Neuropathy target esterase (NTE) is a membrane-bound carboxylesterase activity which is proposed as the target site in nerve tissue for initiation of organophosphate-induced delayed neuropathy. This activity is identified as phenyl valerate hydrolysis which is resistant to treatment with paraxon and sensitive to co-incubation with paraxon and mipafox. NTE preparations were obtained, which did not contain paraxon-sensitive or mipafox-resistant hydrolases, by selective reconstitution of detergent-solubilized NTE from chick embryo brain into asolectin vesicles during gel filtration. The topography of the catalytic site of NTE was then examined by investigating the inhibition of NTE by a series of 3-alkylthio- and 3-arylthio-1,1.1-trifluoro-propan-2-ones. These trifluoromethyl ketones were found to be rapidly reversible, competitive inhibitors of NTE with I50 values 1.3 x 10(-4) M to 4.9 x 10(-8) M. Correlation of I50 values with octanol/water partition coefficients (P), in the range of log P = 1.5 to 5.9. indicated that the optimal lipophilicity for NTE substrates and inhibitors is in the range of log P = 3.0 to 3.4. Electrophilic substitution at the meta position of aromatic rings increased the inhibitory capacity of these inhibitors, whereas substitution at the ortho position reduced inhibitory capacity. These results indicate both that a large hydrophobic pocket is closely associated with the catalytic residue of NTE, and that affinity for the active site is affected by steric and electronic parameters.