Toxicokinetics & Toxicodynamics Toxicokinetics (Determines the no. molecules that

can reach the receptors)• Uptake• Transport• Metabolism & transformation• Sequestration• Excretion

Toxicodynamics (Determines the no. of receptors that can interact with toxicants)

• Binding • Interaction • Induction of toxic effects

Uptake and Elimination 

BiologicalSystemUptake EliminationK1 K2

K1 > K2 : Accumulation & Toxic effect

Toxicokinetics

1. Uptake2. Transport3. Metabolism & Transformation4. Sequestration5. Excretion

Uptake routes

1. Ingestion (toxicity may be modified by enzymes, pH and microbes)

2. Respiration (Air borne toxicants)3. Body surface (Lipid soluble toxicants

such as carbon terta chloride and organophosphate)

Uptake Barriers

1. Cell membrane 2. Cell wall/cuticles/stomata3. Epithelial cells of GI tract 4. Respiratory surface (lung, gill

tracheae)5. Body surface

Uptake of Toxicants

1. Passive diffusion2. Facilitated transport3. Active transport4. Pinocytosis

Uptake by Passive diffusion

Uncharged molecules may diffuse along conc. gradient until equilibrium is reached

Not substrate specific Small molecules of < 0.4 nm (e.g. CO,

N20, HCN) can move through cell pores Lipophilic chemicals may diffuse through

the lipid bilayer

Uptake by Passive diffusion

First order rate process, depends on:– Concentration gradient– Surface area (aveoli = 25 x body surface) – Thickness (fluid mosaic phospholipid bi-layer

ca. 7 nm)– Lipid solubility & ionization(dissolved before

transport, polar chemicals have limited diffusion rate)

– Molecular size (membrane pore size = 4-40 A, allowing MW of 100-70,000 to pass through)

Diffusion governed by Flicks law

  D/dt = KA (Co - Ci) / X  Where:

– dD/dt = rate of transport accross the membrane – K= constant– A= Cross sectional area of membrane exposed to the

compound– Co = Concentration of the toxicant outside the

membrane– Ci = Concentration of the toxicant inside the

membrane– X= Thickness of the membrane

Uptake by Facilitated Transport

Carried by trans-membrane carrier along concentration gradient

Energy independent May enhance transport up to 50,000 folds Example: Calmodulin for facilitated

transport of Ca

Uptake by Active Transport Independent of or against conc. gradient Require energy Substrate –specific Rate limited by no. of carriers Example:

– P-glycoprotein pump for xenobiotics (e.g. OC) – Ca-pump (Ca2+ -ATPase)

Uptake by Pinocytosis

For large molecules ( ca 1 um) Outside: Infolding of cell membrane Inside: release of molecules Example:

– Airborne toxicants across alveoli cells – Carrageenan accross intestine

Transport & Deposition Transport

• Blood• Lymph, haemolymph• Water stream in xylem • Cytoplamic strands in phloem

Deposition Toxicant Target organs

Pb Bone, teeth, brainCd Kidney, bone, gonadOC, PCB Adipose tissue,milkOP Nervous tissueAflatoxin Liver

Metabolism & Transformation Evolved to deal with metabolites and

naturally occurring toxicants Principle of detoxification:

1. Convert toxicants into more water soluble form (more polar & hydrophilic)

2. Dissolve in aqueous/gas phases and eliminate by excretion (urine/sweat) of exhalation

3. Sequestrate in inactive tissues (e.g bone, fat)

P450 system A heme-containing cytochrome protein

located in ER, and is involved in electron transport.

Highly conservative, occur in most plants & animals

Two phases of transformation May increase or decrease toxicity of

toxicants after transformation (e.g turn Benzo[a]pyrene into benzo[a]pyrene diol epoxide, and nitroamines into methyl radicals)

Inducible by toxicants

Induction of P450

Aryl HydrocarbonReceptor

Toxicant

Toxicant-ReceptorComplex

Translocatingprotein

m-RNA for CYP1A

hoursBind at Specific site

Phase I Transformation Mixed Function Oxidase (MFO) System in

smooth ER is responsible (Microsomes) In vertebrates, primarily found in liver

parenchyma cells, but also other tissues (e.g intestine, gill)

In invertebrates, found in hepatopancrease & digestive glands

Lower MFO activities in molluscs Add polar group(s) to increase

hydrophilicity for Phase II transformation

Examples of Phase I Transformation

Hydrolysis  RCOO-R’ + H2O ---------> RCOO-H + R’-OH

Hydroxylation 

NADP NADP+

R-H --------------------------> R-OH + H2O

Examples of Phase I Transformation

Epoxidation O

R-CH==CH-R’ -----------> R---CH ----CH-R’

 

Phase II transformation • Cytochrome P450 II enzyme systems

in cytosol is responsible • Covalent conjugation to water soluble

endogenous metabloites (e.g. sugars, peptides, glucuronic acid, glutathione, phosphates & sulphate)

• May involve deamination, acyclic hydroxylation, aromatic hydroxylation, and dealkylation

• Further increase hydrophilicity for excretion in bile, urine and sweat

Important Phase II enzymes

Glutathion S-transferases (GST)Epoxide Hydrolase (EH) UDP-glucuronosyltransferase (UDP-

GTS) Sulfotransferase (ST).

Examples of Phase II Transformation

Deamination R-NH2 ---------------------------> R=O + NH3

 

Examples of Phase II Transformation Dealkylation

R-CH2-CH3 ----------------------> R + CH3-CH2O

Dehalogenation: R-Cl ---------------------------------> R-H + Cl+

Glutathione-S-transferase (GST)

  O R------R’ ----------------------> HO-R-SG 

R-Cl ------------------------------> R-SG + Cl

GST

GST

Sequestration Animals may store toxicants in inert

tissues (e.g. bone, fat, hair, nail) to reduce toxicity

Plants may store toxicants in bark, leaves, vacuoles for shedding later on

Lipophilic toxicants (e.g. DDT, PCBs) may be stored in milk at high conc and pass to the young

Metallothionein (MT) or phytochelatin may be used to bind metals

Excretion Gas (e.g. ammonia) and volatile (e.g. alcohol)

toxicants may be excreted from the gill or lung by simple diffusion

Water soluble toxicants (molecular wt. < 70,000) may be excreted through the kidney by active or passive transport

Conjugates with high molecular wt. (>300) may be excreted into bile through active transport

Lipid soluble and non-ionised toxicants may be reabsorbed (systematic toxicity)

Tutorial Questions1. Find TWO enzymes/proteins which are

inducible by xenobiotics or metals

2. Molluscs have low P450 activities. They are often used as pollution indicators for metals and xenobiotics. Explain why.

3. Lipophilic compounds may normally have a longer biological half life. Explain why.

4. Why exposure of animals to sub-lethal level of toxicants may increase tolerance of the organisms to the chemical.