Low Endotoxin Recovery

Abstract

Low endotoxin recovery (LER) in biopharmaceutical drug products involves the potential risk of under-determination of endotoxin (LPS) contaminations. Here, the proposed mechanism and the driving forces of the LER-effect are described. To understand LER, it should be clearly distinguished from test-interference. In the case of LER it is hypothesized that the supramolecular state of endotoxin is altered in a two-step process. The detectable endotoxin aggregation state is first destabilized by disturbance of the ionic salt bridges between the LPS molecules. Secondly, the supramolecular structure is changed into a state that does not allow activation of Factor C. It is thought that the endotoxin forms mixed aggregates with matrix components of the sample and the active unit of the LPS. Moreover, such a transition from detectable to non-detectable endotoxin is time dependent. The reaction kinetics can be influenced by the experimental setup as well as by the composition of the tested sample. As the structure of the endotoxin varies within different species, the LER phenomenon is also dependent on the source and origin of endotoxin. Alteration of the aggregation states come along with a shift of the equilibrium state. To this end, a new equilibrium state of endotoxin is formed under LER conditions. Consequently, by target-oriented changing of sample conditions the reaction can be reversed and a detectable state can be restored.