Evidence for the existence of non-esterified cholesterol carried by albumin in rat serum.

Isolation of non-esterified [14C]cholesterol bound to albumin from rat serum, 8 days after i.p. injection of [14C]cholesterol, was achieved by affinity chromatography, using Cibacron blue F3GA bound to Sepharose 4B and by Sephadex G-150 column chromatography. Both methods permit isolation of large quantities of cholesterol-loaded albumin, free of globulins and lipoproteins. The isolated albumin-cholesterol fraction was estimated to be 4.6 mg/100 ml serum, which represents approx. the 24% of the non-esterified cholesterol present in the rat serum. Albumin-cholesterol, cholesterol glucoside, cholesterol hemisuccinate and hydroxylated derivatives of cholesterol produced a biphasic curve of changes in synaptosomal plasma membranes (SPM)-bound (Na+ + K+)-stimulated ATPase activity. Low concentrations of the ligand progressively increased the enzyme activity, while increasing the ligand concentration above that which maximally stimulated the enzyme activity, produced a progressive inhibition. Lipoproteins did not have any effect on the enzyme activity. The fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene-labeled SPM, increased in albumin-cholesterol derivatives-treated SPM, which is consistent with a general decrease in membrane bilayer fluidity. The results provide evidence that the 'albumin-cholesterol' fraction of the serum may directly affect the cell membrane-bound enzyme activity.