How To Test Filter For Sterility

1. Purpose:-

This process describes the method for sterility test

ii. Telescopic:-

This test is performed to ensure the sterility of the sterile raw materials, sterile products, sterile packaging materials and all other materials which are claimed as sterile.

iii. HSE Statement:-

Proper gowning technique should be followed.

iv. Responsibilities:-

Director Quality Control is responsible to ensure that process & formats are followed entirely every bit approved.

ii)

Microbiologist is responsible to perform the test.

iii)

Lab bellboy is responsible to assist the microbiologist for the training of test

5. Materials Required:-

5.ane Membrane Filters:

Membrane filters should be compatible to the materials to be tested.

Pore size is 0.45 μm and the diameter 47 or l mm. They are individually packed and sterilized. The edge of the filters is hydrophobic. Cellulose nitrate filters are ordinarily used in sterility test.

v.2 Sterility Testing Apparatus:

A suitable 3 or 6 manifold membrane filter unit (e.grand. Millipore, Sartorius or equivalent properly wrapped and sterilized) is used with detachable funnels, filter holders and funnel covers.

5.3.1

Fluid Thioglycollate Medium (FTM)

Sterilized and pre-incubated for 2-5 days at 30-35^OC temperature, dispensed in 100 ml quantities

in suitable glass containers.

v.iii.two Soybean Casein Assimilate Medium (SCDM) (TSB)

Sterilized and pre-incubated for 2-5 days at 20-25^OC or at room temperature, dispensed in 100 ml quantities in suitable glass containers.

5.four Diluting Fluids:

five.4.1 Fluid 'A' (Sterilized and Pre-Incubated at 22-30^oC for ii-v Days)

Dissolve 1g of peptic assimilate of animal tissue in water to make i liter, filter or centrifuge to clarify if necessary, and adjust to a pH of vii.1and dispense into containers and sterilize using a validated process.

5.4.one.1 Preparation for Penicillin and Cephalosporin

Aseptically add together to above preparation, if necessary a quantity of sterile Pencillinase enzyme sufficient to inactivate any residual activity on the membranes later on solution of test specimen has been filtered.

5.4.2 Fluid 'D' (Sterilized)

This fluid is used to inactivate preservatives and when production contains oil or lecithin. It contains 0.1% polysorbate 80 and 0.1% peptone.

5.5

General Requirements:-

5.5.1 Sterilized medium size spatulas for measuring vi grams in instance of bulk powder and pocket-sized size spatulas for measuring 300 mg for filled units.

five.v.two Spatulas and scissors wrapped as pair wise in aluminum foil, dry heat at 200^o C for a maximum of 2 hours.

v.5.three Disinfectant: 70% alcohol filtered, five% phenol, three% Savlon, v-ten % Chlorine bleach solution, 4th Ammonium compounds or equivalent.

5.5.four

Sterilized lint free wipes.

5.v.five

Suitable sized sterilized Due south/S containers with Due south/S covers for dumping of 20 units to exist tested.

5.5.6

S/S bowls for disinfectant.

5.5.7

Filled unit opening device.

5.5.8

Sterilized de-capper or vial opener in case of vials or, sterilized ampoule cutter.

v.5.nine

Sterile consummate uniform including masks, surgical gloves etc.

5.5.eleven

Filtration flask (2 liter capacity).

5.five.12

Autoclave-able rubber tubing, silica gel cavalcade and 2 liter filtration flask to the sterility testing unit.

5.v.13

Petri dishes for air count and gloves per procedures.

5.v.fourteen Sterile glass or disposable syringes for liquid testing (optional).

5.five.15 Gas burner.

All materials for testing are sent inside the sterility testing suite through the hatch. Materials are wrapped in aluminum foil and sterilized either by dry heat (200˚C for NLT ii.0 hours) or past autoclaving. Remove the outer wrapping in the hatch and transfer the material inside under LFH. Materials that are not sensitive to UV calorie-free exposure are kept at that place for at least 30 minutes to minimize the outside microbial bioburden. Perform growth promotion test of all medias to exist used in the test as per SOP.

6. Definitions:-

Inside the strictest definition of sterility, an commodity is accounted to sterile when there is complete absenteeism of viable microorganisms. Absolute sterility cannot be practically demonstrated because it is technically unfeasible to prove a negative absolute.

half dozen.2 Sterility a test that critically assesses whether a
sterilized pharmaceutical product is free from contaminating microorganisms'.

6.three Membrane Filtration Method

This procedure is applicable to all the sterile materials, which when in liquid course can pass through the membrane filter of pore size of 0.45 uchiliad. For example Eye drops, Ampoules, Powder vials and sterile bulks.

half dozen.4 Fluid Thioglycollate Medium (FTM):-

FTM is primarily intended for the tillage of anaerobic bacteria.

6.5 Soybean Casein Digest Goop (Tryptic Soy Broth) (TSB):-

TSB is suitable for the culture of both fungi and aerobic bacteria.

6.half-dozen Peptone Water:-

Water-soluble digests or hydrolysates of proteins that are used in the grooming of civilisation media. Peptone h2o is used equally diluting and rinsing fluid in sterility testing.

6.seven Negative Command: - To verify testing conditions a negative control is performed using the chosen diluents in place of examination grooming. At that place must be no growth of microorganisms.

7.0 Period Nautical chart:-

8.0 Procedure:-

viii.one Adopt gowning for sterility test every bit per SOP.

8.2 Sample Requirements:

Collect the sample of product under test as per SOP.

8.3 Sample Preparation:

8.iii.1 Finished Stage Products

8.iii.1.1 Decontaminate vials / ampoules by submerging in sterile five% phenol solution or whatsoever other suitable approved disinfectant solution independent in sterile glass chalice for at to the lowest degree 30 minutes. Identify the container in the hatch. Transfer the sample container under LFH after wiping with sterile cotton duster containing approved disinfectant. Remove the vials from the container under LFH and allow drying under LFH.

8.3.1.2 Aseptically remove the aluminum foil seal with a sterile decapper and let information technology to dry.

vi.3.ane.3 Using a sterile spatula remove (flat side) the safe plug in one wiggle keeping the vials at an angle of 45^Oto the laminar flow bench.

vi.3.1.4Transfer aseptically pulverisation sample from each 20 filled vials into 200 ml of diluting fluid A (Peptone H2o) independent in a spiral capped flask, as follows:

Whole contents of products having filled weight of 250 mg.

300 mg of products having filled weight of 500 mg to 1.0 gm.

6.iii.1.5 Aseptically transfer half contents for each of 20 ampoules into 200 ml of diluting fluid A contained in a screw capped flask.

6.3.1.6 For 1ml volume ampoules whole contents will be used.

6.3.1.7 Replace the cap gently on the flask and spiral the cap tight. (Extensively flame with a Bunsen burner the lip of the flask prior to dumping of the sample & before replacing the cap].

8.3.2 Bulk Products:-

viii.3.2.1 Wipe the exterior of the sample bag with a duster soaked in a suitable disinfectant solution and identify in the hatch 20 minutes prior to testing. Before transferring the samples bag into testing room disinfectant again and place it under LFH and permit to dry.

8.three.two.2 Using a sterile spatula aseptically transfer approx. 6 gm of bulk into a flask containing diluting fluid.

viii.3.ii.3 Replace the cap carefully and spiral the cap tight. (Extensively flame with a Bunsen burner the lip of the flask prior to dumping of pulverisation and before replacing the cap].

8.3.2.4 Swirl gently without splashing the contents of the flask.

8.3.two.5 Set the grooming aside for a minimum of 1 hour to permit complete inactivation of penicillin and for the sample to dissolve completely.

viii.3.3 Sample Filtration:

8.iii.3.ane Set up the Millipore filtration organization nether LFH.

viii.iii.3.2 Remove the outer covering of aluminum foil in the hatch and then transfer the apparatus

under LFH with the inner roofing.

8.three.iii.iii Before the start of the test remove the inner covering of aluminum foil and ready the assembly to brand it ready for filtration process.

8.3.three.4 Centre a sterile membrane filter, aseptically, using a fine sterile flat tipped forceps on each of the filter holder to be used and clench the funnel tight.

8.3.3.five Connect the filtration associates to filtrate collection flask likewise as to vacuum pump. The collecting vessel along with the tubing is steam sterilized for 30 minutes, properly rapped in aluminum foil.

8.3.3.6 Unscrew the flask containing prepared sample. Flame the lip of the flask and pour the unabridged contents into the filtration funnel without dripping the sample down the side of the flask.

8.three.3.7 Filter the sample past using vacuum pump.

viii.three.three.8 Rinse the filter one time with 200 ml of sterile peptone water.

viii.3.three.9 Filter 200 ml of peptone water containing aforementioned quantity of pencillinase as used in case of examination sample and treat information technology as negative control.

eight.3.three.10 Remove the valve on each filter system after each filtration and at the terminate of the unabridged process.

8.3.3.11Open the caps of media flasks every bit FTM & Soyabean Casein Digest Medium / TSB and rut their lips properly.

8.iii.3.12 Remove the filter funnel and pick the filter using a sterile flat tipped forceps, from the base of the filter holder.

8 .three.three.13 Cut the filter in to ii equal halves with the assistance of sterile scissors and dip one piece in FTM and one piece in Soyabean Casein Digest Medium / TSB.

8.3.iii.fourteen Proceed the same way for all other samples.

8.iv Environmental Monitoring During Test

eight.4.ane Betrayal plate of Tryptic Soy Agar (TSA) under LFH for the whole duration of the test in infinite between the annotator and the examination appliance.

8.iv.2 Ane plate in sterility testing adjust.

8.4.three One plate in buffer area and one plate in modify room.

eight.iv.4 Gloves monitoring of the annotator is to be performed at the cease of the test.

8.5 Incubation

eight.five.1 Remove all the media out in the lab, Incubate FTM at thirty – 35 ºC & Soyabean Casein Digest Medium / TSB at 20 – 25 ºC for 14 days.

8.5.2 Incubate all tested plates at xxx – 35 ºC for a minimum of 48 hours.

8.half-dozen Positive Control

Positive command of media is performed only on receiving of new lot or changed lot of media in order to check the efficacy of testing media. Run a positive control by calculation one ml

of unlike bacterial / mold cultures, available in the lab, to a flask of diluting fluid A containing powder equivalent to 20 vials of 250 mg or book equal to twenty ampoules. Separate sample for all products is taken and proceeded in the same way. Go along the filtration in the same style as described above, except that glass filtration assembly is used instead of South/S Millipore filtration assembly in order to avoid any contamination in routine sterility test. Also filtration process is carried out in bacterial sub culturing lab under condom cabinet instead of sterility testing suite. Use the same media and incubation temperature and fourth dimension every bit used for test.

8.7 Estimation of Sterility Test Results

During incubation period observe the test media flasks daily for the bear witness of turbidity due to any microbial growth. Production is sterile if no prove of growth is plant throughout the incubation period. When microbial growth is observed and confirmed microscopically, the article does not come across the requirements of the sterility test. However if the microbial growth tin can exist without a incertitude ascribed to faulty aseptic techniques or materials used in conducting the sterility testing process, the test is invalid and must be repeated. Repeat the test on aforementioned number of units as already tested in previous test.

9. Records

Material/Product Sterility Test Written report

Sterility Test Data.

Surface area Monitoring of Sterility Testing Room

10. References:

ten.ane

U.s.a.

Pharmacopoeia 35 .

10.2 Guidelines on the Test Methods for Ecology Monitoring for Hygienic Dispensing Facilities, Produced by: A Working grouping of the Scottish Quality Assurance Specialist Interest Group, February 2004. (For All Classes at Rest and Operational B, C and D)

eleven. Distribution :-

This SOP has to exist distributed in below mentioned Departments:-

Sr. NO

Distributed to

Received

(Current)

Returned

(Obsolete)

one

Quality Control Department

Quality Management Section

12 Revision History:-

Date Changes