYHD-001 seaweed fermentation broth, 10-20 parts of amino acid, 5-10 parts of trace elements and 5-10 parts of fulvic acid;
the mass ratio of the microelements is 2:2:1, a mixture of ferrous sulfate, zinc sulfate and boric acid.
4. The method of claim 3, wherein the method for preparing the bacillus caldarius LYHD-001 microbial agent comprises the following steps:
(1) Preparing seaweed mix: uniformly mixing the dried kelp and the pickled algae according to the mass ratio of 8:2, cleaning with tap water, and cutting into uniform seaweed mixture small pieces; preferably, the area of the small piece of sheet is 3×3mm;
(2) Preparing a product A: loading seaweed mixture small slices and water into a fermentation tank according to a mass ratio of 1-10:100, adding fermentation nutrients at the same time, and sterilizing at 100-150 ℃ for 15-35 min to obtain a product A;
the fermentation nutrient is prepared from 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 15 parts of sodium chloride, 0.3 part of magnesium sulfate, 0.4 part of monopotassium phosphate, 0.01 part of ferrous sulfate, 0.1 part of yeast powder, 1 part of tween 80 and 1000 parts of distilled water according to the mass ratio, wherein the sterilization temperature is 121 ℃ and the sterilization time is 30 minutes;
(3) Preparing seed liquid: inoculating bacillus californicus LYHD-001 into a liquid culture medium for culturing for 20-30 hours; the volume ratio between the bacillus californicus LYHD-001 and the culture medium is 1-10:100;
the pH 7.0-7.2 of the liquid culture medium is prepared from 0.1 part of sodium alginate, 5 parts of ammonium sulfate, 10 parts of sodium chloride, 200 parts of seaweed mixture, 0.1 part of yeast extract, 50 parts of Vickers salt solution and 1000 parts of distilled water according to the mass ratio;
(4) Preparing a product B, namely adding seed liquid into the product A to obtain the product B, wherein the mass ratio of the seed liquid to the product A is 1-10:100;
(5) Preparing a product C, namely fermenting the product B for 2-6 days under the conditions of ventilation and stirring at the temperature of 20-40 ℃ and the pH of 5-9 to obtain the product C;
(6) Preparing a product D, namely adjusting the pH value of the product C to 5.0-9.0, heating to 30-40 ℃, adding 1% -2% of alginic acid, and preserving for 3-72 h to obtain the product D;
(7) Preparing a product E, namely adjusting the temperature of the product D to 30-50 ℃, adjusting the pH value to 4-9, and filtering to obtain the product E;
(8) The bacillus californicus LYHD-001 microbial agent is prepared by adjusting the temperature of the product E to 30-50 ℃, adding polyglutamic acid, amino acid, fulvic acid and microelements, uniformly mixing, and adjusting the pH value to 4-9.
5. The method according to claim 5, wherein the mass ratio of the seaweed small pieces to the water in the step (2) is 4:100.
6. The method according to claim 5, wherein the seed liquid is prepared in the step (3), wherein the Bacillus calmette-guerin LYHD-001 is inoculated into a liquid medium and cultured for 30 hours to obtain the seed liquid, and the volume ratio of the Bacillus calmette-guerin to the medium is 8:100.
7. The method according to claim 5, wherein the mass ratio of the seed liquid to the product A in the step (4) is 8:100.
8. The method according to claim 5, wherein in the step (5), the temperature is 33 ℃, the pH is 7.0 to 7.2, and the fermentation time is 48 hours; in the step (6), the pH is 7.0, the temperature is 40 ℃, and the mixture is stored for 48 hours after 1% alginic acid is added.
9. The method according to claim 5, wherein the temperature in the step (7) is 35℃and the pH is 6.5, and the temperature in the step (8) is 40℃and the pH is 6.5.
10. Use of the water-soluble seaweed fertilizer as claimed in claim 1 or 2 and the water-soluble seaweed fertilizer obtained by the preparation method as claimed in any of claims 3 to 9 for soil regulation.
CN202211259111.5A 2022-10-14 2022-10-14 Water-soluble seaweed fertilizer containing bacillus californicus and polyglutamic acid as well as production method and application thereof Pending CN116041115A (en)

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CN106220416A (en) * 2016-07-20 2016-12-14 五洲丰农业科技有限公司 Containing addicted to sugar gemma bacillus agent and the production method of amino acid whose liquid Water soluble fertilizer
CN106348873A (en) * 2016-08-29 2017-01-25 五洲丰农业科技有限公司 Method for preparing microelement liquid water-soluble fertilizer through alga composite graded fermentation
CN106365802A (en) * 2016-08-29 2017-02-01 五洲丰农业科技有限公司 Production method of liquid water-soluble fertilizer containing bacillus megatherium fungicide and amino acid
CN106396907A (en) * 2016-08-31 2017-02-15 五洲丰农业科技有限公司 Production method of water-soluble fertilizer containing Bacillusmegaterium fungicide and amino acids

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106187615A (en) * 2016-07-20 2016-12-07 五洲丰农业科技有限公司 Containing addicted to sugar bacillus cereus and the production method of amino acid whose Water soluble fertilizer
CN106220416A (en) * 2016-07-20 2016-12-14 五洲丰农业科技有限公司 Containing addicted to sugar gemma bacillus agent and the production method of amino acid whose liquid Water soluble fertilizer
CN106348873A (en) * 2016-08-29 2017-01-25 五洲丰农业科技有限公司 Method for preparing microelement liquid water-soluble fertilizer through alga composite graded fermentation
CN106365802A (en) * 2016-08-29 2017-02-01 五洲丰农业科技有限公司 Production method of liquid water-soluble fertilizer containing bacillus megatherium fungicide and amino acid
CN106396907A (en) * 2016-08-31 2017-02-15 五洲丰农业科技有限公司 Production method of water-soluble fertilizer containing Bacillusmegaterium fungicide and amino acids

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