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Gross phosphorus (P) fluxes measured in isotopic dilution studies with 33P labeled soils include the biological processes of microbial P immobilization remineralization of immobilized P and mineralization of non microbial soil organic P. In this approach isotopic dilution due to physicochemical processes is taken into account. Our objectives were to assess the effect of inorganic P availability on gross P mineralization and immobilization in soil under permanent grassland and to relate these fluxes to soil respiration phosphatase activity and substrate availability as assessed by an enzyme addition method. We used soils from an 18 year old grassland fertilization experiment near Zurich Switzerland that were collected in two treatments which differed only in the amount of mineral P applied (0 and 17 kg P ha-1 yr-1 in NK and NPK respectively). Water extractable phosphate was low (0.1 and 0.4 mg P kg-1 soil in NK and NPK while hexanol labile (microbial) P was high (36 and 54 mg P kg-1 soil in NK and NPK). Extremely fast microbial P uptake under P limited conditions in NK necessitated the use of a microbial inhibitor when determining isotopic dilution due to physicochemical processes. At the higher inorganic P availability in NPK however isotopic exchange parameters were similar in the presence and absence of a microbial inhibitor. Phosphatase activity was higher in NK than in NPK while soil respiration water extractable organic P and its enzyme labile fraction were not affected by P status. Together the results showed that inorganic P availability primarily affected microbial P immobilization which was the main component of gross P fluxes in both treatments. Gross P mineralization rates (8.2 and 3.1 mg P kg-1 d-1 for NK and NPK) during the first week were higher than reported in other studies on arable and forest soils and at least equal to isotopically exchangeable P due to physicochemical processes confirming the importance of microbial processes in grassland soils.