Involvement of peroxidase in chlorophyll degradation in stored broccoli (Brassica oleracea L.) and inhibition of the activity by heat treatment
ABSTRACT Cell localization of chlorophyll (Chl)-degrading peroxidase and the effects of heat treatment on the activity were determined to elucidate the involvement of the enzyme with yellowing in stored broccoli. Subcellular fractions and intact chloroplasts were prepared by differential and Percoll gradient centrifugation, respectively. In fresh broccoli, only low levels of Chl-degrading peroxidase activity were located in the intact chloroplast, and the highest activity was present in the cytosolic fraction. The Chl content of broccoli without heat treatment decreased greatly after 4 days storage at 15 °C, whereas the content in broccoli treated at 50 °C for 2 h showed almost no change during storage. In microsomal and cytosolic fractions, the activity of C2 (Rf=0.47) isoperoxidase, which is involved in Chl degradation, increased greatly with floret yellowing. The Chl-degrading peroxidase activities in microsomal and cytosolic fractions also increased strongly after 4 days of storage. In addition, the Chl-degrading peroxidase activity as well as C2 isoperoxidase activity was suppressed by heat treatment. These results indicate that heat treatments may be effective in inhibiting senescence in part by suppressing the enhancement of Chl-degrading peroxidase activity in the microsomes and the cytosol.