CN113621667B - Electromagnetic coupling fermentation streptomyces glaucescens Hainan variety microorganism cell - Google Patents
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Abstract
The invention relates to a method for fermenting Streptomyces lilacinus Hainan variety by electromagnetic couplingStreptomyces lavendulae var. hainanensis new var.) The microbial cell comprises an anode carbon rod, a cathode carbon rod, a shake flask culture medium, a fixed resistor, a variable resistor, a proton exchange membrane, wastewater preparation liquid, an external electric field, a workstation, a pH sensor, a dissolved oxygen sensor, a stirrer, an oxygenation device, an external magnetic field and an anode carbon rod for inoculating the Streptomyces lilacinus Hainan variety. The invention can improve the production titer of Streptomyces lilacinus and degrade the wastewater of biological pesticide in the fermentation production process by adopting electromagnetic coupling environment.
Description
Technical Field
The invention belongs to the field of microbial pharmacy, and particularly relates to an electromagnetic coupling fermentation Streptomyces lilacinus Hainan variety microbial cell and application thereof.
Background
The Zhongshengmycin belongs to N-glycoside agricultural antibiotics, contains 6 effective components, is sequentially increased with one beta-lysine, has activity on bacteria, saccharomycetes and filamentous fungi diseases, is an environment resource protection type biological sterilization treatment drug which is prepared by processing and preparing biological bactericides, has good inhibition or effective inactivation effect on various gram rod-shaped positive bacteria, gram rod-shaped negative bacteria and filamentous negative fungi, has very wide application of the Zhongshengmycin in agriculture, and mainly prevents and treats rice fine streak diseases, strip spot diseases, bacterial leaf blight, black rot of Chinese cabbage, soft leather diseases, green pepper red blight, light purple canker ginger blast, angular leaf spot diseases of cucumbers, perforated diseases of peach trees, apricot trees and plum trees, bacterial influenza of kidney beans and the like; the fungus control mainly comprises apple tree ring rot, alternaria leaf spot, wheat scab, melon fusarium wilt, peach tree, apricot tree, plum tree carbuncle sore, citrus tree ulcer carbuncle sore, pus scar carbuncle sore, huang Lusun stem-blight carbuncle sore and the like, and the Zhongshengmycin has low toxicity to people and animals and does not pollute the environment, thus being an ideal antibacterial and antifungal biological pesticide.
Compared with other novel prior art which utilizes the organic production capacity, the microbial cell (MFC) has the characteristics and functional advantages of simplicity and easiness in operation, namely, firstly, the original matrix is directly changed into electric power, and the conversion of high energy is ensured; second, unlike all current bioenergy processing methods, microbial fuel solar cells can operate efficiently in the environment; thirdly, microbial cells do not have to be subjected to waste gas treatment, since the main exhaust component of waste gas generated in the cells is carbon dioxide, and generally none of these has a reusable energy source; fourth, the microbial fuel cell does not have to input much energy into the air, since a single cell microbial cell only needs ventilation for a single air, and can passively replenish the anode gas into the air; fifth, in some countries where there is still a poverty, there is not enough funds to support the development of microbial fuel solar cells, and these developing fuels are more diverse. Although the microbial cell has the advantages, the anode microbial utilization and biological value excavation of the microbial cell are insufficient, the anode microbial cell only transmits redundant electrons generated by metabolism to the cathode chamber through the anode region of the anode chamber and through an externally connected lead, and the redundant electrons react with oxygen and waste water substrates of the cathode chamber to decompose waste water organic matters, and although the microbial cell has the two functions of degrading cathode waste water and generating electricity, metabolic products generated by microbial fermentation of the anode chamber are often ignored, even when the microbial fermentation products of the anode chamber are treated as three wastes, the anode microbial value utilization of the microbial cell is greatly influenced, huge resource waste is caused, and the potential utilization value of the anode microbial of the microbial cell is fully excavated, so that more beneficial experimental evidence and reference value are provided for later research of the microbial cell.
Disclosure of Invention
In order to solve the problem of the prior art that Streptomyces lilacinus is a variety of HainanStreptomyces lavendulae var. hainanensis new var.) The invention provides an electromagnetic coupling fermentation Streptomyces lilacinus Hainan variety microbial cell, which solves the technical problem of low content of antibiotics in the existing strain. The original strain of the invention is Streptomyces lilacinus Hainan variety @Streptomyces lavendulae var. hainanensis new var.)。
More specifically, the invention relates to an electromagnetic coupling fermentation streptomyces lilacinus Hainan variety microbial cell which is characterized by comprising an anode carbon rod, a cathode carbon rod, a shake flask culture medium, a fixed resistor, a variable resistor, a proton exchange membrane, wastewater preparation liquid, an external electric field, a workstation, a pH sensor, a dissolved oxygen sensor, a stirrer, an oxygenation device and an external magnetic field, wherein the anode carbon rod, the pH sensor, the dissolved oxygen sensor and the stirrer are positioned in the shake flask culture medium, the cathode carbon rod is positioned in the wastewater preparation liquid, the anode carbon rod and the cathode carbon rod are electrically connected with the variable resistor through the fixed resistor, the proton exchange membrane is positioned between the shake flask culture medium and the wastewater preparation liquid, the shake flask culture medium and the wastewater preparation liquid are positioned between the external electric field, the shake flask culture medium and the wastewater preparation liquid are positioned between the external magnetic field, the oxygenation device is connected with the shake flask culture medium, and the anode carbon rod is inoculated with the streptomyces lilacinus Hainan variety.
In a preferred embodiment of the invention, the microbial cell further comprises an acid and/or base addition bottle for conditioning the shake flask medium, and a container for automatic feeding of the shake flask medium.
In a preferred embodiment of the invention, the applied magnetic field and the applied electric field are both adjustable.
In a preferred embodiment of the invention, the magnetic induction of the externally applied magnetic field is adjustable in the range of 0.1 to 0.6T, preferably 0.15 to 0.25T.
In a preferred embodiment of the invention, the applied electric field is regulated by a direct current source, the current intensity of which is adjustable in the range of 0-100mA.
The invention also relates to a method for simultaneously producing the Zhongshengmycin and treating the wastewater, which is characterized in that the microbial cell is adopted, wherein the Zhongshengmycin is obtained from a shake flask culture medium, and the wastewater is treated in a wastewater preparation liquid.
In a preferred embodiment of the invention, the wastewater preparation is characterized in that the wastewater for producing kasugamycin by biological fermentation has an initial COD concentration of 600-700mg/mL and an initial ammonia nitrogen concentration of 200-300mg/mL. Preferably, the COD reduction rate of the treated wastewater preparation liquid is more than 95%, preferably more than 98%, and the ammonia nitrogen reduction rate is more than 90%, preferably more than 94%.
In a preferred embodiment of the invention, the shake flask medium is formulated as follows: soluble starch (AR) 1.0%, mgSO 4 ·7H 2 O（AR）0.045%、KH 2 PO 4 （AR）0.10%、KCl（AR）0.045%、NaNO 3 （AR）0.125%、FeSO 4 ·7H 2 O (AR) 0.001%, and distilled water in balance, pH6.8-7.0.
In a preferred embodiment of the invention, the shaking culture medium is stirred at a speed of 120-160rmp, at a temperature of 28-30℃and a ventilation of 1:1-2 (V/V) per minute; the automatic material changing amount is 1/5-1/3 of the liquid volume of the anode chamber every day.
In a preferred embodiment of the invention, the direct current is 30-50mA and the magnetic field is 0.15-0.25T.
Further, inoculating a culture solution of a strain of Streptomyces lilacinus Hainan variant mutagenesis in the medium of claim 6 to 5-15% (V/V), preferably 10%; the stirring speed of the anode chamber is 120-160rmp, preferably 150rmp, the culture temperature is 28-30 ℃, and the ventilation amount per minute is 1:2-1:4 (V/V) of sterilizing air, preferably 1:1; the automatic material changing amount is 1/10-1/3, preferably 1/5 of the liquid volume of the anode chamber every day; the cultivation time is 72-128 hours, preferably 84 hours.
Compared with the prior art, the invention has the following advantages: on one hand, the production titer of the streptomyces lilacinus can be improved under the electromagnetic coupling environment, and on the other hand, the constructed MFC is used for degrading the wastewater of the biological pesticide in the fermentation production process, the construction effect of the biological cell is obviously different from that of the prior biological cell, the device fully excavates the anode microorganism economic benefit of the MFC, and the wastewater in the cathode region of the MFC can be treated simultaneously.
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Further description is provided below with reference to the accompanying drawings.
FIG. 1 shows a schematic structure of an electromagnetic coupling fermentation Streptomyces lilacinus Hainan variety microbial cell, wherein the structure comprises a 1-anode carbon rod, a 2-cathode carbon rod, a 3-shake flask culture medium, a 4-custom resistor, a 5-variable resistor, a 6-proton exchange membrane, a 7-wastewater manual preparation liquid, an 8-externally applied electric field, a 9-workstation, a 10-pH sensor 11-dissolved oxygen sensor, a 12-stirrer, a 13-HCl (0.1 mol/L) micro-addition bottle, a 14-NaOH (0.1 mol/L) micro-addition bottle, a 15-automatic feed supplement bottle, a 16-oxygenation device and a 17-externally applied magnetic field.
FIG. 2 shows the change of the enzymes of the middle-growth fungus and mitochondria of the Hainan variety of Streptomyces lilacinus fermented by non-applied electric and magnetic fields.
FIG. 3. Variation of the enzyme in the middle-growth hormone and mitochondria of the Hainan variety of Streptomyces lilacinus with the addition of 20 mA.
FIG. 4. Variation of the enzyme in the middle-growth of the Hainan variety of Streptomyces lilacinus plus 40 mA.
FIG. 5. Variation of the enzyme in the middle-growth of the Hainan variety of Streptomyces lilacinus plus 80 mA.
FIG. 6. Changes in the activity of the enzymes of the middle-growth of the Hainan variety of Streptomyces lilacinus with the addition of 40mA and 0.1T magnetic field.
FIG. 7. Changes in the activity of the enzymes of the middle-growth of the Hainan variety of Streptomyces lilacinus with the addition of 40mA and 0.2T magnetic field.
FIG. 8 variation of the enzymes of the middle-growth hormone and mitochondria of the Hainan variety of Streptomyces lilacinus under an applied magnetic field of 40mA and 0.4T
Detailed Description
The present invention will be described in further detail with reference to specific embodiments in order to make the objects, technical solutions and advantages of the present invention more apparent. It should be understood that the specific embodiments described herein are for purposes of illustration only and are not to be construed as limiting the invention.
The invention is further illustrated by the following examples.
Example 1: inoculating Streptomyces lilacinus in the anode region of MFC, wherein the anode inoculating solution comprises glucose (AR) 1.5%, cold pressed soybean cake powder (IR) 2.0%, corn powder (FR) 0.15%, corn starch (FR) 1.5%, and NH 4 Cl（AR）0.45%、NaCl（AR）0.35%、MgSO 4 ·7H 2 O（AR）0.03%、KH 2 PO 4 (AR) 0.03%, light CaCO 3 (AR) 0.04%, and tap water, wherein the pH value of the shake flask seed culture medium before sterilization is 6.5-7.5, the fermentation temperature of the Zhongshengmycin is 25 ℃, the ventilation rate is 600-650L/min, the stirring speed is 120rmp, the inoculation amount of Streptomyces lilacinus is 10%, the feeding amount is 3 mL/L.h, the cathode liquid is Shanxi Mukouo kasugamycin fermentation workshop wastewater, the COD initial concentration of the configured wastewater is 650mg/L, and the ammonia nitrogen concentration is 65mg/L. Thus, MFC of Streptomyces lilacinus was constructed.
Example 2:
inoculating Streptomyces lilacinus in the anode region of MFC, wherein the anode inoculating solution is 1.5% glucose (AR), 2.0% cold pressed soybean cake powder (IR), and semen Maydis0.15% of Flour (FR), 1.5% of corn starch (FR) and NH 4 Cl（AR）0.45%、NaCl（AR）0.35%、MgSO 4 ·7H 2 O（AR）0.03%、KH 2 PO 4 (AR) 0.03%, light CaCO 3 (AR) 0.04%, and tap water, wherein the pH value of the shake flask seed culture medium before sterilization is 6.5-7.5, the fermentation temperature of the Zhongshengmycin is 25 ℃, the ventilation rate is 600-650L/min, the stirring speed is 120rmp, the inoculation amount of Streptomyces lilacinus is 10%, the feeding amount is 3 mL/L.h, the trend of change of the F content of the Zhongshengmycin along with the extension of the fermentation time is basically unchanged after the increase, and the F content of the Zhongshengmycin reaches the maximum value after the fermentation time is 24 h. The change trend of the mitochondrial enzyme I content along with the extension of the fermentation time is that the mitochondrial enzyme I content is slowly increased and then decreased, and the mitochondrial enzyme I content reaches the maximum value within about 72 hours of the fermentation time. The trend of the mitochondrial enzyme II content over the extended fermentation time is continuously and slowly increasing. The trend of the mitochondrial enzyme III content with the extension of the fermentation time is that the mitochondrial enzyme III content is gradually increased and then decreased, and the mitochondrial enzyme III content reaches the maximum value about 36 hours of the fermentation time (the specific change is shown in figure 2).
The mitochondrial enzyme assay method for the fermentation process of the strain of Streptomyces lilacinus Hainan variant is characterized by comprising the following main steps: a: pretreatment of a fermentation broth of a strain of Streptomyces lilacinus variant of Hainan: taking 0.5-2mL, preferably 0.9mL, of fermentation liquor in different fermentation periods; centrifugal speed is 15000-20000rmp, preferably 15000rmp; ultracentrifugation for 10-20min, preferably 15min; taking the supernatant as a mitochondrial complex enzyme system sample for testing a strain of Streptomyces lilacinus Hainan variant mutagenesis; the method for determining the mitochondrial complex enzyme I of the strain mutagenized by the Hainan variant of the streptomyces anodinius comprises the following steps: a900. Mu.l of the A-treated test sample was added with 100. Mu.l of a glycylglycine buffer (6 mmol/L NADH 0.2mmol/L, glycylglycine 0.5mmol, cytochrome C, pH8.5, reaction was carried out for 2 minutes, and absorbance was measured at a UV spectrometer at 550 nm. C.A-treated test sample was taken, 100. Mu.l of the A-treated test sample was added with a buffer (0.03M/L Potassium iron hydride 0.2mmol/LPBS,1% bovine serum albumin and 0.6mol/L succinic acid) and reacted for 2 minutes, D.A-treated test sample was taken, 100. Mu.l of the A-treated test sample was added with 900. Mu.l of a buffer (0.3 mmol/L cytochrome C) and reacted at 75mM/L phosphate buffer for 2 minutes, absorbance was measured at 550nm by a UV spectrometer
Example 3:
inoculating Streptomyces lilacinus in the anode region of MFC, wherein the anode inoculating solution comprises glucose (AR) 1.5%, cold pressed soybean cake powder (IR) 2.0%, corn powder (FR) 0.15%, corn starch (FR) 1.5%, and NH 4 Cl（AR）0.45%、NaCl（AR）0.35%、MgSO 4 ·7H 2 O（AR）0.03%、KH 2 PO 4 (AR) 0.03%, light CaCO 3 (AR) 0.04%, and tap water, wherein the pH value of the shake flask seed culture medium before sterilization is 6.5-7.5, the fermentation temperature of the Zhongshengmycin is 25 ℃, the ventilation rate is 600-650L/min, the stirring speed is 120rmp, the inoculation amount of Streptomyces lilacinus is 10%, the feeding amount is 3 mL/L.h, the change condition of the Zhongshengmycin content along with the fermentation time under the action of an external electric field of 20mA is shown in figure 3, the change trend of the Zhongshengmycin F content along with the extension of the fermentation time is basically unchanged after the increase, and the maximum value is reached after the fermentation time is 36h as can be seen from figure 3. The trend of the mitochondrial enzyme I content over the fermentation time is continuously increasing. The trend of the mitochondrial enzyme II content over the extended fermentation time is continuously and slowly increasing. The change trend of the mitochondrial enzyme III content along with the extension of the fermentation time is that the mitochondrial enzyme III content is slowly increased and then decreased, and the mitochondrial enzyme III content reaches the maximum value within about 72 hours of the fermentation time.
Example 4:
inoculating Streptomyces lilacinus in the anode region of MFC, wherein the anode inoculating solution comprises glucose (AR) 1.5%, cold pressed soybean cake powder (IR) 2.0%, corn powder (FR) 0.15%, corn starch (FR) 1.5%, and NH 4 Cl（AR）0.45%、NaCl（AR）0.35%、MgSO 4 ·7H 2 O（AR）0.03%、KH 2 PO 4 (AR) 0.03%, light CaCO 3 (AR) 0.04%, tap water, pH of the shake flask seed culture medium before sterilization of 6.5-7.5, fermentation temperature of zhongshengmycin of 25 ℃, ventilation of 600-650L/min, stirring speed of 120rmp, and inoculation amount of Streptomyces lilacinus ofThe feed amount is 3 mL/L.h, the change condition of the content of the biotin with the fermentation time under the action of an external electric field of 40mA is shown in figure 4, and the change trend of the content of the biotin F with the extension of the fermentation time under the external direct current of 40mA is that the content of the biotin F is increased firstly and then is basically unchanged and then is increased again, and the maximum value is reached after the fermentation time is 36h as can be seen from figure 4. The trend of the mitochondrial enzyme I content over the fermentation time is continuously increasing. The trend of the mitochondrial enzyme II content over the extended fermentation time is continuously increasing. The trend of the mitochondrial enzyme III content over the fermentation time was first to slowly increase.
Example 5:
inoculating Streptomyces lilacinus in the anode region of MFC, wherein the anode inoculating solution comprises glucose (AR) 1.5%, cold pressed soybean cake powder (IR) 2.0%, corn powder (FR) 0.15%, corn starch (FR) 1.5%, and NH 4 Cl（AR）0.45%、NaCl（AR）0.35%、MgSO 4 ·7H 2 O（AR）0.03%、KH 2 PO 4 (AR) 0.03%, light CaCO 3 (AR) 0.04%, and tap water, wherein the pH of the shake flask seed culture medium before sterilization is 6.5-7.5, the fermentation temperature of the Zhongshengmycin is 25 ℃, the ventilation rate is 600-650L/min, the stirring speed is 120rmp, the inoculation amount of Streptomyces lilacinus is 10%, the feeding amount is 3 mL/L.h, the change condition of the Zhongshengmycin content with the fermentation time under the action of an external electric field of 80mA is shown in figure 5, and the change trend of the Zhongshengmycin F content with the extension of the fermentation time is continuously reduced under the external electric field of 80 mA. The trend of the mitochondrial enzyme I content over the fermentation time was continuously decreasing. The trend of the mitochondrial enzyme II content over the extended fermentation time was continuously decreasing slowly. The trend of the mitochondrial enzyme III content over the fermentation time was first slowly decreasing.
Example 6:
inoculating Streptomyces lilacinus in the anode region of MFC, wherein the anode inoculating solution comprises glucose (AR) 1.5%, cold pressed soybean cake powder (IR) 2.0%, corn powder (FR) 0.15%, corn starch (FR) 1.5%, and NH 4 Cl（AR）0.45%、NaCl（AR）0.35%、MgSO 4 ·7H 2 O（AR）0.03%、KH 2 PO 4 (AR) 0.03%, light CaCO 3 （AR）0.04%、Preparing tap water, wherein the pre-sterilization pH value of the shake flask seed culture medium is 6.5-7.5, the fermentation temperature of the Zhongshengmycin is 25 ℃, the ventilation amount is 600-650L/min, the stirring speed is 120rmp, the inoculation amount of Streptomyces lilacinus is 10%, the feeding amount is 3 mL/L.h, the single variable is the direct current setting amount, the external direct current of 40mA is respectively used, the single variable is the magnetic field setting increment, the external magnetic fields of 0T,0.1T,0.2T and 0.4T are respectively used, the two factors are combined in a crossing manner, when the external direct current of 40mA and the external magnetic field of 0.2T are used, the F component content of Zhongshengmycin is the highest, the fermentation time is 84h, and the Zhongshengmycin content is 3.05g/L (see specific figures 6, 7 and 8).
Example 7:
inoculating Streptomyces lilacinus in the anode region of MFC, wherein the anode inoculating solution comprises glucose (AR) 1.5%, cold pressed soybean cake powder (IR) 2.0%, corn powder (FR) 0.15%, corn starch (FR) 1.5%, and NH 4 Cl（AR）0.45%、NaCl（AR）0.35%、MgSO 4 ·7H 2 O（AR）0.03%、KH 2 PO 4 (AR) 0.03%, light CaCO 3 (AR) 0.04%, and tap water, wherein the pH value of the shake flask seed culture medium before sterilization is 6.5-7.5, the fermentation temperature of the Zhongshengmycin is 25 ℃, the ventilation rate is 600-650L/min, the stirring speed is 120rmp, the inoculation amount of Streptomyces lilacinus is 10%, the feeding amount is 3 mL/L.h, the cathode liquid is Shanxi Mukouo kasugamycin fermentation workshop wastewater, the COD initial concentration of the configured wastewater is 650mg/L, and the ammonia nitrogen concentration is 65mg/L. When the external electric field is 40mA and the magnetic field is 0.2T, the COD water outlet concentration of the kasugamycin wastewater at the cathode region is 11.8mg/L (the COD reduction rate is more than 95%, preferably more than 98%), the ammonia nitrogen concentration of the outlet water is 3.83mg/L (the ammonia nitrogen reduction rate is more than 90%, preferably more than 94%), and the COD concentration and the ammonia nitrogen concentration of the outlet water are obviously lower than any combination of the strength of the external electric field and the strength of the magnetic field.
The foregoing is merely illustrative of specific embodiments of the present invention, and the scope of the invention is not limited thereto, but any changes or substitutions that do not undergo the inventive effort should be construed as falling within the scope of the present invention. Therefore, the protection scope of the present invention should be subject to the protection scope defined by the claims.
Claims (9)
1. Electromagnetic coupling fermentation Streptomyces lilacinus Hainan varietyStreptomyces lavendulae var. hainanensis new var.) The microbial cell is characterized by comprising an anode carbon rod, a cathode carbon rod, a shake flask culture medium, a fixed resistor, a variable resistor, a proton exchange membrane, a wastewater preparation liquid, an external electric field, a workstation, a pH sensor, a dissolved oxygen sensor, a stirrer, an oxygenation device and an external magnetic field, wherein the anode carbon rod, the pH sensor, the dissolved oxygen sensor and the stirrer are positioned in the shake flask culture medium, the cathode carbon rod is positioned in the wastewater preparation liquid, the anode carbon rod and the cathode carbon rod are electrically connected with the variable resistor through the fixed resistor, the proton exchange membrane is positioned between the shake flask culture medium and the wastewater preparation liquid, the shake flask culture medium and the wastewater preparation liquid are positioned between the external electric field, the shake flask culture medium and the wastewater preparation liquid are positioned between the external magnetic field, the oxygenation device is connected with the shake flask culture medium, and the anode carbon rod is inoculated with the Streptomyces lilacinus Hainan varietyStreptomyces lavendulae var. hainanensis new var.）。
2. The microbial cell of claim 1, further comprising an acid and/or base addition bottle for conditioning the shake flask medium, and a container for automatic feeding of the shake flask medium.
3. The microbial cell of claim 1, wherein the applied magnetic field and the applied electric field are both adjustable.
4. A microbial cell according to claim 3, wherein the magnetic induction of the externally applied magnetic field is adjustable in the range of 0.1-0.6T.
5. A device according to claim 3, wherein the applied electric field is regulated by a dc power supply and has a current intensity in the range of 0-100mA.
6. A method for simultaneously producing Zhongshengmycin and treating wastewater, which is characterized in that the microbial cell of any one of claims 1 to 5 is adopted, wherein Zhongshengmycin is obtained from a shake flask culture medium, and the wastewater is treated in a wastewater preparation liquid; the shaking culture medium and the wastewater are arranged between the external electric fields, the shaking culture medium and the wastewater are arranged between the external magnetic fields, the external electric fields are regulated by a direct current power supply, the direct current is 30-50mA, and the magnetic field is 0.15-0.25T.
7. The method of claim 6, wherein the wastewater preparation is a wastewater from the biological fermentation of kasugamycin production, the wastewater having an initial COD concentration of 600-700mg/mL and an initial ammonia nitrogen concentration of 200-300mg/mL.
8. The method of claim 6, wherein the shake flask medium is formulated as follows: 1.0% of soluble starch and MgSO 4 ·7H 2 O0.045%、KH 2 PO 4 0.10%、KCl0.045%、NaNO 3 0.125%、FeSO 4 ·7H 2 O0.001%, and distilled water in balance, pH6.8-7.0.
9. The method according to claim 6, wherein the shaking culture medium is stirred at a speed of 120-160rmp, the culture temperature is 28-30 ℃, and the aeration rate per minute is 1:1-2 (V/V) of sterilized air; the automatic material changing amount is 1/5-1/3 of the liquid volume of the anode chamber every day.
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