Many exotoxins of gram positive bacteria, such as superantigens (staphylococcal enterotoxins, toxic shock syndrome toxin-1 [TSST-1], and streptococcal pyrogenic exotoxins) and anthrax toxin are bioterrorism agents that cause diseases by immunostimulation or cytotoxicity. Glycerol monolaurate (GML), a fatty acid monoester found naturally in humans, has been reported to prevent synthesis of gram positive bacterial exotoxins. This study explored the ability of GML to inhibit the effects of exotoxins on mammalian cells and prevent rabbit lethality from TSS. GML (≥10 ug/ml) inhibited superantigen (5 ug/ml) immunoproliferation, as determined by inhibition of 3H-thymidine incorporation into DNA of human peripheral blood mononuclear cells (1 × 106 cells/ml) as well as phospholipase Cγ1, suggesting inhibition of signal transduction. The compound (20 ug/ml) prevented superantigen (100 ug/ml) induced cytokine secretion by human vaginal epithelial cells (HVECs) as measured by ELISA. GML (250 ug) inhibited rabbit lethality due to TSST-1 administered vaginally. GML (10 ug/ml) inhibited HVEC and macrophage cytotoxicity by anthrax toxin, prevented erythrocyte lysis by purified hemolysins (staphylococcal α and β) and culture fluids containing streptococcal and Bacillus anthracis hemolysins, and was non-toxic to mammalian cells (up to 100 ug/ml) and rabbits (250 ug). GML stabilized mammalian cell membranes, as erythrocyte lysis was reduced in the presence of hypotonic aqueous solutions (0 to 0.05 M saline) or staphylococcal α and β-hemolysins when erythrocytes were pretreated with GML. GML may be useful in management of gram positive exotoxin illnesses; its action appears to be membrane stabilization with inhibition of signal transduction.