Method for improving apramycin yield based on streptomyces darkness breeding

Abstract

The invention relates to the technical field of microbial fermentation, and discloses a streptomyces darkens breeding-based apramycin yield increasing method, which comprises: S1, fermentation broth collection: carrying out streptomyces darkens fermentation, and collecting the liquid; s2, homogenizing treatment: treating the fermentation liquor by using a high-pressure homogenizer, and adding an auxiliary agent; s3, acidizing treatment: adding double acids, adjusting the pH value, and keeping the pH value for a period of time; s4, pH adjustment: adjusting the pH to neutral with ammonia water, and detecting calcium and magnesium ions; s5, resin adsorption: adding the pretreated resin twice to adsorb apramycin; and S6, settling, rinsing, purifying with ammonium chloride and hydrochloric acid, and finally analyzing apramycin. Through high-pressure homogenization treatment, streptomyces darkens cells in the fermentation liquor are effectively broken, the reaction of oxalic acid and calcium and magnesium ions is enhanced, and stable oxalate precipitates can be formed in combination with double-acid acidification, so that the removal rate of inorganic salt ions such as calcium, magnesium and the like is increased, and interference to subsequent steps is reduced.

Description

Technical Field

[0001] The invention relates to the technical field of microbial fermentation, and in particular to a method for improving the yield of apramycin based on the breeding of dark Streptomyces. Background Art

[0002] Streptomyces pyogenes is an important type of actinomycetes, widely used in the production of antibiotics. Among them, apramycin is a broad-spectrum antibiotic produced by the metabolism of Streptomyces pyogenes, mainly used to treat bacterial infections in the veterinary field. With the increasing demand for antibiotics, how to improve the production efficiency and extraction purity of apramycin has become an important challenge facing the current microbial fermentation industry.

[0003] There are some problems in the traditional apramycin production process. First, the presence of inorganic salt ions such as calcium and magnesium in the fermentation broth is a major problem. These ions will combine with apramycin to form precipitation or complexes, resulting in apramycin not existing in a free state, reducing the extraction efficiency. In addition, the presence of calcium and magnesium ions will also affect the adsorption capacity of the resin, making it impossible for the resin to fully adsorb apramycin, thereby reducing the utilization rate of the resin and causing a waste of resources.

[0004] Secondly, during the fermentation process, due to the characteristics of dark Streptomyces, mycelial balls are easily formed, which make the fermentation liquid viscous, aggravate the combination of solids and mycelium, and further limit the release of apramycin. This increased viscosity makes the subsequent separation and extraction process difficult, affecting the quality and yield of the final product.

[0005] Finally, unused sugars and other residual materials in the culture medium will interfere with the acidification and extraction process to a certain extent, reducing the resin's adsorption capacity for apramycin. This interference not only reduces the effectiveness of the extraction process, but also increases the amount of resin input in the production process, further increasing production costs. Summary of the invention

[0006] In view of the shortcomings of the prior art, the present invention provides a method for improving the yield of apramycin based on the breeding of dark Streptomyces, which solves the problem that inorganic salt ions such as calcium and magnesium in the fermentation broth will combine with apramycin to form precipitation and affect the numerical adsorption capacity.

[0007] To achieve the above objectives, the present invention is implemented by the following technical scheme: A method for improving the yield of apramycin based on the breeding of dark Streptomyces, comprising the following steps:

[0008] S1. Fermentation broth collection: performing microbial fermentation of Streptomyces pyogenes in an apramycin fermentation tank until the end of the fermentation, and collecting the fermentation broth;

[0009] S2, homogenization: the collected fermentation liquid is homogenized by a multi-stage high-pressure homogenizer with additives added, firstly using a higher pressure for coarse homogenization, and then using a lower pressure for fine homogenization;

[0010] S3, acidification treatment: adding di-acid to the homogenized fermentation liquid, adjusting the pH value and maintaining it for a period of time;

[0011] S4, pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation liquid back to neutral, and detect the calcium and magnesium ion content in the filtrate;

[0012] S5, resin adsorption: after the acidification treatment, adding the pretreated resin to the filtrate twice to adsorb apramycin;

[0013] S6. Purification and analysis: After adsorption, the resin is allowed to stand to settle, residual liquid is removed, and the resin is cleaned by step-by-step rinsing. The saturated resin is then purified using a mixture of ammonium chloride and hydrochloric acid, and the washing liquid is checked for chloride ion-free using silver nitrate solution and nitric acid solution. Finally, ammonia water is used to gradually analyze the apramycin in the resin and the analyzed liquid is collected.

[0014] Preferably, in step S2, the pressure range of the high-pressure homogenization treatment is 30MPa to 50MPa, and the treatment time is 5min to 10min; the pressure range of the low-pressure homogenization treatment is 10MPa to 50MPa, and the treatment time is 3min to 5min.

[0015] Preferably, in step S3, the concentrations of the di-acid oxalic acid and the auxiliary acid are 0.05M to 0.2M, and the adjusted di-acid concentration is 2.8M to 3.0M, and is maintained for 25min to 35min.

[0016] Preferably, in step S4, the concentration of aqueous ammonia is 1.0M to 3.0M, the pH value is adjusted back to a neutral range of 6.0M to 7.5M, the calcium and magnesium ion content of the filtrate is less than 90%, and the detection time is 10min to 15min after the pH value is adjusted back to neutral.

[0017] Preferably, in the step S5, the pre-treatment of the resin includes acid washing of the resin or heat treatment of the resin. In the two additions, 50% to 70% of the resin volume is added for the first time and stirred for 1 hour to 2 hours, and 30% to 50% of the resin volume is added for the second time and stirred for 1 hour to 2 hours. During the adsorption process, the pH value of the filtrate is 6.5M to 7.0M.

[0018] Preferably, in step S6, the step-by-step rinsing comprises the following steps:

[0019] (1) Low concentration washing: Use low concentration washing solution to wash for 10 min to 20 min to remove inorganic salts;

[0020] (2) High concentration washing: Use high concentration washing solution to wash for 5 min to 10 min to ensure the retention of apramycin.

[0021] Preferably, in step S6, the concentrations of the mixed solution of ammonium chloride and hydrochloric acid are 0.05M to 0.10M and 0.10M to 0.30M, respectively.

[0022] Preferably, in step S6, the concentration of the aqueous ammonia is 1.0-3.0 M, and the concentrations of the silver nitrate solution and the nitric acid solution are 0.08 M-0.13 M and 0.70 M-1.20 M, respectively.

[0023] The present invention provides a method for improving the yield of apramycin based on the breeding of dark Streptomyces. It has the following beneficial effects:

[0024] 1. The present invention uses high-pressure homogenization to effectively break up the dark Streptomyces cells in the fermentation broth, thereby enhancing the reaction between oxalic acid and calcium and magnesium ions. Combined with the use of double acid acidification, a stable oxalate precipitate can be formed, thereby improving the removal rate of inorganic salt ions such as calcium and magnesium, and reducing interference with subsequent steps.

[0025] 2. The homogenization and acidification steps adopted in the present invention effectively reduce the viscosity of the fermentation broth, so that the resin can more fully contact the liquid when adsorbing apramycin. The optimized resin treatment process and efficient reaction conditions significantly enhance the adsorption capacity of the resin, ensuring the efficient extraction of apramycin.

[0026] 3. The present invention effectively improves the yield of apramycin by improving the removal of calcium and magnesium ions and increasing the resin adsorption efficiency. This significant improvement not only increases the output of a single tank batch, but also reduces the waste of raw materials in the overall production process and improves production efficiency.

[0027] 4. The present invention reduces idle time and resource waste in production by optimizing the processing flow, thereby shortening the total time of the entire extraction process. This measure of improving equipment utilization, combined with the reduction in production costs, makes the scheme of the present invention have obvious advantages in terms of economy and further enhances market competitiveness. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 The figure is a flow chart of the method steps of the present invention. DETAILED DESCRIPTION

[0029] The technical solution of the present invention will be described clearly and completely below in conjunction with the accompanying drawings of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0030] Embodiment 1:

[0031] Please see attached Figure 1 The present invention provides a method for improving the yield of apramycin based on the breeding of dark Streptomyces, comprising the following steps:

[0032] S1. Fermentation broth collection: performing microbial fermentation of Streptomyces pyogenes in an apramycin fermentation tank until the end of the fermentation, and collecting the fermentation broth;

[0033] S2, homogenization: the collected fermentation liquid is homogenized by a multi-stage high-pressure homogenizer with additives added, firstly using a higher pressure for coarse homogenization, and then using a lower pressure for fine homogenization;

[0034] S3, acidification treatment: adding di-acid to the homogenized fermentation liquid, adjusting the pH value and maintaining it for a period of time;

[0035] S4, pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation liquid back to neutral, and detect the calcium and magnesium ion content in the filtrate;

[0036] S5, resin adsorption: after the acidification treatment, adding the pretreated resin to the filtrate twice to adsorb apramycin;

[0037] S6, purification and analysis: after the adsorption is completed, the resin is allowed to stand to settle, the residual liquid is removed, and the resin is cleaned by step-by-step rinsing, and then the saturated resin is purified by a mixture of ammonium chloride and hydrochloric acid, and the washing liquid is tested for the absence of chloride ions by silver nitrate solution and nitric acid solution, and finally ammonia water is used to gradually analyze the apramycin in the resin and collect the analyzed liquid;

[0038] In step S2, the pressure range of the high-pressure homogenization treatment is 30MPa-50MPa, and the treatment time is 5min-10min; the pressure range of the low-pressure homogenization treatment is 10MPa-50MPa, and the treatment time is 3min-5min;

[0039] In step S3, the concentration of the di-acid oxalic acid and the auxiliary acid is 0.05M to 0.2M, and the adjusted di-acid concentration is 2.8M to 3.0M, and the period of time is 25min to 35min;

[0040] In step S4, the concentration of ammonia water is 1.0M to 3.0M, the pH value is adjusted back to the neutral range of 6.0M to 7.5M, the calcium and magnesium ion content of the filtrate is less than 90%, and the detection time is 10min to 15min after the pH value is adjusted back to neutral;

[0041] In step S5, the pre-treated resin includes acid washing or heat treatment of the resin. In the two additions, 50% to 70% of the resin volume is added for the first time and stirred for 1h to 2h, and 30% to 50% of the resin volume is added for the second time and stirred for 1h to 2h. During the adsorption process, the pH value of the filtrate is 6.5M to 7.0M.

[0042] In step S6, the step-by-step rinsing includes the following steps:

[0043] (1) Low concentration washing: Use low concentration washing solution to wash for 10 min to 20 min to remove inorganic salts;

[0044] (2) High concentration washing: Use high concentration washing solution to wash for 5 min to 10 min to ensure the retention of apramycin;

[0045] In step S6, the concentrations of the mixed solution of ammonium chloride and hydrochloric acid are 0.05M to 0.10M and 0.10M to 0.30M respectively;

[0046] In step S6, the concentration of the ammonia solution is 1.0-3.0 M, and the concentrations of the silver nitrate solution and the nitric acid solution are 0.08 M-0.13 M and 0.70 M-1.20 M, respectively.

[0047] Summary: Example 1 combines high-pressure homogenization with di-acid acidification to ensure thorough removal of inorganic salt ions such as calcium and magnesium in the fermentation broth, maximize the adsorption capacity of the resin, and thus improve the yield and production efficiency of apramycin.

[0048] Example 2: Example without homogenization

[0049] Specific steps:

[0050] Fermentation broth collection: Perform microbial fermentation of dark Streptomyces in an apramycin fermenter until the end of the fermentation, and collect the fermentation broth;

[0051] Acidification: Add di-acid directly to the fermentation broth, adjust the pH to 2.8 and maintain for 30 minutes;

[0052] pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation broth back to neutral (6.0-7.5);

[0053] Resin adsorption: After acidification, the pretreated resin was added to the filtrate twice to adsorb apramycin;

[0054] Purification and analysis: After adsorption, washing and analysis are performed.

[0055] Summary: Compared with Example 1, Example 2 failed to completely remove inorganic salts, resulting in low resin utilization, affecting the overall extraction effect and production efficiency.

[0056] Embodiment 3: Embodiment without using auxiliary agent

[0057] Specific steps:

[0058] Fermentation broth collection: same as in Example 1;

[0059] Homogenization: A multi-stage high-pressure homogenizer is used for homogenization, but no additives are added;

[0060] Acidification: Add di-acid to the homogenized fermentation broth, adjust the pH to 2.5 and maintain for 35 minutes;

[0061] pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation broth back to neutral (6.0-7.5);

[0062] Resin adsorption: Add the pretreated resin to the filtrate twice to adsorb apramycin;

[0063] Purification and analysis: After adsorption, washing and analysis are performed.

[0064] In summary, the homogenization effect of Example 3 is not as good as that of Example 1. The extraction efficiency of apramycin and the utilization rate of resin can be more thoroughly improved by adding auxiliary agents.

[0065] Example 4: Example of a single acid only

[0066] Specific steps:

[0067] Fermentation broth collection: same as in Example 1;

[0068] Homogenization treatment: same as in Example 1;

[0069] Acidification treatment: add only oxalic acid to the homogenized fermentation broth, adjust the pH to 2.8 and maintain for 30 minutes;

[0070] pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation broth back to neutral (6.0-7.5);

[0071] Resin adsorption: Add the pretreated resin to the filtrate twice to adsorb apramycin;

[0072] Purification and analysis: After adsorption, washing and analysis are performed.

[0073] Summary: Compared with Example 1, Example 4 failed to fully utilize the synergistic effect of the bis-acids, which affected the adsorption capacity of the resin and the extraction efficiency of apramycin.

[0074] Example 5: Simplified resin adsorption process example

[0075] Specific steps:

[0076] Fermentation broth collection: same as in Example 1;

[0077] Homogenization treatment: same as in Example 1;

[0078] Acidification treatment: Add di-acid to the homogenized fermentation broth, adjust the pH to 3.0 and maintain for 30 minutes;

[0079] pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation broth back to neutral (6.0-7.5);

[0080] Resin adsorption: only one resin adsorption was performed without sufficient pretreatment;

[0081] Purification and analysis: After adsorption, washing and analysis are performed.

[0082] Summary: Example 5 fails to fully utilize the adsorption capacity of the resin, affecting the extraction efficiency, and has obvious shortcomings compared with Example 1.

[0083] Test experiment:

[0084] Experimental purpose: To compare the effects of different embodiments in the apramycin extraction process, including the removal rate of calcium and magnesium ions, the yield of apramycin and the utilization rate of resin.

[0085] Experimental Materials:

[0086] Dark Streptomyces fermentation broth, dibasic acids (oxalic acid and citric acid), ammonia water, pretreated resin;

[0087] Equipment: multi-stage high-pressure homogenizer, pH meter, centrifuge, filter;

[0088] Experimental process:

[0089] Fermentation broth collection:

[0090] Microbial fermentation of Streptomyces pyogenes was carried out in an apramycin fermenter for 72 hours, and the fermentation liquid was collected.

[0091] Experimental group settings:

[0092] Example 1: Extraction method based on high pressure homogenization.

[0093] Example 2: Extraction method without homogenization.

[0094] Example 3: Enzyme-assisted extraction method.

[0095] Example 4: Double acid enhanced extraction method.

[0096] Example 5: Simplified resin adsorption extraction method.

[0097] Specific steps:

[0098] Homogenization: Homogenization was performed according to the requirements of each embodiment (Examples 1, 3, and 4), and no homogenization was performed in Example 2;

[0099] Acidification treatment: According to the acidification requirements of each example, adjust the pH value and maintain it for a specified time;

[0100] pH adjustment: Use ammonia to adjust the pH of the acidified fermentation liquid back to neutral and record the pH value;

[0101] Resin adsorption: resin adsorption is performed according to the amount of resin input and stirring time of each embodiment;

[0102] Purification and analysis: After adsorption, washing and analysis are performed, and the analysis solution is collected.

[0103] Data Collection:

[0104] Test the removal rate of calcium and magnesium ions.

[0105] The yield of apramycin was tested.

[0106] Test resin utilization.

[0107] Test data:

[0108]

[0109]

[0110] Test result analysis:

[0111] Example 1 showed the best removal rate of calcium and magnesium ions (90%), and the yield of apramycin also reached 98%, indicating that the combination of homogenization and diacid is an efficient extraction method;

[0112] In Example 2, no homogenization was performed, resulting in the lowest removal rate of calcium and magnesium ions, only 30%, and the lowest yield of apramycin, only 75%. This shows the importance of homogenization to the extraction effect;

[0113] In Example 3, enzyme additives were added, and the removal rate of calcium and magnesium ions was 70%, and the yield of apramycin was 85%, which showed certain advantages, but there was still a gap compared with Example 1;

[0114] In Example 4, the removal rate was 85% and the yield of apramycin was 90% through double acid enhanced acidification treatment, which was good, but still failed to achieve the effect of Example 1.

[0115] The removal rate of calcium and magnesium ions in Example 5 was 50%, and the yield of apramycin was 78%, indicating that the simplified resin adsorption process failed to achieve the best extraction effect.

[0116] Summarize:

[0117] Through the above test experiments, it can be clearly seen that Example 1 is superior to other examples in all aspects, ensuring more thorough removal of inorganic salt ions, giving full play to the adsorption effect of the resin, improving the yield of apramycin, thereby increasing the tank batch output, improving equipment utilization, and reducing production costs. These data provide strong support for the technical solution of the present invention.

[0118] Comparative experiment:

[0119] Purpose:

[0120] Comparing the effects of the prior art and the present invention in the process of apramycin extraction, the following aspects are focused on:

[0121] Removal rate of calcium and magnesium ions;

[0122] Yield of apramycin;

[0123] Resin utilization;

[0124] the duration of the extraction process;

[0125] Production costs.

[0126] Technical solutions of the prior art

[0127] step:

[0128] Fermentation broth collection: Microbial fermentation of dark Streptomyces is carried out in an apramycin fermenter until the end of the fermentation, and the fermentation broth is collected.

[0129] Homogenization: No homogenization.

[0130] Acidification: Add oxalic acid alone directly to the unhomogenized fermentation broth, adjust the pH to 2.5, and maintain for 30 minutes.

[0131] pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation broth back to neutral (6.0M-7.5M).

[0132] Resin adsorption: The pretreated resin is directly added to the filtrate to adsorb apramycin.

[0133] Purification and analysis: After adsorption, washing and analysis are performed.

[0134] Technical solution of the present invention

[0135] step:

[0136] Fermentation broth collection: same as the prior art.

[0137] Homogenization: A multi-stage high-pressure homogenizer was used for homogenization, and a surfactant was added as an auxiliary agent. The processing time was 5 minutes.

[0138] Acidification: Add di-acids (oxalic acid and citric acid) to the homogenized fermentation broth, adjust the pH to 2.8 and maintain for 30 minutes.

[0139] pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation broth back to neutral (6.0M-7.5M), and detect the calcium and magnesium ion content in the filtrate.

[0140] Resin adsorption: The pretreated resin was added to the filtrate twice to adsorb apramycin.

[0141] Purification and analysis: After adsorption, let the resin stand to allow it to settle, remove the residual liquid, and perform step-by-step rinsing and purification.

[0142] Experimental process design:

[0143] Fermentation broth collection:

[0144] Microbial fermentation of Streptomyces pyogenes was carried out under the same conditions for 72 hours, and the fermentation liquid was collected.

[0145] Experimental group settings:

[0146] Group A: Existing technology solutions

[0147] Group B: Technical solution of the present invention (same as Example 1)

[0148] Specific steps:

[0149] Homogenization treatment: Group A was not homogenized, while Group B was homogenized using a high-pressure homogenizer (treatment pressure 30 MPa, time 5 minutes);

[0150] Acidification treatment: Group A used oxalic acid alone, and Group B used dual acids (oxalic acid and citric acid), maintaining the same acidification time (30 min);

[0151] pH adjustment: Use the same concentration of ammonia water (1.0M) to adjust the pH value back to 6.0M-7.5M, and detect the calcium and magnesium ion content;

[0152] Resin adsorption: Both groups used the same type of resin for adsorption, and the adsorption time and final adsorption amount were recorded;

[0153] Purification and analysis: Both groups underwent the same cleaning and analysis steps, and the analysis solutions were collected for testing.

[0154] Experimental data table:

[0155]

[0156] Test result analysis:

[0157] Calcium and magnesium ion removal rate:

[0158] The calcium and magnesium ion removal rate of group B (90%) was significantly better than that of group A (30%), indicating the effectiveness of homogenization and double acid acidification.

[0159] Apramycin yield:

[0160] The apramycin yield of group B (98%) was significantly higher than that of group A (75%), indicating that the scheme of the present invention can release and extract apramycin more effectively.

[0161] Resin utilization:

[0162] The resin utilization rate of group B (95%) is much higher than that of group A (60%), which shows the advantage of the scheme of the present invention in improving resin efficiency.

[0163] Extraction time:

[0164] The total time of the extraction process of group B was shorter (5 hours), which saved time and improved production efficiency compared with group A (6 hours).

[0165] Production cost:

[0166] The production cost of group B is 450 yuan, which is lower than 500 yuan of group A, indicating that the scheme of the present invention not only improves the extraction efficiency but also reduces the production cost.

[0167] Summarize:

[0168] Through comparative experiments, the advantages of the scheme of the present invention in various key parameters are clarified. The high-pressure homogenization treatment and double acid acidification combination of Example 1 effectively removes inorganic salts, gives full play to the adsorption capacity of the resin, significantly improves the yield of apramycin, and reduces production time and cost. These data further verify the innovation and effectiveness of the scheme of the present invention.

[0169] Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present invention, and that the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A method for improving the yield of apramycin based on the breeding of dark Streptomyces, characterized in that: The following steps are involved: S1. Fermentation broth collection: performing microbial fermentation of dark Streptomyces in an apramycin fermentation tank until the end of the fermentation, and collecting the fermentation broth; S2, homogenization: the collected fermentation liquid is homogenized by a multi-stage high-pressure homogenizer with additives added, firstly using a higher pressure for coarse homogenization, and then using a lower pressure for fine homogenization; S3, acidification treatment: adding di-acid to the homogenized fermentation liquid, adjusting the pH value and maintaining it for a period of time; S4, pH adjustment: Use ammonia water to adjust the pH value of the acidified fermentation liquid back to neutral, and detect the calcium and magnesium ion content in the filtrate; S5, resin adsorption: after the acidification treatment, adding the pretreated resin to the filtrate twice to adsorb apramycin; S6. Purification and analysis: After adsorption, the resin is allowed to stand to settle, residual liquid is removed, and the resin is cleaned by step-by-step rinsing. The saturated resin is then purified using a mixture of ammonium chloride and hydrochloric acid, and the washing liquid is checked for chloride ion-free using silver nitrate solution and nitric acid solution. Finally, ammonia water is used to gradually analyze the apramycin in the resin and the analyzed liquid is collected.

2. The method for improving the yield of apramycin based on the breeding of dark Streptomyces according to claim 1, characterized in that: In step S2, the pressure range of high-pressure homogenization treatment is 30MPa-50MPa, and the treatment time is 5min-10min. The pressure range of low-pressure homogenization treatment is 10MPa-50MPa, and the treatment time is 3min-5min. The auxiliary agents include but are not limited to polyvinyl alcohol, polyvinyl pyrrolidone, cellulase and protease.

3. The method for improving the yield of apramycin based on the breeding of dark Streptomyces according to claim 1, characterized in that: In step S3, the double acid includes oxalic acid and citric acid, and the concentration thereof is 0.05M to 0.2M. The adjusted double acid concentration is 2.8M to 3.0M, and the period of time is maintained for 25min to 35min.

4. The method for improving the yield of apramycin based on the breeding of dark Streptomyces according to claim 1, characterized in that: In the step S4, the concentration of the aqueous ammonia is 1.0M to 3.0M, the pH value is adjusted back to the neutral range of 6.0M to 7.5M, the calcium and magnesium ion content of the filtrate is less than 90%, and the detection time is 10min to 15min after the pH value is adjusted back to neutral.

5. The method for improving the yield of apramycin based on the breeding of dark Streptomyces according to claim 1, characterized in that: In the step S5, the pretreatment of the resin includes acid washing of the resin or heat treatment of the resin. In the two additions, 50% to 70% of the resin volume is added for the first time and stirred for 1 hour to 2 hours, and 30% to 50% of the resin volume is added for the second time and stirred for 1 hour to 2 hours. During the adsorption process, the pH value of the filtrate is 6.5M to 7.0M.

6. The method for improving the yield of apramycin based on the breeding of dark Streptomyces according to claim 1, characterized in that: In the step S6, the step-by-step rinsing comprises the following steps: (1) Low concentration washing: Use low concentration washing solution to wash for 10 min to 20 min to remove inorganic salts; (2) High concentration washing: Use high concentration washing solution to wash for 5 min to 10 min to ensure the retention of apramycin.

7. The method for improving the yield of apramycin based on the breeding of dark Streptomyces according to claim 1, characterized in that: In the step S6, the concentrations of the mixed solution of ammonium chloride and hydrochloric acid are 0.05M to 0.10M and 0.10M to 0.30M respectively.

8. The method for improving the yield of apramycin based on the breeding of dark Streptomyces according to claim 1, characterized in that: In the step S6, the concentration of the ammonia solution is 1.0-3.0 M, and the concentrations of the silver nitrate solution and the nitric acid solution are 0.08 M-0.13 M and 0.70 M-1.20 M, respectively.

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