N-Reduction of ximelagatran and N-hydroxy-melagatran, amidoxime prodrugs in the reconstituted system consisting of cytochrome b5, NADH-cytochrome b5reductase, and CYP2D
| Activity | ||
|---|---|---|
| Ethyl-melagatran | N-Hydroxy-melagatran | |
| nmol/min/mg protein | ||
| [nmol/min/nmol P450] | ||
| Complete system, pig | 34.40 ± 1.54 3-a | 17.13 ± 0.08 |
| [14.76 ± 0.38] | [7.35 ± 0.02] | |
| Without CYP2D | <DL | 0.38 ± 0.02 |
| Without protein | <DL | 0.07 ± 0.04 |
Complete incubation mixture consisted of 72.5 pmol of cytochromeb5, 0.35 U of NADH-cytochromeb5 reductase, and 5 μg of pig benzamidoxime reductase (CYP2D), 40 μM DLPC, and 1 mM ximelagatran or 2 mMN-hydroxy-melagatran in 100 mM potassium phosphate buffer pH 6.3. After preincubation for 5 min, the reaction was initiated by 1 mM NADH to a total volume of 150 μl. After 30 min, the reaction was stopped by adding 150 μl of cold methanol. Precepitated proteins were sedimented by centrifugation and 15 μl of the supernatant were analyzed by HPLC.
DL, determination limit.
↵3-a Conversion rates are means ± S.D. of four determinations.