124577907 Toxicology Introduction

8/14/2019 124577907 Toxicology Introduction

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Ullmanns Industrial ToxicologyCopyright c 2005 WILEY-VCH Verlag GmbH & Co. KGaA, WeinheimISBN: 3-527-31247-1

Toxicology 3

Toxicology

Wolfgang Dekant, Institute of Toxicology, University of Wuerzburg, Germany

Spiridon Vamvakas, Institute of Toxicology, University of Wuerzburg, Germany

1. Introduction . . . . . . . . . . . . . . 6

1.1. Definition and Scope . . . . . . . . . 6

1.2. Fields . . . . . . . . . . . . . . . . . . . 6

1.3. History . . . . . . . . . . . . . . . . . 8

1.4. Information Resources . . . . . . . 9

1.5. Terminology of Toxic Effects . . . 11

1.6. Types of Toxic Effects . . . . . . . . 13

1.7. DoseResponse: a Fundamental

Issue in Toxicology . . . . . . . . . .

13

1.7.1. Graphics and Calculations . . . . . . 15

1.8. Dose-Response Relationships for

Cumulative Effects . . . . . . . . . . 18

1.9. Factors Influencing

DoseResponse . . . . . . . . . . . . 19

1.9.1. Routes of Exposure . . . . . . . . . . 19

1.9.2. Frequency of Exposure . . . . . . . . 20

1.9.3. Species-Specific Differences in

Toxicokinetics . . . . . . . . . . . . . 21

1.9.4. Miscellaneous Factors Influencing

the Magnitude of Toxic Responses .

22

1.10. Exposure to Mixtures . . . . . . . . 23

2. Absorption, Distribution,

Biotransformation and

Elimination of Xenobiotics . . . . 23

2.1. Disposition of Xenobiotics . . . . . 23

2.2. Absorption . . . . . . . . . . . . . . . 24

2.2.1. Membranes . . . . . . . . . . . . . . . 24

2.2.2. Penetration of Membranes by

Chemicals . . . . . . . . . . . . . . . . 25

2.2.3. Mechanisms of Transport of

Xenobiotics through Membranes . .

26

2.2.4. Absorption . . . . . . . . . . . . . . . 27

2.2.4.1. Dermal Absorption . . . . . . . . . . 27

2.2.4.2. Gastrointestinal Absorption . . . . . 30

2.2.4.3. Absorption of Xenobiotics by the

Respiratory System . . . . . . . . . . 31

2.3. Distribution of Xenobiotics by

Body Fluids . . . . . . . . . . . . . . 33

2.4. Storage of Xenobiotics in Organs

and Tissues . . . . . . . . . . . . . . . 36

2.5. Biotransformation . . . . . . . . . . 37

2.5.1. Phase-I and Phase-II Reactions . . .

37

2.5.2. Localization of the

Biotransformation Enzymes . . . . . 38

2.5.3. Role of Biotransformation in

Detoxication and Bioactivation . . . 38

2.5.4. Phase-I Enzymes and their

Reactions . . . . . . . . . . . . . . . . 39

2.5.4.1. Microsomal Monooxygenases:

Cytochrome P450 . . . . . . . . . . . 39

2.5.4.2. Microsomal Monooxygenases:

Flavin-Dependent Monooxygenases 41

2.5.4.3. Peroxidative Biotransformation:

Prostaglandin-synthase . . . . . . . . 42

2.5.4.4. Nonmicrosomal Oxidations . . . . . 44

2.5.4.5. Hydrolytic Enzymes in Phase-I

Biotransformation Reactions . . . . 44

2.5.5. Phase-II Biotransformation

Enzymes and their Reactions . . . . 45

2.5.5.1. UDP-Glucuronyl Transferases . . . 45

2.5.5.2. Sulfate Conjugation . . . . . . . . . . 46

2.5.5.3. Methyl Transferases . . . . . . . . . . 47

2.5.5.4. N-Acetyl Transferases . . . . . . . . 47

2.5.5.5. Amino Acid Conjugation . . . . . .

47

2.5.5.6. Glutathione Conjugation of

Xenobiotics and Mercapturic Acid

Excretion . . . . . . . . . . . . . . . . 48

2.5.6. Bioactivation of Xenobiotics . . . . 49

2.5.6.1. Formation of Stable but Toxic

Metabolites . . . . . . . . . . . . . . . 50

2.5.6.2. Biotransformation to Reactive

Electrophiles . . . . . . . . . . . . . . 50

2.5.6.3. Biotransformation of Xenobiotics to

Radicals . . . . . . . . . . . . . . . . . 52

2.5.6.4. Formation of Reactive Oxygen

Metabolites by Xenobiotics . . . . . 53

2.5.6.5. Detoxication and Interactions of

Reactive Metabolites with Cellular

Macromolecules . . . . . . . . . . . . 53

2.5.6.6. Interaction of Reactive

Intermediates with Cellular

Macromolecules . . . . . . . . . . . . 55

2.5.7. Factors Modifying

Biotransformation and Bioactivation 58

2.5.7.1. Host Factors Affecting

Biotransformation . . . . . . . . . . .

58

2.5.7.2. Chemical-Related Factors that

Influence Biotransformation . . . . . 62


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4 Toxicology

2.5.8. Elimination of Xenobiotics and their

Metabolites . . . . . . . . . . . . . . . 62

2.5.8.1. Renal Excretion . . . . . . . . . . . . 63

2.5.8.2. Hepatic Excretion . . . . . . . . . . . 64

2.5.8.3. Xenobiotic Elimination by the

Lungs . . . . . . . . . . . . . . . . . . . 65

2.6. Toxicokinetics . . . . . . . . . . . . .

65

2.6.1. Pharmacokinetic Models . . . . . . . 66

2.6.1.1. One-Compartment Model . . . . . . 66

2.6.1.2. Two-Compartment Model . . . . . . 67

2.6.2. Physiologically Based

Pharmacokinetic Models . . . . . . . 68

3. Mechanisms of Acute and Chronic

Toxicity and Mechanisms of

Chemical Carcinogenesis . . . . . . 69

3.1. Biochemical Basis of Toxicology . 69

3.2. Receptor-Ligand Interactions . .

703.2.1. Basic Interactions . . . . . . . . . . . 70

3.2.2. Interference with Excitable Mem-

brane Functions . . . . . . . . . . . . 72

3.2.3. Interference of Xenobiotics with

Oxygen Transport, Cellular Oxygen

Utilization, and Energy Production 73

3.3. Binding of Xenobiotics to

Biomolecules . . . . . . . . . . . . . . 74

3.3.1. Binding of Xenobiotics or their

Metabolites to Cellular Proteins . . 75

3.3.2. Interaction of Xenobiotics or theirMetabolites with Lipid Constituents 76

3.3.3. Interactions of Xenobiotics or their

Metabolites with nucleic Acids . . . 76

3.4. Perturbation of Calcium

Homeostasis by Xenobiotics

or their Metabolites . . . . . . . . . 77

3.5. Nonlethal Genetic Alterations in

Somatic Cells and Carcinogenesis 78

3.6. DNA Structure and Function . . . 79

3.6.1. DNA Structure . . . . . . . . . . . . . 79

3.6.2. Transcription . . . . . . . . . . . . . .

803.6.3. Translation . . . . . . . . . . . . . . . 80

3.6.4. Regulation of Gene Expression . . . 80

3.6.5. DNA Repair . . . . . . . . . . . . . . . 81

3.7. Molecular Mechanisms of

Malignant Transformation and

Tumor Formation . . . . . . . . . . 81

3.7.1. Mutations . . . . . . . . . . . . . . . . 81

3.7.2. Causal Link between Mutation

and Cancer . . . . . . . . . . . . . . . 83

3.7.3. Proto-Oncogenes and Tumor-

Suppressor Genes as Genetic Targets 83

3.7.4. Genotoxic versus Nongenotoxic

Mechanisms of Carcinogenesis . . . 84

3.8. Mechanisms of Chemically

Induced Reproductive and

Developmental Toxicity . . . . . . . 84

3.8.1. Embryotoxicity, Teratogenesis, and

Transplacental Carcinogenesis . . . 85

3.8.2. Patterns of DoseResponse in Ter-

atogenesis, Embryotoxicity, andEmbryolethality . . . . . . . . . . . . 86

4. Methods in Toxicology . . . . . . . 87

4.1. Toxicological Studies: General

Aspects . . . . . . . . . . . . . . . . . 87

4.2. Acute Toxicity . . . . . . . . . . . . . 90

4.2.1. Testing for Acute Toxicity by the

Oral Route: LD50 Test and Fixed-

Dose Method . . . . . . . . . . . . . . 90

4.2.2. Testing for Acute Skin Toxicity . . 92

4.2.3. Testing for Acute Toxicity by Inhala-

tion . . . . . . . . . . . . . . . . . . . .

944.3. Repeated-Dose Toxicity

Studies: Subacute, Subchronic

and Chronic Studies . . . . . . . . . 95

4.4. Ophtalmic Toxicity. . . . . . . . . . 96

4.5. Sensitization Testing . . . . . . . . . 97

4.6. Phototoxicity and

Photosensitization Testing . . . . . 99

4.7. Reproductive and Developmental

Toxicity Tests . . . . . . . . . . . . . 99

4.7.1. Fertility and General Reproductive

Performance . . . . . . . . . . . . . .

1004.7.2. Embryotoxicity and Teratogenicity 100

4.7.3. Peri- and Postnatal Toxicity . . . . . 101

4.7.4. Multigeneration Studies . . . . . . . 101

4.7.5. The Role of Maternal Toxicity in

Teratogenesis . . . . . . . . . . . . . . 102

4.7.6. In Vitro Tests for Developmental

Toxicity . . . . . . . . . . . . . . . . . 102

4.8. Bioassays to Determine the

Carcinogenicity of Chemicals

in Rodents . . . . . . . . . . . . . . . 103

4.9. In Vitro

and In Vivo

Short-termTests for Genotoxicity . . . . . . . . 105

4.9.1. Microbial Tests for Mutagenicity . . 106

4.9.1.1. The Ames Test for Bacterial Muta-

genicity . . . . . . . . . . . . . . . . . 106

4.9.1.2. Mutagenicity Tests in Escherichia

coli . . . . . . . . . . . . . . . . . . . . 111

4.9.1.3. Fungal Mutagenicity Tests . . . . . . 112

4.9.2. Eukaryotic Tests for Mutagenicity . 112

4.9.2.1. Mutation Tests inDrosophila

melanogaster . . . . . . . . . . . . . . 112

4.9.2.2. In Vitro Mutagenicity Tests in

Mammalian Cells . . . . . . . . . . .

112

4.9.3. In VivoMammalian Mutation Tests 114

4.9.3.1. Mouse Somatic Spot Test . . . . . . 114


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Toxicology 5

4.9.3.2. Mouse Specific Locus Test . . . . . 114

4.9.3.3. Dominant Lethal Test . . . . . . . . . 114

4.9.4. Test Systems Providing Indirect

Evidence for DNA Damage . . . . . 114

4.9.4.1. Unscheduled DNA Synthesis (UDS)

Assays . . . . . . . . . . . . . . . . . . 114

4.9.4.2. Sister-Chromatid Exchange Test . .

115

4.9.5. Tests for Chromosome Aberrations

(Cytogenetic Assays) . . . . . . . . . 116

4.9.5.1. Cytogenetic Damage and its

Consequences . . . . . . . . . . . . . 116

4.9.5.2. In Vitro Cytogenetic Assays . . . . . 117

4.9.5.3. In VivoCytogenetic Assays . . . . . 117

4.9.6. Malignant Transformation of

Mammalian Cells in Culture . . . . 118

4.9.7. In Vivo Carcinogenicity Studies of

Limited Duration . . . . . . . . . . . 119

4.9.7.1. Induction of Altered Foci in the

Rodent Liver . . . . . . . . . . . . . . 119

4.9.7.2. Induction of Lung Tumors in

Specific Sensitive Strains of Mice . 120

4.9.7.3. Induction of Skin Tumors in Specific

Sensitive Strains of Mice . . . . . . . 120

4.9.8. Methods to Assess Primary DNA

Damage . . . . . . . . . . . . . . . . . 120

4.9.8.1. Alkaline Elution Techniques . . . . 120

4.9.8.2. Methods to Detect and Quantify

DNA Modifications . . . . . . . . . . 121

4.9.9. Interpretation of Results Obtained in

Short-Term Tests . . . . . . . . . . . . 122

4.10. Evaluation of Toxic Effects on the

Immune System . . . . . . . . . . . . 123

4.11. Toxicological Evaluation of the

Nervous System . . . . . . . . . . . . 124

4.11.1. Functional Observational Battery . 124

4.11.2. Locomotor Activity . . . . . . . . . . 125

4.12. Effects on the Endocrine System .

1265. Evaluation of Toxic Effects . . . . 126

5.1. Acceptable risk, Comparison of

Risks, and Establishing

Acceptable Levels of Risk . . . . . 127

5.2. The Risk Assessment Process . . . 129

5.2.1. Hazard Identification Techniques . 129

5.2.2. Determination of Exposure . . . . . 131

5.2.3. Dose-Response Relationships . . . . 132

5.2.4. Risk Characterization . . . . . . . . . 133

5.2.4.1. The Safety-Factor Methodology . . 133

5.2.4.2. Risk Estimation Techniques forNonthreshold Effects . . . . . . . . . 135

5.2.4.3. Mathematical Models Used in High-

to Low-Dose Risk Extrapolation . . 136

5.2.4.4. Interpretation of Data from Chronic

Animal Bioassays . . . . . . . . . . . 137

5.2.4.5. Problems and Uncertainties in Risk

Assessment . . . . . . . . . . . . . . . 137

5.3. Future Contributions of

Scientifically Based Procedures to

Risk Assessment and Qualitative

Risk Assessment for Carcinogens 141

5.4. Risk Assessment for Teratogens .

145

6. References . . . . . . . . . . . . . . . 146

Abbreviations:

Ah-R arylhydrocarbon receptorAP apurinic/apyrimidinic siteAPS adenosine 5-phosphosulfateBHK baby hamster kidney

BIBRA British Industrial Biological Re-search Association

CoA Coenzym ADDT 1,1-(2,2,2-trichloro-

ethylidene)bis-(4-chlorobenzene)DHHS U.S. Department of Health and

Human ServicesDHP delayed hypersensitive responseECETOC European Chemical Industry

Ecology and Toxicology CentreED effective dose

ELISA enzyme-linked immunosorbentassay

FCA Freunds complete adjuvant

FAD flavine adenine dinucleotideGABA -aminobutyrateGC/MS gas chromatography/mass spec-

troscopyGOT glutamic acid oxalacetic transam-

inase

GSH glutathioneGSSG glutathione disulfideGST glutathioneS-transferaseGTP guanosine 5-triophosphateHGPRT hypoxanthine guanine phospho-

ribosyltransferaseIPCS International Programme on

Chemical SafetyLDH lactate dehydrogenaseLOAEL lowest-observed-adverse-effect

levelLOEL lowest-observed-effect levelMIF migration inhibition factor


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mRNA messenger RNAMTD maximum tolerated doseNADPH nicotinamide dinucleotide phos-

phate (H)NOEL no-observed-effect-levelNTP National Toxicology Program

PAPS 3-phosphoadenosine-5-phos-phosulfate

PG prostaglandinrRNA ribosomal RNASHE Syrian hamster embryoSMART somatic mutation and recombina-

tion testT, or TCDD 2,3,7,8-tetrachlorodibenzodioxinTD tumor doseTK thymidine kinasetRNA transfer RNAUDP uridine diphosphateUDPG uridine diphosphate glucoseUDPGA uridine diphosphate glucuronic

acidUDS unscheduled DNA synthesis

1. Introduction

1.1. Definition and Scope

Chemicals that are used or of potential use incommerce, thehome, theenvironment, and med-ical practice may present various types of harm-ful effects. The nature of these effects is deter-mined by the physicochemical characteristics ofthe agent, its ability to interact with biologicalsystems (hazard), and its potential to come intocontact with biological systems (exposure).

Toxicology studies the interaction betweenchemicals and biological systems to determinethe potential of chemicals to produce adverse ef-fects in living organisms. Toxicology also inves-tigates the nature, incidence, mechanisms of pro-duction, factors influencing their development,and reversibility of such adverse effects. Ad-verse effects are defined as detrimental to thesurvival or the normal functioning of the individ-ual. Inherent in this definition are the followingkey issues in toxicology:

1) Chemicals must come into close structural

and/or functional contact with tissues or or-gans to cause injury.

2) All adverse effects depend on the amount ofchemical in contact with the biological sys-tem (the dose) and the inherent toxicity ofthe chemical (hazard). When possible, theobserved toxic effect should be related tothe degree of exposure. The influence of dif-

ferent exposure doses on the magnitude andincidence of the toxic effect should be quan-titated. Such dose-response relationships areof prime importance in confirming a causalrelationship between chemical exposure andtoxic effect (for details, see Section 1.7).

Research in toxicology is mainly concernedwith determining the potential for adverse ef-fects caused by chemicals, both natural and syn-thetic, to assess their hazard and risk of human

exposure and thus provide a basis for appropri-ate precautionary, protective and restrictive mea-sures. Toxicological investigations should per-mit evaluation of the following characteristicsof toxicity:

1) The basic structural, functional, or biochem-ical injury produced

2) Dose-response relationships3) The mechanisms of toxicity (fundamental

biochemical alterations responsible for theinduction and maintenance of the toxic re-sponse) and reversibility of the toxic effect

4) Factors that modify response, e.g., route ofexposure, species, and gender

For chemicals to which humans may poten-tially be exposed, a critical analysis, based on thepattern of potential exposure or toxicity, may benecessary in order to determine the risk-benefitratio for their use in specific circumstances andto devise protective and precautionary measures.Indeed, with drugs, pesticides, food additives,

and cosmetic preparations, toxicology testingmust be performed in accordance with govern-ment regulations before use.

1.2. Fields

Toxicology is a recognized scientific disciplineencompassing both basic and applied issues. Al-though only generally accepted as a specific sci-entific field during this century, its principles

have been appreciated for centuries. The harm-ful or lethal effects of certain chemicals, mainlypresent in minerals and plants or transmitted


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Toxicology 7

venomous animals, have been known since pre-historic times. In many countries, toxicology asa discipline has developed from pharmacology.Pharmacology and toxicology both study the ef-fect of chemicals on living organisms and haveoften used identical methods. However, funda-

mental differences have developed. Years ago,only the dependence on dose of the studied ef-fects separated pharmacology and toxicology.Pharmacology focused on chemicals with bene-ficial effects (drugs) at lower doses whereas tox-icology studied the adverse health effects occur-ring with the same chemicals at high doses. To-day, the main interest of research in toxicologyhas shifted to studies on the long-term effectsof chemicals after low-dose exposure, such ascancer or other irreversible diseases; moreover,most chemicals of interest to toxicologists arenot used as drugs.

The variety of potential adverse effects andthe diversity of chemicals present in our environ-ment combine to make toxicology a very broadscience. Toxicology uses basic knowledge fromclinical and theoretical medicine and natural sci-ences such as biology and chemistry (Fig. 1).Because of this diversity, toxicologists usuallyspecialize in certain areas.

Any attempt to define the scope of toxicologymust take into account that the various subdisci-plines are not mutually exclusive and frequentlyare heavily interdependent. Due to the overlap-ping mechanisms of toxicity, chemical classes,and observed toxic effects, clear divisions intosubjects of equal importance are often not pos-sible.

The professional activities of toxicologistscan be divided into three main categories: de-scriptive, mechanistic, and regulatory. The de-

scriptive toxicologistis concerned directly withtoxicity testing. Descriptive toxicology still of-ten relies on the tools of pathology and clinicalchemistry, but since the 1970s more mechanism-based test systems have been included in toxic-ity testing [1]. The appropriate toxicity tests inexperimental animals yield information that isextrapolated to evaluate the risk posed by ex-posure to specific chemicals. The concern maybe limited to effects on humans (drugs, indus-trial chemicals in the workplace, or food addi-tives) or may encompass animals, plants, andother factors that might disturb the balance of

the ecosystem (industrial chemicals, pesticides,environmental pollutants).

The mechanistic toxicologist is concernedwith elucidating the mechanisms by whichchemicals exert their toxic effects on living or-ganisms. Such studies may result in the develop-

ment of sensitive predictive toxicity tests usefulin obtaining information for risk assessment (seeChap. 4). Mechanistic studies mayhelp in thede-velopment of chemicals that are safer to use orof more rational therapies for intoxications. Inaddition, an understanding of the mechanismsof toxic action also contributes to the knowl-edge of basic mechanisms in physiology, phar-macology, cell biology, and biochemistry. In-deed, toxic chemicals have been used with greatsuccess as mechanistic tools to elucidate mech-anisms of physiological regulation. Mechanis-tic toxicologists are often active in universities;however, industry and government institutionsare now undertaking more and more research inmechanistic toxicology.

Regulatory toxicologists have the responsi-bility of deciding on the basis of data providedby the descriptive toxicologist and the mecha-nistic toxicologist if a drug or chemical poses asufficiently low risk to be used for a stated pur-

pose. Regulatory toxicologists are often activein government institutions and are involved inthe establishment of standards for the amount ofchemicals permitted in ambient air in the envi-ronment, in the workplace, or in drinking water.Other divisions of toxicology may be based onthe classes of chemicals dealt with or applicationof knowledge from toxicology for a specific field(Table 1).

Forensic toxicology comprises both analyti-cal chemistry and fundamental toxicologic prin-

ciples. It is concerned with the legal aspectsof the harmful effects of chemicals on humans.The expertise of the forensic toxicologist is in-voked primarily to aid in establishing the causeof death and elucidating its circumstances in apostmortem investigation. The field ofclinicaltoxicology recognizes and treats poisoning, bothchronic and acute. Efforts are directed at treatingpatients poisoned by chemicals and at the devel-opment of new techniques to treat these intoxi-cations.Environmental toxicologyis a relativelynew area that studies the effects of chemicalsreleased by man on wildlife and the ecosystemand thus indirectly on human health.


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8 Toxicology

Figure 1. Scientific fields influencing the science of toxicology

Table 1.Areas of toxicology

Field Tasks and objectives

Forensic toxicology diagnoses poisoning by analytical

proceduresPesticide toxicology studies the safety of pesticides,

develops new pesticides

Occupational toxicology assesses potential adverse effects

of chemicals used in the

workplace, recommendsprotective procedures

Drug toxicology studies potential effects of drugs

after high doses, elucidates

mechanisms of sideeffects

Regulatory toxicology develops and interprets toxicity

testing programs and is involved

in controlling the use of chemicals

Environmental toxicology studies the effects of chemicals on

ecosystems and on humans afterlow-dose exposure from the

environment

Drug toxicology playsamajorroleinthepre-clinical safety assessment of chemicals intendedfor use as drugs. Drug toxicology also eluci-dates the mechanisms of side effects observedduring clinical application. Occupational toxi-cologystudies the acute and chronic toxicity ofchemicals encountered in the occupational en-

vironment. Both acute and chronic occupationalpoisonings have exerted a major influence on thedevelopment of toxicology in general. Occupa-

tional toxicology also helps in the developmentof safety procedures to prevent intoxications inthe workplace and assists in the definition of ex-

posure limits. Pesticide toxicologyis involved inthe development of new pesticides and the safetyof pesticide formulations. Pesticide toxicologyalso characterizes potential health risks to thegeneral population caused by pesticide residuesin food and drinking water.

1.3. History

Toxicology must rank as one of the oldest prac-

tical sciences because humans, from the verybeginning, needed to avoid the numerous toxicplants and animals in their environment. Thepresence of toxic agents in animals and plantswas known to the Egyptian and Greek civilisa-tions. The papyrus Ebers, an Egyptian papyrusdating from about 1 500 b.c., and the surviv-ing medical works of Hippocrates, Aristotle,and Theophrastus, published during the period400250b.c., all included some mention of poi-sons.

The Greek and Roman civilizations know-ingly used certain toxic chemicals and extractsfor hunting, warfare, suicide, and murder. Up


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Toxicology 9

to the Middle Ages, toxicology was restrictedto the use of toxic agents for murder. Poisoningwas developed to an art in medieval Italy and hasremained a problem ever since, and much of theearlier impetus for the development of toxicol-ogy was primarily forensic. There appear to have

been few advances in either medicine or toxicol-ogy between the time of Galen(131200 a.d.)and Paracelsus(14931541). The latter laid thegroundwork for the later development of moderntoxicology. He clearly was aware of the doseresponse relationship. His statement that Allsubstances are poisons; there is none that is nota poison. The right dose differentiates a poisonand a remedy, is properly regarded as a land-mark in the development of the science of tox-icology. His belief in the value of experimenta-tion also represents a break with much earliertradition. Important developments in the 1700sinclude the publication of RamazzinisDiseasesof Workers, which led to his recognition as thefather of occupational medicine. The correlationbetween the occupation of chimney sweepersand scrotal cancer by Pottin 1775 is also note-worthy.

Orfila, a Spaniard working at the Universityof Paris, clearly identified toxicology as a sepa-

rate science and wrote the first book devoted ex-clusively to it (1815). Workers of the later 1800swho produced treatises on toxicology includeChristison, Kobert, and Lewin. They increasedour knowledge of the chemistry of poisons, thetreatment of poisoning, the analysis of both xe-nobiotics and toxicity, as well as modes of actionand detoxication. A major impetus for toxicol-ogy in the 1900s was the use of chemicals forwarfare. In World War I, a variety of poisonouschemicals were used in thebattlefields of France.

This provided stimulus for work on mechanismsof toxicity as well as medical countermeasuresto poisoning. Since the 1960s, toxicology hasentered a phase of rapid development and haschanged from a science that was almost entirelydescriptive to one in which the study of mech-anisms has become the prime task. The manyreasons for this include the development of newanalytical methods since 1945, the emphasis ondrug testing following the thalidomide tragedy,the emphasis on pesticide testing following thepublication of Rachel Carsons Silent Spring andpublic concern over environmental pollution anddisposal of hazardous waste.

1.4. Information Resources

Because of the complexity of toxicology as a sci-ence and the impact of toxicological investiga-tions on legislation and commerce, a wide rangeof information on the toxic effects of chemi-

cals is available. No single, exhaustive sourceof toxicological data exists; several sources arerequired to obtain comprehensive informationon a particular chemical. Printed sources areoften quicker and easier to use than computerdata bases, but interactive online searching canrapidly gather important information from thehuge number of sources present.

The information explosion in toxicology hasresulted in a comprehensive volume dedicatedto toxicological information sources:

P. Wexler, P. J. Hakkinen, G. Kennedy, Jr.F. W.Stoss, Information Resources in Toxi-cology, 3rd ed., Academic Press, 1999.

Textbooks.The easiest way to obtain infor-mation on general topics in toxicology and sec-ondary references are a range of textbooks avail-able on the market. Only a few selected booksare listed below:

C. D. Klaasen, Casarett and Doulls Toxicol-

ogy; The Basic Science of Poisons, 6th ed.,McGraw-Hill, New York, 2001.G. D. Clayton, F. E. Clayton (eds): Pattys

Industrial Hygiene and Toxicology, Wiley,New York, 1993.J. G. Hardman, L. E. Limbird, Goodman andGilmans, The Pharmacological Basis of

Therapeutics, 10th ed., McGraw-Hill, NewYork, 2001.W. A. Hayes, Principles and Methods of Tox-icology, 3rd ed., Raven Press, New York,2001.E. Hodgson (Ed.):Textbook of Modern Tox-icology, 3rd ed., Wiley Interscience, 2004.T. A. Loomis, A. W. Hayes,Loomiss Essen-tials of Toxicology, 4th ed., Academic Press,San Diego, 1996.

The huge volume by N. I. Sax and R. J. Lewis,Dangerous Properties of Industrial Materials,7th ed., Wiley, New York, 1999, contains ba-sic toxicological data on a large selection of

chemicals (almost 20 000) and may serve as auseful guide to the literature for compounds notcovered in other publications.


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10 Toxicology

Monographs. The best summary informa-tion on toxicology is published in the formof series by governments and international or-ganizations. Most of these series are summa-rizing the results of toxicity studies on spe-cific chemicals. The selection of these chemi-

cals is mainly based on the extent of their usein industry (e.g. trichloroethene), their occur-rence as environmental contaminants (mercury)or their extraordinary toxicity (e.g. 2,3,7,8-tetra-chlorodibenzodioxin):

American Conference of Governmental In-dustrial Hygienists, Threshold Limit Valuesand Biological Exposure Indices (Cincin-nati, OH). Published annually.MAK-Begrundungen, VCH Publishers,

Weinheim, Federal Republic of Germany.This German series includes detailed infor-mation on the toxicity of chemicals on theGerman MAK list (ca. 150 reports are avail-able; the series is continuously expanded).

The Commission of the European Communi-ties publishes the Reports of the Scientific Com-mittee on Cosmetology and the Reports of theScientific Committee for Food.

The Environmental Protection Agency (EPA)

publishes a huge number of reports and toxico-logical profiles. They are indexed in EPA Pub-lications. A Quarterly Guide.

The European Chemical Industry Ecol-ogy and Toxicology Centre (ECETOC) issuesMonographs (more than 20 have been pub-lished) and Joint Assessments of CommodityChemicals.

The monographs of the International Agencyfor Research on Cancer are definitive evalua-tions of carcinogenic hazards. The Environ-

mental Health Criteria documents of the Inter-national Programme on Chemical Safety (IPCS)assess environmental and human health effectsof exposure to chemicals, andbiological or phys-ical agents. A related Health and Safety Guideseries give guidance on setting exposure limitsfor national chemical safety programs.

The National Institute for OccupationalSafety and Health (NIOSH), has published 50Current Intelligence Bulletins on health haz-ards of materials and processes at work.

The technical report series of the NationalToxicology Program (NTP) reports results oftheir carcinogenicity bioassays, which include

summaries of the toxicology of the chemicalsstudied. A status report indexes both studies thatare under way and those that have been pub-lished. The program also issues an Annual Re-view of Current DHHS [U.S. Department ofHealth and Human Services], DOE [U.S. De-

partment of Energy] and EPA Research relatedto toxicology.

A large number of internet-based resourcesare also available to collect information ontoxic effects of chemicals and methods for riskassessment. Some information sites containinglarge amounts of downloadable information arelisted below:

US Environmental Protection Agency(EPA), Integrated Risk Information System

(IRIS), http://www.epa.gov/iris/index.htmlUS Environmental ProtectionAgency (EPA), ECOTOX Database,http://www.epa.gov/ecotox/

Organisation for Economic Co-operationand Development (OECD), test guidelines,http://www.oecd.org

Agency for Toxic Substances andDisease Registry (ATSDR), toxi-cological profile information sheet

http://www.atsdr.cdc.gov/toxprofiles/

European Chemicals Bureau,http://ecb.jrc.it/

National Toxicology Programm,http://ntp-server.niehs.nih.gov/htdocs/liason/-Factsheets/FactsheetList.html

United Nations Environment Programm,Chemicals http://www.chem.unep.ch/

Journals Results of toxicological researchare published in more than 100 journals. Thoselisted below mainly publish research closely re-lated to toxicology, but articles of relevance mayalso be found in other biomedical journals:

Archives of Environmental Contamination

and Toxicology

Archives of Toxicology

Biochemical Pharmacology

Chemical Research in Toxicology

CRC Critical Reviews in Toxicology

Clinical ToxicologyDrug and Chemical Toxicology

Environmental Toxicology and Chemistry


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Toxicology 11

Table 2.Toxic effects of different chemicals categorized by time scale and general locus of action

Exposure Site Effect Chemical

Acute local lung edema chlorine gas

systemic liver damage carbon tetrachloride

narcosis halothane

Subchronic local sensitization toluene diisocyanate

systemic neurotoxicity hexaneChronic local bronchitis sulfur dioxide

nasal carcinoma formaldehyde

systemic bladder carcinoma 4-amino-biphenyl

kidney damage cadmium

Food and Chemical Toxicology

Fundamental and Applied Toxicology

Journal of the American College of Toxi-

cology

Journal of Analytical Toxicology

Journal of Applied ToxicologyJournal of Biochemical Toxicology

Journal of Toxicology and Environmental

Health

Neurotoxicology and Teratology

Pharmacology and Toxicology

Practical In Vitro Toxicology

Regulatory Toxicology and Pharmacology

Reproductive Toxicology

Toxicology

Toxicology and Applied PharmacologyToxicology and Industrial Health

Toxicology In Vitro

Toxicology Letters

Databases and Databanks. Electronicsources, such as computer data bases or CD-ROM are a fast and convenient way to obtainreferences on the toxicity of chemicals. Sinceon-line searching of commercial data basessuch as STN-International may be expensive,

CD-ROM-based systems are increasingly be-ing used. The major advantages are speed, theability to refine searches and format the results,and non-text search options, such as chemicalstructure searching on Beilstein and ChemicalAbstracts.

Useful information about actual research onthe toxicology of chemicals may be obtained bysearching Chemical Abstracts or Medline withthe appropriate keywords. Specific data bankscovering toxicology are the Registry of Toxic

Effects of Chemical Substances, which givessummary data, statistics, and structures; Toxline

(available in DIMDI) gives access to the litera-ture.

1.5. Terminology of Toxic Effects

Toxic effects may be divided according totimescale (acute and delayed), general locusof action (local, systemic, organ specific), orbasic mechanisms of toxicity (reversible ver-sus irreversible). Acute toxic effects are thosethat occur after brief exposure to a chemical.Acute toxic effects usually develop rapidly aftersingle or multiple administrations of a chemi-cal; however, acute exposure may also produce

delayed toxicity. For example, inhalation of alethal dose of HCN causes death in less thana minute, whereas lethal doses of 2,3,7,8-tetra-chlorodibenzodioxin will result in the deathof experimental animals after more than twoweeks. Chronic effects are those that appearafter repetitive exposure to a substance; manycompounds require several months of continu-ous exposure to produce adverse effects. Often,the chronic effects of chemicals are differentfrom those seen after acute exposure (Table 2).

For example, inhalation of chloroform for a shortperiod of time may cause anesthesia; long-terminhalation of much lower chloroform concentra-tions causes liver damage. Carcinogenic effectsof chemicals usually have a long latency period;tumors may be observed years (in rodents) oreven decades (in humans) after exposure.

Toxic effects of chemicals may also be classi-fied based on the type of interaction between thechemical and the organism. Toxic effects maybe caused by reversible and irreversible interac-

tions (Table 3). When reversible interactions areresponsible for toxic effects, the concentrationof the chemical present at the site of action is


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12 Toxicology

the only determinant of toxic outcome. Whenthe concentration of the xenobiotic is decreasedby excretion or biotransformation, a parallel de-crease of toxic effects is observed.

Table 3.Reversible and irreversible interactions of chemicals with

cellular macromolecules as a basis for toxic response

Mechanism Toxic response Example

Irreversible inhibition

of Esterase neurotoxicity tri-o-cresylphosphate

Covalent bindingto DNA

cancer dimethylnitrosamine

Reversible binding to

Hemoglobin oxygendeprivation in

tissues

carbon monoxide

Cholinesterase neurotoxicity carbamate pesticides

After complete excretion of the toxic agent,toxic effects are reduced to zero (see below).A classical example for reversible toxic effectsis carbon monoxide. Carbon monoxide binds tohemoglobin and, due to the formation of thestable hemoglobincarbon monoxide complex,binding of oxygen is blocked. As a result ofthe impaired oxygen transport in blood fromthe lung, tissue oxygen concentrations are re-duced and cells sensitive to oxygen deprivation

will die. The toxic effects of carbon monoxideare directly correlated with the extent of car-boxyhemoglobin in blood, the concentration ofwhich is dependent on the inhaled concentrationof carbon monoxide. After exhalation of carbonmonoxide and survival of the acute intoxication,no toxic effect remains (Fig. 2).

Figure 2. Reversible binding of carbon monoxide tohemoglobin and inhibition of oxygen transport

Irreversible toxic effects are often caused by

the covalent binding of toxic chemicals to bi-ological macromolecules. Under extreme con-

ditions, the modified macromolecule is not re-paired; after excretion of the toxic agent, the ef-fect persists. Further exposure to the toxic agentwill produce additive effects; many chemicalscarcinogens are believed to act through irre-versible changes (see Section 2.5.6).

Another distinction between types of effectsmay be made according to the general locusof action. Local toxicity occurs at the site offirst contact between the biological system andthe toxic agent. Local effects to the skin, therespiratory tract, or the alimentary tract maybe produced by skin contact with a corrosiveagent, by inhalation of irritant gases, or by in-gestion of tissue-damaging materials. This typeof toxic responses is usually restricted to the tis-sues with direct contact to the agent. However,life-threatening intoxications may occur if vi-tal organs like the lung are damaged. For exam-ple, inhaled phosgene damages the alveoli of thelung and causes lung edema. The massive dam-age to the lung results in the substantial mortalityobserved after phosgene intoxication.

The opposite to local effects aresystemic ef-fects. They are characterized by the absorptionof the chemical and distribution from the port ofentry to a distant site where toxic effects are pro-

duced. Except for highly reactive xenobiotics,which mainly act locally, most chemicals actsystemically. Many chemicals that produce sys-temic toxicity only cause damage to certain or-gans, tissues, or cell types within organs. Selec-tive damage to certain organs or tissues by sys-temically distributed chemicals is termed organ-or tissue-specific toxicity [2]; the organs dam-aged are referred to as target organs (Table 4).

Table 4.Organ-specific toxic effects induced by chemicals that are

distributed systemically in the organism

Chemical Species Target organ

Benzene humans bone marrow

Hexachlorobutadiene rodents damage to

proximal tubulesof the kidney

Paraquat rodents,

humans

lung

Tri-o-cresylphosphate humans nervous system

Cadmium humans kidney

1,2-Dibromo-3-chloropropane humans,

rodents

testes

Hexane rodents,

humans

nervous system

Anthracyclines humans heart


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Toxicology 13

Major target organs for toxic effects are thecentral nervous system and the circulatory sys-tem followed by the blood and hematopoieticsystem and visceral organs such as the liveror the kidney. For some chemicals, both localand systemic effects can be demonstrated; more-

over, chemicals producing marked local toxicitymay also cause systemic effects as secondary re-sponses to major disturbances in homeostasis ofthe organism.

1.6. Types of Toxic Effects

The spectrum of toxic effects of chemicals isbroad, and their magnitude and nature dependon many factors such as the physiocochemicalproperties of the chemical and its toxicokinetics,the conditions of exposure, and the presence ofadaptive and protective mechanisms. The latterfactors include physiological mechanisms suchas adaptive enzyme induction, DNA repair, andothers. Toxic effects may be transient, reversible,or irrversible; some are deleterious and othersare not. Toxic effects may take the form of tissuepathology, aberrant growth processes, or alteredbiochemical pathways. Some of the more fre-

quently encountered types of injury constitutinga toxic response are described in the following.Immune-mediated hypersensitivity reactions

by antigenic materials are toxic effects often in-volved in skin and lung injury by repeated con-tact to chemicals resulting in contact dermati-tis and asthma. Inflammation is a frequently ob-served local response to the application of irri-tant chemicals or may be a component of sys-temic injury. This response may be acute withirritant or tissue damaging materials or chronic

with repetitive exposure to irritants. Necrosis,that is, death of cells or tissues, may be the re-sult of various pathological processes resultingfrom biochemical interactions of xenobiotics, asdescribed in Chapter 3. The extent and patternsof necrosis may be different for different chem-icals, even in the same organ. Chemical tumori-genesis or carcinogenesis (induction of malig-nant tumors) is an effect often observed afterchronic application of chemicals. Due the longlatency period and the poor prognosis for indi-

viduals diagnosed with cancer, studies to pre-dict the potential tumorigenicity of chemicalshave developed into a major area of toxicolog-

ical research. Developmental and reproductivetoxicology are concerned with adverse effectson the ability to conceive, and with adverse ef-fects on the structural and functional integrity ofthe fetus. Chemicals may interfere with repro-duction through direct effects on reproductive

organs or indirectly by affecting their neural andendocrine control mechansims. Developmentaltoxicity deals with adverse effects on the con-ceptus through all stages of pregnancy. Damageto the fetus may result in embryo reabsorption,fetal death, or abortion. Nonlethal fetotoxicitymay be expressed as delayed maturation, de-creased birth weight, or structural malformation.The most sensitive period for the induction ofmalformation is during organogenesis; neurobe-havioral malformations may be induced duringlater stages of pregnancy.

1.7. DoseResponse: a Fundamental

Issue in Toxicology

Inprinciple,apoisonisachemicalthathasanad-verse effect on a living organism. However, thisis not a useful definition since toxic effects arerelated to dose. The definition of a poison thus

also involves quantitative biological aspects. Atsufficiently high doses, any chemical may betoxic. The importance of dose is clearly seenwith molecular oxygen or dietary metals. Oxy-gen at a concentration of 21% in the atmosphereis essential for life, but 100% oxygen at atmo-spheric pressure causes massive lung injury inrodents and often results in death. Some met-als such as iron, copper, and zinc are essentialnutrients. When they are present in insufficientamounts in the human diet, specific disease pat-

terns develop, but in high doses they can causefatal intoxications. Toxic compounds are not re-stricted to man-made chemicals, but also includemany naturally occurring chemicals. Indeed, theagent with the highest toxicity is a natural poisonfound in the bacterium Clostridium botulinum(LD50 0.01 /kg).

Therefore, all toxic effects are products of theamount of chemical to which the organism is ex-posed and the inherent toxicity of the chemical;they also depend on the sensitivity of the biolog-ical system.

The term dose is most frequently usedto characterize the total amount of material to


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14 Toxicology

which an organism is exposed; dose defines theamount of chemical given in relation to bodyweight. Dose is a more meaningful and com-parative indicator of exposure than the term ex-posure itself. Dose usually implies the exposuredose, the total amount of chemical administered

to an organism or incorporated into a test sys-tem. However, dose may not be directly propor-tional to the toxic effects since toxicity dependson the amount of chemical absorbed. Usually,dose correctly describes only the actual amountof chemical absorbed when the chemical is ad-ministered orally or by injection. Under thesecircumstances, the administered dose is identi-cal to the absorbed dose; other routes of appli-cation such as dermal application or inhalationdo not define the amount of agent absorbed.

Different chemicals have a wide spectrum ofdoses needed to induce toxic effects or death. Tocharacterize the acute toxicity of different chem-icals, LD50values are frequently used as a basisfor comparisons. Some LD50 values (rat) for arange of chemicals follow:

Ethanol 12 500

Sodium bicarbonate 4 220

Phenobarbital sodium 350

Paraquat 120

Aldrin 46

Sodium cyanide 6.4

Strychnine 5

1,2-Dibromoethane 0.4

Sodium fluoroacetate 0.2

2,3,7,8-Tetrachlorodibenzodioxin 0.01

Certain chemicals are very toxic and producedeath after administration of microgram doses,while others are tolerated without serious toxic-ity in gram doses. The above data clearly demon-strate that the toxicity of a specific chemical is re-

lated to dose. The dependence of the toxic effectsof a specific chemical on dose is termed doseresponse relationship. Before doseresponse re-lationships can be appropriately used, severalbasic assumptions must be considered. The firstis that the response is due to the chemical ad-ministered. It is usually assumed that the re-sponses observed were a result of the variousdoses of chemical administered. Under exper-imental conditions, the toxic response usuallyis correlated to the chemical administered, since

both exposure and effect arewell defined andcanbe quantified. However, it is not always apparentthat the response is the result of specific chem-

ical exposure. For example, an epidemiologicstudy might result in discovery of an associa-tion between a response (e.g., disease) and oneor more variables including the estimated doseof a chemical. The true doses to which individ-uals have been exposed are often estimates, and

the specificity of the response for that chemicalis doubtful.

Further major necessary assumptions in es-tablishing doseresponse relationships are:

A molecular site (often termed receptor)with which the chemical interacts to producethe response. Receptors are macromolecularcomponents of tissues with which a chemi-cal interacts and produces its characteristiceffect.

The production of a response and the degreeof the response are related to the concentra-tion of the agent at the receptor.

The concentration of the chemical at the re-ceptor is related to the dose administered.Since in most cases the concentration of anadministered chemical at the receptor cannotbe determined, the administered dose or theblood level of the chemical is used as an in-dicator for its concentration at the molecularsite.

A further prerequisite for using the doseresponse relationship is that the toxic responsecan be exactly measured. A great variety of cri-teria or end points of toxicity may be used. Theideal end point should be closely associated withthe molecular events resulting from exposureto the toxin and should be readily determined.However, although many end points are quan-titative and precise, they are often only indirectmeasures of toxicity. For example, changes in

enzyme levels in the blood can be indicativeof tissue damage. Patterns of alterations mayprovide insight into which organ or system isthe site of toxic effects. These measures usu-ally are not directly related to the mechanismof toxic action. The doseresponse relationshipcombines the characteristic of exposure and theinherent toxicity of the chemical. Since toxic re-sponses to a chemical are usually functions ofboth time and dose, in typical doseresponse re-lationships, the maximum effect observed dur-

ing the time of observation is plotted against thedose to give time-independent curves. The time-independent doseresponse relationship may be


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Toxicology 15

used to study doseresponse for both reversibleand irreversible toxic effects. However, in riskassessments that consider the induction of ir-reversible effects such as cancer, the time fac-tor plays a major role and has important influ-ences on the magnitude or likelihood of toxic

responses. Thus, for this type of mechanism oftoxic action, dosetimeresponse relationshipsare better descriptors of toxic effects.

The doseresponse relationship is the mostfundamental concept in toxicology. Indeed, anunderstanding of this relationship is essential forthe study of toxic chemicals.

From a practical point of view, there are twodifferent types of doseresponse relationships.Doseresponse relationships may be quantal (allor nothing responses such as death) or graded.The graded or variable response involves a con-tinual change in effect with increasing dose, forexample, enzyme inhibition or changes in phys-iological function such as heart rate. Gradedresponses may be determined in an individ-ual or in simple biochemical systems. For ex-ample, addition of increasing concentrationsof 2,3,7,8-tetrachlorodibenzodioxin to culturedmammalian cells results in an increase in theconcentration of a specific cytochrome P450 en-

zyme in the cells (for details of mechanisms, seeSection 2.5.4.1). The increase is clearly dose re-lated and spans a wide range (Fig. 3). An exam-ple for a graded toxic effect in an individual maybe inflammation caused by skin contact with anirritant material. Low doses cause slight irrita-tion; as the amount increases, irritation turns toinflammation and the severity of inflammationincreases.

Figure 3. Dose-dependent induction of cytochrome P4501A 1 protein in cultured liver cells treated with 2,3,7,8-tetrachlorodibenzodioxin [3]

In doseresponse studies in a population, aspecific endpoint is also identified and the doserequired to produce this end point is determinedfor each individual in the population. Both dose-dependent graded effects and quantal responses(death, induction of a tumor) may be investi-

gated. With increasing amount of a chemicalgiven to a group of animals, the magnitude ofthe effect and/or the number of animals affectedincrease. For example, if an irritant chemical isapplied to the skin, as the amount of the materialincreases, the numbers of animals affected andthe severity of inflammation increases. Quantalresponses such as death induced by a potentiallylethal chemical will also be dose-dependent. Thedose dependency of a quantal effect in a popula-tion is based on individual differences in the re-sponse to the toxic chemical. A specific amountof the potentially lethal xenobiotic given to agroup of animals may not kill all of them, butas the amount given increases, the proportion ofanimals killed increases.

Althought the distinctions between gradedand quantal doseresponse relationships are use-ful, the two types of responses are conceptuallyidentical. The ordinate in both cases is simplylabeled response, which may be the degree of

response in an individual, or the fraction of apopulation responding, and the abscissa is therange of administered doses.

1.7.1. Graphics and Calculations

Even with a genetically homogenous populationof animals of the same species and strain, theproportion of animals showing the effect willincrease with dose (Fig. 4A). When the num-

ber of animals responding is plotted versus thelogarithm of the dose, a typical sigmoid curvewith a log-normal distribution that is symmetri-cal about the midpoint, is obtained (Fig. 4B).

When plotted on a log-linear scale, the ob-tained normally distributed sigmoid curve ap-proaches a response of 0% as the dose is de-creased, and 100% as the dose is increased, buttheoretically never passes through 0 or 100%.Small proportions of the population at the right-and left-hand sides of the curve represent hypo-

susceptible and hypersusceptible members. Theslope of the dosereponse curve around the 50%value, the midpoint, gives an indication of the


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16 Toxicology

Figure 4. Typical dose response curves for a toxic effectPlots are linear linear (A); log linear (B); and log probit (C) for an identical set of data

ranges of doses producing an effect. A steepdoseresponse curve indicates that the major-

ity of the population will respond over a narrowdose range; a shallow doseresponse curve in-dicates that a wide range of doses is required toaffect the majority of the population. The curvedepicted in Fig. 4B shows that the majority ofthe individuals respond about the midpoint ofthe curve. This point is a convenient descriptionof the average response, and is referred to as themedian effective dose (ED50). If mortality is theendpoint, then this dose is referred as medianlethal dose (LD50).

Death, a quantal response, is simple to quan-tify and is thus an end point incorporated in manyacute toxicity studies. Lethal toxicity is usually

calculated initially from specific mortality levelsobtained after giving different doses of a chem-

ical; the 50% mortality level is used most fre-quently since it represents the midpoint of thedose range at which the majority of deaths oc-cur. This is the dose level that causes death ofhalf of the population dosed. The LD50 valuesare usually given in milligrams of chemical perkilogram of body weight (from the viewpointof chemistry and for comparison of relative po-tencies of different chemicals, giving the LD50in moles of chemical per kilogram body weightwould be desirable). After inhalation, the ref-

erence is to LC50 (LC = lethal concentration),which, in contrast to LD50 values, depends onthe time of exposure; thus, it is usually expressed


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Toxicology 17

as X-hour LC50 value. The LD50 or LC50 val-ues usually represent the initial information onthe toxicity of a chemical and must be regardedas a first, but not a quantitative, hazard indicatorthat may be useful for comparison of the acutetoxicity of different chemicals [3].

Similar doseeffect curves can, however, beconstructed for cancer, liver injury, and othertypes of toxic responses. For the determinationof LD50 values and for obtaining comparativeinformation on doseresponse curves, plottinglog dose versus percent response is not practi-cal since large numbers of animals are neededfor obtaining interpretable data. Moreover, otherimportant information on the toxicity of a chem-ical (e.g., LD05 and LD95) cannot be accu-rately determined due to the slope of sigmoidcurve. Therefore, the doseresponse curve istransformed to a log-probit (probit = probabilityunits) plot. The data in the Fig. 4B form astraight line when transformed into probit units(Fig. 4C). The EC50or, if death is the end point,the LD50 is obtained by drawing a horizontalline from the probit unit 5, which is the 50% re-sponse point, to the doseeffect line. At the pointof intersection a vertical line is drawn, and thisline intersects the abscissa at the LD50 point.

Information on the lethal dose for 90% or for10% of the population can also be derived by asimilar procedure. The confidence limits are nar-rowest at the midpoint of the line (LD50) and arewidest at the two extremes (LD05 and LD95) ofthe doseresponse curve. In addition to permit-ting determination of a numerical value for theLD50 of a chemical with few groups of dosedanimals, the slope of the doseresponse curvefor comparison between toxic effects of differ-ent chemicals is obtained by the probit transfor-

mation [4].The LD50 by itself, however, is an insuf-

ficient index of lethal toxicity, particulary ifcomparisons between different chemicals are tobe made. For this purpose, all available doseresponse information including the slope of thedoseresponse line should be used. Figure 5demonstrates the doseresponse curves for mor-tality for two chemicals.

The LD50 of both chemicals is the same(10 mg/kg). However, the slopes of the doseresponse curves are quite different. Chemical Aexhibits a flat doseresponse curve: a largechange in dose is required before a significant

change in response will be observed. In contrast,chemical B exhibits a steep doseresponsecurve, that is, a relatively small change in dosewill cause a large change in response. The chem-ical with the steep slope may affect a much largerproportion of the population by incremental in-

creases in dose than chemicals having a shallowslope; thus, acute overdosing may be a prob-lem affecting the majority of a population forchemicals with steeper slopes. Chemicals withshallower slopes may represent a problem forthe hyperreactive groups at the left-hand side ofthe doseresponse curve. Effects may occur atsignificantly lower dose levels then for hyperre-active groups exposed to chemicals with a steepdoseresponse.

While the LD50 values characterize the po-tential hazard of a chemical, the risk of an expo-sure is determined by the hazard multiplied bythe exposure dose. Thus, even very toxic chem-icals like the poison of Clostridium botulinumpose only a low risk; intoxications with thiscompound are rare since exposure is low. More-over, acute intoxications with other highly toxicagents such as mercury salts are rarely seen, de-spite detectable blood levels of mercury salts inthe general population, since the dose is also

low. On the other hand, compounds with lowtoxicity may pose a definite health risk whendoses are high, for example, constituents of dietor chemicals formed during food preparation byheat treatment.

Figure 5. Comparison of dose response relationships fortwo chemicals (log probit plot)Both chemicals have identical LD50 values, but differentslopes of the dose response curve


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18 Toxicology

Therefore, for characterizing the toxic risk ofa chemical, besides information on the toxicity,information on the conditions of exposure arenecessary. When using LD50values for toxicitycharacterisation, the limitations of LD50 valuesshould be explicitly noted. These limitations in-

clude methodological pitfalls influenced by

1) Strain of animal used2) Species of animal used3) Route of administration4) Animal housing

and intrinsic factors limiting the use of LD50values

1) Statistical method2) No doseresponse curve3) Time to toxic effect not determined4) No information on chronic toxicity

The most serious limitation on the use ofLD50 values for hazard characterization are thelack of information on chronic effects of a chem-ical and the lack of doseresponse information.Chemicals with low acute toxicity may have car-cinogenic or teratogenic effects at doses that donot induce acute toxic responses. Other limita-tions include insufficient information on toxic

effects other than lethality, the cause of death,and the time to toxic effect. Moreover, LD50values are not constant, but are influenced bymany factors and may differ by almost one or-der of magnitude when determined in differentlaboratories.

1.8. Dose-Response Relationships for

Cumulative Effects

After chronic exposure to a chemical, toxic re-sponse may be caused by doses not showingeffects after single dosing. Chronic toxic re-sponses are often based on accumulation of ei-ther the toxic effect or of the administered chem-ical. Accumulation of the administered chemi-cal is observed when the rate of elimination ofthe chemical is lower than the rate of adminis-tration. Since the rate of elimination is depen-dent on plasma concentrations, after long-term

application an equilibrium concentration of thechemical in the blood is reached. Chemicals mayalso be stored in fat (polychlorinated pesticides

such as DDT) or bone (e.g., lead). Stored chem-icals usually do not cause toxic effects becauseof their low concentrations at the site of toxicaction (receptor). After continuous application,the capacities of the storage tissues may becomesaturated, and xenobiotics may then be present

in higher concentration in plasma and thus at thesite of action; toxic responses result. Besides cu-mulation of the toxic agent, the toxic effect mayalso cumulate (Fig. 6).

Figure 6. Accumulation of toxic chemicals based on theirrate of excretiona) The rate of excretion is equal to the rate of absorption,no accumulation occurs; b) Chemical accumulates due to a

higher rate of uptake and inefficient excretion; the plasma

concentrations are, however, not sufficient to exert toxiceffects; c) The plasma concentrations reached after accu-mulation are sufficient to exert toxicity

For chemicals which irreversibly bind tomacromolecules, the magnitude of toxic re-sponses may be correlated with the total doseadministered. In contrast to chemicals whichact reversibly, the effect is not dependent onthe frequency of dosing. Effect accumulationis often observed with carcinogens and ion-

izing radiation. In Figure 7 accumulation ofeffects is exemplified by the time- and dose-dependent induction of tumors by 4-(dimethyl-amino)azobenzene, a potent chemical carcino-gen [5]. The TD50 values (50% of the treatedanimals carry tumors) are used to characterizethe potency. Identical tumor incidences were ob-served after high doses and a short exposure timeor after low doses and long exposure; the tumorincidence was only dependent on the total doseadministered.

Reversibility of toxic responses also dependson the capacity of an organ or tissue to repairinjury. For example, kidney damage by xeno-


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Toxicology 19

biotics is often, after survival of the acute phaseof the intoxication, without further consequencedue to the high capacity of the kidney for cellproliferation and thus the capacity to repair or-gan damage [6]. In contrast, injury to the centralnervous system is largely irreversible since the

differentiated cells of the nervous system cannotdivide and dead cells cannot be replaced.

Figure 7. Time-dependent induction of tumors after differ-ent daily doses of 4-dimethylaminoazobenzene in rats [5]

1.9. Factors Influencing DoseResponse

In animals and humans, the nature, severity, andincidence of toxic responses depend on a largenumber of exogenous and endogenous factors[7]. Important factors are the characteristics ofexposure, the species and strain of animals used

for the study, and interindividual variability inhumans [8]. Toxic responses are caused by a se-ries of complex interactions of a potentially toxicchemical with an organism. The type and mag-nitude of the toxic response is influenced by theconcentration of the chemical at the receptor andby the type of interaction with the receptor. Theconcentration of a chemical at the site of actionis influenced by the kinetics of uptake and elimi-nation; since these are time-dependent phenom-ena, toxic responses are also time-dependent.

Thus, the toxic response can be separated intotwo phases: toxicokinetics and toxicodynamics(Fig. 8).

Toxicokinetics describe the time dependencyof uptake, distribution, biotransformation, andexcretion of a toxic agent (a detailed descriptionof toxicokinetics is given in Section 2.5). Toxi-codynamics describes the interaction of the toxicagent with the receptor and thus specific inter-

actions of the agent (see below). Toxicokinet-ics may be heavily influenced by species, strain,and sex and the exposure characteristics [913].Differences in toxic response between species,route of exposure, and others factors are oftendependenton influences on toxicokinetics. Sincetoxicodynamics (mechanism of action) are as-sumed to be identical between species, this pro-vides the basis for a rational interspecies ex-trapolation of toxic effects when differences intoxicokinetics are defined.

Figure 8. Toxicokinetics and toxicodynamics as factorsinfluencing the toxic response

1.9.1. Routes of Exposure

The primary tissue or system by which a xeno-biotic comes into contact with the body, andfrom where it may be absorbed in order to exertsystemic toxicity, is the route of exposure. Thefrequent circumstances of environmental expo-sure are ingestion (peroral), inhalation, and skincontact. Also, for investigational and therapeutic

purposes, intramuscular, intravenous, and sub-cutaneous injections may also be routes of ex-posure.

The major routes by which a potentially toxicchemical can enter the body are in descendingorder of effectiveness for systemic delivery in-

jection, inhalation, absorption from the intesti-naltract, and cutaneous absorption. The relation-ship between route and exposure, biotransfor-mation, and potential for toxicity, may be com-plex and is also influenced by the magnitude and

duration of dosing (Table 5).The route of exposure has a major influence

on toxicity because of the effect of route of ex-


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posure on the bioavailability of the toxic agent.The maximum tissue levels achieved, the timeto maximum tissue levels, and thus the durationof the effect are determined by the rate of ab-sorption and the extent of distribution within thesystem.

Table 5.Toxicity of chemicals applied by different routes of expo-

sure (data taken from [13])

Chemical Species Route of application

LD50,mg/kg

DDT rat intravenous 68

rat oral 113

rat skin contact 1931

Atropine sulfate rat intravenous 41

rat oral 620

1-Chloro-2,4-dinitro-

benzene

rat oral 1070

rat intraperitoneal 280rabbit skin contact 130

Dieldrin rat oral 46

rat intravenous 9

rat skin contact 10

Direct injection into veins is usually re-stricted to therapeutic applications, but it is im-portant for the toxicology of intravenously in-

jected drugs in addicts. Chemicals applied byintravenous injection are rapidly distributed towell-perfused organs in the blood and thus mayresult in the rapid induction of toxic effects. Therapid dilution of a chemical after intravenousinjection by venous blood permits even the in-

jection of locally acting or corrosive chemicalswhich are well tolerated. The likelihood of toxi-city from inhaled chemicals depends on a num-ber of factors, of which the physical state andproperties of the agent, concentration, and timeand frequency of exposure are important. Majorinfluences on the absorption and disposition of

xenobiotics are exerted by species peculiaritiessince the anatomy of the respiratory tract andthe physiology of respiration show major differ-ences between rodents and humans. The watersolubility of a gaseous xenobiotic has a major in-fluence on penetration into the respiratory tract.As water solubility decreases and lipid solubil-ity increases, penetration into deeper regions ofthe lung, the bronchioli, and the alveoli becomesmore effective. Water-soluble molecules such asformaldehyde,are effectively scavenged by the

upper respiratory tract and may have toxic ef-fects on the eye and throat. In contrast, gaseswith low water solubility such as phosgene may

penetrate through the bronchii and bronchioli tothe alveoli. Damage to the alveolar surface mayinitiate a series of events that finally results inlung edema. The degree to which inhaled gases,vapors, and particulates are absorbed, and hencetheir potential to produce systemic toxicity, de-

pend on their diffusion rate through the alve-olar mebrane, their solubility in blood and tis-sue fluids, the rate of respiration, and blood flowthrough the capillaries.

Uptake through the alimentary tract repre-sents an important route of exposure for xeno-biotics accumulated in the food chain, for natu-ral constituents of human diet, and, drugs. Ab-sorption from the gastrointestinal tract is de-pendent on the lipophilicity of a chemical, themolecular mass of the xenobiotic, and the pres-ence of certain dietary constituents may influ-ence the extent and rate of absorption. Chemi-cals absorbed from the gastrointestinal tract aretransported to the liver via the portal vein; hep-atic metabolism (hepatic first-pass effect) mayefficiently reduce the concentration of the xeno-biotic available in the systemic circulation afteroral uptake. Compounds undergoing bioactiva-tion in the liver usually exhibit greater toxicitywhen given orally than when absorbed across

the respiratory tract, due to the high proportionof material passing through the liver. In contrast,chemicals causing toxicity to extrahepatic, well-perfused organs such as the kidney often showa lower degree of toxicity to extrahepatic targetorgans when given orally.

Skin contact is an important route of expo-sure in the occupational and domestic environ-ments. Local effects may include acute inflam-mation and corrosion, chronic inflammatory re-sponses, immune-mediated reactions, and neo-

plasia. The percutaneous absorption of materialsmay also be a significant route for the absorptionof systemically toxic materials. Factors influenc-ing the percutaneous absorption of substancesinclude skin site, integrity of skin, tempera-ture, formulation, and physicochemical charac-teristics, including charge, molecular mass, andhydro- and lipophilicity.

1.9.2. Frequency of Exposure

The exposure of experimental animals may becategorized as acute, subacute, subchronic, and


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Toxicology 21

chronic. Acute exposures usually last less than24 h, and all above-mentioned routes of expo-sure may be applied. With chemicals of low tox-icity, repeated exposures may be used. Acute in-halation exposure is usually less than 24 h; fre-quently 48 h is chosen as timescale. Repeated

exposure refers to application of the chemi-cal for less than one month (subacute), one tothree months (subchronic), and more than threemonths (chronic). Chronic exposures to detectspecific toxic effects (carcinogenicity of a chem-ical) may span most of the lifetime of a rodent(up to two years). Repeated exposure may be byany route; the least labor intensive route is oral,by mixing the chemical with the diet; only forspecific chemicals or to simulate likely routesof exposure for humans are application in drink-ing water, by gastric intubation, and by inhala-tion applied. These are more labor-intensive andrequire skilled personnel and/or sophisticatedtechniques and thus are more expensive.

The toxic effects observed after single expo-sure often are different form those seen after re-peated exposure. For example, inhalation of highconcentration of halothane causes anesthesia inanimals and humans. In contrast, long-term ap-plication of halothane in lower doses causes liver

damage in sensitive species The frequency of ex-posure in chronic studies is important for thetemporal characterisation of exposure. Chem-icals with slow rates of excretion may accu-mulate if applied at short dosing intervals, andtoxic effects may result (see Section 1.6). Also,a chemical producing severe effect when givenin a single high dose may have no detectableeffects when given in several smaller doses. In-terspecies and strain differences in susceptibilityto chemical-induced toxicity may be due to het-

erogeneity of populations, species specific phys-iology (for example of the respiratory system),basal metabolic rate, size- and species-specifictoxicokinetics and routes of metabolism or ex-cretion (Table 6). In some cases, animal testsmay give an underestimate, in others an overes-timate, of potential toxicity to humans [14].

1.9.3. Species-Specific Differences in

Toxicokinetics

Species-specific differences in toxic responseare largely due to difference in toxicokinetics

and biotransformation. Distribution and elimi-nation characteristics are quite variable betweenspecies. Both qualitative and quantitative differ-ences in biotransformation may effect the sen-sitivity of a given species to a toxic response(Table 7).

Table 6.Comparative LD50 values for four different chemicals in

different animal species and estimated LD50 for humans

Chemical Species LD50, mg/kg

Paraquat rat 134

mouse 77

guinea pig 41

human 32 48

Ethanol rat 12 500

mouse 8000

guinea pig 5500

human 3500 5000

Acetaminophen rat 3763mouse 777

guinea pig 2968

human 42 800

Aspirin rat 1683

mouse 1769

guinea pig 1102

human 3492

Table 7.Species and sex differences in the acute toxicity of

1,1-dichloroethylene after oral administration and inhalation in rats

and mice (data from World Health Organization, Geneva, 1990)

Species Dosing criteria Estimated LD50/LC50

Rat, male inhalation/4 h 7000 3 2 000 mg/L

Rat, female inhalation/4 h 10 3 00 m g/L

Mouse, male inhalation/4 h 115 m g/L

Mouse, female inhalation/4h 205mg/L

Rat, male gavage 1550 mg/kg

Rat, female gavage 1500 mg/kg

Mouse, male gavage 201 235 mg/kg

Mouse, female gavage 171 221 mg/kg

For example, the elimination half-live of

2,3,7,8-tetrachlorodibenzodioxin in rats is 20 d,and in humans it is estimated to be up to sevenyears [15]. An example for quantitative differ-ence in the extent of biotransformation as a fac-tor influencing toxic response is the species dif-ferences in the biotransformation of the inhala-tion anesthetic halothane. Both rats and guineapigs metabolize halothane to trifluoroacetic acid,a reaction catalyzed by a specific cytochromeP450 enzyme [1618]. As a metabolic interme-diate, trifluoroacetyl chloride is formed, which

may react with lysine residues in proteins andwith phosphatidyl ethanolamine in phospho-lipids (Fig. 9).


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22 Toxicology

This interaction initiates a cascade of eventsfinally resulting in toxicity. The metabolismof halothane in guinea pigs occurs at muchhigher rates than in rats, so guinea pigs aresensitive to halothane-induced hepatotoxic ef-fects and rats are resistant. Qualitative dif-

ferences in biotransformation are responsi-ble for apparent differences in the sensitiv-ity of rats and guinea pigs to the bladdercarcinogenicity of 2-acetylamidofluorene. Inrats, 2-acetylamidofluorene is metabolized by

N-oxidation by certain cytochrome P450 en-zymes. The N-oxide is further converted toan electrophilic nitrenium ion which interactswith DNA in the bladder; this biotransforma-tion pathway explains the formation of blad-der tumors in rats after long-term exposureto 2-acetylamidofluoren. In guinea pigs, 2-acetylamidofluorene is metabolized by oxida-tion at the aromatic ring; since nitrenium ionscannot be formed by this pathway, guinea pigsare resistant to the bladder carcinogenicity of 2-acetylamidofluorene (Fig. 10).

With some chemicals, age may significantlyaffect toxicity, likely due to age related differ-ences in toxicokinetics. The nutritional statusmay modify toxic response, likely by altering

the concentration of cofactors needed for bio-transformation and detoxication of toxic chemi-cals. Diet also markedly influences carcinogen-induced tumor incidence in animals [19] andmay be a significant factor contributing to hu-man cancer incidence.

The toxic response is influenced by the mag-nitude, number, and frequency of dosing. Thus,local or systemic toxicity produced by acute ex-posure may also occur by a cumulative pro-cess with repeated exposures to lower doses;

also, additional toxicity may be seen in repeated-exposure situations. The relationships for cu-mulative toxicity by repetitive exposure com-pared with acute exposure toxicity may be com-plex, and the potential for cumulative toxicityfrom acute doses may not be quantitatively pre-

dictable. For repeated-exposure toxicity, the pre-cise profiling of doses may significantly influ-ence toxicity.

Figure 10. Biotransformation pathways of 2-acetylamido-fluorene in rats and guinea pigs

1.9.4. Miscellaneous Factors Influencing theMagnitude of Toxic Responses

A variety of other factors may affect the na-ture and exhibition of toxicity, depending on theconditions of the study, for example, housingconditions, handling, volume of dosing, vehicle,etc. Variability in test conditions and proceduresmay result in significant interlaboratory variabil-ity in results of otherwise standard procedures.For chemicals given orally or applied to the skin,

Figure 9. Halothane metabolism by cytochrome P450 in rats, guinea pigs, and humans


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Toxicology 23

toxicity may be modified by the presence of ma-terials in formulations which facilitate or retardthe absorption of the chemicals. With respiratoryexposure to aerosols, particle size significantlydetermines the depth of penetration and deposi-tion in the respiratory tract and thus the site and

extent of the toxic effects.

1.10. Exposure to Mixtures

In experimental animals most data on the toxiceffects of chemicals are collected after exposureto a single chemical; in contrast, human expo-sure normally occurs to mixtures of chemicalsat low doses. Moreover, prior, coincidential, andsucessive exposure of humans to chemicals islikely. Interactions between the toxic effects ofdifferent chemicals are difficult to predict, ef-fects of exposure to different chemicals may beindependent, additive, potentiating (ethanol andcarbon tetrachloride), antagonistic (interferencewith action of other chemical, e.g., as seen withantidotes administered in case of intoxications),and synergistic. Ethanol exerts a potentiating ef-fect on the hepatotoxicity of carbon tetrachlo-ride. In rats pretreated with ethanol, the hepa-

totoxic effects of carbon tetrachloride are muchmore pronounced than in control animals. Thispotentiation is due to an increased capacity forbioactivation (see Section 2.4) of carbon tetra-chloride in pretreated rats due to increased con-centrations of a cytochrome P450 enzyme in theliver [20]. Thus, an important considerations forthe assessement of potential toxic effects of mix-tures of chemicals are toxicokinetics and toxico-dynamic interactions. Toxicokinetic interactionsof chemicals may influence absorption, distribu-

tion, and biotransformation, both to active andinactive metabolites. Mixtures of solvents oftenshow a competitive inhibition of biotransforma-tion. Usually, one of the components has highaffinity for a specific enzyme involved in its bio-transformation, whereas another component hasonly a low affinity for that particular enzyme.Thus, preferential biotransformation of the com-ponent with the high affinity occurs. Differentoutcomes of enzyme inhibition are possible: ifthe toxic effects of the component whose meta-

bolism is inhibited is dependent on bioactiva-tion, lower rates of bioactivation will result indecreased toxicity; if the toxic effects are inde-

pendent of biotransformation, the extent of toxi-city will increase due to slower rate of excretion.Toxic effects of mixtures may also not be due toa major component, but to trace impurities withhigh toxicity. For example, many long-term ef-fects seen in animal studies on the toxicity of

chlorophenols are believed to be due to 2,3,7,8-Tetrachlorodibenzodioxin, which was present asa minor impurity in the samples of chlorophe-nols used for these studies.

2. Absorption, Distribution,

Biotransformation and Elimination

of Xenobiotics

2.1. Disposition of XenobioticsThe induction of systemic toxicity usually re-sults from a complex interaction between ab-sorbed parent chemical and biotransformationproducts formed in tissues; the distribution ofboth parent chemical and biotransformationproducts in body fluids and tissues; their bindingand storage characteristics; and their excretion.

The biological effects initiated by a xenobi-otic are not related simply to its inherent toxic

properties; the initiation, intensity, and durationof response are a function of numerous factorsintrinsic to the biological system and the admin-istered dose. Each factor influences the ultimateinteraction of the xenobiotic and the active site(Section 1.9). Only when the toxic chemical hasreached the specific site and interacted with itcan the inherent toxicity be realized. The routea xenobiotic follows from the point of adminis-tration or absorption to the site of action usuallyinvolves many steps and is termed toxicokinet-

ics. Toxicokinetics influence the concentrationof the xenobiotic or its active metabolite at thereceptor. In the doseresponse concept outlinedin Section 1.9 and 1.7, it is generally assumedthat the toxic response is proportional to the con-centration of the xenobiotic at the receptor. How-ever, the same dose of a chemical administeredby different routes may cause different toxic ef-fects. Moreover, the same dose of two differentchemicals may result in vastly different concen-trations of the chemical or its biotransformation

products in a particular target organ. This differ-ential pattern is due to differences in the dispo-sition of a xenobiotic (Fig. 11).


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24 Toxicology

Figure 11. Possible fate of a xenobiotic in the organisms

The disposition of a xenobiotic consists ofabsorption, distribution, biotransformation, andexcretion, which are all interrelated. The com-plicated interactions between the different pro-cesses of distribution are very important deter-minants of the concentration of a chemical atthe receptor and thus of the magnitude of toxicresponse. They may also be major determinants

for organ-specific toxicity.For example, in the case of absorption of axenobiotic through the gastrointestinal tract, thechemical proceeds from the intestinal lumen intothe epithelial cells. Following intracellular trans-port, it passes through the basal membrane andlamina propria and enters the blood or lymphcapillaries for transport to the site of action orstorage. At that site, the xenobiotic is releasedfrom the capillaries, into an interstitial area, andfinally through various membranes to its site of

action, which may be a specific receptor, an en-zyme, a membrane, or many other possible sites.

2.2. Absorption

The skin, the lungs, and the cells lining the al-imentary tract are major barriers for chemicalspresent in the environment. Except for causticchemicals, which act at the site of first contactwith the organism, xenobiotics must cross thesebarriers to exert toxic effects on one or several

target organs. The process whereby a xenobi-otic moves through these barriers and enters thecirculation is termed absorption.

2.2.1. Membranes

Because xenobiotics must often pass throughmembranes on their way to the receptor, it isimportant to understand membrane character-istics and the factors that permit transfer of

foreign compounds. Membranes are initiallyencountered whether a xenobiotic is absorbed


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Toxicology 25

by the dermal, oral, or vapor route. These mem-branes may be associated with several layers ofcells or a single cell. The absorption of a sub-stance from the site of exposure may result frompassive diffusion, facilitated diffusion, activetransport, or the formation of transport vesicles

(pinocytosis and phagocytosis). The process ofabsorption may be facilitated or retarded by avariety of factors; for example, elevated tem-perature increases percutaneous absorption bycutaneous vasodilation, and surface-active ma-terials facilitate penetration. Each area of entryfor xenobiotics into the organism may havespecific peculiarities, but a unifying concept ofbiology is the basic similarity of all membranesin tissues, cells, and organelles.

Figure 12. Simplified model of the structure of a biologicalmembrane

All membranes are lipid bilayers with polarhead groups (phosphatidylethanolamine, phos-phatidylcholine). The polar groups predominateat the outer and inner surfaces of the membrane;

the inner space of the membrane consists of per-pendicularly arranged fatty acids [21]. The fattyacids do not have a rigid structure and are fluidunder physiological conditions; the fluid charac-ter of the membrane is largely dominated by thefatty acid composition. The width of a biologicalmembrane is approximately 79 nm. Figure 12illustrates the concept of a biological membrane(fluid-mosaic model).

Proteins are intimately associated with themembrane and may be located on the surface

or inside the membrane structure, or extendcompletely through the membrane. These pro-teins may also form aqueous pores. Hydropho-

bic forces are responsible for maintaining thestructural integrity of both proteins and lipidswithin the membrane structure. The ratio of lipidto protein in different membranes may vary from5:1 (e.g,. myelin) to 1:5 (e.g., the inner mem-brane of mitochondria). Usually, pore diame-

ters in membranes are

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