ToxicologyAn Introduction

ToxicologyAn IntroductionJohn Duffus

The Edinburgh Centre for Toxicology

John Duffus

The Edinburgh Centre for Toxicology

General considerationsGeneral considerations

�Toxicology is the science which lets us know how substances can harm life by physico-chemical reactions with living cells

�All substances, whether synthetic or natural, can cause harm to people, animals, plants, micro-organisms, and their environment

�Toxicology is the science which lets us know how substances can harm life by physico-chemical reactions with living cells

�All substances, whether synthetic or natural, can cause harm to people, animals, plants, micro-organisms, and their environment

General considerationsGeneral considerations

�There are naturally occurring substances which are as poisonous as or more poisonous than the most toxic synthetic chemicals

�For example - ricin, Clostridium botulinum toxin, saxitoxin

�There are naturally occurring substances which are as poisonous as or more poisonous than the most toxic synthetic chemicals

�For example - ricin, Clostridium botulinum toxin, saxitoxin

Toxicity and doseToxicity and dose

�The toxicity (poisonous nature) of any substance is inversely related to the amount (dose) required to cause harm

�The more that is required, the lower the toxicity

�But But But But - all substances are toxic at a high enough dose

�The toxicity (poisonous nature) of any substance is inversely related to the amount (dose) required to cause harm

�The more that is required, the lower the toxicity

�But But But But - all substances are toxic at a high enough dose

Toxicity and doseToxicity and dose

�Substances that can cause harm following exposure to very small amounts, sometimes no more than a few molecules, are said to be extremely toxic

�Substances that require exposure to many grams before harm results are said to have low toxicity

�Substances that can cause harm following exposure to very small amounts, sometimes no more than a few molecules, are said to be extremely toxic

�Substances that require exposure to many grams before harm results are said to have low toxicity

Toxicity and DoseToxicity and Dose

�Even essential nutrients become toxic if the amount ingested is above a certain acceptable dose

�Some substances such as oxygen are toxic (harmful) at the dose which is essential for life! We are protected by anti-oxidants

�Oxygen is toxic at all levels to obligate anaerobes (toxicity classification?)

�Even essential nutrients become toxic if the amount ingested is above a certain acceptable dose

�Some substances such as oxygen are toxic (harmful) at the dose which is essential for life! We are protected by anti-oxidants

�Oxygen is toxic at all levels to obligate anaerobes (toxicity classification?)

Increasing DoseIncreasing Dose

Threshold Threshold NonstochasticNonstochasticToxicantToxicant

ThresholdThreshold

NoNo--Threshold Threshold Stochastic Stochastic ToxicantToxicant

Essential Essential NutrientNutrient

Increasing Increasing adverse effectadverse effect

0

NOAELNOAEL

Dose-effect curvesDose-effect curves

What is a poison?What is a poison?

�500 years ago, a physician called Paracelsus wrote the following fundamental rule of toxicology:

�“Only the dose required makes the difference between a cure and a poison”

�Note: Paracelsus is the name given to Theophrastus Phillippus Aureolus Bombastus vonHohenheim

�500 years ago, a physician called Paracelsus wrote the following fundamental rule of toxicology:

�“Only the dose required makes the difference between a cure and a poison”

�Note: Paracelsus is the name given to Theophrastus Phillippus Aureolus Bombastus vonHohenheim

What is a poison?What is a poison?

�Water ? - essential for life but people have died from drinking too much pure water, washing out essential salts such as sodium chloride and potassium chloride

�Fresh water fish rarely survive in salt water or salt water fish in fresh water and so both could be classified as “poisons”

�Water ? - essential for life but people have died from drinking too much pure water, washing out essential salts such as sodium chloride and potassium chloride

�Fresh water fish rarely survive in salt water or salt water fish in fresh water and so both could be classified as “poisons”

What is a poison?What is a poison?

�Vitamin A ? - essential for human health; it is fairly easy to exceed the required dose and people have died from eating too much

�It may also cause developmental abnormalities in babies if their mothers consume too much while carrying them during pregnancy

�Vitamin A ? - essential for human health; it is fairly easy to exceed the required dose and people have died from eating too much

�It may also cause developmental abnormalities in babies if their mothers consume too much while carrying them during pregnancy

What is a poison?What is a poison?

�Oxygen ? - essential for aerobic life but destroys essential molecules by oxidation; protective mechanisms involving compounds such as vitamin C, vitamin E, and glutathione have evolved

�Reactive oxygen species (ROS) have been implicated in mutagenesis and carcinogenesis

�Oxygen ? - essential for aerobic life but destroys essential molecules by oxidation; protective mechanisms involving compounds such as vitamin C, vitamin E, and glutathione have evolved

�Reactive oxygen species (ROS) have been implicated in mutagenesis and carcinogenesis

Natural and synthetic chemicals

Natural and synthetic chemicals

�Aflatoxin, from the fungus Aspergillus flavus, is one the most potent carcinogens known, causing liver cancer in people, birds and fish after eating food made from contaminated cereals such as rice

�Aflatoxin, from the fungus Aspergillus flavus, is one the most potent carcinogens known, causing liver cancer in people, birds and fish after eating food made from contaminated cereals such as rice

Natural and synthetic chemicals

Natural and synthetic chemicals

�The distinction between naturally occurring substances and synthetic substances is of little value in considering toxicity

�Pure natural vitamin C and pure synthetic vitamin C are identical molecules and equally good for people

�The distinction between naturally occurring substances and synthetic substances is of little value in considering toxicity

�Pure natural vitamin C and pure synthetic vitamin C are identical molecules and equally good for people

Movement through the environment

Movement through the environment

�Substances in the environment, whether toxic or not, may enter living organisms directly - by inhalation (lungs or gills), ingestion, by skin contact, through wounds, or through the eyes

�Or indirectly from food or through the environmental media - air, soil, sediments, or water

�Substances in the environment, whether toxic or not, may enter living organisms directly - by inhalation (lungs or gills), ingestion, by skin contact, through wounds, or through the eyes

�Or indirectly from food or through the environmental media - air, soil, sediments, or water

Routes of human exposure from the environment

Routes of human exposure from the environment

�Each route of exposure must be considered separately although the effects may be interactive

�Each route of exposure must be considered separately although the effects may be interactive

Routes of direct human exposureRoutes of direct human exposureRoutes of direct human exposure

EyesEyesEarsEarsRespiration through nose and mouthRespiration through nose and mouth

Ingestion through Ingestion through mouth*mouth*

Absorption through skin, Absorption through skin, woundswounds

Exposure of baby in the Exposure of baby in the womb through the placentawomb through the placenta

*Do not forget exposure of babies through *Do not forget exposure of babies through mothermother’’s milk s milk

Different routes -different effects?

Different routes -different effects?

�Different routes of exposure may cause different effects from the same substance�Sometimes only one route of exposure is harmful

�For example, organophosphate pesticides are highly toxic through the lungs but break down in the stomach

�Different routes of exposure may cause different effects from the same substance�Sometimes only one route of exposure is harmful

�For example, organophosphate pesticides are highly toxic through the lungs but break down in the stomach

Exposure of the child in the womb and babies

Exposure of the child in the womb and babies

�The developing child in the womb may be harmed by substances in the mother’s bloodstream which can pass across the placenta into the baby’s blood circulation�An example is methylmercury chloride, found naturally in tuna and swordfish, which can kill a baby in the womb at levels which do not harm the mother

�The developing child in the womb may be harmed by substances in the mother’s bloodstream which can pass across the placenta into the baby’s blood circulation�An example is methylmercury chloride, found naturally in tuna and swordfish, which can kill a baby in the womb at levels which do not harm the mother

Exposure of the child during breast feeding

Exposure of the child during breast feeding

�The breast feeding child may be harmed by substances in mother’s milk�Particularly dangerous in this respect are persistent fat-soluble compounds such as organochlorine pesticides which can accumulate in the breast; they are included in the United Nations list of persistent organic pollutants (POPS), banned in many countries

�The breast feeding child may be harmed by substances in mother’s milk�Particularly dangerous in this respect are persistent fat-soluble compounds such as organochlorine pesticides which can accumulate in the breast; they are included in the United Nations list of persistent organic pollutants (POPS), banned in many countries

NanoparticlesNanoparticles

�Particles smaller than 2.5 µm and particularly nanoparticles, previously called ultrafines (<100 nm), may lodge permanently in the alveoli and cause chronic problems; nanoparticles may move through cell junctions into the body and affect the heart

�Particles smaller than 2.5 µm and particularly nanoparticles, previously called ultrafines (<100 nm), may lodge permanently in the alveoli and cause chronic problems; nanoparticles may move through cell junctions into the body and affect the heart

Phases of poisoningPhases of poisoning

�Exposure (already discussed) is the first phase

�The toxicokinetic phase covers uptake to excretion

�The toxicodynamic phase covers all aspects of the way in which a substance causes harm once it reaches its “target” in the body

�Exposure (already discussed) is the first phase

�The toxicokinetic phase covers uptake to excretion

�The toxicodynamic phase covers all aspects of the way in which a substance causes harm once it reaches its “target” in the body

Phases of poisoningPhases of poisoningPhases of poisoningChemical Exposure Chemical Exposure

PhasePhaseToxicokineticToxicokinetic

PhasePhaseToxicodynamicToxicodynamic PhasePhase

Potentially Potentially Toxic Toxic

SubstancesSubstances

Derivatives Derivatives or or

FormulationFormulation

BiotransformationBiotransformation

AbsorptionAbsorptionCirculationCirculation

Toxicity Toxicity InitiatorInitiator

Tissue BindingTissue Binding

CirculationCirculation

ExcretionExcretion

Reaction at Reaction at Key Sites Key Sites

(receptors)(receptors)

ToxicityToxicity

Metabolism and excretion of fat soluble (lipophilic) substances

Metabolism and excretion of fat soluble (lipophilic) substances

Fat Soluble Fat Soluble SubstancesSubstances

Highly Fat Highly Fat Soluble Soluble

SubstancesSubstances

Binding to blood cells, albumin, Binding to blood cells, albumin, lipoproteins, lymph cells, lymph proteinslipoproteins, lymph cells, lymph proteins Physical localization, Physical localization,

accumulation in fatty accumulation in fatty tissuetissue

Phase 1 and Phase 2 ReactionsPhase 1 and Phase 2 Reactions

Water soluble productsWater soluble products

Blood CirculationBlood Circulation

Excretion in UrineExcretion in Urine

Blood Blood CirculationCirculation

Excretion in Excretion in Bile to the Bile to the

GutGut

Excretion in FaecesExcretion in Faeces

ReabsorptionReabsorptionfrom Gutfrom Gut

Hydrolysis in GutHydrolysis in Gut

Milk, sweat Milk, sweat and other and other secretions, secretions, including including genital genital secretionssecretions

Breathed Out Breathed Out in Airin Air

Acute toxicityAcute toxicity

�Toxicity resulting from short exposure is called acute acute acute acute toxicitytoxicitytoxicitytoxicity

�Toxicity resulting from short exposure is called acute acute acute acute toxicitytoxicitytoxicitytoxicity

Duffus & Worth, ©IUPAC39

Acute Toxicity

Chronic toxicityChronic toxicity

�Toxicity resulting from long term exposure

�Toxicity resulting from long term exposure

Duffus & Worth, ©IUPAC40

Chronic Toxicity

Chronic toxicity follows accumulation of toxicants or

effects

Chronic toxicity follows accumulation of toxicants or

effects�Accumulation by storage in:

�Fat– DDT, PCBs, tetraethyl lead etc

�Bone– Lead ions, calcium ions, strontium ions etc

�Liver and kidney-bind and trap both organic and inorganic toxicants

�Plasma proteins- especially albumin, bind both organics and inorganics; competition for binding may displace a large dose causing toxicity

�Accumulation by storage in:

�Fat– DDT, PCBs, tetraethyl lead etc

�Bone– Lead ions, calcium ions, strontium ions etc

�Liver and kidney-bind and trap both organic and inorganic toxicants

�Plasma proteins- especially albumin, bind both organics and inorganics; competition for binding may displace a large dose causing toxicity

Important aspects of chronic toxicity

Important aspects of chronic toxicity

�Chronic toxicity includes the production of cancer (carcinogenicity) and possibly senile dementia and other diseases of old age

�Note the special problem of brain cells where normal decrease through life may be accelerated by exposure to toxicants

�Chronic toxicity includes the production of cancer (carcinogenicity) and possibly senile dementia and other diseases of old age

�Note the special problem of brain cells where normal decrease through life may be accelerated by exposure to toxicants

Exposure to mixturesExposure to mixtures

�Living organisms are exposed to mixtures of potentially harmful substances

�There are four types of effects chemicals can have on each other: exposure to two or more substances simultaneously may produce effects that are independent, additive, synergistic, or antagonistic

�Living organisms are exposed to mixtures of potentially harmful substances

�There are four types of effects chemicals can have on each other: exposure to two or more substances simultaneously may produce effects that are independent, additive, synergistic, or antagonistic

Effects of exposure to mixturesEffects of exposure to mixtures

�Independent – substances do not interfere with each other or enhance each other’s effect on simultaneous exposure

�Additive - have the same effect independently and any combined exposure produces a total effect equal to the sum of the effects of separate exposure to each substance

�Independent – substances do not interfere with each other or enhance each other’s effect on simultaneous exposure

�Additive - have the same effect independently and any combined exposure produces a total effect equal to the sum of the effects of separate exposure to each substance

Effects of exposure to mixturesEffects of exposure to mixtures

�Synergistic – substances have the same effect or different effects but the final effect observed is greater than the sum of the effects of separate exposure to each substance

�Antagonistic - effect of one substance counteracts the adverse effect of another; exposure to the substances together has less effect than the sum of the effects of independent exposures

�Synergistic – substances have the same effect or different effects but the final effect observed is greater than the sum of the effects of separate exposure to each substance

�Antagonistic - effect of one substance counteracts the adverse effect of another; exposure to the substances together has less effect than the sum of the effects of independent exposures

Chemical SpeciationChemical Speciation

�The chemical species of an element is the specific form in which it exists, defined as to isotopic composition, electronic or oxidation state, and/or complex or molecular structure

�Thus, organic chemistry is the study of the nature and properties of all the chemical species of carbon

�The chemical species of an element is the specific form in which it exists, defined as to isotopic composition, electronic or oxidation state, and/or complex or molecular structure

�Thus, organic chemistry is the study of the nature and properties of all the chemical species of carbon

Why is speciation important?Why is speciation important?�Consider mercury.

�It can exist as the pure metal which is an electrically neutral volatile liquid

�The vapour is fat soluble, the liquid metal is neither fat soluble or water soluble

�It can also exist as Hg+ or Hg2+ ions that are sparingly soluble. However HgCl2 in seawater is largely unionized and therefore fat soluble

�Methylmercury chloride is sparingly water soluble and very fat soluble

�Consider mercury.

�It can exist as the pure metal which is an electrically neutral volatile liquid

�The vapour is fat soluble, the liquid metal is neither fat soluble or water soluble

�It can also exist as Hg+ or Hg2+ ions that are sparingly soluble. However HgCl2 in seawater is largely unionized and therefore fat soluble

�Methylmercury chloride is sparingly water soluble and very fat soluble

Why is speciation important?Why is speciation important?�Mercury (continued)

�Fat soluble mercury vapour and mercuric chloride enter cells easily through the phospholipid membrane

�Liquid mercury is not absorbed by cells.

�Ionized mercurous chloride does not enter cells; unionized mercuric chloride does.

�Fat soluble methylmercury chloride enters cells readily

�Mercury (continued)

�Fat soluble mercury vapour and mercuric chloride enter cells easily through the phospholipid membrane

�Liquid mercury is not absorbed by cells.

�Ionized mercurous chloride does not enter cells; unionized mercuric chloride does.

�Fat soluble methylmercury chloride enters cells readily

Why is speciation important?Why is speciation important?

�Mercury continued

�Fat-soluble mercury species, e.g. mercury vapour, fat-soluble unionized mercuric chloride, and fat-soluble methylmercury chloride are highly toxic

�Liquid mercury and ionized mercurous and mercuric salts have relatively low toxicity

�Mercury continued

�Fat-soluble mercury species, e.g. mercury vapour, fat-soluble unionized mercuric chloride, and fat-soluble methylmercury chloride are highly toxic

�Liquid mercury and ionized mercurous and mercuric salts have relatively low toxicity

The biological dimension -prokaryotes (e.g.bacteria) –

no nuclear membrane

The biological dimension -prokaryotes (e.g.bacteria) –

no nuclear membrane

DNA

Cell Membrane

Tail

Glycogen

Ribosomes

The biological dimension –eukaryotes, e.g., human cells, note the nuclear membrane

The biological dimension –eukaryotes, e.g., human cells, note the nuclear membrane

Toxicity targets in eukaryote cells

Toxicity targets in eukaryote cells37

30 µm

Lysosome

Ribosomes

Golgi apparatus

Plasmicmembrane

Microfilaments

Vacuole

Endoplasmic reticulum (rough)

Endoplasmic reticulum (smooth)

“THE CELL’S POWER PLANT” ATP synthesis

Synthesis of proteins for the cell

and for export

“Waste treatment” : Intracellular

digestion

Synthesizes carbohydrates which are packed with

proteins for cellular use or export

Detoxification. Production of lipids, carbohydrates

Protein synthesis

Nucleus

Directs protein synthesis, regulates metabolism, reproduction. Contains

chromatin, DNA

MitochondriaCytosol

Speciation and biologySpeciation and biology

CellCellmembranemembrane

Cr(III) as[Cr(H2O)6OH]2+

Cr(VI) as CrO42-

(chromate)

Sulfate / phosphateactive transport system

Cytochrome P450 system

Cr (III / IV)- species unknown

Reacts with phosphate and sulfate metabolismenzymes, DNA, RNA and their polymerases?

Cr(VI) as CrO42-

(chromate)

Reactive Oxygen Species (ROS)

Mutations

Nuclear Nuclear membranemembrane

Hypotheses

The EndThe End

�Thank you very much for your attention

�Thank you very much for your attention