XENOBIOTIC METABOLISM

1

Background

Increasingly, humans are subjected to exposure to various foreign

chemicals (xenobiotics)— drugs, food additives, pollutants, etc.

A xenobiotic (Gk xenos “stranger”) is a compound that is

foreign to the body.

2

The principal classes of xenobiotics of

medical relevance

-Drugs

-Chemical carcinogens

-Various compounds that have found their way into our

environment by one route or another, such as

polychlorinated biphenyls (PCBs) and certain insecticides.

3

Xenobiotic metabolism

Metabolism of xenobiotics include two phases:

A-Phase 1, e.g, hydroxylation.

B-Phase 2, conjugation (e.g. glucronic acid).

A-Phase 1, the major reaction involved is hydroxylation, catalyzed

by members of a class of enzymes referred to as monooxygenases

or cytochrome P450s isoforms .

The other reaction include reduction and hydrolysis, which

perform by same enzymes species.

4

B-Phase 2, the hydroxylated or other compounds produced in

phase 1 are converted by specific enzymes to various polar

metabolites by conjugation with glucuronic acid, sulfate, acetate,

glutathione, or certain amino acids, or by methylation.

-The overall purpose of the two phases of metabolism of

xenobiotics is to increase their water solubility (polarity) and thus

excretion from the body.

-Very hydrophobic xenobiotics would persist in adipose tissue

almost indefinitely if they were not converted to more polar forms.

5

Hydroxylation reactionHydroxylation is the chief reaction involved in phase 1. The

responsible enzymes are called monooxygenases or cytochrome

P450s.

Example of the reaction:

RH + O2 +NADPH +H+ →R-OH +H2O +NADP

(RH) represent a very wide variety of xenobiotics, including

drugs, carcinogens, pesticides, petroleum products, and

pollutants (such as a mixture of PCBs), as well as endogenous

compounds, such as certain steroids, eicosanoids, fatty acids,

and retinoids.

-In the above reaction one atom of oxygen enters R-OH and

one atom enters water.

6

-This dual fate of the oxygen accounts for the former naming of

monooxygenases as (mixed function oxidases).

The reaction catalyzed by cytochrome P450 can also be

represented as follows:

Reduced cytochrome P450 Oxidized cytochrome P450

RH + O2→R — OH + H2O

The major monooxygenases in the endoplasmic reticulum

are cytochrome P450s (CYP450)—so named because the enzyme

was discovered when it was noted that preparations of

microsomes that had been chemically reduced and then exposed

to carbon monoxide exhibited a distinct peak at 450 nm.

7

-This enzyme is important is the fact that approximately 50% of

the drugs humans ingest are metabolized by isoforms of

cytochrome P450.

Important points concerning cytochrome P450s.

(1) Like hemoglobin, they are hemoproteins.

(2) They are widely distributed across species.

(3) They are present in highest amount in liver and adrenal.

In liver, in the membranes of the smooth endoplasmic reticulum,

which constitute part of the microsomal fraction, In the adrenal,

in mitochondria as well as in the endoplasmic reticulum.

8

(4) At least six isoforms of cytochrome P450 are present.

(5) NADPH, not NADH, is involved in the reaction mechanism

of cytochrome P450.

(6) Lipids are also components of the cytochrome P450

system..The preferred lipid is phosphatidylcholine,

9

(7) Most isoforms of cytochrome P450 are inducible. For

instance, the administration of phenobarbital or many other

drugs causes three to four fold increase in the amount of

cytochrome P450.

(8) Certain isoforms of cytochrome P450 (e.g, CYP1A1) are

particularly involved in the metabolism of polycyclic aromatic

hydrocarbons (PAHs).

(9) Certain cytochrome P450s exist in polymorphic forms (genetic

isoforms), some of which exhibit low catalytic activity.

10

CONJUGATION REACTIONS

-In phase 1 reactions; xenobiotics are generally converted to more

polar, hydroxylated derivatives.

-In phase 2 reactions; these derivatives are conjugated with

molecules such as glucuronic acid, sulfate, or glutathione.

This renders them even more water-soluble, and they are eventually

excreted in the urine or bile.

11

Five Types of Phase 2 Reactions Are Described

A. GLUCURONIDATION:

The glucuronidation of bilirubin is similar reactions whereby

xenobiotics are glucuronidated.

UDP-glucuronic acid glucuronyl donor (e.g phenol, benzoic

acid).

B. SULFATION:

Some alcohols, arylamines, and phenols are sulfated. The sulfate

donor in these and other biologic sulfation reactions (eg, sulfation

of steroids, glycosaminoglycans glycolipids, and glycoproteins) is

adenosine 3-phosphate- 5-phosphosulfate (PAPS)

12

C. CONJUGATION WITH GLUTATHIONE

Glutathione (γ-glutamyl-cysteinylglycine) is a tripeptide

consisting of glutamic acid, cysteine, and glycine. Glutathione is

commonly abbreviated GSH (because of the sulfhydryl group of

its cysteine, which is the business part of the molecule).

A number of potentially toxic electrophilic xenobiotics (such as

certain carcinogens) are conjugated to the nucleophilic

GSH in reactions that can be as follows:

R + GSH→R—S—G

where R = an electrophilic xenobiotic. The enzymes catalyzing

these reactions are called glutathione S transferases.

13

Some common notes about GHS.

1-If the potentially toxic xenobiotics were not conjugated to GSH,

they would be free to combine covalently with DNA, RNA, or cell

protein and could thus lead to serious cell damage.

2-GSH is therefore an important defense mechanism against

certain toxic compounds, such as some drugs and carcinogens.

3-If the levels of GSH in a tissue such as liver are lowered

then that tissue can be shown to be more susceptible to injury by

various chemicals that would normally be conjugated to GSH.14

**Glutathione has other important functions in human

cells apart from its role in xenobiotic metabolism.

1. It participates in the decomposition of potentially toxic hydrogen

peroxide in the reaction catalyzed by glutathione peroxidase.

2-It is an important intracellular reductant, helping to maintain

essential SH groups of enzymes in their reduced state.

3-Metabolic cycle involving GSH as a carrier has been implicated in

the transport of certain amino acids across membranes in the

kidney.

Amino acid+ GSH → γ-Glutamyl amino acid + Cysteinylglycine

15

4. Acetylation:

Acetylation is represented by

X + Acetyl - CoA→Acetyl -X + CoA

where X represents a xenobiotic. As for other acetylation

reactions, acetyl-CoA (active acetate) is the acetyl donor. These

reactions are catalyzed by acetyltransferases.

e.g. The drug isoniazid, used in the treatment of tuberculosis, is

subject to acetylation

5. Methylation:

A few xenobiotics are subject to methylation by methyltransferases,

employing S-adenosylmethionine as methyl donor.

16

** THE ACTIVITIES OF XENOBIOTIC

METABOLIZING ENZYMES ARE AFFECTED BY

AGE, SEX, & OTHER FACTORS.

There are significant differences in enzyme activities among

individuals, many of which appear to be due to genetic factors. The

activities of some of these enzymes vary according to age and sex.

17

**RESPONSES TO XENOBIOTICS INCLUDE

PHARMACOLOGIC, TOXIC, IMMUNOLOGIC,

& CARCINOGENIC EFFECTS

18

-When the xenobiotic is a drug, phase 1 reactions may produce its

active form or may diminish or terminate its action if it is

pharmacologically active in the body without prior metabolism.

-Certain xenobiotics are very toxic even at low levels

(e.g, cyanide). On the other hand, there are few xenobiotics,

including drugs, that do not exert some toxic effects

if sufficient amounts are administered.

19

1-The first is cell injury (cytotoxicity), which can be severe enough

to result in cell death.

-There are many mechanisms by which xenobiotics injure cells.

The one considered here is covalent binding to cell macromolecules

of reactive species of xenobiotics produced by metabolism. These

macromolecular targets include DNA, RNA, and protein.

20

2-Second, the reactive species of a xenobiotic may bind to a

protein, altering its antigenicity. The xenobiotic is said to act as a

hapten, i.e, a small molecule that by itself does not stimulate

antibody synthesis but will combine with antibody once formed.

The resulting antibodies can then damage the cell by several

immunologic mechanisms that grossly perturb normal cellular

biochemical processes.

21

3-Third, reactions of activated species of chemical carcinogens

with DNA are thought to be of great importance in chemical

carcinogenesis. Some chemicals (eg, benzo[α]pyrene) require

activation by monooxygenases in the endoplasmic reticulum to

become carcinogenic (they are thus called indirect carcinogens).

22

-The enzyme epoxide hydrolase is of interest because it can exert a

protective effect against certain carcinogens. The products of the

action of certain monooxygenases on some procarcinogen substrates

are epoxides.

-Epoxides are highly reactive and mutagenic or carcinogenic or

both.

23

-Epoxide hydrolase—like cytochrome P450, also present in the

membranes of the endoplasmic reticulum—acts on these

compounds, converting them into much less reactive dihydrodiols.

The reaction catalyzed by epoxide hydrolase can be represented as

follows:

24

25