air & media sterilization
TRANSCRIPT
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Presentation on Sterilization of air & media
By: neitho-o Roll: 080217Sem: 7th Dept: Biotechnology
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Sterilization • Term referring to any process that eliminates
(removes) or kills all life forms of microbial life including transmissible agents (fungi, bacteria, viruses, spore forms etc.) present on a surface, contained in a fluid, in medication, or in a compound such as biological culture media
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Sterilization of air• Heat treatment• Ultraviolet light treatment• Filter sterilization• Ozone gas treatment• Chemical treatment
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Heat treatment1.Dry heat sterilization
2. Moist heat sterilization
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Dry heat sterilization
• Circulation of heated air within the chamber of the oven can be of two processes:
a. Gravity convection process: Produces inconsistent temperatures within the chamber and has a very slow turn over.
b. Mechanical convection process: the oven contains a blower that actively forces heated air throughout the chamber. It ensures uniform temperatures and the equal transfer of heat throughout the load.
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Moist heat sterilization
Processes/steps:1.Chamber closed and heated so that steam
forces air out of the vents or exhausts.2. Pressure applied so that the interior
temperature reaches 121 oc3.The temperature maintained for between 15-
30 minutes.
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UV-light treatment• Short wave length(254nm)• Destroys the nucleic acids (disruption of DNA)
of microorganisms resulting in reproductive incapability and dead.
• Used in medical sanitation and sterile work facilities
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Advantages of UV-light treatment
• It is rapid• Low cost • Does not need special operator training
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Disadvantages of UV-light treatment
• Must be operated in a closed system• Needs careful monitoring
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Filter sterilization of air
• Most commonly used sterilization process• fixed pore filter membrane (0.2-0.3
micrometer)widely used• PTFE(hydrophobic) most common
construction membrane used
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Sterilization of fermenter exhaust air
• Fixed pore membrane modules are also used• Pretreatment of exhaust gas is necessary
before it enters the absolute filter.• Pretreated air is then fed to a 0.2 micrometer
hydrophobic filter.
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Basic principles of filter design• Reduction of particle entering the filter is: dN/dx = -KN….(1)Where, N =concentration of particles entering the filter K = constantIntegrating (1) over the length of filterN/No = e-kx ……(2)Where, No = no. of particles entering the filter N = no.of particles leaving the filterTaking log, (2) becomesln(N/No ) = -kx……..(3) { log penetration relationship}Efficiency of filter , E = (No - N)/ No Or, E= 1 - e-kx
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Ozone gas treatment
• Ozone gas is most efficient, very reactive and it can decompose back to oxygen without leaving a trail.
• broad- spectrum• Concentrations of 0.05-0.08 ppm. required
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Advantages of ozone gas treatment
• Faster sterilization• No requirement of chemicals• Improved shelf life of products.• Higher preservation duration
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Media sterilization
Media may be sterilized by filtration , radiation, ultrasonic treatment, chemical treatment or
heat.
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Kinetics of sterilization• Destruction of micro-organisms by steam (moist ) may be described by 1st order
chemical reaction:-dN/dT= kN………….(1)Where, N=no. of viable organism present,t = time of sterilization treatmentK=reaction rate constant of the reaction/specific death rate.Integration of (1)Nt /N0= e-kt …………(2)Where N0= no. of viable organisms present at the start of sterilization treatment Nt = no.of viable organisms present after a treatment period t.Taking log,ln(Nt /N0) = -kt ………(3)
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Relationship between T & k(reaction rate constant)
Arrhenius equationd ln k/dT = E/RT2 -------(4)Where, E = activation energyR= gas constantT= absolute temperatureIntegration of (4)K= Ae -E/RT ----------(5) where A=Arrhenius constantTaking natural log in (5) ln k = ln A – E/RT --------(6)
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Heat sterilization of a pure culture at constant temperature
Combination of eqn (3) & (5) ln(Nt /N0) = A*t*e -E/RT .........(7)
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Del factor ( )or Nabla factor (i.e.,Nt /N0)
= measure of fractional reduction in viable organism count produced by a certain heat and time regime .
= ln(No /Nt) since Kt = A*T*e (-E/RT) = ln(No /Nt)
so, = A*t*e (-E/RT) ………… (8) rearranging (8)
ln t = E/RT + ln ( /A )
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The same degree of sterilization may result from treatment at a higher temperature for a short time as from a low temperature for a long time.
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Deleterious effect of increasing medium sterilization on the yield of product
Destruction of essential media due to heat: Xt /x0 = e-kt
Where, xt = concn. of nutrient after a heat treatment period,t x0 = original concn. of nutrient at the onset of sterilizationk = reactn. rate constant
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Two types of reaction contributing to the loss of nutrient quality during sterilization
• Interactions between nutrient components of the medium
• Degradation of heat liable components.
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Design of Batch Sterilization Process
Information needed for the design:1. Temperature of the fermentation medium2. Number of micro-organism originally present in the
medium.3. Thermal death characteristics of the ‘design
organism’(example Bacillus stearothermophilus)
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Calculation of Del factor in Batch culture
overall= heating+ holding+ cooling i.e.,
=
A*T*e (-E/RT)
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sum of the values of Del factor corresponding to each time increment is equal to the overall Del factor
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Methods of batch sterilization
• Fermentation vessel • Separate Mash cooker
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Design of continuous sterilization processes
• Approach is same as batch sterilization processes
• Includes a time period during which the medium is heated to the sterilization temperature, a holding time at the desired temperature and a cooling period to restore the medium to the fermentation temperature
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Eg. Spiral heat exchanger
• The plant is sterilized prior to sterilization of the medium by circulating hot water through the plant in a closed circuit
• The fermenter and the pipe work between the fermenter and the sterilizer are steam sterilized.
• In coming unsterile medium partially heated before reaching the sterilizer
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Filter sterilization of fermentation media
• Essential/suitable for heat-labile proteins(animal-cell culture)
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Advantages of continuous sterilization over batch sterilization
• Superior maintenance of medium quality• Ease of scale up• Easier automatic control • Reduction of sterilization cycle time.
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Advantages of batch sterilization over continuous sterilization
• Lower capital equipment costs • Lower risk of contamination • Easier manual control• Easier to use with media containing a high
proportion of solid matter.
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Reference : 1. Principles of fermentation technology by P.F. STANBURY A. WHITAKER & S.J.HALL 2. www.creative ozone.com 3. sterilization of air by R.C telling & J .W.S. Ford
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Thank you