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Monday, August 10, 2009

STERILTY TESTING SOP STERILE PHARMACEUTICAL TOPICAL PREPARATIONS

SOP for sterlity testing for steril pharmaceutical eye drops and ointments 

OBJECTIVE :- To check whether the product to be tested is sterile

UNIT OF RESPONSIBILITY
:- QA Executive and trained QA assistants

IMPORTANT POINT/ NOTE/ PRECAUTIONS :-

1 Follow proper entry/exit procedure

2 Follow proper aseptic technique while testing the sample

3 Do not work in laminar air flow unit when UV lamp is on
MATERIAL :-

a) Media : 1) Sterile Soybean Casien Digest Medium with ploysorbate 80 ( if specified in monograph

2) sterile Fluid Thioglycollate Medium with ploysorbate 80 (if specified in monograph)

b) Sterile Isopropyl Myristate obtained by filteration through 02 u membrane filter

c) 01 % w/v sterile peptone water with 2 % v/v polysorbate 80 (if specified in monograph)

d) Manifold unit for 3 or more holder membrane filter funnel with suitable vaccum flask

e) Sterilised membrane filter of 045 u and 02 u

f) Sterile forceps and scissors wrapped in parchment paper/butter paper

g) Sterile 10 ml pipettes

h) sterile empty flasks

PROCEDURE :-


I) METHOD:-

a) MEMBRANE FILTERATION METHOD :-

1 Follow the entry procedure in the aseptic area as mentioned in the SOP
2 Dissolve not less than 1 gm of ointment from not less than 20 tubes/ containers in not
less than 100 ml of sterile isopropyl myristate with a pH of water extract not less than 65, which previously has been rendered sterile by filteration through 02 u membrane filter
3 Warm the sterilised solvent, and if necessary the test material, to not more than 44 degC
( just prior to use)

4 Swirl the flask to dissolve the topical preparation, taking care to expose a large surface of the material to the solvent

5 Following dissolution aseptically transfer the mixture in membrane filter funnel and immediately pass the mixture through the membrane filter of 045 u with the aid of vacuum

6 Keep the filter membrane covered with liquid throughout the filteration for maximum efficiency of the filter Following filteration of the liquid wash the membrane filter with 300 ml of sterile 01 % w/v peptone water

7.Aseptically remove the membrane filter from the filter holders, cut the membrane in two halves immerse one half of the membrane filter with the aid of sterile forceps in 100 ml of Soybean Casien Digest Medium and the other half in 100 ml of sterile Fluid Thioglycollate Medium

8 Keep the positive control by inoculating the culture in Fluid thioglycollate medium and Soybean casien digest medium

9 Keep the negative control by filtering autoclaved 100 ml of 01%w/v peptone water,cut the membrane filter in two halves & immerse one half of the filter in Soybean casien Digest Medium and the other half in sterile Fluid Thioglycollate Medium

10 Incubate the tubes at 20 - 25c and 30 - 33c for growth of fungus and bactreia respectively for not less than 7 days as per IP and 14 days as per USP and BP
11Observe the tubes for growth in terms of turbidity daily till the completion of 7 days as per IP and 14 days as per USP & BP
12 If none of the tubes shows the growth in terms of turbidity within 7days as per IP and as per USP and BP within 14 daysof observation then the test passes for sterility

14 If any of the tubes shows growth in terms of turbidity than preserve the tubes and carry
out the investigation by identifying the organism
15 Repeat the test using double quantity of tubes/ containers and if no growth observed the
test passes for sterility and if growth observed then the test fails for sterility

b) DIRECT INOCULATION METHOD :-

1 Quantity to be used is 1 to 10 gm

2 Quantity to be used for each culture medium 05 to 10 gm

3 Follow the entry procedure in the aseptic area as mentioned in the SOP NO QA/SOP/304/98

4 Take about 20 Samples of topical preparation and take out about 05 gm from each sample into a flask containing 100 ml of 01 % w/v sterile peptone water with 2 % w/v polysorbate 80 and mix the ointment by swirling the flask

5 Transfer aseptically 10 ml of the diluted media to each 100 ml of sterile Soybean casien Digest Medium and 100 ml of sterile Fluid Thioglycollate medium Incubate the media at 20 - 25 deg C and 30 -35 deg C for not less than 14 days for fungus and bacteria respectively

6 Observe the tubes for growth in terms of turbidity daily till the completion of 14 days

7 Keep the positive control by inoculating the culture in Fluid thioglycollate medium and Soybean casien digest medium and Negative control by inoculating an autoclaved 01% peptone water

8 If none of the tubes shows the growth in terms of turbidity within days of observation then
the test passes for sterility

9 If any of the tubes shows growth in terms of turbidity than preserve the tubes and carry
out the investigation by identifying the organism

10 Repeat the test using double quantity of tubes/ containers and if no growth observed the
test passes for sterility and if growth observed then the test fails for sterility

II) GROWTH PROMOTION TEST:-

To each autoclaved load of each lot of medium for its growth promotion qualities by
seperately inoculating test containers of each medium with about 100 vaible micro
organisms of each strain as listed below and incubating according to the conditions
specified The test media are satisfactory if clear evidence of growth appears in all
inoculated media within 7 days as per IP and 14 days as per USPand BP
If the freshly prepared media are not used within 2 days then store them in dark
preferably at 8 - 15 deg CThe sterility is considered invalid if the test medium
shows inadequate growth response
Medium -----------------------------------Test Microorganism ----------Temp------- Condition
Fluid Thioglycollate Medium ------B.subtilis ATCC 6633/S.aereus NCTC 7447---30-35 C-- -Aerobic/Anaerobic
Fluid Thioglycollate Medium ------ -----S.aereus NCTC 7447------------------------30-35 C-- --Aerobic
Soybean Casien Digest Medium -------C.albicans ATCC 10231-----------------------30-35 C-------Aerobic
III) PRODUCT POSITIVE CONTROL TEST :-

After tge sterility testing is over, add to each media tubes of the particular batch tested
around 100 cfu/ml of B subtilis ATCC 6633 and C albicans ATCC 10231 resp in
Fluid thioglycolate medium and Soybean casien Digest medium, incubate furthur for
minimum of 24 hrs at 30-35 deg C and 20-25 deg C resp Observe for growth in the
inoculated media tubes to verify that the media does not contain bacteriostatic or
fungistatic activity

IV) DESTRUCTION :- After the testing is over the media is destroyed by autoclaving in a pressure cooker
for 20 minutes This destroyed media is discarded into the drain
The samples after testing are destroyed by sweezing the ointment out and incernating it
All the observations , results and destruction records are recorded in the attachment 1

PERSONNEL TO BE REPORTED IN CASE OF NON COMPLIANCE :- Department head

Production Manager


FREQUENCY OF OPERATION :- Batchwise
SOP for microbiology in pharmaceuticals manufacturing

Following are some of articles which will be useful for you in further understanding of aspects of sterile dosage form manufacturing

Pharmaceutical Validation

Cleen Room Classification

Pharmaceutical Industry

Pharmaceutical Manufacturing

Clinical Trials

Media fill

Pharmaceutical companies

Regulatory Affairs in Pharmaceutical Industry


Determination of Phenol coeeficient of a disinfectant

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