Purified preparations of human plasminogen often contain up to ten different forms of plasminogen, as demonstrated by zymographic analysis after starch-gel electrophoresis. Of these forms, only six have electrophoretic properties identical to those found in human plasma. The other forms, which move closer to the cathode, are probably altered by proteolytic degradation. By chromatography on DEAE-Sephadex using a pH and ionic gradient, a group separation was obtained of these plasminogen forms. One fraction (DE-A) contained four to six forms with electrophoretic properties consistent with the main plasminogen components of normal plasma. The other fraction (DE-B) contained the altered forms. On isoelectric focusing the isoelectric points of the components of Fraction DE-A range between 6.0 and 6.6, and those of Fraction DE-B between 7.3 and 8.8. The molecular weights of Fraction DE-A and Fraction DE-B were 90 000 and 105 000, respectively, as determined by gel filtration. There is evidence that the apparently higher molecular weight of Fraction DE-B is due to a partial degradation of the plasminogen molecules followed by conformational changes. As shown by gel-filtration experiments, the presence of ε-aminocaproic acid in the solvent brings about an increase in molecular volume of plasminogen. This is much more pronounced for Fraction DE-A than for Fraction DE-B. The amino acid composition and the N-terminal dipeptide have been determined on the plasminogen forms of Fraction DE-A isolated by isoelectric focusing. No significant difference among the different forms were found. The amino acid composition is consistent with the rather low values of the isoelectric points.