Xenobiotics basis

1. DETOXIFICATION MECHANISM
BCH - ANA
By
Miriam Watafua
1

2. OUTLINE
2
 Introduction and definition of xenobiotics and
detoxification
 Classification and examples of xenobiotics
 Route of entry, absorption and excretion
 Detoxification mechanism:
- Phases 1 and 2

3. Introduction and Definition of
Xenobiotics
3
 Xenobiotics: (Greek: xenos - stranger, foreigner) a
compound that is foreign to the body system.
 Man is continuously exposed to several foreign
compounds such as drugs, pollutants, food
additives, cosmetics, pesticides e.t.c.
 Certain unwanted compounds are produced in the
large intestine by bacteria which enter the
circulation.
These include:
- indole from tryptophan,
- cadaverine from lysine,
- tyramine from tyrosine,

4. INTRO.. CONT….
4
 In the normal metabolism of the body, certain
waste compounds (e.g. bilirubin) are formed.
 A vast majority of the foreign compounds or
the unwanted substances produced in the
body, are toxic and, therefore, they should be
quickly eliminated from the body.

5. Definition of Detoxification
5
 The term detoxication or detoxificafion refers
to the series of biochemical reactions
occurring in the body to convert the foreign
(often toxic) compounds to non-toxic or less
toxic, and more easily excretable forms.
 Detoxification - a misnomer? Detoxification is
rather misleading, since sometimes a
“detoxified” product is more toxic than the
original substance (e.g. procarcinogens to
carcinogens).

6. Detoxification Cont….
6
 lt appears that the body tries to get rid of a
foreign substance by converting it into a more
soluble (often polar), and easily excretable
compound, but this may be sometimes
associated with increase in toxicity (e.g.
conversion of methanol to formaldehyde).
 So therefore, in recent years the term
detoxification is replaced by biotransformation
or metabolism of xenobiotics or simply
metabolism of foreign compounds.

7. Classification of Xenobiotics
7
1. Environmental pollutants/chemicals
2. Natural toxins
3. Food additives
4. Drugs
5. Cosmetics
6. Heavy metals

8. Classification Cont…
8
1. Environmental Pollutants:
 The main sources of pollution are from industrial
processes and the deliberate release into the
environment of substances such as pesticides,
insecticides, herbicides, gaseous and water
wastes from industries and man.
 Pesticides are deliberately sprayed onto crops or
agricultural land with the potential for exposure
either via water or air.
 With pesticides and other such pollutants, the
major problem is persistence in the environment
and increase in concentration during passage

9. Environmental pollutants cont...
9
 Pollutants derived from burning fossil fuel,
radio nuclides from fallouts or emissions of
industrial processing (toxic trace elements,
polycyclic aromatic carbons);
 packaging materials and other frequently used
products (such as polychlorinated biphenyls,
washing agents and disinfectants) are all
environmentally implicated xenobiotics.
 Radioactive compounds and rays are also
being exposed to in various fields of work from
the environment

10. Classification cont...
10
2. Natural Toxins:
 Many plants and animals produce toxic
substances for both defensive and offensive
purposes.
 Natural toxins of animals(snakes, frogs,
scorpions), plants, mould and bacterial origin
(toxic metabolites from microorganisms -
enterotoxin, mycotoxins) cause a variety of
toxic effects and are a significant cause of
human poisoning by accidental ingestion or
touch of poisonous plants or animals, and by
insect or animal sting/bite.

11. Natural toxins Cont...
11
 Mycotoxins - Most mycotoxins are from the
genera Aspergillus spp., and the aflatoxins
they produce during growth. Exists as a
mixture of Aflatoxin B1, B2, and G1 and G2
 Also the most common and highly toxic
fungal toxins, eg. aflatoxin B1 - the most
powerful carcinogen known.
 Cyanogenic glycosides – occur in large
quantity in cassava.

12. Classification cont...
12
3. Food Additives:
 This category of xenobiotic is directly ingested.
 However, food additives are usually of low
biological activity. toxic food contaminants might
also be formed within the food itself or within the
human digestive tract by reactions of some food
ingredients and additives (eg. nitrosamines).
 Many additives are added to food to alter the
flavor or colour, prevent spoilage, or in some
other way change the nature of the food stuffs
 Eg Processing aids (EDTA, gum ghatti), favour
and taste modificators (eg. saccharin, vanillin),
colour modificators (sunset yellow, tartrazine),
preservatives (Ascorbic acid, Sodium nitrite) etc.

13. Classification cont...
13
4. Drugs: Drugs are also classified as
xenobiotics because they are foreign to the
body system. Drugs like the antibiotics,
tranquilizers, sedatives etc must undergo
metabolism to either be activated or
eliminated.

14. Classification cont...
14
5. Cosmetics : Nowadays cosmetics are versatile
and used on almost daily basis by both men and
women. Most of the ingredients contained in
these cosmetics are considered xenobiotics. Eg.
the traditional Kohl used in making eye makeup
contain the heavy metal lead which after
prolonged exposure accumulates in the body
system (the brain) and poise some deleterious
effect to the body system especially to infants.
Others include the talc used in making powders,
black henna, body cream and moisturizers,
shampoos and conditioners, fragrance etc

15. Classification cont...
15
6. Heavy metals: People working in mining
industries have a high tendency of being
exposed to heavy metals like arsenic, lead,
mercury, cadmium etc through inhalation or
dermal absorbtion of these metals. Some
environments located close to high
concentration of these metals also get in
contact with them through drinking water and
diet. These metals have a possibility of causing
harmful effects thereby causing deleterious
effects eg lead poisoning when they are
accummulated in the body over a period of
time without being eliminated.

16. 16
Absorption of Foreign Compounds
 Foreign compounds can be taken into the
body from the environment by the oral,
inhalation, intravenous or intradermal
routes and are circulated in the blood.
 However, absorption of foreign compound
has to do with crossing of one or more semi
permeable membranes of the body system.

17. Excretion of Foreign Compounds
17
 Foreign compounds and their metabolites need to
be removed from the body via excretion and this
usually occurs via the kidney (urine), skin (sweat),
in faeces, breast milk, saliva or via expired air.
 This excretion is needed so that adverse effects of
their accumulation are avoided.
 Biliary excretion is initiated in the liver and the
products pass through the gut until they are finally
excreted along with the waste products of the
faeces eg glucuronides.

18. XENOBIOTIC BIOTRANSFORMATION
18
 Biotransformation is the process whereby xenobiotic are
metabolized in the tissues with the subsequent result of
being excreted from the body.
 The bulk of biotransformation activities take place in the
liver.
 There are basically three processes that occur during
biotransformation.
 Activation
 Detoxification
 Modification
 All of the processes are intended at making the foreign
compound more hydrophilic and therefore less toxic but
more readily excretable.

19. 19
 Activation enhances the effectiveness of certain
drugs/compound and at the same time it
enhances the capacity of such compounds to
interact with cellular macromolecules.
e.g Prontosil Sulphanilamide ; Inactive
Active
 Detoxification is the process of making the
compounds less toxic thereby preventing them
from causing cellular damage.
 Modification entails certain changes in the
compound for easy transportation and excretion.

20. Site of Drug/xenobiotic metabolism
20
 The detoxification reactions are carried out
mainly in the liver which is equipped with
the enzyme machinery eg the cytochrome
p450 enzyme system.
 However, kidney, GIT and other organs
may sometimes be involved.
 Intracellular site – Smooth endoplasmic
reticulum (most important). Others are
cytosol and mitochondrion

21. METABOLISM OF XENOBIOTICS
21

22. 22
 Phase 1: The reactions of Phase 1 are
oxidation, reduction and hydrolysis.
 Phase 2: These are the conjugation
reactions, involving compounds such as
glucuronic acid, amino acids (glycine),
glutathione, sulfate, acetate and methyl
group.
 Generally, metabolism of a compound
involves phase 1 as well as phase 2
reactions. For instance, oxidation followed
by conjugation is the most frequent process
in the metabolism of xenobiotics.

23. 23

24. 24
 Oxidation:
 A large number of foreign substances are
detoxified by oxidation.
 These include alcohols, aldehydes, amines,
aromatic hydrocarbons and sulfur
compounds.
 in general, aliphatic compounds are more
easily oxidized than aromatic ones.
 Alcohols: Aliphatic and aromatic alcohols
undergo oxidation to form the corresponding
acids.
CH3OH HCOOH
Methanol Formic acid

25. 25

26. 26

27. 27

28. 28
 Sulphur compounds: Organic sulphur is
oxidized to sulphuric acid.
 Drugs : Meprobamate, a tranquilizer is
oxidized to hydroxymeprobamate and
excreted via the urine.

29. Role of Cytochrome P450
29
 Most of the oxidation reactions of metabolism are
catalyzed by monooxygenase or cytochrome P450
enzyme system
 This enzyme system, also called mixed function
oxidase enzyme, is associated with the
microsomes.
 The usage P450 refers to the absorption peak (at
450 nm), exhibited by the enzyme when exposed
to carbon monoxide.
 Most of the reactions of cytochrome P450 involve
the addition of a hydroxyl group to aliphatic or
aromatic compounds which may be represented
as
RH+ O+ NADPH ROH+ H2O+ NADP+

30. Salient features of Cytochrome P450
30
1.Multiple forms of cytochrome P450 are believed
to exist, ranging from 20-200.
At least 6 species have been isolated and
worked in detail.
2.They are all hemoproteins, containing heme
as the prosthetic group.
3.Cytochrome P450 species are found in the
highest concentration in the microsomes of
liver.
In the adrenal gland, they occur in
mitochondria.
4.The mechanism of action of cytochrome P450
is complex and is dependent on NADPH.

31. 31
5. The phospholipid-phosphatidylcholine
constituent of cytochrome P450 system is
necessary for the action of this enzyme.
6. Cytochrome P450 is an inducible enzyme. Its
synthesis is increased by the administration
of drugs such as phenobarbitol.
7. A distinct specie namely cytochrome P448
(with absorption peak at 448 nm) has been
studied.
lt is specific for the metabolism of polycyclic
aromatic hydrocarbons, hence it is also
known as aromatic hydrocarbon hydroxylase.

32. 32

33. Hydrolysis
33
 The hydrolysis of the bonds such as ester,
glycoside and amide is important in the
metabolism of xenobiotics.
 Several compounds undergo hydrolysis
during the course of their metabolism.
 These include aspirin, atropine,
acetanilide, diisopropylfluorophosphate
and procaine

34. 34

35. PHASE 2: Conjugation
35
 Several xenobiotics undergo metabolism by
conjugation to produce less toxic and/or more
easily excretable compounds.
 Conjugation is the process in which a foreign
compound combines with a substance produced
in the body.
 The process of conjugation may occur either
directly or after the phase I reactions.
 At least 8 different conjugation agents have been
identified in the body.
 These are glucuronic acid, glycine, cysteine (of
glutathione), glutamine, methyl group, sulfate,
acetic acid and thiosulfate.

36. 36
 Glucuronic acid :
 Conjugation with glucuronic acid is the most
common.
 The active form of glucuronic acid is UDP-
glucuronic acid produced in the uronic acid
pathway.
 The microsomal enzyme UDP-
glucuronyltransferase participates in
glucuronide formation.
 A general reaction of glucuronide
conjugation is shown below (X-OH
represents xenobiotic).

37. 37

38. 38
 Certain drugs (e.g. barbiturates) when
administered induce glucuronyltransferase
and this increases the glucuronide
formation.
 Glucuronic acid conjugation may occur
with compounds containing hydroxyl,
carbonyl, sulfhydryl or amino groups.
 A few examples of glucuronide conjugation
are given below.

39. Glucuronide conj. contd
39

40. Conjugation with Glycine
40
 Glycine : Many aromatic carboxylic acids(e.g.
benzoic acid, phenylacetic acid) are conjugated
with glycine.
 Hippuric acid is formed when glycine is conjugated
with benzylCoA.
 The excretion of hippuric acid (Greek : hippos -
horse) was first reported in 1829 in the urine of
cows and horses.

41. Conjugation with Glycine cont...
41

42. Conjugation with glycine cont…
42
 Phenylacaetic acid + Glycine
Phenylaceturic acid
 Cholic acid + Glycine Glycocholic acid

43. Conjugation with Sulphate
43
 Sulphate : The active form of sulphate-3'-
phosphoadenosine 5- phosphosulphate
(PAPS) participates in conjugation
reactions and the enzyme sulfotransferase
is involved in this process.
 Several aliphatic and aromatic compounds
undergo sulfation.

44. Conjugation with sulphate cont...
44

45. Conjugation with acetylCoA
45

46. Conjugation with Glutathione
46
 Glutathione : The tripeptide glutathione
(Glu-Cys-Gly) is the active conjugating
agent.
 A wide range of organic compounds such
as alkyl or aryl halides, alkenes, nitro
compounds and epoxides get conjugated
with cysteine of glutathione.
 The formation of mercapturic acid is an
example of conjugation with glutathione.

47. Conjugation with glutathione
cont..
47

48. Conjugation with Methyl group
48
 The methyl group (-CH3) of S-
adenosylmethionine is frequently used to
methylate certain xenobiotics.
 This is catalysed by the enzyme
methyltransferase.
S-Adenosylmethionine + X-OH S-Adenosylhomocysteine
+XO-CH3

49. Conjugation with thiosulfate
49
 Thiosulfate : The highly toxic cyanides are
conjugated with thiosulfate to form less toxic
thiocyanate.
Cyanide+ Sodium thiosulfate Thiocyanate +
Sodium sulfate

50. Activity of Metabolizing Enzymes:
50
 Most foreign compounds are metabolized before
their elimination/excretion.
 If the enzymes required for the conjugation of a
foreign compound (before excretion) are inactive,
the foreign compound will remain in the human
body (unless there are alternative routes).
 This can be exemplified by the conjugation of
bilirubin with glucuronic acid.
 If the glucuronyl transferase is defective, free
bilirubin crosses the capillary membrane into the
brain causing a kind of damage to the essential
organ.

51. Detoxification by Drugs
51
 lt may be surprising to know that some drugs are
administered to detoxify foreign substances.
 The toxic effects of certain metals such as arsenic,
mercury and cadminum could be overcome by
administering
 BAL (British antilewisite).
 This compound was developed during the World
War ll and was used as a detoxifying agent for
certain war poisons.
 The mechanism of action of BAL is not clearly
known. lt is believed that BAL readily combines with
metals and gets easily excreted into urine.

52. Clinical Relevance of Drug Metabolism
52
 The dose and the frequency of administration
required to achieve effective therapeutic blood
and tissue levels vary in different patients
because of individual differences in drug
distribution and rates of drug metabolism and
elimination.
 These differences are determined by genetic
factors and non genetic variables such as age,
sex, liver size, liver function, body temperature,
nutritional and environmental factors such as
concomitant exposure to inducers or inhibitors
of drug metabolism.

53. 53
 Individual Differences:
 The individual differences in metabolic rate depend
on the nature of the drug or compound itself.
 Thus, within the same population, steady state
plasma levels may reflect a 30-fold variation in the
metabolism of one drug and only a 2-fold variation
in the metabolism of another.
 Genetic Factors:
 Genetic factors that influence enzyme levels
account for some of these differences.
Succinylcholine, for example is metabolized only
half as rapidly in persons with genetically
determined defects in pseudocholinesterase as in
persons with normally functioning cholinesterase.

54. 54
 Diet and Environmental Factors:
 Diet and environmental factors also contribute to
individual variations in drug metabolism.
 Charcoal broiled foods and cruciferous vegetables are
known to induce CYP1A enzymes, whereas grape
fruit juice is known to inhibit CYP3A metabolism of
coadministered drug substrates.
 Cigarette smokers metabolize some drugs more
rapidly than non-smokers because of enzyme
induction.
 Industrial workers exposed to some pesticides
metabolize certain drugs more rapidly than non-
exposed individuals.
 Such differences make it difficult to determine
effective and safe doses of drugs that have narrow
therapeutic indices.

55. 55
 Age and Sex :
 Increased susceptibility to the pharmacologic or toxic
activity of drugs has been reported in very young and
very old patients as compared with young adults.
 Though this may reflect differences in absorption,
distribution and elimination, differences in drug
metabolism cannot be ruled out- a possibility
supported by studies in other mammalian species
indicating that drugs are metabolized at reduced rates
during the prepubertal period and senescence (aging).
 Slower metabolism could be due to reduced activity of
metabolic enzymes or reduced availability of essential
endogenous cofactors.
 Similar trends have been observed in humans but
incontrovertible evidence is yet to be obtained.

56. 56
 Sex dependent variations in drug metabolism
have been well documented in rats but not in
other rodents.
 Male rats metabolize drugs much faster than
mature female rats.
 These differences in drug metabolism have
been clearly associated with androgenic
hormones.
 A few clinical reports suggest that similar sex
dependent difference in drug metabolism also
exist in humans for ethanol, propranolol,
benzodiazepines, estrogens and salicylates.

57. 57
 Enzyme Inducers and Inhibitors
 Enzyme inducers increase the activity of microsomal
ezymes by de novo synthesis of new enzymes eg
phenobarbitone, barbiturates, alcohol, tobacco while
enzyme inhibitors decrease the activity of
microsomal enzymes by binding to the active site of
the enzyme eg disulfiram, isoniazid, cimetidine.
 Consider this for enzyme induction: if phenobarb is
added to warfarin regimen, the metabolism of
warfarin will accelerate within 2-3 days and the dose
of warfarin must be increased to achieve therapeutic
success. If phenobarb is withdrawn, the dose of
warfarin must be decreased since its metabolism
must have returned to normal and failure to do that
will result to fatal haemorrhage.

58. Diseases Affecting Drug Metabolism
 Acute or chronic diseases that affect liver architecture or
function markedly affect hepatic metabolism of some
drugs.
 Such conditions include fat accumulation, alcoholic
hepatitis, active and inactive alcoholic cirrhosis,
hemochromatosis, chronic active hepatitis, biliary
cirrhosis and acute viral or drug-induced hepatitis.
 Depending on their severity, the conditions impair
hepatic drug-metabolizing enzymes particularly
microsomal oxidases and thereby markedly affect drug
elimination.
 For example, the half-lives of chlordiazepoxide and
diazepam in patients with liver cirrhosis or acute viral
hepatitis are greatly increased, with a corresponding
58

59.  Liver cancer has been reported to impair hepatic drug
metabolism in humans.
 For example, aminopyrine metabolism is slower in
patients with malignant hepatic tumours than in normal
controls.
 These patients also exhibit markedly diminished
aminopyrine clearance.
 Studies of liver biopsy specimens from patients with
hepatocellular carcinoma also indicate impaired ability to
oxidatively metabolize drugs in vitro.
 This is associated with correspondingly reduced
cytochrome P450 content.
 Cardiac disease by limited blood flow to the liver may
impair disposition of those drugs whose metabolism is
flow limited. These drugs are so readily metabolized by
59

60.  Pulmonary disease may also affect drug
metabolism as indicated by the impaired hydrolysis
of procainamide and procaine in patients with
chronic respiratory insufficiency and increased half-
life of antipyrine in patients with lung cancer.
 Impairment of enzyme activity or defective
formation of enzymes associated with heavy metal
poisoning or porphyria also results in reduction of
hepatic drug metabolism.
 For example, lead poisoning has been shown to
increase the half-life of antipyrine in humans.
60

61. Biomedical/Clinical Concepts of Xenobiotics Metabolism
61
 Knowledge of the metabolism of xenobiotics is
essential for the understanding of toxicology,
pharmacology and drug addiction.
 The body possesses the capability to get rid of the
foreign substances by converting them into more
easily excreted forms.
 Metabolism is not necessarily associated with the
conversion of toxic into non-toxic compounds. For
instance, methanol is metabolized to a more toxic
formaldehyde.
 Metabolism primarily occurs in the liver through one
or more of the reactions, namely: oxidation,
reduction, hydrolysis and conjugation.

62. Summary
62
1.Detoxification deals with the series of
biochemical reactions occurring in the body
to convert the foreign (often toxic)
compounds to non-toxic or less toxic and
more easily excretable forms. Liver is the
major site of detoxification. ln recent years,
the term detoxification is replaced by
biotransformation or metabolism of
xenobiotics.
2.Metabolism reactions may be divided into
phase I (oxidation, reduction, hydrolysis) and
phase II reactions (conjugation). Oxidation is
a major process of detoxification, involving
the microsomal enzyme cytochrome P450
which is an inducible, NADPH dependent

63. Summary cont..
63
3. Conjugation is a process in which a foreign
compound combines with a substance
produced in the body. The process of
conjugation may occur either directly or after
phase I reactions. At least 8 different
conjugation agents have been identified in
the body: glucuronic acid, glycine, cysteine,
glutamine, methyl group, sulfate, acetic acid
and thiosulfate.