CN111285719A - Method for preparing fish peptide fertilizer based on secondary fermentation technology - Google Patents
Method for preparing fish peptide fertilizer based on secondary fermentation technology Download PDFInfo
- Publication number
- CN111285719A CN111285719A CN202010073077.7A CN202010073077A CN111285719A CN 111285719 A CN111285719 A CN 111285719A CN 202010073077 A CN202010073077 A CN 202010073077A CN 111285719 A CN111285719 A CN 111285719A
- Authority
- CN
- China
- Prior art keywords
- fermentation
- fish peptide
- fish
- peptide fertilizer
- microbial inoculum
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C05—FERTILISERS; MANUFACTURE THEREOF
- C05B—PHOSPHATIC FERTILISERS
- C05B7/00—Fertilisers based essentially on alkali or ammonium orthophosphates
-
- 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
-
- 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
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Pest Control & Pesticides (AREA)
- Fertilizers (AREA)
Abstract
The invention relates to the technical field of microbial fermentation, and discloses a method for preparing a fish peptide fertilizer based on a secondary fermentation technology, which comprises the following steps: 1) pretreating raw materials; 2) primary fermentation: adding No. 1 microbial inoculum for primary fermentation; the No. 1 microbial inoculum is a mixed microbial inoculum of saccharomycetes and lactic acid bacteria; 3) and (3) secondary fermentation: adding No. 2 microbial inoculum for secondary fermentation; pichia pastoris strain producing acid protease with microbial agent No. 2; 4) and (5) post-treatment. Compared with the traditional fermentation process, the method adopts a synergistic fermentation technology, so that the raw material macromolecular protein is orderly converted, the conversion path is controlled, the putrefaction is avoided, and the tolerance is strong. On the product index, the amino acid content is more than 10 percent, almost no peculiar smell exists, and the acceptability is good.
Description
Technical Field
The invention relates to the technical field of microbial fermentation, in particular to a method for preparing a fish peptide fertilizer based on a secondary fermentation technology.
Background
In the process of processing aquatic products, the available part only accounts for 50-70%, about 30% of the available part becomes byproducts, and the byproducts comprise fish heads, fish steaks (spines), minced meat, viscera, fish tails, fish skins and the like, and are rich in nutrients such as protein, fat, amino acid and the like. Among leftover waste proteins, fish solubles are a common waste. Wherein the fish dissolving pulp is a liquid formed by a large amount of water-soluble substances after being steamed, boiled and squeezed in the process of preparing the fish meal. Researches show that the fish soluble paste contains a large amount of beneficial components such as small molecular polypeptide, amino acid, taurine, mineral substances and the like, and retains the specific unknown growth factors of the fish meal. If the waste protein can be used for high-valued resource utilization to prepare products, on one hand, the problems of waste protein disposal and environment are solved, on the other hand, the value of the products is improved through high-valued utilization, and the income is increased.
The fish peptide amino acid liquid fertilizer is a novel fertilizer which aims at the comprehensive utilization of marine biological resources after being used as a seaweed fertilizer, is also a functional fertilizer, and can be used as a base fertilizer, an additional fertilizer, a root irrigation fertilizer, a dropper fertilizer and a foliar spray fertilizer. The fish peptide fertilizer is rich in organic matters, amino acids, polypeptides and the like, and after the fish peptide fertilizer is applied to soil, the content of the organic matters in the soil can be greatly increased, the propagation of soil microorganisms can be rapidly promoted, and the nutrients of the soil are activated to a great extent. The fish peptide biological fertilizer can promote the development of crop roots, improve photosynthesis, promote plant growth, be beneficial to flower bud differentiation and mature in advance, greatly improve the quality, be sprayed on fruit trees, ensure that leaves are emerald green and bright, reduce diseases, reduce flower and fruit dropping, ensure good fruit types, have high sweetness, ensure good quality of melons and vegetables, have high yield, rich nutrition, good taste and bright color, and is a pollution-free environment-friendly green nutritional fertilizer. In addition, the pesticide can play a good role in inhibiting certain plant diseases and insect pests.
In the nineties of the century, domestic enterprises begin to research and develop fish protein fertilizers, but the fish protein fertilizers are in the starting stage, and the industry faces a plurality of development bottlenecks such as insufficient understanding of farmers, different grades, different product quality and the like. In summary, the main problems of the current preparation of fish protein fertilizers are: (1) the research of the fermentation process is seriously lagged, (2) the high-efficiency fermentation strains are extremely deficient.
Disclosure of Invention
In order to solve the technical problems, the invention provides a method for preparing a fish peptide fertilizer based on a secondary fermentation technology. On the product index, the amino acid content is more than 10 percent, almost no peculiar smell exists, and the acceptability is good.
The specific technical scheme of the invention is as follows: a method for preparing a fish peptide fertilizer based on a secondary fermentation technology comprises the following steps: 1) Pretreatment of raw materials: collecting fish peptide fertilizer raw materials, smashing the homogenate, adding an additive, uniformly mixing, and heating.
2) Primary fermentation: adding No. 1 microbial inoculum with the weight percent of 0.2-0.4 of the material for primary fermentation; the fermentation conditions are 32-35 ℃, the fermentation time is 2-4 h: the No. 1 microbial inoculum is a mixed microbial inoculum of yeast and lactobacillus.
3) And (3) secondary fermentation: uniformly stirring after primary fermentation, adding a No. 2 microbial inoculum with the weight percent of 0.1-0.3 of the material for secondary fermentation, wherein the fermentation condition is 32-35 ℃, and the fermentation time is 40-50 h; the Pichia pastoris strain of the No. 2 microbial inoculum acid-producing protease.
4) And (3) post-treatment: after secondary fermentation, uniformly stirring, and sequentially heating, filtering and canning to obtain a finished product.
Compared with the traditional fermentation process, the method adopts a synergistic fermentation technology, so that the raw material macromolecular protein is orderly converted, the conversion path is controlled, the putrefaction is avoided, and the tolerance is strong. On the product index, the amino acid content is more than 10 percent, almost no peculiar smell exists, and the acceptability is good.
Specifically, the No. 1 microbial inoculum and the No. 2 microbial inoculum are respectively adopted in the secondary fermentation process, and the two microbial inocula can be cooperatively matched with each other in tandem to improve the fermentation efficiency. The No. 1 microbial inoculum is a mixed microbial inoculum of saccharomycetes and lactic acid bacteria, a proper acidic environment is created through acid production of bacterial strains in one-time fermentation, the quality of products is guaranteed not to be easily damaged, and meanwhile, active substances generated after metabolism of the bacterial strains are beneficial to plant growth. The No. 2 microbial inoculum is a Pichia pastoris strain which can generate acid protease, and the acid protease generated by the Pichia pastoris strain can hydrolyze macromolecular protein under an acidic condition to change the macromolecular protein into small molecular peptides and amino acids, so that the small molecular peptides and the amino acids are more easily absorbed by plants.
Preferably, in the step 1), the fish peptide fertilizer raw material is one or more of fish soluble pulp, fish meal, marine product leftovers and recovered protein of surimi cleaning wastewater.
The method utilizes fish soluble pulp, fish meal, marine product leftovers, surimi cleaning wastewater recovered protein and the like as production raw materials of the fish peptide fertilizer, has stable sources, fully utilizes waste resources, is more energy-saving and environment-friendly, and the obtained fish peptide fertilizer is easier to degrade and absorb and has richer active substance nutrition.
Preferably, in step 1), the admixture includes molasses and glucose.
Preferably, in the step 1), the mass percentages of the fish peptide fertilizer raw material and the additive are as follows: 40-60% of fish peptide fertilizer raw material, 8-12% of molasses and 3-7% of glucose; the pH of the mixture is 4-5.
Preferably, in the step 1), the mixture is heated to 70-80 ℃ for 0.5-1.5 h.
Preferably, in the step 2), the yeast is Angel saccharomyces cerevisiae, and the lactic acid bacteria are lactobacillus plantarum and lactobacillus acidophilus.
Preferably, in the step 2), the mass ratio of the yeast to the lactic acid bacteria is (40: 60) - (60: 40); the mass ratio of the lactobacillus plantarum to the lactobacillus acidophilus is (40: 60) - (60: 40).
After a large number of experiments in the early stage, the team of the invention discovers that when the No. 1 microbial inoculum is the combination, the combination effect of the No. 1 microbial inoculum and the No. 2 microbial inoculum is the best.
Preferably, in step 3), the pichia pastoris strain is named as ZJU-c, and has been deposited in the general microorganism center of the china microorganism culture collection management committee (address No. 3 of west lu 1 of beijing township sunward north chen) in 2018 at 10 months and 12 days, the deposit number is CGMCC 16580, and the microorganism is classified and named as pichia pastoris.
Compared with other similar strains, the pichia pastoris strain has the characteristic of high-yield acid protease; the pichia pastoris strain is applied to the preparation of fish peptide fertilizer, the enzyme activity produced during fermentation is higher, and the amino acid content in the finally obtained fermentation liquor is higher. On the basis, the effect is better when the bacillus subtilis is matched with the No. 1 microbial inoculum.
Preferably, in step 3), the fermentation broth is supplemented with nutrients to make NaH in the fermentation broth2PO4The concentration of the magnesium sulfate is 11-12g/L, the concentration of the magnesium sulfate is 1.0-1.2g/L, and the concentration of the manganese ions is 0.8-1.0 g/L; the salinity of the culture medium is 22-23, and the pH value is 3-4; the aeration rate is 3-5L/min, and the stirring speed is 300-500 r/min.
Preferably, in the step 4), the heating temperature is 70-80 ℃ and the time is 0.5-1.5 h.
Preferably, in the step 4), rhamnolipid is further added during the stirring process, and the addition amount is 0.5-1.5 wt%.
When the fish peptide fertilizer is used as a foliar fertilizer, the foliar fertilizer needs to be applied to the surfaces of plant leaves, but the surfaces of the plant leaves are usually coated with a layer of wax, so that the fertilizer cannot be effectively attached to the surfaces of the leaves, and the fertilizing effect is poor. Therefore, the rhamnolipid is specially added into the liquid fertilizer, and the rhamnolipid can effectively improve the adhesive force of the fertilizer on the surface of the plant leaves, so that the fertilizing effect is obviously improved.
Compared with the prior art, the invention has the beneficial effects that:
(1) compared with the traditional fermentation process, the method adopts a synergistic fermentation technology, so that the raw material macromolecular protein is orderly converted, the conversion path is controlled, the putrefaction is avoided, and the tolerance is strong. On the product index, the amino acid content is more than 10 percent, almost no peculiar smell exists, and the acceptability is good.
(2) The specific No. 1 microbial inoculum and the specific No. 2 microbial inoculum are selected for fermentation treatment in sequence, and the two microbial inocula can cooperate with each other, so that the fermentation efficiency is greatly improved.
(3) The method utilizes fish soluble pulp, fish meal, marine product leftovers, surimi cleaning wastewater recovered protein and the like as production raw materials of the fish peptide fertilizer, has stable sources, fully utilizes waste resources, is more energy-saving and environment-friendly, and the obtained fish peptide fertilizer is easier to degrade and absorb and has richer active substance nutrition.
Drawings
FIG. 1 is a graph showing the effect of the fish peptide fertilizer obtained in example 1 (left side) and example 5 (right side) as a foliar fertilizer applied to the surface of plant leaves.
Detailed Description
The present invention will be further described with reference to the following examples.
General examples
A method for preparing a fish peptide fertilizer based on a secondary fermentation technology comprises the following steps:
1) pretreatment of raw materials: collecting fish peptide fertilizer raw materials, crushing, homogenizing, adding additives, uniformly mixing, and heating to 70-80 ℃ for 0.5-1.5 h. The fish peptide fertilizer is prepared from one or more of fish soluble pulp, fish meal, marine product leftovers and recovered protein from minced fillet cleaning wastewater. The weight percentage of the fish peptide fertilizer raw material and the additive is as follows: 40-60% of fish peptide fertilizer raw material, 8-12% of molasses and 3-7% of glucose; the pH of the mixture is 4-5.
2) Primary fermentation: adding No. 1 microbial inoculum with the weight percent of 0.2-0.4 of the material for primary fermentation; the fermentation condition is 32-35 ℃, and the fermentation time is 2-4 h; the No. 1 microbial inoculum is a mixed microbial inoculum of yeast and lactobacillus. The yeast is Angel Saccharomyces cerevisiae, the lactobacillus is Lactobacillus plantarum and Lactobacillus acidophilus, and the mass ratio of yeast to lactobacillus is (40: 60) - (60: 40); the mass ratio of the lactobacillus plantarum to the lactobacillus acidophilus is (40: 60) - (60: 40).
3) And (3) secondary fermentation: uniformly stirring after primary fermentation, adding a No. 2 microbial inoculum with the weight percent of 0.1-0.3 of the material for secondary fermentation, wherein the fermentation condition is 32-35 ℃, and the fermentation time is 40-50 h; the Pichia pastoris ZJU-c of the microbial inoculum acidogenic protease No. 2 is preserved in the common microorganism center of China general microbiological culture Collection center in 2018, 10 months and 12 days, the preservation number is CGMCC 16580, and the microorganism classification name is Pichia pastoris.
4) And (3) post-treatment: stirring after secondary fermentation, sequentially heating to 70-80 deg.C for 0.5-1.5h, filtering, and canning to obtain the final product.
Preferably, in the step 3), the nutrient components are supplemented into the fermentation liquor, so that NaH in the fermentation liquor is added2PO4The concentration of the magnesium sulfate is 11-12g/L, the concentration of the magnesium sulfate is 1.0-1.2g/L, and the concentration of the manganese ions is 0.8-1.0 g/L; the salinity of the culture medium is 22-23, and the pH value is 3-4; the aeration rate is 3-5L/min, and the stirring speed is 300-500 r/min.
Preferably, in the step 4), rhamnolipid is further added during the stirring process, and the addition amount is 0.5-1.5 wt%.
Example 1
A method for preparing a fish peptide fertilizer based on a secondary fermentation technology comprises the following steps:
1) pretreatment of raw materials: collecting fish peptide fertilizer raw materials, smashing the homogenate, adding the additive, uniformly mixing, and heating to 75 ℃ for 1 h. The fish peptide fertilizer is prepared from fish soluble slurry. The weight percentage of the fish peptide fertilizer raw material and the additive is as follows: 50% of fish soluble pulp, 10% of molasses and 5% of glucose; the pH of the resulting mix was adjusted to 4.5.
2) Primary fermentation: adding a No. 1 microbial inoculum with 0.3wt% of the material for primary fermentation; the fermentation condition is 32-35 ℃, and the fermentation time is 3 h; the No. 1 microbial inoculum is a mixed microbial inoculum of yeast and lactobacillus. The yeast is Angel Saccharomyces cerevisiae (Angel distiller's yeast), the lactobacillus is Lactobacillus plantarum (produced by Jiangmen golden ocean Biotech Co., Ltd.) and Lactobacillus acidophilus (produced by Jiangmen golden ocean Biotech Co., Ltd.), and the mass ratio of the yeast to the lactobacillus is 50: 50; the mass ratio of the lactobacillus plantarum to the lactobacillus acidophilus is 50: 50.
3) And (3) secondary fermentation: supplementing nutrient components to the fermentation liquor to ensure that NaH is in the fermentation liquor2PO4Has a concentration of 11.55g/L and a concentration of magnesium sulfate of 1.14g/L,manganese ions 0.9 g/L; the salinity of the culture medium is 22.76, and the pH value is 3.5; the ventilation volume is 4L/min, and the mixture is stirred evenly at 400 r/min; then adding a No. 2 microbial inoculum with the weight percent of 0.2 percent of the material for secondary fermentation, wherein the fermentation condition is 32-35 ℃, and the fermentation time is 48 hours; the Pichia pastoris ZJU-c of the microbial inoculum acidogenic protease No. 2 is preserved in the common microorganism center of China general microbiological culture Collection center in 2018, 10 months and 12 days, the preservation number is CGMCC 16580, and the microorganism classification name is Pichia pastoris.
4) And (3) post-treatment: stirring uniformly after secondary fermentation, heating to 75 ℃ in sequence for 1h, filtering and canning to obtain a finished product.
The molecular weight distribution range of the protein/amino acid in the material before and after fermentation is shown as follows:
| molecular weight range | Raw material (%) | Fermentation product (%) |
| Less than 189 | 8.87 | 5.30 |
| 189～500 | 9.16 | 28.56 |
| 500～1000 | 17.17 | 49.68 |
| 1000～2000 | 37.91 | 11.27 |
| 2000～5000 | 25.51 | 5.19 |
| 5000～10000 | 1.37 | 0 |
| Greater than 10000 | 0.01 | 0 |
As can be seen from the data in the table above, the proportion of the material with the molecular weight below 1000 in the fermentation treatment of the invention is obviously increased.
Example 2
A method for preparing a fish peptide fertilizer based on a secondary fermentation technology comprises the following steps:
1) pretreatment of raw materials: collecting fish peptide fertilizer raw materials, crushing, homogenizing, adding additives, uniformly mixing, and heating to 70-80 ℃ for 0.5-1.5 h. The fish peptide fertilizer is prepared from fish meal. The weight percentage of the fish peptide fertilizer raw material and the additive is as follows: 40% of fish meal, 12% of molasses and 7% of glucose; the pH of the resulting mix was adjusted to 5.
2) Primary fermentation: adding No. 1 microbial inoculum with 0.2 wt% of the material for primary fermentation; the fermentation condition is 32-35 ℃, and the fermentation time is 4 h; the No. 1 microbial inoculum is a mixed microbial inoculum of yeast and lactobacillus. The yeast is Angel saccharomyces cerevisiae, the lactobacillus is lactobacillus plantarum and lactobacillus acidophilus, and the mass ratio of the yeast to the lactobacillus is 40: 60; the mass ratio of the lactobacillus plantarum to the lactobacillus acidophilus is 60: 40.
3) And (3) secondary fermentation: supplementing nutrient components to the fermentation liquor to ensure that NaH is in the fermentation liquor2PO4The concentration of the magnesium sulfate is 11.55g/L, the concentration of the magnesium sulfate is 1.14g/L, and the manganese ion concentration is 0.9 g/L; the salinity of the culture medium is 22.76, and the pH value is 3.5; the ventilation volume is 3L/min, and the mixture is stirred evenly at 500 r/min; then adding a No. 2 microbial inoculum with the weight percent of 0.1 percent of the material for secondary fermentation, wherein the fermentation condition is 32-35 ℃, and the fermentation time is 50 h; the Pichia pastoris ZJU-c of the microbial inoculum acidogenic protease No. 2 is preserved in the common microorganism center of China general microbiological culture Collection center in 2018, 10 months and 12 days, the preservation number is CGMCC 16580, and the microorganism classification name is Pichia pastoris.
4) And (3) post-treatment: stirring uniformly after secondary fermentation, heating to 70 ℃ for 1.5h, filtering and canning to obtain a finished product.
Example 3
A method for preparing a fish peptide fertilizer based on a secondary fermentation technology comprises the following steps:
1) pretreatment of raw materials: collecting fish peptide fertilizer raw materials, crushing, homogenizing, adding additives, uniformly mixing, and heating to 70-80 ℃ for 0.5-1.5 h. The fish peptide fertilizer raw material is marine product leftovers. The weight percentage of the fish peptide fertilizer raw material and the additive is as follows: marine product leftover 60%, molasses 8% and glucose 3%; the pH of the resulting blend was adjusted to 4.
2) Primary fermentation: adding No. 1 microbial inoculum with 0.4wt% of the material for primary fermentation; the fermentation condition is 32-35 ℃, and the fermentation time is 2 h; the No. 1 microbial inoculum is a mixed microbial inoculum of yeast and lactobacillus. The yeast is Angel saccharomyces cerevisiae, the lactobacillus is lactobacillus plantarum and lactobacillus acidophilus, and the mass ratio of the yeast to the lactobacillus is 60: 40; the mass ratio of the lactobacillus plantarum to the lactobacillus acidophilus is 40: 60.
3) And (3) secondary fermentation: supplementing nutrient components to the fermentation liquor to ensure that NaH is in the fermentation liquor2PO4The concentration of the magnesium sulfate is 11.55g/L, the concentration of the magnesium sulfate is 1.14g/L, and the manganese ion concentration is 0.9 g/L; the salinity of the culture medium is 22.76, and the pH value is 3.5; the ventilation volume is 3L/min, and the mixture is stirred evenly at 500 r/min; then adding a No. 2 microbial inoculum with the weight percent of 0.3 percent of the material for secondary fermentation, wherein the fermentation condition is 32-35 ℃, and the fermentation time is 40 h; the Pichia pastoris ZJU-c of the microbial inoculum acidogenic protease No. 2 is preserved in the common microorganism center of China general microbiological culture Collection center in 2018, 10 months and 12 days, the preservation number is CGMCC 16580, and the microorganism classification name is Pichia pastoris.
4) And (3) post-treatment: stirring uniformly after secondary fermentation, heating to 80 ℃ in sequence for 0.5h, filtering, and canning to obtain a finished product.
Comparative example 1
Comparative example 1 the same formulation, same batch of starting material as in example 1 was used. The difference is that the step of one-time fermentation is omitted, and the total fermentation time is the same.
The molecular weight distribution range of the protein/amino acid in the material before and after fermentation is shown as follows:
as can be seen from the data in the above table, the ratio of the large molecular weight in the fermentation product of comparative example 1 is significantly higher than that in example 1, indicating that the fermentation was not thorough.
Comparative example 2
Comparative example 2 the same formulation, same batch of starting material as in example 1 was used. The difference from example 1 is that different No. 2 microbial inoculum (Pichia anomala strain CGMCC24056) is adopted.
The molecular weight distribution range of the protein/amino acid in the material before and after fermentation is shown as follows:
| molecular weight range | Raw material (%) | Fermentation product (%) |
| Less than 189 | 8.87 | 3.3 |
| 189～500 | 9.16 | 17.56 |
| 500～1000 | 17.17 | 20.68 |
| 1000～2000 | 37.91 | 25.21 |
| 2000～5000 | 25.51 | 32.19 |
| 5000～10000 | 1.37 | 0.96 |
| Greater than 10000 | 0.01 | 0.10 |
As can be seen from the data in the above table, the ratio of the large molecular weight in the fermentation product of comparative example 2 is significantly higher than that in example 1, indicating that the fermentation was not thorough.
The content of amino acids (determined by the content of free amino acids in the water-soluble fertilizer NYT 1975-2010) in the fermentation products obtained in example 1 and comparative examples 1-2 is as follows:
as can be seen from the data in the above table, the amino acid content of the fermentation product obtained in example 1 is significantly better than that of comparative examples 1-2. Wherein, compared with the comparative example 1, the invention is demonstrated to have remarkable improvement on the secondary fermentation process; compared with the comparative example 2, the specific strain No. 2 of the invention has stronger acid-producing protease capability compared with other similar strains, so that the content of amino acid in a fermentation product can be obviously improved.
Example 4
In order to further verify that the strain of the invention has higher acid protease producing capability compared with the common similar strain, the following experiments are carried out:
fermentation test of 7.5L fermenter
Under the condition of 4L/min of ventilation and 400r/min, 3 percent of inoculation amount is transferred for fermentation for 24 hours, and the formula of a culture medium comprises 50g/L of waste molasses, 50g/L of soybean meal and NaH2PO411.55 g/L; magnesium sulfate 1.14 g/L; salinity of the culture medium 22.76; adjusting pH to 3-4. Finally, the enzyme activity of the crude enzyme solution reaches 9854.4 +/-4.5U/mL, the enzyme activity property of the crude enzyme solution is further researched for obtaining a better research effect of the amino acid liquid fertilizer, and the research result shows that the enzyme activity of the protease of the crude enzyme solution can reach 10542.9 +/-2.7U/mL by adding 0.9g/L manganese ions under the conditions of pH 3.5 and 45 ℃.
Under the same process conditions, the data of the strains according to the invention are compared with those of the commercially available strains in the following table:
| pichia pastoris ZJU-c | Commercial strain (CGMCC24056) | |
| Enzyme activity | 10542.9U/mL | 3453.6U/mL |
Note: the commercially available strain information is as follows: CGMCC-24056, the Latin name Pichia galeiformis, and the Chinese translation name Pichia pastoris.
Definition of enzyme activity: hydrolysis of casein by 1 mL of enzyme solution at 40 ℃ at pH 3.0 to yield 1 microgram of tyrosine per minute corresponds to the consumption of 1 protease activity unit expressed as U/mL.
The comparison of the data shows that compared with the existing similar strains sold in the market, the enzyme activity of the crude enzyme solution obtained after fermentation is improved by about 205%. The bacterial strain disclosed by the invention has obvious advantages in the effects of acid-producing protease and the effects of preparing amino acid liquid fertilizer by fermentation.
Example 5
Compared with example 1, the difference of the example 5 is only that in the step 4), rhamnolipid is also added during the stirring process, and the addition amount is 1%.
The fish peptide fertilizer obtained in example 1 (left side) and example 5 (right side) was applied as foliar fertilizer to the surface of plant leaves, and the effect is shown in fig. 1. As can be seen from FIG. 1, the fish peptide manure without rhamnolipid is in a droplet shape on the surface of the plant leaf and is very easy to run off, while the fish peptide manure with rhamnolipid in example 5 can be fully spread and wetted on the surface of the plant leaf, so that the fish peptide manure can more easily permeate into the interior of the plant leaf.
In addition, the invention also finds that the rhamnolipid added in the fish peptide fertilizer can inhibit fungal diseases, such as botrytis cinerea, rice blast, gibberellic disease, rot, fusarium and other fungal diseases, and has an obvious bacteriostatic action.
The raw materials and equipment used in the invention are common raw materials and equipment in the field if not specified; the methods used in the present invention are conventional in the art unless otherwise specified.
The above description is only a preferred embodiment of the present invention, and is not intended to limit the present invention, and all simple modifications, alterations and equivalents of the above embodiments according to the technical spirit of the present invention are still within the protection scope of the technical solution of the present invention.
Claims (10)
1. A preparation method of a fish peptide fertilizer based on a secondary fermentation technology is characterized by comprising the following steps:
1) pretreatment of raw materials: collecting fish peptide fertilizer raw materials, smashing the fish peptide fertilizer raw materials into homogenate, adding an additive, uniformly mixing, and heating;
2) primary fermentation: adding No. 1 microbial inoculum with the weight percent of 0.2-0.4 of the material for primary fermentation; the fermentation condition is 32-35 ℃, and the fermentation time is 2-4 h; the No. 1 microbial inoculum is a mixed microbial inoculum of saccharomycetes and lactic acid bacteria;
3) and (3) secondary fermentation: uniformly stirring after primary fermentation, adding a No. 2 microbial inoculum with the weight percent of 0.1-0.3 of the material for secondary fermentation, wherein the fermentation condition is 32-35 ℃, and the fermentation time is 40-50 h; the Pichia pastoris strain of the microbial agent No. 2 for producing acid protease;
4) and (3) post-treatment: after secondary fermentation, uniformly stirring, and sequentially heating, filtering and canning to obtain a finished product.
2. The method for preparing the fish peptide fertilizer as claimed in claim 1, wherein in the step 1), the fish peptide fertilizer raw material is one or more of fish pulp, fish meal, marine product leftovers and surimi cleaning water recycled protein.
3. The method for producing a fish peptide fertilizer as claimed in claim 1 or 2, wherein the external additive in step 1) comprises molasses and glucose.
4. The method for preparing the fish peptide fertilizer as claimed in claim 3, wherein in the step 1), the mass percentage of the fish peptide fertilizer raw materials and the additive is as follows: 40-60% of fish peptide fertilizer raw material, 8-12% of molasses and 3-7% of glucose; the pH of the mixture is 4-5.
5. The method for preparing the fish peptide fertilizer as claimed in claim 1, wherein in the step 1), the fish peptide fertilizer is heated to 70-80 ℃ for 0.5-1.5 h; in the step 4), the heating temperature is 70-80 ℃ and the time is 0.5-1.5 h.
6. The method for preparing fish peptide fertilizer of claim 1, wherein in the step 2), the yeast is Angel Saccharomyces cerevisiae, and the lactic acid bacteria are Lactobacillus plantarum and Lactobacillus acidophilus.
7. The method for preparing a fish peptide fertilizer as claimed in claim 6, wherein in the step 2), the mass ratio of the yeast to the lactic acid bacteria is (40: 60) - (60: 40) (ii) a The mass ratio of the lactobacillus plantarum to the lactobacillus acidophilus is (40: 60) - (60: 40).
8. The method for preparing fish peptide fertilizer as claimed in claim 1, 6 or 7, wherein in step 3), the Pichia pastoris strain named ZJU-c is deposited in China general microbiological culture Collection center (CGMCC 16580) 10, 12/2018 and classified as Pichia pastorisPichia pastoris。
9. The method for preparing fish peptide fertilizer as claimed in claim 8, wherein in step 3), the fermentation broth is supplemented with nutrients to make NaH in the fermentation broth2PO4The concentration of the magnesium sulfate is 11-12g/L, the concentration of the magnesium sulfate is 1.0-1.2g/L, and the concentration of the manganese ions is 0.8-1.0 g/L; the salinity of the culture medium is 22-23, and the pH value is 3-4; the aeration rate is 3-5L/min, and the stirring speed is 300-500 r/min.
10. The method for preparing fish peptide fertilizer as claimed in claim 1, wherein rhamnolipid is further added in the stirring process in the step 4), and the addition amount is 0.5-1.5 wt%.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010073077.7A CN111285719A (en) | 2020-01-21 | 2020-01-21 | Method for preparing fish peptide fertilizer based on secondary fermentation technology |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010073077.7A CN111285719A (en) | 2020-01-21 | 2020-01-21 | Method for preparing fish peptide fertilizer based on secondary fermentation technology |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN111285719A true CN111285719A (en) | 2020-06-16 |
Family
ID=71026663
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010073077.7A Pending CN111285719A (en) | 2020-01-21 | 2020-01-21 | Method for preparing fish peptide fertilizer based on secondary fermentation technology |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111285719A (en) |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111792972A (en) * | 2020-07-28 | 2020-10-20 | 广西壮族自治区农业科学院 | Fish peptide liquid microbial fertilizer and production method thereof |
| CN112111547A (en) * | 2020-09-29 | 2020-12-22 | 中国海洋大学 | Fish soluble pulp enzymolysis product and preparation method and application thereof |
| CN115286440A (en) * | 2022-08-23 | 2022-11-04 | 东莞市林氏生物技术股份有限公司 | Fish peptide microbial fertilizer and preparation method thereof |
| CN115812070A (en) * | 2021-06-30 | 2023-03-17 | 华川郡 | Preparation method of fermentation liquor of oncorhynchus masou |
| CN115960732A (en) * | 2023-01-20 | 2023-04-14 | 昆明理工大学 | Pichia anomala strain and microbial agent and application thereof |
| CN116239405A (en) * | 2023-03-06 | 2023-06-09 | 海南省农业科学院农业环境与土壤研究所 | Method and device for manufacturing fish protein organic fertilizer |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103755476A (en) * | 2014-01-17 | 2014-04-30 | 陕西鼎天济农腐殖酸制品有限公司 | Environment-friendly humic acid containing water-soluble fertilizer and production method thereof |
| CN107793244A (en) * | 2017-12-06 | 2018-03-13 | 曹结宾 | A kind of preparation method of organic fertilizer |
| CN108178667A (en) * | 2017-09-11 | 2018-06-19 | 农法自然（上海）农业科技有限公司 | The high nitrogen organic water-soluble fertilizer of fruit tree and its application process |
| CN109971684A (en) * | 2019-04-10 | 2019-07-05 | 天津大成前瞻生物科技研发有限公司 | A kind of lactobacillus-fermented preparation and its activation method |
| CN110655422A (en) * | 2019-11-15 | 2020-01-07 | 湖南省畜牧兽医研究所 | Composting method for promoting rotting and preserving nitrogen and application of organic fertilizer |
-
2020
- 2020-01-21 CN CN202010073077.7A patent/CN111285719A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103755476A (en) * | 2014-01-17 | 2014-04-30 | 陕西鼎天济农腐殖酸制品有限公司 | Environment-friendly humic acid containing water-soluble fertilizer and production method thereof |
| CN108178667A (en) * | 2017-09-11 | 2018-06-19 | 农法自然（上海）农业科技有限公司 | The high nitrogen organic water-soluble fertilizer of fruit tree and its application process |
| CN107793244A (en) * | 2017-12-06 | 2018-03-13 | 曹结宾 | A kind of preparation method of organic fertilizer |
| CN109971684A (en) * | 2019-04-10 | 2019-07-05 | 天津大成前瞻生物科技研发有限公司 | A kind of lactobacillus-fermented preparation and its activation method |
| CN110655422A (en) * | 2019-11-15 | 2020-01-07 | 湖南省畜牧兽医研究所 | Composting method for promoting rotting and preserving nitrogen and application of organic fertilizer |
Non-Patent Citations (2)
| Title |
|---|
| 杨城: "《一株产酸性蛋白酶菌株的筛选、鉴定及发酵条件优化》", 《农业生物技术学报》 * |
| 黄炎坤: "《现代养殖场生产设施与设备》", 30 September 2016, 郑州：中原农民出版社 * |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111792972A (en) * | 2020-07-28 | 2020-10-20 | 广西壮族自治区农业科学院 | Fish peptide liquid microbial fertilizer and production method thereof |
| CN111792972B (en) * | 2020-07-28 | 2022-01-04 | 广西壮族自治区农业科学院 | Fish peptide liquid microbial fertilizer and production method thereof |
| CN112111547A (en) * | 2020-09-29 | 2020-12-22 | 中国海洋大学 | Fish soluble pulp enzymolysis product and preparation method and application thereof |
| CN112111547B (en) * | 2020-09-29 | 2021-11-05 | 中国海洋大学 | Fish soluble pulp enzymolysis product and preparation method and application thereof |
| CN115812070A (en) * | 2021-06-30 | 2023-03-17 | 华川郡 | Preparation method of fermentation liquor of oncorhynchus masou |
| CN115286440A (en) * | 2022-08-23 | 2022-11-04 | 东莞市林氏生物技术股份有限公司 | Fish peptide microbial fertilizer and preparation method thereof |
| CN115960732A (en) * | 2023-01-20 | 2023-04-14 | 昆明理工大学 | Pichia anomala strain and microbial agent and application thereof |
| CN115960732B (en) * | 2023-01-20 | 2023-12-01 | 昆明理工大学 | Pichia glabra strain, microbial agent and application thereof |
| CN116239405A (en) * | 2023-03-06 | 2023-06-09 | 海南省农业科学院农业环境与土壤研究所 | Method and device for manufacturing fish protein organic fertilizer |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111285719A (en) | Method for preparing fish peptide fertilizer based on secondary fermentation technology | |
| CN105586279B (en) | A kind of Trichoderma spp. direct fermentation crop material prepares the method for Trichoderma spp. solid spawn and the product of preparation | |
| CN102617208B (en) | Edible fungus residue organic fertilizer raw materials and preparation method thereof | |
| CN103907897A (en) | Soybean sauce employing multi-strain starter propagation and production technology thereof | |
| CN105110868A (en) | Green environment-friendly leaf fertilizer, and preparation method thereof | |
| CN106316693A (en) | Biological humic acid fertilizer and preparing method thereof | |
| CN103098981B (en) | Dewatered solid combined inoculant for increasing protein level of citrus pulp feed and preparation method of dewatered solid combined inoculant | |
| CN106045626B (en) | Method for producing biological bacterial fertilizer by using amino acid fermentation waste liquid and corn straw | |
| CN103815119A (en) | Method for producing microbial feed by fermenting corn deep processing byproducts | |
| CN102907572A (en) | Banana silage microbial additive and preparation method thereof | |
| CN103992191A (en) | Compound biological bacterial fertilizer | |
| CN101709272A (en) | Method for preparing feed yeast | |
| CN103504123A (en) | Fermented soybean meal with function of complex enzymes and preparation method for fermented soybean meal | |
| CN101361520B (en) | Potato pulp energy fermentation feed capable of replacing bran and preparation method thereof | |
| CN100535104C (en) | Method for preparing feedstuff yeast from maize peel hydrolysis solution | |
| CN108378200A (en) | A kind of earthworm feedstuff base and preparation method thereof | |
| CN102488087A (en) | Biological detoxification method for camellia seed cakes | |
| CN101407762B (en) | Microbial solid inocula, and preparation and use thereof | |
| CN1706271A (en) | Production process of composite biochemical granulated feed | |
| CN106929437A (en) | A kind of preparation method of the yeast culture of wine yeast fermentation | |
| CN104839425A (en) | Method for producing protein feed through solid fermentation of tomato waste residues by using bacterium mixture | |
| CN102337225B (en) | Preparation method of high-nitrogen fresh yeast and extract | |
| CN111286463A (en) | Pichia pastoris strain capable of producing acid protease | |
| CN100535103C (en) | Method for preparing feedstuff yeast from maize peel hydrolysis sugar solution after extracting xylose | |
| CN111394257B (en) | Method for preparing trichoderma harzianum strain by utilizing mushroom residues and fresh cow dung through directional fermentation |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20200616 |