LegalizeCommission Implementing Regulation (EU) 2025/2091 of 17 October 2025 laying down good manufacturing practice for veterinary medicinal products in accordance with Regulation (EU) 2019/6 of the European Parliament and of the Council
V.4.2.Gases used in aseptic processes shall be filtered through a sterilising grade filter (14) (with a nominal pore size of a maximum of 0,22 μm) at the point of use. Where the filter is used on a batch basis (e.g. for filtration of gas used for overlay of aseptically filled products) or as product vessel vent filter, the results of the integrity test shall be reviewed as part of the batch certification/release process. Any transfer pipework or tubing that is located after the final sterilising grade filter shall be sterilised. When gases are used in the process, microbial monitoring of the gas shall be performed periodically at the point of use.
V.4.3.Where backflow from vacuum or pressure systems poses a potential risk to the product, mechanism(s) shall be put in place to prevent backflow when the vacuum or pressure system is shut off.
V.5. Heating and cooling and hydraulic systems
V.5.1.Major items of equipment associated with hydraulic, heating and cooling systems shall, where possible, be located outside the filling room. Appropriate controls shall be implemented to contain any spillage or cross contamination associated with the system fluids.
V.5.2.Appropriate systems shall be put in place to ensure that any leak from these systems that could present a risk to the product are detected (e.g. an indication system for leakage).
SECTION VI
PERSONNEL
VI.1.The manufacturer shall ensure that there are sufficient personnel, suitably qualified, trained and experienced in the manufacture and testing of sterile products and any of the specific manufacturing technologies used in the site’s manufacturing operations.
VI.2.Only the minimum number of personnel required shall be present in cleanrooms. The maximum number of operators in cleanrooms shall be determined and documented. During activities such as initial qualification and the aseptic process simulation the maximum number of operators that can be present in the cleanroom shall be duly considered so as not to compromise sterility assurance.
VI.3.All personnel including those performing cleaning, maintenance, monitoring and those that access cleanrooms shall receive regular training on aspects relevant to the manufacture of sterile products/aseptic manufacturing, including on gowning, the basic elements of microbiology and hygiene, with a specific focus on cleanroom practices, contamination control, aseptic techniques and the protection of sterile products (for those operators entering the grade B cleanrooms and/or intervening into grade A) and the potential consequences to the treated animals if the product is not sterile / fails to meet the required quality specifications. The level of training shall be based on the criticality of the function and the area where the personnel are working.
VI.4.Personnel accessing grade A and B areas shall be trained for aseptic gowning and aseptic behaviour. Compliance with aseptic gowning procedures is to be confirmed by means of an assessment prior to starting their functions and shall be periodically reassessed (at least annually). The assessment process shall involve both visual and microbial assessment (using monitoring locations such as gloved fingers, forearms, chest and hood (facemask/forehead).
VI.5.Unsupervised access to the grade A and grade B areas where aseptic operations are or will be conducted shall be restricted to appropriately qualified personnel, who have passed the gowning assessment and have participated in a successful aseptic process simulation.
Unqualified personnel shall not enter grade B cleanrooms or grade A in operation. If needed in exceptional cases, manufacturers shall establish written procedures outlining the process by which unqualified personnel can be brought into the grade B and A areas. An authorised person from the manufacturer shall supervise the unqualified personnel during their activities and assess the impact of these activities on the cleanliness of the area. Access by these persons shall be assessed and recorded.
VI.6.A process shall be put in place for the disqualification of personnel based on aspects of ongoing assessment and/or identification of an adverse trend from the personnel monitoring programme and/or after being implicated in a failed aseptic process simulation. Once disqualified, retraining and requalification shall be completed before permitting the operator to have any further involvement in aseptic practices. For operators entering grade B cleanrooms or performing intervention into grade A, it is advised that the requalification includes participation in a successful aseptic process simulation.
VI.7.High standards of personal hygiene and cleanness are essential. When a heath condition that may introduce an undue microbial hazard is declared by the relevant personnel or otherwise becomes apparent, access to the cleanroom shall be barred. Health conditions and actions to be taken with regard to personnel that can introduce an undue microbial hazard shall be documented in relevant procedures.
VI.8.Personnel involved in the handling/processing of materials of human/animal origin or of cultures of micro-organisms, other than those used in the current manufacturing process, or in other activities that may have a negative impact to quality (e.g. microbial contamination), shall not enter clean areas unless clearly defined and effective decontamination and entry procedures have been followed and documented.
VI.9.Wristwatches, make-up, jewellery, other personal items such as mobile phones and any other non-essential items shall not be allowed in clean areas. Electronic devices used in cleanrooms, e.g. mobile phones and tablets, that are supplied by the manufacturer solely for use in the cleanrooms, may be acceptable if suitably designed to permit cleaning and disinfection commensurate with the grade in which they are used. The use and disinfection of such equipment shall be included in the contamination control strategy.
VI.10.Cleanroom gowning and hand washing shall be done in accordance with written procedures designed to minimise the contamination of cleanroom clothing and/or the transfer of contaminants to the clean areas.
VI.11.The clothing and its quality shall be appropriate for the process and the grade of the working area. It shall be worn in such a way as to protect the product from contamination. When the required type of clothing needs to protect the operator from the product, it shall also be ensured that the protection of the product from contamination is not compromised.
Garments shall be visually checked for cleanliness and integrity immediately prior to and after gowning. Gown integrity shall also be checked upon exit. Prior to the use of sterilised garments and eye coverings, it shall be checked that they have been subject to the sterilisation process, that they are within their specified hold time and that the packaging has not been tampered. Reusable garments (including eye coverings) are to be replaced if damage is identified, or at a set frequency that is determined during qualification studies. The qualification of garments shall consider any necessary garment testing requirements, including damage to garments that may not be identified by visual inspection alone.
VI.12.A description of clothing typically required for each cleanliness grade is given below:
(a) Grade B (including access/interventions into grade A): — appropriate garments that are dedicated for use under a sterilised suit shall be worn before gowning; — appropriately sterilised, non-powdered, rubber or plastic gloves shall be worn while donning the sterilised garments; — sterile headgear shall enclose all hair (including facial hair) and, where separate from the rest of the gown, it shall be tucked into the neck of the sterile suit; — a sterile facemask and sterile eye coverings (e.g. goggles) shall be worn to cover and enclose all facial skin and prevent the shedding of droplets and particles; — appropriate sterilised footwear (e.g. over-boots) shall be worn; — trouser legs shall be tucked inside the footwear and garment sleeves shall be tucked into a second pair of sterile gloves worn over the pair worn while donning the gown; — the protective clothing shall minimise shedding of fibres or particles and retain particles shed by the body. The particle shedding and the particle retention efficiencies of the garments is to be assessed during the garment qualification; — garments shall be packed and folded in such a way as to allow operators to don the gown without contacting the outer surface of the garment and to prevent the garment from touching the floor.
(b) Grade C: — hair, beards and moustaches shall be covered; — a single or two-piece trouser suit gathered at the wrists and with high neck and appropriately disinfected shoes or overshoes shall be worn; they shall minimise the shedding of fibres and particles; — additional gowning, including gloves and facemask, may be required in grade C areas when performing activities that pose a risk of contamination.
(c) Grade D: — hair, beards and moustaches shall be covered; — a general protective suit and appropriately disinfected shoes or overshoes shall be worn; — appropriate measures shall be taken to avoid any ingress of contaminants from outside the clean area; — additional gowning including gloves and facemask may be required in grade D areas when performing activities that pose a risk of contamination.
VI.13.Cleanroom gowning shall take place in change rooms of an appropriate cleanliness grade to ensure that gown cleanliness is maintained. Outdoor clothing including socks (other than personal underwear) shall not be brought into changing rooms leading directly to grade B and C areas. In addition, a single or two-piece facility trouser suit, covering the full length of the arms and the legs, and facility socks covering the feet, shall be worn before entry to change rooms for grades B and C. Facility suits and socks shall not present a risk of contamination to the gowning area or processes.
VI.14.Every operator entering grade B or A areas shall gown into clean, sterilised protective garments (including eye coverings and masks) of an appropriate size at each entry. The maximum period for which the sterilised gown may be worn before replacement during a shift shall be defined as part of the garment qualification.
VI.15.Gloves shall be regularly disinfected during operations. Garments and gloves shall be changed immediately if they become damaged and present any risk of product contamination.
VI.16.Reusable clean area clothing shall be cleaned in a laundry facility adequately segregated from production operations, using a qualified process ensuring that the clothing is not damaged or contaminated by fibres or particles during the repeated laundry process. Laundry facilities used shall not introduce a risk of contamination or cross-contamination. After washing and before packing, garments shall be visually inspected for damage and visual cleanliness. The garment management processes shall be established as part of the garment qualification programme and shall include a maximum number of laundry and sterilisation cycles.
VI.17.Activities in clean areas that are not critical to the production processes shall be kept to a minimum, especially when aseptic operations are in progress. With a view to avoid excessive shedding of particles and organisms, movement of personnel shall be slow, controlled and methodical. Operators performing aseptic operations shall adhere to aseptic technique at all times to prevent changes in air currents that may introduce air of lower quality into the critical zone. In addition, movement adjacent to the critical zone shall be restricted and the obstruction of the path of the unidirectional (first air) airflow shall be avoided.
SECTION VII
PRODUCTION AND SPECIFIC TECHNOLOGIES
VII.1. Terminally sterilised products (15)
VII.1.1.Preparation of components and materials shall be performed in at least a grade D cleanroom in order to limit the risk of microbial, endotoxin/pyrogen and particle contamination, so that the product is suitable for sterilisation. However, where the product is at a high or unusual risk of microbial contamination (e.g. the product actively supports microbial growth, the product must be held for long periods before filling or the product is not processed mostly in closed vessels), then preparation shall be carried out in at least a grade C environment. Preparation of ointments, creams, suspensions and emulsions shall also be carried out in at least a grade C environment before terminal sterilisation.
By way of derogation from the C grade environment as foreseen above, in exceptional cases, e.g. where the manufacturing process involves the generation of powder/dust that cannot be prevented by reasonable means, preparation of products to be terminally sterilised may be performed in a grade D environment. For the implementation of grade D in this exceptional case, the manufacturer shall be required to perform a risk assessment and apply suitable measures to ensure that there is no negative impact on the quality of the product. This shall be documented as part of the contamination control strategy.
VII.1.2.Primary packaging containers and components shall be cleaned using validated processes to ensure that particle, endotoxin/pyrogen and bioburden contamination is appropriately controlled.
VII.1.3.Filling of products for terminal sterilisation shall be carried out in at least a grade C environment. However, if the product is at an unusual risk of contamination from the environment (for example, the filling operation is slow, the containers are wide necked or are necessarily exposed for more than a few seconds before closing), the product shall be filled in grade A with at least a grade C background, unless additional measures to ensure the absence of a negative impact to the quality of the product are implemented, in which case the filling operation shall take place – as a minimum – in a grade D environment.
VII.1.4.To reduce the level of bioburden and particles prior to filling into the final product container, processing of the bulk solution shall include, where possible, a filtration step with a microorganism retaining filter and a maximum time between preparation and filling shall be set.
VII.1.5.Examples of operations to be carried out in the various grades are given in Table 3.
| Grade A | Filling of products when unusual/high risk of microbial contamination, unless a lower grade can be justified in accordance with Section VII.1.3. |
|---|---|
| Grade C | Preparation of solutions when unusual/high risk of microbial contamination, unless grade D can be justified in accordance with the second subparagraph of Section VII.1.1. Filling of products (other than when grade A is required), unless grade D can be justified in accordance with Section VII.1.3. |
| Grade D | Preparation of solutions and components for subsequent filling. |
VII.2. Aseptic preparation and processing
VII.2.1.The aseptic process shall be documented as part of the contamination control strategy. Specifically, the risks associated with the aseptic process, and any associated requirements, shall be identified, assessed and appropriate controls shall be identified including the acceptance criteria for these controls, requirements for monitoring and the review of their effectiveness. Methods and procedures to control those risks shall be clearly described and implemented. Accepted residual risks shall be formally documented.
VII.2.2.Precautions to minimise microbial, endotoxin/pyrogenic and particle contamination in the site shall be described in the contamination control strategy and shall be implemented during the preparation of the aseptic environment, during all processing stages (including the stages before and after bulk product sterilisation), and until the product is sealed in its final container. The presence of materials liable to generate particles and fibres shall be minimised in cleanrooms.
VII.2.3.Where possible, the use of equipment such as RABS, isolators or other systems shall be used in order to reduce the need for critical interventions (16) into grade A and to minimise the risk of contamination. Robotics and automation of processes may also be considered to eliminate direct human critical interventions (e.g. dry heat tunnel, automated lyophilizer loading, sterilisation in place).
VII.2.4.Examples of operations to be carried out in the various environmental grades are given in Table 4.
| Grade A | — Aseptic assembly of filling equipment. — Connections made under aseptic conditions (where sterilised product contact surfaces are exposed) that are post the final sterilising grade filter. These connections shall be sterilised by steam-in-place whenever possible. — Aseptic compounding and mixing. — Replenishment of sterile bulk product, containers and closures. — Removal and cooling of unprotected (e.g. with no packaging) items from sterilisers. — Staging and conveying of sterile primary packaging components in the aseptic filling line while not wrapped. — Aseptic filling, sealing of containers such as ampoules, vial closure, transfer of open or partially stoppered vials. — Loading of a lyophilizer. |
|---|---|
| Grade B | — Background support for grade A (when not in an isolator). — Conveying or staging, while protected from the surrounding environment, of equipment, components and ancillary items for introduction into grade A. |
| Grade C | — Preparation of solutions to be filtered including sampling and dispensing. |
| Grade D | — Cleaning of equipment. — Handling of components, equipment and accessories after cleaning. — Assembly under HEPA filtered airflow of cleaned components, equipment and accessories prior to sterilisation. — Assembly of closed and sterilised single use systems using intrinsic sterile connection devices (1) |
| (1) For the purposes of this Annex, ‘intrinsic sterile connection device’ means a device that reduces the risk of contamination during the connection process; it can be mechanical or fusion sealing. |
VII.2.5.For products where the final formulation cannot be filtered, the following measures shall be considered as appropriate:
— all product and component contact equipment shall be sterilised prior to use;
— all raw materials or intermediates shall be sterilised and aseptically added;
— bulk solutions or intermediates shall be sterilised.
VII.2.6.The unwrapping, assembly and preparation of sterilised equipment, components and ancillary items with direct or indirect product contact shall be treated as an aseptic process and performed in grade A with a grade B background. The filling line set-up and filling of the product shall be treated as an aseptic process and performed in grade A with a grade B background. Where an isolator is used, the background shall be in accordance with Section III.3.3 of this Annex.
VII.2.7.Preparation and filling of products such as ointments, creams, suspensions and emulsions shall be performed in grade A with a grade B background when the product and components are exposed to the environment and the product is not subsequently filtered (via a sterilising grade filter) or terminally sterilised. Where an isolator or RABS is used, the background shall be in accordance with Section III.3.3 of this Annex.
VII.2.8.Aseptic connections shall be performed in grade A with a grade B background unless subsequently sterilised in place or conducted with intrinsic sterile connection devices that minimise any potential contamination from the immediate environment. Intrinsic sterile connection devices shall be designed to mitigate the risk of contamination. Where an isolator is used, the background shall be in accordance with Section III.3.3 of this Annex.
Aseptic connections shall be appropriately assessed and their effectiveness verified.
VII.2.9.Aseptic manipulations (including non-intrinsic sterile connection devices) shall be minimised through the use of engineering design solutions such as preassembled and sterilised equipment. Whenever feasible, product contact piping and equipment shall be pre-assembled and sterilised in place.
VII.2.10.A list of allowed and qualified interventions, both inherent (17) and corrective, that may occur during production shall be set. The type of inherent and corrective interventions, and how to perform them, shall be first evaluated in accordance with quality risk management principles and the outcome of the aseptic process simulation and be kept up to date.
Interventions shall be carefully designed to ensure that the risk of contamination of the environment, process and product is effectively minimised, including consideration of any impact on air-flows and critical surfaces (18) and products. Engineering solutions shall be used whenever possible to minimise incursion by operators during the intervention. Aseptic technique shall be observed at all times, including the use of sterile tools for manipulations.
Non-authorised/non-qualified interventions shall only be performed in exceptional circumstances, with due consideration of the risks associated with the intervention and with the authorisation of the quality unit. Moreover, the details of the intervention conducted shall be recorded, be thoroughly assessed by the quality department and be duly considered during batch release.
VII.2.11.Interventions and stoppages shall be recorded in the batch record. Each line stoppage or intervention shall be sufficiently documented in batch records with the associated time, duration of the event, and operators involved.
VII.2.12.The duration of each aspect of aseptic preparation and processing shall be minimised as far as possible and validated maximum times shall be set including:
— the holding time between equipment, component, and container cleaning, drying and sterilisation;
— the holding time for the sterilised equipment, components, and containers before use and during filling/assembly;
— the holding time for a decontaminated environment, such as the RABS or isolator before use;
— the time between the start of the preparation of a product and its sterilisation or filtration through a microorganism-retaining filter (if applicable), through to the end of the aseptic filling process. A maximum permissible time for each product shall be set taking into account its composition and the method of storage;
— the holding time for the sterilised product prior to filling;
— the aseptic processing time; and
— the filling time.
VII.2.13.Aseptic operations (including aseptic process simulation) shall be monitored on a regular basis by personnel with specific expertise in aseptic processing to verify the correct performance of operations, including operator’s behaviour in the cleanroom, and to address inappropriate practices if detected.
VII.3. Finishing activities
VII.3.1.Open primary packaging containers shall be maintained under grade A conditions with the appropriate background for the technology as described in Section III.3.3. For vials that are partially stoppered or prefilled syringes, the additional considerations as set forth in Section VII.7.6 apply also.
VII.3.2.Final containers shall be closed by appropriately validated methods.
VII.3.3.Where final containers are closed by fusion, e.g. Blow-Fill-Seal, Form-Fill-Seal, small and large volume parenteral bags, glass or plastic ampoules, the critical parameters and variables that affect seal integrity shall be set and be effectively controlled and monitored during operations.
Glass ampoules, Blow-Fill-Seal units and small volume containers (≤ 100 ml) closed by fusion shall be subject to 100 % integrity testing using validated methods. For large volume containers (> 100 ml) closed by fusion, reduced sampling may be acceptable where scientifically justified and based on data demonstrating the consistency of the existing process and a high level of process control. Visual inspection is not an acceptable integrity test method.
VII.3.4.Samples of products using systems other than fusion shall be taken and checked for integrity using validated methods. The frequency of testing shall be based on the knowledge and experience of the container and closure systems being used. The sampling plan shall be scientifically justified and be based on information such as the supplier’s management, the packaging component specifications and the process knowledge.
VII.3.5.Containers sealed under vacuum shall be tested for maintenance of vacuum after an appropriate pre-determined period prior to certification/release and during shelf life.
VII.3.6.The container closure integrity validation shall take into consideration any transportation or shipping requirements that may negatively impact the integrity of the container (e.g. by decompression or extreme temperatures).
VII.3.7.Where the equipment used to crimp vial caps can generate large quantities of non-viable particle, measures shall be taken to prevent particle contamination, such as locating the equipment at a physically separate station equipped with adequate air extraction.
VII.3.8.Vial capping of aseptically filled products may be undertaken as an aseptic process using sterilised caps or as a clean process outside the aseptic processing area. Where the latter approach is adopted, vials shall be protected by grade A conditions up to the point of leaving the aseptic processing area, and thereafter stoppered vials shall be protected with a grade A air supply (19) until the cap has been crimped. The supporting background environment of grade A air supply shall meet at least grade D requirements.
Where capping is a manual process, it shall be performed under grade A conditions either in an appropriately designed isolator or in grade A with a grade B background.
VII.3.9.Where capping of an aseptically filled product is conducted as a clean process with grade A air supply protection, vials with missing or displaced stoppers shall be rejected prior to capping. Appropriately qualified, automated methods for stopper height detection shall be in place.
VII.3.10.Where human intervention is required at the capping station, appropriate technological and organisational measures shall be used to prevent direct contact with the vials and to minimise contamination. RABS and isolators may be beneficial in assuring the required conditions.
VII.3.11.All filled containers of parenteral products shall be inspected individually for extraneous contamination or other defects. A defect classification including the criticality thereof shall be established during qualification and based on risk and historical knowledge. Factors to consider include, but are not limited to, the potential impact of the defect to the treated animal and the route of administration. A defect library capturing all known types of defects shall be established and be used for the training of production and quality assurance personnel.
Critical defects shall be identified upfront and not during subsequent sampling and inspection of acceptable containers. Any critical defect identified subsequently shall trigger an investigation as it indicates a possible failure of the original inspection process.
Batches with unusual levels of defects, when compared with routine defect numbers for the process (based on routine and trend data) shall be investigated.
VII.3.12.When inspections are performed manually, suitable and controlled conditions of illumination and background shall be ensured. Inspection rates shall be appropriately controlled and qualified. Operators performing the inspection shall undergo visual inspection qualification (whilst wearing corrective lenses, if these are normally worn) at least annually. The qualification shall be performed using appropriate samples from the manufacturer's defect library sets and taking into consideration worst case scenarios (e.g. inspection time, line speed where the product is transferred to the operator by a conveyor system, container size or fatigue) and shall also include eyesight checks. Work conditions shall be adequate to reduce elements of distraction and, in order to minimise operator fatigue, frequent breaks of an appropriate duration shall be taken.
VII.3.13.Where automated methods of inspection are used, the process shall be validated to detect known defects (which may impact product quality or safety). The performance of the automated methods shall be equal to, or better than, manual inspection methods. The performance of the equipment shall be challenged using representative defects prior to start up and at regular intervals throughout the batch.
VII.3.14.Results of the inspection shall be recorded and defect types and numbers trended. Reject levels for the various defect types shall also be trended based on statistical principles. When adverse trends are observed, the impact on batches on the market shall be assessed.
VII.4. Sterilisation
VII.4.1.1.Where possible, finished products shall be terminally sterilised, using a validated and controlled sterilisation process, as this provides a greater assurance of sterility than a validated and controlled sterile filtration process and/or aseptic processing. Where it is not possible for a product to undergo terminal sterilisation, consideration shall be given to using post-aseptic processing terminal heat treatment (20), combined with aseptic process to give improved sterility assurance.
VII.4.1.2.The selection, design and location of the equipment and cycle/programme used for the sterilisation shall be based on scientific principles and data which demonstrate repeatability and reliability of the sterilisation process. All parameters shall be defined and critical parameters shall be controlled, monitored and recorded.
VII.4.1.3.All sterilisation processes shall be validated. Validation studies shall take into account the product composition, the storage conditions and the maximum time between the start of the preparation of a product or material to be sterilised and its sterilisation. Before any sterilisation process is implemented, its suitability for the product and the equipment, and its efficacy in consistently achieving the desired sterilising conditions in all parts of each type of load to be processed shall be validated by physical measurements and, where appropriate, by biological indicators (21). For an effective sterilisation, the process shall be designed to ensure that the whole of the product, as well as the surfaces of the equipment and components are subject to the required treatment.
VII.4.1.4.Particular attention shall be paid when the adopted product sterilisation method is not described in the current edition of the Pharmacopoeia, or when it is used for a product that is not a simple aqueous solution. Where possible, heat sterilisation shall be the method of choice.
VII.4.1.5.Validated loading patterns shall be established for all sterilisation processes and load patterns shall be subject to periodic revalidation. Maximum and minimum loads shall also be addressed as part of the overall load validation strategy.
VII.4.1.6.The validity of the sterilising process shall be reviewed at scheduled intervals based on risk. Heat sterilisation cycles shall be revalidated at least annually for load patterns that are considered worst case. Other load patterns shall be validated at an appropriate frequency that shall be justified as part of the contamination control strategy.
VII.4.1.7.Routine operating parameters shall be established and adhered to for all sterilisation processes, e.g. physical parameters and loading patterns.
VII.4.1.8.Mechanisms shall be put in place to detect a sterilisation cycle that does not conform to the validated parameters. Any failed sterilisation or any sterilisation that deviated from the validated process (e.g. have longer or shorter phases such as heating cycles) shall be investigated.
VII.4.1.9.Suitable biological indicators placed at appropriate locations shall be considered as an additional method to support the validation of the sterilisation process. Biological indicators shall be stored and used according to the manufacturer’s instructions. Where biological indicators are used to support the validation and/or to monitor a sterilisation process (e.g. with ethylene oxide), positive controls shall be tested for each sterilisation cycle. Moreover, if biological indicators are used, strict precautions shall be taken to avoid transferring microbial contamination to the manufacturing or other testing processes. Biological indicator results in isolation cannot be used to override other critical parameters and process design elements.
VII.4.1.10.The reliability of biological indicators is important. Therefore, suppliers shall be qualified and transportation and storage conditions shall be controlled to ensure that the quality thereof is not compromised. Prior to the use of a new batch/lot of biological indicators, the population, purity and identity of the indicator organism of the batch/lot shall be verified. For other critical parameters, e.g. D-value (22), Z-value (23), the batch certificate provided by the qualified supplier may normally be used.
VII.4.1.11.Products, equipment and components that have not been subject to the sterilisation process shall be clearly distinguished from those that have through appropriate means. Equipment such as baskets or trays used to carry products, other items of equipment and/or components shall be clearly labelled (or electronically tracked) with the product name and batch number and an indication of whether or not it has been sterilised. Indicators such as autoclave tape or irradiation indicators may be used, where appropriate, to indicate whether or not a batch (or sub-batch material, component, equipment) has passed through a sterilisation process. It is noted that these indicators show only that the sterilisation process has occurred but are not indicative of product sterility or achievement of the required sterility assurance level.
VII.4.1.12.Sterilisation records shall be available for each sterilisation run. Each cycle shall have a unique identifier. These records shall be reviewed and considered as part of the batch certification/release procedure.
VII.4.1.13.Where required, materials, equipment and components shall be sterilised by validated methods appropriate to the specific material. Suitable protection after sterilisation shall be provided to prevent recontamination.
If sterilised items are not used immediately after sterilisation, these shall be stored using appropriately sealed packaging and a maximum hold time shall be established. Where justified, components that have been packaged with multiple sterile packaging layers need not be stored in a cleanroom if the integrity and configuration of the sterile pack allows the items to be readily disinfected during transfer by operators into grade A (e.g. by the use of multiple sterile coverings that can be removed at each transfer from lower to higher grade). Where protection is achieved by containment in sealed packaging, that packaging process shall take place prior to sterilisation.
VII.4.1.14.The transfer into grade A of sterilised materials, equipment, components and ancillary items in sealed packaging shall be done using appropriate validated methods (for example, airlocks or pass-through hatches) with accompanying disinfection of the exterior of the sealed packaging. The use of rapid transfer port technology (24) may also be considered. The methods used shall be demonstrated to effectively control the potential risk of contamination of the grade A and grade B areas and, likewise, the disinfection procedure shall be demonstrated to be effective in reducing any contamination on the packaging to acceptable levels for entry of the item into the grade B and grade A areas.
VII.4.1.15.Where materials, equipment, components and ancillary items are sterilised in sealed packaging or containers, the packaging shall be qualified for minimizing the risk of particulate, microbial, endotoxin/pyrogen or chemical contamination, and for compatibility with the selected sterilisation method. The packaging sealing process shall be validated. The validation shall consider the integrity of the sterile protective barrier system, the maximum hold time before sterilisation and the maximum shelf life assigned to the sterilised items. The integrity of the sterile protective barrier system for each of the sterilised items shall be checked prior to use.
VII.4.1.16.For materials, equipment, components and ancillary items that are not a direct or indirect product contact part and are necessary for aseptic processing but cannot be sterilised, an effective and validated disinfection and transfer process shall be put in place. These items, once disinfected, shall be protected to prevent recontamination. These items, as well as other items that are potential routes of contamination, shall be included in the environmental monitoring programme.
VII.4.2.1.Each heat sterilisation cycle shall be recorded either electronically or by hardcopy, using equipment with suitable accuracy and precision. The system used shall have safeguards and/or redundancy in its control and monitoring instrumentation to detect a cycle that is not conforming to the validated cycle parameter requirements and to abort or fail such cycle (e.g. by the use of duplex/double probes connected to independent control and monitoring systems).
VII.4.2.2.The position of the temperature probes used for controlling and/or recording shall be determined during the validation having regard to the system’s design and with a view to correctly record and represent routine cycle conditions. Validation studies shall demonstrate the suitability of the system’s control and recording probe locations, and shall include the verification of the function and location of these probes by the use of an independent monitoring probe located at the same position during validation.
VII.4.2.3.The entire load shall reach the required temperature before the measurement of the sterilising time-period starts. For sterilisation cycles controlled by using a reference probe within the load, specific consideration shall be given to ensuring that the load probe temperature is controlled within a defined temperature range prior to the start of the cycle.
VII.4.2.4.After completion of the high temperature phase of a heat sterilisation cycle, precautions shall be taken against contamination of a sterilised load during cooling. Any cooling liquid or gas that comes into contact with the product or sterilised material shall be sterilised. Additional requirements applicable where parametric release has been authorised are laid down in Annex IX.
VII.4.3.1.Moist heat sterilisation can be achieved using steam (direct or indirect contact) or with other systems such as superheated water systems (cascade or immersion cycles) that can be used for containers that may be damaged by other cycle designs (e.g. Blow-Fill-Seal containers, plastic bags).
VII.4.3.2.The items to be sterilised, other than products in sealed containers, shall be dry and packaged in a protective barrier system that allows removal of air and penetration of steam and prevents recontamination after sterilisation. All loaded items shall be dry upon removal from the steriliser. Load dryness shall be confirmed by visual inspection as a part of the sterilisation process acceptance.
VII.4.3.3.For porous cycles (hard goods), time, temperature and pressure shall be used to monitor the process and be recorded. Each sterilised item shall be inspected for damage, packaging material integrity and moisture upon removal from the autoclave. Any item found not to be fit for purpose shall be removed from the manufacturing area and an investigation shall be performed.
VII.4.3.4.For autoclaves capable of performing prevacuum sterilisation cycles, the temperature shall be recorded at the chamber drain throughout the sterilisation period. Load probes may also be used where appropriate, but the controlling system shall remain related to the load validation. For steam in place systems, the temperature shall be recorded at appropriate condensate drain locations throughout the sterilisation period. Validation of porous cycles shall include a calculation of equilibration time (25), exposure time, correlation of pressure and temperature and the minimum/maximum temperature range during the exposure. Validation of fluid cycles shall include temperature, time and/or F0 value (26). Critical processing parameters shall be subject to defined limits (including appropriate tolerances) and be confirmed as part of the sterilisation validation and of the routine cycle acceptance criteria.
VII.4.3.5.Leak tests on the steriliser shall be carried out periodically (normally weekly) when a vacuum phase is part of the cycle, and when the system is returned – post-sterilisation – to a pressure lower than the environment surrounding the steriliser.
VII.4.3.6.When the sterilisation process includes air purging (e.g. porous autoclave loads, lyophilizer chambers), there shall be adequate assurance of air removal prior to and during sterilisation. For autoclaves, this shall include an air removal test cycle (normally performed on a daily basis) or the use of an air detector system. Loads to be sterilised shall be designed to support effective air removal and be free draining to prevent the build-up of condensate.
VII.4.3.7.Distortion and damage of non-rigid containers that are terminally sterilised, such as containers produced by Blow-Fill-Seal or Form-Fill-Seal technologies, shall be prevented by appropriate cycle design and control (for instance setting correct pressure, heating and cooling rates and loading patterns).
VII.4.3.8.Where steam in place systems are used for sterilisation (e.g. for fixed pipework, vessels and lyophilizer chambers), the system shall be appropriately designed and validated to ensure that all parts of the system are subject to the required treatment. The system shall be monitored for temperature, pressure and time at appropriate locations during routine use to ensure all areas are effectively and reproducibly sterilised. These locations shall be demonstrated as being representative of, and correlated with, the slowest to heat locations during initial and routine validation. Once a system has been sterilised by steam in place, it shall remain integral and, where required by the relevant operations, maintained under positive pressure or otherwise equipped with a sterilising vent filter prior to use.
VII.4.3.9.In fluids load cycles where superheated water is used as the heat transfer medium, the heated water shall consistently reach all of the required contact points. Initial qualification studies shall include temperature mapping of the entire load. There shall be routine checks on the equipment to ensure that nozzles (where the water is introduced) are not blocked and drains remain free from debris.
VII.4.3.10.Validation of the sterilisation of fluids loads in a superheated water autoclave shall include temperature mapping of the entire load and heat penetration and reproducibility studies. All parts of the load shall heat up uniformly and achieve the desired temperature for the specified time. Routine temperature monitoring probes shall be correlated to the worst case positions identified during the qualification process.
VII.4.4.1.Dry heat sterilisation utilizes high temperatures of air or gas to sterilise a product or an article. It is of particular use in the thermal removal of difficult-to-eliminate thermally robust contaminants such as endotoxin/pyrogen. The combination of time and temperature to which the product, components or equipment are exposed shall produce an adequate and reproducible level of lethality and/or endotoxin/pyrogen inactivation/removal when operated routinely within the established limits. The process may be operated in an oven or in a continuous tunnel process, e.g. for sterilisation and depyrogenation of glass containers.
VII.4.4.2.Dry heat sterilisation/depyrogenation tunnels shall be configured to ensure that airflow protects the integrity and performance of the grade A sterilising zone by maintaining appropriate pressure differentials and airflow through the tunnel. Air pressure difference profiles shall be assessed. The impact of any airflow change shall be assessed to ensure that the heating profile is maintained. All air supplied to the tunnel shall pass through at least a HEPA filter and periodic tests (at least biannually) shall be performed to demonstrate air filter integrity. In addition, any tunnel parts that come into contact with sterilised components shall be appropriately sterilised or disinfected.
Critical process parameters that shall be addressed during validation and/or routine processing include, but are not limited to:
— belt speed or dwell time within the sterilising zone;
— temperature – minimum and maximum temperatures;
— heat penetration of the material/article;
— heat distribution/uniformity;
— airflows determined by air pressure difference profiles correlated with the heat distribution and penetration studies.
VII.4.4.3.When a thermal process is used as part of the depyrogenation process for any component or product contact equipment/material, validation studies shall be performed to demonstrate that the process provides a suitable Fh value (27) and results in a minimum 3 log10 reduction in endotoxin concentration. When this is attained, there is no additional requirement to demonstrate sterilisation.
VII.4.4.4.During validation, containers spiked with endotoxin shall be used and a full reconciliation performed. Containers shall be representative of the materials normally processed (in respect to composition of the packaging materials, porosity, dimensions, nominal volume). Endotoxin quantification and recovery efficiency shall also be demonstrated.
VII.4.4.5.Dry heat ovens are typically employed to sterilise or depyrogenate primary packaging components, starting materials or active substances but may be used for other processes. They shall be maintained at a positive pressure relative to lower grade clean areas throughout the sterilisation and post sterilisation hold process unless the integrity of the packaging is maintained. All air entering the oven shall pass through a HEPA filter. Critical process parameters that shall be considered in qualification and/or routine processing include, but are not limited to:
— temperature;
— exposure period/time;
— chamber pressure (for maintenance of over pressure);
— air speed;
— air quality within the oven;
— heat penetration of material/article (slow to heat spots);
— heat distribution/uniformity;
— load pattern and configuration of articles to be sterilised/depyrogenated including minimum and maximum loads.
VII.4.5.1.Sterilisation by radiation is used mainly for the sterilisation of heat sensitive materials and products. Ultraviolet irradiation is not an acceptable method of sterilisation. Specific requirements related to the use of ionising radiation sterilisation are laid down in Annex VII.
VII.4.5.2.Validation procedures shall ensure that the effects of variation in density of the product and packages are considered.
VII.4.6.1.This method shall only be used when no other method is practicable. During process validation, it shall be shown that there is no damaging effect on the product and that the conditions and time allowed for degassing are suitable to attain a reduction of any residual ethylene oxide gas and reaction products to defined acceptable limits for the given product or material.
VII.4.6.2.Direct contact between the gas and microbial cells is essential. Therefore, precautions shall be taken to avoid the presence of organisms likely to be enclosed in material such as crystals or dried protein. The nature, porosity and quantity of packaging materials can also significantly affect the process.
VII.4.6.3.Before exposure to the gas, materials shall be brought into equilibrium with the humidity and temperature required by the process. Where steam is used to condition the load for sterilisation, it shall be of an appropriate quality. The time required for this operation shall be balanced against the need to minimise the time before sterilisation.
VII.4.6.4.Each sterilisation cycle shall be monitored with suitable biological indicators, using the appropriate number of test units distributed throughout the load at defined locations that have been shown to be worst case locations during validation.
VII.4.6.5.Critical process parameters to be considered as part of the sterilisation process validation and routine monitoring include, but are not limited to:
— ethylene oxide gas concentration;
— pressure;
— amount of ethylene oxide gas used;
— relative humidity;
— temperature;
— exposure time.
VII.4.6.6.After sterilisation, the load shall be aerated to allow ethylene oxide gas and/or its reaction products to desorb from the packaged product to predetermined levels. Aeration can occur within a steriliser chamber and/or in a separate aeration chamber or aeration room. The aeration phase shall be validated as part of the overall ethylene oxide sterilisation process validation.
VII.4.7.1Solutions or liquids that cannot be sterilised in their final container shall be sterilised by filtration through a sterile sterilising grade filter (with a nominal pore size of a maximum of 0,22 μm that has been appropriately validated to obtain a sterile filtrate) and subsequently aseptically filled into a previously sterilised container. The selection of the filter used shall ensure that it is compatible with the product and in compliance with the marketing authorisation.
VII.4.7.2.Suitable bioburden reduction prefilters and/or sterilising grade filters may be used at multiple points during the manufacturing process to ensure a low and controlled bioburden of the liquid prior to the final sterilising filter. Due to the potential additional risks of a sterile filtration process, as compared with other sterilisation processes, an additional filtration through a sterile sterilising grade filter, as close to the point of fill as possible, shall be considered as part of an overall contamination control strategy.
VII.4.7.3.The selection of components for the filtration system and their interconnection and arrangement within the filtration system, including pre-filters, shall be based on the critical quality attributes of the product, justified and documented. The filtration system shall minimise the generation of fibres and particles, not cause or contribute to unacceptable levels of impurities, or possess characteristics that otherwise alter the quality or the efficacy of the product. Similarly, the filter characteristics shall be compatible with the fluid and not be adversely affected by the product to be filtered. Adsorption of product components and extraction/leaching of filter components shall be evaluated.
VII.4.7.4.The filtration system shall be designed to:
— allow operation within validated process parameters;
— maintain the sterility of the filtrate;
— minimise the number of aseptic connections required between the final sterilising grade filter and the final filling of the product;
— allow cleaning procedures to be conducted as necessary;
— allow sterilisation procedures, including sterilisation in place, to be conducted as necessary;
— permit in-place integrity testing of the 0,22 μm final sterilising grade filter, preferably as a closed system, both prior to and following filtration as necessary. In-place integrity testing methods shall be preferably used to avoid any adverse impact on the quality of the product.
VII.4.7.5.Sterile filtration of liquids shall be validated in accordance with relevant Pharmacopeia requirements. Validation may be grouped by different strengths or variations of a product but shall be done under worst-case conditions. The rationale for grouping shall be justified and documented.
VII.4.7.6.Wherever possible during filter validation, the product to be filtered shall be used for bacterial retention testing (28) of the sterilising grade filter. Where the product to be filtered is not suitable for use in bacterial retention testing, a suitable surrogate product shall be justified for use in the test. The challenge organism used in the bacterial retention test shall also be justified.
VII.4.7.7.Filtration parameters that shall be considered and established during validation include, but are not limited to:
(a) The wetting fluid used for filter integrity testing: — it shall be based on the filter manufacturer’s recommendation or the fluid to be filtered. The appropriate integrity test value specification shall be established; — if the system is flushed or integrity tested in situ with a fluid other than the product, appropriate actions shall be taken to avoid any deleterious effect on product quality.
(b) Filtration process conditions including: — fluid pre-filtration holding time and effect on bioburden; — filter conditioning, with fluid if necessary; — maximum filtration time/total time that the filter is in contact with the fluid; — maximum operating pressure; — flow rate; — maximum filtration volume; — temperature; — the time taken to filter a known volume of bulk solution and the pressure difference to be used across the filter.
VII.4.7.8.Routine process controls shall be implemented to ensure adherence to validated filtration parameters. Results of critical process parameters shall be included in the batch record, including – but not limited to – the minimum time taken to filter a known volume of bulk solution and pressure difference across the filter. Any significant difference from critical parameters during manufacturing shall be documented and investigated.
VII.4.7.9.The integrity of the sterilised filter assembly shall be verified by integrity testing before use (pre-use post sterilisation integrity test or PUPSIT), to check for damage and loss of integrity caused by the filter preparation prior to use. However, it is recognised that PUPSIT may not always be possible after sterilisation due to process constraints (e.g. the filtration of very small volumes of solution). In these cases, an alternative approach may be taken providing that a thorough risk assessment has been performed and compliance is achieved by the implementation of appropriate controls to mitigate any risk of a non-integral filtration system.
Points to consider in such a risk assessment shall include but are not limited to:
— in-depth knowledge and control of the filter sterilisation process to ensure that the potential for damage to the filter is minimised;
— in-depth knowledge and control of the supply chain including contract sterilisation facilities, defined transport conditions and packaging of the sterilised filter (to prevent damage to the filter during transportation and storage);
— in-depth process knowledge such as the specific product type, including particle burden 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), and the implementation of pre-filtration or processing steps prior to the final sterilising grade filter that would remove particle burden prior to the sterile filtration.
In addition, a sterilising grade filter that is used to sterilise a fluid shall be subject to a non-destructive integrity test post-use prior to removal of the filter from its housing. The integrity test process shall be validated and test results shall correlate to the microbial retention capability of the filter established during validation. Examples of tests that are used include bubble point, diffusive flow, water intrusion or pressure hold test.
VII.4.7.10.The integrity of critical sterile gas and air vent filters (that are directly linked to the sterility of the product) shall be verified by testing after use, with the filter remaining in the filter assembly or housing.
VII.4.7.11.The integrity of non-critical air or gas vent filters shall be confirmed and recorded at appropriate intervals. Where gas filters are in place for extended periods, integrity testing shall be carried out at installation and prior to replacement. The maximum duration of use shall be specified and monitored based on risk (e.g. considering the maximum number of uses and heat treatment/sterilisation cycles permitted as applicable).
VII.4.7.12.For gas filtration, unintended moistening or wetting of the filter or filter equipment shall be avoided.
VII.4.7.13.If the sterilising filtration process has been validated as a system consisting of multiple filters to achieve the sterility for a given fluid, the filtration system is considered to be a single sterilising unit and all filters within the system shall satisfactorily pass integrity testing after use.
VII.4.7.14.In a redundant filtration system (where a second redundant sterilising grade filter is present as a backup but the sterilising process is validated as only requiring one filter), post-use integrity test of the primary sterilising grade filter shall be performed and, if demonstrated to be integral, a post-use integrity test of the redundant (backup) filter is not necessary. However, in the event of a failure of the post-use integrity test on the primary filter, post-use integrity test on the secondary (redundant) filter shall be performed, in conjunction with an investigation and risk assessment to determine the reason for the primary filter test failure.
VII.4.7.15.Bioburden samples shall be taken from the bulk product and immediately prior to the final sterile filtration. In case where a redundant filtration set-up is used, the samples shall be taken prior to the first filter. Procedures for taking samples shall be designed so as not to introduce contamination.
VII.4.7.16.Liquid sterilising grade filters shall be discarded after the processing of a single batch and the same filter shall not be used continuously for more than one working day, unless such use has been validated.
VII.4.7.17.Where campaign manufacture of a product has been appropriately justified in the contamination control strategy and validated, the manufacturer shall:
(a) assess and document the risks associated with the duration of filter use for the sterile filtration process for a given fluid;
(b) conduct and document effective validation and qualification studies to demonstrate that the duration of filter use for a given sterile filtration process and for a given fluid does not compromise the performance of the final sterilising grade filter or the filtrate quality;
(c) document the maximum validated duration of use for the filter and implement controls to ensure that filters are not used beyond the validated maximum duration. Records of these controls shall be maintained;
(d) implement controls to ensure that filters contaminated with fluid or cleaning agent residues or otherwise considered defective, are removed from use.
VII.5. Form-Fill-Seal (29)
VII.5.1.Form-Fill-Seal machines used for terminally sterilised products shall comply with the environmental requirements set out in Section VII.1.3 of this Annex, while Form-Fill-Seal machines used in aseptic manufacture shall comply with the environmental requirements set out in table 4 of this Annex.
VII.5.2.Contamination of the packaging films used during the Form-Fill-Seal process shall be minimised by the implementation of appropriate controls regarding components, supply and handling. Due to the criticality of packaging films, procedures shall be implemented to ensure that the films supplied meet defined specifications and are of the appropriate quality, including material thickness and strength, microbial and particulate contamination, integrity of printed information and packaging design, as relevant. The sampling frequency, the bioburden and, where applicable, endotoxin/pyrogen levels of packaging films and associated components shall be addressed as part of the contamination control strategy.
VII.5.3.The operation of the equipment, including set-up, filling, sealing and cutting processes shall be assessed so that critical process parameters can be identified, validated, controlled and monitored appropriately.
VII.5.4.Any product contact gases (e.g. those used to inflate the container or used as a product overlay) shall be appropriately filtered, as close to the point of use as possible. The quality of gases used and the effectiveness of the gas filtration systems shall also be verified periodically in accordance with Section V.4 of this Annex.
VII.5.5.The controls to be identified during the qualification of Form-Fill-Seal processes, which shall be part of the contamination control strategy, include but are not limited to:
— determination of the boundaries of the critical zone;
— environmental control and monitoring, both of the machine and of the background in which it is placed;
— personnel gowning requirements;
— integrity testing of the product filling lines and filtration systems (as relevant);
— duration of the batch or filling campaign;
— control of the packaging films, including any requirements for film decontamination or sterilisation;
— cleaning-in-place and sterilisation-in-place of the equipment as necessary;
— machine operation, settings and alarm management (as relevant).
VII.5.6.Critical process parameters for Form-Fill-Seal shall be established during the equipment qualification and shall include, but are not limited to:
— settings for uniform package dimensions and cutting in accordance with validated parameters;
— setting, maintenance and monitoring of validated forming temperatures (including preheating and cooling), forming times and pressures as relevant;
— setting, maintenance and monitoring of validated sealing temperatures, sealing temperature uniformity across the seal, sealing times and pressures as relevant;
— environmental and product temperature;
— batch-specific testing of package seal strength and uniformity;
— settings for correct filling volumes, speeds and uniformity;
— settings for any additional printing (batch coding), embossing or debossing to ensure that unit integrity is not compromised;
— methods and parameters for integrity testing of filled containers.
VII.5.7.Appropriate procedures for the verification, monitoring and recording of Form-Fill-Seal critical process parameters and equipment operation shall be implemented during production.
VII.5.8.Operational procedures shall describe how forming and sealing issues are detected and rectified. Rejected units or sealing issues shall be recorded and investigated.
VII.5.9.Appropriate maintenance procedures shall be established based on risks, and shall include maintenance and inspection plans for tooling critical to the effectiveness of unit sealing. Any issues identified that indicate a potential product quality concern shall be documented and investigated.
VII.6. Blow-Fill-Seal (30)
VII.6.1.Blow-Fill-Seal equipment used for the manufacture of products that are terminally sterilised shall be installed in at least a grade D environment. The conditions at the point of fill shall comply with the environmental requirements set out in Section VII.1.3 of this Annex.
VII.6.2.Where Blow-Fill-Seal equipment is used for aseptic processing, the following requirements shall apply:
(a) For shuttle type equipment used for aseptic filling, the parison (31) is open to the environment and therefore the areas where parison extrusion, blow-moulding and sealing take place shall meet grade A conditions at the critical zones. In addition, the filling environment shall be designed and maintained to meet grade A conditions for viable and total particle limits both at rest and when in operation.
(b) For rotary-type equipment used for aseptic filling, the parison is generally closed to the environment once formed and therefore the filling environment within the parison shall be designed and maintained to meet grade A conditions for viable and total particle limits both at rest and when in operation.
(c) The equipment shall be installed in at least a grade C environment, provided that grade A/B clothing is used. The microbiological monitoring (including setting of limits and frequencies applied) of operators wearing grade A/B clothing in a grade C area shall be performed in accordance with risk management principles.
VII.6.3.Due to the generation of particles from polymer extrusion and cutting during operation and the restrictive size of critical filling zones of Blow-Fill-Seal equipment, in operation monitoring of total particle for the equipment is not required. However, data shall be available to demonstrate that the design of the equipment ensures that critical zones of the filling process environment meet grade A conditions in operation.
VII.6.4.Viable environmental monitoring of Blow-Fill-Seal processes shall be risk-based and in accordance with Section VIII of this Annex. In operation viable monitoring shall be performed for the full duration of critical processing, including during equipment assembly, with the exception of rotary-type equipment where monitoring of the critical filling zone is not possible.
VII.6.5.The environmental control and monitoring programme shall take into consideration the moving parts and complex airflow paths generated by the Blow-Fill-Seal process and the effect of the high heat outputs of the process (e.g. through the use of airflow visualisation studies and/or other equivalent studies). Environmental monitoring programmes shall also consider factors such as air-filter configuration, air-filter integrity, cooling systems integrity, equipment design and qualification.
VII.6.6.Air or other gases in contact with critical surfaces of the container during extrusion, formation or sealing of the moulded container shall undergo appropriate filtration. The quality of the gas used and the effectiveness of the gas filtration systems shall be verified periodically in accordance with Section V.4 of this Annex.
VII.6.7.Particulate and microbial contamination of the polymer granulate shall be prevented by appropriate design, control and maintenance of the polymer granulate storage, sampling and distribution systems.
VII.6.8.The capability of the extrusion system to provide appropriate sterility assurance for the moulded container shall be validated. The sampling frequency, the bioburden and, where applicable, endotoxin/pyrogen levels of the raw polymer shall be defined and controlled.
VII.6.9.Interventions requiring cessation of filling and/or extrusion, moulding and sealing and, where required, re-sterilisation of the filling machine shall be clearly defined and described in the filling procedure, and included in the aseptic process simulation as relevant.
VII.6.10.The controls identified during qualification of Blow-Fill-Seal equipment shall be in alignment with the site’s contamination control strategy. Aspects to be considered include but are not limited to:
— determination of the boundaries of the critical zone;
— environmental control and monitoring, both of the machine and of the background in which it is placed;
— personnel gowning requirements;
— integrity testing of the product filling lines and filtration systems (as relevant);
— duration of the batch or filling campaign;
— control of polymer granulate, including distribution systems and critical extrusion temperatures;
— cleaning-in-place and sterilisation-in-place of equipment as necessary;
— machine operation, settings and alarm management (as relevant).
VII.6.11.Critical process parameters for Blow-Fill-Seal equipment shall be determined during equipment qualification and shall include, but are not limited to:
— clean-in-place and sterilisation-in-place of product pipelines and filling needles (mandrels);
— setting, maintenance and monitoring of extrusion parameters, including the temperature, speed and extruder throat settings for parison thickness;
— setting, maintenance and monitoring of mould temperatures, including the rate of cooling where necessary for product stability;
— preparation and sterilisation of ancillary components added to the moulded unit, e.g. bottle caps;
— environmental control, cleaning, sterilisation and monitoring of the critical extrusion, transfer and filling areas as relevant;
— batch-specific testing of the package wall-thickness at critical points of the container;
— settings for correct filling volumes, speeds and uniformity;
— settings for any additional printing (batch coding), embossing or debossing to ensure that unit integrity and quality is not compromised;
— methods and parameters for integrity testing of 100 % of all filled containers;
— settings for cutters or punches used to remove waste plastic surrounding filled units (flash removal).
VII.6.12.Appropriate procedures for the verification, monitoring and recording of Blow-Fill-Seal critical process parameters and equipment operation shall be implemented during production.
VII.6.13.Operational procedures shall describe how blowing, forming and sealing issues are detected and rectified. Rejected units or sealing issues shall be recorded and investigated.
VII.6.14.Where the Blow-Fill-Seal process includes the addition of components to moulded containers (e.g. addition of caps to large volume parenteral bottles), these components shall be appropriately decontaminated and added to the process using a clean, controlled process. The following shall apply:
(a) For aseptic processes, the addition of components shall be performed under grade A conditions, to ensure the sterility of critical surfaces, using pre-sterilised components.
(b) For terminally sterilised products, the validation of terminal sterilisation processes shall ensure the sterility of all critical product pathways between the component and moulded container, including areas that are not wetted during sterilisation.
(c) Testing procedures shall be established and validated to ensure the effective sealing of components and moulded containers.
VII.6.15.Appropriate maintenance procedures shall be established based on risk, including maintenance and inspection plans for items critical to unit sealing, integrity and sterility.
VII.6.16.The moulds used to form containers are considered critical equipment. Therefore, any changes or modification to moulds requires an assessment of finished product container integrity, and where appropriate having regard to the outcome of the assessment, shall be supported by validation. Any issues identified that indicate a potential product quality concern shall be documented and investigated.
VII.7. Lyophilisation (32)
VII.7.1.Lyophilisation is a critical process step and all activities that can affect the sterility of the product or material shall be regarded as extensions of the aseptic processing. In particular, the lyophilisation equipment and its processes shall be designed to ensure that product or material sterility is maintained during lyophilisation by preventing microbial and particle contamination between the filling of products for lyophilisation and the completion of lyophilisation process. The control measures shall form part of the contamination control strategy.
VII.7.2.The sterilisation of the lyophilizer and any associated equipment (e.g. trays, vial support rings) shall be validated and the holding time between the sterilisation cycle and use shall be appropriately challenged during the aseptic process simulation. The lyophilizer shall be sterilised regularly, based on system design. In addition, re-sterilisation shall be performed after maintenance or cleaning. Sterilised lyophilizers and any associated equipment shall be protected from contamination after sterilisation.
VII.7.3.Lyophilizers and any associated product transfer and loading/unloading areas shall be designed to minimise operator intervention as far as possible. The frequency of the lyophilizer sterilisation shall be determined based on the design and risks related to system contamination during use. Lyophilizers that are manually loaded or unloaded with no barrier technology separation shall be sterilised before each load. For lyophilizers loaded and unloaded by automated systems or protected by closed barrier systems, the frequency of sterilisation shall be justified and documented as part of the contamination control strategy.
VII.7.4.The integrity of the lyophilizer shall be maintained following sterilisation and during lyophilisation. The filter used to maintain the lyophilizer’s integrity shall be sterilised before each use of the system and the integrity testing results shall be part of the batch certification/release. In addition, the frequency of vacuum/leak integrity testing of the chamber shall be documented and the maximum permitted leakage of air into the lyophilizer shall be specified and checked at the start of every cycle.
VII.7.5.Lyophilisation trays shall be checked regularly to ensure that they are not misshapen or damaged.
VII.7.6.Points to consider for the design of loading (and unloading, where the lyophilised material is still unsealed and exposed), include but are not limited to:
— the loading pattern within the lyophilizer shall be specified and documented;
— the transfer of partially closed containers to a lyophilizer shall take place under grade A conditions at all times and handled in a manner designed to minimise direct operator intervention. Technologies such as conveyor systems or portable transfer systems (e.g. clean air transfer carts, portable unidirectional airflow workstations) shall be used to ensure that the cleanliness of the system used to transfer the partially closed containers is maintained. Alternatively, where supported by validation, trays closed in grade A and not reopened whilst in the grade B area may be used to protect partially stoppered vials (e.g. appropriately closed boxes);
— airflow patterns shall not be adversely affected by transport devices and venting of the loading zone;
— unsealed containers (such as partially stoppered vials) shall be maintained under grade A conditions and shall normally be separated from operators by means of a physical barrier technology or any other appropriate measures;
— where the seating of the stoppers is not completed prior to the opening of the lyophilizer chamber, the product removed from the lyophilizer shall remain under grade A conditions during subsequent handling;
— tools used during loading and unloading of the lyophilizer (e.g. trays, bags, placing devices, tweezers) shall be sterile.
VII.8. Closed systems
VII.8.1.The use of closed systems can reduce the risk of microbial, particle and chemical contamination from the adjacent environment. Closed systems shall be designed to reduce the need for manual manipulations and the associated risks.
VII.8.2.It is critical to ensure the sterility of all product contact surfaces of closed systems used for aseptic processing. Therefore, the design and selection of any closed system used for aseptic processing shall ensure maintenance of sterility. Connection of sterile equipment (e.g. tubing/pipework) used after the final sterilising grade filter shall be connected aseptically (e.g. by intrinsic sterile connection devices).
VII.8.3.Appropriate measures shall be put in place to ensure the integrity of components used in aseptic connections. The means by which this is achieved shall be determined and addressed in the contamination control strategy. In particular, appropriate system integrity tests shall be considered when there is a risk of compromising product sterility. Supplier assessment shall include the collation of data in relation to potential failure modes that may lead to a loss of system sterility.
VII.8.4.The background environment in which closed systems are located shall be determined having regard to the system’s design and the processes undertaken. For aseptic processing and where there is a risk that the system’s integrity may be compromised, the system shall be located in grade A. If the system can be shown to remain integral at every usage (e.g. via pressure testing and/or monitoring) then a lower classified area may be used. Any transfer between classified areas shall be thoroughly assessed in accordance with Section III.2 of this Annex. When the closed system is opened (e.g. for maintenance of a bulk manufacturing line), this shall be performed in a classified area appropriate to the materials (e.g. grade C for terminal sterilisation processes, or grade A for aseptic processing) or be subject to further cleaning and disinfection (and sterilisation in case of aseptic processes).
VII.9. Single use systems (33)
VII.9.1.Single use systems may be used in the manufacture of sterile products as an alternative to reusable equipment. Single use systems can be individual components or be made up of multiple components such as bags, filters, tubing, connectors, valves, storage bottles and sensors. Single use systems shall be designed to reduce the need for manipulations and complexity of manual interventions.
VII.9.2.There are some specific risks associated with single use systems that shall be assessed as part of the contamination control strategy, including but not limited to:
— the interaction between the product and product contact surface (such as adsorption, or leachables (34) and extractables (35));
— the fragile nature of the system compared with fixed reusable systems;
— the increase in the number and complexity of manual operations (including inspection and handling of the system) and connections made;
— the complexity of the assembly;
— the performance of the pre- and post-use integrity testing for sterilising grade filters;
— the risk of holes and leakage;
— the potential for compromising the system at the point of opening the outer packaging;
— the risk of particle contamination.
VII.9.3.Sterilisation processes for single use systems shall be validated and shown to have no adverse impact on the system’s performance.
VII.9.4.Assessment of suppliers of disposable systems including sterilisation is critical to the selection and use of these systems. Therefore, for sterile single use systems, verification of sterility assurance shall be performed as part of the supplier qualification and evidence of sterilisation of each unit shall be checked on receipt.
VII.9.5.The adsorption and reactivity of the product with product contact surfaces shall be evaluated under process conditions.
VII.9.6.The extractable and leachable profiles of the single use systems and any impact on the quality of the product – especially where the system is made from polymer-based materials – shall be evaluated. An assessment shall be carried out for each component to evaluate the extractable profile data. For components considered to be at high risk from leachables, including those that may absorb processed materials or those with extended material contact times, an assessment of leachable profile studies, including safety concerns, shall be taken into consideration. When applying simulated processing conditions, these shall accurately reflect the actual processing conditions and be based on a scientific rationale.
VII.9.7.Single use systems shall be designed to maintain integrity throughout processing under the intended operational conditions. Attention to the structural integrity of the single use components is necessary where these may be exposed to extreme conditions (e.g. freezing and thawing processes) during routine processing or transportation, including verification that intrinsic sterile connection devices (both heat sealed and mechanically sealed) remain integral under these conditions.
VII.9.8.Acceptance criteria shall be established and implemented for single use systems corresponding to the risks or criticality of the products and its processes. On receipt, each piece of single use systems shall be checked to ensure that they have been manufactured, supplied and delivered in accordance with the approved specification. A visual inspection of the outer packaging (e.g. appearance of exterior carton, product pouches), label printing, and review of attached documents (e.g. certificate of conformance and proof of sterilisation) shall be carried out and documented prior to use.
VII.9.9.Critical manual handling operations of single use systems such as assembly and connections shall be subject to appropriate controls and verified during aseptic process simulation.
SECTION VIII
ENVIRONMENTAL AND PROCESS MONITORING
VIII.1. General requirements
VIII.1.1.Each site shall have an environmental and process monitoring programme to monitor the controls designed to minimise the risk of microbial and particle contamination. The programme, which shall form part of the overall contamination control strategy, shall typically consist of the following elements:
— environmental monitoring – total particle;
— environmental and personnel monitoring – viable particle;
— temperature, relative humidity and other specific characteristics;
— aseptic process simulation (only for aseptically manufactured products).
VIII.1.2.The reliability of each of the elements of the monitoring system when taken in isolation is limited. Therefore, the outcome from the each of the elements above-described cannot be considered – on its own – as an indicator of asepsis. However, the results from all the elements of the programme help confirm the reliability of the design, validation and operation of the monitored system.
VIII.1.3.The information from the programme shall be used for routine batch certification/release and for periodic assessment during process review or investigation. While this applies to both terminal sterilisation and aseptic processes, it is acknowledged that the criticality of the impact may differ depending upon the product and process type.
VIII.2. Environmental and process monitoring
VIII.2.1.The purpose of the environmental monitoring programme is twofold:
— to provide assurance that cleanrooms and clean air equipment continue to provide an environment of appropriate air cleanliness, in accordance with design and regulatory requirements;
— to effectively detect excursions from environmental limits, which -in turn- shall trigger an investigation and an assessment of the risks to product quality.
Risk assessments shall be performed in order to establish a comprehensive environmental monitoring programme, including sampling locations, frequency of monitoring, monitoring methods and incubation conditions (e.g. time, temperature(s), aerobic and/or anaerobic conditions). In particular, the risk assessment shall include the determination of critical monitoring locations, those locations where the presence of microorganisms during processing may have an impact on product quality (e.g. grade A, aseptic processing areas and the grade B areas that directly interface with the grade A area).
The risk assessments shall be performed on the basis of the specific characteristics of the process inputs and the final product, the facility, the equipment, the criticality of specific processes and steps, the operations involved, the routine monitoring data, the monitoring data obtained during qualification and the knowledge of typical microbial flora isolated from the environment. Detailed knowledge of those aspects is therefore required for the establishment of the environmental monitoring program. Other relevant information such as air visualisation studies shall also be considered.
The risk assessments shall be reviewed regularly to confirm the effectiveness of the site’s environmental monitoring programme.
VIII.2.2.Routine monitoring of cleanrooms, clean air equipment and personnel shall be performed in operation throughout all critical stages of processing, including equipment set-up.
VIII.2.3.Other characteristics, such as temperature and relative humidity, shall be controlled within ranges that align with product/processing/personnel requirements and support the maintenance of the defined cleanliness standards (e.g. grade A or B).
VIII.2.4.The monitoring of grade A shall demonstrate the maintenance of aseptic processing conditions during critical operations. Monitoring shall be performed at locations posing the highest risk of contamination to the sterile equipment surfaces, the containers, the closures and the product. The selection of monitoring locations and the orientation and positioning of sampling devices shall be appropriate to obtain reliable data from the critical zones.
VIII.2.5.Sampling methods shall not pose a risk of contamination to the manufacturing operations.
VIII.2.6.Appropriate alert levels and action limits shall be set for the results of viable and total particle monitoring. The maximum total particle action limits are described in Table 5 and the maximum viable particle action limits are described in Table 6. However, more stringent action limits may be required based on data trending, the nature of the process or as determined in the contamination control strategy. Both viable and total particle alert levels shall be established based on results of cleanroom qualification tests and periodically reviewed based on ongoing trend data.
VIII.2.7.Alert levels for grade A (total particle only) grade B, grade C and grade D shall be set such that adverse trends (e.g. a number of events or individual events that indicate a deterioration of environmental control) are detected and addressed.
VIII.2.8.Monitoring procedures shall define the approach to trending. Trends shall include, but are not limited to:
— increasing numbers of excursions from action limits or alert levels;
— consecutive excursions from alert levels;
— regular but isolated excursion from action limits that may have a common cause (e.g. single excursions that always follow planned preventative maintenance);
— changes in microbial flora type and numbers and predominance of specific organisms. Particular attention shall be paid to organisms recovered that may indicate a loss of control, deterioration in cleanliness or organisms that may be difficult to control such as spore-forming microorganisms and moulds.
VIII.2.9.The monitoring of grade C and D cleanrooms in operation shall be performed on the basis on data collected during qualification and routine data to allow effective trend analysis. The requirements of alert levels and action limits will depend on the nature of the operations carried out. Action limits may be more stringent than those listed in Table 5 and Table 6.
VIII.2.10.If action limits are exceeded, a root cause investigation, an assessment of the potential impact to the product (including batches produced between the monitoring and the reporting) and implementation of corrective and preventive actions (as appropriate) shall be required.
If alert levels are exceeded, an assessment and follow-up is mandatory, including consideration of an investigation and/or corrective actions to avoid any further deterioration of the environment.
The above shall be reflected in operating procedures.
VIII.3. Environmental monitoring – total particle
VIII.3.1.A total particle monitoring programme shall be established to obtain data for assessing the potential contamination risks and to ensure the maintenance of the environment for sterile/aseptic operations in a qualified state.
VIII.3.2.The limits for environmental monitoring of airborne particle concentration for each graded area are given in Table 5.
| Grade | Maximum limits for total particle ≥ 0,5 μm/m3 | Maximum limits for total particle ≥ 5 μm/m3 | ||
|---|---|---|---|---|
| at rest | in operation | at rest | in operation | |
| A | 3 520 | 3 520 | 29 | 29 |
| B | 3 520 | 352 000 | 29 | 2 930 |
| C | 352 000 | 3 520 000 | 2 930 | 29 300 |
| D | 3 520 000 | not pre-defined (1) | 29 300 | not pre-defined (1) |
| (1) For grade D, in operation limits are not predetermined. The manufacturer shall establish in operation limits based on a risk assessment and on routine data, where applicable. Note 1: The particle limits given in the table for the ‘at rest’ state shall be achieved after a short ‘clean up’ period defined during qualification (guidance value of less than 20 minutes) in an unmanned state, after the completion of operations. Note 2: The occasional indication of macro particle counts, especially ≥ 5 μm, within grade A may be considered to be false counts due to electronic noise, stray light, coincidence loss etc. However, a consecutive or regular counting of low levels may be indicative of a possible contamination event and shall therefore be investigated. Such events may be indicative of an early failure of the room air supply filtration system, an equipment failure, or may also be a signal of poor practices during machine set-up and routine operation. |
VIII.3.3.For grade A, particle monitoring shall be undertaken for the full duration of the critical processing, including equipment assembly.
VIII.3.4.The grade A area shall be monitored continuously (for particles ≥ 0,5 and ≥ 5 μm) and with a suitable sample flow rate (at least 28 litres (1 ft3) per minute) so that all interventions, transient events and any system deterioration is captured. The system shall frequently correlate each individual sample result with alert levels and action limits at such a frequency that any potential excursion can be identified and responded to in a timely manner. Alarms shall be triggered if alert levels are exceeded. Procedures shall define the actions to be taken in response to alarms including the consideration of additional microbial monitoring.
VIII.3.5.It is recommended that a similar system be used for the grade B area although the sample frequency may be decreased. The grade B area shall be monitored at such a frequency and with a suitable sample size to ensure that the programme captures any increase in levels of contamination and a system deterioration. If alert levels are exceeded, alarms shall be triggered.
VIII.3.6.The selection of the monitoring system shall take into account any risk presented by the materials used in the manufacturing operation (e.g. those involving live organisms, powdery products or radiopharmaceuticals) that may give rise to biological, chemical or radiation hazards.
VIII.3.7.In case where contaminants are present due to the processes involved and can potentially damage the particle counter or present a hazard (e.g. live organisms, powdery products and radiation hazards), the frequency and strategy employed shall be adequate to ensure the environmental classification, both prior to and post exposure to the risk. An increase in the monitoring of viable particle shall be considered to ensure a comprehensive monitoring of the process where appropriate. Additionally, monitoring shall be performed during simulated operations at appropriate intervals. The defined approach is part of the contamination control strategy.
VIII.3.8.The size of the monitoring samples taken using automated systems will usually depend on the sampling rate of the system used. It is not necessary for the sample volume to be the same as that used for formal classification of the cleanrooms and of the clean air equipment. The monitoring sample volumes shall be justified.
VIII.4. Environmental and personnel monitoring – viable particle
VIII.4.1.Frequent microbial monitoring using a combination of methods such as settle plates, volumetric air sampling, glove, gown and surface sampling (e.g. swabs and contact plates) shall be required where aseptic operations are performed. Specifically:
— Viable particle monitoring shall be performed within the cleanrooms when normal manufacturing operations are not occurring (e.g. post disinfection, prior to start of manufacturing, on completion of the batch and after a shutdown period) and in associated rooms that have not been used, in order to detect potential incidents of contamination that may affect the controls within the cleanrooms. In case of an incident, additional sample locations may be used as a verification of the effectiveness of a corrective action (e.g. cleaning and disinfection).
— Continuous viable air monitoring in grade A (e.g. air sampling or settle plates) shall be performed for the full duration of critical processing, including equipment assembly (aseptic set-up) and critical processing. A similar approach shall be considered for grade B cleanrooms based on the risk of impact on the aseptic processing. The monitoring shall be performed in such a way that all interventions, transient events and any system deterioration are captured and any risk caused by interventions of the monitoring operations is avoided.
The method of sampling used shall be justified as part of the contamination control strategy and be demonstrated not to have a detrimental impact on grade A and B airflow patterns. Cleanroom and equipment surfaces shall be monitored at the end of an operation.
VIII.4.2.Monitoring of personnel shall be conducted on the basis of a risk assessment, which shall evaluate the locations, type and frequency of monitoring based on the activities performed and the proximity to critical zones. Microbial monitoring of personnel in the grade A and grade B areas is essential. Where operations are manual in nature (e.g. aseptic compounding or filling), an enhanced emphasis shall be placed on microbial monitoring of gowns and the implemented monitoring measures shall be justified within the contamination control strategy.
VIII.4.3.Monitoring shall include the sampling of personnel at periodic intervals during the process. Sampling of personnel shall be performed in such a way that it does not compromise the process. Particular consideration shall be paid to the monitoring of personnel following involvement in critical interventions (as a minimum gloves, but other parts of gown may also need to be monitored as applicable to the process) and on each exit from the grade B cleanroom (gloves and gown).
VIII.4.4.Where monitoring of gloves is performed after critical interventions, the outer gloves shall be replaced prior to continuation of activity. Where monitoring of gowns is required after critical interventions, the gown shall be replaced before further activity in the cleanroom.
VIII.4.5.Regular oversight by the quality unit is required if monitoring is routinely performed by manufacturing personnel.
VIII.4.6.The adoption of suitable alternative monitoring systems such as rapid methods may be considered by manufacturers in order to expedite the detection of microbiological contamination issues and to reduce the risks to the product. These rapid and automated microbial monitoring methods may be adopted after validation has demonstrated their equivalency or superiority to the established methods.
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