Pyrogens- Physical and Biological Properties

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We must understand the physical properties of pyrogens, in order to understand why they behave differently when placed in different environments. This will also help us in preparing depyrogenated vessels and depyrogenated water. Important properties of pyrogens are their heat stability, their size, their molecular weight and their electrostatic properties. But by far the most important property, which is of significance to the parenteral manufacturing industry is the heat stability property.

?Heat Stability: Scientists knew since long that pyrogens are not destroyed by moist heat. Boiling is also not effective. Endotoxins can be destroyed by dry heat at temperatures above 180*c. Dry heat is the most preferred method for depyrogenating glass vessels and other equipment.

?The pyrogens' environment determines their size: The size of pyrogens is dependent on their state of aggregation and this state is dependent on their environment. The endotoxin is arranged in a bilayer, in an aqueous environment, with the hydrophobic components clustered in the center and the hydrophilic components exposed to the surrounding solution. Magnesium and calcium ions stabilize the bacterial endotoxin. In their presence the endotoxin forms bilayers and possesses a size of 100 nm. But if the divalent cations are removed from the environment ,then the bilayered vesicles breakdown micellae, that are 20-70 nm long and about 3-7 nm thick. If detergents are present in the environment, the size of the micellae further reduces to 0.8-1.2 nm diameter and 10-60 nm length. The endotoxin's tendency to aggregate into larger entities in aqueous environment is due to the attraction of its hydrophobic groups for one another. Even when present as large aggregates, endotoxin can pass through 0.22 mm sterilizing filter. Reverse osmosis membranes that have a pore size of 1.0 nm are used to remove the endotoxin and any salts that are present.

?Molecular Weight: Just like the size, the molecular weight of the endotoxin also dependes on its environment.
-Aqueous environment with the presence of divalent cations--- Molecular weight is 1,000,000 Daltons.
-When the divalent cations are removed by chelation from above environment---- Molecular weight of micelle is 300,000 to 1,000,000.
-If a surface active agent is added to the solution containing endotoxins- molecular Weight is about 10,000-20,000.
-All these steps are reversible.
-The state of aggregation affects both solubility and biological activity.
-The Molecular weight of the endotoxin affects all its properties. As the molecular weight increases | ,toxicity in rats increases |
-Rate of clearance from blood increases| , interaction with complement and affinity for cells also increases | but its lethality in mice decreases | and its pyrogenic nature in rabbits also decreases|.

?Electrostatic Properties: At a pH of more than two, endotoxin aggregates are negatively charged and behave as anions and are attracted by divalent cations. This property also helps in the removal of endotoxins by adsorbing them onto cationically charged adsorbents.

? Endotoxins are potent.
? They produce many harmful effects.
? In extreme situations , they may alter organ function.
? They may produce hypotension.
? Endotoxins may produce disseminated intravascular coagulation.
? The above condition may lead to death.

Beneficial Effects:
> Endotoxins may stimulate the mammalian defense system.
> The above may enhance body's capacity to deal with infections and malignant tumors.

?Fever is the most dramatic biological effect of pyrogens.
?Fever is easy to measure.
?This is the most studied and understood phenomena.
?Exogenous pyrogens enter the blood stream.
?They are removed from circulation by phagocytosis.
?The phagocytic cells (peripheral monocytes) are stimulated to synthesize a family of cytokines or endogenous pyrogens.
?Endogenous pyrogens go to the hypothalamus, the thermoregulatory center of the body.
?Here the endogenous pyrogens induce cells to produce prostaglandins.
?The new thermostatic setting signals the peripheral blood vessels to constrict and conserve heat.
?This makes the body to chill.
?The discomfort makes other defense mechanisms to produce heat.
?Result is fever
?Intravenously at as low a dose as 1 ng/kg there may be a reaction.
?In large doses, endotoxins can be lethal.

Thus we must understand the physical and biological properties thoroughly to be capable of eliminating pyrogens from parenterals.

James Swarbrick, Encyclopedia of Pharmaceutical Technology, Volume 1,Third edition, , Informa Health care, Newyork: 2007: pages 3054- 3056

This book post is free from plagiarized material/b>

About the Author

Prof. J. Vijaya Ratna's picture

Dr. Vijaya Ratna Jayanthi serving Andhra University College of Pharmaceutical Sciences as Chairman, Pharmaceutical Technology Department.

Dr. J. Vijaya Ratna did her B.Pharm (1977), M.Pharm (1979), PGDAS (1981) and Ph.D (1998) at Andhra University Campus and won "M.L. Khorana Gold Medal" for standing University FIRST in graduation.


Santosh kumar. JH's picture

Thank you ma'am for an additional book page on Pyrogens for a greater understanding....
P.V.ABHIGNA's picture

Dear mam, It was really wonderful to know so many things about endotoxins...Thankyou very much for the info. "Disemminated intravascular coagulation can lead to death."Can u pls xplain me in brief abt this.I wouls love to kno.


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