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Guidelines for sterility testing of therapeutic goods

13 September 2006

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6. Media for use in sterility testing

600. Relevant section of the BP/Ph Eur: 'Culture Media and Incubation Temperatures'

Composition

601. Fluid Thioglycollate Medium (Medium 1) and Soybean-Casein Digest Medium (Medium 2) are the media generally used for tests for sterility. The composition of these media should be as specified in Annex II. Commercially available dried media, which differ slightly from the specified composition, may be used provided the reconstituted medium has been shown to support the growth of aerobic and anaerobic bacteria and fungi, as described in clauses 611-
615.

602. As noted in clause 402, alternative media types may be appropriate where the nature of the product or method of manufacture may result in the presence of fastidious organisms (eg, vaccines, blood products).

603. Inactivators of antimicrobials may be incorporated into culture media or solutions if indicated by validation studies.

604. Media may be either purchased from an approved supplier or manufactured in-house.

Method of preparation

605. Media  should  be  prepared  according  to  written  procedures  that  are  based  on  a validated sterilisation process.

606. Every batch should be assigned a unique batch number and expiry date and its manufacture documented.

607. Detailed guidance for preparation of Medium 1 and Medium 2 is provided in Annex II.

Tests on the media

608. Before use, each batch of the sterilised media should be tested for pH, sterility and growth promotion.

609. Measure the pH of a sample of each batch of medium. Any batch of medium of pH not within the range prescribed in Annex II for that medium should not be used in tests for sterility.

610. To check for sterility, incubate the media at 30 - 35°C and 20 - 25°C for 14 days. This testing may be performed on 100% of the batch or on representative portions and may be conducted concurrently with the product sterility test. Media which contain visible particulate matter should not be used in tests for sterility. Parametric release of media may be permitted if media is manufactured under the same conditions and controls as products already approved by the competent authority for parametric release (see also clause 708).

611. The ability of media to support the growth of micro-organisms should be tested by addition of small numbers of challenge organisms (see Annex I Guidance on Obtaining Small Numbers of Vegetative Organisms and Spores). After each batch of medium has been sterilised at least two vessels are selected from positions in the steriliser load where the exposure to heat is likely to be maximal.

612. Where the medium under examination is Medium 1 add not more than 100 viable spores/CFU of each of the following species of micro-organisms: Clostridium sporogenes ATCC 19404, Staphylococcus aureus ATCC 6538 and Pseudomonas aeruginosa ATCC 9027.

613. Where the medium under examination is Medium 2 add not more than 100 viable cells of Candida albicans ATCC 10231, Bacillus subtilis ATCC 6633 and Aspergillus niger ATCC 16404 to selected vessels.

614. Other appropriate strains of challenge organisms are listed in Table 5. It is recommended that from time to time the routine strains should be supplemented by using strains collected from the manufacturing environment.

615. Growth of the challenge micro-organisms should be apparent within 48 hours. If growth is not conspicuous in each of the incubated vessels within 3 days in the case of bacteria or 5 days in the case of fungi, the batch should not be used in tests for sterility.

616. Prepared media purchased from external vendors should be accompanied by certification of the growth promotion test performed on each batch of media. The test need not be performed by the sterility testing laboratory provided there is documented control over the conditions used to transport media between the media manufacturer and the testing laboratory.

Shelf-life

617. Media may be stored at 2 – 25°C in suitable sealed containers but must not be used after storage periods that have not been validated. It should be tested for growth promotion every three months.

618. Medium 1 of which more than the upper one-half is pink in colour should not be used in tests for sterility. Vessels of this medium which have become excessively pink may be heated once only in a steam bath, or in freely flowing steam, until the pink colour disappears.

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