Proteinase inhibitors (PI) are expressed in several members of the Solanaceae. According to species, PI can be both developmentally and environmentally regulated. We investigated the presence and developmental regulation of PI in the flowers of Nicotiana plumbaginifolia. By trypsin affinity chromatography a small protein, Np-PI-II, was isolated. This protein had an N-terminus similar to a part of a Nicotiana alata PI-II, and an inhibitory activity towards trypsin. Inhibitory activity is expressed at very early stapes of floral development. No inhibitor is present in the vegetative meristems. The inhibitory activity remains high throughout flower maturation and rapidly declines in the ovary after pollination. In the stigma of mature Nicotiana tabacum flowers trypsin inhibitory activity is detected extracellularly. Proteinase inhibitor activity is not a regular feature of the stigmas of several solanaceous genera.