Plant cysteine proteinase inhibitors have received special attention due its specific inhibition to pest insect enzyme. These inhibitors have three regions of interaction with papain, a Gly, a QVVAG region and a trip residue. However, Prosopis juliflora papain inhibitor showed structural identity with the Kunitz type inhibitors, where two interaction sites, one to papain (Arg64, Glu89, Glu109 and Trp60) and other to trypsin (Arg 64), were found. The objective this work was to determine if the interaction site to papain/trypsin found in P. juliflora is present in trypsin-Kunitz type from Mimosoideae subfamily such as Acacia confusa, Adenanthera pavonina and Leucaena leucocephala seeds and to construct a filogenitc tree using these inhibitors and 7 Kunitz inhibitors from Papilionoideae and Caesalpinioideae subfamily. Primary amino acid sequences were alignment by CLUSTALW program, which was carried out trough of the PAUP program. The results showed that A. confusa trypsin inhibitor had Trp60 and Glu109 that also were present in papain active site in P. juliflora. The identity observed between these inhibitors was of 68.36% or 121 residues suggesting that A. confusa inhibitor could be bifunctional too. A. pavonina inhibitor presented variation where Trp60 was substituted by Ser60, Glu89 was similar to Asp89, and Glu109 remained identical. The identity observed between these inhibitors was of 66.48% or 117 residues. Date reported confirmed that A. pavonina inhibitor was bifunctional. L. leucocephala inhibitor showed variation where Trp60 was substituted by Ala60, Glu89 showed weak similarity to His88, and identical Glu109 was found. The identity observed between these inhibitors was of 43.50% or 77 residues suggesting that L. leucocephala inhibitor could be not bifunctional. Analysis of the primary sequences among P. juliflora, A. confusa, A. pavonina and L. leucocephala showed identity of 33.89% or 61 residues. Homology of data were used to construct the phylogenetic relationship between 11 Kunitz inhibitors obtained from gene bank and the bifunctional character to P. juliflora, A. confusa, A. pavonina and L. leucocephala. Results showed that P. juliflora, A. confusa, A. pavonina inhibitors were more closes and L. leucocephala inhibitor more distant, inside of the Mimosoideae subfamily, this fact could confirm the variation in papain site observed in silico analysis and the possible bifunctionality these proteins.