Phospholipid transfer protein (PLTP) from pig plasma was purified to homogeneity using ultracentrifugation and a combination of hydrophobic-, heparin-Sepharose-, and anti-pig-PLTP-affinity chromatography techniques. The molecular weight of PLTP is 78,000 as estimated by SDS-PAGE and by gel filtration. The effect of pig plasma PLTP on the particle size distribution of either human or pig high density lipoprotein (HDL) was studied by incubating HDL with PLTP. Incubation of human HDL3 or pig HDL with the highly purified preparations of PLTP induced a conversion of the homogeneous HDL into two main populations of particles. The conversion products were isolated by ultracentrifugation at density 1.21 g/ml. The size changes were evident as analyzed by native gradient gel electrophoresis and by high resolution gel filtration. The diameters of the large and small particles formed were 10.8 nm and 7.6-7.9 nm, respectively. In addition, pig HDL conversion products included a third population of particles with a diameter of 11.5 nm. The degree of conversion was dependent on time and PLTP activity. Neither cholesteryl ester transfer protein nor lecithin:cholesterol acyltransferase activity could be detected in the PLTP preparations. The present study demonstrates that purified pig PLTP can act as a conversion factor: it has the ability to convert HDL into populations of large and small particles. The release of apoA-I is an essential part of the conversion process.