Preservatives are added to pharmaceutical products to prevent microbial growth and extend shelf life. An ideal preservative kills microbes rapidly at low concentrations, is non-toxic, stable, and does not interact negatively with the product ingredients. A preservative efficacy test evaluates the ability of a preservative system to inhibit microbial growth when challenged with common test microbes like S. aureus and P. aeruginosa over 28 days. The test involves inoculating product samples, incubating them, and conducting plate counts to determine microbial survival rates over time. This ensures the preservative maintains effective antimicrobial activity as required by pharmacopeia standards.
This presentation is brief introduction about preservatives employed in pharmaceutical dosage forms to prevent formulation from oxidation and microbial attack during storage and patient usage.it includes classification of preservatives, uses and method of analysis of preservatives and also introduction about pharmacopoeial evaluation of preservatives that is preservative activity test (PAT)
Types of spoilage, factors affecting the microbial spoilage of pharmaceutical...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-V Part-1
Types of spoilage, factors affecting the microbial spoilage of pharmaceutical products, source and type of contaminants. Introduction: Defintion Types of Microbial Spoilage:
1. Infection induced due to contaminated pharmaceutical products: Table no. 1.1 Common pathogens spoiling pharmaceutical products:
2. Physicochemical spoilage –
i) Viable growth ii) Gas production
iii) Colouration / Decolouration
iv) Odour formation
v) Taste change
3. Physical Spoilage:
Cracking of emulsion:
Odor changes
4. Biological spoilage:
Microbial Toxins
Microbial Metabolites
5. Chemical spoilage: Table 1.2 Susceptibility of pharmaceutical ingredients to microbial contamination
Factors affecting microbial spoilage
Size of contaminant inoculum
Nutritional factors
Moisture content
pH
Storage temperature
Redox potential
Packaging design
Sources and Types Of Contamination:
Personnel,
Poor facility design,
Incoming ventilation air,
Machinery and other equipment for production,
Raw material and semi-finished material,
Packaging material,
Utilities,
Different media used in the production process as well as for cleaning and Cleanroom clothing.
Evaluation of Bactericidal and Bacteriostatic (Disinfectant). PHARMACEUTICAL ...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-III Part-5 Evaluation of Bactericidal and Bacteriostatic (Disinfectant). The common methods used for evaluation of a disinfectant are as follows,
Tube Dilution Method.
Agar Plate Method.
Filter Paper & Cup Plate Method.
Ditch-Plate Method.
Phenol Coefficient Method.
The official phenol coefficient tests include,
Rideal-Walker Test (RW Test).
Chick-Martin Test.
United States FDA Test for Phenol Coefficient. (FDA Test)
The US Association of Official Agricultural Chemists Test (FDA Test)
A. Rideal-Walker Test:
Kelsey Sykes Method
This presentation is brief introduction about preservatives employed in pharmaceutical dosage forms to prevent formulation from oxidation and microbial attack during storage and patient usage.it includes classification of preservatives, uses and method of analysis of preservatives and also introduction about pharmacopoeial evaluation of preservatives that is preservative activity test (PAT)
Types of spoilage, factors affecting the microbial spoilage of pharmaceutical...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-V Part-1
Types of spoilage, factors affecting the microbial spoilage of pharmaceutical products, source and type of contaminants. Introduction: Defintion Types of Microbial Spoilage:
1. Infection induced due to contaminated pharmaceutical products: Table no. 1.1 Common pathogens spoiling pharmaceutical products:
2. Physicochemical spoilage –
i) Viable growth ii) Gas production
iii) Colouration / Decolouration
iv) Odour formation
v) Taste change
3. Physical Spoilage:
Cracking of emulsion:
Odor changes
4. Biological spoilage:
Microbial Toxins
Microbial Metabolites
5. Chemical spoilage: Table 1.2 Susceptibility of pharmaceutical ingredients to microbial contamination
Factors affecting microbial spoilage
Size of contaminant inoculum
Nutritional factors
Moisture content
pH
Storage temperature
Redox potential
Packaging design
Sources and Types Of Contamination:
Personnel,
Poor facility design,
Incoming ventilation air,
Machinery and other equipment for production,
Raw material and semi-finished material,
Packaging material,
Utilities,
Different media used in the production process as well as for cleaning and Cleanroom clothing.
Evaluation of Bactericidal and Bacteriostatic (Disinfectant). PHARMACEUTICAL ...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-III Part-5 Evaluation of Bactericidal and Bacteriostatic (Disinfectant). The common methods used for evaluation of a disinfectant are as follows,
Tube Dilution Method.
Agar Plate Method.
Filter Paper & Cup Plate Method.
Ditch-Plate Method.
Phenol Coefficient Method.
The official phenol coefficient tests include,
Rideal-Walker Test (RW Test).
Chick-Martin Test.
United States FDA Test for Phenol Coefficient. (FDA Test)
The US Association of Official Agricultural Chemists Test (FDA Test)
A. Rideal-Walker Test:
Kelsey Sykes Method
Microbial spoilage-by S.D.Mankar types, sources of contamination, factors,Ass...someshwar mankar
Types of spoilage, factors affecting the microbial spoilage of pharmaceutical products,
sources and types of microbial contaminants, assessment of microbial contamination and
spoilage.
Microbiological assay-Principles and methods of different microbiological assay.someshwar mankar
Principles and methods of different microbiological assay. Methods for standardization of
antibiotics, vitamins and amino acids. Assessment of a new antibiotic.
Preservation of pharmaceutical products using antimicrobial agents. PHARMACEU...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-VPart-3
Preservation of pharmaceutical products using antimicrobial agents.
Introduction. Ideal Properties of Preservatives:
Antimicrobial Chemical Preservatives
Development of a Preservative System.
Factors affecting efficacy of a preservative: 1. Interaction With components of the formulation
2. Properties of the Preservatives:
3) Effect of Containers.
4) Type of microbes:
5) Influence of pH:
Challenge Test: Efficacy Test of Preservative : Medium used, Choice of test organism:
Preparation of the inoculum:
Procedure:
Interpretation of Results:
Assessment of microbial contamination and spoilage. PHARMACEUTICAL MICROBIOLO...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-VPart-2
Assessment of microbial contamination and spoilage.
Assessment of microbial contamination and spoilage
1. Physical and chemical changes:
2. Assessment of viable microorganisms in non-sterile products:
3. Sterility test:
4. Estimation of pyrogens:
Microbial Limit Tests:
Total Aerobic Microbial Count:
Membrane Filtration.
Plate Count Methods.
Pour Plate Method.
Surface spread Method.
Most Probable Number(MPN)
Sterility testing products (solids, liquids, ophthalmic and other sterile pro...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-6 Sterility testing products (solids, liquids, ophthalmic and other sterile products) according to IP, BP, USP.
Introduction: Test for Sterility. Culture Media. Fluid Thioglycollate Medium (FTM).
Alternative Thioglycollate Medium (ATM).
Soybean Casein Digest Medium (SCDM).
Tests for Culture Media:
Sterility of Media.
Growth Promotion Test.
Test for Bacteriostatic and Fungistatic.
Sterility Test Methods. Methods A: Membrane Filtration.
Method B: Direct Inoculation Pyrogen Test Methods. Rabbit Test. LAL Test.
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T) Unit-IV Part-3
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic: Introduction:
Principle
Microbiological Assay of Cynocobalamin (Vitamin B12):
Tritrimetric Method.
Turbidimetric Method.
Preparation of Standard Cynocobalmine stock solution:
Preparation of Basal Medium Stock Solution:
Test Solution of the material to be assayed Preparation of inoculum: Procedure of Titrimetric method: Turbidimetric Method: Microbiological assay of Amino acids. Assessment of a New Antibiotic.
Introduction:
MIC of an antibiotic is tested either by one of the following ways,
Liquid Dilution Method.
Solid Dilution Method
Evaluation of Bactericidal and BacteriostaticRajsingh467604
What are disinfectants?
As per the definition given by WHO ( World health organization ) : a disinfectant is a chemical agent, which destroys or inhibits growth of pathogenic microorganisms in the non-sporing or vegetative state.
Why Evaluation?
Evaluation of disinfectants is used to check the ability or efficacy of any disinfectant against specific microorganisms to establish its effectiveness.
Evaluation tests of bactericide.
1. RIDEAL WALKER TEST
This test is also known as the phenol coefficient test,in which any chemical is compared with phenol for its antimicrobial activity.
The result is shown in the form of phenol coefficient.
▪ If a phenol coefficient of a given test disinfectant is less than 1, it means that disinfectant is less effective than phenol.
▪ If a phenol coefficient of a given test disinfectant is more than 1, it means that disinfectant is more effective than phenol.
Procedure
1.1 Different dilutions of the test disinfectant and phenol are prepared and 5 ml of each dilution is inoculated with 0.5ml of the 24 hour growth culture of the organisms.
1.2 All tubes(Disinfectant + organisms & phenol + organisms) are placed in a water bath ( at 17.5° C)
1.3 Subcultures of each reaction mixture are taken and transferred to 5ml sterile broth at an interval of 2.5 minutes from zero to 10 mintues.
1.4 Broth tubes are incubated at 37° C for 2 to 3 days & examined for the presence or absence of the growth.
1.5 Then the Rideal Walker coefficient is calculated :
2. CHICK MARTIN TEST.
CHICK MARTIN test is performed in the much similar way as the RIDEAL Walker test but with a little variation.
Principle : This test is carried out in the presence of organic matter like 3% human feces or dried yeast.
Procedure
2.1 Serial dilutions of test solution and phenol is prepared in distilled water.
2.2 To this 3% yeast suspension is also added.
2.3 To this solution the S. typhi is added
2.4 After contact time of 30 mins the above mixture is transferred to the freshly prepared 10 ml of broth.
2.5 The test tubes are incubated at 37°C for 48 hours.
2.6 Presence or absence of the growth is calculated.
Evaluation tests of Bacteriostatic.
1. Tube dilution & Agar plate Method
1.1 The chemical agent is incorporated into nutrient broth or agar medium and inoculated with test micro-organisms.
1.2 These tubes are incubated at 30° TO 35°C for 2 to 3 days and then the results in the form of turbidity or colonies are observed.
1.3 The results are recorded and the activity of the given disinfectant is compared.
2. Cup plate method
2.1 Agar is melted and cooled at 45° Celsius.
2.2 Then inoculated with test micro-organisms and poured into a sterile petri plate.
2.3 In the cup plate method, when the inoculated agar has solidified, holes around 8mm in diameter are cut in the medium with a steel cork borer.
2.4 Now the antimicrobial agents are directly placed in the holes.
Evaluation of the efficiency of sterilization methods.Sterility indicatorsMs. Pooja Bhandare
Evaluation of the efficiency of sterilization methods.Sterility indicators
Sterility criteria: Bioburden ,Sensitivity of microorganisms
Death rate or Survivor curve,D- Value or Decimal reduction time,Z- value or Thermal reduction time, f- value, Q10 Value or Temperature Coefficient, Inactivation Factor:
STERILITY INDICATORS : Physical Indicators, Chemical Indicators
Biological Indicators
1. Physical Indicators: i) Moist heat Indicator ii) Dry heat iii) Radio sterilization iv) Gaseous methods v) Filtration 2.CHEMICAL INDICATORS : I) Browne’s tubes II) WITTNESS TUBES IV) Royce Sachet V) Chemical Dosimeter 3.BIOLOGICAL INDICATORS
Microbial spoilage-by S.D.Mankar types, sources of contamination, factors,Ass...someshwar mankar
Types of spoilage, factors affecting the microbial spoilage of pharmaceutical products,
sources and types of microbial contaminants, assessment of microbial contamination and
spoilage.
Microbiological assay-Principles and methods of different microbiological assay.someshwar mankar
Principles and methods of different microbiological assay. Methods for standardization of
antibiotics, vitamins and amino acids. Assessment of a new antibiotic.
Preservation of pharmaceutical products using antimicrobial agents. PHARMACEU...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-VPart-3
Preservation of pharmaceutical products using antimicrobial agents.
Introduction. Ideal Properties of Preservatives:
Antimicrobial Chemical Preservatives
Development of a Preservative System.
Factors affecting efficacy of a preservative: 1. Interaction With components of the formulation
2. Properties of the Preservatives:
3) Effect of Containers.
4) Type of microbes:
5) Influence of pH:
Challenge Test: Efficacy Test of Preservative : Medium used, Choice of test organism:
Preparation of the inoculum:
Procedure:
Interpretation of Results:
Assessment of microbial contamination and spoilage. PHARMACEUTICAL MICROBIOLO...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-VPart-2
Assessment of microbial contamination and spoilage.
Assessment of microbial contamination and spoilage
1. Physical and chemical changes:
2. Assessment of viable microorganisms in non-sterile products:
3. Sterility test:
4. Estimation of pyrogens:
Microbial Limit Tests:
Total Aerobic Microbial Count:
Membrane Filtration.
Plate Count Methods.
Pour Plate Method.
Surface spread Method.
Most Probable Number(MPN)
Sterility testing products (solids, liquids, ophthalmic and other sterile pro...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-6 Sterility testing products (solids, liquids, ophthalmic and other sterile products) according to IP, BP, USP.
Introduction: Test for Sterility. Culture Media. Fluid Thioglycollate Medium (FTM).
Alternative Thioglycollate Medium (ATM).
Soybean Casein Digest Medium (SCDM).
Tests for Culture Media:
Sterility of Media.
Growth Promotion Test.
Test for Bacteriostatic and Fungistatic.
Sterility Test Methods. Methods A: Membrane Filtration.
Method B: Direct Inoculation Pyrogen Test Methods. Rabbit Test. LAL Test.
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T) Unit-IV Part-3
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic: Introduction:
Principle
Microbiological Assay of Cynocobalamin (Vitamin B12):
Tritrimetric Method.
Turbidimetric Method.
Preparation of Standard Cynocobalmine stock solution:
Preparation of Basal Medium Stock Solution:
Test Solution of the material to be assayed Preparation of inoculum: Procedure of Titrimetric method: Turbidimetric Method: Microbiological assay of Amino acids. Assessment of a New Antibiotic.
Introduction:
MIC of an antibiotic is tested either by one of the following ways,
Liquid Dilution Method.
Solid Dilution Method
Evaluation of Bactericidal and BacteriostaticRajsingh467604
What are disinfectants?
As per the definition given by WHO ( World health organization ) : a disinfectant is a chemical agent, which destroys or inhibits growth of pathogenic microorganisms in the non-sporing or vegetative state.
Why Evaluation?
Evaluation of disinfectants is used to check the ability or efficacy of any disinfectant against specific microorganisms to establish its effectiveness.
Evaluation tests of bactericide.
1. RIDEAL WALKER TEST
This test is also known as the phenol coefficient test,in which any chemical is compared with phenol for its antimicrobial activity.
The result is shown in the form of phenol coefficient.
▪ If a phenol coefficient of a given test disinfectant is less than 1, it means that disinfectant is less effective than phenol.
▪ If a phenol coefficient of a given test disinfectant is more than 1, it means that disinfectant is more effective than phenol.
Procedure
1.1 Different dilutions of the test disinfectant and phenol are prepared and 5 ml of each dilution is inoculated with 0.5ml of the 24 hour growth culture of the organisms.
1.2 All tubes(Disinfectant + organisms & phenol + organisms) are placed in a water bath ( at 17.5° C)
1.3 Subcultures of each reaction mixture are taken and transferred to 5ml sterile broth at an interval of 2.5 minutes from zero to 10 mintues.
1.4 Broth tubes are incubated at 37° C for 2 to 3 days & examined for the presence or absence of the growth.
1.5 Then the Rideal Walker coefficient is calculated :
2. CHICK MARTIN TEST.
CHICK MARTIN test is performed in the much similar way as the RIDEAL Walker test but with a little variation.
Principle : This test is carried out in the presence of organic matter like 3% human feces or dried yeast.
Procedure
2.1 Serial dilutions of test solution and phenol is prepared in distilled water.
2.2 To this 3% yeast suspension is also added.
2.3 To this solution the S. typhi is added
2.4 After contact time of 30 mins the above mixture is transferred to the freshly prepared 10 ml of broth.
2.5 The test tubes are incubated at 37°C for 48 hours.
2.6 Presence or absence of the growth is calculated.
Evaluation tests of Bacteriostatic.
1. Tube dilution & Agar plate Method
1.1 The chemical agent is incorporated into nutrient broth or agar medium and inoculated with test micro-organisms.
1.2 These tubes are incubated at 30° TO 35°C for 2 to 3 days and then the results in the form of turbidity or colonies are observed.
1.3 The results are recorded and the activity of the given disinfectant is compared.
2. Cup plate method
2.1 Agar is melted and cooled at 45° Celsius.
2.2 Then inoculated with test micro-organisms and poured into a sterile petri plate.
2.3 In the cup plate method, when the inoculated agar has solidified, holes around 8mm in diameter are cut in the medium with a steel cork borer.
2.4 Now the antimicrobial agents are directly placed in the holes.
Evaluation of the efficiency of sterilization methods.Sterility indicatorsMs. Pooja Bhandare
Evaluation of the efficiency of sterilization methods.Sterility indicators
Sterility criteria: Bioburden ,Sensitivity of microorganisms
Death rate or Survivor curve,D- Value or Decimal reduction time,Z- value or Thermal reduction time, f- value, Q10 Value or Temperature Coefficient, Inactivation Factor:
STERILITY INDICATORS : Physical Indicators, Chemical Indicators
Biological Indicators
1. Physical Indicators: i) Moist heat Indicator ii) Dry heat iii) Radio sterilization iv) Gaseous methods v) Filtration 2.CHEMICAL INDICATORS : I) Browne’s tubes II) WITTNESS TUBES IV) Royce Sachet V) Chemical Dosimeter 3.BIOLOGICAL INDICATORS
Preservatives are substances (natural or chemical) that are added to pharmaceutical products to prevent any kind of physical, chemical or biological changes. Antioxidants: They are self reducing agents that oxidize themselves and prevent oxidation of the components that are sensitive to oxygen.
Antimicrobial agent is a substance that interferes with the growth and activity of microorganisms.
These agents inhibit or kill microorganisms. Some antimicrobial agents are used to fight against infections and are called Chemotherapeutic Agents
A unique characteristic of an antimicrobial agent is selective toxicity, that is, it will destroy the organism but not affect the host
Factors affecting action of Disinfectants and Factors Affecting Choice Of Ant...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-III. Factors affecting action of Disinfectants and Factors Affecting Choice Of Antimicrobial Agent: Concentration of the disinfectant.
Chemical Structure of the disinfectant.
Formulation of the disinfectant.
Interfering substances in the environment.
pH of the surrounding.
Potentiation and antagonism of the disinfectants.
Surface Tension.
Temperature.
Time of Contact.
Type and no. of microbes present.
FACTORS AFFECTING CHOICE OF ANTIMICROBIAL AGENT:
Properties of chemical agents
Environment
Types of microorganisms
Intended application
Toxicity agents
Culture state
Stability testing of natural products.docxKipaPape
Stability is defined as the capacity of drug to remain within established specification limits to maintain its identity, strength, quality and purity throughout the retest or expiration dating period.
It is the ability of formulations to retain its physical, chemical, microbiological and toxicological parameters same that time of manufacturer.
Precautions in handling pharmaceutical products containing antibioticsUniversity of Sindh
It includes precautions in handling and manufacturing pharmaceuticals containing antibiotics. all these slides have been prepared with the help of different research papers, books, and FDA guidelines.
That presentation is about the stability of the drug, why it's necessary?? What is the shelf life of a drug? Purpose of stability testing and its importance.Also a review article of Sanjay Bajaj et al published in Journal of applied pharmaceutical sciences.
research papers Life Sciences Biotechnology and Pharma Sciencesnagarajukarnatik4hit
our International Journal of Life Sciences Biotechnology and Pharma Sciences (IJLBPS) stands as a beacon of scientific excellence in the realm of life sciences, biotechnology, and pharmaceutical sciences. Published quarterly, this esteemed journal serves as a platform for disseminating high-quality original research, reviews, and short communications that push the boundaries of knowledge and innovation in these dynamic fields.
https://ijlbps.org/
https://ijlbps.org/contactus.php#1
Hurdle technology in Fish PreservationShubham Soni
Hurdle Technology is a kind of combination of Mechanisms to preserve the perishable commodity like Fish and the Fish Products, its even useful in other Industries like Poultry, Agri-Industries etc.
Just Keep Creating Hurdles for Microbes and we all we have a healthy and Hygienic Life...!
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2. INTRODUCTION
• Preservatives are the chemical substances used to improve or
amplify shelf life of drugs by decreasing or lowering the
oxidation of active ingredients or excipients and by reducing
microbial production
• Prevent the growth of unwanted microorganisms in
pharmaceutical products
Ideal properties of preservatives:
Should able to kill all microbes contaminants rapidly
Tasteless and odourless
Not be irritant
Non toxic to the patient
Physically and chemically Stable
Effective at low concentration throughout the life of the medicine
Should be selective in reacting with the contaminants and not the
ingredients of the medicine.
Cost effective
3. A single preservative is not suitable for preservation of all
pharmaceutical formulations.
• Some preservatives are ineffective with some microbe strains and
should be combined with others to be effective.
• Combination of two or more preservatives are used to extend the
range and spectrum of preservation.(Synergic action)
Germall 115+parabens (antibacterial +antifungal)-Tablets,
Phenylethyl alcohol+phenoxetol+ benzalkonium chloride
(wide antimicrobial) -Eye drops and contact lens
Methyl parahydroxybenzoic acid and propyl
parahydroxybenzoic acid (antimicrobial) –Injection
• It must decrease the percentage of the microbes and prevent any
re-growth, They can be:
• –Microbiostatic- that inhibits the growth or multiplication of
microbiota(pathogenic microbes)
• –Microbiocidal in nature- that kills the microbiota
4. NEED FOR PRESERVATIVES
To protect our drug from microbial
attack
To enhance activity and efficacy of
drug
To increase shelf life of our product
To stabilize our product
5. Classification of preservation
Based on mechanism of action:
1. Antioxidants: The agent which prevent oxidantion of API which
otherwise undergo degradation due to oxidation as they are
sensitive to oxygen.
Ex: Vitamin E & C, Butylated hydroxy anisole (BHA), Butylated
hydroxy toluence(BHT).
2. Antimicrobial agents: the agent which active against Gram +ve
and Gram –ve microbes which causes degradatation of
pharmaceutical preparation which are active in low
concentration.
Ex: benzoates, sodium benzoate, sorbates.
3. Chelating agents: the agents which form the complex with
pharmaceutical ingredient and prevent the degradation of
pharmaceutical formulation.
Ex: EDTA, Polyphosphates, citric acid
6. Classification of preservation
Based on sources:
1. Natural preservatives: These preservatives
are obtained from natural sources like plants,
minerals and animal sources, etc
Ex: Neem oil, salt(sodium chloride), lemon,
honey
2. Artificial preservatives: These are man made
by chemical synthesis and active against
various microbes in small concentration.
Ex: Benzoates, sodium benzoate, sorbates,
propionets, nitrites.
10. 1. Interaction with formulation
• Hydrocollids such as methylcellulose, alignates, tragacanth can
interacts with preservatives and diminish their activity.
• Many emulgents are used in pharmaceutical preparations to
produce elegant applications. Interaction may occur between
preservatives and emulsified oil phase and with emulgent
molecules.
• Nature of oil, oil water ratio, type of concentration of emulgent,
influence the concentration of preservatives needed to protect the
system.
• Many tablet additives cause problems in tablet preservations
due to their interaction with added preservatives.
11. 2. Properties of the preservation
• The distribution of preservative must be
homogeneous and more solubility in the
bulk phase is preferable in multi phase
system.
• Some chemicals such as chlorobutol may
hydrolyse on storage if the pH is
unfavourable.
• Preservatives may react with substances
leached from the container and lose its
antimicrobial activity.
12. 3. Effects of containers
• Formulations packed in glass containers can be
expected to retain their preservative content if
closure is airtight.
• Preservatives may penetrate through the plastic
container and interacts with it.
• Rubber also reacts with many preservatives
but is still used for closures.
• Containers or closures may cause contamination
of pathogens.
• Screw–capped containers and corks are the
common source of mould spores.
13. 4. Type of microorganisms
• Plants products may contain pathogenic
microorganisms from the soil. E.g
Clostridium species, Bacillus anthracis.
• These soil microorganisms can cause spoilage
of pharmaceutical products.
• Many products prepared from animal
sources may contain pathogens like
Salmonella typhi.
• Spores of tetanus and gas gangrene have been
isolated from geletin.
14. 5. Influence of pH
• Adjustment of the pH of solution may affect
the chemical stability and the activity of the
preservative.
• The majority of preservatives are less
dependent upon pH, although cationic active
quaternary ammonium compounds are more
active at high pH values.
17. SIDE EFFECTS
• While choosing preservative for drug product
consideration should be made about:
• 1. Concentration
• 2. Toxicity
• 3. Selectivity
• 4. Interaction with formulation, etc.
18. Some preservatives and their Side effects
Sl. no Preservative Side effects
1. Paraben Neurological damage in rats
Potent irritants
2. Formaldehyde
Diazolidinyl urea
Imidazolidinyl urea
Skin irritants
Eye irritants
lung irritants
3. Benzyl alcohol fatal toxic syndrome in low
weight neonates
4. Cetyl alcohol
Stearyl alcohol
Infrequent sensitizers
5. 2-Phenylethanol Irritant to skin, eye and mucous
membranes
6. Benzoic acid Gastro-irritant
7. Chloroxylenol Cross sensitivity
8. Chlorocresol Irritant to skin, eyes and
mucous membranes
9. Hexachlorophene
EDTA
Neurotoxicity
Dose-related bronchoconstriction
19. EVALUATION OF PRESERVATIVES
• The evaluation of preservatives has traditionally involved time-consuming tests:
1. Preservative Efficacy Tests(PET) also called Antimicrobial Effectiveness
Testing(AET)
A number of factors determine the efficacy of a preservative, including the
active component of the product, the formulation in which it is incorporated or
the container/packaging in which the product is enclosed.
Why needed?
Antimicrobial preservatives are added to pharmaceuticals, medical devices,
personal care products, cosmetics and food products to inhibit the growth
of micro-organisms inadvertently introduced during the manufacturing
process or during the use of products with multiple-dose containers.
Guaranteeing that no inappropriate organisms are able to sustain
themselves within the product or its packaging.
Is undertaken to ensure that the preservative maintains its ability to inhibit
micro-organisms and is required by British Pharmacopoeia (BP),
European Pharmacopoeia (Ph. Eur.) and United States Pharmacopoeia
(USP) standards.
20. Preservative Efficacy Test (PET)
• First appeared in USP in 18th revision, 1st sept. 1970.
• Applied to the formulated medicine in its final container to
determine whether it is adequately protected from microbial
spoilage.
• Tests/standards apply only to the product in original, unopened
containers
• This test is done to determine the efficacy of preservative against
microorganism.
• Involve challenging a product with a defined number of colony forming
units (CFU) of a variety of test microorganisms (bacteria, yeasts and
fungi)
• Then monitoring the kill /survival rate at defined time intervals up to 28-
days.
• It is examined by the duplicate plate count method (CFU/ml = number
of colonies per ml plated /Total dilution factor)
• Multiple dosages forms- Parenterals, otics, nasal, oral, topical and
ophthalmic products made with aqueous bases or vehicles.
21. STEPS INVOLVED
• Preservative efficacy test is done in following
steps-
1. Product category
2. Test organism
3. Media culture preparation
4. Preparation of inoculum
5. Procedure
6. Interpretation
23. 2. Test organisms
• Choice of test microorganisms: Preservatives
should be active against as wide as a range of
microorganisms as possible hence the choice should
be of both Gram+ve and Gram-ve bacteria, yeast
and moulds in IP Test.
• Test organism that are recommended
by all of the pharmacopoeias includes:
Staphylococcus aureus ATCC
6538
Pseudomonas aeruginosa ATCC
9027
E. coli ATCC 8739
Fungi/mould, Aspergillus
Brasiliensis ATCC 16404
Yeast, Candida albicans ATCC
10231
24. 3. Media culture
• Soyabean casein digest agar medium (SCDM) & Sabouraud
-Dextrose medium.
• http://www.himedialabs.com/TD/MM011.pdf
25. 4. Preparation of inoculum
• Preparatory to the test, inoculate the medium from a recently
stock culture of each of the specified microorganisms(1ml).
• Escherichia coli ATCC 4352, Pseudomonas aeruginosa ATCC
9027 and Staphylococcus aureus ATCC 6538 are inoculated in
Sabouraud-Dextrose medium and incubated at 32.5 ± 2.5°C
for 3-5 days.
• Candida albicans ATCC 10231, Aspergillus brasiliensis ATCC
16404 are inoculated in Soyabean-Casein digest medium and
incubated at 22.5 ± 2.5°C for 3-5 days for Candida albicans &
3-7 days for Aspergillus brasiliensis .
• After this incubation period, microbial culture is harvested by
washing with sterile saline(0.9% w/v) to obtain a microbial
count of about 10 8 CFU/ mL (collect it in a sterile container
and treat this as Stock solution of test solution)
26.
27. 5. Procedure
Maintaine aseptic condition throughout the process and
use hand glove rinsed with 70% Isopropyl alcohol
Use five Product containers if the volume per container
is sufficient.
Reconstitute(restore) the product container if it is in dry
powder form, as per the instruction on the label.
Standardize the volume of the inoculum to be between
0.5% and 1.0% of the volume of the product and the
concentration to be between 1× 105 and 1×106 CFU/mL
of the product.
Add the inoculum into the product containers using
Sterile Syringe or Pipette.
Determine the initial count of the inoculated containers
by Plate count Method.
Incubate the inoculated containers at 22.5 ± 2.5°C
28. .The container sampling intervals include:
1. Category I products are sampled at 7, 14 & 28 days,
2. Category II-IV products are sampled at 14 & 28 days.
Withdraw samples from containers at the intervals of 7,
14 and 28 Days depending on product categories and
determine the number of CFUs of viable microbes per
mL present at each of these sample containers by the
Plate Count Method utilizing media with suitable in-
activator (neutralizer) in the diluents such as sterile
normal saline (0.9 % w/v) with polysorbate-80 (0.05 % w/v)
and 0.05 % Soya lecithin or with peptone (0.1 % w/v) and
0.05 % Soya lecithin or sterile normal saline (0.9 % w/v)
with 0.05 % polysorbate-80.(sometimes APIs or other
excipients may show preservative effects to neutralised
theses effects in-activator are added)
Express this in terms of Log Reduction and report as
Complies if results are complying to the criteria as given in
Table.
29. 6. Criteria for antimicrobial effectiveness/INTERPRETATION
For Category 1
BACTERIA 3.0 log reduction in 14 days, no increase up to
28days
Yeast No increase from initial count at 14 and 28days
For Category 2
BACTERIA 2.0 log reduction in 14days, no increase up to
28days
YEAST No increase from initial count at 14 and up to
28 days
For Category 3
BACTERIA
1.0 log reduction in 14days, no increase up to
28days
YEAST/MOULDS No increase from initial count at 14 and 28
days
For Category 4
BACTERIA No increase from initial count at 14 and 28
days
YEAST No increase from initial count at 14 and 28days
The criteria for microbial effectiveness are met if the specified criteria are met.
30. OR Interpretation(ANYONE)
1 Bacteria: Not less than 1.0 log
reduction from the initial count at
14 days, and no increase from the
14 days’ count at 28 days.
2 Yeast and Moulds: No increase
from the initial calculated count at
14 and 28 days.
31. Log reduction
• The log reduction achieved by a decontamination process is a
measure of how thoroughly the process reduces
the concentration of a contaminant.
• It is defined as the common logarithm of the ratio of the levels
of contamination before and after the process, so an increment
of 1 corresponds to a reduction in concentration by a factor of
10.
33. OTHER TECHNIQUES
• High sensitive analytical techniques are being
investigate as possible replacement for the
difficulty and time consuming pharmacopoeial
tests.
• These include methods such as
1. ATP bioluminescence
2. Electrical impedance spectrocopy
3. Spectro-fluorimetry
4. Chemiluminescence