Parenteral Products - Evaluation
There are various reasons due to which the evaluation of parenteral products is important such as the safety, efficacy, quality of the parenteral dosage form, stability of the parenteral, and compatibility of the parenteral dosage form with other medicaments. So, it is very important to measure the effectiveness of the parenteral products by evaluating the parenteral.Evaluation of Parenteral Products
There are the following types of tests that are performed for the evaluation of parenteral products.(a) Leaker test
Leakage may occur when there is a gap present in the packing wall due to which gas can enter under the action of pressure or due to the concentration difference that presents across the wall.If there are any small cracks or pores present in the ampoules, it can allow the microbes or any other contaminants to enter the ampoules due to which leakage of the contents to outside the ampoules may occur. Due to this contamination of the sterile products may occur, and the appearance of the package may also be spoiled.
During the storage of parenteral preparations, temperature changes may occur then which may lead to the expansion as well as contraction of ampoules and their contents, by putting stress upon them if any opening is present.
Leaker tests are generally performed to detect ampoules that are not properly sealed. If it is found then it can be taken out or removed from the batch to maintain the sterile conditions of the formulations.
Tip-sealed ampoules are not properly closed as compared to pull-sealed ampoules. At the point of the seal if any cracks remain that results in leakage.
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Procedure
In this process, the leakers are determined visually.
Step 1 – The ampoules are placed into a vacuum chamber.
Step 2 – Now, submerged completely in 0.5-1% methylene blue which is a solution of a colored dye.
Step 3 – Now, the pressure that is applied to the ampoule should be negative. As a result of the pressure outside causing the dye to penetrate when opening, it becomes clear after washing the ampoules.
Step 4 – About 27 inches Hg of vacuum should be released after 30 minutes.
Step 5 – The detection of the leaker is very clear when ampoules are dipped into the bath of dye at the time of autoclaving. Due to this leaker detection and sterilization occur in one operation.
Result – Within the leaker, the color from the dye will be visible.
Disadvantages
The main disadvantage of the pyrogen test is that the smaller diameter and capillaries of 15 microns in not detected properly.
As the rubber closer is not rigid so the bottles and the vials are not subjected to be tested.
(b) Pyrogen test
(i) LAL Bacterial Endotoxin Test
This test is also known as the Limulus amebocyte lysate (LAL) test.The LAL bacterial endotoxin test is used to determine the concentration and the presence of bacterial endotoxins in the drugs as well as in the biological products.
The endotoxins are the type of pyrogens and lipopolysaccharides that are present in the cell wall of gram-negative bacteria.
Pyrogens are generally fever-inducing substances that may be harmful if administered into the human body above the defined concentrations.
The water may also be the source of pyrogens, so it is very important to monitor the water system from time to time with the help of using the bacterial endotoxins test.
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Procedure
The solution of endotoxins that contains preparation is added to the lysate (It is derived from the horseshoe crabs' hemolymph cells).
1) Take the testing mixture for endotoxin content.
2) Mix the mixture with reagents that cause reactions resulting in turbidity, precipitation, or gelation.
3) Now, observe the turbidity, and precipitation, and use it as a quantitative measure to calculate the content of endotoxin.
4) Note that the rate of the reaction depends upon the concentration of endotoxins, pH, temperature, and the presence of clotting enzymes and clottable proteins from the lysate.
5) Express the number of endotoxins in defined endotoxin units (EU).
6) Use the formula k/M to calculate the endotoxin limit for the specified test preparation, where M is the maximum dosage given to an adult per kg/hr.
7) Use a value of K of 5.0 EU/kg for parenteral preparations and 0.2 EU/kg for intrathecal preparations in the expression.
8) Calculate the endotoxin limit for the given test preparation using the calculated values of k and M.
9) Compare the calculated endotoxin limit with the observed endotoxin content to determine if it is within acceptable limits.
10) Repeat the procedure with different samples to ensure consistency of results.
1) Take the testing mixture for endotoxin content.
2) Mix the mixture with reagents that cause reactions resulting in turbidity, precipitation, or gelation.
3) Now, observe the turbidity, and precipitation, and use it as a quantitative measure to calculate the content of endotoxin.
4) Note that the rate of the reaction depends upon the concentration of endotoxins, pH, temperature, and the presence of clotting enzymes and clottable proteins from the lysate.
5) Express the number of endotoxins in defined endotoxin units (EU).
6) Use the formula k/M to calculate the endotoxin limit for the specified test preparation, where M is the maximum dosage given to an adult per kg/hr.
7) Use a value of K of 5.0 EU/kg for parenteral preparations and 0.2 EU/kg for intrathecal preparations in the expression.
8) Calculate the endotoxin limit for the given test preparation using the calculated values of k and M.
9) Compare the calculated endotoxin limit with the observed endotoxin content to determine if it is within acceptable limits.
10) Repeat the procedure with different samples to ensure consistency of results.
(ii) Pyrogen Test for Rabbit Fever Reaction (Sham Test)
This test is performed by selecting the proper animals for the main test.Rabbit test
1) Obtain rabbits for the test.
2) Acclimate the rabbits to their environment for a minimum of 5 days before beginning the test.
3) Record the baseline temperature of each rabbit using a rectal thermometer.
4) Administer the test substance (which may contain pyrogens) to each rabbit either intravenously or intramuscularly.
5) Monitor the rabbits' temperature every 30 minutes for 3 hours after the administration of the test substance.
6) Record any temperature changes and note the time of the changes.
7) If the rabbits' temperature increases by at least 0.5°C within 3 hours after the administration of the test substance, the substance is considered pyrogenic.
8) Administer a pyrogen-free control substance to ensure the rabbits' temperature doesn't change due to other factors.
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Procedure
1) On the experiment day the food should be stopped.
2) Now, the initial temperature of the rabbit should be recorded. If any rabbit shows a temperature greater than 39°C then it should not be included in the experiment.
3) Now, the sample should be injected into the ear vein of the rabbit.
4) Finally, check the temperature after 30 minutes, 1 hour, 2 hours, and 3 hours respectively.
2) Acclimate the rabbits to their environment for a minimum of 5 days before beginning the test.
3) Record the baseline temperature of each rabbit using a rectal thermometer.
4) Administer the test substance (which may contain pyrogens) to each rabbit either intravenously or intramuscularly.
5) Monitor the rabbits' temperature every 30 minutes for 3 hours after the administration of the test substance.
6) Record any temperature changes and note the time of the changes.
7) If the rabbits' temperature increases by at least 0.5°C within 3 hours after the administration of the test substance, the substance is considered pyrogenic.
8) Administer a pyrogen-free control substance to ensure the rabbits' temperature doesn't change due to other factors.
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Procedure
1) On the experiment day the food should be stopped.
2) Now, the initial temperature of the rabbit should be recorded. If any rabbit shows a temperature greater than 39°C then it should not be included in the experiment.
3) Now, the sample should be injected into the ear vein of the rabbit.
4) Finally, check the temperature after 30 minutes, 1 hour, 2 hours, and 3 hours respectively.
Disadvantages
1) There are biological variations that may occur.
2) The process is very expensive.
3) The process is effortful as well as laborious.
4) The process is completely dose-dependent
5) This process is not for the antipyretic drug.
Result – If each rabbit shows an increase in temperature then the test is positive.
If in three rabbits only two show an increase in temperature then repeat the test by using the group of five rabbits, and the test will be considered positive if four out of five rabbits show the increased temperature.
Medium of soya bean casein digest
Medium of fluid thioglycolate
Now, wash the filters to remove any unwanted materials and aseptically cut the membranes into equal parts, and transfer one part into each type of culture medium that is used.
Finally, at the desired condition the media is incubated.
In this method the sample is incubated directly into the media means the sample is added into the media and incubated directly.
Result – If there is not any growth occurring in the media then the test is found to be positive.
To make sure injections are safe, the USP has rules called good manufacturing practice (GMP). These rules say that every injection container should be looked at carefully to see if there are any visible particles. If particles are seen, the container should be thrown away.
To follow these rules, people are inspecting every injection container one by one with good lighting and a black-and-white background to make sure they are safe to use.
The two test methods have been identified by USP.
The first test is called the light obscuration test. It uses a special machine to count and measure the size of tiny particles. The machine shines a bright light and when the particles pass through it, they cast a shadow that can be measured.
If the liquid being tested is not clear and colorless, it cannot pass the light obscuration test. In that case, another test called the microscopic count test needs to be done.
Ten sterile units are weighed to note their weight, then their content is removed.
Each of the ten sterile units is then weighed again, and the weight of the empty unit is subtracted from the weight with the content to get the net weight.
The content of active ingredients in each sterile unit is calculated using a method called the "assay" based on individual monographs.
The dose uniformity is checked by making sure that the amount of active ingredient in the 10 sterile units falls between 35-115% of the label claim, as determined by the content uniformity or weight variation methods.
If the relative standard deviation in the dose uniformity test is equal to or less than 60%, the potency value of each sterile unit is checked.
To check the potency, a test is carried out on 20 more sterile units.
If no more than one sterile unit falls outside the range of 85-115%, no unit falls outside the range of 75-125%, and the relative standard deviation is not more than 7.8%, then the sterile units meet the requirements.
1) There are biological variations that may occur.
2) The process is very expensive.
3) The process is effortful as well as laborious.
4) The process is completely dose-dependent
5) This process is not for the antipyretic drug.
Result – If each rabbit shows an increase in temperature then the test is positive.
If in three rabbits only two show an increase in temperature then repeat the test by using the group of five rabbits, and the test will be considered positive if four out of five rabbits show the increased temperature.
(c) Sterility Test
The sterility test is performed for detecting the presence of microorganisms in the solution and for this many containers are to be taken from the product batch.(i) Membrane Filtration Method
The media that is suitable for sterility tests are:Medium of soya bean casein digest
Medium of fluid thioglycolate
Now, wash the filters to remove any unwanted materials and aseptically cut the membranes into equal parts, and transfer one part into each type of culture medium that is used.
Finally, at the desired condition the media is incubated.
(ii) Direct Inoculation Method
When membrane filtration is not a desired option then this method is used.In this method the sample is incubated directly into the media means the sample is added into the media and incubated directly.
Result – If there is not any growth occurring in the media then the test is found to be positive.
(d) Particulate Evaluation
Small things like lint, rubber, and chemicals that can't dissolve in our body can cause blockages in important parts of our body.To make sure injections are safe, the USP has rules called good manufacturing practice (GMP). These rules say that every injection container should be looked at carefully to see if there are any visible particles. If particles are seen, the container should be thrown away.
To follow these rules, people are inspecting every injection container one by one with good lighting and a black-and-white background to make sure they are safe to use.
The two test methods have been identified by USP.
The first test is called the light obscuration test. It uses a special machine to count and measure the size of tiny particles. The machine shines a bright light and when the particles pass through it, they cast a shadow that can be measured.
If the liquid being tested is not clear and colorless, it cannot pass the light obscuration test. In that case, another test called the microscopic count test needs to be done.
(e) Weight Variation or Uniformity of Content
This test is used for the sterile solids that are used in the parenteral preparations.Ten sterile units are weighed to note their weight, then their content is removed.
Each of the ten sterile units is then weighed again, and the weight of the empty unit is subtracted from the weight with the content to get the net weight.
The content of active ingredients in each sterile unit is calculated using a method called the "assay" based on individual monographs.
The dose uniformity is checked by making sure that the amount of active ingredient in the 10 sterile units falls between 35-115% of the label claim, as determined by the content uniformity or weight variation methods.
If the relative standard deviation in the dose uniformity test is equal to or less than 60%, the potency value of each sterile unit is checked.
To check the potency, a test is carried out on 20 more sterile units.
If no more than one sterile unit falls outside the range of 85-115%, no unit falls outside the range of 75-125%, and the relative standard deviation is not more than 7.8%, then the sterile units meet the requirements.
I hope you have liked the article parenteral products and the evaluation of parenteral products. If you have any points please ask in the comment.
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