Preparation before fermentation
Check that the power supply is normal and that the air compressor, microcomputer system, and circulating water system are working properly.
Check that the valves, fittings, and fastening screws on the system are tight.
Start the air compressor and check the seed tank, fermenter , filter, pipeline, valve, etc. with a pressure of 0.15Mpa for good sealing and leakage. Whether the tank jacket is sealed with the tank (should be tested during the season) to ensure that all valves are closed (except for the valve in front of the solenoid valve).
Check if the water (cooling water) pressure, voltage, and gas (steam) pressure are normally supplied. The inlet water pressure is maintained at 0.12Mpa, allowed to vary from 0.15-0.2Mpa, not more than 0.3Mpa, the temperature should be lower than the fermentation temperature by 10°C; single-phase power supply AC220V±10%, frequency 50Hz, the tank is reliably grounded; input steam pressure It should be maintained at 0.4Mpa, the pressure reduced to 0.24MPa after entering the system; the air compressor pressure value is 0.8Mpa, and the air inlet pressure should be controlled at 0.25-0.30MP (pressure value of air primary filter).
Temperature, dissolved oxygen electrode, PH electrode calibration and calibration, see the touch screen PH, DO calibration help.
Check that each motor is working properly (4 in total). Whether the solenoid valve can be normally sucked in (the whole system has a total of 11 solenoid valves).
The first cleaning after the sterilization fermentation system is installed <br>The cleaning in the tank: the acid and alkali tank, the feeding tank and the seed tank can unload the flange above the tank, and the operator manually cleans it with a clean cloth. Drain the sewage in the tank and wash it several times. The washing tank can be washed by a tap pipe through the hand hole to the inner wall of the tank. When the water level rises to the second impeller of the stirring shaft, the flushing is stopped, and the motor is driven to be stirred and cleaned. The cleaning of each pipeline can be washed first with clean water, and then the corresponding cleaning medium should be used according to the corresponding function (the preconditions for protecting various components in the pipeline should be taken when cleaning the pipeline). For details, please refer to “Exhaustion of air pipeline bacteria". If the fermentation system is not used for a long time or the cultured cells are different from the previous batch, it can be washed with 2% NaOH. The other cans can also be cleaned by the washing method of the fermenter. After the cleaning, the fermentation system should be sterilized.
Sterilization of air lines
The sterilizing filter on the air line uses steam through the pressure reducing valve (the air pressure reducing valve cannot be steam sterilized, so the air pre-filter is not sterilized), the steam filter then enters the sterilizing filter.
The filter element of the air sterilization filter cannot withstand high temperature and high pressure. Therefore, the steam pressure reducing valve should be adjusted to 0.13 MPa and not more than 0.15 MPa.
During the emptying process, the exhaust valve at the lower end of the sterilization filter should be slightly opened to remove the condensate.
The emptying time should last for about 30 minutes. When the equipment is used for the first time or after long-term use, it is best to use intermittent emptying, that is, after the first emptying, it will be removed once every 3 to 5 hours to eliminate spores.
After the air filter is removed, it should be blown dry for about 20 to 30 minutes, then the gas path valve is closed. Keep the air line positive.
Fermentation tank emptying (seed tank, feeding tank, acid and alkali tank emptying as a reference basis)
Before the fermenter is empty, the blowdown valve Q22 should be opened (so that the pressure inside the jacket is not overpressured).
Close Q14, open Q15 slightly, open steam valve Q12, open Q13 to send steam to the tank; open Q18, Q19 through the sampling port to pass steam into the tank.
The inoculating port on the tank, the exhaust valve Q9, and the valves Q24 and Q25 on the sewage line are slightly opened, so that the steam is discharged through these valves. When the humidity reaches 1220C, the timing is started, and the opening degrees of the valves Q9, Q13, and Q19 are adjusted. Keep the temperature inside the tank 1220C~1280C (the pressure is generally 0.11~0.15Mpa), and adjust the air temperature and pressure according to the process.
When the time reaches 30~40 minutes, close Q13, Q15, Q19, then close Q12, Q18, open the valve Q14, Q13 on the air line to cool the air in the tank, and keep the positive pressure in the tank at 0.03MPa-0.05 Between MPa.
When rapid cooling is required, the jacket drain valve Q22 is closed, and the cooling water valve Q27/DC1 is opened to the jacket to be cooled by water, and the cooling water is turned off after reaching the normal temperature. In special cases, intermittent emptying can be used (three to five hours and then empty once to eliminate spores).
When empty, the dissolved oxygen and PH electrodes are taken out and stored properly to prolong their service life.
The fermenter can be eliminated (seed tank, feed tank, acid and alkali tank can be used as a reference basis)
The elimination is the process of sterilizing the medium with steam after the medium is added to the tank.
After the end of the air consumption, close the intake valve Q13, open the Q9 to remove the pressure inside the tank, install the corrected PH and DO electrodes, and add the prepared medium to the tank as soon as possible.
Before the medium enters the tank, it should be gelatinized. The formula of the general medium should be determined according to the process requirements. The final volume of the fermentation liquid is calculated as about 70% of the full volume of the tank (the medium with more foam is about 60%, The medium with less foam can reach 75-80%. Considering the factors of condensed water and inoculum, and whether it is added or not, the initial volume is determined by the production according to the requirements of the process, and it needs to be explored in practice.
Turn on the mechanical stirring device and rotate at low speed to mix the materials in the tank evenly.
Open the jacket drain valve Q22 and steam valve Q21, and preheat the medium in the tank (pre-heating with jacketed steam). When the temperature inside the tank rises to 90 °C (the specific temperature should be changed according to the steam quality), close the jacket inlet valve Q21, close Q14, fully open the inlet valve Q12, open the Q13 steam, open Q18, open slightly Q19 through the sampling port to the tank to pass steam, close Q25, Q26, fully open Q23, micro-open Q24 steam to the tank, micro-open exhaust valve Q9 (three-way steam sterilization), turn off the motor stirring.
When the temperature rises to 121-123 ° C, the tank pressure rises to 0.12 MPa, the opening degree of the steam valves Q9, Q19, Q13, Q24 is controlled, the temperature and the tank pressure are maintained, and the timing is started, and the micro-opening flame inoculating port is discharged outward. Before the time reaches 30 minutes, close the flame inoculating port and close the Q24 (the amount of steam should be artificially opened several times before closing to avoid the medium remaining at the valve), close Q23, close Q19, 18, close Q13, Q12 to stop. Supply steam.
Close the jacket drain valve Q22, open the cooling water valve Q27/DC1 to the jacket to pass the water to cool, open the motor to stir, when the pressure drops to 0.05MPa, open the air valve Q14, Q13 to the tank to pass air, speed up the cooling speed, and keep The tank pressure was 0.05 MPa until the temperature of the medium in the tank dropped to the inoculation temperature. When the temperature in the tank drops to 2-3 °C higher than the temperature required for the fermentation process, the valve Q27/DC1 is closed to stop the cooling water.
Seed tank inoculation, seeding
Vaccination:
The system is inoculated with flame sealing. It should be prepared before inoculation: alcohol cotton, pliers, tweezers, inoculating ring, asbestos net gloves, Z-wrench.
The strain is placed in a conical flask, and the inoculum amount is determined according to the process requirements.
The alcohol cotton is ignited around the inoculation ring, and the bottle mouth is burned on the flame for a while. The Z-shaped wrench is used to unscrew the inoculating port, and the strain is quickly poured into the tank. At this time, the tank should be ventilated to make the inoculating port. There is air discharge (pressure is close to zero, but not zero).
Tighten the inoculum flap on the flame and sterilize. After inoculation, the tank pressure is maintained at 0.03 MPa to 0.07 MPa to adjust the ventilation.
Seeding:
Before the seeding, the condensed water in the steam line should be drained. After the emptying, keep the steam valve slightly open (only the water does not discharge steam) so that the condensed water in the steam line can be emptied at any time.
Special attention should be paid to adjusting the pressure difference between the seed tank and the culture tank when transplanting. The pressure of the seed tank should be higher than 0.1MPa of the culture tank to transplant! Seeding should be carried out immediately after shutting down the steam valve to avoid negative pressure pollution.
Immediately after the transplanting, the seeding valve should be closed, then the steam valve connected to it should be opened for steaming and disinfection, and the seed tank should be cleaned immediately to prevent the fermentation material from sticking to the pipeline valve and the tank wall. Valves and caps for draining condensate.
sampling
Samples can be taken through the sampling port after the inoculation is completed or when sampling is to be carried out in the middle of the fermentation. Before sampling, the sampling pipeline valve should be steam sterilized to prevent miscellaneous bacteria contamination, and the sampling pipeline and valve should also be flushed with steam after sampling.
Operation method: Fully open steam valve Q18, slightly open Q20, when the time reaches 15 minutes, ignite alcohol cotton around the sampling port, close Q20, close Q18, open Q19, open Q20, first discharge the liquid in the sampling tube After that, open the prepared sample bottle on the side of the flame, quickly sample and cover it, close Q20, then open the Q18 cleaning/sterilization sampling port for 5 minutes, and close Q20 and Q18. After the sampling is completed, the PH value of the fermentation broth should be detected by the pH meter, and then the pH should be corrected. When the system is inoculated and the pressure and flow rate are adjusted, the system can enter the fermentation mode for automatic control (temperature, PH, DO). The process parameters are set by the operator himself, and the system will be automatically controlled or manually controlled by the operator according to the set parameters.
Fermentation culture
After the sampling is completed, the PH value of the fermentation broth should be detected by the pH meter, and then the pH should be corrected; when the system is inoculated, the pressure and flow rate can be adjusted, and the system can enter the fermentation mode for automatic control (temperature, PH, DO). The specific process parameters are set according to the production requirements. The system will automatically control or manually control according to the set parameters. The seed is transferred to the seed tank or the fermenter and then controlled by temperature. The tank pressure is generally 0.03-0.05Mpa. The rotation speed is determined according to the process requirements. The temperature of the circulating water is adjusted to control the fermentation temperature. When the ambient temperature is higher than the fermentation temperature, the temperature is lowered by the condensed water. When the temperature of the medium is lower than the fermentation temperature, it is heated with hot water. The PH and DO adjustments are realized by the control system through automatic addition and subtraction by the actuator. The foam alarm is detected by the foam probe and the foam signal is displayed on the touch screen as an indicator light.
Note:
1) The pH electrode should be calibrated before sterilization. The pH control during the culture process is achieved by adding acid and alkali to the peristaltic pump. The acid bottle alkali bottle is first sterilized in the sterilizer.
2) Measurement and control of dissolved oxygen (DO) Calibration of dissolved oxygen electrode: The dissolved oxygen DO value at the constant fermentation temperature before the inoculating, the rotational oxygen and the air amount are turned to the maximum value as 100%.
Dissolved oxygen measurement and control during culture: The control of DO value is achieved by adjusting the air flow rate and adjusting the rotational speed to control the correlation between the rotational speed and dissolved oxygen. Secondly, the intake air amount (manual) control must be adjusted at the same time.
Discharge
The discharge of the device is to discharge the fermentation liquid from the discharge pipe by using the tank pressure, and the tank pressure can be controlled at 0.05 to 0.1 MPa according to the concentration of the fermentation liquid.
After the discharge, take out the dissolved oxygen and pH meter for cleaning and maintenance.
After the discharge is completed, the fermenter and the material pipeline pipe valve should be immediately drained, and the air compressor should be started, and the fermenter should be supplied with gas and stirred to rinse the fermentation liquid in the pipeline.
If the fermenter is not used temporarily, the fermenter is emptied and the water in the tank, jacket and pipe is drained.
α- GPC is a water-soluble substance with two fatty acids removed from the main phospholipid PC (phosphatidylcholine) constituting the cell membrane.α- GPC is a water-soluble substance with two fatty acids removed from the main phospholipid PC (phosphatidylcholine) constituting the cell membrane.
Originally widely existing in organisms, it is one of the body components existing in human breast milk and body fluids. Research in the food industry shows that GPC can be obtained by removing fatty acids (deacyl) in the structure with PC in soybean lecithin as raw material. In the revision of food and drug classification in 2009 α- GPC (sn-glycerol-3-choline phosphate) is defined as food.
Purpose: brain health food. It is clinically used to prevent and treat atherosclerosis, fatty liver, neurasthenia and malnutrition. It can also be used as an additive for food, cosmetics and pharmaceutical excipients. It can be transformed into hemolytic lecithin by phospholipase a.
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