Sterilization

sterile means devoid of life.

Medications that we inject into people should be sterile. Removing the health hazard due to pathogenic organisms is called disinfection.

aseptic means uncontaminated

When a system has organisms but only those that are supposed to be present, the situation is aseptic. In the past, industrial bioprocess could tolerate a very few foreign organisms relative to the desired organisms. Costs of achieving absolute sterility was not justified for processes such as the penicillin fermentation where the production culture is hardy and will outcompete a few foreign organisms and will usually develop a high titer of product anyway. The situation is different today with cell cultures because they grow slowly and are very easily contaminated.

Mistakes in sterilization means lost batches (very seldom reworked) that must go to disposal.
The investment in these bad batches and the cost for waste treatment have provided sufficient incentives for perfecting sterilization, and most companies with good manufacturing practices do not have major headaches with regard to sterilization of bioprocesses that employ microorganisms. As has been pointed out several times, cell cultures are a different story where avoiding contamination is an endless battle.

After achieving sterility, asepsis must be maintained. Equipment that was fully satisfactory for years for easier bioprocesses was nearly worthless for cell cultures because of frequent contamination. The types of valves, pumps, and connections that were fine for antibiotic processes had to be changed. One common modification was elimination of threaded connections or flanges by welding the pipes and tubes.
 

How to mount agitator


European bioreactors often have agitator shafts entering below the liquid level. While steam can be injected into the seals around the shafts to provide a barrier to the entry of foreign organisms, most U.S. companies use top-entry for the agitation as shown in this sketch. Top-entry means long shafts and more severe problems with bearings than with short shafts entering closer to the impeller blades, the seals have no direct contact with the culture fluid and deal with organisms in air instead of in liquid.  Review of seals for shafts

Usually, a fermenter and its medium are sterilized together, and all connecting lines are flushed with steam. A fermenter may be sterilized and then filled with sterile medium. If so, it is common practice to sterilize water and insensitive materials in the fermenter so as to minimize the volume that must be sterilized by other methods such as filtration or passage through a continuous sterilizer. Among the several factors that influence killing are temperature, pH, osmotic pressure, shear, mass transport, and concentrations of extraneous substances that also react with the killing agent. These factors operate synergistically, and temperature plays roles other than simply affecting the kinetics of a reaction.

The following Java applet shows heating and cooling curves for a large fermenter. Note the time above the RED line. This is where you get killing of microorganisms but at the same time destroy much of the heat-sensitive nutritional value. The time at the flat, high temperature determines whether the contents become sterile, but the times above the RED line at temperatures below the flat, high temperature are very bad because of damage to the nutrients.


  • Kinetics of sterilization,
  • Batch sterilization game,
  • Demo of why solids are bad,
  • Continuous Sterilization of Media,
  • Sterilization of Air
  • Thermal pollution will not be on the quiz but provides perspectives for biochemical engineers.