Methods for sterility testing

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Membrane filtration method1,2:

General procedure for all formulation--

How to prepare sample for different dosage forms (membrane filtration):


Solutions: Solution can directly take as a sample most commonly.

Suspensions: Add sufficient quantity of fluid A for rapid filtration.

Oils and oily solutions: if less viscous then directly filter without dilution and if more viscous then dilute with proper diluents.

Ointments and creams: prepare 1%w/v with suitable diluents with the help of heat if required but NMT 40@C.

Sterile Devices: take the sample and shake it for 10 min with 50 ml of suitable broth then instantly pass the mixture through membrane filter which is previously moisten with liquid. Membrane is wash with broth 50 ml for 3-4 times and incubates the test media.

There are some special procedure for surgical cotton, and suture/ligature etc.

Direct inoculation method1,2:

General Procedure--


Observation and interpretation of results1:

Incubation temperature

EP specified 30-32@C for bacteria and 20-25@C for fungi.

USP specified 30-35@C for bacteria and 20-25@C for fungi.

Incubation time

EP specified at least 7 days unless and until specified in any official monograph.

USP specified at least 7 days for steam sterilized product and sterilized by membrane filtration method, but at least 14 days for product sterilized by other methods.

Frequency of observation

Every day; stir the test tube and then observe.

Interpretation of results

  • If no growth or contamination batch passes test.
  • If growth observed and authority will permit to repeat the test then repeat test with greater care.
  • If again growth observed in repeat test, authority will allow repeating unless same microorganism is observe in the two tubes.
  • Generally, one repeat test is allowed for bulk sample and two repeat tests are allowed for final container sample according to WHO.
  • If 2 or more/3 or more and any growth observed in 1st , 2nd, 3rd test respectively then batch will considered an unsuitable (fails sterility test).

References:

  1. Cooper and Gunn's Dispensing Pharmacy. CBS publisher, New Delhi, 12 th Ed: 541-572.
  2. Indian Pharmacopoeia. Govt. of India, ministry of health and family welfare. India, 2007: 53-59.

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About the Author

P.V.ABHIGNA's picture
Author: P.V.ABHIGNA

Comments

Siva Mavuduru's picture

Hi Abhigna What are the materials used for preapartion of membrane filters?
P.V.ABHIGNA's picture

Dear Siva, Here some membrane filters which are composed from various material. Please check PES (Polyethersulfone) membrane Mixed cellulose ester (MCE) membrane filter which compose of cellulose acetate and nitrate. PTFE (polytetrafluorothylene) membrane filter (preffered one) Nylon Membrane Filter Polypropylene(PP) membrane filter CA (Cellulose Acetate) membrane filters and many other types of materials are used. For details please refer following: http://www.membrane-solutions.com/grid_membrane.htm (accessed on 6 Sept' 2010)

ABHIGNA.P.V.

Mrudula Boyapati's picture

hiii will fluid thioglycolate medium support the growth of all kinds of bacteria i.e both aerobic and anerobic. Then why is alternate fluid thioglycolate medium in use...?? regards MRUDULA
P.V.ABHIGNA's picture

Dear Madam, Fluid thio glycollate contains agar. But in alternate fluid thioglycollate medium does not contains agar and thus is it more suitable for the growth of aerobes than fluid thioglycollate medium which has more viscosity, so proper and sufficient aeration will not get. Regards

ABHIGNA.P.V.

TK Indira's picture

Good presentation. Can you please mention the pore size of membrane filters???

T.K. Indira. http://www.pharmainfo.net/tkindira

-- "Our greatest glory is not in never falling, but in rising every time we fall..." Team 'Char'minar.

P.V.ABHIGNA's picture

Dear Indira, There are different pore sizes of membrane filters ranging from 0.2micron to several mm but for bacteria retaintion and sterility testing generally 0.22 micron size is used with diameter 47mm (typical size: sizes also vary based upon need) Thanks

ABHIGNA.P.V.

Santosh kumar. JH's picture

Abhigna, 1. The text of the pictures are difficult to read. 2. How can we ensure that these membrane filters are sterile?? 3. Also pls throw some light on membrane pore size and materials used for preparation of membrane filters.
P.V.ABHIGNA's picture

Dear Santosh, 1.Tex of the picture I tried to make it clear previously. Again I will try to do so that it will be good to read. 2.these filters are sterile preparation and manufacturer ensured for sterility testing while preparing. We are the customer for this article. Simillarly, patient is not going to check the sterility of parentral formulation as he know it is already tested by manufacturer. Similarly here we have to rely. or else if you wish to check. take a minimum samples for the lot what you are purchased and do the same procedure as given in membrane filtration method without prapartion. if growth appear u can say membrane filter is not sterile. It is the simplest method if customer like us want to check but after checking one can't use the same filter. so, proper sampling tech u have to do and u have to rely on these samples as a representative for batch. Hope you got. 3. for pore size pls refer comments Regards,

ABHIGNA.P.V.

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