Anion-exchange high-performance liquid chromatography of human serum apolipoproteins.

The rapid separation of seven urea-soluble apolipoprotein species from delipidated human serum very low density lipoproteins (VLDL) and high density lipoproteins (HDL) has been achieved by high-performance liquid chromatography on an anion-exchange column of Syn-Chropak AX 300. Effluent chromatographic peaks were detected by absorbance at 280 nm in a flow-through cell. Peaks corresponding to apolipoproteins AI1, AI2, AII, CI, CII, CIII1, and CIII2 were identified by amino acid analysis, gel electrophoresis, and isoelectric focusing. Maximum efficient loading of semipreparative columns (250 X 9.0 mm) was established to be ca. 20 mg HDL apolipoprotein. Minimum detectable protein was shown to be ca. 1 microgram on an analytical-scale column (300 X 4.5 mm). Chromatographic resolution is comparable to that of conventional DEAE-cellulose column chromatography. The ratio of apoAI1 to apoAI2 was considerably greater in high-performance liquid chromatography, suggesting that the variants seen in conventional chromatography and isoelectric focusing are in part artifactual.