THE INFLUENCE OF APOPLASTIC ASCORBATE ON THE ACTIVITIES OF CELL WALL-ASSOCIATED PEROXIDASE AND NADH OXIDASE IN NEEDLES OF NORWAY SPRUCE (PICEA-ABIES L)

Cell wall-associated peroxidases (EC 1.11.1.7) were extracted from the current year's needles of Norway spruce trees (Picea abies L.) in two fractions, namely soluble apoplastic peroxidases and covalently wall- bound peroxidases. Peroxidase activities were determined with two subs trates: coniferyl alcohol, which is important for lignification, and N ADH, which is necessary for the production of H2O2. Coniferyl alcohol peroxidase activity was detected in both the soluble apoplastic fracti on and the wall-bound fraction, whereas NADH oxidase activity was foun d only in the soluble apoplastic fraction. Net oxidation of coniferyl alcohol and NADH was inhibited by ascorbate, which reduced the oxidize d intermediates of the peroxidase- and oxidase-catalyzed reactions. Si nce ascorbate itself was oxidized in these reactions, the inhibition w as not persistent and it was released once the ascorbate present in th e assay mixture had been oxidized. Ascorbate delayed the oxidation of NADH 10-fold more efficiently than the oxidation of coniferyl alcohol. Although the level and the redox state of apoplastic ascorbate were l ower in lignifying needles than in mature needles, the concentration, which was 1.17 mM in apoplastic washing fluids, was sufficiently high to inhibit peroxidase activity in vitro. These results suggest that pe roxidases can catalyze lignification only if local differences exist i n the concentration of reduced ascorbate between lignifying and non-li gnifying tissues.