Lesson 8. Sterility Testing

Sterile Products

dosage forms of therapeutic agents that are free of viable microorganisms

bypass the body’s usual defenses

Sterile products must be free from contaminants because they ________________ against infections

ampules, vial, ophthalmic preparations (eye drops) and irrigation solutions

examples of sterile products

Environment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Personnel Traffic Flow

Environment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Facility Design

Environment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Effects from adjacent areas

Environment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Utility Systems

Equipment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Equipment Design

Environment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Decontamination

Equipment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Area Equipment

Product

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Validation

Product

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Product and Material Flow

Environment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Personnel Hygiene

Equipment and Product

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Sterilization

Product

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Depyrogenation

Product

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Product and Materials

Environment and Equipment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Qualification

Equipment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Procedures

Equipment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Procedures

Environment and Equipment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Cleaning and Maintenance

All of the Above

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Personnel Practices and Training

Product

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• Storage Conditions

Environment

Factors Influencing the Sterility of Final Products:

Is it Product, Equipment, or Environment?

• HVAC

Sterility Assurance

• refers to the confirmation of safety for products labeled as sterile

• through the implementation of collective controls that give confidence to the suitability of items labeled as sterile

mitigating microbiological risks

Sterility is important in ______________________________ and provides assurance better than relying solely on the finished product testing. 

microbial decontamination of the equipment, environment, and the product

Sterility starts with the initial ___________________________

Aseptic Processing

METHODS OF PREPARING STERILE PRODUCTS:

• designed to prevent the introduction of viable microorganisms into separately sterilized materials during their assembly into a sealed sterile package

• sterilized materials should be protected from contamination starting from the point of sterilization up to the closure of the primary package

• before assembling the final product, the product itself, its container, and closure are all sterilized

products that are sensitive to high heat, pressure, or radiation

Aseptic Processing is used for:

Terminally-Sterilized Products

METHODS OF PREPARING STERILE PRODUCTS:

• Processing in high-quality environmental conditions followed by a sterilization process

• Sterilization process imparts quantifiable safety level achieved by delivering measurable physical conditions that corresponds to microbial lethality

lowest-risk category of sterile pharmaceutical products

Terminal sterilization is used for:

≤ 10-6 or n.m.t. one nonsterile unit in one million units

Probability of the terminal sterilization process generating a nonsterile unit of ____________________________

Sterility Test

Applied to substances, preparations, or articles which are required to be sterile

no contaminating microorganism

Sterility Test indicates that _____________________ has been found in the sample examined under the conditions of the test

aseptic conditions

Sterility Test is carried out under ___________________

Membrane filtration method

METHODS OF STERILITY TEST

• Filters with nominal pore size of n.m.t. 0.45 µm

Cellulose nitrate filters

filter that is used for aqueous, oily, and weakly alcoholic solutions

Cellulose acetate filters

filter that is used for strongly alcoholic solutions

Direct inoculation method

METHODS OF STERILITY TEST

• Addition of product directly into the culture medium

Fluid Thioglycollate Medium

METHOD SUITABILITY TEST (METHOD VALIDATION)

GROWTH PROMOTION TEST

1.a. Place 5 mL of ___________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately.

1.b. Place 5 mL of ____________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately.

2. Label the test tubes with _____, culture medium, microorganisms, incubation temperature, section & group no., and pullout date.

3.a. Incubate the Fluid Thioglycollate Medium at __________ for 3 days (bacteria) and 5 days (fungi)

3.b. Soybean Casein Digest Medium at _____ for 3 days (bacteria) and 5 days (fungi)

1.a. = ?

Soybean-Casein Digest Medium

METHOD SUITABILITY TEST (METHOD VALIDATION)

GROWTH PROMOTION TEST

1.a. Place 5 mL of ___________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately.

1.b. Place 5 mL of ____________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately.

2. Label the test tubes with _____, culture medium, microorganisms, incubation temperature, section & group no., and pullout date.

3.a. Incubate the Fluid Thioglycollate Medium at __________ for 3 days (bacteria) and 5 days (fungi)

3.b. Soybean Casein Digest Medium at _____ for 3 days (bacteria) and 5 days (fungi)

1.b. = ?

GPT

METHOD SUITABILITY TEST (METHOD VALIDATION)

GROWTH PROMOTION TEST

1.a. Place 5 mL of ___________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately.

1.b. Place 5 mL of ____________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately.

2. Label the test tubes with _____, culture medium, microorganisms, incubation temperature, section & group no., and pullout date.

3.a. Incubate the Fluid Thioglycollate Medium at __________ for 3 days (bacteria) and 5 days (fungi)

3.b. Soybean Casein Digest Medium at _____ for 3 days (bacteria) and 5 days (fungi)

2 = ?

30-35 C

METHOD SUITABILITY TEST (METHOD VALIDATION)

GROWTH PROMOTION TEST

1.a. Place 5 mL of ___________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately.

1.b. Place 5 mL of ____________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately.

2. Label the test tubes with _____, culture medium, microorganisms, incubation temperature, section & group no., and pullout date.

3.a. Incubate the Fluid Thioglycollate Medium at __________ for 3 days (bacteria) and 5 days (fungi)

3.b. Soybean Casein Digest Medium at _____ for 3 days (bacteria) and 5 days (fungi)

3.a. = ?

20-25 C

METHOD SUITABILITY TEST (METHOD VALIDATION)

GROWTH PROMOTION TEST

1.a. Place 5 mL of ___________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately.

1.b. Place 5 mL of ____________________ in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately.

2. Label the test tubes with _____, culture medium, microorganisms, incubation temperature, section & group no., and pullout date.

3.a. Incubate the Fluid Thioglycollate Medium at __________ for 3 days (bacteria) and 5 days (fungi)

3.b. Soybean Casein Digest Medium at _____ for 3 days (bacteria) and 5 days (fungi)

3.b. = ?

Clear visible growth of the microorganisms (turbidity)

GROWTH PROMOTION TEST: Acceptance Criteria

• ______________________________________is present in the culture media.

detection of the test microorganisms.

GROWTH PROMOTION TEST: Interpretation

• Growth Promotion Test shows that Fluid Thioglycollate Medium and Soybean Casein Digest Medium are suitable for the ___________________________.

Pseudomonas aeruginosa and Staphylococcus aureus

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — CONTROL

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

2. Label the test tubes with _______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. Incubate the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for __________.

1.a. =  ?

Bacillus subtilis and Candida albicans

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — CONTROL

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

2. Label the test tubes with _______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. Incubate the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for __________.

1.b. =  ?

Suitability Control

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — CONTROL

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

2. Label the test tubes with _______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. Incubate the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for __________.

2 =  ?

14 days

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — CONTROL

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes, then inoculate the medium with 0.1 mL (90 cfu) of ____________________________ microbial suspension separately.

2. Label the test tubes with _______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. Incubate the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for __________.

3 =  ?

Clear visible growth of the microorganisms (turbidity)

SUITABILITY OF TEST METHOD: Acceptance Criteria

• _____________________________________ is present in the culture media.

normal saline solution

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — TEST PRODUCT

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes and add 1 mL of _________________. Inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes and add 1 mL of ______________________. Inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately

2. Label the test tubes with ______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. _________ the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for 14 days

1.a. = ?

normal saline solution

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — TEST PRODUCT

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes and add 1 mL of _________________. Inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes and add 1 mL of ______________________. Inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately

2. Label the test tubes with ______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. _________ the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for 14 days

1.b. = ?

Suitability Test

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — TEST PRODUCT

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes and add 1 mL of _________________. Inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes and add 1 mL of ______________________. Inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately

2. Label the test tubes with ______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. _________ the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for 14 days

2 = ?

Incubate

METHOD SUITABILITY TEST (METHOD VALIDATION)

SUITABILITY OF TEST METHOD (BACTERIOSTATIS AND FUNGISTASIS) — TEST PRODUCT

1.a. Place 10 mL of Fluid Thioglycollate Medium in each of two test tubes and add 1 mL of _________________. Inoculate the medium with 0.1 mL (90 cfu) of Pseudomonas aeruginosa and Staphylococcus aureus microbial suspension separately

1.b. Place 10 mL of Soybean-Casein Digest Medium in each of two test tubes and add 1 mL of ______________________. Inoculate the medium with 0.1 mL (90 cfu) of Bacillus subtilis and Candida albicans microbial suspension separately

2. Label the test tubes with ______________, culture medium, microorganisms, incubation temperature, section and group no., and pull-out date.

3. _________ the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for 14 days

3 = ?

evaluation of the sterile product (normal saline solution)

SUITABILITY OF TEST METHOD: Interpretation

• Suitability Test demonstrates suitability of the method for the _____________________.

10 mL of Fluid Thioglycollate Medium in a test tube and add 1 mL of normal saline solution

PRODUCT TESTING:

1. Place _________________________________________________________. Do the same with Soybean-Casein Digest Medium.

2. Label the test tubes with ____________ and culture medium

3. Incubate the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for 14 days.

1 = ?

Test Product

PRODUCT TESTING:

1. Place _________________________________________________________. Do the same with Soybean-Casein Digest Medium.

2. Label the test tubes with ____________ and culture medium

3. Incubate the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for 14 days.

2 = ?

14 days

PRODUCT TESTING:

1. Place _________________________________________________________. Do the same with Soybean-Casein Digest Medium.

2. Label the test tubes with ____________ and culture medium

3. Incubate the Fluid Thioglycollate Medium at 30-35 C, and the Soybean Casein Digest Medium at 20-25 C for 14 days.

3 = ?

no evidence

PRODUCT TESTING: Acceptance Criteria

• There should be ________________ of microbial growth

sterile

PRODUCT TESTING: Interpretation

• Sterility test determines if the product tested is _________.