Aquatic Toxicity Tests and their Role in Environmental Protection … 2... · 2017-03-06 ·...
Transcript of Aquatic Toxicity Tests and their Role in Environmental Protection … 2... · 2017-03-06 ·...
1Ministry of the Environment
Aquatic Toxicity Testsand their Role in
Environmental Protection in Ontario
Environmental Chemistry and Toxicology, University of Guelph, Guelph, Ontario
February 2, 2006David Poirier
Physical Chemistry and Litigation Services SectionLaboratory Services Branch
Ontario Ministry of the Environment
2Ministry of the Environment
Introduction
• Toxicity testing 101• Types of toxicity tests• Toxicity testing vs. analytical testing • Quality control (QC) in toxicity testing• Regulations • History of aquatic toxicology in Ontario• Some ways we use toxicity testing in Ontario• Have we made any improvements?• What’s next?
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What is Aquatic Toxicology
• From a regulatory perspective it is the study of:– Environmental contaminants;
• Heat, nutrients, inorganic chemicals, organic chemicals• MIXTURES!
– And their effects;• Lethality, growth, reproduction, genotoxicity, bioaccumulation• Ecosystem changes (species abundance, diversity…)
– On water-dwelling organisms• Bacteria, plants, animals
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What are Aquatic Toxicity Tests? • Laboratory or field exposures of aquatic
organisms to a contaminant (either pure chemical or mixture)
• Use an aquatic organism to detect both the presence and effects of a toxicant
• Chemical analyses of effluents indicate presence, but not environmental/biological effects…
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High Technology vs Low Tech
HIGH analytical complexity
Narrow Chemical focus
LOW analytical complexity
High biological complexity
Broad chemical focus
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Toxicity Testing vs. Analytical Chemistry• Toxicity Testing
– Measures effects on organisms, plants and animals
– Mixtures!– Biological detectors– Animals for Research Act– Infrastructure
• Analytical Chemistry– Measures presence/absence and
concentration– Gives us an idea of what’s causing
effects (weight of evidence approach)
– Various – colorimetric, mass, etc.
Chemistry and biology work hand in hand to identify risk, and cause/source of the hazard.
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How do They Work?• Organisms respond to the three C’s:
1) Concentration (exposure)• How much toxicant is present
2) Complexation• How much of the toxicant is bound to organic or
inorganic ligands3) Competition
• What substances are present to compete at the biological receptor
• E.g., Metals…
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Ca2+
Complexation Ca2+
M2+ + L = Ca2+
Ca2+
M2+
CompetitionM2+ Ca2+
M2+
M2+ ConcentrationM2+ M2+ Ca2+
M2+ M2+ Ca2+
Ca2+
Organism Responds to the Three C's of Toxicity
ML
M metalL ligand
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Complexation M2+
M2+ + L = M2+
M2+
M2+
CompetitionM2+ Ca2+
M2+ M2+
M2+ M2+
ConcentrationM2+ M2+
Organism Responds to the Three C's of Toxicity
ML
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Test Animals/Methods• Select the correct
tools for the job• Look at the
problem/question and decide which of a myriad of methods/test species would be appropriate
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What Makes a Good Test Animal?• Depends on the purpose, but generally:
– Readily available in large numbers;– Lots known about biology/physiology;– Sensitive;– Representative;– Size;
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What Makes a Good Test Animal?• Depends on the purpose, but
generally:– Readily available in large
numbers;– Lots known about
biology/physiology;– Sensitive;– Representative;– Size;
• For compliance/legal/monitoring purposes:– Must be present in Ontario (or ?)– Must represent community to be
protected– “public appeal”– Methods must undergo strict quality
control (QC)• Culture health• Reference testing• Audits• Proficiency testing
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Types of Toxicity Tests• Static Acute Lethality Tests
– Single concentration and LC50 (lethal concentration that kills 50% of the test animals)
• Regulatory compliance tests
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Types of Toxicity Tests• Chronic/Sublethal Tests
– Survival, growth, reproduction endpoints– 7 – 8 days to conduct– Monitoring only
Fathead minnow
Ceriodaphnia dubia
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Sediment Testing • Assists with assessing areas of
historical contamination– Partitioning of contaminants
(water/sediment/biota)
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Sediment TestingChronic exposures
• 4 species, 3 exposure scenarios
• survival, growth, bioaccumulation
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Interpreting Test Results• Survival Effects
– LC50 (concentration estimated to cause mortality to 50% of the test organisms)
• Tells us how much toxicant is required to kill ½ the organisms exposed
• The higher the LC50, the lower the toxicity
– Single Concentration Test• Determines the mortality of organisms exposed to an undiluted
effluent• The higher the percent mortality, the more toxic the effluent
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Quality Control (QC)• Culture Health
– % mortality in cultures– Number of offspring/brood– Time to first brood
• Routine water quality monitoring• Standards Council of Canada
(SCC) or Canadian Association for Environmental Analytical Laboratories (CAEAL) accreditation– Third party audits– Veterinary inspections
• Under the Animals for Research Act• Negative controls with every test• Monthly QC checks
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Quality Control• Reference toxicant testing and control
chartingDaphnia magna NaCl 48 hr LC50 Control Chart
Mean and Limits Represent Antilog of Log LC50 Calculations
3000
4000
5000
6000
7000
8000
9000
10000
11000
12000
Date of Test
LC50
NaC
l (g/
L)
UCL= 10952
UWL=9300
Mean=6706
LWL=4835
LCL=4106
LC50
CV = 12.7%
UCL upper control limit
UWL upper warning limit
LCL lower warning limit
LWL lower warning limit
CV Coefficient of variance
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What do Chemists do For Us? • QC
– Measure culture water parameters– Measure food quality and contamination– Confirm reference toxicant concentrations– Supply internal proficiency evaluation (PE) samples
• Toxicity Assessment– Measure chemicals in toxic samples– Characterize unknown effluents/toxicants
• Sediment Assessment– Measure chemicals in sediments– Prepare and measure chemicals in biota (bioaccumulation)
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Refocus Our Perspective• Why are we doing what we are doing?
– More sensitive analytical techniques, new analytes, metal speciation…– Measure/monitor/reduce environmental contaminants– Balance industry with human and ecosystem health
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Regulations • Our mandate comes from:
– Ontario Environmental Protection Act (EPA)– Ontario Water Resources Act (OWRA)– Federal Fisheries Act (FA)
• “no person shall discharge a contaminant or cause or permit the discharge of a contaminant into the natural environment that causes or is likely to cause an adverse effect”(EPA section 14.1)
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History• 1960s and 70s
– Acute lethality testing with fish– Primarily petroleum, pulp and paper, mining
industries
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History• 1980s – 1990s
– Municipal/Industrial Strategy for Abatement (MISA)
– Monitoring regulation• Effluent Limits Regulations (under the
EPA)• Chemical and toxicological
characterization of effluents from >250 companies representing 10 industrial sectors
– Development of national test methods
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History – Present• Effluent Limits Regulations
– End of pipe controls– Sublethal data assessment
• Development of Provincial Water Quality Guidelines/Objectives (PWQO)
• Spill investigations• Contaminated sites
– Remedial Action Plan Sites (RAPS)– Contaminated sediments
• Toxicity Identification/Reduction Evaluations
• Method Development
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Investigations• Legal
– Belle Island Landfill Leachate• Highly toxic (48 hour LC50 to Daphnia
<0.0001%)• Toxic “soup” (1000s of chemicals
identified)• Assistance
– Ministry of Transportation• Assess toxicity of various bridge cleaning
solutions– Metro Toronto Works
• Help choose an appropriate polymer and dose to assist settling
• Wastewater Treatment Plants (WWTP)• Food Processing Plant Effluents
– Part of a large team looking at effluent characteristics, including toxicity
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Investigations• RAPS• Ecological Risk Assessments (ERA)
– Aquaculture (cage culture facilities)• High copper, zinc, nutrient,
Biological Oxygen Demand (BOD)– Aluminum smelter decommissioning
• Aluminum contamination– Pulp and paper plant
decommissioning• Mercury
– Manufacturing plant decommissioning
• Polychlorinated biphenyls (PCBs) in river sediments and biota
– Randle’s Reef • Polycyclic aromatic hydrocarbons
(PAHs), metals, BOD and ammonia
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Has There Been Improvement?Rainbow Trout Acute Lethality Results from the
Pulp & Paper Sector: 1990
50%50% Nonlethal
Lethal
n = 361
Daphnia Acute Lethality Results from the Pulp & Paper Sector: 1990
56%
44% Nonlethal
Lethal
n = 426
Rainbow Trout Acute Lethality Results from the Pulp & Paper Sector: 1998-Present
94%
6%
NonlethalLethal
n = 143
Daphnia Acute Lethality Results from the Pulp & Paper Sector: 1998-Present
91%
9%
Nonlethal
Lethal
n = 140
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Sublethal Monitoring 1998 – 2002
C. dubia Sublethal Toxicity Results from the Metal Mining Sector
71%
29%
IC25 <100IC25 >100
n = 78
P. promelas Sublethal Toxicity Results from the Metal Mining Sector
19%
81%
IC25 <100IC25 >100
n = 78
IC25 inhibitory concentration
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What’s Next?• Support PWQO and
Canadian Council of Ministers of the Environment (CCME) guideline development– Fill in data gaps (e.g.,
ammonia toxicity in cold water and low pH)
– Concern with mining and WWTP in northern communities
0.38(0.25, 0.53)
0.24(0.16, 0.35)
0.19(0.17, 0.21)
20LC50
(ConfidenceLlimit (CL))(mg/L NH3)
0.43(0.39, 0.48)
0.23(0.21, 0.26)
13LC50 (CL)(mg/L NH3)
150(100, 210)
20
137(93-202)
251(225, 280)
15
162(145, 181)
199(178, 222)
10
20LC50 (CL.)(mg/L total ammonia)
13LC50 (CL)(mg/L total ammonia)
TestTemperature
(oC)
Daphnia magna NaCl Reference 48 hr LC50 Control Chart at 20oC, 15oC and 10oC
3.5
4.5
5.5
6.5
7.5
8.5
9.5
10.5
11.5
12.5
2004
/07/20
2004
/07/22
2004
/07/24
2004
/07/26
2004
/07/28
2004
/07/30
2004
/08/01
2004
/08/03
2004
/08/05
2004
/08/07
2004
/08/09
2004
/08/11
2004
/08/13
2004
/08/15
2004
/08/17
2004
/08/19
2004
/08/21
2004
/08/23
2004
/08/25
Date
NaCl
(g/L
)
LC50 20CMean 20C=5.96LWL 20C=5.33UWL 20C=6.59LC50 15CMean 15C=7.22LWL 15C=4.97UWL 15C=9.47LC50 10CMean 10C=8.6LWL 10C=7.49UWL 10C=9.71
Effects of temperature on toxicity of ammonia to Daphnia magna cultured at different temperatures.
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What’s Next?• Method Development
– In Canada, method development is often done on a national level through guidance from Intergovernmental EcotoxicologyGroup (IGETG)
• Presently includes “tweaking” and harmonizing sediment toxicity test methods, and pH stabilization in trout tests
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What’s Next?• Monitor
persistent and bioaccumulative substances– No well
developed method available
– Rely on field studies ($$$)
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What’s Next?
• Reduce number of animals used in testing– Animals for Research
Act– E.g., cell culture assays
using fish liver or gill cell lines
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What’s Next?• Ecosystem Protection
– The types and numbers of tests used to monitor and protect the environment based on:
• Available methods• Available resources• Perceived importance• Sample size to collect
– Need a plant test!
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Plant Tests• Micro-scale
– Selenastrum microplatetechnique
• Macro-scale– Lemna
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What’s Next?• Screening tools
– Reduce workload in areas of expensive and difficult analyses required by regulation.
– Luminotox• Using algae
– Microtox• Using bacteria
– Rotox• Using rotifers
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Luminotox• Looks at chlorophyl disruption
PEC Photosynthetic enzyme complex
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Rapid Tests• Industry request
– Wish to know if a discharge is toxic without waiting 2 – 4 days for results.
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Modelling• Quantitative Structure Activity Relationships
(QSARS)– Extrapolates expected toxicities of compounds from data
from one compound in the groups and their chemical structural relationships.
• Biotic Ligand Model (BLM)– Predicts copper toxicity (to trout) based on water quality
relationships to toxicity• Toxic Equivalents/Toxic Units (TU)
– Bases toxicity of a compound on it’s relative toxicity to another similar compound
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Conclusions• Toxicity tests have the advantage of looking at
the environmental/biological effects of mixtures
• Choose the right “instrument” for the job• There have been definite improvements in
effluent quality over the past 10 years• Still a long way to go (e.g., chronic toxicity
concerns, bioaccumulation, personal care products in sewage discharges, new toxins, etc.)
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Acknowledgements• The staff of the Aquatic Toxicology Unit
for their dedication• D. Rogers (Kinectrics) for the slides on
the IQ method, and L. Novak (Stantec) for the slides on the Microtox, and algal methods.