阅读说明：本技术 一种生产土霉素的发酵工艺 (Fermentation process for producing oxytetracycline ) 是由 王波政 李蕊红 兰明 李贵红 于 2021-07-15 设计创作，主要内容包括：本发明属于抗生素发酵技术领域,具体涉及一种生产土霉素的发酵工艺；提供了一种生产土霉素的发酵工艺,其进一步优化了发酵及其分离纯化的工艺,使得制得的产品品质有较大的提升,极大的降低了杂质的含量,提高了玉米原料的利用率,采用的技术方案为：将玉米分别粉碎,分离其中碳源和氮源及油脂,制备发酵培养基,然后酸化后,用磷酸盐缓冲液回调pH过滤,再进行脱色,脱色后的脱色液通过超声波间歇搅拌的工艺来结晶,最后清洗,干燥制得成品土霉素。(The invention belongs to the technical field of antibiotic fermentation, and particularly relates to a fermentation process for producing oxytetracycline; the fermentation process for producing the oxytetracycline is provided, and further optimizes the processes of fermentation, separation and purification, so that the quality of the prepared product is greatly improved, the content of impurities is greatly reduced, the utilization rate of corn raw materials is improved, and the adopted technical scheme is as follows: respectively crushing corns, separating a carbon source, a nitrogen source and grease, preparing a fermentation culture medium, acidifying, adjusting the pH back by using a phosphate buffer solution, filtering, decoloring, crystallizing the decolored solution after decoloring through an ultrasonic intermittent stirring process, and finally cleaning and drying to obtain the finished product oxytetracycline.)

1. A fermentation process for producing oxytetracycline is characterized by comprising the following steps:

1) fermentation of terramycin

Threshing the corn cobs, respectively crushing corn kernels and corncobs, separating germs after the corn kernels are crushed, preparing corn steep liquor from the crushed corn kernels, and preparing corn oil from the separated germs and obtaining a byproduct of corn germ meal; preparing the corncob degradation liquid from the corncobs; after the preparation of the raw materials is finished, the specific fermentation process is as follows:

selecting streptomyces rimosus as a zymocyte, and fermenting to prepare a fermentation liquor through spore culture and three-level seed fermentation; the first-stage seed fermentation conditions are as follows: controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 20-35 parts of starch, 20-35 parts of corncob degradation liquid, 5-15 parts of corn germ meal, 5-15 parts of soybean cake powder, 10-25 parts of corn steep liquor, 1.5-4 parts of calcium carbonate, 1-3.5 parts of monopotassium phosphate, 1-5 parts of corn oil, 3-9 parts of composite amino acid powder, 1-5 parts of vegetable oil, 1-5 parts of silicon-based defoaming agent and 1000 parts of sterile water together with 1500 parts of materials;

the secondary seed fermentation conditions are as follows:

controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 30-40 parts of starch, 30-40 parts of corncob degradation liquid, 10-15 parts of corn germ meal, 10-20 parts of soybean cake powder, 15-30 parts of corn steep liquor, 1.5-4 parts of calcium carbonate, 1-3.5 parts of monopotassium phosphate, 1-5 parts of corn oil, 3-9 parts of composite amino acid powder, 1-5 parts of vegetable oil, 1-5 parts of silicon-based defoaming agent and 1200 parts of sterile water 600-;

the fermentation conditions of the third-level seeds are as follows: controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 40-50 parts of starch, 40-50 parts of corncob degradation liquid, 15-25 parts of corn germ meal, 15-25 parts of soybean cake powder, 25-35 parts of corn steep liquor, 1.5-4 parts of calcium carbonate, 1-3.5 parts of monopotassium phosphate, 1-5 parts of corn oil, 3-9 parts of composite amino acid powder, 1-5 parts of vegetable oil, 1-5 parts of silicon-based defoaming agent and 800 parts of sterile water;

2) injecting the oxytetracycline fermentation liquor into an acidification tank, adding oxalic acid for acidification treatment, adjusting the pH value to 1.6-1.9, diluting the acidified oxytetracycline fermentation liquor, performing prefiltration, and performing plate-frame filtration twice to separate out hypha residues and oxytetracycline stock solution;

3) adding a buffer solution with the pH value of 5-7 into the oxytetracycline stock solution, adjusting the pH value back to 2-3.6, and filtering by using an ultrafiltration membrane;

4) adding hydrochloric acid into the oxytetracycline liquid ultrafiltered in the step 2 to adjust the pH value to 1.6-1.9, and then decoloring with decoloring resin to obtain a decoloring liquid;

5) filtering the decolorized oxytetracycline decolorized solution to remove residual resin, heating the decolorized solution to 28-32 ℃, and crystallizing;

6) adding ammonia water into the decolorized solution after temperature rise to adjust the pH value to 4.8-5.4, simultaneously carrying out ultrasonic intermittent stirring, and crystallizing for 40-60min to finish crystallization;

7) and (2) carrying out filter pressing on the crystallization liquid by using a filter press, separating to obtain oxytetracycline wet crystals, and circularly cleaning the oxytetracycline wet crystals by using pure water in the filter press, wherein the weight ratio of the pure water to the oxytetracycline wet crystals is controlled to be 5-10: 1, cleaning and drying to obtain a finished product of oxytetracycline.

2. The fermentation process for producing oxytetracycline of claim 1, wherein the buffer in step 3 is phosphate buffer.

3. The fermentation process for producing oxytetracycline of claim 1, wherein the ultrafiltration membrane is a 1-5 ten thousand molecular weight ultrafiltration membrane.

4. The fermentation process for producing oxytetracycline of claim 1, wherein the decolorizing resin has parameters of volume exchange capacity of 3eq/L and water content of 65-72%.

5. The fermentation process for producing oxytetracycline of claim 1, wherein the ultrasonic frequency is 6.5-35kHz and the intensity is 0.6-4W/cm²。

6. The fermentation process for producing oxytetracycline according to claim 1, wherein the ultrasonic intermittent stirring is ultrasonic stirring for 5-10min at 1-3min intervals.

7. The fermentation process for producing oxytetracycline according to claim 1, wherein the ultrasonic frequency is gradually increased under the control of the ultrasonic intermittent stirring, the first ultrasonic frequency is 6.5-10kHz, and the first ultrasonic frequency is gradually increased by 5-10 kHz.

8. The fermentation process for producing oxytetracycline of claim 1, wherein the washing time in step 7 is 15-25 min.

Technical Field

The invention belongs to the technical field of antibiotic fermentation, and particularly relates to a fermentation process for producing oxytetracycline.

Background

Terramycin has broad-spectrum anti-pathogenic microorganism effect, is a rapid bacteriostatic agent, and has bactericidal effect on certain bacteria at high concentration. The action mechanism is that the medicine can be specifically combined with the A position of ribosome 30S subunit, and the connection of aminoacyl-tRNA at the position is prevented, so that the growth of peptide chain is inhibited, and the protein synthesis of bacteria or other pathogenic microorganisms is influenced. The terramycin has stronger antibacterial activity on staphylococcus aureus, pneumococcus, streptococcus pyogenes, gonococcus, meningococcus, escherichia coli, aerobacter, shigella, yersinia, listeria monocytogenes and the like; in addition, terramycin also has strong effects on rickettsia, mycoplasma, chlamydia, actinomycetes and the like.

The existing oxytetracycline fermentation method adopts a mature technical method in the processes of fermentation and subsequent separation and purification, has the advantages of simple process and low cost, but greatly limits the quality of finished products and simultaneously does not fully utilize fermentation raw materials.

Disclosure of Invention

The invention overcomes the defects of the prior art, provides a fermentation process for producing oxytetracycline, and further optimizes the processes of fermentation, separation and purification, so that the quality of the prepared product is greatly improved, the content of impurities is greatly reduced, and the utilization rate of corn raw materials is improved.

In order to solve the technical problems, the invention adopts the technical scheme that: a fermentation process for producing oxytetracycline is characterized by comprising the following steps:

1) fermentation of terramycin

Threshing the corn cobs, respectively crushing corn kernels and corncobs, separating germs after the corn kernels are crushed, preparing corn steep liquor from the crushed corn kernels, and preparing corn oil from the separated germs and obtaining a byproduct of corn germ meal; preparing the corncob degradation liquid from the corncobs; after the preparation of the raw materials is finished, the specific fermentation process is as follows:

selecting streptomyces rimosus as a zymocyte, and fermenting to prepare a fermentation liquor through spore culture and three-level seed fermentation; the first-stage seed fermentation conditions are as follows: controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 20-35 parts of starch, 20-35 parts of corncob degradation liquid, 5-15 parts of corn germ meal, 5-15 parts of soybean cake powder, 10-25 parts of corn steep liquor, 1.5-4 parts of calcium carbonate, 1-3.5 parts of monopotassium phosphate, 1-5 parts of corn oil, 3-9 parts of composite amino acid powder, 1-5 parts of vegetable oil, 1-5 parts of silicon-based defoaming agent and 1000 parts of sterile water together with 1500 parts of materials;

the secondary seed fermentation conditions are as follows:

controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 30-40 parts of starch, 30-40 parts of corncob degradation liquid, 10-15 parts of corn germ meal, 10-20 parts of soybean cake powder, 15-30 parts of corn steep liquor, 1.5-4 parts of calcium carbonate, 1-3.5 parts of monopotassium phosphate, 1-5 parts of corn oil, 3-9 parts of composite amino acid powder, 1-5 parts of vegetable oil, 1-5 parts of silicon-based defoaming agent and 1200 parts of sterile water 600-;

the fermentation conditions of the third-level seeds are as follows: controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 40-50 parts of starch, 40-50 parts of corncob degradation liquid, 15-25 parts of corn germ meal, 15-25 parts of soybean cake powder, 25-35 parts of corn steep liquor, 1.5-4 parts of calcium carbonate, 1-3.5 parts of monopotassium phosphate, 1-5 parts of corn oil, 3-9 parts of composite amino acid powder, 1-5 parts of vegetable oil, 1-5 parts of silicon-based defoaming agent and 800 parts of sterile water;

2) injecting the oxytetracycline fermentation liquor into an acidification tank, adding oxalic acid for acidification treatment, adjusting the pH value to 1.6-1.9, diluting the acidified oxytetracycline fermentation liquor, performing prefiltration, and performing plate-frame filtration twice to separate out hypha residues and oxytetracycline stock solution;

3) adding a buffer solution with the pH value of 5-7 into the oxytetracycline stock solution, adjusting the pH value back to 2-3.6, and filtering by using an ultrafiltration membrane;

4) adding hydrochloric acid into the oxytetracycline liquid ultrafiltered in the step 2 to adjust the pH value to 1.6-1.9, and then decoloring with decoloring resin to obtain a decoloring liquid;

5) filtering the decolorized oxytetracycline decolorized solution to remove residual resin, heating the decolorized solution to 28-32 ℃, and crystallizing;

6) adding ammonia water into the decolorized solution after temperature rise to adjust the pH value to 4.8-5.4, simultaneously carrying out ultrasonic intermittent stirring, and crystallizing for 40-60min to finish crystallization;

7) and (2) carrying out filter pressing on the crystallization liquid by using a filter press, separating to obtain oxytetracycline wet crystals, and circularly cleaning the oxytetracycline wet crystals by using pure water in the filter press, wherein the weight ratio of the pure water to the oxytetracycline wet crystals is controlled to be 5-10: 1, cleaning and drying to obtain a finished product of oxytetracycline.

The buffer solution in the step 3 is phosphate buffer solution.

The ultrafiltration membrane is an ultrafiltration membrane with the molecular weight of 1-5 ten thousand.

The parameters of the decolorizing resin are volume exchange capacity of 3eq/L and water content of 65-72%.

The ultrasonic frequency is 6.5-35kHz, and the intensity is 0.6-4W/cm²。

The ultrasonic intermittent stirring is ultrasonic stirring for 5-10min at an interval of 1-3 min.

The ultrasonic frequency is gradually increased under the control of the ultrasonic intermittent stirring, the first ultrasonic frequency is 6.5-10kHz, and the ultrasonic frequency is gradually increased by 5-10kHz each time.

The cleaning time in the step 7 is 15-25 min.

Compared with the prior art, the invention has the following beneficial effects.

1. The invention fully utilizes corn as the raw material of a seed culture medium, firstly, corn kernels are separated from corncobs, corn germs of the separated corn kernels are separated out for preparing corn oil, corn germ meal is produced during oil preparation, meanwhile, a part of the corn kernels are used for preparing corn steep liquor, and a part of the threshed corn cobs are soaked, steamed and saccharified to prepare corn cob degradation liquid for supplementing a carbon source in the culture medium.

2. According to the method, the phosphoric acid buffer solution is added into the oxytetracycline stock solution, the pH value is adjusted back, and meanwhile, an ultrafiltration membrane filtration method is used, so that organic impurities are reduced, endotoxin is removed, the service life and the use efficiency of the resin can be prolonged when the decolorizing resin is used for decolorizing, and the decolorizing solution is more stable.

3. The method of ultrasonic intermittent stirring adopted by the invention is used for crystallization, wherein the intermittent ultrasonic stirring is adopted to increase the strength in a gradient manner, so that the purpose of adding seed crystals can be achieved in the early stage, meanwhile, the granularity of the crystals is more uniform along with the increase of the strength, and the water-soluble impurities are less and are easier to filter and dry by matching with the pure water cleaning process in the later stage.

Detailed Description

The invention is further illustrated by reference to the following examples.

Example 1

A fermentation process for producing oxytetracycline is characterized by comprising the following steps:

1) fermentation of terramycin

Threshing the corn cobs, respectively crushing corn kernels and corncobs, separating germs after the corn kernels are crushed, preparing corn steep liquor from the crushed corn kernels, and preparing corn oil from the separated germs and obtaining a byproduct of corn germ meal; preparing the corncob degradation liquid from the corncobs; after the preparation of the raw materials is finished, the specific fermentation process is as follows:

selecting streptomyces rimosus as a zymocyte, and fermenting to prepare a fermentation liquor through spore culture and three-level seed fermentation; the first-stage seed fermentation conditions are as follows: controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 40h, wherein the culture medium comprises the following components in parts by weight: 35 parts of starch, 35 parts of corncob degradation liquid, 15 parts of corn germ meal, 15 parts of soybean cake powder, 25 parts of corn steep liquor, 4 parts of calcium carbonate, 3.5 parts of monopotassium phosphate, 5 parts of corn oil, 9 parts of composite amino acid powder, 5 parts of vegetable oil, 5 parts of silicon-based defoaming agent and 1500 parts of sterile water;

the secondary seed fermentation conditions are as follows:

controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 40h, wherein the culture medium comprises the following components in parts by weight: 40 parts of starch, 40 parts of corncob degradation liquid, 15 parts of corn germ meal, 20 parts of soybean cake powder, 30 parts of corn steep liquor, 4 parts of calcium carbonate, 3.5 parts of monopotassium phosphate, 5 parts of corn oil, 9 parts of composite amino acid powder, 5 parts of vegetable oil, 5 parts of silicon-based defoaming agent and 1200 parts of sterile water;

the fermentation conditions of the third-level seeds are as follows: controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 40h, wherein the culture medium comprises the following components in parts by weight: 50 parts of starch, 50 parts of corncob degradation liquid, 25 parts of corn germ meal, 25 parts of soybean cake powder, 35 parts of corn steep liquor, 4 parts of calcium carbonate, 3.5 parts of monopotassium phosphate, 5 parts of corn oil, 9 parts of composite amino acid powder, 5 parts of vegetable oil, 5 parts of silicon-based defoaming agent and 800 parts of sterile water;

2) injecting the oxytetracycline fermentation liquor into an acidification tank, adding oxalic acid for acidification treatment, adjusting the pH value to 1.6-1.9, diluting the acidified oxytetracycline fermentation liquor, performing prefiltration, and performing plate-frame filtration twice to separate out hypha residues and oxytetracycline stock solution;

3) adding a phosphate buffer solution with the pH value of 5-7 into the oxytetracycline stock solution, adjusting the pH value back to 2-3.6, and filtering by using an ultrafiltration membrane with the molecular weight of 1-5 ten thousand;

4) adding hydrochloric acid into the oxytetracycline liquid ultrafiltered in the step 2 to adjust the pH value to 1.6-1.9, and then decolorizing with decolorizing resin with volume exchange capacity of 3eq/L and water content of 65-72%, wherein the decolorizing resin is the obtained decolorizing liquid;

5) filtering the decolorized oxytetracycline decolorized solution to remove residual resin, heating the decolorized solution to 28-32 ℃, and crystallizing;

6) adding ammonia water into the decolorized solution after temperature rise to adjust pH to 4.8-5.4, simultaneously performing ultrasonic intermittent stirring, firstly controlling ultrasonic frequency at 6.5kHz and intensity at 1W/cm²Ultrasonic stirring for 10min, intermittently stirring for 1min, and increasing ultrasonic frequency to 15kHz with intensity of 2W/cm²Ultrasonic stirring for 10min, intermittence for 2min, increasing the ultrasonic frequency to 25kHz, and increasing the intensity to 3W/cm²Ultrasonically stirring for 10min, pausing for 1min, then standing for crystallization for 25min, and finishing crystallization;

7) and (2) carrying out filter pressing on the crystallization liquid by using a filter press, separating to obtain oxytetracycline wet crystals, and circularly cleaning the oxytetracycline wet crystals by using pure water in the filter press, wherein the weight ratio of the pure water to the oxytetracycline wet crystals is controlled to be 10: 1, washing for 25min, and drying after washing to obtain a finished product of oxytetracycline.

Example 2

A fermentation process for producing oxytetracycline is characterized by comprising the following steps:

1) fermentation of terramycin

Threshing the corn cobs, respectively crushing corn kernels and corncobs, separating germs after the corn kernels are crushed, preparing corn steep liquor from the crushed corn kernels, and preparing corn oil from the separated germs and obtaining a byproduct of corn germ meal; preparing the corncob degradation liquid from the corncobs; after the preparation of the raw materials is finished, the specific fermentation process is as follows:

selecting streptomyces rimosus as a zymocyte, and fermenting to prepare a fermentation liquor through spore culture and three-level seed fermentation; the first-stage seed fermentation conditions are as follows: controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 30h, wherein the culture medium comprises the following components in parts by weight: 20 parts of starch, 20 parts of corncob degradation liquid, 5 parts of corn germ meal, 5 parts of soybean cake powder, 10 parts of corn steep liquor, 1.5 parts of calcium carbonate, 1 part of monopotassium phosphate, 1 part of corn oil, 3 parts of composite amino acid powder, 1 part of vegetable oil, 1 part of silicon-based defoaming agent and 1000 parts of sterile water;

the secondary seed fermentation conditions are as follows:

controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 30h, wherein the culture medium comprises the following components in parts by weight: 30 parts of starch, 30 parts of corncob degradation liquid, 10 parts of corn germ meal, 10 parts of soybean cake powder, 15 parts of corn steep liquor, 1.5 parts of calcium carbonate, 1 part of monopotassium phosphate, 1 part of corn oil, 3 parts of composite amino acid powder, 1 part of vegetable oil, 1 part of silicon-based defoaming agent and 600 parts of sterile water;

the fermentation conditions of the third-level seeds are as follows: controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 30h, wherein the culture medium comprises the following components in parts by weight: 40 parts of starch, 40 parts of corncob degradation liquid, 15 parts of corn germ meal, 15 parts of soybean cake powder, 25 parts of corn steep liquor, 1.5 parts of calcium carbonate, 1 part of monopotassium phosphate, 1 part of corn oil, 3 parts of composite amino acid powder, 1 part of vegetable oil, 1 part of silicon-based defoaming agent and 500 parts of sterile water;

2) injecting the oxytetracycline fermentation liquor into an acidification tank, adding oxalic acid for acidification treatment, adjusting the pH value to 1.6-1.9, diluting the acidified oxytetracycline fermentation liquor, performing prefiltration, and performing plate-frame filtration twice to separate out hypha residues and oxytetracycline stock solution;

3) adding a phosphate buffer solution with the pH value of 5-7 into the oxytetracycline stock solution, adjusting the pH value back to 2-3.6, and filtering by using an ultrafiltration membrane with the molecular weight of 1-5 ten thousand;

4) adding hydrochloric acid into the oxytetracycline liquid ultrafiltered in the step 2 to adjust the pH value to 1.6-1.9, and then decolorizing with decolorizing resin with volume exchange capacity of 3eq/L and water content of 65-72%, wherein the decolorizing resin is the obtained decolorizing liquid;

5) filtering the decolorized oxytetracycline decolorized solution to remove residual resin, heating the decolorized solution to 28-32 ℃, and crystallizing;

6) adding ammonia water into the decolorized solution after temperature rise to adjust pH to 4.8-5.4, and simultaneously performing ultrasonic intermittent stirring, wherein the ultrasonic frequency is controlled to be 20kHz, and the intensity is 1W/cm²Ultrasonic stirring for 5min, pausing for 5min, then increasing the ultrasonic frequency to 25kHz, and the intensity to be 2W/cm²Ultrasonic stirring for 6min, intermittent stirring for 10min, increasing ultrasonic frequency to 30kHz, and increasing intensity to 3W/cm²Ultrasonic stirring for 10min, intermittent stirring for 1min, controlling frequency to 35kHz, and controlling intensity to be 3W/cm²Ultrasonically stirring for 5min, standing for crystallizing for 5min to complete crystallization;

7) and (3) carrying out filter pressing on the crystallization liquid by using a filter press, separating to obtain oxytetracycline wet crystals, and circularly cleaning the oxytetracycline wet crystals by using pure water in the filter press, wherein the weight ratio of the pure water to the oxytetracycline wet crystals is controlled to be 5: 1, cleaning for 15min, and drying after cleaning to obtain a finished product of oxytetracycline.

Example 3

A fermentation process for producing oxytetracycline is characterized by comprising the following steps:

1) fermentation of terramycin

Threshing the corn cobs, respectively crushing corn kernels and corncobs, separating germs after the corn kernels are crushed, preparing corn steep liquor from the crushed corn kernels, and preparing corn oil from the separated germs and obtaining a byproduct of corn germ meal; preparing the corncob degradation liquid from the corncobs; after the preparation of the raw materials is finished, the specific fermentation process is as follows:

selecting streptomyces rimosus as a zymocyte, and fermenting to prepare a fermentation liquor through spore culture and three-level seed fermentation; the first-stage seed fermentation conditions are as follows: controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 33h, wherein the culture medium comprises the following components in parts by weight: 25 parts of starch, 22 parts of corncob degradation liquid, 7 parts of corn germ meal, 5 parts of soybean cake powder, 21 parts of corn steep liquor, 2.5 parts of calcium carbonate, 1.9 parts of monopotassium phosphate, 2 parts of corn oil, 4 parts of composite amino acid powder, 3.3 parts of vegetable oil, 1.5 parts of silicon-based defoaming agent and 1450 parts of sterile water;

the secondary seed fermentation conditions are as follows:

controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 35 parts of starch, 35 parts of corncob degradation liquid, 12 parts of corn germ meal, 17 parts of soybean cake powder, 18 parts of corn steep liquor, 3 parts of calcium carbonate, 2 parts of monopotassium phosphate, 3 parts of corn oil, 4.5 parts of composite amino acid powder, 3.6 parts of vegetable oil, 3 parts of silicon-based defoaming agent and 1150 parts of sterile water;

the fermentation conditions of the third-level seeds are as follows: controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 48 parts of starch, 45 parts of corncob degradation liquid, 24 parts of corn germ meal, 20 parts of soybean cake powder, 32 parts of corn steep liquor, 3 parts of calcium carbonate, 3 parts of monopotassium phosphate, 3 parts of corn oil, 7 parts of composite amino acid powder, 2 parts of vegetable oil, 3 parts of silicon-based defoaming agent and 750 parts of sterile water;

2) injecting the oxytetracycline fermentation liquor into an acidification tank, adding oxalic acid for acidification treatment, adjusting the pH value to 1.6-1.9, diluting the acidified oxytetracycline fermentation liquor, performing prefiltration, and performing plate-frame filtration twice to separate out hypha residues and oxytetracycline stock solution;

3) adding a phosphate buffer solution with the pH value of 5-7 into the oxytetracycline stock solution, adjusting the pH value back to 2-3.6, and filtering by using an ultrafiltration membrane with the molecular weight of 1-5 ten thousand;

4) adding hydrochloric acid into the oxytetracycline liquid ultrafiltered in the step 2 to adjust the pH value to 1.6-1.9, and then decolorizing with decolorizing resin with volume exchange capacity of 3eq/L and water content of 65-72%, wherein the decolorizing resin is the obtained decolorizing liquid;

5) filtering the decolorized oxytetracycline decolorized solution to remove residual resin, heating the decolorized solution to 28-32 ℃, and crystallizing;

6) adding ammonia water into the decolorized solution after temperature rise to adjust pH to 4.8-5.4, and simultaneously performing ultrasonic intermittent stirring, wherein the ultrasonic frequency is controlled at 15kHz, and the intensity is 1W/cm²Ultrasonic stirring for 10min, pausing for 5min, then increasing the ultrasonic frequency to 20kHz, and the intensity to be 2W/cm²Ultrasonic stirring for 10min, pausing for 5min, increasing ultrasonic frequency to 25kHz, and increasing intensity to 3W/cm²Ultrasonically stirring for 10min, standing for crystallizing for 5min to complete crystallization;

7) and (3) carrying out filter pressing on the crystallization liquid by using a filter press, separating to obtain oxytetracycline wet crystals, and circularly cleaning the oxytetracycline wet crystals by using pure water in the filter press, wherein the weight ratio of the pure water to the oxytetracycline wet crystals is controlled to be 6: 1, cleaning for 20min, and drying after cleaning to obtain a finished product of oxytetracycline.

Example 4

A fermentation process for producing oxytetracycline is characterized by comprising the following steps:

1) fermentation of terramycin

Threshing the corn cobs, respectively crushing corn kernels and corncobs, separating germs after the corn kernels are crushed, preparing corn steep liquor from the crushed corn kernels, and preparing corn oil from the separated germs and obtaining a byproduct of corn germ meal; preparing the corncob degradation liquid from the corncobs; after the preparation of the raw materials is finished, the specific fermentation process is as follows:

selecting streptomyces rimosus as a zymocyte, and fermenting to prepare a fermentation liquor through spore culture and three-level seed fermentation; the first-stage seed fermentation conditions are as follows: controlling the temperature to be 26-34 ℃, the pressure to be 0.01-0.05MP, and the culture time to be 35h, wherein the culture medium comprises the following components in parts by weight: 30 parts of starch, 33 parts of corncob degradation liquid, 11 parts of corn germ meal, 6 parts of soybean cake powder, 18 parts of corn steep liquor, 2.5 parts of calcium carbonate, 2 parts of monopotassium phosphate, 3 parts of corn oil, 4 parts of composite amino acid powder, 3 parts of vegetable oil, 1.5 parts of silicon-based defoaming agent and 1300 parts of sterile water;

the secondary seed fermentation conditions are as follows:

controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 35 parts of starch, 35 parts of corncob degradation liquid, 12 parts of corn germ meal, 15 parts of soybean cake powder, 20 parts of corn steep liquor, 3 parts of calcium carbonate, 2 parts of monopotassium phosphate, 3 parts of corn oil, 4.5 parts of composite amino acid powder, 3.5 parts of vegetable oil, 2 parts of silicon-based defoaming agent and 1000 parts of sterile water;

the fermentation conditions of the third-level seeds are as follows: controlling the temperature at 26-34 ℃, the pressure at 0.01-0.05MP and the culture time at 30-40h, wherein the culture medium comprises the following components in parts by weight: 45 parts of starch, 45 parts of corncob degradation liquid, 25 parts of corn germ meal, 22 parts of soybean cake powder, 32 parts of corn steep liquor, 2.5 parts of calcium carbonate, 3 parts of monopotassium phosphate, 3 parts of corn oil, 6 parts of composite amino acid powder, 3 parts of vegetable oil, 3 parts of silicon-based defoaming agent and 750 parts of sterile water;

2) injecting the oxytetracycline fermentation liquor into an acidification tank, adding oxalic acid for acidification treatment, adjusting the pH value to 1.6-1.9, diluting the acidified oxytetracycline fermentation liquor, performing prefiltration, and performing plate-frame filtration twice to separate out hypha residues and oxytetracycline stock solution;

3) adding a phosphate buffer solution with the pH value of 5-7 into the oxytetracycline stock solution, adjusting the pH value back to 2-3.6, and filtering by using an ultrafiltration membrane with the molecular weight of 1-5 ten thousand;

4) adding hydrochloric acid into the oxytetracycline liquid ultrafiltered in the step 2 to adjust the pH value to 1.6-1.9, and then decolorizing with decolorizing resin with volume exchange capacity of 3eq/L and water content of 65-72%, wherein the decolorizing resin is the obtained decolorizing liquid;

5) filtering the decolorized oxytetracycline decolorized solution to remove residual resin, heating the decolorized solution to 28-32 ℃, and crystallizing;

6) adding ammonia water into the decolorized solution after temperature rise to adjust pH to 4.8-5.4, and simultaneously performing ultrasonic intermittent stirring, wherein the ultrasonic frequency is controlled at 15kHz, and the intensity is 1W/cm²Ultrasonic stirring for 5min, pausing for 5min, then increasing ultrasonic frequency to 20kHz, and the intensity is 2W/cm²Ultrasonic stirring for 5min, pausing for 5min, increasing ultrasonic frequency to 25kHz, and increasing intensity to 3W/cm²Ultrasonic stirring for 5min, pausing for 5min, controlling frequency to 30kHz, and controlling intensity to be 4W/cm²Ultrasonically stirring for 5min, standing for crystallizing for 5min to complete crystallization;

7) and (3) carrying out filter pressing on the crystallization liquid by using a filter press, separating to obtain oxytetracycline wet crystals, and circularly cleaning the oxytetracycline wet crystals by using pure water in the filter press, wherein the weight ratio of the pure water to the oxytetracycline wet crystals is controlled to be 8: 1, cleaning for 22min, and drying after cleaning to obtain a finished product of oxytetracycline.

The above embodiments are merely illustrative of the principles of the present invention and its effects, and do not limit the present invention. It will be apparent to those skilled in the art that modifications and improvements can be made to the above-described embodiments without departing from the spirit and scope of the invention. Accordingly, it is intended that all equivalent modifications or changes be made by those skilled in the art without departing from the spirit and technical spirit of the present invention, and be covered by the claims of the present invention.