Two forms of alpha-1-antitrypsin (.alpha.1-AT), .alpha.1-AT(I) and .alpha.1-AT(II), were separately obtained from human sera which was pooled and stored for 3-6 mo. at -20.degree. C by a combination of ion-exchange chromatography, gel-filtration and affinity chromatography. The N-terminal amino acid sequences of .alpha.1-AT(I) and .alpha.1 AT(II) were tentatively determined as Glu-Asp-Pro-Gln-Gly-Asp-Ala-Ala-Gln-Glu-Thr-Asp-Thr-Ser-His-(His)- and Glu-Thr-Asp-Thr-Ser-His-His-Asp-Gln-Asp-Ser-Pro-Thr-Phe-Asn-Lys-Leu-Thr-Pro-Asn-Leu-Ala-Glu-Phe-, respectively. The amino acid sequence of .alpha.1-AT(II) corresponds to the sequence beginning at the 10th glutamic acid residue from the N-terminus of .alpha.1-AT(I). .alpha.1-AT(II) may arise by cleavage of the Gln-Glu bond in .alpha.1-AT(I) by an enzyme either originally present in the serum or produced by contaminating microorganisms. Based on a combination of the N-terminal sequences of .alpha.1-AT(I) and .alpha.1-AT(II), the sequence of the N-terminal 33 residues of .alpha.1-AT is tentatively proposed. .alpha.1-AT isolated from fresh human plasma had the same N-terminal amino acid sequence as .alpha.1-AT(I). The C-terminal amino acid sequences of the 3 .alpha.1-AT were identified as -Gln-Lys by the carboxypeptidase digestion method. Amino acid analysis of performic acid-oxidized .alpha.1-AT showed that this protein contained 1 mol of cysteine residue/mol of protein, which was linked to a free cysteine through a disulfide bond. Most of the .alpha.1-AT preparations inhibited bovine trypsin at a molar ratio of 1:1. Certain preparations bound more than 1 mol of trypsin/mol of inhibitor.