An endotoxin unit (EU) is a unit of the biological activity of bacterial endotoxin, defined against a reference standard, not a unit of mass. Endotoxin is lipopolysaccharide (LPS) from the outer membrane of Gram-negative bacteria. Different LPS preparations have very different potencies per nanogram, so the pharmacopeias define the unit by activity relative to a reference standard endotoxin rather than by weight. One EU is equivalent to one International Unit (IU), and as a rough mass equivalent one EU corresponds to about 0.1 ng of the reference E. coli endotoxin.
What 0.5 EU/mL means on a certificate of analysis
The USP limit for bacteriostatic water is not more than 0.5 EU per mL; for the WFI-grade water it is made from, the limit is 0.25 EU/mL. A certificate of analysis usually reports the result as “less than” a value (for example <0.25 EU/mL), which means endotoxin was not detected at the sensitivity of the test used, rather than a measured amount. At the limit, a 30 mL vial would contain at most 15 EU in total. See how to read a COA.
How it is measured
The bacterial endotoxins test (USP <85>) uses Limulus amebocyte lysate (LAL), an extract of horseshoe-crab blood cells that clots in the presence of endotoxin. The gel-clot method gives a pass or fail at the labeled sensitivity of the lysate (commonly 0.03 to 0.25 EU/mL); the turbidimetric and chromogenic methods give a numerical result. Recombinant reagents that replace the animal-derived lysate are now recognized as well (USP <86>). See endotoxin testing: the LAL test and the 0.5 EU/mL limit.
Why sterilization does not remove endotoxin
Endotoxin is heat-stable. Autoclaving kills bacteria but leaves their LPS behind, and 0.2 µm filtration removes cells but not the much smaller LPS molecules. Endotoxin in a water product is therefore controlled by preventing Gram-negative growth in the water system in the first place (distillation or reverse osmosis followed by hot circulation) and confirmed by testing the finished product. Glassware is depyrogenated by dry heat at about 250 °C, which is not something that can be done to a solution. This is why “sterile” and “nonpyrogenic” are two separate claims on the label: see sterile vs. nonpyrogenic.
Related
USP water grades explained · Is bacteriostatic water sterile? · Glossary
All BAC Water Supply products are supplied for laboratory research use only and are not for human or veterinary use.