Chapter 7: STERILIZATION

Introduction Sterilization is the process to eliminate microorganisms from a medium or equipment. It is used to: Presterilize equipments Sterilization of feedstocks Maintenance of sterile operating conditions

Batch Sterilization Uses steam to eliminate living organisms. Heat losses, heating and cooling are major steps and is a time consuming process. Air is evacuated and replaced with steam. It wastes energy and overcooks the medium. There is no conservation of energy, therefore not economical for implementation in large scale. Done in an autoclave. When the pressure from the steam has been built up, a venting valve for the outlet of air chamber is closed and the inlet valve allows the steam to enter the chamber. The pressure of the chamber is increased to 105 kPa (15 psig). At this point, the sterilization time begins to count down. Sterilization of 1 L media at 121°C requires 20 min. Larger volumes may require longer retention time.

Autoclave

Continuous sterilization Direct use of live steam by injection into the medium. Heat exchanger is eliminated. The medium stays in a loop for a predetermined holding time until the entire medium is sterile. The problem with directly injecting steam is dilution of media since it is initially cold. However it has better heat economy since it substitutes heat exchangers. It has a holding coil for detention long enough to kill all the microorganisms. The medium from a make-up vessel flowing through the exchanger is held in the coil, and then goes through the heat exchanger, heating more unsterile medium while becoming cool itself, as it is collected in a sterile bioreactor.

Continuous sterilization

Dry heat sterilization Used for equipments that can withstand high temperature and dry heat but cannot withstand wet or steam autoclave. Often used for glasswares as it dries and sterilizes in one operation. Pipettes must be wrapped in dust proof aluminium foil or placed in metal pipette cans. Disposable items are not recommended for dry heat sterilization.

Sterilization with Filtration Certain media components are susceptible to heat. It denatures if heated. Therefore they must be added to the media after autoclaving. To do so, filtration is necessary using a 0.22 μm pore size filter that is appropriate to the solvent used.

Chemical Sterilization Chemical agents are used to sterilize heat-sensitive equipments. Application of an oxidizing agent agent such as 10% chlorox for 20 min or longer proves the system free from contamination. Excess amounts of chemical agents have to be removed, otherwise organisms won’t be able to grow under toxic environment.