Route of exposure, mode of action and modifying factors

1. Route of exposure, mode of action and modifying factors Important considerations – interaction of animals and plants with xenobiotics Exposure Uptake Transport Storage Metabolism Excretion

2. LEAF EPIDERMIS (stomata) Uptake barriers are important. For example: • Exposure vs. uptake of toxicant from air and rain in plants • Atmospheric concentration is important, but what more important is what gets into the plant • Stomata might be completely closed/regulated by CO2, temperature, humidity light, water, nutrients (potassium in particular) • When the stomata are closed the toxicant might not be able to get into the plant • Pollutants like ozone on the other hand may affect the membrane material to form toxic substance, such as aldehydes, ketones and various free radicals INTRACELLULAR SPACE SPONGY PARENCHYMA PALISADE PARENCHYMA EXCRETION

3. In animals the exposure may be • Dermal • skin • Water pollution e.g. swimming in a polluted lake • Inhalation • lungs • air pollutants e.g. ozone, lead, methyl mercury • Ingestion • intestine • pollutant in food; e.g. PCBs in fish we eat • Immediate and long term effects are directly linked to the mode of entry into the body • The most common place of entry in animals is the skin through follicles, sweat glands, and wounds • Toxicant has to pass through a series of membranes (tissue, cell, capillary

4. Uptake / how to get through a membrane • Passive diffusion • Facilitated diffusion • Active transport • Lipophilicity (Kow) of a compound is important, because it affects adsorption properties

6. Transport • Lymphatic system • Blood system • Transport is usually achieved through blood protein that binds the toxicant – e.g. a lipoprotein • Storage • Liver • Lungs • Kidneys • Bone • Adipose (fat) tissue • Toxicant may be stored in one reservoir but later transported elsewhere depending on physiological conditions • Storage site may or may not be the site of the toxic effect • Lead (Pb) is stored in bone, but it’s toxic effect is in the liver • DDT is stored in adipose tissue but it’s effect is through prolonged gradual release

7. Metabolism • Toxicants are metabolized by • Lungs • Gastrointestinal tract • Skin • Kidneys • Central role : liver • The liver contains many non-specific enzymes that give it the ability to metabolize a broad spectrum of organic molecules • Two phases: • Phase I involves the addition of reactive polar groups through oxidation, reduction, or hydrolysis; sometimes this makes a non-toxic chemical more toxic • Phase II involves conjugation with an endogenous substance to form a complex secondary metabolite which is more water soluble so it can be excreted

8. General mechanism of action of xenobiotics • Intrinsic activity • Activated metabolites • Interact with specific site of action to initiate toxic effect • Effect can be anywhere in the body • Oxidative metabolism • Central nervous system • DNA replication (cancer and reproduction) • Protein synthesis • Protein function • Effect can be • Be terminated by • storage • transformation • Excretion • Permanent

9. Disruption or destruction of cellular structure • Structural damage to tissue • Plants • SO2, O3, NO2 and fluoride are for example phytotoxic • Feedback: SO2 weakens stomata which lets even more SO2 into cells • Animals • SO2, O3 can cause irritation and damage to the respiratory system pulmonary edema • fluid buildup in alveoli due to membrane disruption • Industrial solvents can cause narcosis • General symptomatic response to toxicants • Results from membrane alterations / ion-transport • logKow (log([octanol]/[water])) is used to asses hydrophobicity of toxicants (correlates with toxicant’s ability to cross lipid membranes)

10. Narcosis • Alteration of physical/chemcial proterties of lipid bilayer • Permeability • Fluidity • Interact with pumps and receptors • Affect enzyme function

11. Binding to cellular constituents • Xenobiotics my directly bind to a cellular component and inhibit it’s normal function • Carbon monoxide (CO) binds to hemoglobin (Hb) in red blood cells and prevent the Hb from binding O2 • Cadmium; highly toxic heavy metal that binds to metallothionein. This occurs primarily in the kidneys where Ca-metallothionein accumulates and is highly toxic to tubular cells Effect on Enzymes • Enzymes are proteins that are involved in biochemical catalysis i.e. they help transform chemical structures. The purpose of a catalyst is to increase the rate of a reaction • Optimal function of enzymes depends on co-factors • There are organic and inorganic co-factors = coenzymes • vitamin K • Mg, Mn, Zn, Ca, Fe, Cu, K, and Na

12. Enzyme Enzyme Enzyme S-Pb-S SH + Pb inactive A pollutant may inactivate the active site of an enzyme • Pb binds to –SH groups in cystein residues of proteins forming disulfide-lead bridges and poisoning the enzyme • Organophosphates inhibit acetylchlolinesterase

13. Inactivation of cofactors • Fluoride inactivates the enolase cofactor Mg • Mg cofactor • Fluoride complexes with Mg and phosphate to form FMgPO4 complex and prevents the enzyme from functioning by scavenging it’s cofactor

14. Replacement of co-factors • Be competes with Mg and Mn • Cd can replace Zn e.g. superoxide dismutase • toxic synthesis • --SH or --NH2 groups; hydrogens are replaced with CH2COOH in alkylhalide toxicants • e.g. sodium fluoroacetate • Concerted to fluorocitrate, which irreversibly binds to aconitase poisoning the enzyme

15. Secondary effects • Presence of toxicant causes production of toxic substances • Chemicals that trigger ore repress immune responses • histidine -> histamine (in Mast cells through decarboxylation) • Released to trigger immune response • Histamine is a powerful vasodialator • Increases permeability of blood vessels • Reduce blood pressure and lead to anaphylactic shock • Most extreme is vascular collapse • Antihistamines inhibit the function of histamines by structure • Carbon tetrachloride • Leads to massive release of epinephrine from sympathetic nerve • Epinephrine is a powerful hormone involved in sugar metabolism • Inhibits glucagon secretion • => inhibition of glucose uptake in muscles • Increases blood pressure • Liver damage

16. Chelation • Atom of metal is chemically bound / sequestered by ring shaped molecules • O, N, or S act as electron donor to the metal • This forms chelate rings that are extremely stable • E.g. the Fe in hemoglobin and the Mg chlorophyll are examples of chelated metals Chlorophyll a

17. Receptor mediated toxicity • Hormones are small inorganic or organic molecules that are used by the body to signal physiological states or illicit responses • Responses are very precise and specific to individual tissues • Exceedingly small quantities are needed to illicit response • Steroid hormones • lipid soluble; • androgens = testosterone and andosterone • estrogens = estradiol, estriol, estrone • Toxicants can mimic the shape of hormones • block the hormone binding site • illicit the response in lieu of the presence of the hormone • are called hormone/endocrine disruptors

18. Factors influencing the activity of toxicants • There are many pollutants in the environment • Their toxicity is influenced by • Partitioning, fate, and transport • physiochemical properties • mode of exposure • Time • Environmental factors • Interaction • Biological factors • Nutrition, starvation • Genetics • Proteins (Mixed Functional Oxidases) • Lipids • vitamins • Sex / effect on males vs. females • Disease • Behavior • Chemodynamics • Bio-availabiltiy

19. 2,3,7,8-TCDD • Not very toxic to invertebrate animals • Highly toxic to vertebrates • Ingredient in agent orange • Two modes of toxicity: • Planar molecule that fits into the active site of a receptor protein (Ah – aryl hydrocarbon receptor) • Sort of ‘velcros’ / stacks with aromatic structures in proteins Ukrainian opposition leader Viktor Yushchenko was poisoned by TCDD

20. Activity of a toxicant depends on chemical conformation • Conformation determines how much a chemical mimics a natural ligand (show table of DDT LC50s) Examples • dioxins • Irreversibly bind to DNA - mutagen • Lipid soluble – bioaccumulates • Can interact with receptors • PCBs • May act as an estrogen • Coplanar 3,3’,4,4’,5,5’-hexachlorobiphenyl (HCB) resembles 2,3,7,8-TCDD => is toxic by a similar mechanism