CYTOCHROME-P450 ISOZYMES INVOLVED IN AROMATIC HYDROXYLATION AND SIDE-CHAIN N-DESISOPROPYLATION OF ALPRENOLOL IN RAT-LIVER MICROSOMES

Alprenolol 4-hydroxylation and N-desisopropylation in liver microsomes from male Wistar rats were kinetically analyzed to be biphasic. In th e 4-hydroxylation at a low substrate concentration (5 mu M), significa nt strain [Wistar>Dark Agouti (DA)] and sex (male>female) differences were observed, and the differences decreased at a high substrate conce ntration (1 mM). In the N-desisopropylation, only a strain difference (Wistar>DA) was observed at the low substrate concentration. Cytochrom e P450BTL (P450BTL, corresponding to CYP2D2) in a reconstituted system with 5 mu M alprenolol had high 4-hydroxylase activity, which was abo ut 10 times that of P450ml corresponding to CYP2C11, and N-desisopropy lase activity at a similar extent to P450ml. The two microsomal activi ties at 5 mu M alprenolol were efficiently decreased by antibodies aga inst P450BTL and by sparteine, a typical substrate of the CYP2D subfam ily. Polyclonal antibodies against P450ml and P450PB-1 (corresponding to CYP3A2) partially suppressed only N-desalkylation at 5 mu M, wherea s they reduced the two activities at 1 mM. P450ml showed a high N-desi sopropylase activity at a substrate concentration of 1 mM, where the s ex difference was not observed. Furthermore, P450PB-2 corresponding to CYP2C6, which is one of the major P450 isozymes in female rats, also had 4-hydroxylase and N-desalkylase activities. These results suggest that a CYP2D isozyme(s) is the primary enzyme in alprenolol 4-hydroxyl ation and N-desisopropylation in a lower substrate concentration range , and that the involvement of some male-specific P450 isozyme(s) other than CYP2C11 or CYP3A2 may cause the sex difference in the 4-hydroxyl ation. In a higher substrate concentration range, CYP2C11 is thought t o play a major role particularly in N-desisopropylation in male rats. In female rats, some major constitutive P450 isozyme(s) with a relativ ely high K-m value (e.g., CYP2C6) may be involved in the metabolism of alprenolol, resulting in the disappearance of the sex difference.