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Biol 354 Midterm Notes

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University of Waterloo
BIOL 354
Gerald Tetreault

Lecture 1 Toxicology: study of harmful effects incurred by living organisms from exposure to substances which are foreign to them Xenobiotic: any substance which is foreign to an organism they can be useful or toxic Anthropogenic: caused/produced by humans; describes origin of a compound which would otherwise not occur normally Toxicant: poisonous agent, produces biological effects ex. PCBs, PAHs, metals Toxin: type of toxicant in which its produced by a living organism ex. snake venom, mycotoxin, tetrodotoxin (sodium channel blocker which paralyzes the muscles while the victim remains fully conscious and dies from asphyxiation) Paracelsus -> the dose makes the poison Ecotoxicology: study of the impacts of pollutants upon the structure and function of living organisms and ecological systems -one must first be exposed to a toxicant for there to be any effect 1. release into environment 2. exposure to target organism 3. taken up by target organism 4. response in target organism Inorganic = metals, ammonia Organic = dioxins, furans, PAHs Chimney Sweep -> scrotal cancer from PAHs formed from incomplete combustion demonstrated by Percival Pott Mad Hatters -> Hg to cure beaver pelts, mercury vapour is fat soluble and accumulates in the nerve lining; it blocks the degradation of catecholamines such as epinephrine -> causes sweating, excited state -pollutants are not necessarily toxicants Lecture 2 – Contaminants from Source to Receptor to Impact Fate of contaminants – origins, concentrations, partitioning and persistence of contaminants Effect of contaminants – evaluate biotic and abiotic consequences Rachel Carson -> environmental revolution, evidence of harmful effects of chemicals on human and wildlife SSRI= selective serotonin uptake inhibitor -> largely reduced male fish (Japanese Medaka) approaches, circling, copulation even though it slightly increased egg production (fluoxetine in muscle and adipose tissue) 1. Generation of a contaminant 2. Release to environment 3. Transport to a biotic receptor 4. Exposure at sufficient concentration and duration to produce a 5. Biological response Direct – point source discharge ex. industrial, sewage discharge Indirect – indirect point source discharge ex. agricultural or urban runoff, car exhaust Exposure Routes 1. Oral – administration by mouth or diet -unit expressed as a relative weight to the organism; 1mg/L = 1 ppm=1mg/kg 1ug/L=1ug/kg=1ppb 2. Injection – dose administered intravenous, subcutaneous or intra-peritoneal 3. Topical – applied to skin 4. Respiratory – inhalation from lungs or gill uptake (unit expressed as a relative weight to volume of air or water) Measuring Toxicity Bioassay – determines the effects of a toxicant on organisms Test organisms – rainbow trout, water flea, fathead minnow, earthworms (soil) or plants -requires sufficient exposure time to allow a response to occur from a range of doses and the dose must be enough to develop a response within a given exposure time period Classification of Toxic Reponses 1. Lethal – death 2. Sub-lethal – molecular, biochemical, individual species or community levels a. Individual i. biochemical -> enzyme inhibition (ex. acetylcholinesterase inhibition by organophosphates) ii. Hormonal -> changes in cortisol levels iii. Physiological -> changes in somatic growth or reproduction b. Species -> change in population structure c. Community -> change in species abundance and diversity Timing of Response -depends on experimental conditions, test organism, dose/concentrations 1. Acute – severe stimulus (high potency/concentration of toxicant) to quickly bring about a response -short term; seen within 96 hours -common endpoint in routine lethal toxicity testing (calculation of 96Hr LC50) 2. Sub Acute – less severe stimulus than acute, response takes longer and may become chronic (less than 10% of organism’s life) 3. Chronic – slowly brings about a response, long-term test of at least 10% of organism’s life span 4. Cumulative – response to repeated exposure to a contaminant often from human occupations (farming) -similar to chronic, but doesn’t have to occur over at least 10% of organism’s life -exposure levels may or may not fluctuate over time -if acting on a common biological pathway (additive), multiple chemicals could produce a cumulative response such as several pesticides that have a similar mechanism of action 5. Delayed – response does not occur until well after exposure Ex. Diethylstilbestrol (DES) in daughters may develop vaginal cancer in adulthood -> 100x greater risk developing vaginal cancer, used as morning after pill, was used in chicken feed, 125mg=700 BC pills, blocks the uptake of testosterone in cancer cells, -triclosan = dangerous anti-bacteria in soap (affects fish swimming ability, muscle strength, disrupts hormone balance, increase infertility) The next generation’s sufferings from exposure to toxicants… -increased birth defects -reduced fertility -reduced sperm counts -earlier puberty onset in girls Bioassay Results 1. Keep time constant and vary dose or concentration a. LC50 = lethal concentration for 50% of organisms b. LD50 = lethal dose for 50% of organisms 2. Keep dose or concentration constant and vary time a. LT50 = lethal time for 50% of organisms Other endpoints: enzyme induction, inhibition of growth/metabolism of enzyme, EC50 or ED50 (effect dose or concentration) -want to keep all conditions constant other than the toxicant concentration or time 1. Biotic – species, sex, age, size, nutrition 2. Abiotic – exposure route, partitioning, hardness, pH, oxygen, temperature, light Lecture 3 – Sources & Fate of Contaminants Pollutant Sources 1. Accidental spills – nuclear accidents, oil spills, fires, train crashes 2. Intentional spills – deliberate or routine and from either direct or indirect sources -salt, fertilizers, pesticides -urban storm water/runoff -atmospheric entry of aerosols/CFCs (chlorofluorocarbon) -combustion/incineration – PAH and PCB formation -landfill contaminants leak into soil and groundwater -septic tank leaks Sewage Treatment Plants -sewage has decomposable organic matter, microbes, nutrients, pharmaceuticals, etc. -discharge large volumes of imperfectly treated waste, often with high ammonia concentrations 1. Primary treatment -> removal of solids through a settling process, wastewater pumped to detention ponds 2. Secondary treatment -> redox reactions with BOD, COD (chemical oxygen demand) and bacteria via activated sludge or biological filters 3. Tertiary -> chemical removal of specific components – nitrification (NH3 -> NO2 -> NO3), sand filtration, chlorination then de-chlorination with sodium bisulphite (NaHSO3) Problems with STPs -toxins/toxicants can kill bacteria in the sludge during secondary treatment -residual solids must be disposed of in landfills, farmland or incinerated -sludge can contain lipophilic contaminants and metals -effluent may contain toxins, toxicants and other biologically active compounds which plants are not designed to remove -in many areas, stormwater is not fully separated from sewage -> downstream there can be raw sewage in drinking water source Important Physical and Chemical Properties Help predict the fate in the absence of other information -water solubility -pKa, Ka -octanol-water partition coefficient -sediment-water partition coefficient -vapour pressure, Henry’s Law (Kh function of temperature, solublity) -bioconcentration Temperature fluctuates the most in the environment, but pressure is also important Equilibrium Partitioning 0 law of thermal equilibrium -> no net heat flow in either direction, temperatures are equal 1 law thermodynamics – conservation of energy, it is not created or destroyed nd 2 law thermodynamics – systems seek to minimize their free energy Example. At equilibrium partition coefficient C1/C2 = Cair/Cwater = K ->awatio of the concentration of benzene in air and water Note that K is temperature-dependent. Ka = [A-][H+]/[HA] pKa=-log(Ka) (charge(pH-pKa)) weak acid/base: % ionized = 100/(1 +10 ) where charge =1 for bases and -1 for acids vapour pressure = solubility in air Cairmol/volume = n/V Looking at the ideal gas law: PV=nRT At equilibrium C air/V = P/RT Henry’s Law Constant H=p/C =(partial pressure in air)/(conc. in water) w Contaminants with low water solubility: H=P/S = vwpour pressure/water solubility -water solubulity (Sw)= max amount of substance that can dissolve in water at a given temperature or pH -solubility of a gas in a liquid is proportional to the pressure of that gas above the liquid for a given temperature H=K aw /Cair water H’=(P/RT)/C water P/RTC water/S wT KawH/RT Therefore, K aw dependent on vapour pressure as well as water solubility and temperature. Kow is the measure of hydrophobicity; diffusion from high chemical potential (µ) to low K =C /C at equilibrium ow octanolwater Determining K ow -equal volume of distilled water and octanol, add known volume of chemical and mix well Kow = concentration of chemical in octanol/Concentration of chemical in water High K ow= lipophilic/hydrophobic and low mobility Low K = lipophobic/hydrophilic ow -when pH < pKa, the compound is primarily neutral (more likely to be in octanol) and the apparent K ow decreases at high pH because the compound turns into its ionized form and is more water soluble -can graph contaminants on a polarity (lipophilic to hydrophilic) vs volatility chart Kp= water partition coefficient = Csoil water (mg/Kg)/(mg/L) = L/kg -can multiple by soil density of 2.5kg/L to become dimensionless -organic chemicals partition into the organic component of soils, sediments, etc. -Kp can be related to the organic carbon partition coefficient by knowing the fraction of organic carbon K = fK where f=0.02-0.05 most often p oc Fugacity models -> can incorporate the bioconcentration and bioaccumulation of chemicals in biota, require info. on diet and lipid content -generally, partition coefficients between fish and water are similar to K ,owish are about 5% lipid by volume and octanol is used as a lipid surrogate -contaminants partition in the environment based on: chemical properties and properties of the environment Long-Range Transport -contaminants can travel far from their origins via air and water -stratosphere suffering from ozone depletion due to CFCs -arctic -> contamination of wildlife and people Lecture 4 – Inorganic Pollutants -oldest water found is 2.6 billion years old! Inorganic pollutants include: metals, metalloids (As), non-metallic inorganic (ammonia), inorganic gases (CO 2 -when defining inorganic vs organic compounds, organic compounds have at least one C-H bond -metals can be required by the organism, toxic or both depending on the metal and its concentration 3 -heavy metals are defined as having a density greater than iron (7.874 g/cm ) -metalloids have some properties of both metals and non-metals (Se, As, B, Si, Ge, Sb, Te, Po) Common Metals & Metalloids of Concern -aluminum (Al), arsenic (As), antimony (Sb) -Barium (Ba), Beryllium (Be), Boron (B) -Copper (Cu), Cadmium (Cd), Chromium (Cr), Cobalt (Co) -Iron (Fe), Lead (Pb), Manganese (Mn), Molybdenum (Mo) -Mercury (Hg), Nickel (Ni), Strontium (Sr) -Selenium (Se), Silver (Ag), Tin (Sn), Thallium, (Tl), Vanadium (V), Zinc (Zn) Aluminum Under low pH it becomes Al which kills aquatic species Arsenic Used in a variety of things such as preservatives, gold mining -> carcinogenic and highly toxic, metalloid, found in agriculture, lumber processing Cadmium Tanning process, Cr is carcinogenic, heavy metal, may be found in processed sludge from anthropogenic waste, lumber processing Copper Control algal growth, easily binds to dissolved organics in solution (reducing bioavailability), toxic at high concentrations, tanning process, lumber processing Lead Used for batteries, causes anemi2+and neurological dysfunction with chronic exposure, present in natural waters at Pb , heavy metal Mercury Methylation by sulfate-reducing microorganisms (in both aerobic and anaerobic sediments) increases bioavailability, heavy metal, found in agriculture, fluorescent bulbs, thermometers, dental fillings Nickel At high concentrations it is both toxic and carcinogenic Selenium By-product of mining, volcanism, modification by microorganisms makes them less toxic than in their pure form, metalloid Zinc Protective coatings and prevents corrosion, less toxic than most metals Inorganic NOx, SO 2eact in atmosphere to produce low pH precipitation or acid rain gases Nutrients Nitrate can cause methemoglobinemia (reacts with hemoglobin, cannot carry oxygen); ammonia toxicity is dependent on pH Excess nutrients -> eutrophication -> algal bloom -> oxygen depletion -> fish kills PAHs Has 2+ aromatic rings Includes naphthalene, Benzo[a]pyrene, pyrene ; forms from the incomplete combustion of organic matter, metabolized by vertebrates with CYP but accumulates in invertebrates -metals such as copper, iron and zinc are often required 3+ 2+ 2+ -some metals act as enzyme cofactors, such as: Fe (hemoglobin), Mg (chlorophyll), Cu (mitochondria) -Na+ and K+ used for osmoregulation and membrane potential, Ca2+ for muscle contraction -classification of metals based on preferential binding to certain ligands Class A Metals -complex with O>N>S -oxygen seeking ex. Ca2+, Na+, Mg2+, Pb -associate with carboxyls (-COOH), carbonyls (-C=O), alcohols (-OH) and phosphates (H PO o2 P4 ) 4- -target phospholipids in cell membranes and nucleic acids (DNA/RNA), ATP and ADP Class B Metals -complex with S>N>>O -sulfur/nitrogen seeking ex. Cu2+, Cd2+, Hg2+ -associate with sulfhydryl (-SH), disulfides (-S-S), thioethers (-S-R) and amine (-NH2) -target amino acids, proteins and free nucleotides Example. Yellowknife’s Giant Mine -> $1 billion to remediate arsenic-loaded contamination -gold mine from 1940s to 2004, over 200,000 tonnes Arsenic trioxide (water soluble dust product) released over the years -to release the gold it had to be roasted at very high temperatures and released As-gas as a by-product -no known way to remove AsO from3environment; can freeze it with CO (carbon monoxide) to prevent seepage -AsO 3an cause kidney and liver failure, cancer; a lethal dose is only 120mg Sources of Metal Pollution -natural leaching -lead-acid batteries, bullets, weights, lead and tin-based paints, toys, boats -metals increase in abundance due to human activity, metal mining and weathering of metal-bearing rock; can be found in soil, sediment, surface water -agriculture may contain Hg or As, as well as volcanic sources Neurotoxin: rapidly absorbed into bloodstream and accumulates in soft tissues and bones; affects central nervous system, cardiovascular system, kidneys, immune system, reduced cognitive capacity -lumber -> pressure treated wood may contain Cr, Cu and As -tannery -> leather processing uses Cu2+ and Cd2+ Metal Toxicity -bind to molecules in organisms and alter their function ex. bind to enzyme, blocking active site -interferes with the uptake of essential metals -generation of reactive oxygen species; can oxidize the lipid bilayer leading to decreased membrane fluidity or damage DNA Organometals -more likely to enter organisms 1. tributyl tin (TBT) -> antifouling compound -used in water environments -paints contain TBT -prevents build-up of material on bottom of boats 2. Tetraethyl lead -> used in gasoline -most of Pb in city dust from this -organometallic compounds show enhanced toxicity and bioconcentration over the ionic forms 3. Methyl mercury -> from biomethylation -ability to bioaccumulate -Hg has no biological function -methylation occurs at the bottom of lakes; often hydrodams increase methylation (anoxic zones) -SO 2-reducing bacteria transform inorganic Hg into methyl mercury which is lipophilic 4 -neurotoxin Ammonia + -ammonia NH (un3onized), ammonium NH (ionized) 4 -most tests look at total ammonia -common cause of fish kills, burn of plants Sources: landfills, fish farms, refrigerants, pulp and paper mill, STPs, farms -toxicity mostly from NH 3 -% NH i3crease with increase in temperature and pH (over pH 7.4 mostly ammonia) 1. free diffusion – determined by pressure gradient (NH ) 3 2. active transport - of NH into water in exchange for Na + 4 3. ammonium (NH ) ca4not diffuse through membrane -ammonia toxicity causes neural damage, disturbance of electrochemical gradients, enzymatic reactions, intracellular pH changes Other inorganics: Chlorine (highly toxic and reactive), cyanide (produced in gold mining and metals processing; affects oxidative phosphorylation used to produce ATP in mitochondria) Lecture 5 – Organic Pollutants -generally low persistence except for persistent organic pollutants (POPs), persistent bioaccumulative and toxic (PBTs) -many modes of action, compounds being invented and mass produced Includes… -petroleum hydrocarbons -polycyclic aromatic hydrocarbons (PAHs) ex. benzo[a]pyrene BaP -polychlorinated biphenyls (PCBs) -dioxins and furans ex. TCDD -volatile organic compounds (VOCs) -chlorinated/brominated compounds -pesticides -pharmaceuticals (not regulated with environmental health in mind) Petroleum Hydrocarbons -crude oil (unrefined) - made up of numerous aromatic (ring structure) and aliphatic (non-ring) compounds between C5 to C34 -highly lipophilic -abundant in environment due to human activity -low MW compounds are volatile but more toxic (can enter membranes more easily) -high MW compounds absorb onto sediments/soils and have reduced bioavailability but are persistent and eventually weather over time PAHs (polycyclic aromatic hydrocarbons) -consist of 2 or more aromatic rings ex. naphthalene -natural sources from volcanoes, forest fires but in low levels (products of combustion) -anthropogenic sources include fossil fuels, smoke stacks, automobile exhaust, tobacco, metal refining -16 major PAHs, but there are over 50 -low water solubility, lipophilic (absorbed and stored in lipids) -low volatility for large PAHs -rapidly partitions in sediments -metabolized and excreted rapidly by higher vertebrates (relatively unreactive) -main environmental sinks in soils and sediments which alters bioavailability (less available) PCBs (polychlorinated biphenyls) -stable, inflammable, non-conducting, water insoluble, chemically inert -use has been reduced, but are very persistent and hydrophobic -209 isomers (congeners) -no natural formation, all anthropogenic -chronic effects, interferes with endocrine system of vertebrates Dioxins and Furans -chlorinated organics -low water solubility, highly lipophilic, bioaccumulate -byproduct of chlorination and bleaching processes -polychlorinated dibenzodioxins (PCDPs) = dioxins -polychlorinated dibenzofurans (PCDFs) = furans -72 different isomers -persistent & hydrophobic -not readily metabolized and excreted due to high chlorination, making it have a high bioconcentration -specific isomers are very toxic to mammals ex. 2,3,7,8-TCDD as they can interfere with the endocrine system and it is a contact carcinogen Sources include: pulp and paper mills (chlorine bleach), metal refineries, manufacturing of PCBs, pentachlorophenol, pesticides ex. 2,4-D and 2,4,5-T Toxic Equivalent (TEQ) = ∑C * TEF (max 1) -> measures the toxicity of dioxins and furans relative to the n n most toxic form: 2,3,7,8-TCDD -fish tend to have lower gonad sizes, liver had increased function (higher TEQ) Lecture 6 Case Study: Pulp and paper mill – wood composed of cellulose, hemicellulose, lignin, wood extractives -lignin is removed and cellulose used to make pulp -the Kraft process removes the greatest amount of lignin using bleaching which requires Cl and ClO 2 -effluent doesn’t mix well with water making fish exposed to high levels -some dioxins partition into the sediment where they may affect life after the mill is gone Ex. White sucker fish -> Great Lakes, benthivores, small niche, live up to 20 years, lots of historical data Experiment -> determine if TEQs remain elevated above reference levels in white sucker following process changes in pulp mill in Terrace Bay, ON -> Mountain Bay as a reference and Moberal Bay is exposed -> obtain sediment cores to view dioxins partitioned Pesticides -biological/chemical agent that kills unwanted pests -want it to kill quickly, be effective, break down into non-toxic products, cheap manufacture -closest is glyphosate (Round Up) Include: herbicides, insecticides, fungicides, rodenticides -warfarin kills rodents,
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