Determination of stereochemical configuration of the glycerol moieties in glycoglycerolipids by chiral phase high-performance liquid chromatography

This study reports a simple and sensitive method for determining the absolu te configuration of the glycerol moieties in glycoglycerolipids. The method is based on chiral phase high-performance liquid chromatography (HPLC) sep arations of enantiomeric di- and monoacylglycerols released from glycosyldi - and monoacylglycerols, respectively, by periodate oxidation followed by h ydrazinolysis. The released di- and monoacylglycerols were chromatographed as their 3,5-dinitrophenylurethane (3,5-DNPU) and bis(3,5-DNPU) derivatives , respectively. The derivatives were separated on two chiral phases of oppo site configuration, (R)- and (S)-1-(1-naphthyl)ethylamine polymers for diac ylglycerols and N-(R)-1-(1-naphthyl)ethylaminocarbonyl-(S)-valine and N-(S) -1-(1-naphthyl)etliylamino-carbonyl-(R)-valine for monoacylglycerols. Clear enantiomer separations, which permit the assignment of the glycerol config uration, were-achieved for sn-1,2 (2,3)-diacyl- and sn-1 (3)-monoacylglycer ols generated from linseed oil triacylglycerols by partial Grignard degrada tion on all the chiral stationary phases employed. Using the method, we hav e determined the glycerol configuration in the glycosyl-diacylglycerols (mo nogalactosyl-, digalactosyl-, and sulfquinovo-syldiacylglycerols) and glyco sylmonoacylglycerols (monogalactosyl-, digalactosyl-, and sulfoquinovosylni onoacylglycerols) isolated from spinach leaves and the coralline red alga C orallina pilulifera. The results clearly showed that the glycerol moieties in all the glycoglycerolipids examined have S-configuration (sn-1,2-diacyl- and sn-1 -monoacylglycerols). The new method demonstrates that chiral phas e HPLC provides unambiguous information on the configuration of the glycero l backbone in natural glycosyldi- and monoacylglycerols, and that the two-s tep liberation of the free acylglycerols does not compromise glycerol chira lity.