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EU GMP Annex 1 - The new draft and implications for sterile product
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Technical Report · April 2020

DOI: 10.13140/RG.2.2.14468.68487

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EU GMP Annex 1: The New Draft and the
Implications for Sterile Products Manufacturing

White Paper

Author:
Dr Tim Sandle

1
Abstract
EU GMP Annex 1 is the primary document governing the manufacture, control and release of
sterile pharmaceutical products (both terminally sterilised and aseptically filled medicines).
The Annex will shortly undergo a comprehensive update, as signalled by a new draft
issued in February 2020. This white paper assesses many of the key points within the draft,
focusing on those areas that have a direct impact upon sterile product manufacturing.

Contents
Introduction 3

What is EU GMP Annex 1? 3

Key Changes 4

New Structure 4

Contamination Control Strategy 4

Centrality of the Pharmaceutical Quality System 5

Premises 5

Personnel 8

Microbiological Environmental Monitoring 9

Quality Control 10

Filtration 10

Product Inspection 10

Cleaning and Disinfection 11

Utilities 12

Summary 12

References 13

2
Introduction
Annex 1 of EudraLex “The Rules Governing Medicinal Products in the European Union” forms part of Volume 4 of the
European guidelines covering the manufacture and release of sterile products, which are subject to special
requirements (1).

The Annex is going through a revision process, with the final version expected in 2020. This review process began with
the issuing of a draft in December 2017. This draft was subject to over 6,000 comments submitted by professional
organisations. After a review of these, across a two-year period, the European Medicines Agency (EMA), issued a
new draft (described as ‘version 12’) (2). The new draft addresses some of the issues in the first version and contains
some additional requirements. The scope for the review of version 12 is more limited, with specific areas marked for
consultation and the group of professional bodies invited to comment has been narrowed. This signals that much of the
draft content will be in the final version, and that the final version will be issued within a relatively short timeframe.

The trigger points for the revision to the Annex included emergence of new technologies, and what regulatory
inspectors have highlighted as inadequate root cause analysis and ineffective CAPA. Other points of regulatory concern
included poor implementation of ICHQ9 (which relates to quality risk management) (3). A further reason for revision
was to address various points of ambiguity in the interpretation of the Annex 1 requirements.

This white paper provides an overview of the draft Annex 1 and considers the focal points for sterile products
manufacture, control and release.

What is EU GMP Annex 1?

There are two major, global guidance documents for
sterile products manufacture: the FDA guidance, last
revised in 2004 (4), and Annex 1 of EU GMP. Annex 1 of
EudraLex “The Rules Governing Medicinal Products in the
European Union” forms part of Volume 4 of the European
guidelines (1) (there is also a WHO guidance. The purpose
of these guidances, is to detail the controls required
around the manufacture of sterile products. Special
requirements are needed in order to minimise risks of
microbiological, particulate and pyrogen contamination
of sterile products; and also, to provide guidance as to
how sterile products are best protected. Annex 1 includes
key areas like personnel training, equipment qualification,
cleanroom design and environmental monitoring.
The scope of Annex 1 relates to pharmaceutical
companies who manufacture products within the
European Union and those companies who import
products into the European Union (including a post-
Brexit United Kingdom).
Annex 1 of EU GMP has undergone no major revision
since 2007 and no change whatsoever since 2009
(where there was a minor point of clarification about the
required air supply grading for oversealing - Grade A air
supply). The lack of an update through the intervening
years was especially notable in the context of updates to
cleanroom technology and the appearance of new types
of rapid microbiological methods. As part of the revision
process, the current title of “Annex 1: Manufacture of
Sterile Medicinal Products” is now tweaked to read
“Annex 1: Manufacture of Sterile Products”.

3
Key Changes
There are five broad areas of change to the draft, in terms
of tone and emphasis. These are reflected at various
intervals in the document (often through repeated
occurrences). These areas are:
1. The global acceptance and implementation of ICH Q9
(Quality Risk Management) and Q10 (Pharmaceutical
Quality System), is not reflected in the current Annex.
The new draft contains many references to Quality Risk
Management (QRM) in particular, emphasising that
QRM should be used as a proactive tool.
2. There is the requirement for a formal, holistic
contamination control strategy. The expectation is for a
formal document which reflects the site-wide strategy
for minimising contamination control with respect to
sterile manufacturing. Such a strategy, as outlined below,
is for each organisation to fully-understand and review
design, procedural, technical and organisational controls.
3. There have been advances in sterile manufacturing
technology, especially with RABS and isolators. There
have also been advances with rapid microbiological
methods, which the draft Annex acknowledges
4. There was some ambiguity with the current version
and these needed correction or clarification
5. Annex 1 is often beyond sterile manufacturing,
including aspects of non-sterile manufacturing.
The scope of the new draft has been modified and
broadened to reflect this.
6. In particular the references to Quality Risk Management
are more extensive, with requirement to use QRM to
review existing products and processes; new products
and processes (where risk assessment needs to link
to Quality by Design); and to address problems with
procedures and processes, with the expectation that
QRM becomes a key part of deviation management.
This reflects a significant shift in regulatory thinking
towards a more risk-centric approach.
New Structure
The new draft is divided into eleven parts, which are
presented across 52 pages (which means that it is two
pages longer than the 2017 version and considerably
more than the current Annex’s 16 pages).
1. Scope
2. Principle
3. Pharmaceutical Quality System (PQS)
4. Premises
5. Equipment
6. Utilities
7. Personnel
8. Production and Specific Technologies
9. Viable and Non-viable Environmental and
Process Monitoring
10. Quality Control (QC)
11. Glossary
4
Contamination Control
Strategy
A substantial part of the draft Annex is given over to
each organisation having a detailed, facility-specific
contamination control strategy (abbreviated to CCS). To
be effective this needs to be an approach that can assess
seemingly isolated contamination events holistically and
which is capable of putting appropriate corrective and
preventive actions (CAPA) in place. This requirement
is intended to signal a new paradigm in terms of
contamination control, shifting the risk review process to
one that assesses the impact of a contamination event in
a far wider context.
The main elements of such a control strategy are:
• Design of both the plant and process
• Equipment and facilities
• Personnel
• Utilities
• Raw materials control – including in-process controls
• Product containers and closures
• Vendor approval – such as key component suppliers,
sterilisation of components and single use systems,
and services
• For outsourced services, such as sterilisation,
sufficient evidence should be provided to the
contract giver to ensure the process is operating
correctly
• Process risk assessment
• Process validation
• Preventative maintenance – maintaining equipment
and premises (planned and unplanned maintenance)
to a standard that will not add significant risk of
contamination
• Cleaning and disinfection
• Monitoring systems - including an assessment of the
feasibility of the introduction of scientifically sound,
modern methods that optimise the detection of
environmental contamination
• Prevention – trending, investigations, corrective and
preventive actions (CAPA), root cause determination
and the need for more robust investigational tools
• Continuous improvement based on information from
the above systems
With each of these different elements it is notable
that they are not confined to biocontamination, since
reference is also made to subvisible particles (the classic
appearance tests) and the overall appearance of the
pharmaceutical product.
Centrality of the Pharmaceutical Quality System
The first main section of the draft refers to the Pharmaceutical Quality System, a quality system that each manufacturer
should have in place. While general matters are covered in Chapter 1 of EU GMP, the draft Annex makes reference to
specific aspects for sterile products manufacture. These include:

• The proactive use of risk management

• Regular review of risk assessments

• Rationales in place to address different categories of risk arising from risk assessments

• Employing staff with sufficient expertise to undertake risk assessments

• Use of effective root cause and CAPA

• Those tasked with releasing products must be fully conversant with risks and quality issues

• The first two bullet points signal the continuing reference to risk management and risk assessment throughout the
document

The reference to risk assessment should not simply confined to manufacturing, it needs to extend to packaging (primary
and secondary) and to the distribution of the finished medicinal product, thereby embracing the requirements of Good
Distribution Practice (GDP).

Premises

The main changes under the premises section relate to cleanroom design and operation. With the overall design of
cleanrooms, the draft stresses the importance of ‘quality-by-design’, with particular reference made to lay-out and
process flow, where the process steps are reflective of an intent to maintain contamination control. This design concept
extends to the necessity of conducting all non-essential processes outside clean areas. Reference is made to Annex 15 of
the EU GMP guide (“Qualification and Validation”) in relation to ensuring that equipment has been suitably qualified.

The text makes reference to the specific series of tests required when a cleanroom is qualified. These tests are (where
relevant to the design):

• Installed filter leakage and integrity testing • Microbial airborne and surface contamination

• Airflow measurement - Volume and velocity • Temperature measurement

• Air pressure difference measurement (with the new • Relative humidity measurement
draft the specification changes from 10 to 15 pascals
• Recovery testing
to a minimum of 10 Pascals)
• Containment leak testing
• Airflow direction and visualisation

5
Notably here is the task of microbiological monitoring. part of the acceptance of the adjusted volume includes an
This is something that has been discussed in relation to airflow visualisation to be conducted. For isolators that are
the update of the ISO 14698 biocontamination standards, ‘closed’ the Annex states that the air direction need not
about adding a microbiological monitoring aspect to the necessarily be unidirectional.
assessment of cleanrooms.
The revised Annex has added information relating to
With particle assessments, the Annex keeps the requirement environmental controls in relation to pressure. In addition
to count airborne particulates equal to or greater than 0.5 to revising the limit for pressure (as indicated above), there
and 5.0 μm (which continues to create a difference with the is a requirement that all areas with differential pressure
FDA aseptic processing guidance, which refers to particles have warning alarms in place, where the warning signals a
of ≥0.5 μm only). However, for Grade A zone and Grade B potential problem with air supply.
at rest, classification only needs to be for particles equal
Barrier Technology
to or greater than 0.5 μm. The cleanroom classification
expectations apply to the ‘at rest’ and ‘in operation’ states
(with the ‘in operation’ state there is a suggestion that for
aseptic processing areas that the exercise is undertaken
during media fills, in order to represent worst-case). For
selecting particle locations the draft Annex states that the
ISO 14644 methodology is to be followed, along with the
additional expectation that, for aseptic processing areas,
sample locations are positioned so that all critical processing
zones like the point of fill and stopper bowls are included and
based on a documented risk assessment. The Annex does not go as far as mandating the use of
barrier technology; however, there is a recommendation
Whilst there are no changes to requalification intervals
that manufacturers consider adopting “appropriate
(Grades A and B six-monthly and Grade C and D annually)
technologies”, such as Restricted Access Barriers Systems
additional text has been added stipulating that re-
(RABS) or isolators. The recommendation extends to
qualifications should be undertaken following any remedial
consideration of robotic systems, which will reduce the
works needed on equipment or where the facility requires
need for human intervention. With the emphasis upon
work or where new equipment is added to the cleanroom,
barrier technology the Annex requires that any alternative
as assessed through change control. Other reasons for
must be robustly risk assessed.
undertaking a re-qualification include change of room use
and to reassess areas following a loss of power. With isolator technology used for aseptic processing, the
text states that pass through of items should be minimised
Outside of classified cleanrooms, reference is made to
and ideally go through a rapid decontamination chamber.
‘controlled but not classified areas’. Here the movement
Gloves are recognised as the weakest point with an isolator
of material from controlled but not classified to Grade C
system, so consequently there is the requirement for glove
needs to be based on risk management principles, with
leak testing at a minimum interval of before and after
means that the level of cleaning and disinfection and the
each batch (the leak assessment also extends to the main
control of materials needs to be commensurate with the
isolator unit as well). In the text, the frequency of glove leak
level of risk assessed.
testing is detailed as “a minimum at the beginning and end
For the first time the draft Annex provides guidance for of each batch.” Also, with gloves, there is reference to the
Grade D particles, albeit for ‘at rest’ rather than for ‘in importance of selecting the correct isolator gloves; those
operation’. Here a value of 29,000 particles for the ≥0.5 with good mechanical and chemical resistance. The period
μm size particle is provided as the limit. The current Annex of time between the replacement of gloves now requires
simply lists Grade D particles as ‘separately determined’, justifying in the contamination control strategy.
with no further guidance supplied.
For the preparation of isolators for filling, the new draft
In terms of other critical parameters, the draft Annex stresses the importance of cleaning prior to disinfection
offers more scope with unidirectional airflow velocities. (since the presence of residues can inhibit the ability of
With air speed, while the range of 0.36 – 0.54 m/s disinfectants to traverse microbial cell walls, which is
remains the general requirement at working height; necessary for the killing of microorganisms). In keeping
however, there is scope for companies to operate clean with cleaning, the ‘clean hold time’ also requires qualifying.
air devices outside of this range should be designed, There is a requirement to verify that the disinfectant used,
provided that this is scientifically justified and detailed in relation to any remaining residues, does not have an
in the contamination control strategy. With the related adverse impact upon the product.
aspect of airflow visualisation (‘smoke’) studies these
A further change, with the set-up of filling line contained
now need to be conducted in both the ’at rest’ and ‘in
within barrier technology, is that a sporicidal disinfection
operational’ states (previously the reference was just to
process is used following set-up, as mechanism to
‘in operation’). The draft Annex requires that following
reduce any microbial contamination that may have been
an adjustment to air velocities for qualified devices that
presented during the assembly process.

6
Specific points relating to aseptic
processing
The current Annex requires all connections for aseptic
processing (such as vessel to manifold) to be performed
under Grade A. The draft acknowledges advancements
with sterile processing technology, permitting aseptic
connections that use intrinsic sterile connection devices,
designed to minimise any potential contamination from
the immediate environment, to be performed in lower
classified environments provided that the connection
device has been appropriately validated to show no
ingress of microbial contamination.
The control of operation time is discussed in several
places, perhaps with the view that times outside what is
ordinarily seen can lead to an increased contamination
risk. Time is referred to with:
• Pre-fill time should be assessed as part of media fills
• A time limit required for aseptic assembly
• Maximum exposure time of sterilised containers and
closures prior to closure. This infers oversealing times
need to be set. This has an impact with regards to
ensuring crimping is conducted expediently
• For items sterilised “in house” (such as by
autoclaving), these need to be stored in Grade A
or B, using appropriately sealed packaging and a
maximum hold period must be established
• The Annex sets out the main elements for Grade A
processing, which allows manufacturers to risk assess
the essential elements. These are:
• Maintaining the critical processing zone
• The aseptic assembly of filling equipment
• Aseptic connections (these should be sterilised by
steam-in-place whenever feasible).
• Special focus on aseptic compounding and mixing
• The risks around the replenishment of sterile product,
containers and closures
• Concerns around the removal and cooling of items
from heat sterilisers
• Staging and conveying of sterile primary packaging
components
• Aseptic filling, sealing, transfer of open or partially
stoppered vials, including interventions
• Loading and unloading of a lyophiliser changes are designed to strengthen controls around
These elements could be cross-referred back to
the contamination control strategy and implied risk
assessment.
7
For transfer of items and equipment into aseptic
processing areas, there is added emphasis
upon unidirectional airflow and sterilisation or
decontamination. Where possible, items should be
sterilised and passed into the area through double-ended
sterilisers (such as a through a double-door autoclave
or depyrogenation oven/tunnel). Where sterilisation
on transfer of the items is not possible, pass through
hatches equipped with unidirectional airflow and with a
method of transfer disinfection, in place should be used.
With equipment sterilisation, as required for aseptic
processing, the text requires all direct and indirect
contact parts to be sterilised.
Single-use systems are technologies used in manufacture
of sterile products as alternatives to reusable equipment.
The systems are designed to maintain integrity
throughout processing under the intended operational
conditions and their application can help to reduce
contamination risk, e.g. aseptic connections. The use of
single-use systems and technologies continues to be
encouraged within the draft Annex, albeit with the caveat
that the interaction between the product and product
contact surface (notably adsorption, leachable and
extractables) is carefully understood.
Other revisions to the Annex which impact on
operations conducted within cleanrooms include
additional information about media fills (aseptic process
simulations), where more ‘time based’ criteria have been
added (such as assessing filling machine hold times and
sterilised equipment hold times as part of the exercise).
The importance of simulating all events remains (given
that events, especially interventions are more important
for the success or failure of a media fill than simply run
time). There has also been a change to acceptable failure
limits in media fill studies, based on filling run size. The
formally specified statistical approach of >99.9% at 95%
confidence interval is no longer applicable to determine
acceptable failure limits. Instead the expectation is
zero, irrespective of the media fill size. This will create
tighter limits for those manufacturers who have yet to
adopt more stringent, especially for large batch sizes.
A further point with media fill failures is that, following
investigation and root cause analysis, the previous note
about running one media fill has changed to a near
requirement to run three media fills.
Greater detail is provided for assessing the success of
autoclave operations, such as the requirement to inspect
sterilised packaging for its integrity and dryness. Such
sterile products manufacture.
Personnel
The biggest section within the draft is dedicated to
personnel. The section includes a requirement to restrict
the numbers of personnel permitted to enter cleanrooms.
The recommendation is that personnel numbers,
especially in sterile manufacturing areas, is minimised
based on risk assessment. This acknowledges that the
biggest risk of microbial contamination in cleanrooms
derives from people, principally through shedding and by
touching critical surface.
In relation to controlling access to cleanrooms there
is reference made to the importance of on-going
quality assurance oversight. To minimise additional
risk from people, there is a reference to such activities
being completed outside of the cleanroom, such as by
inspection windows or through the use of CCTV.
As expected, training plays a significant part in the
personnel section. This extends to everyone entering the
cleanroom, including cleaning and maintenance staff.
The minimum basis for a training program is set out as
to include: hygiene, cleanroom practices, contamination
control, aseptic techniques, and potential safety
implications to the patient of a loss of product sterility
and in the basic elements of microbiology. For access
into the higher-grade cleanrooms (Grades A and B) to
work on aseptic processing lines, personnel are expected
to have completed aseptic process qualifications (such
as through a media fill). An exception is made for
maintenance staff under exceptional circumstances,
although procedures need to be in place to describe how
this will be controlled.
With qualifying operators, the language of the 2020 draft
refers to ‘qualified’ and ‘unqualified’ operators (it is also
possible for qualified operators to become disqualified
if there are concerns around microbial contamination).
Part of the qualification is a gown test and the movement
through a well-designed cleanroom system that seeks to
minimise contamination. The requirements for entering
8
changing areas for access into Grade B and C areas have
been strengthened. The draft states that outdoor clothing
(other than personal underwear) cannot be brought into
changing rooms. In terms of what is not permitted to be
taken into clean areas, the Annex now calls out mobile
devices (reflecting the ubiquity of smartphones and the
like) unless these have been supplied by the organisation
and are shown to be suitable for cleanroom used, and
covered by a cleaning and disinfection process. With
entry into Grade B changing areas, there needs to be a
separate way in and a separate way out (the requirement
to add this to Grade C areas has been dropped). This is in
order to reduce cross-contamination.
With gowns, the text also requires that:
• Gowns must be suitable to prevent shedding and
the environment in which operators work and the
movements they may be undertaking needs to be
taken into account
• The gown worn needs to be of a suitable size
• Gowns must not affect the product (this is a
reference to the risk of fibres being released from
gowns and ending up in the product)
• That gowns are assessed for their suitability (e.g. no
signs of damage) prior to be being worn and once
donned, prior to entering the cleanroom. There is a
comment that visual inspection may not be sufficient
to assess gown integrity. However, no further advice
is afforded, and this is something for which there is
no obvious solution for, prior to entering a cleanroom
• Gowns must be processed in dedicated laundry
facilities and the process of cleaning, evaluating and
sterilising gowns must be qualified
For cleanroom operators entering aseptic processing
areas a gowning test is required (which is a combination
of visual assessment and microbiological monitoring).
The new draft expands the list of recommended locations
on an operator’s gown that require monitoring as part
of the gowning qualification: hands, arms, chest and
forehead. Each one of these locations presents a different
microbial contamination risk, in terms of the types of
organisms and the route of contamination transfer. In
addition, microbiological limits are presented for gown
plates for the first time (these are afforded the same
maximal values as finger plates). Further with gowning,
the new draft now requires the maximum time that a
gown can be worn for to be defined.
All staff working in cleanrooms are expected to
have knowledge of hygiene, cleanroom practices,
contamination control, aseptic techniques, and potential
Microbiological Environmental Monitoring
For viable monitoring expanded information is provided
in relation to sample site selection, stating that this needs
to be risk based and, where applicable, determined
through a review of airflow visualisation studies.
A change has been made to the EU GMP Grade A limit;
which changes from 1 CFU to ‘no growth’. This change is
both a reflection of the expectation that microorganisms
are not typically recovered from Grade A environments
(and that every recovery requires an investigation) and
with the different types of techniques that could be
applied as replacements to the classic culture-based
methods, such as the use of rapid and automated
microbiological methods are permitted provided the
facility has demonstrated their equivalency or superiority.
9
safety implications to the patient of a loss of product
sterility and in the basic elements of microbiology. As
well as requiring that personnel are suitably qualified
to work in cleanrooms, the new draft of the Annex
states that each facility must have staff who are
specifically experienced in microbiology, environmental
monitoring regime and with conducting microbiological
investigations.
With aseptic operations specifically, the revised draft
places strong emphasis upon how operators are trained,
especially in relation to any interventions with Grade A.
It is stressed that practices must not disrupt Grade A
unidirectional airflow in terms of movement or with the
placing of objects that might cause air disruption. To
aid with operator training a recommendation is made
that airflow visualisation studies constitute part of the
operator training program.
The Annex also sets out the expectation for continuous
monitoring in relation to aseptic processing. This is
mandatory for Grade A and recommended for Grade B.
By continuous monitoring this means air samples (either
settle plates or volumetric air samplers).
Trending is an important consideration and the Annex
provides information about setting alert and action
levels and for looking for data patterns. Where action
level excursions occur, there is a necessity to investigate,
determine root cause and to set alert and action levels.
Trending is not only with numbers of colony forming
units; trending also applies to microbial speciation. For
microbial identification the draft Annex stipulates that all
isolates from Grade A and B areas should be identified,
and recommends that those isolated from Grade C and
D areas are identified. The Annex does not appear to
acknowledge that it is not always possible to obtain
an accurate identification result although the intent is
presumably for the microbiologist to try to obtain a
genus or species level outcome.
To strengthen monitoring, a reference is made to
qualifying some aspects of the technologies used,
including verifying that contact plates are not providing
false counts due to the presence of disinfectant residues.
Most users overcome this through the use of a neutraliser
being added to the agar.
A limitation is placed on monitoring (and testing) being
no substitute for a robust sterility assurance system. With
this there is an emphasis that testing is too imprecise to
detect a weakness with the sterility assurance system.
As the draft states: “Exclusively monitoring or testing
does not give assurance of sterility.” Instead, design and
control matter far more for patient safety.
Quality Control
Microbiological tests also feature in an expanded ‘quality on sampling from the beginning, middle and end of the
control’ section. These tests include the examination of batch, plus following any significant intervention. For
raw materials, where the specification applied should terminally sterilised products the worst case locations
be based on the required microbial quality as defined relate to areas like cold spots; and with lyophilised
in the contamination control strategy. Further tests products, samples need to be taken from different freeze-
include assessing intermediate bioburden of intermediate dryer loads.
product and a bioburden test conducted immediately
before the sterilising filter (for aseptically filled products)
or prior to terminal sterilisation. In both cases, the
samples should be representative of the worst-case
conditions, such as at the end of a process hold time.

The quality control section contains a discussion

around the sterility test. While full limitations with the
sterility test are not discussed in detail, the Annex does
emphasise that the value of the sterility test is diminished
if sterilisation and environmental controls are not in place.
To strengthen the sterility test, in terms of making the
test more representative, the draft text provides guidance

Filtration
Sterile filtration is a critical step for aseptically filled
products, where products are passed through a
μ0.22 μm filter, with pressure and time controlled
(following validation of the filter, where a bacterial
challenge has been used in the presence of product). A
key measure is an assessment of the product bioburden
prior to filtration.
The issue of pre-use post sterilisation integrity testing
(PUPSIT) is set to apply to all large volume products.
For small volume products, a risk assessment can be
used, assessing the filter sterilisation process in terms of
having controls in place ensuring that the risk of damage
to the filter is minimised. There is also a requirement to
understand the supply chain, such as having oversight
of contract sterilisation facilities; modes of transport;
packaging of the sterilised filter, designed to prevent
damage to the filter during transportation and storage.
Further controls around filtration include process
knowledge including the specific product type, including
microbial challenge and whether there exists any risk of
impact on filter integrity values, such as the potential to
alter integrity testing values and therefore prevent the
detection of a non-integral filter during a post-use filter
integrity test. Finally, pre-filtration and processing steps,
prior to the sterilising filter, must be in place to remove a
microbial challenge and clarify the product prior to the
sterile filtration.

Product Inspection
Sterile, medicinal pharmaceutical products need to be impact of transportation, a further reference to Good
sterile, apyrogenic and particle free. This latter point Distribution Practices (GDP). The third point is that
relates to visual inspection, a subject that has not been microbial ingress studies (or alternative methods) should
covered in any great detail by the Annex before. Each be utilised to determine the acceptable stopper height
facility is required to have a list of critical deficiencies displacement.
– such as particle, hairs and turbidity – and to subject
operators to regular assessment. The assessment should
be under practical conditions, with control of inspection
time, line speed, and component size. To capture
operator fatigue the test should be executed at the end
of the shift.

There are several references made to container integrity.

The first is that containers sealed under vacuum
should be tested for maintenance of vacuum after an
appropriate, pre-determined period and during shelf life
(which relates to a stability studies). The second is that
container closure integrity testing needs to consider the

10
Cleaning and Disinfection

The references to disinfection have been expanded. The need to rotate between two different disinfectants remains,
although the fact that one of these should be a sporicidal agent is a new one. Reference is also made to disinfectant
qualification, for both cleanrooms and for transfer disinfection (the act of introducing items into cleanrooms or between
cleanrooms of different grades). Not only is disinfectant efficacy testing described as important the Annex infers that this
is a type of testing that needs to be carried out by a facility independently. One reason for this is because the different
types of surfaces found in different facilities, together with regional variations with the microbial flora. With the discussions
about disinfectants in section 5, references are made to the need to assess the bioburden of non-sterile disinfectants and
to assign expiry dates.

Further requirements are that:

• Validation studies relate to the specific manner in
which disinfectants are use (by this there will be
differences between sprays, sprays followed by
wiping, and pre-saturated wipes, for example)
• Validation needs to be extended to the in-use expiry
periods of prepared solutions
• It remains that disinfectants used in Grade A and
B areas needs to be sterile; to this point the text
has been revised to state that disinfectants used in
Grade C and D areas may need to be sterile (this
generalisation is not especially helpful)
• There is a requirement that disinfectants that are not
purchased ready-made and which are thus made up
in-house as assessed for microbial contamination
(the time point is not specified, but presumably
this means end-of-use to represent ‘worst case’).
For ready-made disinfectants, assessment can be
undertaken by a review of certificates of analysis
• Monitoring the effectiveness of disinfectants through
environmental monitoring does appear in the earlier
draft; however, reference to “spore-resistant strains”
has been replaced with “organisms resistant to the
disinfection regime currently in use”, which is more
scientifically accurate since a microorganism does
not always need to enter into a spore state to be
difficult to kill. By resistance this does not refer to
‘acquired resistance’ (for which there is little scientific
support) but to the fact of innate resistance, where
some microorganisms are more resistant (such as
due to a factor of their cell wall, for example) to a
given disinfectant

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Utilities

The section on utilities has a reference to heating systems
for the first time. There is also an added requirement
for the trending of critical utility parameters. This
is expressed as an important step so alarms can be
reviewed to determine if they are specific or common
causes, thereby enabling the appropriate engineering
action to be taken. With alarm-based monitoring
particular emphasis is placed on Water-for-Injections
systems where is stated technology for monitoring Total
Organic Carbon and conductivity is put in place, at
locations determined by risk. This is because this type of
process analytical technology that permits continuous
monitoring is seen as superior to the type of discrete
monitoring that might be conducted by a chemistry
department.
The main change with water systems follows on from the
update to the European Pharmacopeia, permitting the
production of WFI through methods other than distillation.
Due to unspecified concerns with biofilm growth on
filters and the potential risk of endotoxin, the draft Annex
makes a recommendation that further techniques such
as nanofiltration and ultra-filtration are considered in
conjunction with operating reverse osmosis membranes.
As part of design controls the Annex requires that water
remains in a state of turbulent flow through distribution
systems (this minimises the risk of microbial adhesion
and hence biofilm development) and that a sanitisation
method is used. As an example, holding water at 70°C or
above is provided (although there are other forms water
control, such as chemical treatments). Another control
measure for water systems is with filtration. In section
6.11 the new text states: “Where WFI storage tanks are
equipped with hydrophobic bacteria retentive vent
filters, the filters should be sterilised and the integrity
of the filter tested before installation and after removal
following use.”
Where microbial counts occur, especially in response
to an upward trend, a normal recourse is to disinfect
the system (by heat or by chemicals). In relation to this,
the draft Annex states: “To minimise the risk of biofilm
formation, sterilisation or disinfection or regeneration
of water systems should be carried out according to
a predetermined schedule and when microbial counts
exceed action limits.”

Summary
The latest draft Annex 1 expands considerably upon the current text. This white paper has considered many of the key
aspects, focusing on the changes that impact sterile products manufacture. No review can cover every aspect, and
readers are encouraged to read the proposed Annex in detail for themselves.

In summary, the key takeaways from the latest draft are:

• The expectation for each facility to have in place a formal, holistic contamination control strategy, focused on
minimising contamination control with respect to sterile manufacturing

• Additional requirements for cleanroom classification (beyond ISO requirements)

• A major focus on risk-based approaches

• Recommendations for the wider use of barrier technology

• A strong focus on personnel controls, such as gowning, and training

Such stipulations are unlikely to alter when the final version appears and it is recommended that sterile product
manufacturers invest some time in considering the impact of these changes upon their procedures and processes.

12
References
1. Eudralex The Rules Governing Medicinal Products in the European Union, Volume 4 EU Guidelines to Good
Manufacturing Practice Medicinal Products for Human and Veterinary Use. Annex 1 Manufacture of Sterile Medicinal
Products, European Commission, Brussels, Belgium, 2009:
https://ec.europa.eu/health/sites/health/files/files/eudralex/vol-4/2008_11_25_gmp-an1_en.pdf

2. EMA (2020) Second targeted stakeholders’ consultation on the revision of Annex 1, on manufacturing of sterile
medicinal products, of Eudralex volume 4, at: https://ec.europa.eu/health/medicinal_products/consultations/2020_
sterile_medicinal_products_en

3. ICH Q9 Quality Risk Management. ICH Harmonised Tripartite Guideline, Step 4, Geneva, Switzerland. 2005.

About the author

Dr Tim Sandle
Dr. Tim Sandle has over twenty-five years’ experience of microbiological research and
biopharmaceutical processing. He is a member of several editorials boards and he has
written over six-hundred book chapters, peer reviewed papers and technical articles
relating to microbiology. Dr. Sandle works for a pharmaceutical manufacturer in the UK,
and is a visiting tutor at both the University of Manchester and UCL.

13
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