High density lipoproteins (HDL) in the plasma from cholestatic rats were studied by gel filtration, polyacrylamide gel electrophoresis and electron microscopy. HDL from cholestatic rats separated into two subfractions, abnormally enlarged particles and smaller ones, on gel filtration through Bio-Gel A 1.5 m. Polyacrylamide gel electrophoresis in 8 M urea revealed that the relative proportion of C-III-3 peptide in C-apoproteins was higher in the two HDL subfractions from cholestatic rats than in normal HDL. Polyacrylamide gel electrophoresis in sodium dodecyl sulfate showed that the abnormally enlarged HDL contained the 'arginine-rich' apoprotein, whereas a band corresponding to 'arginine-rich' apoprotein was only faintly visible in the smaller HDL subfraction. Normal HDL contained much smaller amount 'arginine-rich' apoprotein than the enlarged HDL. In electron microscopy, both of the two subfractions of cholestatic HDL appeared spherical. The abnormally enlarged HDL were consisted of particles with a diameter of 17 +/- 4.5 nm (mean +/- S.D.). The other subfraction contained smaller particles with a diameter (about 10 nm) similar to normal HDL.