The correlation between the natural levels of GST and the tolerance to the insecticide decamethrin (dMT), as well as the interaction between the molecules of affinity purified enzyme and the insecticide were investigated in order to collect further information on the obscure role of the Glutathione S-transferase system (GST) as a mechanism of defence against pyrethroids. The studies were carried out, comparatively, on the larvae and pupae developmental stages of the coleopteran Tenebrio molitor, which exhibit varying natural levels of GST activity. No stage dependent susceptibility of the insect against pyrethroid insecticides was found during the first 24 h, however 48 h after treatment, the KD50 dose increased significantly due to the recovery of some individuals from the larvae stage. Simultaneous injection of decamethrin with compounds which inhibit GST activity in vitro, resulted in an increased tolerance, which was more pronounced in the pupae stage. Inhibition studies combined with competitive fluorescence spectroscopy and high pressure liquid chromatography (HPLC) showed that the insecticide binds probably to the active site of the enzyme inhibiting its activity towards CDNB in a competitive manner, but is not conjugated with GSH. According to this, GST offers a passive protection towards pyrethroid insecticides by binding to their molecule in a sequestering mechanism.