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The fractionation of lipoproteins of normal and hyperlipidemic subjects on a column of 2% agarose was compared with ultracentrifugation and paper electrophoresis procedures. The following results were obtained. (a) Plasma lipoproteins were eluted successively from the column in the four overlapping peaks of chylomicrons, very low density lipoproteins, low density lipoproteins, and high density lipoproteins. (b) Very low density lipoproteins and high density lipoproteins (d > 1.063, containing nonlipoprotein proteins) showed continuous progressive changes in lipid composition as these fractions emerged, while low density lipoproteins showed a relatively constant lipid composition. (c) A discontinuous transition of lipid composition was observed when consecutive ultracentrifugal fractions were placed on the column. (d) The "trail" of pre-beta lipoprotein seen on paper electrophoresis was shown to consist of particles whose molecular sizes range between chylomicrons and pre-beta lipoproteins. A reverse relationship was observed between electrophoretic mobilities of "trail" components and their particle size. (e) Gel with an agarose content of 2% seemed to fractionate chylomicrons and very low density lipoproteins more effectively than other lipoprotein classes.

Procedures are described for the isolation of lipoproteins from human serum by precipitation with polyanions and divalent cations. A mixture of low and very low density lipoproteins can be prepared without ultracentrifugation by precipitation with heparin and either MnCl(2) alone or MgCl(2) plus sucrose. In both cases the precipitation is reversible, selective, and complete. The highly concentrated isolated lipoproteins are free of other plasma proteins as judged by immunological and electrophoretic methods. The low density and very low density lipoproteins can then be separated from each other by ultracentrifugation. The advantage of the method is that large amounts of lipoproteins can be prepared with only a single preparative ultracentrifugation. Polyanions other than heparin may also be used; when the precipitation of the low and very low density lipoproteins is achieved with dextran sulfate and MnCl(2), or sodium phosphotungstate and MgCl(2), the high density lipoproteins can subsequently be precipitated by increasing the concentrations of the reagents. These lipoproteins, containing small amounts of protein contaminants, are further purified by ultracentrifugation at d 1.22. With a single preparative ultracentrifugation, immunologically pure high density lipoproteins can be isolated from large volumes of serum.

It has been proposed that particles within tubules and vesicles of the Golgi apparatus of liver cells are precursors of very low density lipoproteins in blood plasma. To characterize these particles we isolated a cell fraction rich in Golgi apparatus and associated particles from rat liver in quantities sufficient for analysis. Particles freed from the membranes of the Golgi apparatus and floated at d = 1.006 were studied by chemical analysis, immunodiffusion, and paper electrophoresis. The lipid composition of the Golgi particles was similar to that of very low density lipoproteins from the same rats. The protein content was about 10% of dry weight for both the Golgi particles and plasma very low density lipoproteins. The Golgi particles formed lines of identity with plasma very low density lipoproteins during immunodiffusion against antiserum to plasma very low density lipoproteins. On paper electrophoresis, however, many Golgi particles remained near the origin, with only a few migrating to the pre-beta position. It was concluded that the lipoproteins in the Golgi apparatus are the precursors of plasma very low density lipoproteins.

Dogs were given infusions of norepinephrine and subsequent additional infusions of propranolol and nicotinic acid over a 4-hr period. Under different physiological conditions, alpha-lipoproteins of three different electrophoretic mobilities were identified by means of paper electrophoresis; they were designated alpha-lipoproteins X, Y, and Z. During norepinephrine infusion, alpha-lipoprotein Y fell from 45% (of all lipoproteins) to 14%. There was a reciprocal rise in alpha-lipoprotein Z. On the other hand, alpha-lipoprotein X was not significantly changed. There was evidence that alpha-lipoprotein Y was progressively transformed into alpha-lipoprotein Z by increasing plasma FFA concentrations. The percentages of both alpha-lipoproteins Y and Z returned to original values after the dogs were given either nicotinic acid or propranolol. The alterations in the alpha-lipoprotein peaks Y and Z were rapid, being noted within 5 min of change in plasma FFA concentration. However, there appeared to be a threshold of plasma FFA concentration of 1200 micro Eq/liter, below which no changes in alpha-lipoproteins were noted. It was concluded that alpha-lipoprotein Y is rapidly, progressively, but reversibly transformed into alpha-lipoprotein Z by binding to plasma FFA above a threshold level of 1200 micro Eq/liter. However, alpha-lipoprotein X does not appear to be involved in the binding of plasma FFA.