Biotransformations hydroxylation sites

Using the parent compound depletion method, pyrethroid metabolic rate constants (i.e., Umax and K, hast, etc.) for hydroxylation by cytochrome P450 enzymes or hydrolysis by carboxylesterases were developed by Scollon et al. (2009). The sources of the enzymes were rat and human microsomes. The pyrethroids they studied included bifenthrin, S-bioallethrin, bioresmethrin, p-cyfluthrin, cypermethrin, cis-permethrin, and frans-permethrin. The depletion method considers multiple hydroxylations as a single biotransformation at sites on either the acid or alcohol moieties, or on a combination of both. The metabolic pathways (Tables D1-D15 and E1-E15 of Appendices D and E, respectively) require Umax, Am, and values for the individual hydroxylated and hydrolyzed products. It is interesting that only bioresmethrin and cypermethrin per se were found to actually be hydrolyzed. 

In summary, utilizing biotransformation techniques, eight out of the 28 carbon atoms in MFA were successfully hydroxylated. In Fig. (14), arrows indicate the sites of hydroxylation. It is noteworthy that despite the hindered nature of C14, cultures UC 5059, UC 11141, and UC 11144 were able to introduce a hydroxyl to give 23 in a 10-15% yield. 

The mass shifts of some fragment peaks in the MS-MS spectra of KR-60436 and its metabolites are summarized in Table 10.4 [14], From the interpretation of the data, four biotransformation processes can be recognized, each related to a specific site in the stracture (see the structirre in Table 10.4) (A) hydroxylation (mass shift of +16 Da), (B) double-bond formation (-2 Da), (C) loss of the hydroxyethyl side chain (-44 Da), and (D) demethylation (-14 Da). Combination of these processes takes place as well, e.g., in M2, where both double-bond formation and hydroxylation takes place. From the MS-MS data, it can be concluded that hydroxylation at (A) blocks the fragmentation of the pyridine ring, i.e., no loss of 84 Da is observed, and that double-bond formation at (B) blocks the formation of the fragment at m/z 150. The identification of the metabolites was as follows Ml and M2 contained a hydroxy-group in (A). M2, M4, M6, and M7 contained a donble bond in (B). M3 and M4 were demethylated at D. In M5 and M7, the hydroxyethyl group was lost [14]. 

Decreasing the electron density at the site of a GYP450-mediated biotransformation will generally attenuate aromatic hydroxylation but have little impact upon N- and O-dealkylation processes. 

In plants, however, almost all naturally occurring coumarins are 7-hydroxy derivatives, marking the possible site of the initiation of coumarin biotransformation. It seems that the electron charge theory is not congruent with any relationship between the wide occurrence of the 7-hydroxylated plant coumarins and their metabolism. It is, therefore, likely that the hydroxy group is formed before the ring closure [51,52] and the hydroxylated products do not originate from the parent compound. 

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