Air sterilization

Air sterilization, Methods of sterilization, depth filter, Membrane cartridge filters, sterilization of culture media

1. R. NITHYA, M. Sc., M. Phil ., Ph. D ASSISTANT PROFESSOR DEPARTMENT OF BIOTECHNOLOGY SRI ADI CHUNCHNAGIRI WOMENS COLLEGE, CUMBUM THENI DT, TAMIL NADU AIR STERILIZATION

2. STERILIZATION OF AIR Industrial fermentation are carried out under vigorous and continuous aeration. For an effective fermentation, the air should be completely sterile. Free from all microorganisms and suspended particles Quantity of suspended particles in air = 20 - 10,000/m3 Microorganisms may range from 10 - 2000/m3 . Among Microbes in air, fungal spores(50%) and Gram negative bacteria (40%) dominate.

3. METHODS OF STERILIZATION Air or other gases can be sterilized by Filtration Heat UV radiation gas scrubbing Heat and filtration are most commonly used. Sterilization by Heat Air was passed over electrically heated elements and sterilized Expensive method

4. STERILIZATION BY HEAT

5. STERILIZATION BY FILTRATION Most commonly used method in fermentation industries. Depth filters Membrane cartridge filters DEPTH FILTERS Air is passed through a glass wool containing depth filters the particles are trapped and removed

6. DEPTH FILTERS

7. DEPTH FILTERS Filtration primarily involves physical effects inertia blocking gravity electrostatic attraction Diffusion Glass wool filters are subjected to seam sterilization and reused. Limitation During steam sterilization the glass wool shrinks and solidifies . Nowadays glass fiber cartridges are being used

8. GLASS FIBER SHEET

9. MEMBRANE CARTRIDGE FILTERS These are removable pleated membrane filters made up of cellulose ester nylon or polysulfone Smaller in size Simpler for operation and replacement

10. MEMBRANE CARTRIDGE FILTERS

11. LIMITATION OF AIR STERILIZATION There is no filter that can remove bacteriophages . Bacteriophages are capable of crippling the industrial fermentation. E.g. Bacteriophages interfere in the production of glutamic acid by Corynebacterium glutamicum

12. STERILIZATION OF CULTURE MEDIA The constituents of culture media, water and containers contribute to contamination by vegetative cells and spores. The media must be free from contamination. Sterilization is done by Applying heat Physical methods Chemical treatment Radiation

13. Heat sterilization - Most widely used technique. - Vegetative cells are destroyed at lower temperature in a short time - around 60 C in 5 -10 minutes. - Spores are destructed at higher temperature. It required longer time ( around 80 C for15-20 minutes ). -Spores of Bacillus stearothermophilus are the most heat resistant.

14. Heat sterilization

15. PHYSICAL METHODS The physical methods such as Filtration Centrifugation and Adsorption are in use. Filtration Certain components like Vitamins, blood components, antibiotics of culture media are heat labile. These components are completely dissolve and then subjected to filter sterilization.

16. FILTRATION

17. LIMITATIONS Application of high pressure infiltration is unsuitable for industries. Some media components may be lost from the media during filtration Combination of filtration and heat sterilization are applied sometimes. For example Water used for media preparation is filtered. Concentrated nutrient solution is subjected to heat sterilization. Then the filtered water is used for dilution of the media.

18. CHEMICAL METHODS AND RADIATION These chemical methods ( using disinfectants ) and radiation methods ( using UV rays, Y rays, X rays ) are not commonly used for media sterilization

19. BATCH STERILIZATION The culture media are subjected to sterilization at 121 C in batch volumes in the bioreactor. Two methods Direct method – Sterilization is done by injecting steam into the medium. The steam should be pure and free from all chemical additives Indirect method - Injecting the steam into interior coils

20. DISADVANTAGES OF BATCH STERILZATION Damage to culture media Alteration in nutrients, change in pH and discolouration of culture media are common. High energy consumption Sterilization takes few hours (2- 4 hrs) for the entire contents of the bioreactor to attain the requisite temperature (i.e. 120 C). Another 20- 60 minutes for the actual sterilization. Then cooling taken 1 – 2 hrs. All this process involve the wastage of energy.

21. CONTINUOUS STERILIZATION - Carried out at 140 C for a very short period of time ranging from 30 – 120 seconds. - The time required is shorter at higher temperature. - It is carried out by directly injecting the steam or by means of heat exchangers. - In both cases the temperature is very quickly raised to 140 C and maintained for 30 – 120 seconds.

22. Different stages in continuous sterilization process in relation to temperature are Exchanger, Heater, Heat maintenance unit, Recovery of residual heat, Cooling and Fermenter.

23. HEAT EXCHANGERS Three types of heat exchangers are used. First exchanger - raises temperature to 90 - 120 C within 20-30 seconds Second exchanger - raises temperature to 140 C and Maintains for 30 -120 seconds. Third exchanger - brings down the temperature by Cooling in the next 20 - 30 seconds.

24. Advantages of continuous sterilization Main advantage is that about 80- 90 % of the energy is conserved. Disadvantage Certain compounds in the medium precipitate due to high temperature. e.g. calcium phosphate, calcium oxalate

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