Endotoxin is lipopolysaccharide (LPS) from the outer membrane of Gram-negative bacteria. It is a potent activator of Toll-like receptor 4, triggering inflammatory signalling at picogram-per-millilitre concentrations, and it is measured in endotoxin units (EU), where roughly 10 EU corresponds to 1 ng of reference LPS.
Why endotoxin matters in peptide research
It is a separate specification because ordinary purity testing cannot see it. A peptide can be 99% pure by HPLC and still carry enough endotoxin to dominate an immunological readout, since LPS is not a peptide-related impurity and does not appear on the chromatogram. In cell culture, macrophage and immune assays, contamination produces cytokine responses that are easily misattributed to the test peptide — a well-documented source of irreproducible results.
Two contamination routes dominate: recombinant expression in bacterial hosts, where LPS co-purifies from the host itself, and water or glassware in otherwise synthetic workflows. Endotoxin is heat-stable and survives autoclaving, so it must be excluded rather than destroyed. Testing is by LAL assay, reported on the certificate of analysis only when specifically requested. See reading a COA and third-party testing.