Soil storage method may alter enzymatic activity being storage conditions of the soil samples prior to analysis decisive for the results. Studies made on freshly collected soils are generally preferred. However it is always not possible due to practical reasons since for example sampling is often restricted to short period of the year or because a great quantity of microbiological analyses must be made on time and by few people. On this context, soil storage methods are needed, being cold at 4°C the most widely used although sometimes alternative storage methods are also utilized. The aim of this study is to evaluate the effect of two alternative storage methods of soil samples (freezing at –20°C and air drying conservation methods) in comparison to cold at 4°C on the enzymatic activities (dehydrogenase, phosphatase, β-glucosidase and urease soil enzymes). Samples of two forest ecosystems (pine and holm oak forest stand) were taken in two different season of the year (winter and spring 2009). Results showed that enzymatic activities differed when freezing or air drying conservation methods were used in comparison with cold soil samples. Generally, alternative soil storage methods presented lower enzymatic activity than cold at 4°C. However, these changes depend on season and sampling location.