The 5,000-litre fermentation tank must be cleaned before use: remove the electrodes from the fermentation tank, select the appropriate setting, and clean the tank.

Check for airtightness: open the valve (exhaust valve) and keep it open throughout the experiment; switch on the main power supply, turn on the steam generator water supply to fill the steam generator, and switch off the water supply when the red light goes out; Switch on the air compressor to fill the air receiver to a pressure of 2 kg (0.2 MPa), then switch off the air compressor. Introduce a measured quantity of deionised water into the fermenter via the inlet port. Open the fermenter’s inlet valve, then close it. Turn the fermenter’s sampling knob to the ‘nozzle’ position to prevent leakage. If the pressure remains stable for 30 minutes, the airtightness is satisfactory.

Sterilisation and air purging of the fermentation tank: After 30 minutes, open the exhaust valve and adjust the pressure to stabilise at 0.03 MPa; before opening the steam generator valve, first open the steam outlet valve on the fermentation tank; open the steam generator valve and the fermentation tank inlet valve, then close the inlet valve to prevent reverse suction. When the temperature reaches 121°C, maintain this for 30 minutes, ensuring the pressure inside the tank remains at a specific level (excessive pressure is hazardous, whilst insufficient pressure makes it difficult to raise the temperature). At this point, the fermentation tank is ready for use; commence the fermentation process. After 30 minutes, release the pressure; close the steam generator valve and the fermentation tank’s steam inlet valve, then open the inlet valve (to prevent tank deformation).

Procedure for using a 5,000-litre fermentation tank:

Calibrate the pH electrode and dissolved oxygen electrode for the 5,000-litre fermentation tank.

Sterilise the 5,000-litre fermentation tank. Add the culture medium to the tank as required, seal it according to specifications, and place small fermentation tanks (5 L) into a large sterilisation tank for sterilisation (115°C, 30 minutes); steam sterilisation (121°C, 30 minutes) is used for large fermentation tanks (10 L and above).

Once the 5,000-litre fermenter has cooled, place it on the fermentation bench and ensure it is properly installed; turn on the cooling water, switch on the air pump, connect the ventilation tubing to commence aeration, and adjust the air inlet knob to achieve an appropriate airflow rate, maintaining a tank pressure of 0.05 MPa; set the temperature, pH, stirring speed, etc., and calibrate the oxygen electrode slope to 100% at the specified speed and airflow rate.

Once the 5,000-litre fermenter has reached a stable temperature and all parameters are correct, connect the pre-shaken seed culture, start the fermentation timer, and begin recording the various parameters.

After fermentation, clean the vessel and electrodes. Insert the pH electrode into a 3M potassium chloride conical flask for storage, and cover the dissolved oxygen electrode probe with a protective cap for future use.