The water-soluble venom of Bothrops asper Garman (San Juan Evangelista, Veracruz, Mexico) showed 15 polypeptide bands on polyacrylamide-gel electrophoresis. This material exhibited phospholipase, hyaluronidase, N-benzoyl-L-arginine ethyl hydrolase, N-benzoyl-L-tyrosine ethyl hydrolase and phosphodiesterase activity, but no alkaline phosphatase or acid phosphatase activity. Fractionation on Sephadex G-75 afforded seven protein fractions, which were apparently less toxic than the whole venom (LD50 = 4.3,ug/g mouse wt.). Subsequent separation of the phospholipase-positive frac-tion (II) on DEAE-cellulose with potassium phosphate buffers (pH 7.55) gave several fractions, two being phospholipase-positive (11.6 and 11.8). These fractions were further purified on DEAE-cellulose columns with potassium phosphate buffers (pH 8.6). Frac-tion II.8.4 was rechromatographed in the same DEAE-cellulose column, giving a pure protein designated phospholipase 1. The fraction II.6.3 was further separated by gel disc electrophoresis yielding two more pure proteins designated phospholipase 2 and phos-pholipase 3. Analysis of phospholipids hydrolysed by these enzymes have shown that all three phospholipases belong to type A2. Amino acid analysis has shown that phospho-lipase A2 (type 1) has 97 residues with a calculated mol.wt. of 10978 + 1 1. Phospholip-ase A2 (type 2) has 96 residues with a mol.wt. of 10959 + 11. Phospholipase A2 (type 3) has 266 residues with 16 half-cystine residues and a calculated mol.wt of 29042+31. Automated Edman degradation showed the N-terminal sequence to be: Asx-Leu-Trp-Glx-Phe-Gly-Glx-Met-Met-Ser-Asx-Val-Met-Arg-Lys-Asx-Val-Val-Phe-Lys-Tyr-Leu-for phospholipase A2 (type 2).