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Presented by:B.Kavita 3 rd year B.pharmacy

GUIDED BY:

Mrs. Ch.Supriya ;

NIRMALA COLLEGA OF PHARMACY

TEST FOR STERILITY FOR PARENTRALS

Asso. professor

Department of Biotechnology

Aseptic processing : in this components of the final dosage form are sterilize separately and the finished article assembled in an aseptic manner.

Prevents introduction of viable micro-organisms into components.

Requirements: 1- Air environment free from viable micro-organisms to permit

effective maintenance of air supply unit.

2- Adequately equipped trained operating personnel. The facility include primary and secondary barrier system.

Composition: w/v

Pancreatic Digest of Casein 15.0 g

Yeast Extract (water-soluble) 5.0 g

Glucose monohydrate/anhydrous 5.5 g/5.0 g

Sodium chloride 2.5 g

L- Cystine 0.5 g

Sodium thioglycollate 0.5 g

0.1% Resazurin Sodium Solution (freshly prepared)

1.0 mL

Granulated Agar (moisture not more than 15%) 0.75 g

Purified Water 1000 mL

Polysorbate 80 5.0 mL

pH after sterilization (measured at room temperature): 7.1± 0.2 particularly suitable for the cultivation of anaerobic organisms (incubation temperature 30–35°C)

Medium 1 (Fluid Thioglycollate Medium)

Mix L-cystine, NaCl, Dextrose, Yeast extract, Pencreatic

digest of casein with purified Water.

Dissolve Sodium thioglyocollate or thioglycollic acid in solution

if filtration is necessary, heat the solution without boiling and

filter hot through moistened filter paper

Add Resazurin sodium solution,mix,place in suitable vessel and subject for sterilization process.

(If necessary, add 1N NaOH to adjust pH to 7.1)

Fluid thioglycollate medium incubated at 32.5oC. Shows Pink colour change after incubation.

Medium 2 (Soybean-Casein Digest Medium)

Composition w/v

Pancreatic Digest of Casein 17.0 g

Pepsin Digest of Soybean Meal 3.0 g

Glucose monohydrate/anhydrous 2.5 g /2.3 g

Sodium chloride 5.0 g

Di potassium hydrogen phosphate 2.5 g

Purified Water 1000 mL

Polysorbate 80- 5.0 mL

pH after sterilization (measured at room temperature): 7.3±0.2

which will support the growth of both aerobic bacteria (incubation temperature 30–35°C) and fungi (incubation temperature 20–25°C)

Method of preparation:

Mix solids with purified water

Heat the solution

Cool to room temperature, and add sufficient quantity of 0.1 N NAOH to adjust the PH of the medium between 7.1 to 7.5 .transfer to suitable

container and kept for sterilization. Store at temperature 20 C – 25 oC. Incubated at 22.5oC.

Membrane filtration Appropriate for : (advantage)

Filterable aqueous preparations

Oily ,Alcoholic preparations

Preparations miscible with or soluble in aqueous or

oily (solvents with no antimicrobial effect)

All steps of this procedure are performed aseptically

in a Class 100 Laminar Flow Hood

Membrane filter 0.45μ porosity

Filter the test solution

After filtration remove the filter

Cut the filter in to two halves

First halves (For Bacteria) Second halves (For Fungi)

Transfer in 100 ml culture media(Fluid Thioglycollate medium)

Incubate at 30-350 C for not less then 7 days

Transfer in 100 ml culture media(Soyabeans-Casein Digest medium)

Incubate at 20-250 C for not less then 7 days

Observe the growth in the media Observe the growth in the media

Suitable for samples with small volumesvolume of the product is not more than 10% of the

volume of the medium suitable method for aqueous solutions, oily liquids,

ointments and creamsDirect inoculation of the culture medium suitable

quantity of the preparation to be examined is transferred directly into the appropriate culture medium & incubate for not less than 14 days.

Positive controls

small numbers of suitable microorganisms to grow in media in the presence of the sample is measured.

The microorganism used for positive control tests with a product containing an antimicrobial agent must.

indicates a satisfactory inactivation

To be sensitive to that agent

Culture media is examined during and after at the end of incubation.

The following observations are possible:

1) 1) No evidence of growth Pass the test for sterility.

2) 2)Re-testing is performed with twice no. of sample if:

3) 3) No evidence of growth Pass the test for sterility. There

is evidence of growth Re-testing is performed same no. of

sample, volume & media as in original test No evidence

of growth Pass the test for sterility.

4) 4)There is evidence of growth isolate & identify the organism.