The complete primary structure of five chymotrypsin/elastase isoinhibitors isolated from Ascaris lumbricoides was determined by conventional methods. These structures represent the first sequence set for the Ascaris inhibitor family. All five isoinhibitors are single-chain polypeptides crosslinked by five disulfide bridges. Isoinhibitor 1 consists of 63 amino acid residues and has glycine at the N-terminal and histidine at the C-terminal. Isoinhibitors 2-5 all have arginine at the N-terminal, differ at positions 25 and 40, and have different C-terminal regions. Isoinhibitors 2 and 4 have asparagine at positions 25 and serine at position 40, whereas isoinhibitors 3 and 5 have lysine and threonine at these positions, respectively. The different C-terminal regions of isoinhibitors 2-5 account for their varying lengths. Isoinhibitor 1 has no sequence heterogeneity. Frequent repetitions of various dipeptides and one tripeptide are evident along the peptide chain of isoinhibitors 2-5. None of the isoinhibitors contains the aromatic amino acids phenylalanine or tyrosine. Comparison of the amino acid sequence of isoinhibitor 1 with the sequence of isoinhibitors 2-5 shows that they differ at a minimum of 16 positions. The primary structures of isoinhibitors 1-5 from Ascaris do not demonstrate a great degree of homology when compared with the sequence of presently known proteinase inhibitors. However, these isoinhibitors share with a very large number of inhibitor families the presence of half-cystine in the P3 position.