The document discusses pyrogen testing and the Limulus Amebocyte Lysate (LAL) test. It begins by explaining that pyrogen testing is used to detect bacterial toxins in vaccines and drugs that could cause fever. The LAL test uses a lysate extracted from horseshoe crab blood cells to detect endotoxins. There are three main types of LAL tests: gel clot, turbidimetric, and chromogenic. The gel clot test detects endotoxins based on clotting. The turbidimetric test measures turbidity increases spectrophotometrically. The chromogenic test also uses spectrophotometry to measure color changes from a cleaved substrate. The document concludes
This document provides information on microbiological assays for vitamins B2 and B12. It discusses the underlying principles, which involve measuring the growth response of test microorganisms to different concentrations of the vitamin being assayed. Two common methods are described: the cylinder-plate method and the turbidimetric tube assay method. Specific details are given on reagents, preparation of inoculum, procedures, and interpretation of results for assays of vitamins B2 and B12 using Lactobacillus species as the test microorganisms.
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 document discusses microbiological assays for testing antibiotics. It describes biological assays using microorganisms and the two main types of microbiological assays: agar diffusion assays and turbidimetric assays. The agar diffusion assay uses discs containing antibiotics placed on an agar plate seeded with bacteria, and measures zones of inhibition of bacterial growth. The turbidimetric assay measures inhibition of bacterial growth in liquid culture containing known concentrations of antibiotics.
Microbiological assays use microorganisms to determine the potency of drugs. There are two main methods - the cylinder-plate method which measures inhibition zone diameters, and the turbidimetric method which measures absorbance changes in liquid cultures. Standard curves are prepared using known concentrations of a reference standard. Test samples are run alongside at assumed concentrations and their potency determined by comparing results to the standard curve. Proper preparation of media, buffers, microorganism cultures and standards is required for accurate and reproducible assays.
The document discusses various quality control methods for herbal drugs as outlined by WHO guidelines. It describes tests for powder fineness, foreign matter, macroscopic and microscopic examination, thin layer chromatography, determination of ash, extractable matter, water and volatile matter, volatile oils, bitterness value, haemolytic activity, and tannins. The methods provide quantitative and qualitative analysis of herbal drugs to ensure appropriate quality standards.
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic...Ms. Pooja Bhandare
The document discusses methods for assessing new antibiotics through microbiological assays. It describes how the minimum inhibitory concentration (MIC) can be determined using either liquid or solid dilution methods. The liquid dilution method involves setting up a series of test tubes with doubling dilutions of the antibiotic being tested and incubating with a test microorganism. The solid dilution method incorporates the antibiotic dilutions into an agar plate which is then inoculated. After incubation, the MIC is the lowest concentration that inhibits microbial growth. These assays help establish the efficacy and potency of new antibiotics.
The document discusses pyrogen testing and the Limulus Amebocyte Lysate (LAL) test. It begins by explaining that pyrogen testing is used to detect bacterial toxins in vaccines and drugs that could cause fever. The LAL test uses a lysate extracted from horseshoe crab blood cells to detect endotoxins. There are three main types of LAL tests: gel clot, turbidimetric, and chromogenic. The gel clot test detects endotoxins based on clotting. The turbidimetric test measures turbidity increases spectrophotometrically. The chromogenic test also uses spectrophotometry to measure color changes from a cleaved substrate. The document concludes
This document provides information on microbiological assays for vitamins B2 and B12. It discusses the underlying principles, which involve measuring the growth response of test microorganisms to different concentrations of the vitamin being assayed. Two common methods are described: the cylinder-plate method and the turbidimetric tube assay method. Specific details are given on reagents, preparation of inoculum, procedures, and interpretation of results for assays of vitamins B2 and B12 using Lactobacillus species as the test microorganisms.
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 document discusses microbiological assays for testing antibiotics. It describes biological assays using microorganisms and the two main types of microbiological assays: agar diffusion assays and turbidimetric assays. The agar diffusion assay uses discs containing antibiotics placed on an agar plate seeded with bacteria, and measures zones of inhibition of bacterial growth. The turbidimetric assay measures inhibition of bacterial growth in liquid culture containing known concentrations of antibiotics.
Microbiological assays use microorganisms to determine the potency of drugs. There are two main methods - the cylinder-plate method which measures inhibition zone diameters, and the turbidimetric method which measures absorbance changes in liquid cultures. Standard curves are prepared using known concentrations of a reference standard. Test samples are run alongside at assumed concentrations and their potency determined by comparing results to the standard curve. Proper preparation of media, buffers, microorganism cultures and standards is required for accurate and reproducible assays.
The document discusses various quality control methods for herbal drugs as outlined by WHO guidelines. It describes tests for powder fineness, foreign matter, macroscopic and microscopic examination, thin layer chromatography, determination of ash, extractable matter, water and volatile matter, volatile oils, bitterness value, haemolytic activity, and tannins. The methods provide quantitative and qualitative analysis of herbal drugs to ensure appropriate quality standards.
Microbiological Assay of Vitamin & Amino acid Assessment of a New Antibiotic...Ms. Pooja Bhandare
The document discusses methods for assessing new antibiotics through microbiological assays. It describes how the minimum inhibitory concentration (MIC) can be determined using either liquid or solid dilution methods. The liquid dilution method involves setting up a series of test tubes with doubling dilutions of the antibiotic being tested and incubating with a test microorganism. The solid dilution method incorporates the antibiotic dilutions into an agar plate which is then inoculated. After incubation, the MIC is the lowest concentration that inhibits microbial growth. These assays help establish the efficacy and potency of new antibiotics.
This document presents information about anti-bacterial activity screening techniques. It discusses bacteriostatic agents, which prevent bacterial growth without killing bacteria, and assessments of bacteriostatic activity including serial dilution methods in fluid and solid media as well as cup plate and gradient plate methods. It also discusses bactericidal agents, which kill bacteria, and assessments of bactericidal activity including end-point or extinction time methods such as the Phenol coefficient test and varying the concentration or contact time of disinfectants. The document was presented to Dr. Chaluvaraju KC by Mr. Pradeep on the topic of anti-bacterial activity and screening techniques.
The document discusses three main methods for the bacterial endotoxin test - gel clot, turbidimetric, and chromogenic. The gel clot method is the simplest but least quantitative, while turbidimetric and chromogenic methods allow for more automation and precision using spectrophotometry. All three methods use Limulus amebocyte lysate and detect endotoxins through coagulation reactions. The choice of method depends on factors like testing volumes, sample properties, required sensitivity, and compliance needs. Photometric methods have advantages of automation and precision but higher costs, while gel clot is inexpensive but less quantitative.
The document discusses three methods for isolating pure cultures of microbes: streak plate technique, pour plate technique, and spread plate technique. The streak plate technique involves spreading microbial culture on media with a sterilized inoculating needle. The pour plate technique mixes culture with liquid agar before pouring into plates, trapping some microbes beneath the surface. The spread plate technique spreads diluted culture samples on the agar surface and is best for isolating pure colonies as it only produces surface colonies.
Process of implementing and developing technical standards based on the consensus of different parties that include firms, users, interest groups, standards organizations and governments
Microbial Assay of Antibiotics
STANDARD PREPARATION AND UNITS OF ACTIVITY
Preparation of media
Buffer solutions
Standard solution
Sample solution
Test organisms
Preparation of inoculum Method -1
Method 2
Method 3
Method 4
Determination of Inoculum
Apparatus
Assay design
Assay method
cylinder plate method
One level assay with standard curve
Estimation of potency
Turbidimetric method
Sterility testing is required for all products labeled as sterile to ensure they have been effectively sterilized. Tests are conducted using specific culture media and procedures to detect any viable bacteria, fungi, or yeasts. The USP outlines sterility testing methods for various pharmaceutical products and devices, including membrane filtration and direct inoculation. Interpretation of results involves incubating samples and checking for any microbial growth over time, with growth indicating test failure.
This document discusses and compares herbal drugs and synthetic drugs. It notes that herbal drugs are derived from plants and have been widely used throughout history. They tend to have fewer side effects and be more affordable than synthetic drugs, though they may lack dosage instructions and have interactions. Synthetic drugs are chemically produced substances like marijuana, cocaine, and methamphetamines. They have more consistent quality but can cause issues like violent behavior and memory problems. Both have benefits and risks, and differ in aspects like potency, costs, and targets in the body. Commonly used herbal medicines include glucosamine for joint pain.
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
This document discusses pyrogen testing methods. Pyrogens are fever-inducing substances, mainly lipopolysaccharides of bacterial origin. The rabbit pyrogen test, introduced in 1942, involves measuring temperature increases in rabbits injected with a test solution. If temperature increases exceed thresholds, the sample fails the test. The Limulus amebocyte lysate (LAL) test directly measures endotoxins using a lysate from horseshoe crab blood. Both tests are used to ensure medical products are free of pyrogens.
The document discusses the scope of pharmaceutical microbiology. It covers several key points:
1) Pharmaceutical microbiology involves the study of microorganisms like viruses, bacteria, fungi, and protozoa that are important for fields like medicine, pharmaceuticals, and biotechnology.
2) Areas of pharmaceutical microbiology include production of antibiotics, vaccines, enzymes and other drugs using microbes, ensuring sterility of drugs to prevent contamination, and using microbes and their components in medical research.
3) Important applications are using microbes or their products to produce therapeutic agents, vaccines, enzymes for fermentation, and ensuring sterility of medical devices, cosmetics and other products through microbiological quality control.
Cloning is the asexual reproduction of plants through vegetative propagation techniques like taking cuttings, division, or grafting. This allows the production of exact genetic copies of plants. When taking cuttings for cloning, a portion of the plant with stem and leaves is placed in a sterile medium and grown into a new plant. Clones have advantages like allowing reproduction of superior cultivars, disease resistance, and overcoming issues with seeds like low germination rates or long juvenile periods before flowering.
Object
Theory
Definition
Types
Filter paper method
Cup plate method
Ditch plate method
Method of inoculation
Phenol coefficient test
Material required
Procedure
Enumeration is counting of microorganisms present in a sample.
This is done to know the intense of presence of the spoilers in the spoiled food.
To detect which type of organism is responsible for the spoilage.
Mostly this is done two important methods.
Viable count
Total count
VIABLE COUNT:
A viable cell count allows one to identify the number of actively growing or dividing cells in a sample.
The plate count method or spread plate method relies on bacteria growing a colony on a nutrient medium.
Number of colonies can be counted.
Plate count agar is used for general count
MacConkey agar is used for Gram negative organisms.
TOTAL COUNT:
The initial analysis is done by mixing serial dilution of sample in liquid nutrient agar which is then poured into bottles.
The bottles are then sealed and laid on their sides to produce a slopping agar surface.
The colonies are then counted by eye.The total number of colonies are said as Total Viable Count. The initial analysis is done by mixing serial dilution of sample in liquid nutrient agar which is then poured into bottles.
The bottles are then sealed and laid on their sides to produce a slopping agar surface.
The colonies are then counted by eye.The total number of colonies are said as Total Viable Count.
Pour plate method:
The same procedure is done for this till serial dilution.
The serially diluted sample is then mixed with the molten nutrient agar.
Then poured onto the sterile petridish.
Incubated under appropriate temperature amd the colonies where counted.
ConclusionThe enumeration of these spoiled food samples are important to encounter the type of microbe is causing the spoilage.
And hence this is used to prevent the same type of spoilage.
This can be avoided by making the environmental changes which inhibits the organism which is responsible for the spoilage.
This document discusses bio-insecticides, which are organic formulations used to control insects that damage crops. Bio-insecticides use microorganisms or their toxins, including bacteria, viruses, fungi, protozoa, and nematodes. These organisms act as natural insecticides by producing toxins or by infecting and killing insects. The document categorizes different types of bio-insecticides and provides examples and modes of action for each type. It also outlines advantages like specificity and safety compared to chemical pesticides, as well as disadvantages like slower action and susceptibility to environmental factors.
This document discusses various physical parameters and methods used to evaluate crude drugs, including ash values, swelling factor, extractive values, and bioassay. It describes determining total ash value, acid insoluble ash value, sulphated ash value, and water soluble ash value. Methods are provided for measuring swelling factor and water soluble, alcohol soluble, and ether soluble extractive values. Finally, it outlines using bioassay to evaluate drug activity through tests on living organisms.
This document summarizes the Limulus Amebocyte Lysate test (LAL test), which is used to detect endotoxins from gram-negative bacteria. It discusses that endotoxins are part of the cell wall of gram-negative bacteria and are released when the bacteria die. The LAL test was developed in the 1960s and works by detecting the clotting reaction that occurs when horseshoe crab blood cells, called amebocytes, come into contact with endotoxins. The document describes the three techniques used in the LAL test - gel clot, turbidimetric, and chromogenic - and provides details on how the test is performed and interpreted.
Role of biomarkers and dna fingerprinting in herbal drug standardisationRoshni Ann
1. DNA contains the genetic instructions that determine an organism's characteristics. DNA is organized into genes located on chromosomes within cells.
2. The document discusses several techniques used for DNA fingerprinting, including microsatellites, restriction fragment length polymorphisms, amplified fragment length polymorphism, and random amplified polymorphic DNA.
3. DNA fingerprinting can be used to identify plant species and strains, helping to standardize herbal drugs and ensure quality. It has applications in forensics, ancestry tracing, and tracing the evolution of plants and microorganisms.
The document discusses various methods for testing the efficacy of disinfectants, including:
1. Koch's method, which tests the ability of a disinfectant to kill Bacillus anthraces spores.
2. Rideal Walker and Chick-Martin tests, which determine the phenol coefficient of a disinfectant by comparing its bactericidal effects to phenol under clean and dirty conditions.
3. In-use and capacity use dilution tests assess the ability of a disinfectant to kill microbes when diluted in conditions mimicking actual use over time in the presence of organic matter.
4. No single test can reliably determine a disinfectant's efficacy
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.
This document presents information about anti-bacterial activity screening techniques. It discusses bacteriostatic agents, which prevent bacterial growth without killing bacteria, and assessments of bacteriostatic activity including serial dilution methods in fluid and solid media as well as cup plate and gradient plate methods. It also discusses bactericidal agents, which kill bacteria, and assessments of bactericidal activity including end-point or extinction time methods such as the Phenol coefficient test and varying the concentration or contact time of disinfectants. The document was presented to Dr. Chaluvaraju KC by Mr. Pradeep on the topic of anti-bacterial activity and screening techniques.
The document discusses three main methods for the bacterial endotoxin test - gel clot, turbidimetric, and chromogenic. The gel clot method is the simplest but least quantitative, while turbidimetric and chromogenic methods allow for more automation and precision using spectrophotometry. All three methods use Limulus amebocyte lysate and detect endotoxins through coagulation reactions. The choice of method depends on factors like testing volumes, sample properties, required sensitivity, and compliance needs. Photometric methods have advantages of automation and precision but higher costs, while gel clot is inexpensive but less quantitative.
The document discusses three methods for isolating pure cultures of microbes: streak plate technique, pour plate technique, and spread plate technique. The streak plate technique involves spreading microbial culture on media with a sterilized inoculating needle. The pour plate technique mixes culture with liquid agar before pouring into plates, trapping some microbes beneath the surface. The spread plate technique spreads diluted culture samples on the agar surface and is best for isolating pure colonies as it only produces surface colonies.
Process of implementing and developing technical standards based on the consensus of different parties that include firms, users, interest groups, standards organizations and governments
Microbial Assay of Antibiotics
STANDARD PREPARATION AND UNITS OF ACTIVITY
Preparation of media
Buffer solutions
Standard solution
Sample solution
Test organisms
Preparation of inoculum Method -1
Method 2
Method 3
Method 4
Determination of Inoculum
Apparatus
Assay design
Assay method
cylinder plate method
One level assay with standard curve
Estimation of potency
Turbidimetric method
Sterility testing is required for all products labeled as sterile to ensure they have been effectively sterilized. Tests are conducted using specific culture media and procedures to detect any viable bacteria, fungi, or yeasts. The USP outlines sterility testing methods for various pharmaceutical products and devices, including membrane filtration and direct inoculation. Interpretation of results involves incubating samples and checking for any microbial growth over time, with growth indicating test failure.
This document discusses and compares herbal drugs and synthetic drugs. It notes that herbal drugs are derived from plants and have been widely used throughout history. They tend to have fewer side effects and be more affordable than synthetic drugs, though they may lack dosage instructions and have interactions. Synthetic drugs are chemically produced substances like marijuana, cocaine, and methamphetamines. They have more consistent quality but can cause issues like violent behavior and memory problems. Both have benefits and risks, and differ in aspects like potency, costs, and targets in the body. Commonly used herbal medicines include glucosamine for joint pain.
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
This document discusses pyrogen testing methods. Pyrogens are fever-inducing substances, mainly lipopolysaccharides of bacterial origin. The rabbit pyrogen test, introduced in 1942, involves measuring temperature increases in rabbits injected with a test solution. If temperature increases exceed thresholds, the sample fails the test. The Limulus amebocyte lysate (LAL) test directly measures endotoxins using a lysate from horseshoe crab blood. Both tests are used to ensure medical products are free of pyrogens.
The document discusses the scope of pharmaceutical microbiology. It covers several key points:
1) Pharmaceutical microbiology involves the study of microorganisms like viruses, bacteria, fungi, and protozoa that are important for fields like medicine, pharmaceuticals, and biotechnology.
2) Areas of pharmaceutical microbiology include production of antibiotics, vaccines, enzymes and other drugs using microbes, ensuring sterility of drugs to prevent contamination, and using microbes and their components in medical research.
3) Important applications are using microbes or their products to produce therapeutic agents, vaccines, enzymes for fermentation, and ensuring sterility of medical devices, cosmetics and other products through microbiological quality control.
Cloning is the asexual reproduction of plants through vegetative propagation techniques like taking cuttings, division, or grafting. This allows the production of exact genetic copies of plants. When taking cuttings for cloning, a portion of the plant with stem and leaves is placed in a sterile medium and grown into a new plant. Clones have advantages like allowing reproduction of superior cultivars, disease resistance, and overcoming issues with seeds like low germination rates or long juvenile periods before flowering.
Object
Theory
Definition
Types
Filter paper method
Cup plate method
Ditch plate method
Method of inoculation
Phenol coefficient test
Material required
Procedure
Enumeration is counting of microorganisms present in a sample.
This is done to know the intense of presence of the spoilers in the spoiled food.
To detect which type of organism is responsible for the spoilage.
Mostly this is done two important methods.
Viable count
Total count
VIABLE COUNT:
A viable cell count allows one to identify the number of actively growing or dividing cells in a sample.
The plate count method or spread plate method relies on bacteria growing a colony on a nutrient medium.
Number of colonies can be counted.
Plate count agar is used for general count
MacConkey agar is used for Gram negative organisms.
TOTAL COUNT:
The initial analysis is done by mixing serial dilution of sample in liquid nutrient agar which is then poured into bottles.
The bottles are then sealed and laid on their sides to produce a slopping agar surface.
The colonies are then counted by eye.The total number of colonies are said as Total Viable Count. The initial analysis is done by mixing serial dilution of sample in liquid nutrient agar which is then poured into bottles.
The bottles are then sealed and laid on their sides to produce a slopping agar surface.
The colonies are then counted by eye.The total number of colonies are said as Total Viable Count.
Pour plate method:
The same procedure is done for this till serial dilution.
The serially diluted sample is then mixed with the molten nutrient agar.
Then poured onto the sterile petridish.
Incubated under appropriate temperature amd the colonies where counted.
ConclusionThe enumeration of these spoiled food samples are important to encounter the type of microbe is causing the spoilage.
And hence this is used to prevent the same type of spoilage.
This can be avoided by making the environmental changes which inhibits the organism which is responsible for the spoilage.
This document discusses bio-insecticides, which are organic formulations used to control insects that damage crops. Bio-insecticides use microorganisms or their toxins, including bacteria, viruses, fungi, protozoa, and nematodes. These organisms act as natural insecticides by producing toxins or by infecting and killing insects. The document categorizes different types of bio-insecticides and provides examples and modes of action for each type. It also outlines advantages like specificity and safety compared to chemical pesticides, as well as disadvantages like slower action and susceptibility to environmental factors.
This document discusses various physical parameters and methods used to evaluate crude drugs, including ash values, swelling factor, extractive values, and bioassay. It describes determining total ash value, acid insoluble ash value, sulphated ash value, and water soluble ash value. Methods are provided for measuring swelling factor and water soluble, alcohol soluble, and ether soluble extractive values. Finally, it outlines using bioassay to evaluate drug activity through tests on living organisms.
This document summarizes the Limulus Amebocyte Lysate test (LAL test), which is used to detect endotoxins from gram-negative bacteria. It discusses that endotoxins are part of the cell wall of gram-negative bacteria and are released when the bacteria die. The LAL test was developed in the 1960s and works by detecting the clotting reaction that occurs when horseshoe crab blood cells, called amebocytes, come into contact with endotoxins. The document describes the three techniques used in the LAL test - gel clot, turbidimetric, and chromogenic - and provides details on how the test is performed and interpreted.
Role of biomarkers and dna fingerprinting in herbal drug standardisationRoshni Ann
1. DNA contains the genetic instructions that determine an organism's characteristics. DNA is organized into genes located on chromosomes within cells.
2. The document discusses several techniques used for DNA fingerprinting, including microsatellites, restriction fragment length polymorphisms, amplified fragment length polymorphism, and random amplified polymorphic DNA.
3. DNA fingerprinting can be used to identify plant species and strains, helping to standardize herbal drugs and ensure quality. It has applications in forensics, ancestry tracing, and tracing the evolution of plants and microorganisms.
The document discusses various methods for testing the efficacy of disinfectants, including:
1. Koch's method, which tests the ability of a disinfectant to kill Bacillus anthraces spores.
2. Rideal Walker and Chick-Martin tests, which determine the phenol coefficient of a disinfectant by comparing its bactericidal effects to phenol under clean and dirty conditions.
3. In-use and capacity use dilution tests assess the ability of a disinfectant to kill microbes when diluted in conditions mimicking actual use over time in the presence of organic matter.
4. No single test can reliably determine a disinfectant's efficacy
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.
ANTIBIOTIC SENSITIVITY TEST
Tube dilution and agar plate method.
Filter paper and cup plate method.
Ditch-plate method.
Phenol coefficient method.
Kelsey Sykes method.
The document outlines various methods used to test the efficacy of disinfectants, including carrier tests, suspension tests, and practical tests. Carrier tests involve contaminating a thread with bacteria and exposing it to disinfectants. Suspension tests measure a disinfectant's ability to kill bacteria suspended in its solution. Practical tests evaluate disinfectants under real-world conditions. The document also describes the phenol coefficient test, which compares a disinfectant's effectiveness to that of phenol, and the filter paper test, which detects zones of bacterial inhibition around treated disks.
This document summarizes testing methods used to evaluate the efficacy of disinfectants. It describes two main types of tests: suspension tests where bacterial cultures are exposed to disinfectant solutions to see if organisms are killed, and in-use tests where disinfectant samples are collected from hospitals and labs and cultured to detect any contamination. Suspension tests provide either qualitative or quantitative results, with quantitative tests determining the level of organisms killed. An ideal disinfectant is one that can kill at least 99.999% of germs as determined by these standardized evaluation methods.
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Evaluation of Antimicrobial potency of Disinfectants,ANTI-INEFFECTIVE.pptx
1. Evaluation of Antimicrobial potency of
Disinfectants, Anti-infective and antibiotics(.Phenol
Coefficient Method,Tube dissolution Method,Agar
Plate Method )
PRESENTED BY AREEBA SHAFIQ
M.PHILL PHARMACEUTICS SCHOLAR (ISLAMIA UNIVERSITY
BAHAWALPUR,PAKISTAN)
2. Contents
Disinfectants
Evaluation of Antimicrobial Potency of Disinfectants
Anti-Infective and Antibiotics
Evaluation of Antimicrobial Potency of Anti-Infective and
Antibiotics .
3. Disinfectants
• A disinfectant is a chemical substance or compound used to
inactivate or destroy microorganisms on inert surfaces.
• Disinfection does not necessarily kill all microorganisms, especially
resistant bacterial spores; it is less effective than sterilization, which
is an extreme physical or chemical process that kills all types of
life.
5. Need for testing the disinfectants
A disinfectant must be tested
• To know the required effective dilution.
• To know the time taken for the onset of action.
• Periodical monitoring of its activity. As disinfectants are known to
loose their action on long standing & in the presence of organic
matter their efficacy must be tested periodically.
6. Evaluation of Antimicrobial potency of Disinfectant
Phenol Coefficient Method
• The measure of the disinfecting power of a substance, determined by dividing the figure
indicating the degree of dilution of the disinfectant that kills a microorganism within a
given time by that indicating the degree of dilution of phenol killing the microorganism
under similar conditions.
Two types of phenol coefficient tests are done:
1. Rideal Walker method
2. Chick Martin test
7. 1.Rideal Walker Method
Phenol coefficient test is suitable for testing disinfectants miscible
with water and which exert their antimicrobial action in manner
similar to that of phenol.
• Test Organism: Salmonella typhi
• Standard disinfectant: Phenol
8. • Different dilutions of the test disinfectants and phenol are prepared and 5ml of each
dilution is inoculated with 0.5 ml broth culture of the organisms for 24 hr.
• All tubes (disinfectants + organisms and phenol + organisms)are placed in 17.5 °C water
bath.
• Subcultures of each reaction mixture are taken and transferred to 5ml sterile broth after
2.5, 5, 7.5 and 10 min.
• The broth tubes are incubated at 37 °C for 48 to 72 hr and are examined for presence or
absence of growth.
9. • If a phenol coefficient or Rideal-Walker coefficient of a given test disinfectant
is 1, it means that disinfectant has same effectiveness as phenol.
• If a phenol coefficient or Rideal-Walker coefficient of a given test disinfectant
is less than 1, it means that disinfectant is less effective than phenol.
• If a phenol coefficient or Rideal-Walker coefficient of a given test disinfectant
is more than 1, it means that disinfectant is more effective than phenol.
• If the phenol coefficient of the test disinfectant is 20 it means that the
disinfectant is 20 times more active than phenol.
10. 2.Chick Martin
• Chick martin test incorporates the presence of organic matter as the test is not carried
out in the water but yeast suspension or 4% dried human feces.
• The total time of the test is 30 minutes.
• Both S. typhi and S. aureus cultures are used to test the efficacy of disinfectants.
• The calculation method is the same as that Rideal Walker test.
11. • To calculate for phenol coefficient, the number indicating the degree of dilution of
the disinfectant in which it kills the microorganism within a given time is divided
by the number indicating the degree of dilution of phenol in which the latter kills
the microorganism in the same period of time and under the same conditions.
• A phenol coefficient that is greater than 1 indicates that the disinfectant is more
effective than phenol. In contrast, a phenol coefficient that is lower than 1 means
the disinfectant is less effective than phenol.
13. Benefits
Cheaply carried out and quickly Analyzable
Effectiveness of the disinfectant can be expressed quantitatively.
14. Anti –Infective Agent
An agent that is capable of acting against infection, either by inhibiting the spread of an
infectious agent or by killing the infectious agent .
Antibiotics
Antibiotics are medicines that fight bacterial infections in people and animals. They
work by killing the bacteria or by making it hard for the bacteria to grow and multiply.
Evaluation of Antmicrobial Potency of Anti-
Infective Agent and Antibiotics
15. Need for testing Antibiotics
Help find out which antibiotic will be most effective in treating
your infection.
To obtain more realistic and precise measurement of potency to
overcome the antibiotic resistance problem.
16. 1.Agar Plate Method
1.Preparation of Inoculum
From each inoculated agar plate, a minimum of four colonies were touched with
a sterile loop and transferred into a tube containing normal saline (0.9%) or in
broth and density of each microbial suspension was adjusted equal to that of 108
cfu/ml (standardized by 0.5McFarland standard) and was used as the inoculum.
2.Inoculation of Test Plate
The test is performed by applying a bacterial inoculum of approximately 1–
2×108CFU/mL to the surface of a large (150 mm diameter) Mueller-Hinton agar
plate.
17. 3. Application of Disks
Then, filter paper discs (about 6 mm in diameter), containing the test
compound(Antibiotics ) at a desired concentration, are placed on the agar
surface with sterile Forceps or a suitable disc dispenser.
4.Incubation
Plates are incubated for 16–24 h at 35°C prior to determination of results.
18. 5.Zone of Inhibition
Generally, antimicrobial agent diffuses into the agar and inhibits
growth of the test microorganism . Measuring the diameter of zone of
Inhibition by using a ruler or Sliding Caliper.
19. Benefits
Easy to perform
Inexpensive
Allow Visibility of growth
Correct Inoculum
20.
21. 2. Tube Dilution Method
• Used to determine minimal concentration of antibiotic to inhibit or kill
the microorganism.
• Achieved by dilution of antibiotic in either agar or broth media.
Minimum Inhibitory Concentration:
The lowest concentration of drug that inhibit the growth of the bacteria
isolated from the patient.
22. In this method we use Sterile Muller Hinton Broth
We make 2 folds dilution of Antibiotics in the broth i.e 2µg/ml,4µg/ml
,8µg/ml,16µg/ml and so on.
Then we add broth culture (0.1ml) of test organism to the Prepared
dilutions.
One tube without antibiotic served as the organism control.
After adding test organism we incubate the tubes at 37ͦC for 24 hours .
Examine tubes for visible signs of bacterial growth. The tube exhibiting no
visible growth and containing the least amount of antibiotic was
considered the minimal inhibitory concentration (MIC).
23.
24. Benefits
Standard method for determining level of Microbial Resistance to
an antimicrobial agent .
The MIC provides the ability to precisely determine the
concentration of antibiotic required to inhibit growth of a pathogen.