ChE 473 Lecture - 21-24 Biosafety and Sterilization PDF

ChE 473: Biochemical Eng I; Lecture 21-24 May 2017
Dept of Chem Eng, BUET

ChE 473: Lecture 21-24
ChE 473: Biochemical Eng I

Biosafety and Sterilization
Lecture 21-24
May 2017
Course Teacher:
Dr. Mohidus Samad Khan
www.mohidkhan.com 1
References
Russell, Hugo & Ayliffe’s Principles and

Practice of Disinfection, Preservation &
Sterilization, 5th Ed, 2013, Willey-Blackwell.
Kolhe, Shah and Rathore (2013), Sterile

Product Development – Formulation,
Process, Quality and Regulatory
Considerations.
Class Lectures and Reading Materials

Course: ChE 473. Dept of Chem Eng, BUET.  Dr. Mohidus Samad Khan 2016 ChE 473: Lecture 21-24
© Dr. Mohidus Samad Khan. 2017 1

Biosafety Laboratory

Course: ChE 473. Dept of Chem Eng, BUET.  Dr. Mohidus Samad Khan 2016 ChE 473: Lecture 21-24 3
• Biosafety Level 1 Lab (BSL 1) :
– i) Work with agents with Little Potential Hazard to lab personnel and
the environment
– ii) Using Fume Hoods and Biosafety Cabinets are preferred
– iii) Contaminated samples need to be disposed
– iv) similar to a high school biology lab
– i) Work with agents with Moderate Potential Hazard to lab personnel
and the environment
– ii) 100% air filtration required
– iii) Contaminated samples need to be disposed
– iv) Access to the laboratory is limited to the trained personnel.


– i) BSL 3 is required to work on Infectious Disease Research
– ii-iv) BSL2 requirements
– v) Continuous Power supply.
– Biosafety Level 4 Lab (BSL 4) :

– i) Work with Dangerous Agents that pose a high risk of Aerosol
Transmitted Infections
– ii-v) BSL 3 requirements.

Cleanroom Standard
Ref: Wikipedia: Cleanroom
Video Demonstration : (8:02 min)

Semiconductor Technology at TSMC, Taiwan

Cleanroom Requirements
• Active Air Monitoring:
– microbes in the air in the clean areas that may be sampled using
devices such as slit-to-agar or surface air system samplers.
• Surface Monitoring:
– contact or touch plates or swabs that can sample surfaces such as
walls, floors, or
• Passive Air Monitoring:

– bacterial culture plates (“settle plates”) exposed to the environment
allowing any airborne bacteria above the plate to fall onto it.

Sterilization


Sterilization
• “Sterilization” is an absolute term meaning the ability to
destroy all life forms. Sterilization, by its very nature,
damages or alters the materials subjected to the process.
• Relevant Sectors
• Medical
• Pharmaceutical
• Biotechnology
• Food
• Circuits, etc.

Sterilization
• Heat Sterilization
• Moist Heat/Steam Sterilization
• Dry Heat Sterilization
• Gas/Chemical Sterilization
• Alkylating Agents
• Oxidizing Agents
• Filtration Sterilization
• Filtration of Liquid, Solid or Gas
• Irradiation Sterilization
• Gamma Radiation
• E-Beam,
• Beta Radiation.
• Gas Plasma Sterilization

Heat Sterilization

Heat Sterilization
• Heat is now the major sterilization
method in use worldwide employing
pressurized steam.
• Heat has also been used in combination
with other microbicidal agents, including
chemicals and ionizing radiation.
• Aim - To deliver sufficient heat to
inactivate all of the organisms that might
be present
• Critical Factors – Temperature, Pressure
and Time

Heat Sterilization: Techniques

• Moist Heat/Stem Sterilization -
• The heat from the steam kills microorganisms, and
the effectiveness of the sterilization depends both
on the amount of time the items in the load are
exposed to the steam and the efficiency of steam
penetration into all product surfaces and spaces.
• Dry Heat Sterilization -

• Dry heat sterilization may be accomplished
using ovens specially designed to handle
components of pharmaceuticals.

Steam Sterilization: Autoclave
14

Steam Sterilization: Autoclave

Autoclave: Cycles
http://www.rpi.edu/dept/chem-eng/Biotech-
Environ/Projects00/sterilize/batch.html
http://www.intertechequip.com/DRY_
HEAT_STERILIZER.html


Steam Sterilization: Types

Steam Sterilization: Uses
18

Kinetics of Heat Inactivation
http://academic.pgcc.edu/~kroberts/Lecture/Chapter%209/heat.html
The decimal reduction time (D) represents the time it takes
for 90% of a population of microbes to be killed by a
particular method of microbial control, such as autoclaving.

Kinetics cont’d
At Constant
Temperature


Kinetics cont’d
• Factors to Characterize Thermal Inactivation
• D-value - The D-value (decimal reduction time (DRT)), is defined as the
time in minutes at a particular constant temperature to reduce the viable
population by 1-log10, that is to 10% of the initial value, or by 90%.
• z-value - The z-value is defined as the temperature (°C) to bring about a
10-fold reduction in D-value; it is obtained from the slope of the curve in
which the D-value on a logarithmic scale is plotted against temperature
on an arithmetic scale.
• F-value - The F-value expresses a heat treatment at any temperature as
equivalent to that effect produced by a certain number of minutes at
121°C; F0 is the F-value when z is 10°C.
Kinetics cont’d


Steam Sterilization: T-P Relationship
23
Thermal Death Kinetics of Microorganisms
N = Number of viable organism

k = Thermal death rate constant
A = Arrhenius Constant
E = Activated Energy

Thermal Death Kinetics Cont’d
http://www.rpi.edu/dept/chem-eng/Biotech-
Environ/Projects00/sterilize/batch.html
Dry Heat Sterilizer

• Sterilization by dry heat is less efficient than by
moist heat.
• Definitions of D-value and z-value given for moist
heat above apply equally here.
• The F-value concept utilized in steam-sterilization
processes has an equivalent in dry-heat
sterilization, although its application has been
limited.
• Dry heat as a means of sterilization is reserved for
those products and materials that contain little or
no water and cannot be saturated with steam
during the heating cycle.
• A hot-air oven is usually employed to effect dry-
heat sterilization.

Dry Heat Sterilizer Cont’d

Dry Heat Sterilizer Cont’d
http://www.outpatientsurgery.net/surgical-facility-administration/infection-
control/infection-prevention--01-02


Thermal Processing of Foods

• Thermal processing of high pH (low acid) and
high water activity (Aw) foods - Pasteurization
• Destroy or inactivate microorganisms
• Improve shelf-life
• Heat Sterilization: Combination Treatment
• Heat Sterilization + Acidification
• Heat Sterilization + Chill Storage
• Heat Sterilization + High Hydrostatic Pressure.
• Etc.

Filtration Sterilization


Filtration Sterilization
• Old technique: Cloth Filtration: 400 BC; Sand Filtration: 3-4 century AD
• Can be used for Liquid, Solid and Gas
• Filtration Media: Key Characteristics
• Efficient removal of particles above a stated size;
• Acceptably high flow rate;
• Resistance to clogging;
• Steam Sterilisable;
• Flexibility and mechanical strength;
• Low potential to release fibers or chemicals into the filtrate;
• Low potential to sorb materials from the liquids being sterilized;
• Non-pyrogenic; and
• Biologically inert.

Cells: Dimension
Ref: click4biology.info; 2013

Relative sizes:
1. Molecules (1nm).
2. Cell membrane thickness (10nm).
3. Virus (100nm).
4. Bacteria (1000nm).
5. Organelles (less 10,000nm).
6. Cells (<0.10 mm).

Figure: Staphyllococcus aureus retained on the surface of a

0.45 micrometer cast Durapore™® membrane
Filter Devices
Table: Filter Device Properties


Filter Devices: Performance

• Log Reduction Value (LRV)
• C = Concentration of Microorganisms
(CFU/ml)
• V = Liquid Volume
• Higher LRVs mean larger
reductions in bioburden and
higher filtrate sterility
assurance.

Membrane Filters: Flow Direction
Figure: Comparison between (a) normal and (b) cross-flow filtration.


Membrane Filters: Pore Size Distribution
Figure: Typical pore size distribution curves for 0.45 m-rated cellulose
membranes obtained from mercury intrusion tests.

Membrane Filters: Properties of Polymers


Gas Sterilization

Gas Sterilization
• Evolution of Gas Sterilization
• Chlorine Gas (sulfuric acid + sea salt) to
remove odor from Cemetry (18th
Century)
• Chlorine gas produced from
Chlorhydric Acid
• Disinfecting Hospitals with
Formaldehyde (19th Century)
• Modern Days:
• Gaseous Sterilization of Hospitals with
Ethylene Oxide
• Low temperature sterilization based on
Hydrogen Peroxide, Peracetic Acid,
Ozone, and Chlorine Oxide

Properties of Gaseous Sterilants

Gas Sterilization: Formaldehyde


Gas Sterilization: Hydrogen Peroxide

Gas Sterilization: Comparison


Irradiation Sterilization

Irradiation Sterilization
• Irradiation, or Ionization Radiation,
is a type of ‘Cold’ Sterilization
• Radiation Sterilization:
• Gamma Radiation
• E-Beam,
• Beta Radiation.
• Sources:
• Gamma Radiation (Cobalt 60 (60Co))
• Electron Beam (E-Beam, high energy
electrons)


Irradiation Sterilization: Units
47
End


ChE 473 Lecture - 21-24 Biosafety and Sterilization PDF

Încărcat de

Informații document

Titlu original

Drepturi de autor

Formate disponibile

Partajați acest document

Partajați sau inserați document

Opțiuni de partajare

Vi se pare util acest document?

Este necorespunzător acest conținut?

Drepturi de autor:

Formate disponibile

ChE 473 Lecture - 21-24 Biosafety and Sterilization PDF

Încărcat de

Drepturi de autor:

Formate disponibile

ChE 473: Biochemical Eng I; Lecture 21-24 May 2017

Dept of Chem Eng, BUET

ChE 473: Biochemical Eng I

Russell, Hugo & Ayliffe’s Principles and

Kolhe, Shah and Rathore (2013), Sterile

Class Lectures and Reading Materials

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 1

Dept of Chem Eng, BUET

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 2

– Biosafety Level 4 Lab (BSL 4) :

Dept of Chem Eng, BUET

Video Demonstration : (8:02 min)

© Dr. Mohidus Samad Khan. 2017 3

• Passive Air Monitoring:

Dept of Chem Eng, BUET

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 4

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 5

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 6

Heat Sterilization: Techniques

• Dry Heat Sterilization -

Dept of Chem Eng, BUET

Steam Sterilization: Autoclave

© Dr. Mohidus Samad Khan. 2017 7

Steam Sterilization: Autoclave

Dept of Chem Eng, BUET

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 8

Steam Sterilization: Types

Dept of Chem Eng, BUET

Steam Sterilization: Uses

© Dr. Mohidus Samad Khan. 2017 9

Kinetics of Heat Inactivation

Dept of Chem Eng, BUET

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 10

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 11

Steam Sterilization: T-P Relationship

Thermal Death Kinetics of Microorganisms

N = Number of viable organism

© Dr. Mohidus Samad Khan. 2017 12

Thermal Death Kinetics Cont’d

Dry Heat Sterilizer

© Dr. Mohidus Samad Khan. 2017 13

Dry Heat Sterilizer Cont’d

Dept of Chem Eng, BUET

Dry Heat Sterilizer Cont’d

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 14

Thermal Processing of Foods

Dept of Chem Eng, BUET

Dept of Chem Eng, BUET

© Dr. Mohidus Samad Khan. 2017 15

Dept of Chem Eng, BUET

Ref: click4biology.info; 2013

© Dr. Mohidus Samad Khan. 2017 16

Figure: Staphyllococcus aureus retained on the surface of a

Dept of Chem Eng, BUET