Serine peptidase inhibitors (serpins) form a superfamily of proteins covering abroad spectrum of different biological functions. Here we describe the inhibitory characterization of leviserpin, the first serpin from the sugar cane weevil Sphenophorus levis. Leviserpin was able to inhibit bovine trypsin by the formation of the covalent complex serpin-peptidase, demonstrated by SDS-PAGE and mass spectroscopy analysis. We also have determined the cleavage site at the reactive center loop, by the analysis of the polypeptides released from de C-terminus of leviserpin. Moreover we investigated the mRNA expression of leviserpin in different stages of S. levis development. Thus the specificity of leviserpin, in addition with its mRNA coding being transcribed through all lifecycle of the insect, can suggest a possible role in defense mechanism by regulating the action of prophenoloxidase (proPO) activating enzyme.