Characteristics of dibucaine inhibition of “usual” and “atypical” forms of BChE.

<p>DN values were calculated in assays performed at various dibucaine concentration at 25°C using 5mM BTC (A) or PTC (B) as substrate with two types of 400-fold diluted serum samples from heterozygous individuals with usual UU and UA phenotype. Dibucaine Number (DN) = (1 - (butyrylcholinesterase activity in presence of inhibitor/butyrylcholinesterase activity in absence of inhibitor)) x 100. Assays were performed in triplicate, standard error for all points is less than 3% of the values.</p

The effect of serum dilution on BChE activity.

<p>The activity was estimated based on the mean of hydrolysis rate (OD/min) determined from at least 5 spectrophotometer reads (every 1 minute) to ensure linearity and regularity of slopes. Assays were done in triplicate, error bars represent mean ± standard error (Student’s t-test, p<0.05). <b>A.</b> The effect of BTC concentration on the BChE activity in 100, 400, 4000 -fold diluted serum samples. Inset shows a Lineweaver–Burk replot of the results presented in Fig 2A at [BTCh] = 0.1, 0.5 and 1 mM. <b>B.</b> The effect of 5mM and 2.5 mM BTC concentration on the BChE activity in variously diluted serum samples. <b>C.</b> BChE activity in 100, 400, 2000 -fold diluted eleven randomly selected serum samples from our serum collection samples.</p

Optimization of the BChE assay linearity.

<p>Measurement of TNB ion concentration at 412 nm produced in Ellman's reaction in the presence of 5 mM BTC and 40 (5), 100 (2), 200 (1), 400 (0.5), 600 (0.3), 800 (0.25)-fold diluted serum. Volume (μl) of serum in the reaction mixture (200 μl total volume) is shown above in brackets. Assays were performed in triplicate, standard error for all points is less than 2.5% of the values.</p