Metabolism of nitrogen functionalities (e.g., amines, amides) is important because such functional groups are found in many natural products (e.g., morphine, cocaine, nicotine) and in numerous important drugs (e.g., phenothiazines, antihistamines, tricyclic antidepressants, j-adrenergic agents, sympathomimetic phenylethylamines, benzodiazepines).159 The following discussion divides nitrogen-containing compounds into three basic classes:
1. Aliphatic (primary, secondary, and tertiary) and alicyclic (secondary and tertiary) amines
2. Aromatic and heterocyclic nitrogen compounds
The susceptibility of each class of these nitrogen compounds to either a-carbon hydroxylation or N-oxidation and the metabolic products that are formed are discussed.
The hepatic enzymes responsible for carrying out a-carbon hydroxylation reactions are the CYP mixed-function oxidases. The N-hydroxylation or N-oxidation reactions, however, appear to be catalyzed not only by CYP but also by a second class of hepatic mixed-function oxidases called amine oxidases (sometimes called N-oxidases).160 These enzymes are NADPH-dependent flavoproteins and do not contain CYP.161,162 They require NADPH and molecular oxygen to carry out N-oxidation.
Tertiary Aliphatic and Alicyclic Amines. The oxidative removal of alkyl groups (particularly methyl groups) from tertiary aliphatic and alicyclic amines is carried out by hepatic CYP mixed-function oxidase enzymes. This reaction is commonly referred to as oxidative N-dealkylation.163 The initial step involves a-carbon hydroxylation to form a carbinolamine intermediate, which is unstable and undergoes spontaneous heterolytic cleavage of the C-N bond to give a secondary amine and a carbonyl moiety (aldehyde or ketone).164,165 In general, small alkyl groups, such as methyl, ethyl, and isopropyl, are removed rapidly.163 N-dealkylation of the t-butyl group is not possible by the carbinolamine pathway because a-carbon hydroxylation cannot occur. The first alkyl group from a tertiary amine is removed more rapidly than the second alkyl group. In some instances, bisdealkylation of the tertiary aliphatic amine to the corresponding primary aliphatic amine occurs very slowly.163 For example, the tertiary amine imipramine
(Tofranil) is monodemethylated to desmethylimipramine (desipramine).166,167 This major plasma metabolite is pharmacologically active in humans and contributes substantially to the antidepressant activity of the parent drug.168 Very little of the bisdemethylated metabolite of imipramine is detected. In contrast, the local anesthetic and antiarrhyth-mic agent lidocaine is metabolized extensively by N-deethy-lation to both monoethylglycylxylidine and glycyl-2,6-xyli-dine in humans.169,170
Numerous other tertiary aliphatic amine drugs are metabolized principally by oxidative N-dealkylation. Some of these include the antiarrhythmic disopyramide (Norpace),171,172 the antiestrogenic agent tamoxifen (Nolvadex),173 diphenhydramine (Benadryl), 174,175chlorpromazine (Thorazine),176,177 and (+)-a-propoxyphene (Darvon).178 When the tertiary amine contains several different substituents capable of undergoing dealkylation, the smaller alkyl group is removed preferentially and more rapidly. For example, in benzpheta-mine (Didrex), the methyl group is removed much more rapidly than the benzyl moiety.179
An interesting cyclization reaction occurs with methadone on N-demethylation. The demethylated metabolite normethadone undergoes spontaneous cyclization to form the enamine metabolite 2-ethylidene-1,5-dimethyl-3,3-diphenyl-pyrrolidine (EDDP).180 Subsequent N-demethylation of EDDP and isomerization of the double bond leads to 2-ethyl-5-methyl-3,3-diphenyl-1-pyrroline (EMDP).
Many times, bisdealkylation of a tertiary amine leads to the corresponding primary aliphatic amine metabolite, which is susceptible to further oxidation. For example, the bisdesmethyl metabolite of the H1-histamine antagonist brompheniramine (Dimetane) undergoes oxidative deami-nation and further oxidation to the corresponding propionic acid metabolite.181 Oxidative deamination is discussed in greater detail when we examine the metabolic reactions of secondary and primary amines.
Like their aliphatic counterparts, alicyclic tertiary amines are susceptible to oxidative N-dealkylation reactions. For example, the analgesic meperidine (Demerol) is metabolized principally by this pathway to yield normeperidine as a major plasma metabolite in humans.182 Morphine, N-eth-ylnormorphine, and dextromethorphan also undergo some N-dealkylation.183
Direct N-dealkylation of t-butyl groups, as discussed below, is not possible by the a-carbon hydroxylation pathway. In vitro studies indicate, however, that N-t-butyl-norchlorocyclizine is, indeed, metabolized to significant amounts of norchlorocyclizine, whereby the t-butyl group is lost.184 Careful studies showed that the t-butyl group is removed by initial hydroxylation of one of the methyl groups of the t-butyl moiety to the carbinol or alcohol product.185 Further oxidation generates the corresponding carboxylic acid that, on decarboxylation, forms the N-isopropyl derivative. The N-isopropyl intermediate is dealkylated by the normal a-carbon hydroxylation (i.e., carbinolamine) pathway to give norchlorocyclizine and acetone. Whether this is a general method for the loss of t-butyl groups from amines is still unclear. Indirect N-dealkylation of t-butyl groups is not observed significantly. The N-t-butyl group present in many jS-adrenergic antagonists, such as terbutaline and
salbutamol, remains intact and does not appear to undergo any significant metabolism.186
Alicyclic tertiary amines often generate lactam metabolites by a-carbon hydroxylation reactions. For example, the tobacco alkaloid nicotine is hydroxylated initially at the ring carbon atom a to the nitrogen to yield a carbinolamine intermediate. Furthermore, enzymatic oxidation of this cyclic carbinolamine generates the lactam metabolite cotinine.187,188 Formation of lactam metabolites also has been reported to occur to a minor extent for the antihistamine cyprohep-
tadine (Periactin)189,190 and the antiemetic diphenidol (Vontrol).191
N-oxidation of tertiary amines occurs with several drugs.192 The true extent of N-oxide formation often is complicated by the susceptibility of N-oxides to undergo in vivo reduction back to the parent tertiary amine. Tertiary amines such as H1-histamine antagonists (e.g., orphenadrine, tripel-ennamine), phenothiazines (e.g., chlorpromazine), tricyclic antidepressants (e.g., imipramine), and narcotic analgesics (e.g., morphine, codeine, and meperidine) reportedly form
N-oxide products. In some instances, N-oxides possess pharmacological activity.193 A comparison of imipramine N-oxide with imipramine indicates that the N-oxide itself possesses antidepressant and cardiovascular activity similar to that of the parent drug.194,195
Secondary and Primary Amines. Secondary amines (either parent compounds or metabolites) are susceptible to oxidative N-dealkylation, oxidative deamination, and N-oxidation reactions.163'196 As in tertiary amines, N-dealkylation of secondary amines proceeds by the carbinolamine pathway. Dealkylation of secondary amines gives rise to the corresponding primary amine metabolite. For example, the ^-adrenergic blockers propranolol46'47 and oxprenolol197 undergo N-deisopropylation to the corresponding primary amines. N-dealkylation appears to be a significant biotransformation pathway for the secondary amine drugs methamphetamine198,199 and ketamine,200,201 yielding amphetamine and norketamine, respectively.
The primary amine metabolites formed from oxidative dealkylation are susceptible to oxidative deamination. This process is similar to N-dealkylation, in that it involves an initial a-carbon hydroxylation reaction to form a carbinolamine intermediate, which then undergoes subsequent carbon-nitrogen cleavage to the carbonyl metabolite and ammonia. If a-carbon hydroxylation cannot occur, then oxidative deamination is not possible. For example, deamination does not occur for norketamine because a-carbon hydroxylation cannot take place.200,201 With methamphetamine, oxidative deamination of primary amine metabolite amphetamine produces phenylacetone.198,199
In general, dealkylation of secondary amines is believed to occur before oxidative deamination. Some evidence indicates, however, that this may not always be true. Direct deamination of the secondary amine also has occurred. For example, in addition to undergoing deamination through its desisopropyl primary amine metabolite, propranolol can undergo a direct oxidative deamination reaction (also by a-carbon hydroxylation) to yield the aldehyde metabolite and isopropylamine (Fig. 3.9).202 How much direct oxidative deamination contributes to the metabolism of secondary amines remains unclear.
Some secondary alicyclic amines, like their tertiary amine analogs, are metabolized to their corresponding lactam derivatives. For example, the anorectic agent phen-metrazine (Preludin) is metabolized principally to the lactam product 3-oxophenmetrazine.203 In humans, this lactam metabolite is a major urinary product. Methylphenidate (Ritalin) also reportedly yields a lactam metabolite, 6-oxori-
talinic acid, by oxidation of its hydrolyzed metabolite, rital-inic acid, in humans.204
Metabolic N-oxidation of secondary aliphatic and alicyclic amines leads to several N-oxygenated products.196 N-hydroxylation of secondary amines generates the corresponding N-hydroxylamine metabolites. Often, these hydroxylamine products are susceptible to further oxidation (either spontaneous or enzymatic) to the corresponding ni-trone derivatives. N-benzylamphetamine undergoes metabolism to both the corresponding N-hydroxylamine and the nitrone metabolites.205 In humans, the nitrone metabolite of phenmetrazine (Preludin), found in the urine, is believed to be formed by further oxidation of the N-hydroxylamine intermediate N-hydroxyphenmetrazine.203 Importantly, much less N-oxidation occurs for secondary amines than oxidative dealkylation and deamination.
Primary aliphatic amines (whether parent drugs or metabolites) are biotransformed by oxidative deamination (through the carbinolamine pathway) or by N-oxidation. In general, oxidative deamination of most exogenous primary amines is
carried out by the mixed-function oxidases discussed previously. Endogenous primary amines (e.g., dopamine, norepinephrine, tryptamine, and serotonin) and xenobiotics based on the structures of these endogenous neurotransmitters are metabolized, however, via oxidative deamination by a specialized family of enzymes called monoamine oxidases (MAOs).206
MAO is a flavin (FAD)-dependent enzyme found in two isozyme forms, MAO-A and MAO-B, and widely distributed in both the CNS and peripheral organs. In contrast, CYP exists in a wide variety of isozyme forms and is an NADP-dependent system. Also the greatest variety of CYP isozymes, at least the ones associated with the metabolism of xenobiotics, are found mostly in the liver and intestinal mucosa. MAO-A and MAO-B are coded by two genes, both on the X-chromosome and have about 70% amino acid sequence homology. Another difference between the CYP and MAO
families is cellular location. CYP enzymes are found on the endoplasmic reticulum of the cell's cytosol, whereas the MAO enzymes are on the outer mitochondrial membrane. In addition to the xenobiotics illustrated in the reaction schemes, other drugs metabolized by the MAO system include phenylephrine, propranolol, timolol and other ^-adrenergic agonists and antagonists, and various phenylethylamines.206 Structural features, especially the a-substituents of the primary amine, often determine whether carbon or nitrogen oxidation will occur. For example, compare amphetamine with its a--methyl homologue phentermine. In amphetamine, a--carbon hydroxylation can occur to form the carbinolamine intermediate, which is converted to the oxidatively deami-nated product phenylacetone.67 With phentermine, a-carbon hydroxylation is not possible and precludes oxidative deamination for this drug. Consequently, phentermine would be expected to undergo N-oxidation readily. In humans, p-hydroxylation and N-oxidation are the main pathways for biotransformation of phentermine.207
Indeed, N-hydroxyphentermine is an important (5%) urinary metabolite in humans.207 As discussed below, N-hydroxylamine metabolites are susceptible to further oxidation to yield other N-oxygenated products.
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