Phosphatidate, phosphatidic acid analysis, separation

The nature of the separation achieved with a lipid extract from rat kidney is shown in Figure 2.5 [166]. In spite of the abrupt changes in solvent composition at various points, little base-line disturbance is apparent, and each of the main simple lipid and phospholipid classes is clearly resolved in only 20 minutes. Only the highly acidic lipids, phosphatidic acid and to a lesser extent phosphatidylserine, do not give satisfactory peaks. There is no "solvent peak" at the start of the analysis, as is often seen with other detectors, and BHT added as an antioxidant evaporates with the solvent so does not interfere. After a further 10 minutes of elution to regenerate the column, the next sample can be analysed. 

Reversed-phase HPLC is capable of separating the individual phosphatides according to the acyl substitution. Details of the HPLC analysis are found in Chapter 7. Enzymatic techniques allow selective hydrolysis of either the phosphorus-containing component or the fatty acids in the beta position, so that the resulting fragments can be analyzed by GC, GC-MS, and other methods (35-37). 

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