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Measured as percent loss at 4 h in rat hepatocytes.
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In vitro metabolism assay procedure: In vitro metabolic stability of analogs in plated rat hepatocytes (Sprague-Dawley) obtained from Cedra Corporation. Metabolic activity was determined in triplicate using a total volume of 0.2 mL containing 0.25 μM NCE incubated in rat hepatocyte and matrigel blank microtiter plates. The plates were maintained at 37 °C throughout the study. Samples were removed from wells at 0, 2, and 4 h, and NCE samples were analyzed by HP-LC/MS/MS with reverse-phase chromatography. To improve analytical efficiency, compounds were grouped together (post-incubation) into a multi-compound assay. Samples from like time-points containing the different compounds were combined and an internal standard (1.1 ng/mL stock) was added. Results for each compound were expressed as the ratio of the compound response area over the internal standard response area. Percent loss was calculated by dividing the 2 and 4 h ratios by the 0 h ratio.
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