Extent of CYP2A13-catalyzed nicotine (CN) metabolism and loss of enzyme activity in the absence and presence of cyanidea
Time | Nicotine Metabolismb | % Activity Remaining | ||||
|---|---|---|---|---|---|---|
| – CN | + CN | – CN | + CN | |||
| min | % | |||||
| 0 | 0 | 0 | 100c | 100c | ||
| 0.5 | 36 ± 3c,d | 14 ± 3e | 97 ± 3c | 103 ± 5e | ||
| 1 | 67 ± 7c | 19 ± 2e | 89 ± 2c,d | 95 ± 4d,e | ||
| 2 | 89 ± 4c,d,f | 28 ± 5c | 78 ± 2c,d,g | 96 ± 10c,d | ||
| 5 | 95 ± 1d,e,f | 53 ± 3d,e,f | 66 ± 2d,e,g | 88 ± 6d,e,g | ||
| 10 | 95 ± 0e,f | 78 ± 6c,d,f | 53 ± 1d,e,g | 77 ± 6c,d,g | ||
| 16 | 95 ± 1e,f | 94 ± 2d,e,f | 47 ± 2d,e,g | 76 ± 13d,e,g | ||
↵ a CYP2A13 was incubated with 25 μM [5-3H]-(S)-nicotine at 30°C in the presence or absence of 1 mM KCN. At the indicated times, aliquots of the reaction were removed and either analyzed by radioflow HPLC for nicotine metabolites (Fig. 5) or for coumarin 7-hydroxylation in a secondary reaction as described under Materials and Methods. The extent of nicotine metabolism and the percent activity remaining was calculated based on control samples (+ nicotine, – NADPH)
↵ b Nicotine metabolism is the sum of total nicotine metabolites compared with total nicotine at time 0
↵ c Values are means ± S.D. of four independent experiments
↵ d Values are significantly lower than previous time point, P ≤ 0.05
↵ e Values are means ± S.D. of three independent experiments
↵ f Values are less than those at time 0, P ≤ 0.005
↵ g Values are less than those at time 0, P ≤ 0.01