While investigating the immunological activities of Echinacea preparations, the question
of microbiological contamination arises, because microorganisms or parts and products
of them may influence the test results. Especially LPS from gram-negative bacteria
and lipopeptides from gram-negative and gram-positive bacteria may stimulate the immune-system
already at very low concentrations.
Arabinogalactan-proteins (AGPs) from Echinacea purpurea [1] were tested in the LAL-test, using the gel-clotting-method, before and after
LPS-removal. Since this test responses not only to LPS, but also to β-1,3-glucans [2], it seemed possible that there could also be a response to other
polysaccharide containing polymers such as AGPs.
After removal of LPS by two different methods (affinity chromatography and treatment
with sodium hydroxide), absence of LPS and bacterial lipopeptides was proven by testing
the LPS-free AGP preparation for interaction with toll-like receptors (TLR) 2- or
TLR4-transfected HEK293 cells. There was no reactivity of AGP preparations with these
receptors, indicating that LPS and lipopeptides have been successfully removed.
After blocking of possible glucan-activity in the LAL-test by a special buffer, the
LPS-free AGP preparations still led to coagulation. The remaining activity in the
test therefore has to be considered as the activity of AGPs. Thus, AGPs are able to
activate the LAL-test in a µg-concentration range. This is lower, but considerable
activity compared to the activity of glucans (ng-concentration range) or LPS (pg-concentration
range).
Acknowledgement: The authors thank the Madaus AG, Köln, for financial support of this work.
References: 1. Classen, B. et al. (2000), Carbohydr. Res. 327: 497–504. 2. Blaschek, W. et al. (1992), Pharm. Pharmacol. Lett. 1: 118–122.
