Neuropathy target esterase (NTE) activity is operatively defined in this paper as the phenyl valerate esterase activity resistant to 40 microM paraoxon but sensitive to 250 microM mipafox. Molecular exclusion column chromatography with Sephacryl S-300 of the soluble (S) fraction from chick sciatic nerve demonstrated two NTE activity peaks. The first eluted with the front, thus indicating a mol. wt. of over 700 kDa (peak Vo), while the second peak eluted with kd = 0.36, suggesting a mol. wt. of about 100 kDa. The curve of total phenyl valerate (PVase) activity inhibition with paraoxon (0.19-200 microM) shows that at a concentration of 40 microM the esterases highly sensitive to paraoxon are inhibited in the Vo and 100-kDa peaks. The NTE activity in these two peaks in turn represented 31% and 44% of the 40 microM paraoxon resistant activity, respectively. The mipafox inhibition curves (1.0-250 microM) revealed different sensitivities to mipafox, with I50 values (t = 30 min) of approximately 1.47 and 63 microM, for Vo and 100-kDa peaks respectively. Mipafox sensitivity of the Vo and 100-kDa peaks correlates with the two components, that had been deduced from the kinetic properties of the S-fraction.