1). The incubations

proceeded for 1 h, at 37 °C. Four readings of each concentration were recorded at intervals of 60 s at 37 °C and 450 nm with constant stirring at 600 rpm in a UV/visible HP 8453 spectrophotometer. The absorbance used to calculate the AZD2281 clinical trial enzyme activity was the average per min of these 4 readings. The concentrations of protein samples were evaluated using the Bradford method (1976) before the enzyme evaluations because all of the enzyme activities were reported in terms of μmol/min/g of protein. Calpain activity in the chicken brain and neuroblastoma cells was analyzed as described elsewhere (Emerick et al., 2010), but before the assay, tissue homogenate was incubated with mipafox (0.01 mM) or (+)-methamidophos (10 mM) or (−)-methamidophos (100 mM) for one hour, at 37 °C. CaCl2 in a concentration of 4 mM was added in the follow proportion: 1 g of tissue or 1 ml of cells (1 × 107/ml)/0.01 ml of OP in ethanol/1 ml of CaCl2. The concentrations of OPs used were based on the NTE inhibition with concentration for each compound selleck screening library causing at least 80% NTE inhibition. Inhibitor concentrations capable of inhibiting 50% of enzyme activity (IC50) were determined using the equation of the line graph of the log of % activity versus the concentration of inhibitor (semilog plots). The semilog plots

are not shown to avoid repetitions of results. The regression coefficients of these lines were calculated using the method of least squares. Differences in biochemical see more analyses were examined for statistical significance by one way ANOVA (Analysis Of Variance) followed by Tukey’s test for multiple comparisons. These tests were performed in Microsoft Office Excel 2007 for Windows. The definition of significance was p < 0.05 for all statistical analyses. All biochemical data are presented as the averages of three samples done in triplicate (n = 3). All biochemical data are expressed as means ± the standard deviation (SD). Control values for NTE and AChE activities

in hens and humans are presented in Table 1. All of the coefficients of variation remained below 20%. AChE activity was not evaluated in the hens’ erythrocytes because a previous study showed that this activity could not be detected (Wilson and Henderson, 1992). The potencies of the isomers of methamidophos against NTE and LNTE differed. The inhibition curves of NTE in hens and humans are depicted in Fig. 2A, C, E and G and IC50 values are reported in Table 2. These data indicate that the (+)-methamidophos form was a more potent inhibitor of NTE than the (−)-methamidophos form. The (−)-methamidophos isomer exhibited an IC50 value approximately 5.6 times greater than did the (+)-methamidophos isomer for the LNTE activity of hen and approximately 4 times that observed for the inhibition of human LNTE activity. The percentage activity versus inhibitor concentrations exhibited high inverse regression coefficients for all NTE activities ( Table 2).