VALIDATION OF ASEPTIC FILLING PROCEDURE WITH MEDIA FILL
Definition of Media Fill and Requirements of the Guidelines: – According to all guidelines the process simulation with media fill is state of the art for the validation of aseptic manufacturing process. Media fill means that a microbiological nutrient media will be filled into a container closure system (ampule, vials etc) instead of the product under simulation of aseptic standard procedure. The filled container closure systems are incubated under defined parameters and finally checked for microbiological contamination. This is to demonstrate that rooms, equipment and personnel are able to manufacture a product with very low contamination rate.
All manufacturing procedures in pharmaceutical industry must be validated. This requirement is stated in the European Pharmacopoeia:18 “Process validation include checks on the process are regularly carried out by means of process simulation tests using microbial growth media which are then incubated and examined for microbial contamination (media fill tests).“
The EU GMP Guide11 provides more details on this issue: – “Validation of aseptic processing should include a process simulation test using a nutrient medium (media fill) … The process simulation test should imitate as closely as possible the routine manufacturing process and include all the critical subsequent manufacturing steps.“
DESCRIPTION OF MEDIA FILL
Choice of the Media Fill:- A microbiologically appropriate nutrient is to be used. Within pharmaceutical industries Soya-bean casein digest medium (aerobe microorganisms) and thioglycollate (anaerobe microorganisms) have been verified. Both media relate to the quality control of pharmaceutical medicinal products because of their use in sterility testing. The growth
promotion proprieties for a lot of microorganisms such as bacteria, yeasts and moulds have been proved very often. The anaerobe simulation is restricted for filling lines which are used for products filled in an atmosphere where oxygen is excluded. The absence of germs of Bovine Spongiforme Encephalopathy (BSE) is to be mandatorily demonstrated in case of every nutrient.
Media Fill and Filling Volume:- Some points regarding the filling volume have to be considered:-
- all surfaces and stoppers must be wetted;
- microbiological growth must be ensured;
- this means that at least half of the nominal volume of the container should be used,
- the filling volume has to be taken into account the physiological preconditions of the microorganisms
- [Aerobic conditions: half the nominal volume; anaerobic conditions: full nominal volume].
Preparation of Media Fill:- The nutrient medium must be processed, handled, and filled in a manner that precisely simulates the normal manufacturing process. Thus, the normal manufacturing process has to be analysed. In media fill validation, dried nutrient medium base and Water for Injection (WFI) are used as the compounding starting materials. Bioburden counts and determinations of endotoxin content, as well as analysis of pathogenic contaminants, for e.g. Pseudomonas aeruginosa or Echerichia coli, are performed as in-process controls on the raw materials. Media fill should be prepared considering the instructions of the manufacturer regarding the usual manufacturing process (for example using the sterile filter systems as appropriate).
The preparation should be performed analogously as the drug product (facility, monitoring
activities, equipment, materials, personnel).
Environmental monitoring, comprising:-
- airborne counts,
- particle counts, and
- hygiene status of personnel and materials.
- Prior to filtration, the pH-value of the nutrient broth is checked and in-process controls (IPC) on identity, clarity, and bioburden are conducted.
- Samples are controlled for analytical and microbiological controls like that of any other product.
- Holding and process times are documented and may be prolonged for validation purposes.
- The bulk solution is sterile-filtered using the same filter material as in normal aseptic processing.
- Filter integrity is checked prior to and after use.
- Environmental monitoring is conducted at this processing step.
- Prior to filling, primary containers are sterilised and depyrogenized, the filling line is cleaned and sterilised (CIP/SIP) or transfer lines and dosage pumps are sterilised separately.
- Environmental monitoring is performed during the filling procedure.
- A change in personnel is carried out during filling, as are several, pre-defined investigation and in process controls, for e.g. filling volume.
Worst Case Simulation:- The simulation should consider such conditions which simulate the highest risk (worst case) of maximum expected and permitted loads. Examples for worst case conditions are defined in ISO 13408.28 PIC/S26 requires simulation of all interventions which may occur during a shift (refilling of closures, adjustments of filling needles) Not only should these kinds of interventions be regarded, but also their frequency (FDA Draft Guidance12). For vial dimension and filling speed the worst condition is the biggest vial with the longest filling time, the widest-neck vial and the smallest vial with the highest speed.
Interventions:- All interventions and measures of the usual process should be simulated in media fill. For example manual control of the filling volume, interventions in class A / 100, change in personnel, performance of environmental monitoring. Even technical interruptions should be considered (lack of air system, stopping of the machine). The approach is done during risk analysis with cooperation from the following departments: production, quality, validation, quality assurance.
Number of Filled Units and Duration of the Filling Process:- According to FDA Guidance24 the minimum number of filled units of media fill is 3,000 units (to reach a confidence level of 95% for demonstration of a contamination rate of less than 0.1%). For batch sizes smaller than 3,000 units, smaller numbers are acceptable (requirements are given in ISO 1340828 and EU GMP Guide Annex 111). For small batch sizes (for example products used for clinical trials) at least the actual batch size should be simulated during media fill. For very large batches, it is recommended to simulate media fill with 1% till 10% of the actual daily batch size. The vials with the smallest and the biggest size should be regarded in media fill. The units in media fill shall be enough to simulate worst case conditions.
Duration of Process, Holding Times and Stopping Times:- Time limits should be established for each phase of aseptic processing. Time limits should include for example the period between the start of bulk product, compounding and its filtration, filtration processes, product exposure while on the processing line, and storage of sterilised equipment, containers and closures. Bioburden and endotoxin load should be assessed when establishing time limits for stages such as formulation processing stage. The total duration of the procedure consists of the time needed for the preparation of the bulk, time between the beginning of the preparation and the end of the sterile filtration. PIC/S26 recommendation gives information about the duration of a media fill run. The whole filling time should be simulated, but it is possible to stop the machine to avoid excessive numbers of filled units. PIC/S26 recommends simulating the process per shift and per filling line and not only per filling line. This is integrated in the EU GMP guide Annex 111 and also in the FDA Draft Guidance 2003.
Holding times (e.g. materials, bulk, equipment) up to the beginning of the filling process etc. are usually defined within the manufacturing process. The validation of such times is performed during media fill and then simulates worst case conditions. For small batch sizes (for example products for clinical trial) at least the actual batch size should be simulated during media fill (GMP Guide Annex 111). ISO 13408-128 requires at last 5,000 units for the primary qualification of the process with three or more runs. The number of units for media fill is increased in FDA draft Guidance12 (at least 5,000 units) compared with the current FDA Guidance.
Selection of Units:- It is recommendable to incubate all units of media fill. In any case the thorough documentation of all filled units is necessary. It is possible to select damaged units prior to incubation according to routine processing. But the accurate reconciliation of all units is a general requirement. It is not acceptable to select positive units after incubation because the checking reveals defects for example in the container closure system. The FDA draft Guidance12 clarifies that intervention in the aseptic manufacturing process during media; that is to say an interruption of the aseptic barrier does not mean that those units have to be incubated, but it must be assured (SOP) that during routine manufacturing process such units are rejected.
Although no guideline mentions that the samples for fertility testing should not be taken prior to incubation of media fill, it is recommended to perform the fertility test after the evaluation of the media fill.
Acceptance Criteria in Media Fill
Warning Limits and Action Limits:- All guidelines regard any positive unit during media fill as a potential problem and the source of contamination should be evaluated thoroughly. ISO 13408-128 contains a table which gives a statistical approach of the evaluation of media fill. Warning limits and action limits have to be defined. Acceptance criteria for the successful validation of an aseptic manufacturing process is a contamination rate of 0.1% in at least 3,000 filled units with a confidential level of 95%. The sterilisation assurance level SAL is 10-3. The current discussion of the contamination rate aims at negative detection of contaminated units. A contamination rate of 0.1% will no longer be tolerated by FDA’s inspectors. Any positive unit has to be examined thoroughly and could be a reason for the failed media fill. FDA’s acceptance of this probability in test results (0.1% contamination rate) does not mean that an aseptically processed lot of drug product purporting to be sterile may contain one non-sterile unit per thousand count. According to PIC/S26 the contamination rate should be ideally zero but the statistic approach refers to ISO 13408-1.
Action Limits / Procedure in the Case of Failed Simulations:- Measures for analysing the cause of contamination and an investigation thereafter have to be established. On exceeding the action limit, a requalification is immediately required. According to ISO 13408-128 an investigation should be performed in case of exceeding the warning limit (1 contaminated unit up to 10, 250 units) and the run has to be repeated. If the warning limit is exceeded again, it implies that the media fill has failed and the complete primary qualification has to be repeated (three consecutive runs of media fill must be successful). In the case of requalification (usually every 6 months one successful media fill) exceeding of the warning limit in two consecutive runs has to be evaluated as exceeding the action limit. This is clearly said in the ISO 13408-128 and in PIC/S:26 “Exceeding the action limit means that a thorough investigation into the failure has to be performed and a complete requalification must be initiated. All produced batches since the failure must be quarantined until the cause for failure of the media fill is identified.” PIC/S26 recommends that all produced batches since the last successful process simulation have to be taken into account. Table below illustrates the limits of first qualification and requalification in media fill.
First Qualification and Requalification: Warning and Action Limits in Media Fill According to ISO 13408-128
In case of any contamination, it is necessary to identify the corresponding microorganism. The FDA Draft Guidance12 recommends the following:-
- 5,000 – 10,000 filling units in media fill: 1 contaminated unit (investigation required including consideration of a repeat media fill)
- < 10,000 filling units in media fill: 1 contaminated unit (investigation required 2 contaminated units are considered as cause for revalidation following investigation
Incubation Conditions: – The container filled with media fill shall be incubated for 14 days at temperatures which allow the growth of a wide microbiological spectrum. The FDA Draft Guidance12 recommends a temperature range from 20 to 35 °C and maintenance within 2.5 °C of the target temperature. A link to sterility testing is recommendable so as to have a rationale for the choice of these temperatures. Usually the containers are stored for 7 days at 20 to 25 °C followed by 7 days at 30 to 35 °C (the temperatures are defined in the European Pharmacopoeia). The incubation conditions have to be documented. (PDA32) The change in incubation temperature requires a change in storing of the filled containers. Here, during change in storing it is recommendable to inspect the containers and to wet the whole surface of the container with media fill.
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