Separation of Lipids in Gravimetric Acetone Method for Plasma Total Protein

In the method of Bierry and Vivarro for the estimation of plasma total protein as modified by Guillaumin, Wahl and Laurencin, proteins are precipitated by adding 3 ml of plasma to 10 ml of acetone, centrifuging and washing the precipitate twice with acetone. Commenting on this method, Peters and van Slyke 1 state, “This is the most rigidly exact of plasma protein methods, because the acetone treatment removes lipoids. … In the other methods … the lipoids are precipitated with the proteins, and their nitrogen content probably causes a slight plus error in the protein figure.”

Boyd 2 has shown that when cold, fat solvents are used to extract plasma lipids, a minimal initial volume of solvent is required per volume of plasma if complete extraction is to be obtained. With Bloor's extracting fluid, 3 :1 alcoholether which is generally conceded to be the best lipid extractor, a minimum of 20 volumes of solvent per one volume of plasma were found necessary. On checking the protein method of Bierry and Vivarro as outlined by Peters and van Slyke, 1 it was found, as was to be expected, that lipids are not completely removed by the acetone. A few typical examples are given in Table I to show that lipid values in alcohol-ether extracts by Boyd's method 2 are appreciably and consistently higher than lipid values of the combined acetone mother liquors in the protein method.

The acetone protein method does not, therefore, remove all the lipids and the presumption is that the portion left is weighed later with the proteins. In plasma with normal amounts of protein and lipids, this would result in a plus error of 2 to 3% for the protein fraction.