Removal of Fatty Acids from Serum Albumin by Charcoal Treatment

The fatty acid contents of 26 different serum albumin preparations representing different species and obtained from various commercial sources have been determined. Some samples had surprisingly little fatty acid contamination, but it was found that other samples contained between 2 and 3 moles of acid per mole of protein, in confirmation of earlier reports. Treatment of these samples with charcoal at low pH resulted in the virtually complete removal of fatty acids. The conditions for such treatment were investigated as a function of the type of fatty acid, pH, and the amount of charcoal required. Charcoal treatment removed at least 99% of albumin-bound radioactive fatty acids under optimal conditions, whereas the amount of protein adsorbed onto charcoal was 4.9%. The protein that remained in solution following separation of the charcoal appeared to be native, as judged by a number of criteria, including analytical ultracentrifugation, optical rotatory dispersion, the ability to bind fatty acids, and the ability to bind the fluorescent dye, 8-anilinonaphthalene-1-sulfonic acid, as well as several other fluorescence parameters. Fluorescence spectra of human serum albumin samples indicated that impurities are sometimes present which can be removed by charcoal at neutral pH. A possible explanation for the gentleness of charcoal treatment is offered. Acid-charcoal treatment is a much more rapid method of removing lipid impurities than other methods previously described.