Sterile Products

 Sterile products are dosage forms of

therapeutic agents that are free from viable
microorganisms.
 Sterility is an absolute term, means the
absence of living microorganisms.
1. Microbe is a microscopic organisms such as
a bacteria, fungus, protozoa or virus.
2. Pyrogens are metabolic by-products of live
or dead microorganisms that cause a pyretic
response (i.e., a fever) upon injection.
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 There are three terms used interchangeably to
describe these products—Parenteral, Sterile,
and Injectable.
 Parenteral and injectable basically have the
same meaning and are used interchangeably.
 Sterile dosage forms encompass
parenteral/injectable dosage forms as well as
other sterile products such as topical
ophthalmic products, irrigating solutions,
wound-healing products, and devices.
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 Definition
 A product introduced in a manner that
circumvents the body’s most protective barriers,
the skin and mucous membranes, and, therefore,
must be “essentially free” of biological
contamination.
 Modifier words “essentially free” added to this
definition because most small-volume (≤100 mL per
container) sterile products are produced where the
finished product is not terminally sterilized, but rather
is aseptically processed.
 Difference in sterility assurance is far greater
(generally at least 3 logs) for terminally sterilized
products compared to aseptically processed
products. 4
 A dosage form is said to be sterile when it is free from
(1)Microorganisms (2)Spores (3)Pyrogens &
(4)Pathogens
 Pharmaceutical dosage forms that are sterile include;
1. Parenterals
2. Topical Opthalmics
3. Topical wound healing medications
4. Irrigation fluids
5. Sterile devices
6. Pulmonary drug delivery (Not all)
 Sterile product compounding requires cleaner facilities,
personnel training and testing, as well as a sound of
knowledge of sterilization and stability principles and
practices.
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 1- Parenterals
 Parenteral (Gk, para enteron, beside the intestine)
dosage forms differ from all other drug dosage forms,
because they are injected directly into body tissue
through the primary protective systems of the human
body, the skin & mucous membranes.
 Must be exceptionally pure and free from physical,
chemical, and biological contaminants.
 These requirements place heavy responsibility on
pharmaceutical industry to practice current good
manufacturing practices (cGMPs) in manufacture of
parenteral dosage forms & on pharmacists and other
health care professionals to practice good aseptic
practices (GAPs) in dispensing parenteral dosage forms
for administration to patients.
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 As these Bypass the highly efficient first line of body
defense – skin & mucous membranes, they must be;
i. Free from microbial contamination.
ii. Free from toxic components.
iii. Possess exceptionally high level of purity.
 Components and processes involved in preparation of
these products must be selected & designed to
eliminate as much as possible contaminations of all
types like;
i. Physical origin.
ii. Chemical origin.
iii. Microbiological origin.
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 Preparations must be sterile to enter veins
and arteries, otherwise anaphylactic shock
or other abnormalities may result.
 Sterility: Complete destruction of all living
organisms & their spores or their complete
removal from formulation.
 Aseptic Technique: Technique for
preparation and manipulation of compounded
sterile products and parenteral preparations
that prevents contamination.
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 Contamination: Any effect or action that has
a negative impact on a product's integrity
making it unfit for use;
i. Chemical composition
ii. pH
iii. Sterility (microorganism contamination)
iv. Pyrogenicity
v. Physical appearance
vi. Particulate matter (e.g. dust, glass or
precipitation)

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 2- Ophthalmic Preparations
 Preparations for eye, although not
introduced into the internal body cavities,
but are placed with the tissues which are
extremely sensitive to contamination.
 Due to this reason, similar standards are
required for ophthalmic preparations as for
the parenterals.

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 3- Irrigating Solutions
 Irrigating solutions are also required to meet
the same standards as parenteral solutions,
b/c during an irrigation procedure,
substantial amounts of these solutions can
enter the bloodstream directly through;
i. Open blood vessels of wounds.
ii. Abraded mucous membranes.

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 4- Pulmonary Drug Delivery
 Previously Nasal/pulmonary administration of drugs
was primarily employed for local drug effects.
 Potential nasal route for systemic delivery was
discovered after observation that nasally
administered sympathomimetic & antihistaminic drug
for local action has significant systemic effects.
 Nasally administered small dose display a rapid
absorption that is comparable to intravenously
administered drugs.
 Drugs can be administered by pulmonary route
utilizing the techniques of aerosol inhalation and
intratracheal instillation or Insufflation.
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 Types
 Sterile products are most frequently;
1. Solutions ready for injection
2. Dry, soluble products ready to be combined with a
solvent just prior to use
3. Suspensions ready for injection
4. Dry, insoluble products ready to be combined
with a vehicle just prior to use
5. Emulsions
6. Liquid concentrates ready for dilution prior to
administration
7. Solid pellets for tissue implantations (not
common).
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 Control of a process to minimize
contamination for a small quantity of sterile
product can be achieved easily.
 As the quantity of product increases,
problems of controlling the process to prevent
contamination go on multiplying.
 Preparation of sterile products has become a
highly specialized area in Pharmaceutical
industry which requires a superior level of;
i. Established Standards
ii. Attitude of Personnel
iii. Process control
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 The organizational divisions normally
responsible for the preparation of sterile
products in the Pharmaceutical Industry
are;
1. Product Development
2. Production
3. Q.C.
4. Packaging

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 Particulate
 Simply airborne particles are solids suspended in
the air.
 A body having finite mass and internal
structure; a minute portion, piece, fragment,
or amount; a tiny or very small bit; a grain,
speck.
 Particulate matter in injections and parenteral
infusions consists of mobile undissolved particles,
other than gas bubbles, unintentionally present in
the solutions.
 Size of contaminants and particles are usually
described in microns. 18
 Air, whether it is from outside or re-
circulated, acts as a vehicle for bacterial and
gaseous contaminants brought in by the
movement of people, material, equipment
etc.
 Since many of these air borne contaminants
are harmful to products & people, their
removal is necessary on medical, legal,
social or financial grounds.

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 PM implies an ideal; a solid or collection of solids,
observed as a single solid. Solid PM or a collection of solids
are certainly the most prevalent nature of PM;
 However, many other and alternate conditions (nonideal) may
be considered as PM by the observer and may affect the
quality of the formulation.

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 Sources of Particles
 There are two main sources of particulates.
1. External sources
2. Internal sources
1- External sources
 These consist of the following:
i. Outside air introduced into the room.
• This is typically the largest source of external
particulates.
ii. Infiltration through doors, windows and other
penetrations through the clean room barriers.

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Control Action
i. Make-up air filtration
ii. Room pressurization
iii. Sealing of all penetrations into the space.
2- Internal sources
 Internal sources consist of the following:
i. People in the clean area – people are potentially
the largest source of internally generated
particulates (Touch contamination & Generation of
particulates from shedding cells or hair)
ii. Clean room surface shedding
iii. Process equipment
iv. Material and ingredients
v. Manufacturing processes

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 People are a major source of contamination in cleanroom. People
activities producing number of particles per minute are presented below;

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Control Action
 Design airflow path to shield humans from
surroundings.
 Use of air showers [to continually wash
occupants with clean air].
 Using hard-surfaced, non-porous materials such
as polyvinyl panels, epoxy painted walls, & glass
board ceilings.
 Proper gowning procedures, head wear mask
etc.
 A super clean environment with controlled
temperature & relative humidity has now become
an essential requirement for a wide range of
applications in Pharmaceutical Plants.
 More critical in case of Sterile products. 25
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Types of Sterile Products Processing
1) Terminally sterilised
 → Prepared, filled and sterilised

2) Sterilised by filtration

3) Aseptic preparation

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 1. Terminally sterilised
 Usually involves filling & sealing product
containers under high-quality environmental
conditions.
 Products are filled & sealed in this type of
environment to minimize microbial & particulate
content of in-process product & to help ensure that
subsequent sterilization process is successful.
 In most cases, the product, container & closure
have low bio-burden, but they are not sterile.
 Product in its final container is then subjected to a
sterilization process such as heat or irradiation.

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 2. Sterilisation by Filtration
 Previously sterilized container are taken.
 Filters having nominal pore size 0.22 μm or less
are used for filtration.
 Remove bacteria and moulds but Not viruses &
Mycoplasmas.
 Double filter layer or second filtration is required
sometime.
 No fibre shedding should be permitted or asbestos
filters.
 Filter integrity testing is required.

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 3. Aseptic Preparation
 In an aseptic process, drug product, container &
closure are first subjected to sterilization methods
separately, as appropriate, and then brought together.
 As there is no process to sterilize product in its final
container, it is critical that containers be filled &
sealed in an extremely high-quality environment.
 Before aseptic assembly into a final product, individual
parts of the final product are generally subjected to
various sterilization processes.
 Any manual or mechanical manipulation of sterilized
drug, components, containers, or closures prior to
or during aseptic assembly poses risk of contamination
and thus necessitates careful control.
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 Clean rooms
 Cleanrooms provide for the control of airborne
contamination to levels appropriate for
accomplishing contamination-sensitive activities.
 Following are the fields where clean rooms are
applicable;
1. – Aerospace
2. – Microelectronics
3. – Pharmaceuticals
4. – Medical devices
5. – Healthcare (Hospitals)
6. – Food
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 Clean rooms
 Production of sterile preparations should be carried out
in clean areas, entry to which should be through
airlocks for personnel and/or for goods.
 Clean areas should be maintained to an appropriate
standard of cleanliness & supplied with air that has
passed through filters of an appropriate efficiency.
 Various operations of component preparation,
product preparation, filling & sterilization should be
carried out in separate areas within a clean area.
 Clean areas for production of sterile products are
classified according to the required characteristics of
the air, in grades A, B, C & D or Classes 100, 10,000
& 100,000.
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 A clean room is defined as a room in which the
concentration of airborne particles is controlled.
 Clean rooms have a defined environmental control of
particulate & microbial contamination, and are
constructed, maintained, and used in such a way
as to minimize introduction, generation & retention
of contaminants inside the room & and in which other
relevant parameters e.g., temperature, humidity,
and pressure, are controlled as necessary.

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 Clean room Classification
 Clean room classifications are established by
measurement of number of particles of 0.5 micron
& larger that are contained in 1 ft³ 0r 1 m³ of
sampled air.
 Level of airborne particulate cleanliness represents
maximum allowable concentrations (in particles
per cubic metre/fit of air) for considered sizes of
particles.
 Generally class 100 to 100,000 rooms are used in
the pharmaceutical industry.
 Rooms may be classified as clean as class 1 or 10
for other applications, particularly in the microchip or
semiconductor industry. 37
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 Particles & Occupancy states
Particle: Solid or liquid object which, for purposes of
classification of air cleanliness, falls within a threshold size
in the range from 0.1 to 5μm.
As built: condition where installation is complete with all
services connected & functioning but with no production
equipment, materials, or personnel present.
At rest: condition where installation is complete with
equipment installed & operation in a manner agreed
upon by customer & supplier, but with no personnel
present.
Operational: condition where installation is functioning
in the specified manner, with the specified number of
personnel and working in the manner agreed upon.

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 “As-built” condition “At-rest” condition

“Operational” condition
ISO Air Classification System
 ex

WHO GMP US 209E US Customary ISO/TC (209) EEC GMP

ISO 14644
Grade A M 3.5 Class 100 ISO 5 Grade A
Grade B M 3.5 Class 100 ISO 5 Grade B
Grade C M 5.5 Class 10 000 ISO 7 Grade C
Grade D M 6.5 Class 100 000 ISO 8 Grade D
WHO Technical Report Series, No. 902, 2002
Annex 6
 Manufacturing Environment
Classification of Clean Areas
 Classified in terms of airborne particles (Table)

“At rest” - Production equipment installed and operating

“In operation” - Installed equipment functioning in defined
operating mode and specified number of personnel present
 Four Grades of Clean Areas
 Grade D (equivalent to Class 100,000, ISO 8): Clean area
for carrying out less critical stages in manufacture of
aseptically prepared products e.g. handling of components
after washing.
 Grade C (equivalent to Class 10,000, ISO 7): Clean area for
carrying out less critical stages in manufacture of aseptically
prepared products e.g. preparation of solutions to be filtered.
 Grade B (equivalent to Class 100, ISO 5): Background
environment for Grade A zone, e.g. Clean room in which
laminar flow workstation is housed.
 Grade A (equivalent to Class 100 (US Federal Standard
209E), ISO 5 (ISO 14644-1): Local zone for high risk
operations e.g. product filling, stopper bowls, open vials,
handling sterile materials, aseptic connections, transfer of
partially stoppered containers to be lyophilized.
 Conditions usually provided by laminar air flow workstation. 44
 Limits for Microbial Contamination
 Each grade of cleanroom has specifications for viable and
non-viable particles
1. Non-viable particles are defined by the air
classification
2. Limits for viable particles (microbiological
contamination) are given below;
Grade Air sample Settle plates (90mm Contact plates Glove print
(CFU/m3) diameter) (55mm (5 fingers)
(CFU/4hours) diameter) (CFU/glove)
(CFU/plate)
A <1 <1 <1 <1
B 10 5 5 5
C 100 50 25 -
D 200 100 50 -
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 All pharmaceutical facilities belong to one or
other class of clean room.
 General acceptance is:

A. Tabletting facilities - Class 100,000

B. Topical & oral liquids - Class 10,000
C. Injectables class - Class 100

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 Critical Area (Grade A)
1. Processing area where sterilized products and
materials as well as their surfaces are directly
exposed to the environment (The local zone for
high risk operations).
 Environmental conditions should be specified to be
suitable for the virtual elimination of contamination
risks & preservation of the sterility of products.
 Processes conducted in this area are sterilization
activities (sterile connections, addition of sterile
materials) Filling zone, open ampoules and vials,
making aseptic connections.
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 Critical Area (Grade A)
 Provided by a LAF work station with a
homogeneous air speed in a range of 0.36 – 0.54
m/s (guidance value). 0.45m/s ± 20%
 A unidirectional air flow and lower velocities may be
used in closed isolators and glove boxes.
2. Per-cubic-meter content of particles ≥ 0.5 μm in
diameter in the critical area should be below 3,500
under both operating & non-operating conditions.
 This level of air cleanliness is designated as Grade
A, Class 100, M 3.5 or ISO-5 according to
domestic & international standards on air quality.

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3. Intervention of personnel into the critical area
should always be kept to a minimum.
4. Count of airborne particles & microorganisms
should be regularly monitored by appropriate
procedures at sites which are critical for ensuring
sterility of pharmaceutical products.
5. Powder filling operations may generate higher
counts of airborne particles than the specifications.
 Count of airborne particles should be obtained by
sampling air at different locations or monitoring the
count in same room while no powder filling operation
is going, & causes of the deviation should be
identified to maintain air quality in the room at a
required level.
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Direct Support Area (Grade B)
1. Direct support area is a working area for
personnel who operate machines installed in
the critical area & for those who supervise the
operation of machines.
 For aseptic preparation and filling, this is the
background environment for the grade A zone.
 This area also serves as a route for the transfer of
sterilized products, materials, and equipment to
the critical area or for moving sterilized products
from the critical area.
 Appropriate measures need to be implemented to
protect sterilized products or materials from direct
exposure to the environment. 52
2. The per-cubic-meter count of particles (diameter: ≥
0.5 μm) in the direct support area should be
controlled below 350,000 and 3,500 under
operating and non-operating conditions,
respectively.
 These levels of air cleanliness are designated as
Grade B, Class 100, M 3.5 or ISO-5.
3. The count of airborne particles and microorganisms
should be regularly monitored by appropriate
procedures in the direct support area.
 The frequency and method of monitoring should be
carefully selected based on evaluation results of
product contamination risks in the critical area.
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Indirect Support Areas (Grade C or D)
1. Indirect support area is an area used for
processing materials & products prior to
sterilization & hence materials & products are
directly exposed to environment.
 Clean areas for carrying out less critical stages in
the manufacture of sterile products
 Example for indirect support areas include an area
for preparing drug solution prior to sterilization
and an area for washing & cleaning sterilization
equipment & apparatuses.
2. Cleanliness of indirect support area needs to be
controlled by establishing specifications for
acceptable airborne particle count by taking into
account the required level of contamination control
and type of works performed in the area. 54
3. Air cleanliness of indirect support area may be either of
the following two grades.
a) One of the grades specifies that per-cubic-meter particle
content (diameter: ≥ 0.5μm) should not exceed
3,500,000 & 350,000 under operating & non-operating
conditions, respectively.
These levels of cleanliness are designated as Grade C,
Class 10,000 M 5.5 or ISO-7 (standard under operating
conditions) according to domestic and international
standards on air quality.
b) Other grade specifies that per-cubic-meter particle
content (diameter: ≥ 0.5μm) should not exceed
3,500,000 under non-operating conditions. This level of
cleanliness is designated as Grade D, Class 100,000 M
6.5 or ISO-8. 55
4. Weighing & preparation processes should
preferably be conducted in Grade C or cleaner
areas.
 If powder handling might elevate the airborne
particle count above the specification, air quality
should be maintained below the specification by
accurately determining the particle count that
may cause contamination in the area, and for the
determination, air should be sampled, for
example, at multiple locations and/or under
powder-free conditions.

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Type 1: Unidirectional (Laminar flow)

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Type 2: Non-unidirectional flow (Turbulent flow)

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 Types of Manufacture of Sterile
Preparations
 Manufacturing operations are divided into 3
categories:
1. In which preparation is sealed in its final container
and terminally sterilized.
2. Preparation is sterilized by filtration.
3. Preparation can be sterilized neither by filtration nor
terminally & consequently must be produced from
sterile starting materials in an aseptic way.
 In Terminally sterilized products solutions should
generally be prepared in a grade C environment in
order to give low microbial and particulate counts,
suitable for immediate filtration and sterilization.
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 Solution preparation could be allowed in grade D
environment if additional measures are taken to
minimize contamination, such as use of closed
vessels.
 For parenterals, filling should be done in a laminar
airflow workstation (grade A) in a grade C
environment.
 Preparation of other sterile products, e.g.,
ointments, creams, suspensions & emulsions, &
filling of containers should generally be done in
grade C environment before terminal sterilization.
 For other sterile filtered products prepared from
sterile materials in an aseptic way the handling of
starting materials & all further processing should be
done in a grade A or B area with a grade B or C
background respectively. 65
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