Toxicity Testing

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Toxicity Testing

• Modern History of Toxicology
• Toxicology is Easy - Discussion
• Shape of Dose-Response Curve
• Linear vs Sigmoid
• Steep vs Flat
• Why LC50?

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Cycle of Response to Toxics
Public response/ outcry
Toxic effect of one substance documented
Legislation to regulate toxic effects
More studies of other chemicals Improved
analytical accuracy
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Modern History of Toxicology

• Industrial Revolution
• The Jungle, Wiley Bill
• 1900 - vital amines, rat bioassays
• 1919 - Alice Hamilton, Occup. Health
• 1920s- Prohibition, Bootleg, Ginger Jake
• 1930-40 - WWII Chemistry, NIH Founded

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Modern History of Toxicology

• 1938 - Sulfaniliamide Disaster, US FDA
• 1947 - FIFRA, 1st non-food or drug toxics law
• 1955 - Delaney Amendment
• 1962 - Silent Spring by Rachel Carson
• 1980s - Vastly improved analytical sensitivity
• 1996 - Delaney Replaced by FQPA
• (Food Quality Protection Act)

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Discussion - Toxicology is Easy
Toxicology is the science of poisons and deals
with how substances make organisms ill or kills
them. Thus, preventing toxic effects simply
involves identifying the concentration of a
substance that causes those effects and
preventing exposures at or above that
concentration.
What challenges or complexities may cloud the
simplicity of this commonly held perception?
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Shape of the Dose-Response Relationship
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Why LC50?
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Response
0
Low
High
Concentration (or Dose)
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Acute Toxicity Test Design

• Test Material (toxicant)
• Pure
• Commercial formulation
• Mixtures of known concentration
• Carriers/solvents
• Unknown mixtures (eg. sediment,effluent)

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• 2. Test Organism
• Most sensitive
• Most representative
• Wild species
• Rear in lab
• Known physiology
• Bred for uniformity
• Certified disease free
• Known susceptible strain

Ex Daphnia Fathead minnows Rats/Mice Animal
Cell cultures Algal cell cultures Duckweed Meal
worms Earthworms Frogs/tadpoles
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• 3. Exposure Method/Apparatus
• Oral dose/gavage
• Diet
• Intraperitoneal injection
• Inhalant
• Dermal
• Dry vial
• Static vs Flow-thru aquaria

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• 4. Experimental Design
• Sample size
• Unbiased allocation of subjects
• Test environment (temp, 02, pH, light cycle,
  food, etc)
• Negative controls (untreated, solvent/carrier)
• Positive controls (toxin with well known effect)
• Baseline measurements (size, test envt, etc)

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• 5. Range-Finding Test
• 10X progression of toxin concentration
• 3-5 individuals per concentration
• 5 concentrations plus control(s)

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• 6. Definitive Test
• Expand on meaningful conc. From range test
• 5 conc. Plus control(s)
• Geometric progression of conc. (2X or higher)
• 1 conc. Kills lt 35 1 conc. Kills gt 65
• 10 individuals per concentration
• Replicates?
• 96h, no food

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• 7. Endpoint (what response to measure?)
• Death (LC50, LD50)
• Paralysis, loss of equilibrium (EC50)
• Other sub-lethal endpoints (EC50)

Pop. growth rate Indiv. growth rate Foraging
behavior Escape behavior Learning/cognitive
Bone formation Protein production Enzyme
activity Chromosome breakage RNA synthesis
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8. Calculations

• Plot mortality vs log conc. (or dose)
• Do not include control data in curve fit
• If control mortality exceeds 10, correct w/
  Abbotts formula
• Do probit analysis for accurate LC50
• Calc. 95 confidence intervals

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(Lindane)
Diff. toxicity to diff. spp. Diff. Toxicity via
diff routes of exposure why?
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Yikes! those are my tax dollars!
What about transgenerational effects??
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Scale of Toxicological Endpoints
Increasing reality Increasing cost Increasing
uncertainty