New Colorimetric Method for Determination of Bile Acids in Blood.

Abstract

An exhaustive comparative study of the hitherto published methods for the determination of bile acids and salts in the blood demonstrated that they entail a very large loss of these substances by adsorption in the precipitation of the proteins, and in the removal of the bile pigments by charcoal. This fact could have been anticipated, if it is recalled that the bile acids are the most surface-active substances that are present in the blood. We found that by precipitating the proteins with an alcoholic solution of the alkaline ZnSO₄-NaOH reagent used in the Hagedorn-Jensen blood sugar method, and by avoiding the use of charcoal, a quantitative recovery (90 to 95 per cent) of bile salts added to blood plasma could be effected. The new color reaction is a modification of the well known Liebermann-Burchard reaction for cholesterol.

The method is as follows: To a mixture consisting of 75 cc. of a freshly prepared 0.45 percent ZnSO₄ solution, 15 cc. of 0.1 N NaOH solution and 50 cc. of redistilled alcohol, 5 cc. of plasma are added, drop by drop with shaking. The mixture is allowed to stand over night and filtered. The filtrate is evaporated to dryness on the water bath. The protein precipitate is washed on the filter with 50 cc. of boiling water. These hot water washings are used to dissolve out the bile salts from the residue of the alcoholic filtrate, leaving undissolved the bile pigments and some of the lipoids. The filtered aqueous solution is evaporated to dryness, and the residue is washed several times with ether to remove the remaining traces of bile pigments and cholesterol.