endotoxin

An endotoxin is the most important pyrogen in injectable manufacturing: it is a piece of the outer wall of Gram-negative bacteria. Unlike a secreted toxin, it is built into the bacterium's own membrane and is shed when the cell divides or dies, which means killing the bacteria does not remove it — the dead cell still leaves its endotoxin behind.

Chemically, endotoxin is a lipopolysaccharide (LPS): a fatty lipid anchor (lipid A) joined to a long chain of sugars. The lipid A portion is the dangerous part. Injected into the blood, even tiny amounts are recognized by the immune system as a sign of bacterial invasion, triggering the same cascade that produces fever, and at higher doses inflammation, low blood pressure and septic shock.

Two properties make endotoxin a formulation nightmare. It is extraordinarily heat-stable, surviving an autoclave cycle that easily kills the living bacteria, so it must be destroyed by much harsher dry heat or excluded from the start. And it is potent at vanishingly small amounts, so parenterals carry strict endotoxin limits — typically expressed in endotoxin units per millilitre or per dose, scaled to how much a patient receives per kilogram of body weight.

Detection is done in vitro. The Limulus amebocyte lysate (LAL) test, based on a clotting reaction from horseshoe-crab blood cells, is the classical method; recombinant reagents are increasingly used as an animal-free alternative. The test gives a sensitive, quantitative readout of how much endotoxin a product contains.