Molecular requirements of endotoxin (ET) actions: changes in the immune adjuvant, TNF liberating and toxic properties of endotoxin during alkaline hydrolysis.
The effect of alkaline pH and alkaline hydrolysis on the physico-chemical and biological properties of endotoxin (ET) isolated from Serratia marcescens ATCC 13477 by the Biovin procedure was studied. Major emphasis was put on the ion exchange column chromatography and immune adjuvant activity (ADA) of the alkali treated samples. To measure changes in some endotoxicity parameters, Limulus lysate clotting (LAL), chick embryo lethality, Shwartzman skin reactivity and in vitro TNF release were measured. The toxic properties of ET, with the unique exception of the Shwartzman skin reactivity, rapidly diminished during alkaline treatment. As immunogen CBre3, a recombinant HIV glycoprotein which spans the C terminus of gp 120 and the N terminus of gp 41, was used in CD-1 mice, alkali treated and immediately neutralized ET samples (zero time) were inactive as adjuvants, in some cases immunosuppression could be clearly seen. But if the alkaline hydrolysis was continued for 6 h, the ADA became higher than it had been for the starting ET sample. Further alkaline hydrolysis eliminated the ADA of the samples. Both NaOH and propylamine acted similarly on the ET preparation. Reaction kinetic studies of the NaOH detoxification indicated the cleavage of ester bound acyl groups with low binding energy. Chemical analyses of the samples revealed that changes occurred in the fatty acid composition, characterized by a loss of approximately half of the 3-OH myristic acid content.
Somlyo
B
B
Center for Oral Health Research, University of Pennsylvania 19104.
Csanky
E
E
Shi
X M
XM
Zhang
Y L
YL
Kovats
E
E
Bona-Liptak
E
E
Nowotny
A M
AM
Tripodi
D
D
Nowotny
A
A
D000276:Adjuvants, Immunologic / Q000494:pharmacology*; D000818:Animals; D002642:Chick Embryo; D004731:Endotoxins / Q000032:analysis / Q000494:pharmacology* / Q000633:toxicity; D005227:Fatty Acids / Q000032:analysis; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D011817:Rabbits; D012790:Shwartzman Phenomenon; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
D000276:Adjuvants, Immunologic; D004731:Endotoxins; D005227:Fatty Acids; D014409:Tumor Necrosis Factor-alpha