PMSF is a serine protease inhibitor that irreversibly inhibits serine proteases by sulfonylation of the serine residue in the active site of the protease. It is widely used in protocols for the isolation of proteins.

PMSF is inactivated in aqueous solutions. The rate of inactivation increases with increasing pH and is faster at 25°C than at 4°C. The half-life of a 20mM aqueous solution of PMSF is ~35 minutes at pH 8.0. This short half-life means that aqueous solutions of PMSF can be safely discarded after they have been rendered alkaline (pH >8.6) and stored for several hours at room temperature.