The concept of sterilisation
"Sterilisation is any chemical or physical process that results in the elimination of any microbiotic living form, whether pathogenic or not, including fungal spores."
Sterile material
"A material can be considered sterile is the time when the probability of finding a microorganism on it is less than one in a million."
Over time, with the advancement of technology and new scientific knowledge, the concept of sterilisation has gradually changed. With the new knowledge came the realisation that the result of sterility was determined by the way in which materials were prepared prior to sterilisation operations, and last but not least, by the effective operation of the machines that were used.
In the light of all this, it can be said that sterilisation is the result obtained at the end of a procedure that uses advanced technology to be carried out in the best possible way; in this way it is able to guarantee a condition such that the survival of micro-organisms is highly unlikely.
Unfortunately, statistics cannot state the absolute certainty of sterility, which is why it is absolutely essential to define codified procedures, which are used in the treatment of the material in order to minimise any possible risk of contamination or alteration of the final result.
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Sterilitarisation methods
Sterilitarisation can be performed by physical means, using heat or radiation.
Thermal methods (heat)
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Thermal methods are based on the transfer of heat from a fluid (which can be air or water) to the objects that are to be sterilised. In this procedure, the heat denatures the proteins of the micro-organisms and thus kills them.
How effective the thermal medium is depends on the amount of heat that can be transferred to the micro-organisms, more precisely on the temperature of the fluid used; the specific heat of the fluid itself; and the latent heat that may be released as a result of the change of state.
When heat is used, sterilisation can be by dry heat or moist heat.
Sterilisation is based on the transfer of heat from the fluid (which may be air or water) to the objects to be sterilised.
Damp heat sterilisation in an autoclave
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The moist heat sterilisation method is a technique that uses saturated steam, in which microorganisms are eliminated through the denaturation of their proteins and other biomolecules.
Autoclave sterilisation is in fact the most widely used method, as it is very cost-effective, has no toxic effects and has a good penetration capacity.
Autoclave sterilisation operation
The operation of the autoclave is similar to that of a pressure cooker. We all know that water boils at 100 degrees with a pressure of 760 mmHg; the autoclave increases the pressure and in doing so the water boils at temperatures above the classic 100 degrees. In this way, the autoclave exploits this principle to achieve the destruction of microorganisms at a significantly higher temperature in relatively less time.
The mechanism of how autoclave sterilisation takes place is actually very simple: air is removed from a container that is hermetically sealed, this operation allows the water in it to evaporate, as a consequence, since the steam cannot disperse, since the container is hermetically sealed, there is an increase in pressure in the container itself; this increased pressure causes the temperature at which the water evaporates to increase with respect to the normal boiling point. For sterilisation to be effective, it is essential that the steam penetrates all parts of the materials it is sterilising, remaining in contact with these for a certain time, ensuring that no air pockets are present. Bacteria and virus spores are eliminated after 30 minutes that have been exposed to saturated steam at a temperature of 130 degrees.
We must always remember that in addition to temperature, another factor to consider is the time of exposure to steam not water. The higher the temperature, the shorter the exposure time required.
Reference standard
The European Technical Commission has determined through the study of leading experts in microbiology and sterilisation, a document prEN 13060 that defines 3 classes of sterilisation cycle:
- Type B ( where B stands for Big sterilizers) with this cycle you have the sterilisation and drying of all types of loads defined by the mornings;
- Type N ( where N stands for Naked) with this cycle you can sterilise all solid materials except for hollow materials that do not have to be bagged, in this way no storage of instruments is allowed;
- Type S ( where S stands for specified) is a cycle which is in the middle between Type B and Type N, the manufacturer must declare what type of loads can be sterilised, specifying whether this cycle is capable of drying the loads they have declared.
Load categories
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Based on resistance to exposure to saturated steam, material categories have been defined:
- solids without hollow spaces;
- type A hollow solids, are those solids that have deep, narrow hollow spaces;
- type B hollow solids, are those solids that have shallow, hollow spaces;
- porous solids, are those particularly complicated loads that have the ability to retain air before the sterilisation cycle and retain moisture afterwards.
The simplest materials to sterilise are the unwrapped solids, while the more complex ones are the packed porous materials.
Classes of Laboratory Autoclaves
Autoclaves are divided into classes according to their ability to handle different material loads.
Class B autoclaves
Also known as fractionated vacuum autoclaves, they have the capacity to sterilise any type of material in the load; the vacuum pump performs several vacuum strokes, thus guaranteeing complete elimination of air.
There are two types of fractionated vacuum autoclaves on the market:
- autoclaves with direct value reproduction;
- autoclaves with steam generator.
Class N autoclaves
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also known as thermodynamic, they are so called because the evacuation of air from the sterilisation chamber is carried out by means of a thermodynamic device, with these autoclaves the air in the turbines, contra-angles and bagged instruments is not guaranteed;
S-class autoclaves
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also known as pre-vacuum autoclaves, in these the air is removed using a vacuum pump; performance is superior to thermodynamic, but not sufficient to guarantee proper air evacuation;
Thermal sterilisation methods (radiation)
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UV sterilisation (ultraviolet radiation)
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UV sterilisation has a high sterilising power, however, it has a low penetration capacity of <1 mm, which makes its application rather limited. The light that kills microorganisms is generated by a mercury vapour lamp and this is emitted at a wavelength of 254nm.
UV rays can be absorbed by many materials such as, glass, plastic, turbid liquids. Unfortunately, they are not able to sterilise powders or solutions, or rather, the possibility is very low, which is why they are used to sterilise air or surfaces in rooms that need to be sterilised.
The sterilisation occurs through the creation of reactive species (called radicals) within the bacterial cell, thus changing the fundamental biochemical reactions, which cause microorganisms to die or be unable to reproduce. Again, the effect is related to the intensity of the radiation and the exposure time. The major contraindication for operators is the possibility of serious eye damage.
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Microwave sterilisation
Microwave sterilisation is used in many areas due to its ease of execution and low cost. The operation of microwave systems is based on two effects, thermal and non-thermal.
Thermal effect microwave sterilisation comes from the ability to create very fast molecular vibrations, which in turn create an increase in temperature that leads to an alteration in the vital and functional capacities of the microorganisms.
Non-thermal effect microwave sterilisation, on the other hand, is determined by the energy that is carried by electromagnetic waves and that is transferred to the matter that is affected. In this way, certain amino acids are transformed, ions are transported and affect metabolism, electrical signals are altered, all of which leads to an acceleration of DNA synthesis that alters RNA transcription, resulting in chromosome aberrations. The effect described so far is very important, because it has been shown that microbial reduction is greater if, at the same temperature reached, there is also a non-thermal effect.
This type of non-thermal microwave sterilisation is compatible with all materials, glass, plastic, metal, rubber, ceramics and does not alter their quality.
In addition, the sterilising action is obtained in a very short time, from 90 to 150 seconds, but safety is achieved with 4-minute cycles.