The Flavin-Containing Monooxygenase FMO

like CYP, are located in the endoplasmic reticulum and are involved in the oxidation of numerous organic xenobiotics containing nitrogen, sulfur, or phosphorus heteroatoms as well as some inorganic ions. 

TABLE 10.2. Substrates Oxidized by Flavin-Containing Monooxygenases 

Sec- and fert-acyclic and cyclic amines A-Alkyl and N,A-dialkylarylamines Hydrazines Primary amines 

Thiols and disulfides Cyclic and acyclic sulfides Mercapto-purines, -pyrimidines, -imidazoles Dithio acids and dithiocarbamides Thiocarbamides and thioamides 

Reactivation of the enzyme is considered to be the rate-limiting step in the reaction. Because oxidation of the substrate occurs more rapidly than the regeneration of the active enzyme, the Vmax values are relatively similar for a variety of substrates even though the Km values differ. 

Tertiary amines such as trimethylamine and dimethylamine had long been known to be metabolized to A -oxides by a microsomal amine oxidase that was not dependent on CYP. This enzyme, now known as the microsomal flavin-containing monooxygenase (FMO), is also dependent on NADPH and 02, and has been purified to homogeneity from a number of species. Isolation and characterization of the enzyme from liver and lung samples provided evidence of clearly distinct physicochemical properties and substrate specificities suggesting the presence of at least two different isoforms. Subsequent studies have verified the presence of multiple forms of the enzyme. 

Most FMO substrates are also substrates for CYP. Since both enzymes are microsomal and require NADPH and oxygen, it is difficult to distinguish which enzyme is responsible for oxidation without the use of techniques involving specific inactivation or inhibition or one or the other of these enzymes while simultaneously examining the 

Toxicologically it is of interest that the FMO enzyme is responsible for the oxidation of nicotine to nicotine F-N-oxide, whereas the oxidation of nicotine to cotinine is catalyzed by two enzymes acting in sequence CYP followed by a soluble aldehyde dehydrogenase. Thus nicotine is metabolized by two different routes, the relative contributions of which may vary with both the extrinsic and intrinsic factors outlined in Chapter 9. 

---

Figure 7.9 Examples of oxidations catalyzed by the flavin-containing monooxygenase (FMO).

Monooxygenations are those oxidations in which one atom of molecular oxygen is reduced to water while the other is incorporated into the substrate. Microsomal monooxygenation reactions are catalyzed by nonspecific enzymes such as the flavin-containing monooxygenases (FMOs) or the multienzyme system that has cytochrome P450s (CYPs) as the terminal oxidases. 

The assessment of clearance is complicated by the numerous mechanisms by which compounds may be cleared from the body. These mechanisms include oxidative metabolism, most commonly by CYP enzymes, but also in some cases by other enzymes including but not limited to monoamine oxidases (MAO), flavin-containing monooxygenases (FMO), and aldehyde oxidase [45, 46], Non-oxidative metabolism such as conjugation or hydrolysis may be effected by enzymes such as glucuronyl transferases (UGT), glutathione transferases (GST), amidases, esterases, or ketone reductases, as well as other enzymes [47, 48], In addition to metabolic pathways, parent compound may be excreted directly via passive or active transport processes, most commonly into the urine or bile. 

Monooxygenation of xenobiotics are catalyzed either by the cytochrome P450 (CYP)-dependent monooxygenase system or by flavin-containing monooxygenases (FMO). 

In addition to cytochrome P-450 enzymes, another enzyme that mediates phase I oxidations is flavin-containing monooxygenase (FMO), likewise contained in the endoplasmic reticulum. It is especially effective in oxidizing primary, secondary, and tertiary amines. Additionally, it catalyzes oxidation of other nitrogen-containing xenobiotic compounds, as well as those that contain sulfur and phosphorus, but does not bring about hydroxylation of carbon atoms. 

Phase I oxidation generally is described as the addition of an oxygen atom (e.g., as an hydroxyl moiety) to the parent molecule. Phase I oxidation is carried out by multiple enzyme pathways, including the various isoforms of the cytochrome P450 (CYP) family and the non-P450 biotransformation enzymes such as flavin-containing monooxygenase (FMO) and monamine oxidase (MAO). 

Oxidative biotransformations, which constitutes the major portion of Phase I reactions, can be catalyzed by either cytochrome P450s (CYP450) or nonmicrosomal enzymes such as flavin-containing monooxygenases (FMOs), monoamine oxidase (MAOs), alcohol dehydrogenase, and aldehyde dehydrogenase. As listed in Table 5.1,... 

This approach, as well as other approaches using chemical inhibitor or inhibitory monoclonal antibodies, assumes that the probe substrates behave like the test compound. A simplified approach is to study the test compound in expressed enzymes only and then assume hepatic metabolism is covered by the several expressed enzymes that have been assayed (Venkatakrishnan et al., 2001). In other words, this simplified approach uses the sum of various supersomal clearances instead of the clearance in liver microsomes as 100% of hepatic metabolic clearance. While in most cases several major CYPs may have covered the majority of hepatic metabolism, other enzymes could also contribute significantly to the overall metabolism. Examples of non-CYP microsomal enzymes often involved in drug metabolism include flavin-containing monooxygenase (FMO), UDP—glucuronosyltransferase (UGT), sulfotransfer-ase (SULT), and many others (Parkinson and Ogilvie, 2008). 

Flavin-containing monooxygenases (FMOs) are a class of flavin monooxygenases that contain tightly boxmd FAD and require NADPH as coenzyme. These enzymes catalyze the monooxygenation of different heteroatoms, with the natural role of participating in the detoxification of drugs and xenobiotics [22,53]. Most of the FMOs are membrane associated, which makes it difficult to obtain them for further application. The human proteome contains five FMO isoforms, FMOl-5. FMOS seems to be the dominant enzyme in the himian body. 

Microsomal flavin-containing monooxygenases. As well as the cytochromes P-450 MFO system, there is also a system, which uses FAD. This flavin-containing monooxygenase or FMO enzyme system is found particularly in the microsomal fraction of the liver, and the monomer has a molecular weight of around 65,000. Each monomer has one molecule of FAD associated with it. The enzyme may accept electrons from either NADPH or NADH although the former is the preferred cofactor. It also requires molecular oxygen, and the overall reaction is as written for cytochromes P-450 ... 

Larsen-Su, S. and Williams, D. E., Dietary indole-3-carbinol inhibits FMO activity and the expression of flavin-containing monooxygenase form 1 in rat liver and intestine, Drug Metab. Disposition, 24, 927, 1996. 

---