How does a steam autoclave work?

E. coli

Steam autoclaves are designed to use high pressure and high temperature steam to kill microorganisms. They are also used to render biohazardous materials inactive. For the steam autoclave to be effective, the materials to be sterilized must be saturated with steam. If there isn't enough steam or air pockets exist in the materials, decontamination will not take place.

Sterilization involves absolute destruction of all microorganisms including bacteria that are resistant, bacterial spores, protozoans, prions, viruses and fungi that are present in fluids, on surfaces of materials, in medication or culture media. Absolute destruction means just that. There is no such thing as “partial sterilization”. To achieve a sterilized state can be difficult as well as be difficult to prove. Under a number circumstances, steam sterilization is the preferred method of sterilization and works by denaturing proteins. Denaturing proteins involves protein coagulation that comes about by a change in protein conformation in the presence of heat. Chemicals have the ability to kill microorganisms, however, they may not be totally effective and may leave undesirable or potentially toxic residues behind. Nevertheless, chemical sterilants are used when heat would damage materials. Ionizing radiation and UV can be used and they disrupt or modify DNA to prevent replication, however, they may not produce the desired effect and validation is a bit tricky. Ultimately, moist heat sterilization tends to be superior and the preferred method for sterilization.

Most microorganisms are killed at temperatures above 80 degrees C. Prions on the other hand, require higher temperatures and longer times to deactivate them. In the process of moist heat sterilization, steam molecules condense on cooler microorganisms. The steam molecules then transfer 2500 joules per gram of steam heating the microorganisms to a temperature that they will be killed. Other heating methods that transfer lower heat of dry gases tend to create boundary layer effects. This ultimately creates an insulation effect which ends up protecting microorganisms.

To achieve the maximum effect of steam, it must be saturated. With
moist heat sterilization procedures, the temperature and pressure can easily be monitored making it easy to determine if sterilization has occurred. There is no doubt that steam sterilization produces a high level of sterility and that's why it is the most often used form of sterilization in hospitals and laboratories. Moist heat autoclaving is the fastest and most reliable form of sterilization.


How the Steam Autoclave Works

Essentially an autoclave is a pressure cooker that uses steam under pressure as its sterilizing agent. The increase in pressure (above atmospheric pressure) enables steam to reach higher temperatures. The extra pressure brings the boiling temperature of water higher. In fact, around 20 degrees C higher. This effectively increases its heat content and killing ability. This comes from its latent heat of evaporation. The latent heat of evaporation is the amount of heat that is required to convert boiling water to steam. This conversion heat is quite large compared to the amount of heat that is required to make water hot.

To get one liter of water to boil requires 80 kcal/mol, however, converting boiling water to steam requires 540 kcal/mol. That means that steam at 100 degrees C has almost 7 times more heat than boiling water.

The nice property about steam is that is has the ability to penetrate objects that have a cooler temperature. As the steam encounters a cooler surface, it condenses into water on that object. This effectively decreases the amount of steam present. This negative pressure in steam draws more steam to the cooler area. Condensation will continue to occur so long as the temperature of the condensing surface is less than that of steam. This allows for rapid heating of surfaces, penetration of dense materials, denaturing proteins and microorganism sterilization.


Thermal Death Time

Thermal death time (TDT) is used to determine how long it takes to kill specific microorganisms at a specific temperature and specific suspension. Let it suffice to say that death rate is directly proportional to the concentration of microorganisms at any given time. If you increase the temperature of your sterilization process, this has the effect of decreasing thermal death time. Lowering the temperature increases thermal death time. Higher temperatures for shorter periods of time are preferred.

Beyond temperature and time, thermal death time is affected by the materials being sterilized. For example, oily materials slow down steam penetration and therefore increase thermal death time. Materials that are highly acidic or basic tend to decrease thermal death times. Thermal death times are available for a number of microorganisms and specific suspensions they may be in. Keep in mind that thermal death time may not be accurate but a good place to start.

Autoclaving is accepted as being the most effective and most efficient way of sterilization. Moist heat autoclaves work on a time and temperature relationship. Higher temperatures are important for more rapid killing of microorganisms. Longer sterilization times are required for larger loads, large liquid volumes, and dense materials. Temperatures and pressures most often used in a moist heat autoclave are 115 degrees C at 10 psi (pounds per square inch), 121 degrees C at 15 psi and 132 degrees C at 27 psi. Moist heat autoclaving works well for glassware, biological media, surgical dressings, biohazardous waste and much more.


Autoclaves: Pressure and Temperature

Pressure and temperature affect boiling. Water boils when the water molecules contains enough energy to escape the liquid and form water vapor or what is called steam above it. As the water gets hotter, water molecules contain more energy and can more easily escape the liquid. On the other hand, pressure is important as well. The higher the pressure above the water, the more difficult it is for the water molecules to break free of the liquid and vice versa.

Consider going up in altitude. The higher you go, the lower the atmospheric pressure. Boiling water at this higher elevation occurs at a lower temperature because it takes less heat to boil the water. It is much easier for liquid water to go into water vapor because the pressure above the liquid is much less. In fact, if you were on top of Mount Everest, water boils at around 70 degrees C not 100 degrees C.

The boiling point of water at one atmosphere (sea level, 760mmHg) is 100 degrees C. Raising or lowering the pressure by around 28mmHg changes the boiling point by 1 degree C.


Vapor Pressure

In the case of a closed container such as an autoclave, the process of evaporation will continue until there are as many molecules returning to the liquid as there are escaping. When this occurs, the vapor is said to be saturated. At higher temperatures, more water molecules can escape and the corresponding saturated vapor pressure is greater. In the case of an open container, vapor pressure is actually a partial pressure along with the other components in the air above. In this case, the temperature at which the vapor pressure is equal to the atmospheric pressure is referred to as the boiling point.


Summary

Steam autoclaves use high pressure and high temperature steam to kill pathogens. In order for the steam autoclave to be effective, the materials to be sterilized must be saturated with steam. Sterilization refers to absolute destruction of all microorganisms that are present in fluids, on surfaces of materials, in medication and culture media. There is no such thing as “partial sterilization”. Steam sterilization works by denaturing proteins. Denaturing proteins involves protein coagulation which is a change in the conformation of the protein rendering it inactive.

Basically the steam autoclave is a pressure cooker that uses steam under pressure as its sterilizing agent. The increase in pressure above one atmosphere allows steam to reach a higher temperature. The extra pressure brings the boiling temperature of water higher bringing about more killing power.

Thermal death time is information used to determine how long it takes to kill specific microorganisms at a specified temperature. The death rate is directly proportional to the amount of microorganisms present at any given time.

Moist heat autoclaving is the most often used form of sterilization. It is the fastest and most reliable form of sterilization.


Further Readings

The World of Autoclaves

http://esf.uvm.edu/uvmsafety/labsafety/biosafety/autoclaves/autoclaves.html

Hyperphysics and Thermodynamics

http://hyperphysics.phy-astr.gsu.edu/Hbase/Kinetic/vappre.html#c2

The Logarithmic Nature of Thermal Death Time Curves

http://www.jstor.org/stable/30082910

Enzymatic detergent treatment protocol that reduces protease-resistant prion protein load and infectivity from surgical-steel monofilaments contaminated with a human-derived prion strain

http://vir.sgmjournals.org/content/88/10/2905.full

Photo Credits and Information

http://commons.wikimedia.org/wiki/File:Diverse_e_Coli.png

description: English: These are E. coli

date: 4 August 2009

Source: Own work

Author Mattosaurus

I, the copyright holder of this work, release this work into the public domain. This applies worldwide.
In some countries this may not be legally possible; if so:
I grant anyone the right to use this work for any purpose, without any conditions, unless such conditions are required by law.


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