Lignification in Zinnia elegans L. stems is characterized by a burst in the production of H(2)O(2), the apparent fate of which is to be used by xylem peroxidases for the polymerization of p-hydroxycinnamyl alcohols into lignins. A search for the sites of H(2)O(2) production in the differentiating xylem of Z. elegans stems by the simultaneous use of optical (bright field, polarized light and epi-polarization) and electron-microscope tools revealed that H(2)O(2) is produced on the outer-face of the plasma membrane of both differentiating (living) thin-walled xylem cells and particular (non-lignifying) xylem parenchyma cells. From the production sites it diffuses to the differentiating (secondary cell wall-forming) and differentiated lignifying xylem vessels. H(2)O(2) diffusion occurs mainly through the continuous cell wall space. Both the experimental data and the theoretical calculations suggest that H(2)O(2 )diffusion from the sites of production might not limit the rate of xylem cell wall lignification. It can be concluded that H(2)O(2) is produced at the plasma membrane in differentiating (living) thin-walled xylem cells and xylem parenchyma cells associated to xylem vessels, and that it diffuses to adjacent secondary lignifying xylem vessels. The results strongly indicate that non-lignifying xylem parenchyma cells are the source of the H(2)O(2) necessary for the polymerization of cinnamyl alcohols in the secondary cell wall of lignifying xylem vessels.