Toxicant Disposition and Metabolism

Jan ChambersCenter for Environmental Health Sciences

College of Veterinary Medicinechambers@cvm.msstate.edu

Definitions

• Disposition– Absorption—passage across membrane.– Distribution—circulation in blood stream.– Storage—sequestration.– Excretion—elimination from body.

• Metabolism– Enzyme-mediated change in chemical structure.– Change in size, configuration, polarity, reactivity.

Disposition

Absorption• Portals of entry

– Digestive tract (oral route of exposure).– Respiratory tract (respiratory route of exposure).– Skin (dermal route of exposure).

• Mechanisms of entry– Diffusion across membranes (kinetic energy).– Filtration across membranes (hydrostatic energy).– Active transport (specific carrier protein, metabolic

energy).– Endocytosis (pinocytosis, phagocytosis) (receptors,

metabolic energy).

Membrane

Cell

Digestive Tract

Brush Border

Respiratory System•

Alveolus

Skin

Skin

Absorption--Summary

• Majority of toxicants diffuse through membranes.• Majority of toxicants/xenobiotics of biological

interest (e.g., drugs) are lipophilic.• Therefore, a general requirement for toxicant

absorption is lipophilicity (non-polar, non-charged).

• Toxicant size is generally less relevant than lipophilicity.

Distribution

• Once absorbed, circulate in the blood stream.

• Serum is aqueous medium.• Lipophilic toxicants are not readily

dissolved or suspended in the serum.• Bound to serum proteins (e.g., albumin).• Must exit blood and cross membranes to

reach biological targets.

Distribution

serum protein serum capillary membrane interstitial fluid [cell membrane cytosol (organelle membrane interior of organelle)] target molecule (i.e., receptor, enzyme, channel, DNA).

Circulatory System

Hepatic Portal Vein and EnterohepaticCirculation

Storage

• Storage sites: – fat stores and fatty tissues (e.g., liver) for

lipophilic toxicants; partition into fat; e.g., polychlorinated biphenyls (PCB’s).

– bone for divalent cations resembling calcium; active transport using calcium transporter; e.g., lead.

• Stored toxicant is biologically inactive until mobilized.

Excretion

• Major routes: kidney (urine), digestive tract (feces).

• Minor routes: respiratory tract, tears, sweat.• Urine and feces are aqueous media.• Lipophilic toxicants cannot partition into urine or

feces; therefore, cannot be excreted—bioaccumulation [(e.g., PCB’s , organochlorine insecticides, persistent organic pollutants (POP’s)].

Kidney

Nephron

Enterohepatic Circulation

Metabolism

Metabolism/Biotransformation• Chemical alteration to structure.• Enzyme-mediated (enzyme is protein, chemical

catalyst, lowers activation energy for reaction).• Outcome: changes physicochemical

characteristics of toxicant:– Ability to be stored or excreted (half-life; potential for

bioaccumulation).– Reactivity with targets (toxic potential; bioactivation or

detoxication).

In general: lipophilic (readily absorbed/stored) hydrophilic (readily excreted).

Enzyme Reaction Schematic

Reaction Pathways

Categories of Biotransformation Reactions

• Phase 1: adds or uncovers a reactive group.– Makes more polar; more likely to be excreted, but may

not be truly water soluble.– More chemically reactive; possibly more toxic.– More likely to undergo Phase 2 metabolism.

• Phase 2: adds an endogenous ligand.– Usually makes more polar and usually water soluble.– Usually ligand interacts with reactive group, so

decreases toxicity.– May act on parent toxicant or Phase 1 metabolite.

Locations of Metabolism

• Most active tissue: liver.• Moderately active tissues: kidney, skin, intestine.• Therefore, oral route of exposure leads to greater

toxicant metabolism than respiratory or dermal routes.– If toxicant is bioactivated, oral route leads to greater

toxicity than other routes.– If toxicant is detoxified, oral route leads to lesser

toxicity than other routes

Phase 1 Reactions

• Oxidation.– Monooxygenases:

• Cytochromes P450 (CYP; P450).• Flavin-containing monooxygenases (FMO).

– Dehydrogenases:• Alcohol dehydrogenase.• Aldehyde dehydrogenase.

• Hydrolysis.– Hydrolases (esterases, amidases).– Hydratases..

• Other.

Cytochromes P450

• Enzyme family, with broad substrate specificities.• Most significant of all toxicant oxidation reactions.• Adds one atom of molecular oxygen to substrate,

other atom becomes a reactive oxygen species (with potential for oxidative damage within the cell).

• Very important in detoxication of many toxicants.• Most important for bioactivations:

– carcinogens (e.g., polycyclic aromatic hydrocarbons, PAH’s; benzene; vinyl chloride; aflatoxin).

– neurotoxicants (e.g., organophosphate insecticide oxons).– hepatotoxicants (e.g., carbon tetrachloride).

P450 Reaction Cycle

P450: Epoxidation

P450: N-Oxidation

P450: Desulfuration/Dearylation

P450: O-Demethylation

Hydrolysis

• Addition of water to break a bond and perhaps the molecule.

• Hydrolases (e.g., esterases, amidases): split molecule into two metabolites.

• Hydratases: hydrates a bond, but molecule remains intact.

• Usually detoxications.

Organophosphate Hydrolysis

Epoxide Hydration

Other: DDT Dehydrochlorinase

Phase 2 Reactions

• Conjugation reactions, adding an endogenous ligand to a reactive moiety.– Makes more water-soluble and usually detoxifies.

• Sulfate.• Glucuronic acid (a sugar).• Glutathione (a peptide).

– Makes less water-soluble and more readily absorbed.• Metal methylation; e.g., inorganic to methyl mercury.

Sulfate Conjugation

Glucuronide Conjugation

Glutathione (GSH) Conjugation

Reaction Pathways

Levels of Enzymes

• Vary with age.• Vary with sex.• Inhibition—drug interactions, insecticide

synergists.• Induction—alcohol, PAH’s, PCB’s, drugs.

SUMMARY

• Lipophilic toxicants can get in, but don’t leave.

• Phase 1 metabolites more likely to leave but may be highly toxic reactive metabolites.

• Phase 2 metabolites readily excreted.