Method for increasing fermentation yield of natamycin
By adding amino acids, citric acid protein and meat and bone meal during the fermentation process, the problem of low fermentation yield of natamycin is solved, which significantly increases the yield and reduces economic costs, making it suitable for industrial applications.
Patent Information
- Application Number
- CN202411798661.3
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-09
- Publication Date
- 2025-02-07
AI Technical Summary
The fermentation yield of natamycin is relatively low, which limits its industrial production and application development.
The addition of amino acids, citrate protein and meat and bone meal during the fermentation process significantly increased the production of natamycin by Streptococcus brûlée.
The production of natamycin is significantly increased by adding amino acids and citrate protein, and the fermentation time is extended by adding meat and bone meal, which increases the yield. The method is safe and economical, and is suitable for large-scale industrialization.
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Abstract
Description
Technical Field
[0001] The invention relates to a method for improving the fermentation yield of natamycin and belongs to the technical field of natamycin production. Background Art
[0002] Natamycin, also known as streptomycin, is a polyene antibiotic. It is a white to milky white, almost odorless and tasteless crystalline powder with a molecular formula of C 33 H 47 NO 13 , with a molecular weight of 665.73, contains a basic group and an acidic group in the molecule, can dissolve in both acidic and alkaline conditions, can specifically inhibit yeast and mold, and has good physical and chemical stability. As a natural harmless biological preservative, it can safely and efficiently inhibit food spoilage.
[0003] The biosynthesis of natamycin is very complex, and there are many factors that affect the fermentation yield of natamycin, including culture medium composition, process conditions, and metabolic regulation of the fermentation process. Most of the research on natamycin is based on the process conditions or metabolic regulation that affect the fermentation of natamycin. The purpose of efficiently producing natamycin is achieved by controlling external conditions and adjusting the nutritional components of the culture medium, providing a theoretical and application basis for the industrial production of such antibiotics and scientific research. However, the types of each nutrient element in the culture medium are rich, and different microorganisms have different absorption capacities for different nutrients, and the absorption and utilization capacities for the same nutrient are not necessarily the same. Natamycin still has the problem of low yield and slow development of industrial production, which limits the development of natamycin. Therefore, providing a method for improving the fermentation yield of natamycin is very important for the industrial production and industrial application of natamycin. Summary of the invention
[0004] In view of the shortcomings of the prior art, the object of the present invention is to provide a method for increasing the yield of natamycin by fermentation, by adding amino acids, citric acid dregs protein and meat and bone meal during the fermentation process, so as to significantly increase the yield of natamycin produced by Streptomyces chrysoporus.
[0005] In order to solve the above technical problems, the technical solution provided by the present invention is: A method for increasing the fermentation yield of natamycin comprises the following steps: (1) Shake flask seed culture: Under sterile conditions, spores of Streptomyces chrysanthemus are spread on an agar medium and cultured in an incubator. The cultured Streptomyces chrysanthemus is then prepared into a spore suspension, which is then pipetted into a triangular flask in the seed culture medium and placed in a constant temperature oscillator for culture to obtain a seed solution. (2) Fermentation culture in a fermenter: The seed liquid is inoculated into the fermentation medium and stirred for culture. During the culture process, glucose, amino acids, citric acid gluten solution, and meat and bone meal are added to obtain a fermentation liquid containing natamycin.
[0006] Furthermore, in step (1), the composition and content of the seed culture medium are: glucose 8-10 g / L, yeast powder 4-6 g / L, peptone 4-6 g / L, sodium chloride 4-5 g / L, oatmeal powder 0.3-0.5 g / L, and pH 7.2-7.4.
[0007] Furthermore, in step (1), the culture temperature is 30° C., the rotation speed is 250 rpm, and the culture time is 18 h.
[0008] Furthermore, in step (2), the composition and content of the fermentation medium are: glucose 20-25 g / L, yeast powder 5-7 g / L, peptone 13-17 g / L, sodium chloride 1-3 g / L, oatmeal powder 0.3-0.7 g / L, magnesium sulfate 0.5-1.5 g / L, and pH 7.2-7.4.
[0009] Furthermore, in step (2), the stirring speed of the stirring culture is 220-500 rpm, the tank pressure is 0.1 MPa, the culture temperature is 30±0.5°C, the ventilation ratio is 1-1.5 vvm, and the dissolved oxygen is controlled at 30-50%.
[0010] Furthermore, in step (2), the pH of the fermentation process is 6.0, the fermentation time is 140 h, and the fermentation temperature is 30°C.
[0011] Furthermore, in step (2), glucose is fed throughout the fermentation process, and the concentration of glucose in the fermentation medium is ≤20 g / L.
[0012] Furthermore, in step (2), the amino acids are tryptophan, arginine, isoleucine, and γ-aminobutyric acid.
[0013] Furthermore, in step (2), the amino acid addition rate is 4 mL / h, and the addition time is from 40 h to 80 h of fermentation.
[0014] Furthermore, in step (2), the mass fraction ratio of the citric acid gluten, tryptophan, arginine, isoleucine and γ-aminobutyric acid is (50-80): (8-16): (5-15): (1-3): (10-40).
[0015] Furthermore, in step (2), the flow rate of the citric acid grain protein is 4 mL / h, and the flow addition time is from 40 h to 80 h of fermentation.
[0016] Furthermore, in step (2), after 90 hours of fermentation, 200 mL of a meat and bone meal solution with a concentration of 0.2 g / L was added.
[0017] Furthermore, in step (2), the concentration of natamycin in the fermentation broth is 17.62-20.21 g / L.
[0018] Compared with the prior art, the present invention has the following beneficial effects: (1) The present invention significantly increases the yield of natamycin produced by Streptomyces chrysoporus by adding amino acids and citric acid dregs protein to the fermentation medium.
[0019] (2) The present invention increases the content of dissolved phosphorus in the culture medium by adding meat and bone meal during the fermentation process. High content of dissolved phosphorus and amino nitrogen helps to slow down the autolysis of mycelium, prolong the fermentation time, and increase the fermentation yield.
[0020] (3) The added substances of the present invention are safe and cheap, have no adverse effect on the subsequent use of natamycin, reduce economic costs, increase industrial benefits, and the method is simple to operate, which is conducive to large-scale industrialization. BRIEF DESCRIPTION OF THE DRAWINGS
[0021] Figure 1 This is a morphological picture of the bacteria in the fermentation liquid of Example 1 under a microscope; Figure 2 This is a morphological picture of the bacteria in the fermentation liquid of Comparative Example 1 under a microscope; Figure 3 This is a morphological picture of the bacteria in the fermentation liquid of Comparative Example 2 under a microscope. DETAILED DESCRIPTION
[0022] Example 1 A method for increasing the fermentation yield of natamycin comprises the following steps: (1) Shake flask seed culture: under sterile conditions, spores of Streptomyces chrysanthemi are spread on an agar medium and cultured in an incubator. The cultured Streptomyces chrysanthemi are then prepared into a spore suspension, which is then pipetted into a triangular flask in the seed culture medium and cultured in a constant temperature oscillator at 30°C and 250 rpm for 18 hours to obtain a seed solution. The seed culture medium contains 8-10 g / L glucose, 4-6 g / L yeast powder, 4-6 g / L peptone, 4-5 g / L sodium chloride, 0.3-0.5 g / L oatmeal powder, and a pH of 7.2-7.4. (2) Fermentation culture in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum of 10%, with a liquid volume of 8L, and stirred at a stirring speed of 450rpm and a culture temperature of 30°C. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 8:50:15:3:40, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. After 90h of fermentation, 200mL of 0.2g / L meat and bone meal was added, and the fermentation time was 140h. The fermentation was terminated after 140h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 19.35g / L.
[0023] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0024] Example 2 A method for increasing the fermentation yield of natamycin comprises the following steps: Step (1) is the same as step (1) of Example 1.
[0025] (2) Fermentation in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum of 10%, with a liquid volume of 8L, and stirred at a stirring speed of 450rpm and a culture temperature of 30°C. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 12:60:5:1:40, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. After 90h of fermentation, 200mL of 0.2g / L meat and bone meal was added, and the fermentation time was 140h. The fermentation was terminated after 140h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 17.62g / L.
[0026] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0027] Example 3 A method for increasing the fermentation yield of natamycin comprises the following steps: Step (1) is the same as step (1) of Example 1.
[0028] (2) Fermentation culture in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum size of 10%, and the liquid volume was 8L. The culture was stirred at a stirring speed of 450rpm and a culture temperature of 30℃. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 8:40:5:1:5, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. After 90h of fermentation, 200mL of 0.2g / L meat and bone meal was added, and the fermentation time was 140h. The fermentation was terminated after 140h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 18.21g / L.
[0029] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0030] Example 4 A method for increasing the fermentation yield of natamycin comprises the following steps: Step (1) is the same as step (1) of Example 1.
[0031] (2) Fermentation culture in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum size of 10%, and the liquid volume was 8L. The culture was stirred at a stirring speed of 450rpm and a culture temperature of 30℃. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 8:50:5:3:20, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. After 90h of fermentation, 200mL of 0.2g / L meat and bone meal was added, and the fermentation time was 140h. The fermentation was terminated after 140h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 19.27g / L.
[0032] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0033] Example 5 A method for increasing the fermentation yield of natamycin comprises the following steps: Step (1) is the same as step (1) of Example 1.
[0034] (2) Fermentation culture in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum size of 10%, and the liquid volume was 8L. The culture was stirred at a stirring speed of 450rpm and a culture temperature of 30°C. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 12:80:15:1:20, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. After 90h of fermentation, 200mL of 0.2g / L meat and bone meal was added, and the fermentation time was 140h. The fermentation was terminated after 140h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 20.21g / L.
[0035] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0036] Example 6 A method for increasing the fermentation yield of natamycin comprises the following steps: Step (1) is the same as step (1) of Example 1.
[0037] (2) Fermentation culture in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum size of 10%, and the liquid volume was 8L. The culture was stirred at a stirring speed of 450rpm and a culture temperature of 30℃. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 16:60:5:2:40, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. After 90h of fermentation, 200mL of 0.2g / L meat and bone meal was added, and the fermentation time was 140h. The fermentation was terminated after 140h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 18.60g / L.
[0038] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0039] Example 7 A method for increasing the fermentation yield of natamycin comprises the following steps: Step (1) is the same as step (1) of Example 1.
[0040] (2) Secondary seed culture: Inoculate the cultured seed solution at a 10% inoculation rate into a 50L fermentation tank, with a liquid volume of 20L, a culture temperature of 30°C, and ferment for 10 hours to obtain the secondary seed solution; (3) Fermentation in a 500L fermenter: The cultured secondary seed liquid was inoculated into the fermentation medium at an inoculum size of 10%, with a liquid volume of 400L, and stirred at a stirring speed of 220rpm and a culture temperature of 30°C. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 12:80:15:1:20, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. After 90h of fermentation, 8L of meat and bone meal at 0.2g / L was added during the culture process, and the fermentation time was 140h. The fermentation was terminated after 140h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 22.83g / L.
[0041] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0042] Comparative Example 1 Step (1) is the same as step (1) of Example 1.
[0043] (2) Fermentation in a 10L fermenter: Inoculate the cultured seed liquid into the fermentation medium at a rate of 10%, fill the medium with 8L of liquid, and perform stirring culture at a stirring speed of 450rpm and a culture temperature of 30°C. Glucose is added throughout the fermentation process, and the glucose concentration in the fermentation liquid does not exceed 20g / L. The fermentation is terminated after 110h of cultivation, and the natamycin content in the fermentation liquid is determined by HPLC to reach 15.39g / L; The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0044] Comparative Example 2 Step (1) is the same as step (1) of Example 1.
[0045] (2) Fermentation culture in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum rate of 10%, and the liquid volume was 8L. The culture was stirred at a stirring speed of 450rpm and a culture temperature of 30°C. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation liquid did not exceed 20g / L. After 90h of fermentation, 200mL of 0.2g / L meat and bone meal was added. The fermentation was terminated after 140h of culture to obtain a fermentation liquid containing natamycin. The natamycin content in the fermentation liquid was determined by HPLC to be 17.18g / L.
[0046] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0047] Comparative Example 3 Step (1) is the same as step (1) of Example 1.
[0048] (2) Fermentation culture in a 10L fermenter: The cultured seed liquid was inoculated into the fermentation medium at an inoculum size of 10%, with a liquid volume of 8L, and stirred at a stirring speed of 450rpm and a culture temperature of 30°C. After 40h of fermentation, tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid solution were added, wherein the mass ratio of tryptophan, citric acid protein, arginine, isoleucine, and γ-aminobutyric acid was 8:50:15:3:40, and the flow rate was 4mL / h. The addition was stopped after 80h of fermentation. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation broth did not exceed 20g / L. The fermentation was terminated after 110h of culture to obtain a fermentation broth containing natamycin. The natamycin content in the fermentation broth was determined by HPLC to be 17.56g / L.
[0049] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0050] Comparative Example 4 Steps (1)-(2) are the same as steps (1)-(2) of Example 7.
[0051] (3) Fermentation in a 500L fermenter: The cultured secondary seed liquid was inoculated into the fermentation medium at a rate of 10%, with a liquid volume of 400L, and stirred at a speed of 220rpm and a culture temperature of 30°C. Glucose was added throughout the fermentation process, and the glucose concentration in the fermentation liquid did not exceed 20g / L. The fermentation time was 140h. The fermentation was terminated after 140h of cultivation to obtain a fermentation liquid containing natamycin. The natamycin content in the fermentation liquid was determined by HPLC to be 16.05g / L.
[0052] The composition and content of the fermentation medium are: 20 g / L glucose, 6 g / L yeast powder, 15 g / L peptone, 2 g / L sodium chloride, 0.5 g / L oatmeal powder, 1 g / L magnesium sulfate, pH 7.4.
[0053] Experimental Example 1 The natamycin content and fermentation time in the fermentation broth of Examples 1-6 and Comparative Examples 1-2 are listed in Table 1.
[0054] Table 1. Natamycin in fermentation broth and fermentation time As can be seen from Table 1, the present invention produces natamycin using Streptomyces chrysoporus by adding tryptophan, citric acid gluten, arginine, isoleucine and γ-aminobutyric acid during the fermentation process and adding meat and bone meal in the later stage of fermentation. The yield is higher than 17.6 g / L and the fermentation time is 140 h. It can be seen that the yield of natamycin obtained by the method of the present invention is high and the fermentation time is long.
[0055] Experimental Example 2 The natamycin content and fermentation time in the fermentation broth of Example 1 and Comparative Examples 1-3 are listed in Table 2.
[0056] Table 2. Natamycin in fermentation broth and fermentation time As shown in Table 2, in Comparative Example 1, amino acids, citric acid dregs and meat and bone meal were not added, the fermentation time was 110 hours, and the fermentation yield was 15.39 g / L. In Comparative Example 2, meat and bone meal was added, but amino acids and citric acid dregs were not added, the fermentation time was 140 hours, and the fermentation yield was 17.18 g / L. It can be seen that the addition of meat and bone meal can prolong the fermentation time and increase the fermentation yield of natamycin. In Comparative Example 3, amino acids and citric acid dregs were added, but meat and bone meal was not added, the fermentation time was 110 hours, and the fermentation yield was 17.56 g / L. It can be seen that the addition of amino acids can increase the fermentation yield of natamycin. Therefore, the present invention effectively increases the yield of natamycin produced by Streptomyces chrysospora by adding tryptophan, citric acid dregs, arginine, isoleucine and γ-aminobutyric acid solution during the fermentation process and adding meat and bone meal in the late fermentation period, and prolongs the fermentation time.
[0057] Experimental Example 3 The natamycin content and fermentation time in the fermentation broth of Example 7 and Comparative Example 4 are listed in Table 3.
[0058] Table 3. Natamycin in fermentation broth and fermentation time As shown in Table 3, the content of natamycin obtained in Example 7 is 22.83 g / L, and the fermentation time is 140 h. The content of natamycin obtained in Comparative Example 4 is 16.05 g / L, and the fermentation time is also 140 h. It can be seen that under the same fermentation time, after expanded culture, the method of the present invention still significantly improves the yield of natamycin produced by Streptomyces chrysospora.
[0059] Experimental Example 4 Take the fermentation broth of Example 1 and Comparative Examples 1-2 and observe the bacterial morphology under a microscope. The observation results are shown in the attached Figure 1-3 .
[0060] From the attached Figure 1-3 It can be seen that after adding meat and bone meal in Example 1, the bacteria in the fermentation broth maintained a good mycelial state, and a small amount of new mycelium was produced. In Comparative Example 1, no meat and bone meal was added, and the bacteria in the fermentation broth produced mycelial autolysis. In Comparative Example 2, meat and bone meal was added, and the mycelial state in the fermentation broth was better. It can be seen that the method of the present invention helps to slow down the autolysis of mycelium, prolong the fermentation time, and increase the fermentation yield by adding meat and bone meal.
Claims
1. A method for increasing the fermentation yield of natamycin, characterized in that: The steps include: (1) Shake flask seed culture: Under sterile conditions, spores of Streptomyces chrysanthemus are spread on an agar medium and cultured in an incubator. The cultured Streptomyces chrysanthemus is then prepared into a spore suspension, which is then pipetted into a triangular flask in the seed culture medium and placed in a constant temperature oscillator for culture to obtain a seed solution. (2) Fermentation culture in a fermenter: The seed liquid is inoculated into the fermentation medium and stirred for culture. During the culture process, glucose, amino acids, citric acid gluten solution, and meat and bone meal are added to obtain a fermentation liquid containing natamycin.
2. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), the stirring speed of the stirring culture is 220-500 rpm, the tank pressure is 0.1 MPa, the culture temperature is 30±0.5°C, the ventilation ratio is 1-1.5 vvm, and the dissolved oxygen is controlled at 30-50%.
3. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), the pH of the fermentation process is 6.0, and the fermentation time is 140 hours.
4. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), glucose is added throughout the fermentation process, and the concentration of glucose in the fermentation medium is ≤20 g / L.
5. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), the amino acids are tryptophan, arginine, isoleucine, and γ-aminobutyric acid.
6. The method for increasing the fermentation yield of natamycin according to claim 5, characterized in that: In step (2), the amino acid addition rate is 4 mL / h, and the addition time is from 40 h to 80 h of fermentation.
7. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), the mass fraction ratio of the citric acid gluten, tryptophan, arginine, isoleucine and γ-aminobutyric acid is (50-80): (8-16): (5-15): (1-3): (10-40).
8. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), the flow rate of the citric acid grain protein is 4 mL / h, and the flow addition time is from 40 h to 80 h of fermentation.
9. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), after 90 hours of fermentation, 200 mL of a 0.2 g / L meat and bone meal solution was added.
10. The method for increasing the fermentation yield of natamycin according to claim 1, characterized in that: In step (2), the concentration of natamycin in the fermentation broth is 17.62-20.21 g / L.