Sterility testing products (solids, liquids, ophthalmic and other sterile products) according to IP, BP, USP. PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-6
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
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:
Principles and methods of different microbiological assay, methods for standa...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IV Part-2 Principles and methods of different microbiological assay, methods for standardization of antibiotics.
Introduction: Principles Advantages of Microbial Assay: Disadvantages of Microbial Assay: MICROBIOLOGICAL ASSAY OF ANIBIOTICS PRINCIPLE Media used for antibiotics assay Standard Preparation. Buffer Solutions Preparation of the Sample Solution: Test Organisms Preparation of inoculum: Methods of preparation of test organism suspension: Assay Methods: Method A: Cup-plate or Cylinder Plate Method.
Method B: Turbidimetric or Tube assay Method
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
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:
Principles and methods of different microbiological assay, methods for standa...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IV Part-2 Principles and methods of different microbiological assay, methods for standardization of antibiotics.
Introduction: Principles Advantages of Microbial Assay: Disadvantages of Microbial Assay: MICROBIOLOGICAL ASSAY OF ANIBIOTICS PRINCIPLE Media used for antibiotics assay Standard Preparation. Buffer Solutions Preparation of the Sample Solution: Test Organisms Preparation of inoculum: Methods of preparation of test organism suspension: Assay Methods: Method A: Cup-plate or Cylinder Plate Method.
Method B: Turbidimetric or Tube assay 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.
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)
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
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
Designing of aseptic area including design, construction, service, flow chart,source of contamination, method of prevention of it,clean area classification as per USPDA.
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.
Designing of aseptic area, laminar flow equipment: Study of different source ...Ms. Pooja Bhandare
Designing of aseptic area, laminar flow equipment: Study of different source of contamination in aseptic area and methods of prevention, clean area classification. PHARMACEUTICALMICROBIOLOGY (BP303T)Unit-IVPart-1
Introduction: Designing of Aseptic Area . i) The clean-up area,
ii) The compounding area,
iii) The aseptic area,
iv) The quarantine area and
v) The packaging/labelling area.
Flow diagram of aseptic area. Floors, walls and ceilings, Doors, windows and services Personnel and protective clothing Cleaning and disinfection. Air Supply. Laminar flow equipment. Vertical laminar air flow bench
Horizontal laminar air flow bench
High Efficiency Particulate Air (HEPA) Filter. Operating Instructions Uses of Laminar Air Flow.Advantages of Laminar Air Flow.Limitations of Laminar Air Flow. Air flow pattern Unidirectional airflow
Non-unidirectional airflow
Combined airflow
Different Sources of Contamination in an Aseptic Area
1) Personnel:
2) Buildings and Facilities
3) Equipment and Utensils:
4) Raw Materials
5) Manufacturing Process:
Methods of Prevention of Contamination Clean Area Classification
Disinfectant - Pharmaceutical microbiology (Second year b.pharm) (3rd semester)Kiran Shinde
Prof.Mr.Kiran K. Shinde (M.Pharm), Assistant professor (VNIPRC)
Pharmaceutical microbiology (Second year b.pharm) (3rd semester)
Introduction
Classification & mode of action of disinfectant
Factors affecting disinfectant, antiseptics & their evaluation
Evaluation of bacteriostatic & bactericidal
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.
this presentation gives informationabout microbial assay of vitamins B2 and B12. it is based upon the guidelines of indian pharmacopoeia. this presentation highlights the principle, process and applications of microbial assay
VIRUS PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-2Study of morphology, ...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-2Study of morphology, classification, reproduction/replication and cultivation of Virus. Introduction, Def General characteristics of Viruses: small size characteristic shapes, obligate intracellular parasites no built-in metabolic machinery no ribosomes
only one type of nucleic acid
do not grow in size. Morphology of Virus: Helical, Polyhedral (Icosahedral) Viral Envelop, Complex virus, Classification of virus. Viral Replication LIFE CYCLE OF BACTIRIOPHAGES Lytic cycle: Attachment, Penetration, Biosynthesis, Maturation and Release of progeny Phage Particles. The Lysogenic Cycle, Cultivation of virus : Animal inoculation, Embryonated eggs or chick embryo method and Tissue culture or cell culture: Organ cultures Explant culture and Cell culture. Types of cell culture
1.Primary cell culture: 2. Diploid cell culture (Semi-continuous cell lines):3. Heteroploid cultures (Continuous cell lines):
MULTIPLICATION OF HUMAN VIRUS:1. Attachment of Viral Particles 2. Penetration 3. Uncoating 4. Replication Of Viral Nucleic Acids And Translation Of The Genome 5) Maturation Or Assembly Of Virions. ) 6. Release Of Virions Into The Surrounding Environment
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.
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)
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
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
Designing of aseptic area including design, construction, service, flow chart,source of contamination, method of prevention of it,clean area classification as per USPDA.
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.
Designing of aseptic area, laminar flow equipment: Study of different source ...Ms. Pooja Bhandare
Designing of aseptic area, laminar flow equipment: Study of different source of contamination in aseptic area and methods of prevention, clean area classification. PHARMACEUTICALMICROBIOLOGY (BP303T)Unit-IVPart-1
Introduction: Designing of Aseptic Area . i) The clean-up area,
ii) The compounding area,
iii) The aseptic area,
iv) The quarantine area and
v) The packaging/labelling area.
Flow diagram of aseptic area. Floors, walls and ceilings, Doors, windows and services Personnel and protective clothing Cleaning and disinfection. Air Supply. Laminar flow equipment. Vertical laminar air flow bench
Horizontal laminar air flow bench
High Efficiency Particulate Air (HEPA) Filter. Operating Instructions Uses of Laminar Air Flow.Advantages of Laminar Air Flow.Limitations of Laminar Air Flow. Air flow pattern Unidirectional airflow
Non-unidirectional airflow
Combined airflow
Different Sources of Contamination in an Aseptic Area
1) Personnel:
2) Buildings and Facilities
3) Equipment and Utensils:
4) Raw Materials
5) Manufacturing Process:
Methods of Prevention of Contamination Clean Area Classification
Disinfectant - Pharmaceutical microbiology (Second year b.pharm) (3rd semester)Kiran Shinde
Prof.Mr.Kiran K. Shinde (M.Pharm), Assistant professor (VNIPRC)
Pharmaceutical microbiology (Second year b.pharm) (3rd semester)
Introduction
Classification & mode of action of disinfectant
Factors affecting disinfectant, antiseptics & their evaluation
Evaluation of bacteriostatic & bactericidal
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.
this presentation gives informationabout microbial assay of vitamins B2 and B12. it is based upon the guidelines of indian pharmacopoeia. this presentation highlights the principle, process and applications of microbial assay
VIRUS PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-2Study of morphology, ...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-2Study of morphology, classification, reproduction/replication and cultivation of Virus. Introduction, Def General characteristics of Viruses: small size characteristic shapes, obligate intracellular parasites no built-in metabolic machinery no ribosomes
only one type of nucleic acid
do not grow in size. Morphology of Virus: Helical, Polyhedral (Icosahedral) Viral Envelop, Complex virus, Classification of virus. Viral Replication LIFE CYCLE OF BACTIRIOPHAGES Lytic cycle: Attachment, Penetration, Biosynthesis, Maturation and Release of progeny Phage Particles. The Lysogenic Cycle, Cultivation of virus : Animal inoculation, Embryonated eggs or chick embryo method and Tissue culture or cell culture: Organ cultures Explant culture and Cell culture. Types of cell culture
1.Primary cell culture: 2. Diploid cell culture (Semi-continuous cell lines):3. Heteroploid cultures (Continuous cell lines):
MULTIPLICATION OF HUMAN VIRUS:1. Attachment of Viral Particles 2. Penetration 3. Uncoating 4. Replication Of Viral Nucleic Acids And Translation Of The Genome 5) Maturation Or Assembly Of Virions. ) 6. Release Of Virions Into The Surrounding Environment
Similar to Sterility testing products (solids, liquids, ophthalmic and other sterile products) according to IP, BP, USP. PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-6
Sterility Testing is defined as a testing which confirms that products are free from the presence of viable microorganisms. Sterility testing is very important for medical devices, pharmaceuticals, preparations, tissue materials and other materials that claim to be sterile or free from viable microorganisms.
In Process Quality Control Tests (IPQC) For Parenteral or Sterile Dosage FormsSagar Savale
These are the tests performed between QA and QC and provides for the authorization of approved raw materials for manufacturing based on actual laboratory testing generally called as IPQC such as physical, chemical, microbiologic and biologic tests.
Similar to Sterility testing products (solids, liquids, ophthalmic and other sterile products) according to IP, BP, USP. PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-6 (20)
Pharmaceutical Inorganic chemistry UNIT-V Radiopharmaceutical.pptx
Isotopes Types of decay
Alpha rays, which could barely penetrate a piece of paper
Beta rays, which could penetrate 3 mm of aluminium
Gamma rays, which could penetrate several centimetres of lead
Units of Radioactivity:
Measurement of Radioactivity
The measurement of nuclear radiation and detection is an important aspect in the identification of type of radiations (, , ) and to assay the radionuclide emitting the radiation, suitable detectors are required. The radiations are identified on the basis of their properties.
e.g. Ionization effect is measured in Ionization Chamber, Proportional Counter and Geiger Muller Counter.
The scintillation effect of radiation is measured using scintillation detector and the photographic effect is measured by Autoradiography.
Gas Filled Detectors:
Ionization Chamber:
Proportional Counters:
Geiger-Muller Counter
Properties of α, β, γ radiations
Half –life of Radioelement
Sodium Iodide (I131)
Handling and Storage of Radioactive Material:
Storage of Radioactive Substances –
Precautions For Handling Radioactive Substances
Labelling of Radioactive Substances
Pharmaceutical Application Of Radioactive Substances
Major extra and intracellular electrolytes. Pharmaceutical Inorganic chemistr...Ms. Pooja Bhandare
Major extra and intracellular electrolytes. Pharmaceutical Inorganic chemistry UNIT-II (Part-II)
Electrolyte: Intracellular fluid
Interstitial fluid
Plasma (Vascular fluid)
Anionic electrolytes- HCO₃⁻, Cl⁻, SO₄²⁻, HPO₄²⁻
Cationic electrolytes- Na⁺, K⁺, Ca²⁺, Mg²⁺
Concentration of important Electrolytes:
Electrolytes used in the replacement therapy: Sodium
chloride*, Potassium chloride, Calcium gluconate* and Oral Rehydration Salt
(ORS), Physiological acid base balance.
Acids, Bases And Buffers Pharmaceutical Inorganic chemistry UNIT-II (Part-I)
Acids, Bases are defined by Four main theories,
1.Traditional theory / concept
2.Arrhenius theory
3.Bronsted and Lowry theory
4.Lewis theory
Importance of acids and bases in pharmacy
Buffers: Buffer action
Buffer capacity Buffers system
Types of Buffers : Generally buffers are of two types:
1. Acidic buffers
2. Basic buffers
There are some other buffer system:
3. Two salts acts as acid-base pair. Ex- Potassium hydrogen phosphate and potassium dihydrogen phosphate.
4. Amphoteric electrolyte. Ex- Solution of glycine.
5. Solution of strong acid and solution of strong base. Ex- Strong HCl with KCl Mechanism of Buffer action: Mechanism of Action of acidic buffers: Buffer equation-Henderson-Hasselbalch equation:
Standard Buffer Solutions Preparation of Buffer Solutions: Buffers in pharmaceutical systems or Application of buffer: Stability of buffers Buffered isotonic solution Types of Buffer Isotonic solution
1. Isotonic Solutions:
2. Hypertonic Solutions:
3. Hypotonic Solution:
Measurement of Tonicity: 1. Hemolytic method: 2. Cryoscopic method or depression of freezing point:
Methods of adjusting the tonicity:
Class I methods:
In this type, sodium chloride or other substances are added to the solution in sufficient quantity to make it isotonic. Then the preparation is brought to its final volume withan isotonic or a buffered isotonic diluting solution.
These methods are of two types:
Cryoscopic method
Sodium chloride equivalent method.
Class II methods:
In this type, water is added in sufficient quantity make the preparation isotonic. Then the preparation is brought to its volume with an isotonic or a buffered isotonic diluting solution.
These methods are of two types:
White-Vincent method
Sprowls method.
Limt test Pharmaceutical Inorganic chemistry UNIT-I (Part-III) Limit Test.
Limit tests:- Factors affecting limit tests:
Specificity of the tests
Sensitivity
Control of personal errors (Analyst errors)
Test in which there is no visible reaction
Comparison methods
Quantitative determination
Limit test for Chloride: Principle, Procedure, observation and result.
Limit test for Sulphate: Principle, Procedure, observation and result
Limit test for Iron: Principle, Procedure, observation and result.
Limit test for Heavy metal: Principle, Procedure, observation and result.
Limit test for Lead: Principle, Procedure, observation and result.
Limit test for Arsenic: Principle, Gutzet test Procedure, detail in Gutzet Apparatus. observation and result.
Modifies Limit test for Chloride: Principle, Procedure, observation and result.
Modified Limit test for sulphate: Principle, Procedure, observation and result.
Types and Sources of impurities.pptx Pharmaceutical Inorganic chemistry UNIT-...Ms. Pooja Bhandare
Types and Sources of impurities. Pharmaceutical Inorganic chemistry UNIT-I (Part-II) Impurities:
Impure Chemical Compound
Pure Chemical Compound.
Types of impurities: Organic Impurity, Inorganic impurity, Residual solvent, Sources of Impurities in Pharmaceuticals
The different sources of impurities in pharmaceuticals are listed below:
Raw material used in manufacture
Reagents used in manufacturing process
Method/ process used in manufacture or method of manufacturing
Chemical processes used in the manufacture
Atmospheric contamination during the manufacturing process
Intermediate products in the manufacturing process
Defects in the manufacturing process
Manufacturing hazards
Inadequate Storage conditions
Decomposition of the product during storage
Accidental substitution or deliberate adulteration with spurious or useless materials.
Test for purity: Pharmacopoeia prescribes the “Test for purity” for pharmaceutical substances to check their freedom from undesirable impurities.
Pharmacopoeia will decide and fix the limit of tolerance for these impurities.
For certain common impurities for which pharmacopoeia prescribes the test of purity are:
Colour, odour, taste
Physicochemical constants (Iodine value, saponification value, melting point, refractive index etc.)
Acidity, alkalinity, pH
Humidity (Estimation of moisture)
Cations and anions
Insoluble Constituent or Residue.
Ash, Water insoluble ash
Arsenic or lead
Loss on drying
Loss on ignition.
Effect of Impurities
Introduction of Inorganic Chemistry, History of Pharmacopoeia.pptxMs. Pooja Bhandare
Introduction of Inorganic Chemistry, History of Pharmacopoeia, Pharmaceutical Chemistry, Inorganic Chemistry:
IMPORTANTS OF INORGANIC CHEMISTRY, Introduction of Pharmacopoeia, Types of Pharmacopoeia, History of pharmacopoeia, HISTROY OF INDIAN PHARMACOPOEIA
Content of pharmacopoeia Introduction including general Notices
Monographs of the official drugs
Appendices
Polyploidy, mutation and hybridization with reference to medicinal plants. PH...Ms. Pooja Bhandare
Polyploidy, mutation and hybridization with reference to medicinal plants. PHARMACOGNOSY & Phytochemistry-I (BP405T)Unit-IIPart-4
Polyploidy reference to medicinal plants.
Types Of Polyploidy
A. Euploidy
a.Autopolyploidy
b. Allopolyploidy
B. Aneuploidy
1. Causes Of Polyploidy
2. Non-disjunction in mitosis
3. Non-reduction in meiosis
4. Polyspermy
5. Endo-replication or Endo- reduplication.
Factors Promoting Polyploidy
1. Physical factor
2. Chemical factor
3. Biological factor
Physical factor:-
Temperature :- heat temperature & cold temperature
Centrifugation
X-rays
Gamma rays
Cosmic rays
Ionizing & non-ionizing radiations
UV-radiations
Chemical factor:-
Alkylating agents:- nitrogen & sulphur mustard
Acridines
Proflavins
Nitrous acid
Colchicines[6]
Colchicines (Poisonous alkaloids):-
Biological factor
Mode of reproduction
Mode of fertilization
Breeding system present (Hybridization)
Growth habit of the plant
Size of chromosomes
Application Of Polyploidy
Mutation breeding
Seedless fruits production
Bridge crossing
Ornamental & forage breeding
Disease resistance through aneuploidy
Industrial application of polyploidy
mutation reference to medicinal plants
Type of mutations:
1. Spontaneous and induced mutations.
2. Recessive and dominant mutations.
3. Somatic and germinal mutations.
4. Forward, back and suppressor mutation.
5. Chromosomal, genomic and point mutations
Application Of Mutation:
Hybridization reference to medicinal plants
The following steps are involved in hybridization of plant:
Choice Of Parents:.
Selfing Of Parents
Emasculation:.
Bagging:
Crossing Or Cross Pollination
Labelling
Collection Of Hybrid Seeds
Significance of Hybridization
PHARMACOGNOSY & Phytochemistry-I (BP405T)Unit-IIPart-2.FACTORS AFFECTING CULTIVATION
1. Altitude
2.Temperature
3. Rainfall
4. Day Length and Day Light
5. Soil
6. Soil Fertility
7. Fertilizers and Manures
a) Chemical fertilizers
(b) Manures
(c) Biofertilizers
8. Pests and Pests Control
a. Microbes
b) Insects
C) Non insect pests
d) Weeds
9. Other Factors that Affect the Cultivated Plants
a. Air Pollution
b. Herbicide
Cultivation and collections of drugs of natural origin..pptxMs. Pooja Bhandare
PHARMACOGNOSY & Phytochemistry-I (BP405T)Unit-IIPart-1Cultivation and collections of drugs of natural origin.
Advantages of cultivation
Methods of Plant Propagation
1.Sexual method (seed propagation)
2. Asexual method
Methods of sowing the seeds
Broadcasting Dibbling Miscellaneous
Special treatment to seeds
Asexual method.
Asexual method of vegetative propagation consists of three types:
a) Natural methods of vegetative propagation.
b) Artificial methods of vegetative propagation.
c) Aseptic method of micropropagation (tissue-culture).
COLLECTION OF CRUDE DRUGS
HARVESTING OF CRUDE DRUGS
DRYING OF CRUDE DRUGS
(1) natural (sun drying) and (2) artificial
Artificial Drying
Drying by artificial means includes drying the drugs in
(a) an oven; i.e. tray-dryers;
(b) vacuum dryers and
(c) spray dryers.
GARBLING (DRESSING)
PACKING OF CRUDE DRUGS
STORAGE & PRESEVATION OF CRUDE DRUGS
Quality control of Drugs of Natural Origin. PHARMACognosy & Phytochemistry-I ...Ms. Pooja Bhandare
Quality control of Drugs of Natural Origin PHARMACognosy & Phytochemistry-I (BP405T)Unit-I Part-3.
CONTENTS
Adulteration
Evaluation of adulteration
Morphological / Organoleptic evaluation
Microscopic evaluation
Quantitative evaluation
Physical evaluation
Chemical evaluation
Biological evaluation
Adulteration is of two types:
Indirect or Unintentional adulteration
Direct or Intentional adulteration
Intentional adulteration may be due to the following reasons
adulteration using manufactured substances
substitution using inferior commercial varieties
substitution using exhausted drugs
substitution of superficially similar inferior natural substance
adulteration using the vegetative part of the same plant
addition of toxic materials
adulteration of powders
addition of synthetic principles
Evaluation of Crude Drugs
1. ORGANOLEPTIC EVALUATION
2. MICROSCOPICAL EVALUATION
Stomatal index Vein-islet number
Veinlet termination number
Palisade ratio
Quantitative Microscopy (Lycopodium Spore Method)
3.CHEMICAL EVALUATION
4. Physical Evaluation
I. Solubility
II. Optical Rotation
III. Refractive Index
III. Specific Gravity
IV Viscosity
V. Melting Point
VI. Moisture Content
VII. Ultraviolet Light
VIII. Ash Values
Total ash
Acid-insoluble ash
The water-soluble ash
IX. Extractive Values
X. Foreign Organic Matters
5. BIOLOGICAL EVALUATION
Toxicity
Oxytocic activity
Microbiological assays
Classification of Crude Drugs. HARMACognosy & Phytochemistry-I (BP405T)Unit-I...Ms. Pooja Bhandare
Classification of Crude Drugs.PHARMACognosy & Phytochemistry-I (BP405T)Unit-I Part-2.
A method of classification should be:
a) simple,
b) easy to use, and
c) free from confusion and ambiguities.
TYPES OF CLASSIFICATION.
1.Alphabetical classification
2.Taxonomical classification
3.Morphological classification
4.Pharmacological classification
5.Chemical classification
6.Chemotaxonomical classification
7. Serotaxanomical Classification
Animal Cell Culture: Growth of animal cells in culture. PHARMACEUTICAL MICROB...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-VPart-4
Animal Cell Culture: Growth of animal cells in culture.
Introduction: Histroy, The culture media used for animal cell culture are classified as,
Natural, Artificial, Synthesized
Natural Culture Media:
a. Blood Plasma:
b. Blood Serum:
c. Tissue Extracts:
Artificial Media
Some common examples of artificial media are,
Minimal Essential Medium (MEM),
CMRL 1066,
RPMI 1640.
Synthetic media re classified as,
Serum Containing Media.
Serum Free Media.
a. Serum Containing Media:
b. Serum Free Media:
Physicochemical Parameters needed for growth animal cell culture:
General procedure for cell Culture.
Isolation of the tissue:
Disaggregation of the Tissue:
Mechanical disaggregation
b. Enzymatic Disaggregation
. Trypsin based disaggregation or trypsinization:
Warm trypsinization:
Cold trypsinization:
Drawbacks of trypsin disaggregation:
B. Collagenase based disaggregation:
C. Chelating Agents:
3. Seeding of Culture:
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
Classification and mode of action of disinfectants PHARMACEUTICAL MICROBIOLOG...Ms. Pooja Bhandare
PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-III Classification and mode of action of disinfectants. DISINFECTANT
Definition: Ideal properties of disinfectants: CLASSIFICATION OF DISINFECTANTS: Based on consistency 1. Liquid (E.g., Alcohols, Phenols) 2.Gaseous (Formaldehyde vapor, Ethylene oxide). Based on spectrum of activity 1. High level disinfectant
2. Intermediate level disinfectant
3. Low level disinfectant .Based on mechanism of action: 1.Action on membrane2.Denaturation of cellular proteins 3.Damage to nucleic acids 4.Oxidation of essential sulfhydryl groups of enzymes 5.Alkylation of amino-, carboxyl- and hydroxyl group. MODE OF ACTION AND APPICATION OF DISINFECTANT
Acid and alkalies
Halogens
Heavy metals
Phenols and its derivatives
Alcohol
Aldehydes
Dyes:
Quaternary ammonium compounds
Detergents and soaps.
A workshop hosted by the South African Journal of Science aimed at postgraduate students and early career researchers with little or no experience in writing and publishing journal articles.
Francesca Gottschalk - How can education support child empowerment.pptxEduSkills OECD
Francesca Gottschalk from the OECD’s Centre for Educational Research and Innovation presents at the Ask an Expert Webinar: How can education support child empowerment?
How to Make a Field invisible in Odoo 17Celine George
It is possible to hide or invisible some fields in odoo. Commonly using “invisible” attribute in the field definition to invisible the fields. This slide will show how to make a field invisible in odoo 17.
A review of the growth of the Israel Genealogy Research Association Database Collection for the last 12 months. Our collection is now passed the 3 million mark and still growing. See which archives have contributed the most. See the different types of records we have, and which years have had records added. You can also see what we have for the future.
Safalta Digital marketing institute in Noida, provide complete applications that encompass a huge range of virtual advertising and marketing additives, which includes search engine optimization, virtual communication advertising, pay-per-click on marketing, content material advertising, internet analytics, and greater. These university courses are designed for students who possess a comprehensive understanding of virtual marketing strategies and attributes.Safalta Digital Marketing Institute in Noida is a first choice for young individuals or students who are looking to start their careers in the field of digital advertising. The institute gives specialized courses designed and certification.
for beginners, providing thorough training in areas such as SEO, digital communication marketing, and PPC training in Noida. After finishing the program, students receive the certifications recognised by top different universitie, setting a strong foundation for a successful career in digital marketing.
Exploiting Artificial Intelligence for Empowering Researchers and Faculty, In...Dr. Vinod Kumar Kanvaria
Exploiting Artificial Intelligence for Empowering Researchers and Faculty,
International FDP on Fundamentals of Research in Social Sciences
at Integral University, Lucknow, 06.06.2024
By Dr. Vinod Kumar Kanvaria
Read| The latest issue of The Challenger is here! We are thrilled to announce that our school paper has qualified for the NATIONAL SCHOOLS PRESS CONFERENCE (NSPC) 2024. Thank you for your unwavering support and trust. Dive into the stories that made us stand out!
Model Attribute Check Company Auto PropertyCeline George
In Odoo, the multi-company feature allows you to manage multiple companies within a single Odoo database instance. Each company can have its own configurations while still sharing common resources such as products, customers, and suppliers.
Sterility testing products (solids, liquids, ophthalmic and other sterile products) according to IP, BP, USP. PHARMACEUTICAL MICROBIOLOGY (BP303T)Unit-IIIPart-6
1. PHARMACEUTICAL MICROBIOLOGY (BP303T)
Unit-III
Part-6
Sterility testing products (solids, liquids, ophthalmic and other
sterile products) according to IP, BP, USP.
Name: Ms. Pooja Deepak Bhandare
Assistant Professor
G H RAISONI UNIVERSITY
SCHOOL OF PHARMACY
2. Introduction
• Sterilization is the process of complete removal of microorganisms
from any surface or product.
• Sterility hence can be defined as “Complete freedom from
microorganisms”.
• The tests for sterility are conducted to make sure there is complete
absence of viable forms of microorganisms on a pharmaceutical
product.
• The products which are to be strictly supervised for absence of the
microorganisms are those who come directly in contact with the
systemic circulation e.g. ophthalmic preparations, parenteral
injections, implants, bandages, surgical dressings, needles, surgical
instruments etc.
• The test for sterility has to be conducted in aseptic environment so as to
avoid the contamination of the product.
3. • A pyrogen can be defined as “Fever Producing Substance”, which
induces “Fever (Hyperpyrexia)”.
• Bacterial endotoxins are the most common type of “Exogenous
Pyrogen”.
• Bacteria, viruses, malaria parasites and fungi on entering in the body
releases certain chemicals that act as Pyrogen.
• During the sterilization process these microorganisms die but presence of
pyrogens in the products may cause a febrile reaction.
• Test for sterility ensures absence of microorganisms while the
Pyrogen test ensures absence of pyrogens in the pharmaceutical
products.
• The lipopolysaccharide (LPS) from gram negative bacterias is a very
common example of pyrogen.
• The common pyrogen tests are,
• LAL Test (Limulus Amoebecyte Lysate Test).
• Sham Test (Rabbit Test).
4. Test for Sterility.
• Principle:
• A viable microorganism when provided with favorable conditions and
nutrients shows growth indicating its presence in the product.
• The probability of detecting the microbes in a product increases with
the number of the testings done.
• The external surfaces of the ampoules and vials or the containers
should be cleaned with the suitable antimicrobial substance and test be
carried out in an aseptic environment in order to avoid any contamination
of microbes in the products under testing.
5. Culture Media.
• Following culture media are used for conducting the sterility tests,
1. Fluid Thioglycollate Medium (FTM).
2. Alternative Thioglycollate Medium (ATM).
3. Soybean Casein Digest Medium (SCDM).
6. 1. Fluid Thioglycollate Medium (FTM)
• For use with Clear fluid products.
• Mainly intended for “Anaerobic Bacteria” but can also detect Aerobic
Bacterias.
• It is an indicator culture medium as it contains “Resazurin” a dye which
becomes pink on contact with oxygen.
• While using not more than one tenth of the upper portion of the medium
should have pink color.
• If more than one third of the medium is looking pink (Showing presence of
oxygen) it can be simply restored by reheating in the water bath until color
disappears.
• pH: 7.1 ± 0.2.
• Sterilization: Autoclaving @ 121℃ for 20 mins.
7. 2. Alternative Thioglycollate Medium (ATM):
• For use with turbid or viscous products and devices with small layers.
• It is an anaerobic medium i.e. made for cultivation of anaerobes and hence
anaerobic conditions must be followed while preparing and using.
• pH: 7.1 ± 0.2.
• Sterilization: Autoclaving @ 121℃ for 20 mins.
8. 3. Soybean Casein Digest Medium (SCDM):
• Suitable for cultivation of both fungi and aerobic
bacterias.
• pH: 7.3 ± 0.2.
• Sterilization: Autoclaving @ 121℃ for 20 mins.
9. Sr No. Component FTM ATM SCDM
1 L-Cysteine 0.5 gm 0.5 gm --
2 Sodium CHloride 2.5 gm 5.5 gm 5. gm
3 Dextrose (Monohydrate or
Anhydrous)
5.5 gm / 5.0 gm 5.5 gm / 5.0 gm 2.5 gm / 2.3 gm
4 Yeast Extract (Water Soluble) 5.0 gm 5.0 gm --
5 Pancreatic Digest of Casein 15.0 gm 15.0 gm 17.0 gm
6 Sodium Thioglycollate /
Thioglycolic Acid
0.5 gm / 0. ml 0.5 gm / 0.3 ml --
7 Papic digest of Soybean meal -- -- 3.0 gm
8 Dipotassium Hydrogen
Phosphate
-- -- 2.5 gm
9 Resazurin Sodium 1.0 ml -- --
10 Distilled water upto 1000 ml 1000 ml 1000 ml
11 pH after Sterilization by
Autoclave @ 121℃ for 20
mins.
7.1 ± 0.2 7.1 ± 0.2 7.3 ± 0.2
Table 2.1: Composition of the Culture Medias used for Sterility Test.
10. Tests for Culture Media:
• The media to be used in sterility testings should pass the following tests,
1. Sterility of Media.
2. Growth Promotion Test.
3. Test for Bacteriostatic and Fungistatic.
1. Sterility of Media:
• Incubate the portions of FTM and ATM @ 30-35℃ and SCDM at 20-25℃ and
incubate them for not less than 14 days.
• The presence of microbial growth fails the test, while absence of growth passes
the test
11. 2.Growth Promotion Test:
• This test is used to check the capability of the medium to promote the bacterial
growth.
• Medium sample from each autoclaved medium container is taken and inoculated with
about 100 test organisms of each “Test Strain” and incubated as per the requirements
of the given strain.
• The sample passess the test if it shows satisfactory growth of the organisms.
• The test is considered failed on inadequate growth of the microbes in the sample.
12. Medium Test Microorganisms Incubation Parameters
Temperature Duration in Days Conditions
FTM 1.Clostridium Sporogenes 30-35℃ 3 Anaerobic
2.Staphylococcus aureus 30-35℃ 3 Aerobic
3.Pseudomonas aeruginosa. 30-35℃ 3 Aerobic
ATM 1.Bacteroides vulgatus 30-35℃ 3 Anaerobic
2.Clostridium Sporogenes. 30-35℃ 3 Anaerobic
3.Bacillus subtilis. 30-35℃ 3 Aerobic
SCDM 1.Aspergillus brasiliensis 20-25℃ 5 Aerobic
2.Candida albicans 20-25℃ 5 Aerobic
3.Bacillus subtilis. 30-35℃ 3 Aerobic
Table 2.2: List of Test Microorganisms for the Growth Promotion Test of Media.
13. 3) Test for Bacteriostatic and Fungistatic:
• Prepare cultures of the test bacterias and fungi.
• Inoculate the required volume of culture media with 100 viable microorganisms.
• Add the required amount of the preparation under test to the half of the
containers containing medium and microbes.
• The half of the containers are kept as control.
• After specified time the growth of microbes is visually compared in test and control
containers.
• If the test preparation is found to be bacteriostatic or fungistatic it can be neutralized
using “Neutralising agents” as specified.
14. Table 2.2: List of Neutralizing substances for antimicrobial compounds.
Sr No. Antimicrobial
agent
Neutralizing Agent.
1 Penicillin Penicillinase
2 Streptomycin Streptomycin
phosphotransferase
3 Cephalosporins Cephalosporinase.
4 Sulfonamides P Amino benzoic Acid
15. Sterility Test Methods.
• Sterility tests can be carried out using one of the following
methods,
1. Methods A: Membrane Filtration.
2. Method B: Direct Inoculation.
16. 1. Method A: Membrane Filtration.
• This method is preferred when substance to be tested is,
a) an oil,
b) an ointment that can be put into solution,
c) a non bacteriostatic solid not readily soluble in the culture medium and
d) a soluble powder or a liquid that possesses inherent bacteriostatic and fungistatic properties.
• For liquid products where the volume > 100ml or more
• Precautions:
• A laminar sterile airflow cabinet – to avoid accidental contamination.
• Working conditions monitored regularly by sampling the air and surfaces of the working area.
17. • Apparatus:
• A suitable unit consists of a closed reservoir and a receptacle between which a
properly supported membrane of appropriate porosity is placed.
• Nominal pore size not > 0.45µm and diameter of approx. 47mm.
• Flow rate is adjusted to 55-75 ml of water / minute.
• Pressure 70 mm of Hg.
• Cellulose nitrate filter for aqueous, oily and weakly alcoholic solutions.
• Cellulose acetate solution for strong alcoholic solutions.
18. • Dilution Fluids:
• Fluid A: Dissolve 1g of peptic digest of animal tissue in water to make 1
liter, filter / centrifuge, adjust to pH 7.1±0.2, dispense into flasks in 100-
ml quantities and sterilise at 121º for 20 minutes.
• Fluid B: If the test sample contains lecithin or oil, use fluid A to each
liter of which has been added 1 ml of polysorbate 80, adjust to pH
7.1±0.2, dispense into flasks in 100-ml quantities and sterilise at 121º for
20 minutes.
• Quantities of sample to be used:
19. Table 3.1 Quantities of sample to be used:
Preparation Quantity in each container
of the preparation
Minimum quantity to be used for each culture medium
Liquids Less than 1 ml. Total contents of container.
1-40 ml Half of the container but not less than 1 ml.
40 ml to 100 ml. 20 ml
More than 100 ml 10% of the container but not less than 20 ml.
Antibiotic liquids 1 ml
Other soluble preparations The whole contents of each container to provide not less than 200mg.
Insoluble preparations ointments & creams. The whole contents of each container to provide not less than 200mg.
Solids Less than 50mg Total container.
50 mg - 300 mg Half of the container but not less than 50 mg.
300 mg - 5g 150 mg
More than 5 gm 500 mg
Catgut and other surgical sutures for veterinary use. Three sections of a strand each 30 cm long
Surgical dressings/ cotton/ gauze (In packages) 100 mg per package
Sutures and other individually packed single use material. Whole device
Other medical devices Whole device or material cut into pieces or diassembled
20. Method of test:
1) For aqueous solution: -
• Prepare each membrane by aseptically transferring fluid A + media + preparation being
examined. -
• Alternatively, first, combined quantities of preparation + prescribed in the two media.
• Draw the liquid rapidly through a filter with aid of vacuum.
• If the solution being examined has antimicrobial properties, wash the membrane by filtering
100ml of sterile fluid A, three times, or quantities should be sufficient to allow growth of small
inoculum of organisms.
• After filtration, aseptically remove membrane from holder, cut the membrane in half, immerse
one half of membrane in 100ml of soyabean-casein digest medium and incubate at 20º to 25º for
not < 7days.
• Similar, other membranes in FTM (30º to 35º).
21. 2) For liquids immiscible with aqueous vehicles and suspensions: -
• Same as above but add sufficient quantity of fluid A to achieve rapid filtration.
• Sometimes use sterile enzyme preparations such as penicillinase or cellulase to aid in
dissolving insoluble substances.
• If lecithin is there, use fluid B for diluting.
3) For oils and oily solutions:
• Low viscous, filter without dilution through dry memb.
• Viscous oils – dilute as per needed using sterile diluent like isopropyl myristate –
filter by applying pressure or suction gradually.
• Wash the membrane at least 3 times sterile fluid B (100 ml) or to heat not > than 45º
and use warm solutions for washing membrane.
22. 4) For ointments and creams:
• Dilute ointments in a fatty base and emulsions of the w/o type to give a fluid concentration of
1%w/v, by heating (if necessary), not > than 40º with a suitable sterile diluent.
5) For soluble solids:
• Dissolve substance in a suitable sterile solvent for each medium and carry out test described
under aqueous solutions.
6) For sterile devices:
• Aseptically pass a sufficient volume of fluid B through each of not less than 20 devices so that
not less than 100ml is recovered from each device.
• Collect fluids in sterile containers and filter the entire volume through a membrane filter funnel,
and follow the test as under for aqueous solutions.
23. 2. Method B Direct Inoculation
•The quantities of substance or preparation being examined
which is to be used for inoculation for culture media varies
according to quantity in each container.
24. 1) For aqueous solutions and suspensions:
• Remove liquid from test containers with a sterile pipette or with a sterile syringe
or a needle
• Aseptically transfer specified volume of the material from each container to a
vessel of the culture medium
• Mix the liquid with the medium but do not aerate excessively
• Incubate the inoculated media for not less than 14 days, unless otherwise
specified in the monograph.
• Incubate at 30º to 35º in case of fluid thioglycollate medium and at 20º to 25º in
case of soyabean casein digest medium.
• If a test sample renders the medium turbid, it is difficult to examine presence or
absence of microbial growth by visual examination.
• Transfer suitable portions of the same medium to fresh vessels between the third
and seventh days after the test is started.
• Continue incubation of transfer vessels for not less than 7 additional days after the
transfer and for a total of not less than 14 days
25. 2) For oils and oily solutions:
• In media, add 0.1% w/v of (4 –tert -octylphenoxy) polyethoxy ethanol, 1% w/v of polysorbate
80 or other suitable emulsifying agent, in an appropriate conc.
• Oil containing cultures should be shaken gently each day.
3) For ointments: -
• Preparation is diluted tenfold in a sterile diluent such as fluid B or any other aqueous vehicle
capable of dispersing test material homogeneously throughout the fluid mixture.
• Mix 10ml of fluid mixture with 80 ml of the medium and proceed the same as aqueous
solutions & suspension.
4) For solids: -
• Transfer the required quantity of the material to the medium.
26. 5) For sterile devices: -
• For articles of such size and shape as permit complete immersion in not more than
1000ml of culture medium test the intact article, using the appropriate media and
incubate same as 1).
• For transfusion or infusion assemblies or where the size of an item makes immersion
impracticable, flush the lumen of each of 20 units with a sufficient quantity of fluid
thioglycollate medium and soyabean casein medium to yield a recovery of not less than
15ml of each medium, and incubate with not less than 100 ml of each of two media.
• Where the presence of the specimen being tested interferes with the test because of
bacteriostatic action, rinse the article thoroughly with minimum amount of fluid A.
Recover the rinsed fluid and test as described for sterile devices under method A.
27. Observation and Interpretation of Results.
• At interval during incubation period, and at its conclusion, examine media for
macroscopic evidence of microbial growth.
• If no evidence of growth is found, the preparation being examined passes the test for
sterility.
• If evidence of microbial growth is found, reserve the containers, and it is demonstrated
that it is not due to preparation, hence the tests for sterility are invalid and may be
recommenced.
• Perform a retest using the same number of samples, volumes to be tested and the media
• if no evidence of microbial growth is then found, the preparation being examined passes the test
for sterility.
• If evidence of microbial growth is found, isolate and identify the organisms.
• If they are not readily distinguishable from those growing in the containers reserved in the first
test, the preparation being examined fails the test for sterility.
• If they are readily distinguishable from those growing in the containers reserved in the first test,
perform a second retest using twice the number of samples.
• If no evidence of microbial growth is found in the second retest, the preparation being examined
passes the test for sterility.
• If evidence of growth of any micro-organisms is found in second retest, the preparation being
examined fails the test for sterility.
28. Pyrogen Test Methods.
• Pyrogen tests can be carried out using one of the following
methods,
• Rabbit Test.
• LAL Test.
29. 1. Rabbit Test.
• For this test, three healthy rabbits are selected each weighing at least 1.5
kg.
• No rabbit should be selected if:
• 1. It has a normal temperature greater than 49.8°C.
• 2. It was used in a positive test during the last two weeks.
31. • The pyrogen testing is performed in an air-conditioned room.
• The food and water is withheld from the rabbit overnight.
• A clinical thermometer is inserted in the rectum of each rabbit to a depth of
not less than 7.5 cm.
• Two readings of the temperature of rabbit in normal conditions should be taken
at the interval of half an hour before the start of the test and the mean of both
should be calculated to determine the initial temperature.
• The equipment, injectors and needles used in the test should be pyrogen-
free.
• These should be washed with water for injection and then heated at 260°C for
two hours.
• The injection is warmed to 38°C before injecting to the rabbits.
• 0.5 to 1.0 ml per kg dose should be injected through the ear vein.
• Six readings of temperature are recorded at an interval of half an hour.
32. • Pyrogen Test Results:
• The response of each rabbit is detected by the difference of initial temperature and the
highest temperature recorded.
• The response of all three rabbits gives the sum of responses and can be concluded as:
• i) If the sum of responses does not exceed 1.4°C and any rabbit shows the response less than 0.6° C,
the product passes the test.
• ii) If the sum of responses is greater than 1.4 °C or any rabbit shows the response 0.6 or greater,
continue the test using 5 rabbits.
• iii) If the test is done using 5 rabbits, then if the sum of responses of all 5 rabbits is greater than
3.7°C and the individual response of not more than three rabbits is greater than 0.6°C, the product
passes the test.
33. 2. LAL Test.
• Also called “Limulus Amoebocyte Lysate Test” and “Bacterial Endotoxin
Test”.
• The LAL test reagent is prepared by lysing amoebocyte blood cells from
• American Horseshoe crab Limulus polyphemus and other species like
Tachyleus tridentus, Tachypleus gigas and Carcinoscropius rotundicauda.
34. • The horseshoe crab has blue colored blood.
• The amebocyte blood cells of horseshoe crab show defensive clotting
in presence of bacterial endotoxins.
• When a test material containing a pyrogen is added to the LAL reagent
it forms turbidity or a gel consistency indicating presence of
endotoxins.
• The test is 100 times more sensitive than the rabbit test.
• LAL reagent is available commercially and is a popular alternative to
animal tests.