Analysis of phospholipids

Abstract 1. 1. The well-known method of successive chemical hydrolysis of phospholipid mixtures according to DAWSON 1 has been modified in several respects and can be applied to a whole phospholipid mixture as well as to fractions obtained by silica gel column prefractionation. 2. 2. In contrast to known methods, the combination of silica gel column fractionation and step-wise hydrolysis presented here makes it possible to discriminate between diacyl- and monoacylglycerophosphatides. 3. 3. The improvements in the technique of step-wise hydrolysis proposed here are the following: by modification of the mild alkaline hydrolysis, a quantitative deacylation with a minimum of additional breakdown of the polar part of the phosphatides is achieved; by using a Folch partition after alkaline hydrolysis no interference by emulsions or interfacial fluff is encountered, even though the total sphingolipid content of the sample may be as high as 60–70%; a better paperchromatographic separation of the glycerophosphate esters has been made possible by the use of relatively small quantities of alkali; an increase in accuracy, again compared with the known procedure has been achieved by determining the phosphorus content of the various fractions, and of the upper and lower layers after partition separately, and using the paper-chromatographic separation of the glycerophosphate esters only as a means to determine the ratio of the components; finally, by modification of the acid hydrolysis, losses of phosphate esters due to the washings necessary when using trichloroacetic acid-mercuric chloride are avoided.