Biotransformation mixed-function oxidases

Cytochrome P450 2C9 is a mixed-function oxidase localized in the endoplasmic reticulum which is responsible for the biotransformation of several nonsteroidal anti-inflammatory diugs, S-warfarin, several sulfonylurea antidiabetics and other diugs. 

The biotransformation systems involved in insecticide metabolism have been studied in the R and S populations to determine any differences which might be potential contributory factors to or results of insecticide resistance. In addition, the possibility of mixed-function oxidase induction has been investigated. Specifically, the studies have encompassed a seasonal study of microsomal mixed-function oxidase (mfo) components, and studies of aldrin, dieldrin and DDT metabolism. 

Hepatic mixed-function oxidase activities demonstrated seasonal trends, with higher specific activities in the cold weather months in both populations with few differences in enzyme activities or cytochrome levels between the two populations. Metabolism of aldrin, dieldrin and DDT was similar between the two populations. R fish have larger relative liver size and, therefore, a greater potential for xenobiotic metabolism. However, biotransformation appears to be of minor importance in chlorinated alicyclic insecticide resistance in mosquitofish barriers to penetration appear to be of greater importance and an implied target site insensitivity appears to be the most important factor in resistance. 

Considerable interest has developed concerning the nature of the mixed function oxidase system in fish and the role that this system may play in the development of toxic responses in these animals. Studies have shown that components of the mixed function oxidase system are present in relatively high concentrations in fish liver (4, 5, 6) and that the enzyme systems in this organ are capable of many of the biotransformation reactions already described for the mammalian liver (7, 8, 9). The presence of this complement of enzymes in the livers of many fishes suggests that this organ too may be particularly sensitive to insult from sub lethal concentrations of many waterborne toxicants. For this reason, methods to evaluate liver function in fish may be particularly useful in identifying the sublethal effects of certain classes of toxicants. 

Lipid-soluble xenobiotics are commonly biotransformed by oxidation in the drug-metabolizing microsomal system (DMMS). For each description below, choose the component of the microsomal mixed-function oxidase system with which it is most closely associated ... 

Microsomal activation for bacterial assays typically has involved crude supernatant fractions (S9) from livers of rats induced by polychlorobiphenyl mixtures, usually Aroclor 1254. These S9 mixtures contain a spectrum of mixed function oxidases and other enzymes active in biotransformation. In such mixtures, a given compound might be activated to become more mutagenic, may be inactivated, or may remain unaffected. All three types of response have been observed with various water residues (9, 14). 

This enzyme system, also known as the mixed function oxidase system (see Chapter 4 Drug Biotransformation), uses NADPH as a cofactor in the metabolism of ethanol (Figure 23-1, right). 

T.A. Popov, andB.J. Blaauboer, Biotransformation of pesticides. The role of mixed function oxidase system in pesticide toxicity, in Toxicology of Pesticides in Animals, ed., T.S.S. Dikshith, CRC Press, Boca Raton, FL, 41, 1991. 

For most drugs, oxidative biotransformation is performed primarily by the mixed-function oxidase enzyme system, which is present predominantly in the smooth endoplasmic reticulum of the liver. This system comprises (1) the enzyme NADPH cytochrome P450 reductase (2) cytochrome P450, a family of heme-containing proteins that catalyze a variety of oxidative and reductive reactions and (3) a phospholipid bilayer that facilitates interaction between the two proteins. Important exceptions to this rule are ethyl alcohol and caffeine, which are oxidatively metabolized by enzymes primarily present in the soluble, cytosolic fraction of the liver. 

Biotransformation of non-polar, non-volatile toxicants is a two-phase biochemical reaction. In the first phase (Phase I), the body s enzyme system introduces a polar group into the toxicant (e.g., by oxidation, reduction, or hydrolysis). See Figure 9.28. The enzymes responsible for these transformations are part of a mixed function oxidase system (MFO) of smooth endoplasmatic reticulum present in liver parenchyma cells and in other tissues (e.g., intestine and gill). 

Like benzo[r ]pyrene, benz[a]anthracene may cross the gastrointestinal lining, pulmonary endothelium, or percutaneous barriers. Benz[a]anthracene is biotransformed to five dihydrodiols and a number of phenolic metabolites by P450 mixed-function oxidases. Detectable levels of benz[a]anthracene can be observed in most internal organs from minutes to hours after administration. Regardless of route of administration, once metabolized, hepatobiliary excretion and elimination through feces is the major route. 

The principal biotransformation of toxicants is catalyzed by the microsomal mixed function oxidase system (MFO). A deficiency of essential fatty acids generally depresses MFO activities. This is also true with protein deficiency. The decreased MFO has different effects on the toxicity of chemicals. For example, hexobarbital and aminopyrine are detoxified by these enzymes and are thus more toxic to rats and mice with these nutrient deficiencies. On the other hand, the toxicity of aflatoxin is lower in such animals because of their depressed bioactivation of this toxicant. MFO activities are decreased in animals fed high levels of sugar. 

Storage of vinyl chloride is limited by the rapid metabolism and subsequent excretion. Vinyl chloride is biotransformed by cytochrome P450-mixed function oxidase systems (CYP 2E1), with the two primary metabolites being chloroethylene oxide and chloroacetaldehyde. These materials are further converted to chloroethanol and monochloroacetic acid. Metabolites are primarily excreted in urine. When rats were exposed to vinyl chloride at 100 ppm for 5 h, 70% of the absorbed dose was excreted as urinary metabolites within 24 h. The half-life for urinary excretion in rats was 4h. With an increase in dose via either inhalation or ingestion, the proportion exhaled increased and urinary and fecal elimination decreased. 

Biotransformation Enzyme induction Glutathione S Transferases Mixed Function Oxidases Hydrolases DNA Repair Enzymes... 

The examples provided above give only a brief overview of the variety of enzymatic functions that alter, biotransform, and biodegrade xenobiotics. In many instances numerous enzymes are known, as in the case of the mixed function oxidases. In order to provide a concrete example of a system of detoxification enzymes that is widely distributed, we have chosen the organophosphate acid anhydrolases — enzymes that may aid in the understanding of organophosphate intoxication and may also provide a means for the detoxification and bioremediation of these materials. 

In silico estimation of metabolism is still an area of intense study and development. Accurate prediction of intrinsic clearance is still not possible with the currently available methods [15]. Most of the progress in this area has been focused on the mixed function oxidase cytochrome P450 enzyme family. Advances in this area have been focused on three areas (1) prediction of the cytochrome P450 (CYP) enzyme isotype that is responsible for the major metabolism, (2) prediction of the chemical site of a molecule that is most likely to undergo biotransformation by oxidative metabolism, and (3) structure-based docking studies of CYP enzyme substrate complexes. ... 

Further biotransformations of A" VPA involve both the liver microsomal CYP enzymes and the fatty acid (3-oxidation pathway (Figure 33.29). The mixed-function-oxidase system metabolizes the unsaturated metabolite to a -butyrolactone derivative through a chemically reactive entity that is a mechanism-based inhibitor of CYP. The alkylation of the prosthetic heme by means of the radical occurs prior to formation of the epoxide. Thus, the epoxide is not involved in the CYP inhibition. 

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