Separation and characterization of adjuvant oligosaccharide oleate ester derived from product mixture of mannitol-oleic acid esterification

Nearly 30 years after intense investigations of mannide monooleates for use as vaccine adjuvants, a novel adjuvant-active saccharide oleate ester was isolated and identified from the product mixture synthesized from mannitol and oleic acid. The mixture, which contained many kinds of mannide mono- and dioleates and their derivatives, was fractionated by liquid chromatography (LC), and the fraction with the highest adjuvanticity was obtained. Gel permeation chromatography (GPC) showed that it consisted of one major compound with an average molecular weight (MW) 2850. Infrared (IR) absorption and proton nuclear magnetic resonance spectra suggested it had oligosaccharide moieties and oleate domains. These findings suggested that it was an oligosaccharide oleate ester of the average MW 2850. The molecular ratio of oleate chains per monosaccharide unit was approximately 0.8. The ester induced both IgG1 and IgG2a antibody responses in mice in a dispersed form without base oil. This ester thus appears to be one of the adjuvant-active compounds largely contributing to the excellent adjuvanticity of mannide oleate mixture broadly used as vaccine emulsifier. These results and previous findings suggest that the fundamental adjuvanticity of this ‘oligo’ saccharide acylate ester was in accord with the hydrophil-lipophil balance (HLB) theory, similarly to other saccharide acylate esters. It is now expected that this compound will be useful as novel vaccine adjuvant which may induce both Th1 and Th2 type immune responses with low or no toxicity, not only as an vaccine emulsifier but in an aqueous suspension form.