The activity of ascorbate oxidase (AO) purified from green zucchini squash (Cucurbita pepo medullosa var. Milano) decreased on freeze-drying. In rehydrated samples, enzyme activity was strictly correlated with the addition of lyoprotectants, the freeze temperature, the enzyme concentration and the storage time. The addition of lyoprotectants such as sucrose, mannose, glucose or polyvinylpyrrolidone (PVP) improved enzyme stabilization. A notable loss of enzymic activity was observed when freezing was performed by immersion in liquid nitrogen in comparison to samples frozen at -20 or - 75 degrees C, Moreover, sugar-based lyoprotectants provided better stabilization when the freeze temperature was -75 degrees C. PVP species were most effective in stabilizing AO when samples were frozen in liquid nitrogen. The decrease of enzyme activity on freeze-drying was inversely related to the enzyme concentration . A decrease in enzyme activity was observed in all samples after rehydration at longer storage times, the decrease being less evident in samples containing the lyoprotectants mentioned earlier. Freeze-thawing experiments showed a similar trend considering the presence of lyoprotectant and the enzyme concentration. Samples without additives showed the highest residual activities after the first freeze-thaw cycle but samples were much more prone to a gradual decrease of residual activity with subsequent freeze-thawing cycles.