John D. Stark, Ph.D.

Washington State University

Puyallup

Northwest Biosolids Management Association Conference

Suncadia

August 27th, 2012

Toxicology

The study of poisons

Toxicologists evaluate chemicals for their toxic effects (Toxicodynamics) and their uptake, incorporation, and elimination from the body (Toxicokinetics)

Everything is toxic in the right quantity

The dose makes the poison

The dose-response concept

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Threshold

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no effect

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Probability

LD50 versus LC50

The LD50 is the amount of toxicant per body weight or amount per organism – example: 10 milligrams/kg

The LC50 is the amount of chemical in an environmental media containing an organism such as water or air – example: 10 milligrams/liter of water

The lower the LD or LC50 is, the more toxic the chemical is because it takes less chemical to kill 50% of

a population

Acute mortality LD50’s of some common chemicals (oral - rat)

Botulism toxin <1 mg/kg Ricin <1 mg/kg Aflatoxin B1 9 mg/kg Sodium Fluoride 180 mg/kg Tylenol 338 mg/kg Diazinon 350 mg/kg Aspirin 1,500 mg/kg Malathion 2,800 mg/kg Table Salt 3,750 mg/kg Ethanol 10,600 mg/kg

Notice the importance of body weight

Acute versus chronic exposures

Acute exposures are exposures that can last from seconds to a few days

Chronic exposures last from a few days to a life time

Endpoints evaluated in toxicity evaluations Mortality

Reproduction

Weight gain – body size

Reduction of life span

Effects on nervous system

Changes in behavior

Changes in organ function

Cancer

Genetic mutations

others

The No Observable Effect Concentration or Dose (NOEC)

The lowest concentration that causes a negative effect

The Lowest Observable Effect Concentration or Dose (LOEC)

The lowest concentration of dose that causes a negative effect

Example of NOEC and LOEC Daphnia exposed to adjuvant R-11

R-11

Concentration (mg/l)

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Hormesis Favorable biological responses to low exposures to

toxicants and other stressors. A pollutant or toxicant showing hormesis thus has the opposite effect in small doses as in large doses

Hormesis doesn’t happen for all chemical or even happen often. However, hormesis indicates that exposure to some chemicals at low doses can actually be beneficial. example - alcohol

How do poisons get in the body?

Breathing

Dermal

Ingestion

How do organisms deal with poisons

Detoxification

Storage

elimination

But if the dose is too high, then damage occurs

Common poisons Arsenic

Cyanide

Mercury

Copper

Zinc

Pesticides

Flame retardants

botulinum toxin

Natural toxicants – mold spores, certain types of algae,

Arsenic

Arsenic disrupts the production of energy at the cellular level. Arsenic uncouples oxidative phosphoralation, thus inhibiting energy-linked reduction of NAD+, mitochondrial respiration, and ATP synthesis.

Arsenic sources and exposure

Naturally occurring in soil – gets into ground water

Smelting plant operations

Applications of arsenic pesticide in the past to crops

Cyanide

Cyanide inhibits cytochrome oxidase at the cellular level, preventing cells from using oxygen. This impairs the function of vital organs.

Sources:

Occurs naturally in some plants; synthesized for use in industry, mining.

Mercury Mercury irreversibly inhibits selenium-dependent

enzymes. Due to the body's inability to degrade catecholamines (e.g. Epinerphrine) a person suffering from mercury poisoning may experience profuse sweating, tachycardia (persistently faster-than-normal heart beat), increased salivation, and hypertension (high blood pressure). Methyl mercury is the most toxic form.

Sources: naturally occurring in rock and soils, released by human activity – smelting, mining, burning of coal, etc.

Copper Highly toxic in aquatic environments and has effects in fish, invertebrates, and amphibians, with all three groups equally sensitive to chronic toxicity Copper bioconcentrates in many different organs in fish and mollusks Copper sulfate and other copper compounds are effective algaecides. Toxic effects in birds include reduced growth rates, lowered egg production, and developmental abnormalities. While mammals are not as sensitive to copper toxicity as aquatic organisms, toxicity in mammals includes a wide range of animals and effects such as liver cirrhosis, necrosis in kidneys and the brain, gastrointestinal distress, lesions, low blood pressure, and fetal mortality. Sources: Pesticides, combustion of fossil fuels, automobile brake dust, mining, metal production, wood production and phosphate fertilizer production.

Cadmium Cadmium is highly toxic to wildlife; it is cancer-causing and teratogenic and potentially mutation-causing, with severe sublethal and lethal effects at low environmental concentrations Associated with increased mortality, and it effects respiratory functions, enzyme levels, muscle contractions, growth reduction, and reproduction It bioaccumulates at all trophic levels, accumulating in the livers and kidneys of fish Crustaceans appear to be more sensitive to cadmium than fish and mollusks Cadmium can be toxic to plants at lower soil concentrations than other heavy metals and is more readily taken up than other metals

Cadmium continued

Uses: smelting of other metals such as zinc, lead and copper. Used in nickel-cadmium rechargeable batteries and for metal plating. paints, plastics, catalytic converters, and metal solders. The main industries that use cadmium are metal smelting, electronics, nuclear power, paint pigment production, and other metal working and refining companies.

Sources: Industrial waste, coal or oil combustion. Released from car exhaust, metal processing industries, catalytic converters, battery and paint manufacturing, and waste hauling and disposal activities.

Zinc Survival, growth and reproduction can be adversely affected by zinc in aquatic plants and animals. Terrestrial invertebrates: Reduced survival, growth, and reproduction Vertebrates: Birds: Mortality, pancreatic degradation, reduced growth, and decreased weight gain. Mammals: cardiovascular, developmental, immunological, liver and kidney problems, neurological, hematological (blood problems), pancreatic, and reproductive problems

Zinc continued

Sources: Mining, smelting metals (including zinc, lead and cadmium), steel production, burning coal, galvanized fences, roofs, tire wear.

Pharmaceuticals Many pharmaceuticals enter the environment. Most of

these come from human activity such as flushing unused products down toilets and the excretion of these products through urination, and production of live stock.

Over 3000 prescription pharmaceuticals are registered for use in the U.S. and only an extremely small sub-set of these have been monitored in water.

Common pharmaceuticals found in our surface waters and salt water

Antimicrobials – antibiotics

Anti-depressants

Pain killers/anti-inflammatory products

Estrogens

Caffeine

Sources: animal husbandry operations have been estimated to

represent a significant source, and they are derived primarily from spills from anaerobic manure storage lagoons, manure fertilization of farm fields, run-off from farm fields, discharge from aquaculture operations, and dust. Excretion by animals.

leaking septic systems, waste water treatment plants, landfills.

Botulinum toxin is a protein and neurotoxin produced by the bacterium, Clostridium botulinum. Botulinum toxin can cause botulism, a serious and life-threatening illness in humans and animals. Botulinum toxin causes paralysis

Tobacco-Nicotine

Nicotine - a neurotoxin –causes nerve excitation – can be beneficial – alzheimers

Tars – carcinogens

Tobacco use leads to disease of the heart and lungs. Smoking is a major risk factor for heart attacks, strokes, chronic obstructive pulmonary disease including emphysema, and chronic bronchitis. Cancer of the lungs, larynx, mouth and pancreas. Can cause high blood pressure.

Alcohol A psychoactive drug- a depressant of the central

nervous system. Has a complex mode of action and affects multiple systems in the brain, the most notable one being its agonistic action on the GABA receptor

Acute and chronic effects

Acute – drunkenness

Chronic – liver disease, brain damage,

Ethanol's toxicity is caused by its primary metabolite acetaldehyde and secondary metabolite acetic acid. Acetaldehyde is toxic -- Symptoms of exposure to this compound include nausea, vomiting, headache. These symptoms may not happen immediately. It has a general narcotic action and large doses can even cause death by respiratory paralysis. It may also cause drowsiness, delirium, hallucinations and loss of intelligence. Acetaldehyde derived from the consumption of ethanol binds to proteins to form adducts that are linked to organ disease. Acetaldehyde generated endogenously from alcoholic beverages is a Group I human carcinogen.

Other chemicals of concern

1. PCB’s 2.PAH’s 3.Phthalates

PCB’s PCBs are mutation-causing, cancer-causing, and teratogenic Readily absorbed through the gut, respiratory system, and skin in mammals and will concentrate in the liver, blood, muscle, adipose (fatty) tissue, and skin In aquatic systems, increased toxicity is associated with increasing exposure, younger developmental stages, crustaceans, and lower chlorinated biphenyls Toxic effects in avian species included the following: morbidity, tremors, upward pointing beaks, muscular incoordination, and hemorrhagic areas in the liver

PCB’s continued Sublethal effects include delayed reproduction and chromosomal aberrations; courtship and nestbuilding behavioral impairments in birds reduced hatchability in chicken eggs; and decline in sperm concentration in American Kestrels Growth reduction in brook trout; reduced egg survival and reduced fertilization success in flounder, minnows, sea urchins; and complete reproductive failure in brook trout Cancer-causing effects and biochemical perturbations were observed in trout liver cells and marine fishes; with anemia, hyperglycemia, and altered cholesterol metabolism in brown trout fed diets with 10 PPM PCBs

PAH’s Highly potent carcinogens that can produce tumors in some organisms at even single doses Effects on benthic invertebrates include inhibited reproduction, delayed emergence, sediment avoidance, and mortality Fish exposed to PAH contamination have exhibited fin erosion, liver abnormalities, cataracts, and immune system impairments leading to increased susceptibility to disease

Phthalates Phthalates are a class of widely used industrial compounds known technically as dialkyl or alkyl aryl esters of 1,2-benzenedicarboxylic acid They have many uses such as components of vinyl, nail polish, perfume, hairspray etc. Phthalates have been in widespread use over the past several decades because of their many beneficial chemical properties They are now ubiquitous, not just in the products in which they are intentionally used, but also as contaminants in just about everything Approximately 1 billion pounds of phthalates are produced per year produced worldwide

Diethylhexyl phthalate (DEHP) highly toxic to the reproductive system of male offspring of rats

Pregnant female were exposed and effects measured in their

offspring DEHP induced high levels of testicular and epididymal

abnormalities, including atrophy and agenesis. A striking effect of DEHP was noted in 8-day old pups. Several males from different litters displayed hemorrhagic testes that were visible by gross examination of the inguinal region.

Toxicity of phthalates

Exposure to phthalates increases the sensitivity of the immune system, leading to heightened allergic responses This occurrs after exposure to concentrations well below EPA’s “low effect level” of 19 mg/kg/day Takano, H, R Yanagisawa, K-I Inoue, T Ichinose, K Sadakano, and T Yoshikawa. 2006 Di-(2-ehylhexyl) Phthalate Enhances Atopic Dermatitis-Like Skin Lesions in Mice Environmental Health Perspectives 114: 1266-1269.

Mechanisms of phthalate ester toxicity in the female reproductive system Environmental Health Perspectives, Feb, 2003 Lovekamp-Swant and Davis

In vivo, DEHP (2 g/kg) causes decreased serum estradiol levels, prolonged estrous cycles, and no ovulations in female adult rats.

Bisphenol A Used to make plastics, resins, expoxys. BPA exhibits estrogen-like properties, raising concerns about its presence in consumer products and foods contained in such products. Banned in baby bottles in the U.S. Bisphenol A is a weak endocrine disruptor, which can mimic estrogen and may lead to negative health effects. Early developmental stages appear to be the period of greatest sensitivity to its effects, and some studies have linked prenatal exposure to later physical and neurological difficulties.

Flame retardants (polybrominated diphenyl eters -PBDEs) are chemicals used in thermoplastics, thermosets, textiles and coatings that inhibit or resist the spread of fire. Nearly all Americans tested have trace levels of flame retardants in their body. Have been shown to have significant reproductive toxicity inhumans. Children who had higher cord blood concentrations of PBDEs scored lower on tests of mental and motor development at 1–4 and 6 years of age.

Pesticides

Chemicals designed to kill organisms

Examples: Insecticides, herbicides, rodenticides, fungicides, bactericides, algacides, etc.

Insecticides

Insecticides – first were arsenic-based, then nicotine. DDT, organophosphates, carbamates and pyrethroids were developed later

Now, - natural products – spinosad, neem and synthetic nicotines

DDT and the Chlorinated Hydrocarbons

Examples – DDT, DDD, DDE, chlordane, lindane, aldrin, dieldrin, - axonic poisons –keep sodium channel open causing nerve excitation followed by paralysis

Organophosphates and Carbamates

Work in the nervous system by binding an enzyme, acetylcholine esterase. Causing repeated firing of nerves until they run out of energy and cease to function. Results in heart failure and organ shutdown.

Examples:

Ops – malathion, diazinon, guthion

Carbamates – Sevin (carbaryl), carbofuran

Pyrethroids

Work like DDT- axonic poisons –keep sodium channel open causing nerve excitation followed by paralysis

Examples – permethrin, bifenthrin, resmethrin, cyermethrin, deltamethrin

Neonicitinoids Imidacloprid, acetamiprid, clothianidin,

dinotefuran, thiacloprid, thiamethoxam

Acts like nicotine fitting in nicotinic receptor on axon causing nerve firing and eventual paralysis

Natural products

Neem

Spinosad

Herbicides Various modes of action - plant hormone mimics, lipid

biosynthesis inhibitors, amino acid inhibitors.

Examples – Roundup (Glyphosate), Casoron, 2,4-D

Roundup - inhibits an enzyme involved in the synthesis of the aromatic amino acids

Casoron inhibits cellulose synthesis

2,4-D – a synthetic plant hormone that causes uncontrolled growth

Fungicides Metabolic poisons, inhibitors of cell division, protein

synthesis inhibitors, nucleic acid synthesis inhibitors

Examples – chlorothalonil (Bravo, Daconil), Copper sulfate, Mancozeb (Dithane),

Rodenticides Blood thinners - warfarin

Metal phosphides - The acid in the digestive system of the rodent reacts with the phosphide to generate the toxic phosphine gas

Calciferols (vitamins D) – causes increases in calcium in the body - free calcium levels are raised sufficiently that blood vessels, kidneys, and the stomach wall and lungs are mineralised/calcificated (formation of calcificates, crystals of calcium salts/complexes in the tissues, damaging them), leading further to heart problems

Mixture toxicity

Conclusions