Phosphatidylinositols acidic solvent extraction

The first hurtle is to reproducibly extract lipids from a matrix. The most common lipids extraction methods are those of Bligh and Dyer [42] and Eolch [43]. Recent analysis of these two methods has shown that the Eolch method tends to have a greater total recovery of lipid [44]. A variety of other solvent mixtures has been compared and may offer fewer hazards with similar recoveries [45]. These extraction methods are designed to recover the principal lipid classes, but may not be as useful for recovery of lipids that have unique charge characteristics. For example, fatty acids, phosphatidic acids, and lyso-phosphatidic acids usually require acidic solvents to facilitate recovery from an aqueous solution while neutral lipids may not be sufficiently soluble in an organic solvent [46]. Other complexities include solvent manipulations required to extract more polar lipids like the phosphatidylinositol phosphates. 

Perhaps the best approach to extraction of the phosphatidylinositols and their various phosphorylated derivatives from a cellular preparation is through use of the Bligh-Dyer technique or some modification of it. However, it is of paramount importance that the chloroform-methanol-water (1 2 0.8, v/v) mixture, as an example, contain an acid, usually 1 N HC1 as the water component. Otherwise, there will be a decreased recovery of the inositol phospholipids in the final chloroform extract. This is directly attributable to the fact that these inositol-containing phospholipids, as already mentioned above, are found naturally as the Ca2+, Mg2+, K1, and/or Na+ salts. If the solvent is not acidic, these salts essentially will remain in a water-rich frac-... 

Extraction parameters such as solvent type, mixture ratios, metal ion concentration, pH of the aqueous phase, extraction time, and temperature influence the recovery of extracted lipids and must be validated to ensure reliable results. For example, the recovery of the acidic lipids PA and phosphatidylglycerol (PG) can be less than 30% in classic Folch and Bligh Dyer extraction, where these lipids can become bound to proteins tightly (17). Lipids bound to proteins covalently are only released under appropriate conditions, which depend on the type of lipid-protein linkage. For example, ceramides bound to protein of the comi-fled envelop in the human skin (18) can be extracted after mild alkaline hydrolysis of the ester linkage between hpid and protein. Special conditions are required for extraction of more polar lipids such as gangliosides, lysophospholipids and lysosphin-golipids, or phosphatidylinositol-phosphates. 

Phospholipids The phospholipid content of crude sunflower oil ranges between 0.5% and 1.2%. Oils extracted by solvent generally have a higher content of phosphlipids than those obtained by pressing. Major phospholipids are phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and phosphatidic acid. Most are hydratable and may be removed from the cmde oil through a water-degumming process (See Section 5.3.1.)... 

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