United States
Environmental Protection
Agency
Great Lakes
National Program Office
77 West Jackson Boulevard
Chicago, Illinois 60604
EPA 905-R93-006
December 1993
*>EPA Assessment and
Remediation
Of Contaminated Sediments
(ARCS) Program
BIOLOGICAL AND CHEMICAL
ASSESSMENT OF CONTAMINATED
GREAT LAKES SEDIMENT
•) United States Areas of Concern
9 ARCS Priority Areas of Concern
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BIOLOGICAL AND CHEMICAL ASSESSMENT OF
CONTAMINATED GREAT LAKES SEDIMENT
Project Officers:
Christopher G. Ingersoll, Denny R. Buckler,
Eric A. Crecelius1, and Thomas W. LaPoint2
National Fisheries Contaminant Research Center
U.S. Fish and Wildlife Service
4200 New Haven Road
Columbia, MO 65201
1Battelle Northwest Laboratory
Marine Science Laboritories
439 West Sequim Road
Sequim, WA 98382
2current address: Clemson University
One TIWET Drive
Pendelton, SC 29670
Funded by USEPA Great Lakes National Program Office
USFWS Contract Number DW14933874-1
Battelle Contract Number 89934235-0
Project Officer:
David C. Cowgill
Great Lakes National Program Office
U.S. Environmental Protection Agency
77 West Jackson Boulevard
Chicago, IL 60604
U.S. Environmental Protection Agency
to' :
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NOTICE
Mention of trade names or commercial products does not constitute
endorsement or recommendation for use.
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ABSTRACT
Section 118 (c)(3) of the Clean Water Act (CWA), as amended by the Water
Quality Act of 1987, authorized the USEPA Great Lakes National Program Office
(GLNPO) to carry out a 5-year project dealing with the Assessment and
Remediation of Contaminated Sediments (ARCS) at selected Great Lakes Areas of
Concern (AOC). Sediment assessment procedures performed by the National
Fisheries Contaminant Research Center (NFCRC) on samples from Indiana Harbor,
Buffalo River, and Saginaw River AOCs included: Elutriate Toxicity Tests
(Chapters 2 and 3), Whole Sediment Toxicity Tests (Chapter 4), Benthic
Community Structure (Chapter 5), and Mutagenicity (Chapter 6) and Genotoxicity
(Chapter 7) Assays.
Sediment samples collected from Indiana Harbor were severely
contaminated compared to either Buffalo or Saginaw River based on sediment
toxicity and chemistry. While sediment samples from Saginaw River were
generally less contaminated compared to Buffalo River, considerable spatial
variability in contamination was evident at all three AOC. About 41% of the
elutriates samples prepared from these sediments were designated as toxic
using either Daphnia magna or Microtox assays. Interpretation of toxicity
using Selenastrum capricornutum was complicated by variable nutrient and
inorganic carbon concentrations in the elutriate samples.
About 68% of the whole sediment samples were toxic to Hyalella azteca.
Chironomus riparius. or Chironomus tentans in 10-d to 28-d exposures.
Fourteen- and 28-d exposures with Hyalella azteca which monitored effects on
survival, body length, and sexual maturation, identified a higher proportion
fv of toxic samples compared to either 14-d exposures with Chironomus riparius or
10-d exposures with Chironomus tentans.
Oligochaetes and chironomids were the dominant taxa at each AOC
'?, indicating impacted benthic communities. Average number of midges with mouth
part deformities ranged from 45 to over 77% in samples from the AOCs.
Artificial substrate samplers which were colonized in situ sampled a more
s diverse taxa compared to benthic grab samplers.
^ Over 90% of organic extracts from sediment samples were classified as
^ mutagenic or genotoxic using Ames and Mutatox assays. Toxicity of the organic
extracts complicated interpretations in the Ames assay, but was not a problem
-, in the Mutatox assay.
^ Each sediment sample contained a complex mixture of inorganic and
organic contaminants. Additional studies are required to determine specific
contaminants that may be causing adverse effects (e.g., sediment spiking and
Toxicity Identification Evaluations).
Data from these studies are to~ be evaluated with the Sediment Quality
Triad which will integrate data from laboratory exposures, benthic community
structure, and chemical analyses to provide complementary evidence for the
degree of pollution-induced degradation in aquatic communities at each AOC.
Results of the Triad evaluations will be discussed in a later report.
This report was submitted in fulfillment of USFWS Contract Number
DW14933874-1 by the National Fisheries Contaminant Research Center, U.S. Fish
and Wildlife Service and Battelle Contract Number 89934235-0 by Battelle
Northwest Laboratories under partial sponsorship of the U.S. Environmental
Protection Agency. This report covers a period from 03/01/89 to 11/10/92, and
work was completed as of 11/10/92.
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CONTENTS
Abstract [[[
List of Figures [[[ v
List of Tables [[[ viii
List of Appendices [[[ xiii
Acknowledgment [[[ xi-v
Introduction [[[ xv
Conclusions and Recommendations ......................................... xvii
Chapter 1: Overview
Ingersoll, C.G., D.R. Buckler, E.A. Crecelius,
and T.W. La Point .......................................... 1-1
Chapter 2: Elutriate Toxicity Tests: Daphnia magna and Microtox
Coyle, J.J., E.A. Crecelius, andC.G. Ingersoll ............. 2-1
Chapter 3: Elutriate Toxicity Tests: Selenastrum capricornutum
Hall, N.E., T.W. La Point, P.R. Heine, and J.F. Fairchild .. 3-1
Chapter 4: Whole Sediment Toxicity Tests
Nelson, M.K. , J.J. Coyle, L.B. King, N.E. Kemble,
E.A. Crecelius, and I.E. Greer ............................. 4-1
Chapter 5: Benthic Community Structure Evaluations
Canfield, T.J., T.W. La Point, M.C. Swift, G.A. Burton,
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LIST OF FIGURES
CHAPTER 1: OVERVIEW
Figure 1.1 Map of Indiana Harbor AOC.
Figure 1.2 Map of Buffalo River AOC.
Figure 1.3 Map of Saginaw River AOC.
CHAPTER 2: ELUTRIATE TOXICITY TESTS: DAPHNIA MAGNA AND MICROTOX
Figure
Figure
Figure
Figure
Figure
2.1 Concentrations
prepared from
2.2 Concentrations
Indiana Harbor
samples.
2.3 Concentrations
prepared from
2.4 Concentrations
prepared from
2.5 Concentrations
prepared from
of select metals in elutriates and pore waters
Indiana Harbor whole sediment samples.
of unionized ammonia in elutriates prepared from
Buffalo River, and Saginaw River whole sediment
of select metals in elutriates and pore waters
Buffalo River whole sediment samples.
of select metals in elutriates and pore waters
Saginaw River (first survey) whole sediment samples.
of select metals in elutriates and pore waters
Sagiaaw River (third survey) whole sediment samples.
CHAPTER 3: ELUTRIATE TOXICITY TESTS: SELENASTRUM CAPRICORNUTUM
Figure 3.1 Toxicity of elutriate samples to £. capricornutum.
Figure 3.2 Hypothetical dose-response curves for stimulatory and non-
stimulatory samples.
CHAPTER 4: WHOLE SEDIMENT TOXICITY TESTS
Figure 4.1 Increase in body length amphipods for the 28-d exposures.
Figure 4.2 Relation of summed total metal concentrations to the survival of
each species.
Figure 4.3 Relation of summed total PAH concentrations to the survival of
each species.
Figure 4.4 Comparison of Hyalella azteca test sensitivity.
CHAPTER 5: BENTHIC COMMUNITY STRUCTURE EVALUATIONS
Figure 5.1 Artificial substrate samplers used to collect aquatic
invertebrates at select stations from each Area of Concern.
Figure 5.2 Relation of the summed total simultaneously extracted metals (Cd,
Cr, Cu, Ni, Pb, Zn) concentration (jxM/dry g) to mean total
invertebrate abundance (number/m2) across all Areas of Concern.
Figure 5.3 Relation of the summed total PAH concentration (/ig/dry g OC) to
mean total invertebrate abundance (number/m2) across all Areas of
Concern.
Figure 5.4 Relation of the summed total PCB concentration (ng/dry g OC) to
mean total invertebrate abundance (number/m2) across all Areas of
Concern.
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5.5
5.6
Figure
Figure
Figure
Figure
Figure
Figure 5.10
5.7
5.8
5.9
Figure
Figure
Figure
5.11
5.12
5.13
Figure 5.14
Figure
Figure
Figure
5.15
5.16
5.17
5.18
Figure
Figure
Figure 5.20
5.19
Figure 6.1
Figure 6.2
Figure 6.3
Figure 6.4
Relation of the summed total simultaneously extracted metals (Cd,
Cr, Cu, Ni, Pb, Zn) concentration (/^M/dry g) to mean Oligochaeta
abundance (number/m2) across all Areas of Concern.
Relation of the summed total simultaneously extracted metals (Cd,
Cr, Cu, Ni, Pb, Zn) concentration (/*M/dry g) to mean Chironomidae
abundance (number/m2) across all Areas of Concern.
Relation of the summed total simultaneously extracted metals (Cd,
Cr, Cu, Ni, Pb, Zn) concentration (/iM/dry g) to mean Bivalvia
abundance (number/m2) across all Areas of Concern.
Relation of the summed total simultaneously extracted metals (Cd,
Cr, Cu, Ni, Pb, Zn) concentration (jiM/dry g) to mean Gastropoda
abundance (number/m2) across all Areas of Concern.
Relation of the summed total PAH concentration (/zg/dry g OC) to
mean Oligochaeta abundance (number/m2) across all Areas of
Concern.
Relation of the summed total PAH concentration (/ig/dry g OC) to
mean Chironomidae abundance (number/m2) across all Areas of
Concern.
Relation of the summed total PAH concentration (^g/dry g OC) to
mean Bivalvia abundance (number/m2) across all Areas of Concern.
Relation of the summed total PAH concentration (jig/dry g OC) to
mean Gastropoda abundance (number/m2) across all Areas of Concern.
Relation of the summed total PCB concentration (/^g/dry g OC) to
mean Oligochaeta abundance (number/m2) across all Areas of
Concern.
Relation of the summed total PCB concentration (yug/dry g OC) to
mean Chironomidae abundance (number/m2) across all Areas of
Concern.
Relation of the summed total PCB concentration (jug/dry g OC) to
mean Bivalvia abundance (number/m2) across all Areas of Concern.
Relation of the summed total PCB concentration (/ig/dry g OC) to
mean Gastropoda abundance (number/m2) across all Areas of Concern.
Percentage composition of the invertebrate taxa collected using
artificial substrates and a ponar grab at Indiana Harbor, IN,
August 1990.
Percentage composition of the invertebrate taxa collected using
artificial substrates and a ponar grab at Buffalo River, NY,
October 1989.
Percentage composition of the invertebrate taxa collected using
artificial substrates and a ponar grab at Saginaw River, MI, June
1990.
Comparison of the total amounts of invertebrates collected with
artificial substrates and a ponar grab at Saginaw River, MI, June
1990.
CHAPTER 6: AMES MUTAGENICITY ASSAY
Toxicity Scores for AOC Stations.
Mutagenicity Scores for AOC Stations.
Summary of Toxicity Score Results for AOCs.
Summary of Mutagenicity Score Results for AOCs.
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CHAPTER 7: MUTATOX GENOTOXICITY ASSAY
Figure 7.1 Genotoxicity of 2-arainoanthracene determined with the activated
Mutatox Assay (Photobacterium/rat hepatic S9).
Figure 7.2 Genotoxicity of benzo(a)pyrene determined with the activated
Mutatox Assay (Photobacterium/rat hepatic S9).
Figure 7.3 Genotoxicity of freshwater sediment extracts from Indiana Harbor
determined with the activated Mutatox Assay (Photobacterium/rat
hepatic S9).
Figure 7.4 Genotoxicity of freshwater sediment extracts from Indiana Harbor
determined with the activated Mutatox Assay (Photobacterium/rat
hepatic S9): single data set from Station 6.
Figure 7.5 Genotoxicity of freshwater sediment extracts from Buffalo River
determined with the activated Mutatox Assay (Photobacterium/rat
hepatic S9).
Figure 7.6 Genotoxicity of freshwater sediment extracts from the first survey
of Saginaw River determined with the activated Mutatox Assay
(Photobacterium/rat hepatic S9).
Figure 7.7 Genotoxicity of freshwater sediment extracts from the third survey
of Saginaw River (core samples) determined with the activated
Mutatox Assay (Photobacterium/rat hepatic S9).
Figure 7.8 Genotoxicity of freshwater sediment extracts from the third survey
of Saginaw River (grab sample) determined with the activated
Mutatox Assay (Photobacterium/rat hepatic S9).
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LIST OF TABLES
CHAPTER 1: OVERVIEW
Table 1.1 USEPA GLNPO ARCS Toxicity-Chemistry Work Group.
CHAPTER 2: ELUTRIATE TOXICITY TESTS: DAPHNIA MAGNA AND MICROTOX
Table 2.1 Sample numbers and sample station locations for sediments
collected from three Great Lakes Areas of Concern.
Table 2.2 Dissolved oxygen, pH, and survival of Daphnia magna during a 48-h
reference toxicant test with Instant OceanR salinity.
Table 2.3 Elutriate water quality for each sample.
Table 2.4 Mean water quality for 48-h acute toxicity tests with Daphnia
ma RH a.
Table 2.5 Concentrations of metals in elutriates prepared from Indiana
Harbor sediment samples.
Table 2.6 Concentration of metals in Indiana Harbor sediment porewater
samples.
Table 2.7 Acute toxicity of Indiana Harbor, Buffalo River, and Saginaw River
elutriates to Daphnia magna and Photobacterium phosphoreum
(MicrotoxR).
Table 2.8 Concentrations of metals in elutriates prepared from Buffalo River
sediment samples.
Table 2.9 Concentration of metals in Buffalo River sediment porewater
samples.
Table 2.10 Concentrations of metals in elutriates from sediment samples
collected during the first survey of Saginaw River.
Table 2.11 Concentration of metals in pore water from sediments collected
during the first survey of Saginaw River.
Table 2.12 Concentrations of metals in elutriates prepared from sediments
collected during the third survey of Saginaw River.
Table 2.13 Concentration of metals in pore water from sediment samples
collected during the third survey of Saginaw River.
Table 2.14 Acute toxicity of elutriates prepared from stored whole sediment
samples from Indiana Harbor, Buffalo River, and Saginaw River to
Photobacterium phosphoreum in MicrotoxR tests.
CHAPTER 3: ELUTRIATE TOXICITY TESTS: SELENASTRUM CAPRICORNUTUM
Table 3.1 Algal culture medium.
Table 3.2 Quality assurance for algal assays.
Table 3.3 Concentrations in non-filtered elutriate samples.
Table 3.4 Water quality of elutriates tested with the algal assays.
CHAPTER 4: WHOLE SEDIMENT TOXICITY TESTS
Table 4.1 Chemical names for compounds analyzed in sediment samples.
Table 4.2 Concentrations of total organic carbon, percent solids, and
organometals for Indiana Harbor sediment samples.
Table 4.3 Concentrations of total metals in Indiana Harbor sediment samples.
VI11
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Table 4.4 Simultaneously extracted metals and the acid volatile sulfides in-
Indiana Harbor and Buffalo River sediment samples.
Table 4.5 Particle size distribution for Indiana Harbor sediment samples.
Table 4.6 Concentrations of polynuclear aromatic and otber semivolatile
compounds in Indiana Harbor sediment samples.
Table 4.7 Concentrations of dioxins and furans in Indiana Harbor sediment
samples.
Table 4.8 Polychlorinated biphenyls in Indiana Harbor and Buffalo River
sediment samples.
Table 4.9 Pesticide ranges of analytical detection limits for sediment
samples.
Table 4.10 Overlying water quality for Indiana Harbor whole sediment toxicity
tests.
Table 4.11 Toxicity responses of Hyalella azteca. Chironomus riparius. and
Chironomus tentans to Indiana Harbor whole sediment samples.
Table 4.12 Concentrations of total organic carbon, percentage solids, and
organometals in Buffalo River sediment samples.
Table 4.13 Concentrations of total metals in Buffalo River sediment samples.
Table 4.14 Particle size distribution for Buffalo River sediment samples.
Table 4.15 Concentrations of polynuclear aromatic and other semivolatile
compounds in Buffalo River sediment samples.
Table 4.16 Concentrations of dioxins and furans in Buffalo River sediment
samples.
Table 4.17 Overlying water quality for Buffalo River whole sediment toxicity
tests.
Table 4.18 Toxicity responses of Hyalella azteca. Chironomus riparius. and
Chironomus tentans to Buffalo River whole sediment samples.
Table 4.19 Concentrations of total organic carbon, percentage solids, and
organometals in sediment samples from the first survey of Saginaw
River.
Table 4.20 Concentrations of metals in sediment samples from the first survey
of Saginaw River.
Table 4.21 Simultaneously extracted metals and the acid volatile sulfides in
sediment samples from the first and third surveys of Saginaw
River.
Table 4.22 Particle size distribution for sediment samples from the first
survey of Saginaw River.
Table 4.23 Concentrations of polynuclear aromatic and other semivolatile
compounds in sediment samples from the first survey of Saginaw
River.
Table 4.24 Concentrations of dioxins and furans in sediment samples from the
first survey of Saginaw River.
Table 4.25 Overlying water quality for Saginaw River whole sediment toxicity
tests, first survey.
Table 4.26 Toxicity responses of Hyalella azteca and Chironomus riparius to
Saginaw River whole sediment samples, first survey.
Table 4.27 Concentrations of total organic carbon, percent solids, and
organometals in sediment samples from the third survey of Saginaw
River.
Table 4.28 Concentrations of total metals in sediment samples from the third
survey of Saginaw River.
IX
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Table 4.29 Particle size distribution of sediment samples from the third
survey of Saginaw River.
Table 4.30 Concentrations of polynuclear aromatic and other semivolatile
compounds in sediments from the third survey of Saginaw River.
Table 4.31 Concentrations of dioxins and furans in sediment samples from the
third survey of Saginaw River.
Table 4.32 Concentrations of pesticides in sediment samples from the third
survey of Saginaw River.
Table 4.33 Polychlorinated biphenyl concentrations and analytical detection
limits for sediment samples from the third survey of Saginaw
River.
Table 4.34 Overlying water quality for Saginaw River whole sediment toxicity
tests, third survey.
Table 4.35 Toxicity responses of Hyalella azteca. Chironomus riparius. and
Chironomus tentans to Saginaw River whole sediment samples, third
survey.
Table 4.36 Categorical comparison matrix of Chironomus tentans survival data
for whole sediment toxicity tests of Saginaw River, third survey.
Table 4.37 Low and high molecular weight polynuclear aromatic hydrocarbons
and total polychlorinated biphenyls normalized to percentage total
organic carbon all Areas of Concern, tested in whole sediment
exposures.
Table 4.38 Recommended threshold concentrations for polynuclear aromatic
hydrocarbons and total polychlorinated biphenyls.
Table 4.39 Recommended metals threshold concentrations.
Table 4.40 Comparisons of recommended threshold concentrations and test
organism responses from whole sediment exposures.
Table 4.41 Comparison of test species sensitivity.
CHAPTER 5: BENTHIC COMMUNITY STRUCTURE EVALUATIONS
Table 5.1 Mean abundance values for major benthic invertebrate groups
collected from Indiana Harbor, IN, August 1989.
Table 5.2 Percentage contribution of each taxa to the total number of taxa
collected from Indiana Harbor, IN, August 1989.
Table 5.3 Mean abundance values for aquatic Oligochaeta species collected
from Indiana Harbor, IN, August 1989.
Table 5.4 Mean abundance values for Chironomidae genera collected from
Indiana Harbor, IN, August 1989.
Table 5.5 Mean abundance values for Mollusca genera and species collected
from Indiana Harbor, IN, August 1989.
Table 5.6 Percentage contribution of each taxa to the total number of taxa
collected from Buffalo River, NY, October 1989.
Table 5.7 Mean abundance values for aquatic Oligochaeta species collected
from Buffalo River, NY, October 1989.
Table 5.8 Mean abundance values for Chironomidae genera collected from
Buffalo River, NY, October 1989.
Table 5.9 Mean abundance values for Mollusca genera and species collected
from Buffalo River, NY, October 1989.
Table 5.10 Percentage contribution of each taxa to the total number of taxa
collected from the first survey of Saginaw River, MI, December
1989.
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CHAPTER 7: MUTATOX GENOTOXICITY ASSAY
Table 7.1 Genotoxicity of arylamines and polycyclic aromatic hydrocarbons in
sediment extracts determined with the activated Mutatox Assay
(Photobacterium/rat hepatic S9).
Table 7.2 Genotoxicity of progenotoxic chemicals in complex mixtures deter-
mined with the activated Mutatox Assay (Photobacterium/rat hepatic
S9).
Table 7.3 Comparison of the Mutatox Assay and Ames Test.
xii
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Appendix 4.1
Appendix 4.2
Appendix 6.1
Appendix 6.2
Appendix 6.3
Appendix 6.4
Appendix 6.5
Appendix 6.6
Appendix 6.7
Appendix 6.8
Appendix 6.9
Appendix 6.10
Appendix 6.11
Appendix 6.12
Appendix 6.13
LIST OF APPENDICES
CHAPTER 4: WHOLE SEDIMENT TOXICITY TESTS
Concentrations of total metals for control sediment.
Concentrations of polynuclear aromatic and other
semivolatile compounds for control sediment.
CHAPTER 6: AMES MUTAGENICITY ASSAY
Mean number of revertants, no S9, Indiana Harbor.
Mean number of revertants, S9, Indiana Harbor.
Mean number of revertants, no S9, Buffalo River.
Mean number of revertants, S9, Buffalo River.
Mean number of revertants, no S9, first survey of Saginaw
River.
Mean number of revertants, S9, first survey of Saginaw
River.
Mean number of revertants, no S9, third survey of Saginaw
River.
Mean number of revertants, S9, third survey of Saginaw
River.
Mean number of revertants, no S9, core samples, third survey
of Saginaw River.
Mean number of revertants, S9, core samples, third survey of
Saginaw River.
Mean number of revertants, no S9, Quality Control and
Quality Assurance samples.
Mean number of revertants, S9, Quality Control and Quality
Assurance samples.
Mean number of revertants, direct-acting and promutagens.
xiii
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ACKNOWLEDGMENTS
In addition to the members of the GLNPO ARCS Work Groups and Activities
Integration Committee, we would like to thank J.B. Hunn for assistance on
Quality Control and Quality Assurance and the following individuals for their
input on the project:
CHAPTER 2: ELUTRIATE TOXICITY TESTS: DAPHNIA MAGNA AND MICROTOX
C.W. Apts, E.L. Brunson, M.R. Ellersieck, L.R. Farley, M.C. Gray, I.E. Greer,
E.K. Henry, N.E. Kemble, B.K Lasorsa, S.L. Kiesser, L.B. King, and K. Srigley
Werner
CHAPTER 3: ELUTRIATE TOXICITY TESTS: SELENASTRUM CAPRICORNUTUM
A.L. Alert, L. Arroyo, A. Barber, E.L. Brunson, L. Burnett, S.A. Burch, R.C.
Clark, J.J. Coyle, F.J. Dwyer, M.R. Ellersieck, L.R. Farley, W.R. Gala, M.C.
Gray, E.K. Henry, M.S. Kaiser, D. Papoulias, D.S. Ruessler, J.A. Thomas
CHAPTER 4: WHOLE SEDIMENT TOXICITY TESTS
C.W. Apts, E.L. Brunson, S.A. Burch, O.A. Cotter, M.R. Ellersieck, L.R.
Farley, M.C. Gray, E.K. Henry, S.L. Kiesser, B.K. Lasorsa, D. Remington, L.S.
Sappington, J.A. Thomas, and K. Srigley Werner
CHAPTER 5: BENTHIC COMMUNITY STRUCTURE EVALUATIONS
M.R. Ellersieck, O.K. Hardesty, R.D. Hurtubise, P. Lovely, B.C. Poulton, D.S.
Ruessler, L.S. Sappington, J.A. Thomas, M.J. Tomasovic, and D.W. Whites
CHAPTER 6: AMES MUTAGENICITY ASSAY
L. Arroyo, G.S. Carey, K.P. Feltz, J.A. Lebo, B.L. Steadman, and D.E. Tillitt
CHAPTER 7: MUTATOX GENOTOXICITY ASSAY
A.A. Bulich
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INTRODUCTION
Section 118 (c)(3) of the Clean Water Act (CWA), as amended by the Water
Quality Act of 1987, authorized the USEPA Great Lakes National Program Office
(GLNPO) to carry out a 5-year project dealing with the Assessment and
Remediation of Contaminated Sediments (ARCS) in the Great Lakes. Annex 14 of
the Great Lakes Water Quality Agreement of 1978, as amended, between the
United States and Canada stipulated the cooperating parties identify the
nature and extent of sediment pollution in the Great Lakes, develop methods to
assess effects, and evaluate the technological capability of programs to
remedy such pollution. Information from these activities is to be used to
guide the development of Remedial Action Plans (RAPs) for the identified Great
Lakes Areas of Concern (AOC) as well as Lake-wide Management Plans (Ross et
al. 1992).
Areas of Concern include major municipal and industrial centers on Great
Lakes rivers, harbors, and connecting channels where beneficial uses are
impaired (International Joint Commission 1987, 1988a, 1988b) . Toxic
contamination of bottom sediments is often a major problem in these areas.
Additionally, areas exist where impairment of beneficial uses (e.g.,
navigation, swimming, fishing) of water or biota have been documented. There
are currently 42 AOC: 25 in U.S. waters, 12 in Canadian waters, and 5
international connecting channels. The CWA designates five AOC for priority
consideration: Saginaw River, MI; Sheboygan Harbor, WI; Indiana Harbor, IN;
Ashtabula River, OH; and Buffalo River, NY. This report describes research
designed to evaluate the toxicity in AOCs where sediments do not have to be
removed to maintain navigational channels, but present a hazard to the
ecosystem if left in place.
In order to implement the ARCS program, a management framework was
established by GLNPO, which included an Activities and Integration Committee
and four technical Work Groups: (1) Toxicity-Chemistry, (2) Risk Assessment-
Modeling, (3) Engineering-Technology, and (4) Communications-Liaison. The
goal of the Toxicity-Chemistry Work Group was to assess the nature and extent
of bottom sediment contamination at selected Great Lakes AOC. The Risk
Assessment-Modeling Work Group was charged with evaluating the ecological
impacts resulting from contaminated sediments and developing techniques for
evaluating various remedial alternatives. The Engineering-Technology Work
Group evaluated procedures for removal and remediation of contaminated
sediments and the Communications-Liaison Work Group facilitated flow of
information between Work Groups and to the public. The Activities Integration
Committee had oversight over the entire ARCS program and coordinated Quality
Control and Quality Assurance (Ross et al. 1992).
The sediment assessment procedures conducted by the National Fisheries
Contaminant Research Center (NFCRC) on samples from Indiana Harbor, Buffalo
River, and Saginaw River included: Elutriate Toxicity Tests (Chapters 2 and
3), Whole Sediment Toxicity Tests (Chapter 4), Benthic Community Structure
(Chapter 5), and Mutagenicity (Chapter 6) and Genotoxicity (Chapter 7) Assays.
A separate project by Wright State University (WSU) provided additional
toxicity information on splits of select sediment samples from each AOC
(Burton et al. 1992). The NFCRC and WSU projects were designed to evaluate
sediment evaluation procedures suggested by both the International Joint
Commission (International Joint Commission 1988a, 1989) and ASTM (ASTM E 1383-
92).
xv
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REFERENCES CITED IN THE INTRODUCTION
American Society For Testing and Materials. 1992. E 1383-92 Standard Guide For
Conducting Sediment Toxicity Tests with Freshwater Invertebrates. ASTM 1992
Annual Book of Standards. Volume 11.04, ASTM, Philadelphia, PA.
Burton, G.A., L. Burnett, P. Landrum, M. Henry, S. Klaine, and M. Swift. 1992.
USEPA GLNPO ARCS Final Report: A Multi-Assay/Multi-Test Site Evaluation of
Sediment Toxicity. March 31, 1992.
International Joint Commission. 1987. Report on Great Lakes Water Quality.
Appendix A. Progress in Developing Remedial Action Plans for Areas of Concern
in the Great Lakes Basin. Report to the International Joint Commission Great
Lakes Water Quality Board, Windsor, Ontario.
International Joint Commission. 1988a. Procedures for the Assessment of
Contaminated Sediment Problems in the Great Lakes. Sediment Subcommittee and
its Assessment Work Group Report to the International Joint Commission Great
Lakes Water Quality Board, Windsor, Ontario.
International Joint Commission. 1988b. Options for the Remediation of
Contaminated Sediments in the Great Lakes. Sediment Subcommittee and its
Remedial Options Work Group Report to the International Joint Commission Great
Lakes Water Quality Board, Windsor, Ontario.
International Joint Commission. 1989. Guidance on Characterization of Toxic
Substances Problems in Areas of Concern in the Great Lakes Basin. Report from
the Surveillance Work Group to the International Joint Commission Great Lakes
Water Quality Board, Windsor, Ontario.
Ross, P.E., G.A. Burton, Jr., E.A. Crecelius, J.C. Filkins, J.P. Giesy, Jr.,
C.G. Ingersoll, P.F. Landrum, M.J. Mac, T.J. Murphy, J.E. Rathbun, V.E. Smith,
H.E. Tatem, R.W. Taylor. Assessment of Sediment Contamination at Great Lakes
Areas of Concern: The ARCS Program Toxicity-Chemistry Work Group Strategy. In
Press.
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CONCLUSIONS AND RECOMMENDATIONS
GENERAL
Sediment samples collected from Indiana Harbor were severely
contaminated compared to either Buffalo or Saginaw River based on
sediment toxicity and chemistry. While sediment samples from Saginaw
River were generally less contaminated compared to Buffalo River,
considerable spatial variability in contamination was evident at all
three AOCs.
Each sediment sample contained a complex mixture of inorganic and
organic contaminants. Additional studies are required to determine
specific contaminants that may be causing adverse effects (e.g.,
sediment spiking and Toxicity Identification Evaluations).
Multiple assessment procedures are needed to thoroughly evaluate
sediment contamination. Data from these studies are to be evaluated
with the Sediment Quality Triad which will integrate data from
laboratory exposures, benthic community structure, and chemical analyses
to provide complementary evidence for the degree of pollution-induced
degradation in aquatic communities at each AOC. Results of the Triad
analyses will be discussed in a later report.
Additional research is needed dealing with factors that control
bioavailability of sediment-associated contaminants (e.g., organic
carbon, particle size, acid volatile sulfides). Further investigations
are needed on factors that control the desorption rates of sediment-
sorbed contaminants.
CHAPTER 2: ELUTRIATE TOXICITY TESTS: DAPHNIA MAGNA AND MICROTOX
Elutriate toxicity tests were conducted with Daphnia magna (48-h
exposures) and Microtox (5- and 15-min exposures). About 33% of the 30
elutriate samples tested with daphnids were identified as toxic (i.e.,
EC50 less than the full-strength elutriate). Forty-one percent of the
39 samples tested with Microtox were identified as toxic. These results
indicate that the Microtox test was a more sensitive tool for evaluating
the toxicity of sediment elutriate samples compared to the Daphnia magna
test.
Toxicity and chemical analyses of elutriates prepared from Buffalo and
Saginaw River sediment samples indicated that sediment contamination at
these AOCs may not be as extensive relative to Indiana Harbor.
Sediment resuspension should be kept to a minimum if contaminated
sediments are removed during remediation based on the toxicity and
contamination of the elutriate samples.
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Microtox and Daphnia maqna toxicity tests with elutriates are useful
tools for assessing the potential hazards of contaminated sediments.
While the elutriate test can be used to evaluate potential effects of
open-water disposal of dredged material, the test may not be appropriate
for evaluating in situ effects of contaminated sediments on aquatic
organisms.
CHAPTER 3: ELUTRIATE TOXICITY TESTS: SELENASTRUM CAPRICORNUTUM
Elutriate toxicity tests were conducted using a 24-h Hcarbon
assimilation algal assay. Interpretation of toxicity using Selenastrum
capricornutum was complicated by variable nutrient and inorganic carbon
concentrations in the elutriate samples. All of the elutriate samples
tested stimulated carbon assimilation of Selenastrum capricornutum in
one or more of the dilutions.
Attempts to modify the algal medium to provide unlimited nutrients were
not successful. An algal medium which supports greater growth potential
should be developed in order to evaluate the toxicity of environmental
samples with high concentrations of nutrients to algae.
To evaluate the effect of storage on toxicity, additional elutriates
were prepared from select whole sediment samples stored for 12 to 16
months after collection. Storage of whole sediments for over a year did
not dramatically change toxicity of elutriate samples in the Selenastrum
capricornutum or in the Microtox tests.
An approach for evaluating toxicity in samples that may be stimulatory
is presented. This approach needs to be evaluated using reference
toxicants.
CHAPTER 4: WHOLE SEDIMENT TOXICITY TESTS
Whole sediment toxicity tests were conducted with amphipod Hyalella
azteca. and the midges Chironomus riparius and Chironomus tentans.
About 68% of the sediment samples were toxic to Hyalella azteca,
Chironomus riparius. or Chironomus tentans in 10-d to 28-d exposures.
Fourteen- and 28-d exposures with Hyalella azteca which monitored
effects on survival, body length, and sexual maturation, identified a
higher proportion of toxic samples compared to either 14-d exposures
with Chironomus riparius or 10-d exposures with Chironomus tentans.
Chronic sediment toxicity tests need further development to better
determine effects of complex chemical mixtures. These tests should
incorporate sublethal responses using representative benthic organisms.
Toxic responses observed in the whole sediment tests need to be compared
to benthic community effects and sediment chemistry to determine if the
test are predictive.
xvni
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CHAPTER 5: BENTHIC COMMUNITY STRUCTURE EVALUATIONS
• Oligochaetes and chironomids were the dominant taxa at each AOC
indicating impacted benthic communities.
• Average number of midges with mouth part deformities ranged from 45 to
over 77% in samples from the AOCs.
• Results of variance partitioning estimates indicates station and
replicates accounted for most of the explained variability.
• Artificial substrate samplers which were colonized in situ sampled a
more diverse taxa compared to benthic grab samplers.
• Since most of the variance in benthic community estimates was between
stations and replicates, future studies should sample more stations and
replicates using a smaller grab sampler.
f Frequency of deformities in chironomids exposed in laboratory toxicity
tests (Chapter 4) should be measured and compared to the frequency of
deformities in the chironomids collected from the field.
• Assessments of benthic community should use both artificial substrate
and grab samplers.
• Additional research is needed evaluating specific contaminant, abiotic,
and biotic factors controlling invertebrate distributions in sediments.
CHAPTER 6: AMES MUTAGENICITY ASSAY
t In order to assess the mutagenicity of Great Lakes sediments,
unfractionated organic extracts of sediments were screened for
mutagenicity using the Ames Salmonella/microsome assay.
• Extracts of sediment samples from every station were both toxic and
mutagenic to the bacteria. Toxicity was controlled by varying rat
hepatic S9 concentrations and extract dilution.
• Of the four bacterial strains tested, only strain TA98 detected
mutagenicity at every station. Although extracts were not fractionated,
chemical analysis indicated that as a class, PAHs comprise the greatest
percentage by weight of total identified organics. Sediment
mutagenicity is likely attributable to PAH compounds.
• Fractionation of samples before testing may allow a greater degree of
discrimination and aid in identifying compounds causing the observed
mutagenicity.
• Methods should be developed to test elutriate and porewater samples to
better evaluate bioavailability of sediment associated contaminants.
Studies utilizing passive membrane bag samplers may be useful for this
purpose.
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CHAPTER 7: MUTATOX GENOTOXICITY ASSAY
Organic extracts of sediment samples were evaluated with the new
activated Mutatox Genotoxicity Assay using rat hepatic S9 for exogenous
metabolic activation and a dark mutant strain of the luminescent
bacterium Photobacterium phosphoreum for detection of environmental DNA-
damaging substances (genotoxins).
A genotoxic response was indicated when the test chemical restored the
luminescent state in bacteria. The degree of light increase indicated
the relative genotoxicity of the sample.
Toxicity of the organic extracts complicated interpretations in the Ames
assay, but was not a problem in the Mutatox assay.
Samples from 27 of 28 stations exhibited evidence of genotoxins, 23 of
28 stations (82%) were designated genotoxic, four were suspect (14%),
and one was negative (3%).
The activated Mutatox Assay was a sensitive, specific, predictive, and
rapid test for detecting the presence of genotoxins in complex
environmental samples.
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CHAPTER 1; OVERVIEW
Ingersoll, C.G., D.R. Buckler, E.A. Crecelius, and T.W. La Point
Determining the effects of contaminants in sediment is a challenging new
area in environmental toxicology. Mounting evidence exists of environmental
degradation in areas where water quality criteria are not exceeded, yet
organisms in or near sediments are adversely affected (Chapman 1986).
Historically, evaluations of contaminant effects have emphasized surface
waters, not sediments. Most assessments of water quality focus on water-
soluble compounds and sediment is considered to be a safe repository of sorbed
contaminants (Maki et al. 1984). This approach emphasizes testing organisms
in the water column without considering the fate of chemicals in sediment.
The assessment of sediment quality is often limited to chemical
characterizations. However, quantifying contaminant concentration alone
cannot provide enough information to adequately evaluate potential adverse
effects, interactions among chemicals, or the time-dependent availability of
contaminants to aquatic organisms (Dillon and Gibson 1986). Because of poorly
understood relationships between concentrations of contaminants in sediment
and their bioavailability, determining effects of contaminated sediments on
aquatic organisms requires use of benthic community surveys and controlled
toxicity and bioaccumulation tests (Seelye and Mac 1984).
The goal of the National Fisheries Contaminant Research Center (NFCRC)
project was to assess the nature and extent of bottom sediment contamination
at selected Great Lakes Areas of Concern (AOC) in the United States (Ross et
al. 1992). Up to 12 stations were sampled from each of three AOCs (Saginaw
River, MI; Buffalo River, NY; and Indiana Harbor, IN; Table 2.1). Stations
were selected by the GLNPO ARCS Toxicity-Chemistry Work Group (Table 1.1).
Personnel from USEPA Grosse lie Large Lakes Research Station (LLRS), Grosse
lie, MI collected and shipped sediment samples to NFCRC for the sediment
assessments. Personnel from Battelle Marine Science Laboratories, Sequim, WA
performed physical and chemical analyses on the sediment samples. Personnel
from the USFWS National Fisheries Center-Great Lakes, Ann Arbor, MI conducted
fish sediment bioaccumulation studies and fish tumor surveys (Mac et al.
1992). NFCRC personnel assessed sediment contamination in each sample using
elutriate and whole sediment toxicity tests, benthic community surveys, and
mutagenicity and genotoxicity assays. The NFCRC project was conducted in
cooperation with a Wright State University project that provided additional
toxicity information for select stations at each AOC (Burton et al. 1992).
Data from these studies are to be evaluated with both the Sediment
Quality Triad (Triad; Chapman 1986) and the Apparent Effects Threshold (AET;
Barrick et al. 1988) approaches in order to assess the biological hazard of
contaminants associated with sediments in Great Lakes AOC. Triad and AET
analyses will be discussed in a later report. The Triad will integrate data
from: (1) laboratory exposures (e.g., sediment toxicity); (2) benthic
community structure (e.g., species abundance); and (3) chemical and physical
analyses (e.g., organic and inorganic contaminants, grain size) and will
provide strong, complementary evidence for the degree of pollution-induced
degradation in aquatic communities (Chapman 1986). The data obtained from the
studies will also be used to generate Apparent Effects Threshold (AET) values.
An AET is defined as the sediment concentration of a given chemical above
which statistically significant biological effects (e.g., sediment toxicity)
1-1
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are always expected. If any chemical exceeds its AET for a particular
biological indicator, an adverse biological effect is predicted for that
indicator. If all chemical concentrations are below their AET for a
particular biological indicator, then no adverse effect is predicted. The AET
has been applied to contaminated sediment in marine environments (e.g., the
Puget Sound, Barrick et al. 1988), however this approach has yet to be applied
to freshwater ecosystems.
DESCRIPTION OF AREAS OF CONCERN AND STATION SELECTION
Indiana Harbor
The Indiana Harbor and the Grand Calumet River (Figure 1.1) receive
heavy industrial and municipal inputs creating severe water and sediment
quality problems. There are 39 permitted outfalls on the Grand Calumet River
and Indiana Harbor Canal. Contaminant problems are aggravated by the lack of
industrial pretreatment and the small size of the upstream drainage basin
(Brannon et al. 1989). Toxic substances are of particular concern in this AOC
because most of the flow in this system enters Lake Michigan. Although source
control during the past decade has improved water quality, sediments and
ground water in the area continue to be a repository for many contaminants
including polychlorinated biphenyls (PCBs), polycyclic aromatic hydrocarbons
(PAHs), polychlorinated dioxins and furans, chlorinated pesticides, heavy
metals, and many other pollutants (International Joint Commission 1987).
Surface sediments are contaminated as are sediments buried by deposition.
A total of 50 USEPA Superfund sites have been identified and listed in
the Indiana Harbor-Grand Calumet River AOC (Simmers et al. 1989). Benthic
invertebrates collected from the area are indicative of pollution tolerant
organisms (e.g., oligochaetes). Carp (Cyprinus carpio) collected from the
Grand Calumet River had deformities including eroded fins, swollen eyes,
deformed lower jaws, and concentrations of PCBs and chlordane exceeding U.S.
Food and Drug Administration action levels (Simmers et al. 1989).
Channelization and historic dredging have also reduced diversity of habitat
and diversity of aquatic organisms at the AOC. No dredging activities have
occurred since 1972 because of concerns regarding severely contaminated
sediment. For additional detail on the Indiana Harbor-Grand Calumet River AOC
see International Joint Commission (1987), Brannon et al. (1989), and Simmers
et al. (1989).
Buffalo River
The Buffalo River (Figure 1.2) has had documented pollution problems
(e.g., bacteria, oil, phosphorus, chlorine, phenol, mercury, and organic dyes)
since the 1940s (International Joint Commission 1987). Updated municipal
wastewater treatment facilities and controls on industrial discharges have
reduced concentrations of many waterborne pollutants, however the major
problem in the Buffalo River continues to be contaminated sediments.
Sediments are contaminated with heavy metals, industrial organic chemicals,
PCBs, and PAHs (International Joint Commission 1987). Surface sediments are
contaminated as are sediments buried by deposition. Fisheries resources and
populations of benthic invertebrates are severely impaired and fish
consumption advisories limit public use of the river. An increased frequency
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of fish tumors and other deformities has also been reported. River sediments
at some locations are contaminated with cyanide and metals to a level that
prohibits open lake disposal of dredge materials. Channelization and dredging
of the Buffalo River have also reduced diversity of habitat and the diversity
of aquatic organisms. For additional details on the Buffalo River AOC see
International Joint Commission (1987) and Lee et al. (1969).
Saqinaw River
The Saginaw River (Figure 1.3) receives industrial and municipal
wastewater discharges from four major urban areas (Flint, Saginaw, Bay City,
and Midland). There are 127 wastewater treatment facilities and 87 industries
that discharge directly into the Saginaw River and Saginaw Bay (Brandon et al.
1989). Eutrophication (e.g., phosphorus) and toxic materials have been
identified as causing degraded conditions in Saginaw River and Saginaw Bay.
Contaminants in sediment include heavy metals, PCBs, polybrominated biphenyls,
and DDT. Surface sediments are contaminated as are the sediments buried by
deposition. Sediments are most severely contaminated around Saginaw and Bay
City. Over 100 hazardous waste sites and 13 USEPA Superfund sites are located
in the Saginaw River basin (International Joint Commission 1987). Pollutants
of concern in fish collected from the river include hexachlorobenzene, furans
and dioxins, diphenyl ethers, styrenes, and terphenyls. Fish consumption
advisories have been issued due to elevated levels of PCBs in tissues.
Benthic communities have been effected by both toxic materials and
eutrophication. Toxic materials may also be affecting reproductive success of
fish-eating birds in Saginaw Bay. For additional details on the Saginaw River
AOC see International Joint Commission (1987) and Brandon et al. (1989).
Station Selection
Stations were selected by the GLNPO ARCS Toxicity-Chemistry Work Group
(Tables 1.1 and 2.1). Attempts were made to select stations representing a
gradient in sediment contamination. Additionally, an effort was made to
provide some degree of geographical coverage of the AOCs. Chemical,
biological, and geophysical characterizations (water, sediment, biota) for
each AOC had been conducted on a limited basis by state and federal agencies,
universities, and International Joint Commission studies. These data were
available through reports, journal articles, and computer database systems
(International Joint Commission 1987, Brannon et al. 1989, Simmers et al.
1989, Lee et al. 1989, Brandon et al. 1989).
GENERAL METHODS
Sediment Collection, Handling, and Storage
Contaminants in field-collected sediment may include carcinogens,
mutagens, and other potentially toxic compounds. Testing of sediments was
started before chemical analyses were completed so contact with sediment was
minimized by using gloves, laboratory coats, safety glasses, and respirators.
Samples were manipulated and tested under negative pressure.
Sediments were collected and shipped to participating laboratories
between August 1989 and July 1990 by personnel from the USEPA Large Lakes
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Research Station, Grosse lie MI (Table 1.1). Station location was determined
with a global positioning system. A stainless steel Ponar grab (23- x 23-cm)
was used to collect benthic invertebrates at each station (Chapter 5). All
benthos samples were collected before samples for toxicity testing were
collected to minimize disturbance of the sediments and associated
invertebrates. Sediments from the first surveys of Indiana Harbor, Buffalo
River, and Saginaw River were collected with the Ponar grab. Sediments from
the third survey of Saginaw River were collected with a Van Veen (41- x 51-cm)
grab sampler. The grab samplers collected material from the upper 10 to 20 cm
of the sediment. Multiple casts of the grab samplers were made and an attempt
was made to avoid sampling the same location twice. Sediment was placed in 5-
L plastic buckets with plastic liners for transport to shore. Total volume
collected ranged from 15 to over 100 L/station. On shore, the sediment was
poured into a cement mixer, a plastic cover was placed over the mouth of the
mixer, and the sediments were homogenized for 15 min. The cement mixer was
washed with soap and site or city water between samples. After
homogenization, sub-samples of the sediment were placed in high density
polyethylene (HDPE) containers. Sediment samples were shipped on ice to the
laboratories via over-night courier. Upon arrival, the sediments were
inventoried and held in the dark at 4°C.
Core samples were collected from select stations during the third survey
of the Saginaw River (Table 2.1) using a portable electro-mechanical vibrator
(Model M3000, Wacker Corporation, Milwaukee, WI). A flexible shaft was
attached to a custom-made stainless steel core head via a Model H45 vibrator
head. The core tubes were 7.6 cm diameter and penetrated to a depth of up to
1.8 m. See Chapter 3 for additional details on coring procedures.
Sediment samples were generally stored for no more than three weeks
before the start of testing. Elutriate toxicity tests with Microtox were
conducted upon receipt of the sediments and again at the start of the whole
sediment toxicity tests to document potential changes in sediment toxicity
with storage (Chapters 2, 3, and 4). The control sediment was a fine silt-
and clay-particle size soil obtained from an undisturbed agricultural area.
This control sediment has been used previously to evaluate contaminated
sediments (Ingersoll and Nelson 1990, ASTM E 1383-92).
Physical and Chemical Characterization of Sediments
Sediment physical characterization included organic carbon, percentage
water, and particle size (Chapter 4). Sediment chemical characterization
included total metals (Ag, As, Cd, Cr, Cu, Fe, Hg, Mn, Ni, Pb, Se, Zn),
organometals (butyltins and methyl mercury), acid volatile sulfide (AVS) and
simultaneously extracted metals (SEM), chlorinated pesticides, total
polychlorinated biphenyls (PCBs), polychlorinated dioxins and furans, and
polycyclic aromatic hydrocarbons (Chapter 4). Metal concentrations in
elutriate and porewater samples were also measured for select samples (Chapter
2).
Quality Control and Quality Assurance
All studies were conducted in basic accordance with Good Laboratory
Practice outlined in the Federal Register (160.120; 40 CFR Ch. 1; 7-1-85
edition; subpart G - "Protocol for and Conduct of a Study") and were in basic
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accordance with NFCRC guidelines for the provisions of good care and humane
handling of test animals during culturing and experimentation. Data are
permanently archived by NFCRC and summary data have been transmitted to the
GLNPO Project Officer. Data verification reports can be obtained from Dr.
Brian Shumacher (USEPA, P.O. Box 93478, Las Vegas, Nevada 89193-3478, 702/798-
2100).
Precision is a term that describes the degree to which data generated
from replicate measurements differ. Accuracy is the difference between the
value of the measured data and the true value. The acceptability of tests was
determined by estimating precision (e.g., replication) and by the response of
organisms to control sediment or dilution water (e.g., survival).
Determining the accuracy of tests with field samples is not possible since the
true values are not known. No procedures have been developed to directly
measure the accuracy of sediment tests. However, accuracy was indirectly
estimated by evaluating the sensitivity of organisms with reference chemicals.
Performance audits were conducted by the NFCRC Quality Assurance Officer and
GLNPO personnel. Specific reference toxicants, analytical standards,
calibration checks, and quality check frequency are described in Chapters 2 to
7.
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REFERENCES CITED IN CHAPTER 1
American Society For Testing and Materials. 1992. E 1383-92 Standard Guide For
Conducting Sediment Toxicity Tests with Freshwater Invertebrates. ASTM 1992
Annual Book of Standards. Volume 11.04, Philadelphia, PA.
Barrick, R., S. Becker, L. Brown, H. Beller, and R. Pastorok. 1988. Sediment
Quality Values Refinement: 1988 Update and Evaluation of Puget Sound AET, Vol.
I, PTI, Bellevue, WA.
Brandon, D.L, C.R. Lee, J.G. Skogerboe, J.W. Simmers, and H.E. Tatem. 1989.
Information Summary Area Of Concern: Saginaw River, Michigan. Miscellaneous
Paper D-89-xx, U.S. Army Engineer Waterways Experiment Station, Vicksburg, MS.
Brannon, J.M., D. Gunnison, D.E. Averett, J.L. Martin, R.L. Chen, andR.F.
Athow, Jr. 1989. Analyses of Impacts of Bottom Sediments From Grand Calumet
River and Indiana Harbor Canal on Water Quality. Miscellaneous Paper D-89-1,
U.S. Army Engineer Waterways Experiment Station, Vicksburg, MS.
Burton, G.A., L. Burnett, P. Landrum, M. Henry, S. Klaine, and M. Swift. 1992.
USEPA GLNPO ARCS Final Report: A Multi-Assay/Multi-Test Site Evaluation of
Sediment Toxicity. March 31, 1992.
Chapman, P.M. 1986. Sediment Quality Criteria for the Sediment Quality Triad-
An Example. Environ. Contam. Toxicol. 5:957-964.
Dillon, T.M. and A.B. Gibson. 1986. Bioassessment Methodologies for the
Regulatory Testing of Freshwater Dredged Material. Miscellaneous Paper EL-86-
6, U.S. Army Engineer Waterways Experiment Station, Vicksburg, MS.
Ingersoll, C.G. and M.K. Nelson. 1990. Testing Sediment Toxicity with Hyalella
azteca (Amphipoda) and Chironomus riparius (Diptera). In W.G. Landis and W.H.
Van der Schalie, eds., Aquatic Toxicology and Risk Assessment: Thirteenth
Volume. ASTM STP 1096. American Society For Testing and Materials,
Philadelphia, PA, pp. 93-109.
International Joint Commission. 1987. Report on Great Lakes Water Quality.
Appendix A. Progress in Developing Remedial Action Plans for Areas of Concern
in the Great Lakes Basin. Report to the International Joint Commission Great
Lakes Water Quality Board, Windsor, Ontario.
Lee, C.R., D.L. Brandon, J.W. Simmers, H.E. Tatem, and J.G. Skogerboe. 1989.
Information Summary Area Of Concern: Buffalo River, New York. Miscellaneous
Paper EL-89-xx, U.S. Army Engineer Waterways Experiment Station, Vicksburg,
MS.
Mac et al. 1992. USEPA GLNPO ARCS Final Report: Sediment Bioaccumulation and
Fish Tumor Surveys. In preparation.
Maki, A.W., K.L. Dickson, and W.A. Brungs. 1984. Introduction. InK.L.
Dickson, A.W. Maki, and W.A. Brungs, eds., Fate and Effects of Sediment-Bound
Chemicals in Aquatic Systems. Pergamon Press, NY, pp. xv-xxi.
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Ross, P.E., G.A. Burton, Jr., E.A. Crecelius, J.C. Filkins, J.P. Giesy, Jr.,
C.G. Ingersoll, P.P. Landrura, M.J. Mac, T.J. Murphy, J.E. Rathbun, V.E. Smith,
H.E. Tatem, R.W. Taylor. Assessment of Sediment Contamination at Great Lakes
Areas of Concern: The ARCS Program Toxicity-Chemistry Work Group Strategy. In
Press.
Seelye, J.G. and M.J. Mac. 1984. Bioaccumulation of Toxic Substances
Associated with Dredging and Dredged Material Disposal: A Literature Review.
EPA 905/3-84-005, Chicago, IL.
Simmers, J.W., C.R. Lee, D.L. Brandon, H.E. Tatem, and J.G. Skogerboe. 1989.
Information Summary Area Of Concern: Grand Calumet River, Indiana.
Miscellaneous Paper EL-89-xx, U.S. Army Engineer Waterways Experiment Station,
Vicksburg, MS.
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Lake Michigan
N
Figure 1.1 Indiana Harbor sediment sampling stations.
complete description of sampling numbers.
See Table 2.1 for a
-------
If-
N
1 Mile
lackawanna. NY
Flow
Figure 1.2 Buffalo River sediment sampling stations. See Table 2.1 for a
complete description of the sampling numbers.
-------
Saginaw
Bay
N
1 Mile
Figure 1.3 Saginaw River sediment sampling stations. See Table 2.1 for a
complete description of the sampling numbers.
-------
Table 1.1. USEPA GLNPO ARCS Toxicity-Chemistry Work Group.
Name
P.E.
G.A.
D.C.
E.A.
J.C.
R.G.
J.P.
C.G.
P.F.
M.J.
T.J.
J.E.
V.E.
H.E.
Ross
Burton, Jr.
Cowgill
Crecelius
Filkins
Fox
Giesy, Jr.
Ingersoll
Landruro
Mac
Murphy
Rathbun
Smith
Tatera
Affiliation
The Citadel; Charleston, SC
Wright State University; Dayton, OH
USEPA-GLNPO; Chicago, IL
Battelle; Sequim, WA
USEPA-LLRS, Grosse He, MI
USEPA-GLNPO; Chicago, IL
Michigan State University; East Lansing, MI
USFWS; Columbia, MO
NOAA; Ann Arbor, MI
USFWS; Ann Arbor, MI
DePaul University; Chicago, IL
AScI, Grosse He; MI
AScI, Grosse lie; MI
USCOE; Vicksburg, MS
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CHAPTER 2; ELUTRIATE TOXICITY TESTS; DAPHNIA MAGNA AND MICROTOXR
Coyle, J.J., E.A. Crecelius, and C.G. Ingersoll
INTRODUCTION
Contaminated sediments are a widespread problem in the maintenance of
acceptable water quality in freshwater, coastal, and offshore marine
environments (Gannon and Beeton 1969, EPA/COE 1977, Shuba et al. 1978, NRC
1989). The increased concern of resource managers and the scientific
community over the ecological role and significance of contaminated sediments,
and the lack of established correlations between concentrations of sediment-
associated contaminants and biological effects has lead to the development of
sediment bioassessment procedures (EPA/COE 1977, Prater and Anderson 1977).
Tests that measure bioconcentration and biological effects are used in these
sediment assessment procedures rather than chemical analyses to determine the
presence or absence of certain pollutants. Typically, these procedures
involve evaluating the liquid, suspended particulate, and solid phases of
sediment samples for potential biological effects and bioconcentration (Shuba
et al. 1978). Thus, organisms that inhabit the water column are exposed to
the liquid and suspended particulate phases, and benthic and epibenthic
organisms are exposed to the solid phase.
The elutriate test was recommended for use by the U.S. Army Corps of
Engineers and the U.S. Environmental Protection Agency as part of a process
developed to evaluate proposals to discharge dredged materials into ocean
waters, and to evaluate the potential of the dredged materials to impact ocean
ecology (EPA/COE 1977). The Corp of Engineers (1978) evaluated and
recommended the elutriate test as a means of predicting effects of open-water
disposal of dredged materials and suggested that the elutriate test may be
useful in testing other solid waste. In elutriate tests, usually the aqueous
phase of a 4:1 water to sediment mixture is separated and aquatic organisms
are exposed to various dilutions of the aqueous phase for a specified period.
Results of elutriate tests depend on temperature, composition of
dilution water, extent of sediment contamination, condition of the test
organisms and other factors (Shuba et al. 1978). Elutriate tests provide
information on the potential toxicity of specific constituents that may be
released from contaminated sediments to the water column under various field
conditions, e.g., the open water disposal of dredged materials (Shuba et al.
1978, Palermo 1986), but they do not necessarily reflect the toxicity of the
in-place sediments (Seelye and Mac 1984). Thus, results of elutriate tests
can be used to support inferences about the potential toxicity of the
contaminated sediments from which the elutriates are prepared and to identify
the biologically active constituents of the contaminated sediment. In
addition to assessing the potential hazard of contaminated sediments to
aquatic organisms, the results of elutriate tests can be used to compare the
relative toxicity of contaminated sediments from different locations (Shuba et
al. 1978) and to study the biological availability of contaminants associated
with sediments (Seelye and Mac 1984).
In the present study, elutriate tests were used to assess the toxicity
of sediment samples collected from three Great Lakes Areas of Concern (AOC).
The AOCs include major municipal and industrial centers on Great Lakes rivers,
harbors and connecting channels where beneficial uses (e.g., drinking,
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swimming, fishing, navigation) are impaired, or where environmental quality
criteria are exceeded to the point that such impairment is likely (IJC 1988).
There are currently 42 AOCs — 25 in U.S. waters, 12 in Canadian waters, and
five in international connecting channels (IJC 1988). Five AOCs have been
specifically designated for "priority consideration" (FWPCA 1987)— Saginaw
Bay (MI), Sheboygan Harbor (WI), Indiana Harbor (IN), Ashtabula River (OH) and
Buffalo River (NY). Toxicity tests were conducted with elutriates prepared
from sediments collected from three of these five AOCs — Indiana Harbor,
Saginaw River, and Buffalo River. The tests were conducted with Daphnia
maqna and Photobacterium phosphoreum (Microtox"). Selenastrum capricornutum
were also exposed to these elutriates in acute toxicity tests (Chapter 3).
Results of these studies will be used to determine the hazard of contaminated
sediments to Great Lakes aquatic resources beyond the context of navigational
dredging and dredged material disposal. Eventually, data generated from the
studies will be used in two assessment techniques for evaluating the
biological hazard of sediment-associated contaminants in the Great Lakes AOCs
— the Sediment Quality Triad method (Chapman and Long 1983, Long and Chapman
1985) and the Apparent Effects Threshold method (Seller et al. 1986, Barrick
et al. 1988).
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MATERIALS AND METHODS
SEDIMENT COLLECTION, STORAGE AND HANDLING
Summary of Samples Tested
The sediment samples tested were collected within each AOC from Stations
selected by the Great Lakes National Program Office (GLNPO) Toxicity and
Chemistry Work Group (see Table 1.1 in Chapter 1). The samples were collected
by personnel of the U.S. Environmental Protection Agency's (USEPA) Large Lakes
Research Station, Grosse lie, MI. About 10 L of bulk sediment grab samples or
four L of bulk core samples were collected from seven Stations in Indiana
Harbor, IN in August 1989; from 10 Stations in Buffalo River, NY in October
1989; and from 12 Stations in Saginaw River, MI in October 1989 and June 1990
(Table 2.1).
Sampling Methods
A contaminant-free silt-clay sized particle soil collected from
Florissant, Missouri was used as a control sediment. This soil has been
previously used in sediment toxicity tests as a control sediment (Adams et al.
1985, Ingersoll and Nelson 1990).
A 23- X 23-cm standard Ponar sampler was used to collect the grab
samples. To insure that true surficial sediments were collected, care was
taken to avoid sampling from the same spot when successive Ponar casts were
required to obtain a desired sample volume. The core samples were collected
with a portable light-weight (8.1 kg) device assembled by Large Lake Research
Station personnel. The coring device was constructed of a Wacker Model M3000,
three horse-power electro-mechanical vibrator (Wacker Corporation, Milwaukee,
WI) and a flexible shaft attached to a custom-made stainless steel core head
via a Model H45 vibrator head. The core head accepted 7.6 cm diameter core
tubes. This coring device did not produce sufficient power to collect cores
more than 1.5 to 1.8 m long. To collect a core sample, (1) the sampling
vessel was three-way-anchored and its position determined, (2) a core tube was
slid into the core head and secured, (3) the coring apparatus was lowered into
the water and the vibrator head was turned on, (4) the apparatus was allowed
to penetrate the sediment until it stopped or until all of the core tube had
entered the sediment, (5) the apparatus was raised onto the deck of the
vessel, and (6) the core tube was removed from the vibrator head and the core
sample was removed from the core tube. The core samples were transported to
shore and separated into different fractions according to depth and identified
with X2, X3, 01, or 02 extensions to the sample number (Table 2.1). The
samples were placed in 10-L high density polyethylene screw-topped containers,
and manually homogenized by stirring for 2 to 3 min. The samples were then
packed in synthetic ice, and shipped to the National Fisheries Contaminant
Research Center (NFCRC) via over-night courier.
Sample Receipt
Upon receipt, the samples were inventoried. Sediment temperatures upon
receipt ranged from 8 to 21°C. The samples were then placed into 18.9 L
plastic secondary containers equipped with water-tight sealing lids, labelled,
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and stored (< 5 d) in the dark in a walk-in cooler at 4°C until the elutriates
were prepared (generally less than two weeks).
Sediment Homoaenization
All sediment samples were mixed at room temperature of about 20°C with
a modified 30.5 cm bench-top drill press (Dayton Model 3Z993). The drill-
press was modified by (1) extending the motor-support column 35 cm, and (2)
installing an adjustable speed, reversible, 1/2 HP, direct current motor
(General Electric Model 5BPB56KAA100) regulated with a SCR direct current
motor controller (Dayton Model 2M171C). This motor controlled the speed of
the auger between 0 and 500 RPM and direction of rotation. A limit-switch-
controlled, AC motor (Dayton, Model 4Z063A) raised and lowered the drill press
platform. Sediments were mixed with a stainless steel auger (7.6 cm diameter,
38 cm overall length, 25.4 cm bit length) obtained from Augers Unlimited,
Exton, PA. TeflonR spatulas and a 40 x 2 x 0.6 cm stainless steel bar was
used to remove sediment that adhered to the auger and sample containers.
The samples were mixed in the original shipping containers. A container
was placed on the platform of the sediment homogenizer directly under the
auger. The platform was then slowly raised to immerse the auger, which was
turning at minimum speed, into the sample. The platform was raised to a point
where the auger was about 2 cm from the bottom of the sample container. Then,
the auger speed was gradually increased to about 350 RPM. The sample was
allowed to mix for 10 min while the container was manually rotated on the
platform. The auger speed was reduced, the homogenizer platform was lowered,
and the stainless steel bar was used to probe the sides and bottom of the
sample container to loosen unmixed sediment. The unmixed sediment was moved
to the center of the sample container with the stainless steel bar, the
platform was raised, and the sample was mixed for an additional 10 min.
Between samples, the auger and other tools that were in contact with the
sample were (1) were rinsed with well water, (2) washed with soap and water,
and (3), sequentially rinsed with acetone, well water, 10% HC1, well water,
and deionized water (three rinses).
Elutriate and Pore Water Preparation
Elutriates were prepared immediately after homogenization by mixing one
part of sediment with four parts of reconstituted dilution water. The
dilution water (hardness 134 mg/L as CaCC^; alkalinity 60-65 mg/L as CaCO3; pH
7.8 to 8.0; conductivity 300 /unhos/cm; sulfate 72 mg/L) was reconstituted
according to procedures of Ingersoll et al. (1990). To prepare the
elutriates, a 200 ±0.05 g sub-sample of each homogenized sediment was removed
from the containers with a disposable polypropylene syringe (60 cc). The
syringe was modified by drilling a 0.9 cm hole in the tip of the syringe
barrel. The sub-sample was placed into a tared 1000-mL polypropylene
centrifuge bottle and weighed on a calibrated electronic pan balance (A&D
Engineering Inc. Model FX-3000). Then 800 g of the dilution water was added
to the tared centrifuge bottle, cap, and sediment sample.
The contents of the centrifuge bottle were then mixed end over end for
30 min in an extraction apparatus. This apparatus consisted of a 31- x 23- x
18-cm stainless steel box with a removable top and six 10- x 10- x 18-cm
compartments. One centrifuge bottle was placed into each compartment. The
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box was attached with a flexible bushing to the motor drive of a 1/10 HP, 14
RPM, shaded pole gear motor (Dayton Model 3M136A) supported by a metal frame
constructed of 2.54 cm tubing. The motor rotated the box and samples (end
over end) at 12 RPM on two lubricated pillow-block bearing assemblies.
After the 30-min mixing, the samples were centrifuged at 5,200 RPM
(7,000 x G) with a large capacity, refrigerated centrifuge (International
Equipment Co. Model PR-7000) equipped with a Model 966 rotor. The overlying
water was decanted through a U.S. Standard # 50 stainless steel sieve into a
3-L glass bottle. The elutriate samples were not filtered. Sub-samples of
each elutriate sample were removed from the 3-L bottles and stored (< 5 d) in
one-L amber bottles equipped with Teflon"-lined screw-on tops in the dark at
4°C until they were tested (generally less than one week).
Porewater samples were prepared by Battelle's Marine Sciences
Laboratory, Sequim Washington from about 40 L of sediment samples. Aliquots
of the 40-L samples were extracted in acid-cleaned 500-mL Teflon jars by
centrifugation in a modified clothing extractor at 2000 RPM for 15 min. The
pore water was decanted into clean 150-mL glass centrifuge tubes and then
centrifuged again at 2000 RPM for one h. Then the pore water was pipetted
without filtration into 500-mL acid-cleaned Teflon bottles, acidified to pH 2
with HN03, and stored at room temperature for metal analyses.
TEST PROCEDURES
Daphnia maana Tests
Daphnia roaqna were exposed in static acute toxicity tests for 48 h to
the full-strength (100%) elutriates and 50%, 25%, and 12.5% volumetric
dilutions of the 100% elutriates; an elutriate prepared from the control
sediment; and to a negative control of dilution water. The treatments were
not replicated. Graduated cylinders and pipettes were used to prepare
dilutions of each elutriate. The 100% elutriates and the dilution water were
aerated before dilutions were prepared to ensure that initial oxygen
concentrations were adequate among treatments at the beginning of each test.
Daphnids were cultured according to procedures described by the USEPA
(1984) as modified by Ingersoll et al. (1990). Adult daphnids were isolated
from laboratory cultures on the day before a test was started. On the day the
test was started (Day 0), daphnids (<24 h old) were removed from the isolated
culture and placed into 250 mL beakers containing 200 mL of culture water.
Before a test was started groups of the young daphnids were acclimated to the
dilution water over a 4-h period. The daphnids were placed first into a
solution containing 50% culture water and 50% dilution water for 2 h and then
into a solution containing 75% dilution water and 25% culture water for 2 h.
Smooth large-bore glass tubes were used to transfer ten of the daphnids, in
groups of five each, from the 75:25% acclimation solution directly into each
250-mL test beaker containing 200-mL of test solution. The treatments were
stocked in the following order: dilution water control, control sediment
elutriate (if tested), and 12.5, 25, 50, and 100% elutriate. Test temperature
was maintained at 20°C with a ventilated temperature-controlled water bath. A
16:8 (light:dark) photoperiod with a light intensity of about 25 to 30
footcandles was maintained with fluorescent and incandescent light bulbs
suspended above the water baths. Mortality was recorded in all treatments at
24 and 48 h of the test. Lack of mobility in response to prodding with a
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blunt probe during a 5-s observation period was the criterion used to
determine death.
Microtox1* Tests
All acute toxicity tests with the luminescent bacteria, (Photobacterium
phoaphoreum) were performed with a Model 500 Microtox" instrument at 15°C
(Microbics 1988). The instrument measured the light output of the luminescent
bacteria before and after exposure to dilutions of osmotically-adjusted
elutriate samples. Metabolic inhibition in the luminescent organisms occurs
if a sample is toxic, and the subsequent reductions in light output are used
to derive a dose-response curve from which the effective concentration (i.e.,
EC50) of the sample is determined.
The Microtox" tests were simplified through partial automation. A RS232
interface was used to transmit measured light intensities from the Microtox"
instrument to a Compaq" III (Model 40) personal computer. A manufacturer
supplied computer program (written in BASIC) was used to determine the Median
Effective Concentrations (EC50) of the sample. The computer program estimated
the EC50 value from linear regression equations obtained by plotting log
concentration vs. log gamma (normalized ratio of light lost during test to
light remaining at time of measurement). No light normalization was performed
for the 100% percent test (described below).
Elutriates samples were tested using either "100%" or "Standard" test
methods. Less toxic samples were tested undiluted in the 100% Test by
substituting sodium chloride for the Microbics" Osmotic Adjusting Solution
(MOAS) to osmotically adjust the sample. Elutriate dilutions up to 45% were
evaluated in the Standard Tests. Color corrections were not necessary for any
of the elutriate samples tested. For sediment samples evaluated with the 100%
method, the full-strength (100%) elutriate and 45, 22.5, 11.3, 5.6% dilutions
were tested. Occasionally, primary dilutions of the 100% elutriate were
prepared for samples showing high toxicity. In tests were a primary dilution
was required, 11.0, 5.5, 2.7, and 1.3% dilutions of the full strength
elutriates were evaluated. Acute toxicity values for each sample were
estimated as 5-and 15-min ECSOs, (the percent dilution of the elutriates that
produced a 50% reduction in light output of the luminescent bacteria within 5
or 15 min). In all toxicity tests, duration of exposure is an important
factor that influences effects. Generally, Photobacterium phosphoreum in the
Microtox8 test system react rapidly to organic compounds and toxicity is
frequently elicited within 5 min; however, metals take longer to elicit
toxicity in the Microtox system and require at least 15 min to stabilize
(MicrotoxR, 1988). The 5-min ECSOs were estimated as intermediate values to
provide information on the rate at which toxicity developed. Because the
sediment samples contained a mixture of organic and inorganic compounds, these
5-min ECSOs should not be used to evaluate their toxicity in the Microtox"
test.
Standard elutriate tests evaluated 45, 22.5, 11.3, and 5.6% dilutions of
the full-strength samples. Standard tests were conducted in the following
manner:
1) clean cuvettes were placed in the Reagent Well and in incubator block
Wells Al through A5 and Bl through B5 of the Microtox" instrument,
2) one tnL of deionized water was placed in the cuvette in the Reagent Well
to reconstitute the bacteria samples,
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3) 500 /il of diluent (2% sodium chloride solution) was added to the
cuvettes in Wells Bl through B5 with a micropipet,
4) one tnL of the diluent was added to the cuvettes in Wells Al through A4,
5) 250 fil of MOAS (22% sodium chloride to osmotically adjust the sample)
and 2.5 mL of elutriate or phenol standard were pipetted into the
cuvette in Well A5 and mixed by aspiration with a micropipet,
6) one mL of this osmotically adjusted elutriate or phenol standard was
used to prepare 50% serial dilutions in the cuvettes in Well A4, A3, A2,
and,
7) the dilutions were mixed by aspiration with a micropipet and allowed 5
min to reach test temperature of 15°C.
Vials of MicrotoxR Reagent (Photobacterium phosphoreuml were stored
frozen at -20°C until they were reconstituted on the day of the test. The
reagent was added to the 1 mL of cooled reconstitution solution in the Reagent
Well cuvette, swirled, and returned to the Well, where it was aspirated 20
times with a 500 /il micropipette. Ten (10) /il of this reagent solution was
pipetted into the cuvettes in Wells Bl through B5 and thoroughly mixed by
repeated aspiration with a 250 /il micropipette. The diluted reagent solutions
were held for 15 min to allow the temperature to stabilize to 15°C.
To begin a test, the reagent sample from Well Bl was placed in the
turret Well to calibrate the instrument. Illumination of the "Ready" light on
the display panel indicated that the instrument was properly calibrated. The
reagent samples in Wells B2, B3, and B4 were then placed in the turret, and
the initial light levels (I0) were recorded by the interfaced personal
computer. Then 500 /il of the samples in Wells Al (Blank), A2, A3, A4, and A5
were transferred, respectively, to the Bl, B2, B3, B4, and B5 Reagent Wells.
The samples and reagents were mixed by aspiration with the 500 /il micropipette
at least twice. Then light outputs of test cuvettes Bl through B5 were
recorded at 5 and 15 min. Dilutions lower than 5.6% were tested in the same
manner except that the serial dilutions were prepared from a 50% solution of
the full-strength elutriate sample.
The "100%" test methods used for undiluted elutriate samples differed
from the "Standard" test methods. The manufacturer-supplied computer program
could not be used in the 100% test, so light intensity values were recorded
manually and the ECSOs were calculated by manually entering the data into the
computer program. Further, the 100% test differed from the Standard test in
that initial I0 values were not recorded for each dilution. Thus, to
calculate the ECSOs, the initial I0 value obtained for sample Al was assumed
to be the same as the I0 values for samples A2 through A5. This assumption
may have diminished the accuracy of the test because the sample dilutions may
not have had the same initial luminescence. However, the potential loss of
accuracy associated with the 100% test was accepted in order to evaluate
greater than 45% dilutions of elutriate samples for toxicity.
The 100% test was performed in the following manner:
1) the lyophilized bacteria were reconstituted and placed in the Reagent
Well as previously described,
2) one mL of diluent was added to cuvettes Al, A2, A3, and A4,
3) analytical reagent grade sodium chloride (50 mg) and 2.5 mL of the
elutriate were added to cuvette A5 and mixed by aspiration to dissolve
the sodium chloride,
4) 500 fil of the sample was removed from cuvette A5 and discarded, and
another 1.0 mL was removed and mixed with the diluent in cuvette A4,
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5) one mL of the sample in A4 was used to prepare 50% serial dilutions in
Wells A3 and A2, and
6) then 1.0 mL of the sample in A2 was withdrawn and discarded.
The temperature of the samples were allowed to stabilized for 5 min to 15°C.
Ten pi of reconstituted reagent was mixed with the samples in Wells Al, A2,
A3, A4, and A5. After five min the instrument was calibrated by placing
cuvette Al (reagent blank) in the turret. Calibration light intensities were
recorded after 5 and 15 min for the samples in cuvettes Al through AS.
Microtox" tests were conducted to determine if the toxicity of
elutriates prepared from stored sediment samples changed over time.
Elutriates were prepared from mixed sediment samples each time stored sediment
samples were homogenized. The elutriates were tested with the same methods
described above with one exception. While original elutriates were stored for
about 1 week at 4°C in the dark before they were tested, subsequent elutriates
were tested within 24 h after they were prepared.
Water Quality Characterization
Immediately after preparation, water quality characteristics of the
dilution water and 100% elutriate samples were determined (APHA et al., 1975).
Dissolved oxygen (mg/L) was measured with a YSI Model 54-A oxygen meter.
Conductivity (/unhos/cm, corrected to 25°C) was measured with a YSI Model 33 S-
C-T conductivity meter. The pH and alkalinity (mg/L as CaCC>3) was measured
with an Orion 940E ionalyzer and a 81-72BN pH probe. Total hardness (mg/L as
CaCC>3) was determined by burette titration. Ammonia (mg/L) was measured with
an Orion 940E ionalyzer and a 95-12 ammonia electrode. Turbidity (NTU) was
measured with a Cole-Palmer Model 8391-35 turbidity meter. Unionized ammonia
was determined by converting the total ammonia measured in the samples to
unionized ammonia, and then correcting for pH and temperature (Thurston et al.
1974). After preparation of the dilution water and 100% elutriates, samples
for chloride (mg/L) were placed in 250 mL I-CHEM bottles, labeled, and stored
at 4 + 3°C until analysis with an Orion 940E ionalyzer and a 94-17B electrode.
The pH, dissolved oxygen, and conductivity were measured at the beginning and
end of each daphnid test in the 100 and 25% treatments, and in the dilution
water control.
About 500 mL of each 100% elutriate sample were placed in Teflon
bottles, acidified to pH 2 with redistilled hydrochloric acid, and shipped via
over-night courier to Battelle Marine Sciences Laboratory, Sequim, Washington
for metal analyses.
Elutriate and porewater samples were analyzed for Ag, As, Cd, Cr, Cu,
Hg, Ni, Pb, SB, and Zn. With the exception of Hg and Zn in elutriates, all
porewater and elutriate samples were analyzed for metals by direct injection
graphite furnace atomic absorption without sample preparation. The Zn in
elutriates was quantified by flame atomic absorption. The Hg in elutriates
were analyzed by cold vapor atomic fluorescence with sub-ng/L detection
limits. Organics prevalent in many of the samples were broken down before Hg
analysis by use of a bromine monochloride/UV oxidation procedure (Bloom and
Crecelius 1983).
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Quality Control and Quality Assurance
The precision of a test describes the degree to which data obtained from
replicate measurements differ. The accuracy of data is the difference between
measured values and true values. The accuracy of data obtained from toxicity
tests with field samples cannot be determined since the true values are not
known. Currently, no methods exist to directly measure the accuracy of
sediment toxicity tests. However, indirect estimates of the accuracy of data
from the elutriate toxicity tests were obtained by measuring the sensitivity
of test organisms to waterborne reference toxicants. Test acceptability was
determined by using methods described for waterborne acute and chronic
toxicity tests (USEPA 1985a).
Elutriates prepared from the AOC sediments, control sediment elutriates
and dilution water controls were tested. The control sediment was a fine soil
containing silt-clay sized particles which has previously been used in
sediment toxicity testing (Adams et al. 1985, Ingersoll and Nelson 1990). To
determine the acceptability of the toxicity tests, responses (e.g., survival)
of test organisms to elutriates prepared from sediments collected at the AOCs
were compared to their responses to the control sediment elutriate and the
dilution water control.
The ECSOs in the daphnid tests were calculated with either probit,
binomial, or moving average methods as appropriate (Stephen 1977). A
reference toxicant test with Instant Ocean" was conducted to evaluate the
sensitivity of daphnids used in the elutriate toxicity tests. The daphnids
were exposed in a 48-h static acute toxicity test to concentrations (1.5 to
33.0 g/L) of Instant Ocean" dissolved in well water. Results of this
reference toxicant test are shown in Table 2.2.
n
All Microtox tests were conducted with equipment, supplies, and
reagents that were recommended by, or obtained from the manufacturer. A 90
mg/L solution of phenol prepared in deionized water was used to conduct
reference toxicant tests at the beginning of each days test run. The EC50s
for the reference toxicant tests averaged 19.7% and ranged from 17.8% to 22.5%
for the full-strength phenol solution, which was within the manufacturer's
established acceptable range of 14 to 29%. These reference toxicant tests
were conducted concurrently with the first AOC sample elutriates (E-l)
MicrotoxR tests.
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RESULTS
INTER-AOC ELUTRIATE WATER QUALITY SUMMARY
Generally, a wide range in elutriate water quality characteristics was
measured across the AOCs. Ranges in elutriate water quality measurements
were: pH, 6.6 to 8.0; alkalinity, 80 to 400 mg/L as CaCOj; hardness, 115 to
283 mg/L as CaCC^; dissolved oxygen, 1.8 to 7.1 mg/L; conductivity 335 to 951
/unhos/cm; unionized ammonia, 0.01 to 0.59 mg/L; chloride 8.9 to 26.2 mg/L; arid
turbidity, 14.3 to 198.0 NTU (Table 2.3). Ranges of water quality
measurements for elutriates prepared from control sediment were: hardness, 118
to 120 mg/L as CaCC^; oxygen, 7.9 to 9.4 mg/L; conductivity, 238 to 259
^mhos/cm; chloride, 9 to 11 mg/L; turbidity, 29 to 54 NTU; pH, 7.0;
alkalinity, 44 mg/L as CaCOj; and unionized ammonia, 0.01 mg/L (Table 2.3).
Mean pH, alkalinity, hardness, conductivity, and chloride measurements were
relatively similar in elutriates across the AOCs (Table 2.3). Mean dissolved
oxygen was lowest in Indiana Harbor elutriates; was intermediate in elutriates
from Buffalo River and Saginaw River (third survey); and was highest in
elutriates from the first survey of Saginaw River (Table 2.3). Mean unionized
ammonia and turbidity in elutriates from Indiana Harbor and the first survey
of Saginaw River were similar, and were slightly higher than unionized ammonia
and turbidity in elutriates from Buffalo River and the third survey of Saginaw
River (Table 2.3).
INDIANA HARBOR
Elutriate Water Quality Characterization
Generally, measured water quality parameters were similar in the control
elutriate and Indiana Harbor elutriates. Exceptions were that alkalinity in
the Indiana Harbor elutriates was three to nine times higher than in the
elutriate prepared from control sediment; unionized ammonia ranged from 0.1 to
0.6 mg/L in the Indiana Harbor elutriates and was much lower (0.01 mg/L) in
the control elutriate; and turbidity was about two to four times higher in
Indiana Harbor elutriates than in the control elutriate (Table 2.3). Measured
water quality was also relatively similar among the Indiana Harbor elutriates
treatments except that alkalinity was about 4 times higher in elutriate sample
IH-01-10 compared to other elutriate samples (Table 2.3). During the daphnid
test with Indiana Harbor elutriates the pH, dissolved oxygen and conductivity
were similar across treatments (Table 2.4).
Elutriate samples from Indiana Harbor contained elevated total metal
concentrations compared to the control elutriate (Table 2.5). Generally, the
concentrations of metals in samples IH-01-04 and IH-01-10 were similar and
lowest; metal concentrations in samples IH-01-03, IH-01-05, IH-01-06, and IH-
01-08 were intermediate; and metal concentrations in sample IH-01-07 were the
highest (Table 2.5).
Total metal concentrations were also elevated in porewater samples
prepared from Indiana Harbor sediments. Pore water from sample IH-01-06
contained the lowest metal concentrations and samples IH-01-03 and IH-01-04
contained metal concentrations that were intermediate between concentrations
in samples IH-01-06 and IH-01-07 (Table 2.6).
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Toxicitv
Daphnia maana Tests —
All the Indiana Harbor elutriates, except the elutriate from sample IH-
01-06, were toxic to Daphnia maana. Elutriates from samples IH-01-03, IH-01-
05, IH-01-07, and IH-01-08 were the most toxic, and 48-h ECSOs for daphnids
(as percent dilutions of the 100% elutriates) ranged from 0.7 to 25.0%;
elutriates from samples IH-01-04 and IH-01-10 elicited intermediate toxicity
to daphnids and 48-h ECSOs ranged from 39.4 to 70.7%; and the undiluted (100%)
elutriate from sample IH-01-06 was not acutely toxic to daphnids (Table 2.7).
Microtox* Tests—
All of the Indiana Harbor elutriates were toxic to Photobacterium
phosphoreum in the MicrotoxR test. Elutriates from samples IH-01-06, IH-01-
08, and IH-01-10 were the most toxic with 15-min ECSOs ranging from 4.3 to
6.8% of the 100% elutriates (Table 2.7). Sample IH-01-03 elicited
intermediate toxicity with an EC50 of 20.3%, and samples IH-01-05, IH-01-04,
and IH-01-07 were the least toxic in the Microtox" test with ECSOs that ranged
from 27.3 to 34.4% dilutions. With the exception of sample IH-01-07, Indiana
Harbor elutriates elicited a slow response in the MicrotoxR tests, and 5-min
ECSOs were about half the 15-min ECSOs (Table 2.7).
BUFFALO RIVER
Elutriate Water Quality Characterization
The pH, alkalinity, hardness, dissolved oxygen, and conductivity of
elutriates prepared from Buffalo River sediments were generally similar. The
exceptions among the sample elutriates were that alkalinity was slightly lower
in samples BR-01-02 and BR-01-03; hardness was slightly lower in samples BR-
01-09; chloride was slightly higher in samples BR-01-01 and BR-01-02; and
turbidity was slightly lower in samples BR-01-01, BR-01-02, and BR-01-10
(Table 2.3). Unionized ammonia in elutriates from sample BR-01-01 was 68 to
137 times higher compared to elutriates from the other Buffalo River samples
which contained relatively low (0.01 to 0.02 mg/L) unionized ammonia
concentration (Table 2.3). During the daphnid test with Buffalo River
elutriates the pH, dissolved oxygen and conductivity were similar across
treatments (Table 2.4).
A wide range of metal concentrations was observed in Buffalo River
elutriates (Table 2.8). Zinc and Pb concentrations were higher in Buffalo
River elutriates compared to the other metals, and concentrations of Cr, Cu,
Ni, Pb, and Zn were highest in sample BR-01-06 (Table 2.8).
Buffalo River sediment pore waters contained low /tg/L concentrations of
metals, and concentrations of As, Cr, and Cu were higher compared to the other
metals (Table 2.9).
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Toxicitv
Daphnia maona Tests—
Elutriates prepared from Buffalo River sediments were generally less
toxic to daphnids than Indiana Harbor elutriates. Exceptions were the
elutriates from sample BR-01-03 for which the 48-h EC50 was 76.2%, and the
elutriate from sample BR-01-06 for which the 48-h ECSOs was 0.11% of the full-
strength elutriate (Table 2.7).
MicrotoxR Tests—
MicrotoxR indicated that the elutriate from sample BR-01-06 was the most
toxic with a 15-min EC50 of 6.7% of the full-strength elutriate (Table 2.7).
Elutriates from samples BR-01-01, BR-01-03, and BR-01-05 elicited intermediate
toxicity in the Microtox" test and 15-m ECSOs ranged from 40.5 to 65.8%
dilutions of the full-strength elutriates (Table 2.7). The remaining
elutriates from samples BR-01-02, BR-01-04, BR-01-07, BR-01-08, BR-01-09, and
BR-01-10 were not toxic in the Microtox" test (Table 2.7). Buffalo River
elutriates elicited a rapid response in the Microtox" tests, and 5- and 15-min
ECSOs were similar with the exception of sample BR-01-06 for which the 15-min
EC50 was about three times lower than the 5-min EC50 (Table 2.7).
SAGINAW RIVER (FIRST SURVEY)
Elutriate Water Quality Characterization
Measured water quality was generally similar in elutriates prepared from
sediments collected during the first survey of Saginaw River (Table 2.3).
Compared to the elutriate prepared from control sediment, alkalinity was two
to five times higher; ammonia was 6 to 283 times higher; chloride was 2 to 4
times higher; and turbidity was 3 to 8 times higher in the Saginaw River
elutriates (Table 2.3). During the daphnid test with Saginaw River (first
survey) elutriates the pH, dissolved oxygen and conductivity were similar
across treatments (Table 2.4). Elutriates prepared from Saginaw River
sediment samples collected during the first survey contained low ^g/L metal
concentrations, and concentrations of As, Cu, Pb, and Zn were higher compared
to the other metals (Table 2.10). Elutriate sample SR-01-06 contained the
highest concentrations of Cd, Cr, Cu, Ni, Pb, and Zn (Table 2.10). Pore water
from samples SR-01-03 and SR-01-10 contained low (< 4.0 /*g/L) metal
concentrations and metal concentrations in pore water from sample SR-01-06
were higher and ranged from < 1.0 to 36.0 /ig/L (Table 2.11).
Toxicitv
Daphnids maana Tests—
Generally, Saginaw River elutriates were not toxic to daphnids.
Exceptions were elutriates from SR-01-06 and SR-01-07 which elicited 48-h
ECSOs for daphnids of 12.5 to 25% dilutions of the full-strength elutriates
(Table 2.7).
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Microtox* Tests—
The Microtox" test indicated that the elutriate from sample SR-01-06
collected during the first survey of Saginaw River was toxic with an EC50 of
13.0% of the full-strength elutriate (Table 2.7). The remaining elutriates
from first survey samples SR-01-02, SR-01-03, SR-01-04, SR-01-07, SR-01-09,
and SR-01-10 were not toxic (EC50 > 100%). Essentially no differences were
observed between 5- and 15-min EC50 values for the Saginaw River (first
survey) elutriates.
SAGINAW RIVER (THIRD SURVEY)
Elutriate Water Quality Characterization
The water quality of elutriates from sediment samples collected during
the third survey of Saginaw River was generally similar (Table 2.3).
Elutriates prepared from core samples contained higher unionized ammonia
concentrations (0.1 to 0.3 mg/L) compared to the elutriates prepared from grab
samples (Table 2.3). During the daphnid test with Saginaw River elutriates
(third survey) the pH, dissolved oxygen and conductivity were similar across
treatments (Table 2.4). With the exception of Zn concentrations that ranged
from <7 to 148 jig/L, metals concentrations in elutriates prepared from
sediment grab samples collected during the third survey of Saginaw River were
< 10.5 /ig/L (Table 2.12). With the exception of Hg, concentrations of metals
in elutriates prepared from sediment core samples were generally higher than
in those prepared from grab samples (Table 2.12). Metal concentrations in
pore water from sediment samples collected during the third survey of Saginaw
River ranged from 0.002 /tg/L for Hg to 10.7 M9/L for Zn (Table 2.13).
Toxicity
Daphnia maona Tests—
None of the elutriates prepared from grab samples collected during the
third survey of Saginaw River were toxic to daphnids (Table 2.7). Elutriates
prepared from core samples were not test with daphnids.
MicrotoxR Tests—
Only one of the elutriates (SR-03-16) prepared from grab samples
collected during the third survey of Saginaw River were toxic in the MicrotoxR
test (Table 2.7). However, the Microtox" tests conducted with elutriates
prepared from the core samples indicated that samples SR-03-02-X2 and SR-03-
06-X2 were the most toxic with ECSOs that ranged from 31.0 to 34.0% dilutions
of the full-strength elutriates; the elutriate from core sample SR-03-06-01
elicited intermediate toxicity (EC50 of 54%); and the remaining elutriates
from third survey Saginaw River core samples elicited MicrotoxR ECSOs > 77%
(Table 2.7). Essentially no differences were observed between 5- and 15-min
EC50 values for the Saginaw River (third survey) elutriates prepared from all
grab samples and most of the core samples.
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MICROTOX* SEDIMENT STORAGE EVALUATIONS
With few exceptions, the MicrotoxR test indicated that the toxicity of
whole sediment samples did not change during storage up to 26 d in the
laboratory at 4°C. Indiana Harbor samples were stored the longest (26 d).
The remaining samples were stored between 21 and 26 d. Of the 19 samples
tested, only 2 elicited a different toxicity over time (Table 2.14). After 14
d of storage, elutriates prepared from sample IH-01-07 (Indiana Harbor)
elicited increased toxicity compared to the toxicity observed in initial
testing, and elutriates prepared from sample BR-01-03 (Buffalo River) did not
elicit toxicity but had been toxic in initial testing. All other samples
exhibited no changes in toxicity.
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DISCUSSION
Photobacterium phosphoreum (MicrotoxR) and Daphnia maqna have been
previously used to measure sediment toxicity (Burton et al. 1989, Giesy et al.
1990, Giesy and Hoke 1989, Ankley et al. 1989). In the present study, these
two organisms were used in elutriate toxicity tests to screen sediment samples
from each AOC for acute toxicity. The toxic effects observed in the Daphnia
maqna and MicrotoxR acute toxicity tests represent integrated responses to the
interaction (i.e., additive, synergistic or antagonistic) of multiple
contaminants that were present in the elutriates. Without applying Toxicity
Identification Evaluation (TIE) procedures (Mount and Anderson-Carnahan 1938,
Mount and Anderson-Carnahan 1989, Mount 1989) the toxicity observed in these
tests cannot be attributed to any particular chemical constituent of the
elutriates.
The rotary procedure used to prepare the elutriates for this study was
similar to elutriate preparation procedures that have been shown to produce
reasonable estimates of bioavailable contaminants in sediment samples (Daniels
et al. 1989). The elutriates from each AOC were analyzed for metals but not
for organic contaminants, and some of the elutriates contained elevated
concentrations of heavy metals. Albeit no analyses for organic contaminants
were performed on the elutriates, organic compounds were present in the whole
sediment samples from which the elutriates were prepared. Undoubtedly, some
fraction of these organic contaminants desorbed into the elutriate sample
during preparation. Ionic contaminants may desorb from sediment particles
agitated in water, and the concentration of these contaminants in the water
phase is dependent on chemical desorption processes which are classified as
either fast (taking place in minutes to hours) or slow (days to weeks)
(Karichoff 1980). In sediment systems, chemical desorption rates are
dependent on partition coefficients, organic carbon content of the sediment,
ionic strength of the aqueous phase, and contact time between the chemical and
the sediment. Thus, the elutriates tested in these studies probably contained
complex mixtures of both inorganic and organic compounds which singly or in
combination contributed to the observed toxicity.
INDIANA HARBOR
Daphnia maqna
With the exception of elutriate sample IH-01-06 which was not toxic at
full-strength (100%), all the elutriates prepared from Indiana Harbor
sediments were highly toxic to daphnids. The elutriate test used in the
present study is an efficient indicator of soluble constituents that may be
released from disturbed sediments and become available to organisms in the
water column. Contaminant-induced biological effects in aquatic organisms are
a function of biologically available contaminant concentrations and exposure
time. Daphnid ECBOs (as percent dilutions of the full-strength elutriates)
for the elutriates that were toxic ranged from 0.7 to 70.7% indicating that
the elutriates contained increasing concentrations of toxic components, or
toxic components that were differentially available to the organisms. These
results were confirmed by the presence of elevated heavy metal concentrations
in the elutriate samples. Furthermore, elutriate sample IH-01-07 which
contained the highest concentrations of metals also elicited the highest
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toxicity to daphnids (EC50 of 0.7%). Apparently contaminant availability was
lower in elutriate sample IH-01-06 compared to the other elutriates, since
this sample elicited no toxicity in daphnids, but it contained concentrations
of metals similar to some of the other samples which were toxic. The Indiana
Harbor elutriates contained elevated unionized ammonia concentrations that
probably also contributed to the observed responses; however, concentrations
of unionized ammonia in most of the elutriates were below concentrations (0.5
to 2.8 mg/L NH3) reported to be acutely toxic to daphnids (USEPA 1985b).
All of the elutriates probably contained toxic components other than
metals, since a wide array of contaminants such as organometals, polynuclear
aromatic hydrocarbons (PAHs), polychlorinated biphenols (PCBs), dioxins, and
pesticides were measured in the whole sediment samples from which the
elutriates were prepared. Refer to Chapter 4 in this report for information
on concentrations of contaminants in the whole sediment samples from Indiana
Harbor.
MicrotoxR
Indiana Harbor elutriates elicited substantial toxicity in the Microtox"
tests which was consistent with the elevated concentrations of metals in the
elutriate samples. The toxicity observed in the Microtox" test with Indiana
Harbor elutriates indicated that spatial differences in contaminant loads
exist within the AOC. Elutriates prepared from sample IH-01-08 collected from
the Lake George Branch, sample IH-01-10 collected from Grand Calumet Branch,
and sample IH-01-07 collected near the "Forks" (See Figure 1.1 in Chapter 1)
area elicited the highest toxicity. Generally, samples collected closer to
Indiana Harbor exhibited less toxicity. This distribution of toxicity may be
related to contaminant discharge patterns, the dredging and removal of
contaminated sediments from the main channel of the harbor, or differences in
contaminant cycling and distribution processes among the sampling stations
The presence of ammonia in the samples may not have influenced the
Microtox" tests because the sensitivity of Photobacterium phosphoreum to
ammonia is low. In porewater toxicity identification evaluations, MicrotoxR
was unresponsive to samples whose toxicity to C. dubia and fathead minnows was
associated primarily with ammonia (Ankley 1990). .
Subsequent testing of two elutriates prepared from sample IH-01-07
indicated that the sample was more toxic than it had been shown to be in the
initial elutriate test. This discrepancy cannot be explained, so the
MicrotoxR results for elutriate sample IH-01-07 must be cautiously
interpreted.
BUFFALO RIVER
Daphnia maona
Compared to Indiana Harbor, sediments in Buffalo River probably contain
lower concentrations of toxic components. Only Buffalo River elutriate
samples BR-01-03 (EC50 76.2%) and BR-01-06 (EC50 0.11%) were toxic to
daphnids, and the other elutriate samples were not toxic at full-strength.
These conclusions were supported in part by the concentrations of metals that
were measured in the elutriate samples. Whereas measured metal concentrations
were generally low and similar among the elutriate samples, concentrations of
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Pb, for example, in samples BR-01-03 and BR-01-06 were much higher, and were
1.0 to 3.0 times higher than the National Acute Water Quality Criteria (EPA
1986) for Pb (82.0 fxg/L, normalized to 140 mg/L hardness).
The Pb concentration in elutriate sample BR-01-05 was two times higher
than the National Water Quality Criteria, but the full-strength elutriate was
not toxic to daphnids. Apparently, there were differences in contaminant
availability to the daphnids or antagonistic interactions among the Buffalo
River elutriate samples tested. Differences in contaminant availability or
contaminant interactions may have resulted in the pattern of mortality that
prevented the calculation of an EC50 for daphnids exposed to dilutions of
elutriate BR-01-07. Despite having unionized ammonia concentrations exceeding
those reported to be acutely toxic to daphnids (0.5 to 2.8 mg/L NH3, USEPA
1985b), the elutriate prepared from BR-01-01 sediment was not toxic to Daphnia
macrna. The physical and chemical characteristics of sample BR-01-01 may have
reduced ammonia toxicity.
Daphnids in the Buffalo River elutriate tests were probably also exposed
to other contaminants in addition to the metals, such as organometals, PAHs,
PCBs, dioxins, and pesticides because they were measured in the whole sediment
samples. Refer to Chapter 4 in this report for information on concentrations
of contaminants in the whole sediment samples from Buffalo River.
Microtox
Elutriate BR-01-06 exhibited the highest MicrotoxR toxicity (15-min
EC50, 6.7%), followed by BR-01-01 (15-min EC50,40.5%), BR-01-03 (15-min EC50,
44.2%) and BR-01-05 (15-min EC50, 65.8%). Compared to Station BR-01-06,
elutriates prepared from sediments collected upstream (BR-01-05) and
downstream (BR-01-07) were considerably less toxic. This spatial distribution
of toxicity implies the presence of a localized source of pollution near
Station BR-01-06.
The toxic effects observed in the Microtox" tests appeared to be related
to the concentrations of select metals in the elutriates. With the exception
of Station BR-01-01, elutriates that exhibited Microtox" toxicity (BR-01-03,
BR-01-05 and BR-01-06) contained concentrations of Cu, Cd, and Pb equal to or
greater than the Chronic National Water Quality Criteria for these metals.
The concentrations of Cu and Cd measured in BR-01-01 approached the Chronic
Water Quality Criteria for these metals and may have been sufficient to elicit
the observed MicrotoxR toxicity. The increasing toxicity observed in BR-01-06
between 5- and 15-min readings suggest that the primary toxic agent is one or
more metals (Elnabarawy et al. 1988). However, as previously noted, the
toxicity measured in these elutriates cannot be attributed to individual
constituents of the elutriates (e.g., metals). Reported Microtox" 15-min
ECSOs for single metals are much higher for Hg, (29 ^g/L), Cu (250 j^g/L), Cd
(17000 pg/L) than those measured in the Buffalo River elutriates tested
(Elnabarawy et al. 1988).
SAGINAW RIVER (FIRST AND THIRD SURVEYS)
Daphnia maana
Sediment samples from Saginaw River contain lower concentrations of
contaminants compared to Indiana Harbor or Buffalo River. Only elutriate
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samples SR-01-06 (EC50 >12%) and SR-01-07 (EC50 >12) prepared from Saginaw
River (first survey) sediments were toxic to daphnids. All other elutriates
from the first and third survey of Saginaw River were not toxic to daphnids at
full-strength. These conclusions were supported by the generally higher
concentrations of metals and probably other toxic components (see Chapter 4)
not measured in elutriate SR-01-06 and SR-01-07 prepared from Saginaw River
(first survey) sediment samples.
MicrotoxR
All samples collected from Saginaw River during the first and third
surveys were evaluated with the 100% MicrotoxR assay because historical
information indicated that Saginaw River sediments contained low
concentrations of contaminants. The toxicity of effluents have been
determined within an order of magnitude using either the 100% or Standard
MicrotoxR tests and the 100% Microtox" test has been recommended for use with
fairly non-toxic samples (Tarkepea and Hansson 1989).
The low Microtox toxicity observed in the Saginaw River elutriates were
consistent with concentrations of metals measured in the samples. The
elutriate prepared from grab sample SR-01-06 was the only sample from either
the first or third survey that exhibited toxicity. This sample contained
concentrations of Cu and Cd higher than the Acute and Chronic Water Quality
Criteria for these metals. Further, the concentration of Pb was nearly 10
times higher than the Chronic Water Quality Criterion value. However, the
concentrations of Cu and Cd in SR-01-06 were lower than the ECSOs determined
for the individual metals (Elnabarawy et al. 1988) indicating that interactive
toxicity between these metals, or toxicity from other contaminants not
measured probably occurred. The presence of a contaminant source near Station
SR-01-06 is suggested since it was the only Saginaw River sample that
exhibited Microtox" toxicity.
Grab samples were collected from Station 06 in the Saginaw River during
both the first and third surveys, but measured metal concentrations and
Microtox1* toxicities differed for the two surveys. Metal concentrations in
elutriate sample SR-01-06 were 2 to 20 times higher than in SR-03-06 and
Microtox1* toxicity observed in these two samples was consistent with their
metal concentrations. These results indicate that considerable differences in
contaminant loading may exist in Saginaw River.
The results of Microtox tests on elutriates from core samples indicate
that historical patterns of contaminant distribution exist in the Saginaw
River. For example, while core sample SR-03-02-X2 (20 to 50 cm deep) was more
toxic than sample SR-03-02-X3 (50 to 88 cm deep), core samples SR-03-05-01 and
SR-03-05-02 did not exhibit increasing toxicity with depth. Further, at
Station 06, where four core samples were taken, differences in toxicity in
relation to core depth were observed. While core SR-03-06-01 (0 to 61 cm
deep) was more toxic than sample SR-03-06-02 (61 to 122 cm deep), sample SR-
03-06-X2 (an oily core sample taken between 13 to 33 cm deep) was the most
toxic core tested. The sample SR-03-06-X3, taken just below the oily core
between 33 to 81 cm deep elicited only slight toxicity. The heterogenous
distribution of sediment toxicity has been reported previously (Stemmer et al.
1990, Burton 1991). In addition to historical contamination, local
disturbances and re-suspension related to shipping and dredging activities in
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the Saginaw River may have contributed to the observed variation in toxicity
with core depth.
The Microtox" test was the only test conducted with elutriates prepared
from stored sediment samples. Microtox" elutriate test results demonstrated
(with few exceptions) that the toxicity of the stored samples did not change
appreciably during storage for up to 26 d, although sediment storage times no
longer than 2 weeks are recommended (ASTM E 1383-92 1991). Apparently,
changes in toxicity of the stored sediment samples as indicated by the
Microtox" test were minimized by maintaining the samples at 4°C in their
original, screw-topped shipping containers.
INTER-AOC TOXICITY COMPARISONS
Elutriates prepared from whole sediment samples are efficient indicators
of sediment-associated contaminants that may be released to the water column
(Shuba et al. 1977). The elutriates and pore waters evaluated in the present
study were centrifuged rather than filtered before they were tested. The
samples were centrifuged to minimize the possibility of reducing toxicity and
subsequently underestimating the potential toxicity of the sediments samples
from each AOC.
The chemical constituents of sediments are associated with the
interstitial water or surfaces of the solid or particulate phase (Engler
1980). These constituents are available at varying degrees for release to the
water column, ranging from essentially no release of insoluble components
associated with mineral lattices to the highly available components of the
interstitial water. The chemical constituents present in elutriates generated
in the present study probably represent interstitial water components and
loosely-bound, easily-exchangeable components of the solid phase sediment.
Apparently, the elutriate preparation process was highly efficient in
extracting many metals from field collected sediment samples. The elutriate
preparation methods used in the present study required mixing a 1:4 ratio of
sediment to water for 30 min. Mixing the sediments with oxygenated water may
have been sufficient to oxidize metal-AVS complexes. Once released from the
AVS-complexes, the metals were maintained in solution by the addition of HC1
to the samples. Assuming that pore water comprises 20 to 50% of field
collected sediments samples, elutriate preparation resulted in a 9- to 21-fold
dilution of the pore water present in the original samples. Despite this
dilution, elutriate samples contained higher metal concentrations (except Hg)
than full-strength porewater samples.
In contrast to other metals, Hg concentrations were frequently higher in
porewater samples compared to those in elutriates (IH-01-04, BR-01-01, SR-01-
06, and all samples from the third survey of Saginaw River). Hg has a strong
affinity for organic substances in soils and natural waters (Lindqvist et al.
1991). Whereas the elutriate preparation process was highly efficient in
extracting cationic metals from the sediment, apparently the process was less
effective in extracting Hg from the sediment. The lower Hg concentrations
measured in the elutriates probably represent a dilution of Hg which was
present in sediment pore water.
Elutriate toxicity tests have been recommended as a means of evaluating
the potential ecological effects of open-water disposal of dredged materials
(EPA/COE 1977). However, conclusions derived from elutriate toxicity tests in
relation to potential ecological effects at open-water sites where dredged
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materials are disposed may not be relevant for in-place contaminated sediments
in inland or coastal waters. For example, potential ecological effects at
open water disposal sites are believed to be minimal because of settling and
dilution that prevent contaminants released from the sediment from reaching
toxic levels; however, ecological effects may develop in inland or coastal
waters where dilution may not be sufficient and contaminants may be
continually released from sediments due to wave action, activity of organisms,
and human disturbances.
Responses observed in the MicrotoxR and daphnid elutriate tests indicate
that sediments in Indiana Harbor are more toxic than those in Buffalo River
which are more toxic than sediments in Saginaw River. Although the responses
observed cannot be related to any particular component in the elutriates
tested due to interactive toxicity that can occur in complex contaminant
mixtures, there were trends in the relation between concentrations of
individual metals and the responses observed. For example, Cu concentrations
in all the Indiana Harbor elutriates exceeded both the Acute and Chronic
National Water Quality Criteria; all the elutriates from Indiana Harbor
contained Pb concentrations that exceeded the National Acute Water Quality
Criteria; and concentrations of Hg and Cd (except sample IH-01-04) in the
Indiana Harbor elutriates exceeded the National Chronic Water Quality Criteria
(Figure 2.1). Also, unionized ammonia concentrations in all the Indiana
Harbor elutriates far exceeded the National Acute Water Quality Criteria
(Figure 2.2). The elutriates from Buffalo River were less toxic than those
from Indiana Harbor, and only three of the Buffalo River elutriates contained
concentrations of Cu and Pb that exceeded the National Acute Water Quality
Criteria and none contained concentrations of Cd and Hg that exceeded the
Acute Criteria (Figure 2.3). Moreover, unionized ammonia in all the Buffalo
River elutriates (except sample BR-01-01) were below the National Acute
Criteria (Figure 2.2). The Saginaw River elutriates were less toxic than
elutriates from Indiana Harbor and Buffalo River with only one sample
containing Cu and Cd concentrations that exceeded National Acute and Chronic
Criteria, although some of the samples exceeded the Chronic Criteria for Pb
and Hg (Figure 2.4). Unionized ammonia in the Saginaw River elutriates were
elevated but were lower than those in the Indiana Harbor elutriates (Figure
2.2) that were more toxic.
Thirty-three percent of the 30 elutriate samples tested with daphnids
were conservatively identified as toxic, i.e., ECSOs of less than the full-
strength elutriate were determined. Fifty-three percent of the 39 samples
tested with Microtox" were conservatively identified as toxic. These results
indicate that the Microtox* test is more sensitive than Daphnia maqna tests
for sediment bioassessments. The Microtox" test has been previously reported
to be more sensitive than the D. maana test for assessing sediment toxicity
(Giesy and Hoke 1989). However, because bacteria and protozoans did not
exhibit a clear, interpretable response in elutriate tests, their use in
sediment testing was discouraged (Shuba et al. 1977). Results of the present
study indicate that the daphnid and MicrotoxR elutriate tests are useful tools
for assessing sediment toxicity.
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Karichoff, S.W. 1980. Sorption kinetics of hydrophobic pollutants in natural
sediments. In R.A. Baker, ed., Contaminants and Sediments, Volume 2,
Analytical Chemistry and Biology. Ann Arbor Science Publications Inc.
Long, E.R. and P.M. Chapman. 1985. A sediment quality triad: Measures of
sediment toxicity and infaunal community composition in Puget Sound. Mar.
Pollut. Bull. 16:405-415.
Lindqvist, O., K. Johansson, M. Aastrup, A. Anderson, L. Bringmark, G.
Hovsenius, L. Hakanson, A. Iverfeldt, M. Meili, and B. Timm. 1991. Mercury
2-22
-------
in the Swedish environment: Recent research on causes, consequences and
corrective methods. In O. Lindqvist, ed., Special Issue. Air and Soil
Pollut. 55:109-129. Kluwer Academic Publishers, The Netherlands.
Micorbics" Incorporated. 1988. A Mircrotox Manual: How to run toxicity
tests using the MicrotoxR model 500. 2232 Rutherford Road, Carlsbad, CA
92008.
Mount, D.I., and L. Anderson-Carnahan. 1988. Methods for aquatic toxicity
identification evaluations: Phase I toxicity characterization procedures.
EPA/600/3-88/034. U.S. EPA Environmental Research Laboratory, National
Effluent Toxicity Assessment Center, Duluth, MN.
Mount, D.I., and L. Anderson-Carnahan. 1989. Methods for aquatic toxicity
identification evaluations: Phase II toxicity identification procedures.
EPA/600/3-88/035. U.S. EPA Environmental Research Laboratory, National
Effluent Toxicity Assessment Center, Duluth, MN.
Mount, D.I. 1989. Methods for aquatic toxicity identification evaluations:
Phase III toxicity confirmation procedures. EPA/600/3-88/036. U.S. EPA
Environmental Research Laboratory, National Effluent Toxicity Assessment
Center, Duluth, MN.
NRC (National Research Council). 1989. Contaminated marine sediment —
assessment and remediation. Committee on Contaminated Marine Sediments,
Marine Board, Commission on Engineering and Technical Systems. National
Acedemy Press, Washington, D.C.
Palermo, M.R. 1986. Development of a modified elutriate test for estimating
the quality of effluent from confined dredged matgerial disposal areas.
Tecnical Report D-86-4, US Army Engineer Waterways Experiment Station,
Vicksburg, MS.
Prater, B.L. and M.A. Anderson. 1977. A 96-h sediment bioassay of Duluth and
Superior Harbor basins (Minnesota) using Hexaqenia limbata. Asellus communis,
Daphnia maqna. and Pimephales promelas as test organisms. Bull. Environ.
Contam. Toxicol. 18:159-169.
Seelye J. G., and M.J. Mac. 1984. Bioaccumulation of toxic substances
associated with dredging and dredged material disposal: A literature review.
Final Report EPA-905/3-84-005. Great Lakes National Program Office, U.S.
Environmental Protection Agency, Chicago, IL.
Shuba, P.J., J.H. Carroll, and K.L. Wong. 1977. Biological assessment of the
soluble fraction of the standard elutriate test. Technical Report D-77-3,
U.S. Army Waterways Experimental Station, CE, Vicksburg, MS.
Shuba, P.J., H.E. Tatem, J.H. Carrol. 1978. Biological assessment of methods
to predict the impact of open-water disposal of dredged material. Technical
Report D-78-50, Environmental Laboratory, U. S. Army Engineer Waterways
Experimental Station, Vicksburg MS.
2-23
-------
Stephan, C.E. 1977. Methods for calculating an LC50. In F.L. Mayer and J.L.
Hamelink, eds., Aquatic toxicology and hazard evaluation. ASTM STP 634.
American Society for Testing and Materials, Philadelphia, PA, pp. 65-84.
Thurston, R.V, R.C. Russo and K. Emerson. 1974. Aqueous ammonia equilibrium
calculations. Technical Report No. 74-1 (MSU-FBL TR 74-1), Fisheries Bioassay
Laboratory, Montana State University, Bozeman, MT.
Tarkepea, M, and M. Hansson. 1989. Comparison between two Microtox test
procedures. Ecotoxicol. Environ. Saf. 18:204-210.
U.S. Environmental Protection Agency/Corps of Engineers Technical Committee on
Criteria for Dredged and Fill Material. 1977. "Ecological Evaluation of
Proposed Discharge of Dredged Material into Ocean Waters; Implementation
Manual for Section 103 of Public Law 92-532 (Marine Protection, Research and
Sanctuaries Act of 1972)," July 1977 (Second Printing April 1978).
Environmental Effects Laboratory, U. S. Army Engineer Waterways Experiment
Station, Vicksburg, MI.
U.S. Environmental Protection Agency. 1984. Interim procedures for
conducting the Daphnia maqna toxicity assay. Environmental Research
Laboratory, Duluth, MN and Environmental Monitoring System Laboratory, Office
of Research and Development, Las Vegas, NV.
U.S. Environmental Protection Agency. 1985a. Methods for measuring the acute
toxicity of effluents to aquatic organisms. EPA 600/4-85-013. USEPA EMSL,
Cincinnati, OH.
U.S. Environmental Protection Agency. 1985b. Ambient water quality criteria
for ammonia. EPA 440/5-85-001. U.S. EPA, Office of Water Regualations and
Standards, Criteria and Standards Division. Washington, D.C.
U.S. Envionmental Protection Agency. 1986. Quality Criteria for Water 1986.
EPA 440/5-86-001. U.S. EPA Office of Water Regulations and Standards,
Washington, DC.
2-24
-------
Table 1. Sample numbers and sample station locations for sediments collected from three Great Lakes
Areas of Concern (AOC) and tested at the National Fisheries Contaminant Research Center (NFCRC).
AOC
and sample
number1
Indiana Harbor
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Buffalo River.
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Saainaw River,
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
Water depth
fm)
. Indiana
ND2
ND
ND
ND
ND
ND
ND
New York
5.5
4.6
6.1
ND
6.7
6.1
6.1
ND
0.9
ND
Michiaan f first survev)
ND
0.9
ND
0.6
ND
2.7
ND
Core depth
(m)
NA3
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
Location of
samolina station
Latitude
41°
41°
41°
41°
41°
41°
41°
42°
42°
42°
42°
42°
42°
42°
42°
42°
42°
43°
43°
43°
43°
43°
43°
43°
40.42'
40.07'
39.65'
39.30'
38.75'
38.81'
38.38'
51.42'
52.18'
52.66'
51.97'
51.71'
51.79'
51.63'
51.69'
51.68'
51.94'
37.42'
37.25'
36.84'
36.74'
36.46'
33.76'
32.66'
Longitude
87°
87°
87°
87°
87°
87°
87°
78°
78°
78°
78°
78°
78°
78°
78°
78°
78°
83°
83°
83°
83°
83°
83°
83°
26.35'
26.17'
27.09'
27.58'
28.34'
28.84'
28.28'
52.04'
52.28'
53.06'
52.14'
52.04'
51.25'
50.93'
50.73'
50.02'
49.05'
50.53'
50.58'
51.40'
52.16'
53.17'
54.57'
53.05'
-------
Table 1 continued
AOC
and sample
number1
Water depth Core depth
(m\ fml
Sacrinaw River, Michiaan ( third survevl
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
6.
2.
1.
0.
0.
6.
0.
2.
2.
1.
1.
0.
0.
0.
0.
4
0
8
7
9
7
8
0
0
8
8
7
7
7
7
NA
NA
NA
NA
NA
NA
NA
0.2-0.
0.5-0.
0.0-0.
0.6-1.
0.0-0.
0.6-1.
0.1-0.
0.3-0.
5
9
6
2
6
2
3
8
Location of
samolina station
Lati
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
.tude
45.
37.
36.
36.
35.
36.
34.
37.
37.
36.
36.
36.
36.
36.
36.
30'
42'
72'
74'
33'
70'
55'
42'
42'
72'
72'
74'
74'
74'
74'
Lonaitude
83
83
83
83
83
83
83
83
83
83
83
83
83
83
83
o
0
o
o
o
o
o
o
0
0
o
0
0
o
0
47.
50.
51.
52.
53.
51.
54.
50.
50.
51.
51.
52.
52.
52.
52.
48'
53'
67'
16'
88'
80'
16'
53'
53'
67'
67'
16'
16'
16'
16'
The Large Lakes Research Station Number includes the site code, survey number, transect number, and
the sample fraction number (core samples only).
ND
not Determined
NA = Not applicable
-------
Table 2. Dissolved Oxygen (DO), pH, and survival of Daphnia maana during a 48-h reference toxicant test with Instant
Ocean1* salinity.
shown.
Instant oceanR
salinitv (0/001
Control
(well water)
33.0
19.8
11.8
7.1
4.2
2.5
1.5
The 48-h EC50 was 25.3 with 95% confidence limits
DO (ma/L) DH Sui
0-h 48-h 0-h 48-h 24-h
7.9 8.2 8.4 8.3 100
7.1 8.0 8.0 8.1 0.0
0.0
0.0
7.5 8.1 8.2 8.2 100.0
100.0
100.0
7.9 8.0 8.3 8.3 100.0
of 21.6 - 13
rvival (%>
48-h
100
80.0
100.0
100.0
100.0
-------
Table 3. Measured elutriate water qualtiy for each area of concern (AOC) and the mean with standard error (in parentheses
across stations for each AOC.
AOC and
sample
number
Indiana Harbor
Control
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Means
Buffalo River
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Means
Saginaw River
(First Survey)
Control
SR-01-02
SR-01-03
SR-01-04
SR-01-06
OH
6.98
7.64
7.78
7.89
7.26
7.51
7.16
7.42
7.5
(0.11)
9.17
7.98
7.08
7.01
7.38
7.24
7.20
7.19
7.05
7.21
7.45
(0.21)
7.02
7.43
7.48
7.57
7.55
Alkalinity, Hardness,
(mg/L as (mg/L as D.O.,
CaCC^l CaCCM fma/L)
44.00
120.00
160.00
112.00
160.00
146.00
160.00
400.00
163
(36.62)
108.00
80.00
82.00
116.00
102.00
112.00
116.00
146.00
134.00
106.00
110.20
(6.43)
44.00
168.00
210.00
154.00
118.00
118.00
155.00
185.00
150.00
168.00
115.00
160.00
183.00
154
(9.32)
160.00
160.00
150.00
160.00
160.00
150.00
150.00
180.00
160.00
160.00
159.00
(2.77)
120.00
208.00
256.00
200.00
120.00
7.90
3.50
4.00
4.00
3.6
3.30
1.80
3.20
3.91
(0.62)
4.40
8.20
4.90
3.40
4.00
3.20
3.00
3.00
3.00
3.20
4.03
(0.51)
9.40
3.10
2.90
3.20
3.80
Unionized
Conductivity, ammonia, Chloride
(^mhos/cm) (ma/Li (mcr/L)
259.46
335.14
432.43
362.16
486.49
383.78
443.24
951.35
456.76
(74.91)
464.86
400.00
347.03
367.57
335.14
367.57
378.38
410.81
421.62
378.38
387.14
(12.10)
237.84
507.03
498.38
432.43
356.76
__
0.12
0.09
0.24
0.27
0.37
0.09
0.59
0.25
(0.07)
1.37
0.03
0.02
0.02
0.02
0.02
0.01
0.02
0.02
0.01
0.15
(0.14)
0.01
0.05
0.07
0.08
1.75
10.58
26.21
10.58
8.25
24.16
10.58
11.88
18.88
15.14
(2.46)
43.80
33.10
14.30
29.20
15.40
19.70
14.90
18.20
15.60
15.40
21.96
(3.18)
8.90
34.60
20.20
18.90
16.70
, Turbidity,
(NTU)
53.50
143.00
101.00
96.40
78.50
95.00
198.00
139.80
113.15
(15.99)
72.20
14.302
112.00
115.00
91.00
156.00
121.60
154.00
160.00
39.80
103.. 59
(15.70)
29.00
57.00
63.00
69.00
64.20
-------
Table 3 continued..
AOC and
sample
number
Saginaw River
SR-01-07
SR-01-09
SR-01-10
Mean
Saginaw River
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02 -X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Mean
DH
(first
7.52
7.54
7.71
7.48
(0.07)
(third
6.91
6.65
6.96
7.09
7.14
7.16
7.36
7.52
7.74
7.74
7.47
7.81
8.00
7.49
7.81
7.39
(0.10)
Alkalinity,
(mg/L as
CaCOn
survey)
164.00
146.00
106.00
138.75
(17.59)
survey >
104.00
128.00
110.00
92.00
126.00
130.00
162.00
144.00
136.00
132.00
168.00
140.00
116.00
148.00
108.00
129.60
(5.50)
Hardness,
(mg/L as
CaCO3>
196.00
192.00
176.00
183.50
(16.08)
150.00
180.00
170.00
160.00
170.00
180.00
190.00
200.00
190.00
172.00
192.00
160.00
160.00
192.00
172.00
200
(3.82)
D.O.,
(ma/Li
3.30
3.10
4.80
4.20
(0.77)
3.40
3.50
4.00
7.10
3.20
3.00
2.50
4.30
5.70
4.20
4.50
4.40
4.30
2.70
3.70
4.03
(0.30)
Conduct ivity ,
(^mhos/cm)
430.27
410.81
340.54
401.76
(31.31)
335.14
400.00
378.38
345.95
400.00
400.00
486.49
513.51
486.49
410.81
518.92
410.81
421.62
513.51
454.05
431.71
(15.58)
Unionized
ammonia,
(mg/L)
0.09
0.08
0.04
0.27
(0.21)
0.01
0.01
0.01
0.01
0.02
0.03
0.09
0.20
0.25
0.16
0.16
0.28
0.32
0.17
0.21
0.13
(0.03)
Chloride,
(mg/L>
20.40
21.30
17.30
19.79
(2.53)
9.50
15.60
15.60
14.50
14.10
15.90
27.50
17.50
18.10
16.60
22.10
19.60
20.50
24.60
19.20
18.06
(1.15)
Turbidity,
(NTU)
66.90
36.00
27.70
51.60
(6.24)
36.20
17.60
27.50
14.20
40.90
66.20
75.50
100.00
38.30
72.50
75.00
75.00
23.00
159.00
25.00
56.39
(10.01)
-------
Table 4. Mean and standard error of the mean (in parentheses) water qualtiy
during 48-hour acute toxicity tests with Daphnia magna exposed to elutriates
from each Area of Concern (AOC).
AOC and
sample
number
DH
D.O.,
(ma/L\
Conductivity,
(umhos/cm)
Indiana Harbor
Control
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Buffalo River
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Saginaw River
Control
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
(first
7.
8.
8.
8.
7.
8.
8.
8.
18
26
31
31
33
11
30
26
64
39
24
31
32
14
23
29
17
25
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
.05)
.01)
.06)
.01)
.05)
.08)
.02)
.03)
.25)
.0)
.17)
.12)
(0.13)
(0
(0
(0
(0
(0
.14)
.12)
.10)
.16)
.13)
8
8
8
8
8
8
8
8
9
9
9
9
9
9
9
9
9
9
.45
.20
.35
.13
.18
.05
.50
.45
.33
.85
.90
.70
.70
.10
.63
.60
.43
.50
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
.03)
.19)
.09)
.19)
.13)
.41)
.07)
.17)
.12)
.25)
.37)
.24)
.17)
.11)
.22)
.23)
.17)
.19)
276.
330.
369.
332.
374.
363.
391.
567.
356.
313.
329.
352.
332.
353.
324.
338.
334.
320.
75
81
19
16
87
52
08
57
76
51
73
70
43
01
32
74
23
00
(13.25)
(13.26)
(29.52)
(14.56)
(29.40)
(18.31)
(23.71)
(113.96)
(36.66)
(34.47)
(41.05)
(31.78)
(31.94)
(23.82)
(5.58)
(17.30)
(16.61)
(7.23)
survey )
98
27
36
34
91
24
17
15
(0
(0
(0
(0
(0
(0
(0
(0
.25)
.22)
.24)
•19)
.22)
.17)
.23)
.21)
9
9
9
9
9
9
9
9
.05
.08
.10
.17
.05
.43
.30
.48
(0
(0
(0
(0
(0
(0
(0
(0
.40)
.19)
.28)
.35)
.29)
.49)
.45)
.51)
294.
403.
481.
400.
339.
356.
372.
334.
87
51
62
27
19
21
16
05
(19.02)
(33.23)
(76.94)
(45.39)
(20.39)
(62.33)
(29.76)
(11.09)
-------
Table 4 continued.
AOC and
sample
number
PH
D.O. ,
(ma/L)
Conductivity,
(umnos/cm)
Saginaw River (third survey)
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
7.
8.
11
08
08
11
08
06
06
00
14
11
08
01
16
87
11
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
09)
11)
12)
11)
14)
14)
15)
11)
08)
11)
14)
07)
07)
16)
07)
8.22
8.22
8.14
8.10
8.24
8.16
8.26
7.95
9.33
8.90
8.58
8.44
9.08
7.30
8.83
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(1.
(0.
(0.
(0.
(0.
(1.
(0.
(0.
55)
52)
57)
48)
62)
61)
59)
02)
87)
91)
72)
86)
04)
96)
99)
321.
355.
352.
327.
343.
352.
386.
420.
377.
340.
409.
341.
380.
450.
400.
30
68
43
57
79
43
60
54
57
54
46
19
54
00
54
(9.
(22
(20
(13
(20
(22
(35
(60
(35
(38
(45
(31
(31
(57
(28
16)
.31)
.33)
.63)
.41)
.09)
.91)
.35)
.63)
.27)
.78)
.00)
.34)
.39)
.06)
-------
Table 5. Concentrations of metals in elutriates prepared from Indiana Harbor sediment samples.
Sample number
FLOR-01-01
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Ag As
<0.03 <1.8
<0.03 15
<0.03 5.5
0.21 9.8
0.26 10.2
1.3 152
<0.03 24.8
0.45 13.0
Metal
Cd
0.05
3.3
1
2.9
2.1
69
5.8
2.6
concentration (/*g/L)
Cr
1.1
61
37
73
86
4700
196
73
Cu
2.6
65
28
42
36
1329
70
23
Hg
0.0011
0.0383
0.0233
0.0345
0.0772
0.6670
0.1203
0.0046
Ni
2.5
23
8.8
18
15
650
46
8.8
Pb
12
489
293.9
391.9
343.9
11756
2449
637
Zn
6.9
517
183
535
348
10870
814
248
TABLE-2 . 5
-------
Table 6. Concentration of metals in Indiana Harbor sediment porewater samples. The ranges for replicates are shown in
parentheses.
Metal concentration fua/L)
Sample
IH-01-03
IH-01-04
IH-01-06
IH-01-07
Aa
<0.05
<0.05
<0.05
<0.05
As
2.3
3.7
(2.9-4.4)
<1.0
3.2
Cd
1.1
1.0
0.19
14.3
Cr
13.2
28.6
(27.3-29.8)
2.46
350.0
Cu
21.4
21.3
(19.1-23.4)
5.3
126.3
Ha
0.02
0.03
(0.02-.03)
0.004
0.06
Ni
10.4
7.5
(5.8-9.2)
5.9
171.4
Pb
96.8
92.8
(89.5-96.0)
5.3
1284.0
Zn
60.8
50.3
(49.6-50.9)
6.3
1081.0
-------
Table 7. Acute toxicity of Indiana Harbor, Buffalo River, and Saginaw River elutriates to Daohnia macrna and
Photobacteria phosphorum (MicrotoxR) . The 48-h ECSO's for daphnids and 5 and 15 min ECSO's for MicrotoxR (95%
confidence intervals in parentheses) are percent dilutions of full-strength elutriate samples. Survival of daphnids in
dilution water controls ranged from 80 to 100%.
Sample
number
Daphnia maqna
(48 h>
EC50 and 95% confidence
MicrotoxR
(5 min)
interval
MicrotoxR
f!5 min)
Indiana Harbor
Control
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
>100
1.0 (0.008 - 9.6)
70.7 (25 - 100)
25.0 (18 - 100)
>100 (NDa)
0.7 (0.001 - 3.1)
5.4 (2.5 - 11.0)
39.4 (30.2 - 53.0)
>100
33.6 (29.0 - 39.0)
58.7 (27.6 - 124.8)
48.4 (38.2 - 61.2)
9.0 (8.3 - 9.8)
37.2 (27.3 - 50.8)
12.1 (11.5 - 12.7)
12.9 (12.2 - 13.7)
>100
20.3 (18.7 - 21.9)
31.5 (27.1 -36.6)
27.3 (25.4 - 29.3)
4.3 (3.2 - 5.8)
34.4 (23.6 - 50.2)
6.0 (5.4 - 6.7)
6.8 (5.8 - 7.9)
Buffalo River
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
>100 (ND)
>100 (ND)
76.2 (50 - 100)
>100 (ND)
>100 (ND)
0.11 (0.001 - 12.5)
ND
<100 (ND)
>100 (ND)
>100 (ND)
33.4 (23.6 - 47.4)
>100 (ND)b
40.8 (38.5 - 43.3)
>100 (ND) b
63.5 (37.3 - 108.3)
19.3 (17.1 - 21.8)
55.5 (41.7 - 73.8)
>100 (ND)b
>100 (ND)b
>100 (ND)b
40.5 (25.4 - 64.4)
>100 (ND)b
44.2 (32.8 - 59.5)
>45 (ND)
65.8 (52.5 - 82.4)
6.7 (6.0 - 7.5)
>45 (ND)
>100 (ND)b
>100 (ND)b
>100 (ND)b
Saainaw River, first survey
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
>100 (ND)
>100 (ND)
>100 (ND)
>12.5 - 100
>12.5 - 100
>100 (ND)
>100 (ND)
>100 (ND)b >100 (ND)b
>100 (ND)b >100 (ND)b
>100 (ND)b >100 (ND)b
14.1 (12.0 - 16.6) 13.0 (9.5 -17.7)
>100 (ND)b >100 (ND)b
>100 (ND)b >100 (ND)b
>100 (ND)b >100 (ND)b
-------
Table 7 continued
EC50 and 95%
Sample
number
Saqinaw River,
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Daphnia
(48 h)
confidence interval
MicrotoxR
(5 min)
MicrotoxR
(15 min)
third survey
>100
>100
>100
>100
>100
>100
>100
ND
ND
ND
ND
ND
ND
ND
ND
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
>100
>100
>100
>100
>100
>100
>100
43.1
ND
80.5
88.1
36.1
76.5
30.5
71.3
100
>100
>100
>100
>100
95.0
>100
34.0
96.8
77.3
ND
53.7
>100
31.1
95.7
(ND)b
(ND)b
(ND)b
(ND)b
(ND)b
(37.2 -242. 9)b
(ND)b
(26.7 - 43.4)b
(0.26 - >100)b
(56.3 - 105. 9)b
(42.3 - 68.3)b
(ND)b
(22.4 - 43.1)b
(64.0 - 143. l)b
a Not determined
100 % elutriates testesd
-------
Table 8. Concentrations of metals in elutriates prepared from Buffalo River sediment samples.
Metal concentration (/*g/L)
Sample Number
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Ag
<0.03
<0.03
0.33
<0.03
0.12
0.04
0.04
<0.03
<0.03
<0.03
As
27
0.7
15
8
8.4
17
12
12
9.8
2.6
Cd
1.5
<0.15
2.2
0.83
2.9
2
1.2
0.61
0.44
<0.15
Cr
22
1.4
44
10
24
31
10
2.3
1.3
0.9
Cu
14
3.2
34
16
30
60
17
11
7.4
4.8
Hg
0.005
0.004
0.01
0.004
0.009
0.10
0.007
0.006
0.003
0.002
Ni
6.6
<5
5.5
<5
7.7
11
9.9
<5
<5
<5
Pb
45
8.5
115
40
156
238
62
25
23
13
Se
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
Zn
122
22
150
60
161
306
67
28
28
17
-------
Table 9. Concentration of metals in Buffalo River sediment porewater samples. The ranges for replicates are shown in
parentheses.
Metal concentration (ua/L)
Sample
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Aa
<0.05
<0.05
<0.052
<0.05
<0.052
As
14.5
1.8
(1.2-2.3)
1.7
<1.0
1.7
cd
0.1
<0.07
<0.07
<0.07
<0.07
Cr
2.5
0.7
0.3
0.2
0.27
Cu
3.3
1.9
(1.3-2.6)
1.65
1.65
2.3
Ha
*0.1
(+0.003)
0.009
(.0098-. 0096)
0.004
0.002
0.002
Ni
<4.0
<4.0
<4.0
<4.0
<4.0
Pb
5.7
1.8
(1.6-2)
0.8
<0.7
<0.7
Zn
0.9
<0.93
<0.93
<0.93
<0.93
-------
Table 10. Concentrations of metals in elutriates from sediment samples collected during the first Survey of Saginaw
River.
Metal concentration (/xg/L)
Sample number
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
Aa
<0.09
<0.09
<0.09
<0.09
<0.09
<0.09
<0.09
As
5.9
8.6
5.6
4.6
3.4
5.3
2.2
Cd
0.17
0.11
0.35
7.4
0.24
0.11
<0.06
Cr
3.1
3.0
4.8
253.0
2.6
1.2
1.2
Cu
7.6
5.6
10.4
98.0
7.2
3.6
<1.2
Ha
0.0057
1.00
0.003
0.005
0.002
0.002
0.008
Ni
<5.0
<5.0
<5.0
124.0
<5.0
<5.0
<5.0
Pb
11.2
11.8
15.1
39.2
11.2
5.0
3.4
Zn
28.0
28.0
50.0
139.0
33.0
28.0
17.0
-------
Table 11. Concentration of metals in porewater from sediments collected during the first survey of Saginaw River.
Metal concentration (ua/Ll
Sample number
SR-01-03
SR-01-06
SR-01-10
Aa As
<0.05 <1.0
<0.052 <1.0
<0.052 1.7
Cd
<0.07
0.84
<0.066
Cr
0.8
24.4
0.33
Cu
<0.57
19.8
2.6
Ha
0.
0.
0.
0034
006
002
Ni
<4.0
97.8
<4.0
Pb
<0
36
0.
.7
.2
81
Zn
<0.
19.
<0.
93
0
93
-------
Table 12. Concentrations of metals in elutriates prepared from sediments collected during the third survey of Saginaw
River. The ranges for replicates are shown in parentheses.
Metal concentration
Samole number
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Aa
<0.085
<0.085
<0.085
0.096
<0.085
0.12
0.12
1.09
0.19
(.15-. 24)
0.54
0.29
0.73
0.07
1.36
0.13
As
1.7
2.6
2.6
2.8
(2.6-2
2.3
3.2
5.2
66.2
9.0
42.1
63.3
7.6
5.8
4.1
6.7
Cd
<0.18
<0.18
0.29
0.34
•9)
0.52
0.52
0.43
8.53
0.16
(.15-. 19)
4.51
1.01
6.11
0.069
14.1
0.44
Cr
2.85
1.22
3.52
2.98
4.34
6.37
3.80
334.0
9.36
(9.2-9.5)
83.0
34.0
334.0
3.1
597.0
14.6
Cu
4.5
3.00
4.51
4.88
6.01
9.39
10.5
122.0
9.56
(9.2-10)
35.0
16.0
61.0
12.0
107.0
17.0
Ha
0.0004
0.0004
0.001
0.001
0.0009
0.002
0.001
0.01
0.002
(.0021-
0.003
0.005
0.005
0.003
0.006
0.005
(ua/Ll
Ni
<4.3
4.9
<4.3
5.7
(4.9-7.4)
<4.3
6.1
4.9
16.0
<4.3
.0024)
13.5
6.1
95.8
<4.3
209.0
8.6
Pb
9.7
5.5
9.7
8.2
(7.6-8.9)
18.6
25.4
32.2
166.0
12.3
52.9
35.5
49.9
11.4
133.0
17.3
Zn
9.4
<7.2
31.0
31.0
53.0
127.0
148.0
148.0
9.4
75.0
39.0
16.0
9.4
302.0
24.0
-------
Table 13. Concentration of metals in porewater from sediment samples collected during the third survey of Saginaw River.
The ranges for replicates are shown in parentheses.
Metal concentration (ua/L)
Sample
SR-03-01
SR-03-02
SR-03-06
SR-03-08
Aa
<0.46
<0.46
<0.46
<0.46
As
1.42
0.84
1.60
3.23
(2.3-3.9)
Cd
<0.17
. <0.17
<0.17
<0.17
Cr
0.71
0.88
1.9
2.66
(2.5-2.8)
Cu
2.7
1.1
3.0
3.56
(3.0-4.3)
Ha
0.002
0.0031
0.003
0.0045
(0.003-0.
Ni
3.15
3.15
4.90
4.90
006)
Pb
3.5
3.5
3.5
4.86
(4.1-6.4)
Zn
1.35
2.44
10.7
2.88
(2.3-3.3)
-------
Table 14. Acute toxicity of elutriates prepared from stored sediments from Indiana Harbor, Buffalo River, and Saginaw Riv
to Photobacter ium phosphoreum in Microtox"
percent dilutions of full-strength elutriate samples. The
stored up to 5 d; E2's from samples stored 14-19 d; and E3 '
Sample
number
Elutriate Days of Percent samp!
number storaae dilution tes
El elutriates were prepared from sedimen
a from samples stored 17-26 d.
le MicrotoxR ECSO's
sted 5 min
and 95% confidence intervals
15 min
Indiana Harbor
IH-01-03
IH-01-03
IH-01-03
IH-01-04
IH-01-04
IH-01-04
IH-01-06
IH-01-06
IH-01-06
IH-01-07
IH-01-07
IH-01-07
Buffalo River
BR-01-01
BR-01-01
BR-01-03
BR-01-03
BR-01-07
BR-01-07
BR-01-08
BR-01-08
BR-01-09
BR-01-09
Saoinaw River
SR-01-03
SR-01-03
SR-01-03
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
El
E2
El
E2
El
E2
El
E2
(first
El
E2
E3
4
19
26
4
19
26
5
19
26
5
19
26
1
15
1
15
2
15
3
15
3
15
survey)
1^
142
223
45.0
45.0
45.0
45.0
45.0
45.0
45.0
45.0
45.0
45.0
45.0
11.2
45.0
45.0
45.
100.
45.
100.
100.
100.
100.
100.
100.
100.
100.
0
0
0
0
0
0
0
0
0
0
0
33.6
31.0
32.7
58.7
61.8
51.5
9.0
10.1
9.0
37.2
1.3
3.8
33.4
42.0
40.8
>100
55.5
70.7
>100
>100
>100
>100
>100
>100
>100
(29.0
(24.5
(28.0
(27.6
(22.8
(10.4
(8.3
(9.4
(8.7
(27.3
(0.7
(3.5
(23.6
(32.6
(38.5
(NC)
(41.7
(48.7
(NC)
(NC)
(ND)
(ND)
(ND)
(ND)
(ND)
- 39.
- 39.
- 38.
- 124
- 167
- 254
- 9.8)
-10.7)
0)
1)
4)
.8)
•9)
•7)
- 9.2)
- 50.8)
- 2.5)
- 4.3)
- 47.
- 54.
- 43.
- 73
- 102
4)
0)
3)
.8)
•5)
20.3
14.3
18.8
31.5
34.1
33.5
4.3
5.0
4.7
34.4
0.6
3.3
40.5
49.6
44.2
>45
91.8
>100
>100
>100
>100
>100
>100
>100
(18.7 -
(13.7 -
(17.8 -
(27.1 -
(28.0 -
(25.0 _
(3.2 -
(4.3 -
(3.7 -
(23.6 -
(0.2 -
(2.8 -
(25.4 -
(44.0 -
(32.8 -
>100
-------
Sample
number
SR-01-06
SR-01-10
SR-01-10
SR-01-10
Saainaw River.
SR-03-01
SR-03-03
SR-03-03
SR-03-02
SR-03-02
SR-03-02
SR-03-05
SR-03-05
SR-03-05
SR-03-06
SR-03-06
SR-03-06
SR-03-08
SR-03-08
SR-03-08
SR-03-16
SR-03-16
SR-03-16
SR-03-24
SR-03-24
SR-03-24
Elutriate Days of
number storage
E3
El
E2
E3
(third
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
E3
22
2
14
22
survey 1
1
14
17
1
14
17
1
14
17
2
14
17
2
14
21
2
14
21
2
14
21
Initial teal
concentratic
45.
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
100
0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
.0
t Microtox" ECSO's and 95% confidence intervals
3ns4 5
20.7
>100
100
>100
>100
92.2
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
min
(20.0 - 21.4)
(ND)
(ND)
(ND)
(NC)
(61. - 14.)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
15 mi
12.0
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
95.0
92.3
>100
>100
>100
>100
.n
(6.0 - 24.0)
(ND)
(ND)
(ND)
(NC)
(NC)
(NC)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(ND)
(37.2 - 242.9)
(63.1 - 135.1)
(ND)
(ND)
(ND)
(ND)
-------
1329 126
_J
TJ>
W
c
jo
tp
5
+•«
0)
O
c
O
O
75
4->
O
O
.a
(Q
C
CO
TJ
C
Flor
1284
11757 2449
300
200
100
69 14.3
Acute
WQC
Chronic
WQC
*******
0.1
Acute
WQC
Hg
0.67
Acute
WQC
Chronic
WQC
Flor 03 04 05 06 07 08 10
*******
• Acute
WQC
Chronic
WQC
Ftor
Ftor 03
04
05
06
07
08
10
* ******
*******
Station
Figure 2.1 Concentrations of select metals (total) in elutriates and pore waters prepared from
Indiana Harbor whole sediment samples. Asterisks indicate samples that elicited effects
in the daphnid or Microtox" test and the horizontal lines show the National Acute or Chronic
Water Quality Criteria, adjusted to 140 mg/L hardness for Cu, Cd, and Pb (EPA 1986).
Toxicity tests were not conducted with porewater samples.
Elutriates
Porewaters
-------
z
0.3
0.25
O)
O 0.2
E
"§0.15
JN
C
O
C
=> 0.1
0.05
0
Stations
0.37 O.S9 1.37
I
ft
sf- -:-
!/*
1.75
Flor03 04 05 06 07 08 10
Grand Calumet River
01020304050607080910 Flor 020304060709 10 01020506081624
Buffalo River Saginaw River (01) Saginaw River (03)
Figure 2.2 Concentrations of unionized ammonia in elutriates prepared from Indiana Harbor, Buffalo
River, and Saginaw River whole sediment samples. The horizontal line shows the National
Acute Water Quality Criteria for unionized ammonia (EPA 1986).
Acute WQC
-------
Buffalo River Metal Concentrations (ug/L)
-------
O)
CO
c
O
c
0)
O
c
O
O
a
-------
_J 20
7»
—' 15
0)
0 10
O
g
O
Q)
CO
o
>,
0)
3
CO
o3
if
03
I
CO
Acute
WQC
Cu
Chronic
WQC
114
z
20
10
Acute
"WQC
Pb
24
2.4
0.002 -
0.001
Station
Elutriates
Porewaters
01 02 05 06 08 16 24
Acute
WQC
02 05 06 08
16 24
Figure 2.5 Concentrations of select metals (total) in elutriates and pore waters prepared from Saginaw
River (third survey) whole sediment samples. Asterisks indicate samples that elicited effects
in the daphnid or MicrotoxR test and the horizontal lines show the National Acute or Chronic
Water Quality Criteria, adjusted to 140 mg/L hardness for Cu, Cd, and Pb (EPA 1986).
Toxicity tests were not conducted with porewater samples.
-------
Table 2.1 Sample numbers and sample station locations for sediments collected from three Great Lakes
Areas of Concern (AOC) and tested at the National Fisheries Contaminant Research Center
(NFCRC).
AOC
and sample
number
Water depth
(m)
Core depth
(m)
Location of
samolina station
Latitude
Lonaitude
Indiana Harbor. IN
IH-01-03*
IH-01-04*
IH-01-05
1H-01-06*
IH-01-07*
IH-01-08
IH-01-10
Buffalo River. NY
BR-01-01*
BR-01-02
BR-01-03*
BR-01-04
BR-01-05
BR-01-06
BR-01-07*
BR-01-08*
BR-01-09*
BR-01-10
Saainaw River, MI
SR-01-02
SR-01-03*
SR-01-04
SR-01-06*
SR-01-07
SR-01-09
SR-01-10*
ND2
ND
ND
ND
ND
ND
ND
5.5
4.6
6.1
ND
6.7
6.1
6.1
ND
0.9
ND
(first survey)
ND
0.9
ND
0.6
ND
2.7
ND
NA3
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
NA
41°
41°
41°
41°
41°
41°
41°
42°
42°
42°
42°
42°
42°
42°
42°
42°
42°
43°
43°
43°
43°
43°
43°
43°
40.42'
40.07'
39.65'
39.30'
38.75'
38.81'
38.38'
51.42'
52.18'
52.66'
51.97'
51.71'
51.79'
51.63'
51.69'
51.68'
51.94'
37.42'
37.25'
36.84'
36.74'
36.46'
33.76'
32.66'
87°
87°
87°
87°
87°
87°
87°
78°
78°
78°
78°
78°
78°
78°
78°
78°
78°
83°
83°
83°
83°
83°
83°
83°
26.35
26.17
27.09
27.58
28.34
28.84
28.28
52.04
52.28
53.06
52.14
52.04
51.25
50.93
50.73
50.02
49.05
50.53
50.58
51.40
52.16
53.17
54.57
53.05
-------
Table 2.1 continued
AOC
and sample
number
Saainaw River, MI
Grab samples
SR-03-01*
SR-03-02*
SR-03-05*
SR-03-06*
SR-03-08*
SR-03-16*
SR-03-24*
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Water depth Core depth
(rtO (m)
(third survey)
6.4
2.0
1.8
0.7
0.9
6.7
0.8
2.0
2.0
1.8
1.8
0.7
0.7
0.7
0.7
NA
NA
NA
NA
NA
NA
NA
0.2-0.5
0.5-0.9
0.0-0.6
0.6-1.2
0.0-0.6
0.6-1.2
0.1-0.3
0.3-0.8
Location of
samplina station
Latitude
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
43°
45.30'
37.42'
36.72'
36.74'
35.33'
36.70'
34.55'
37.42'
37.42'
36.72'
36.72'
36.74'
36.74'
36.74'
36.74'
Lonqitude
83°
83°
83°
83°
83°
83°
83°
83°
83°
83°
83°
83°
83°
83°
83°
47.48
50.53
51.67
52.16
53.88
51.80
54.16
50.53
50.53
51.67
51.67
52.16
52.16
52.16
52.16
1 The Large Lakes Research Station Number includes the site code, survey number, transect number, and
the sample fraction number (core samples only).
2 ND = Not determined
3 NA = Not applicable
* Indicate samples tested in whole sediment tests
-------
Table 2.2 Dissolved Oxygen (DO), pH, and survival of Daphnia maana during a 48-h
reference toxicant test with Instant Ocean" salinity. The 48-h EC50 was :
with 95% confidence limits of 21.6 to 13.0 0/00 salinity for the survival
shown .
Instant ocean"
salinity fO/001
Control
(well water)
33.0
19.8
11.8
7.1
4.2
2.5
1.5
DO (ma/D pH
0-h 48-h 0-h 48-h
7.9 8.2 8.4 8.3
7.1 8.0 8.0 8.1
ND ND ND ND
ND ND ND ND
7.5 8.1 8.2 8.2
ND ND ND ND
ND ND ND ND
7.9 8.0 8.3 8.3
Survival (%l
24-h 48-h
100
0.0
0.0
0.0
100
100
100
100
100
.0 80.0
.0 100.0
.0 100.0
.0 100.0
ND = Not determined
-------
Table 2.3 Measured water gualtiy for elutriate samples prepared from sediments collected from each area of concern
(AOC) and the mean with standard error (in parentheses) across stations for each AOC.
AOC and
sample
Indiana Harbor
Control
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Means
Buffalo River
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Means
Saainaw River
Control
SR-01-02
SR-01-03
SR-01-04
SR-01-06
PH
6.98
7.64
7.78
7.89
7.26
7.51
7.16
7.42
7.5
(0.11)
9.17
7.98
7.08
7.01
7.38
7.24
7.20
7.19
7.05
7.21
7.45
(0.21)
Alkalinity,
(mg/L as
CaCO^)
44.00
120.00
160.00
112.00
160.00
146.00
160.00
400.00
163
(36.62)
108.00
80.00
82.00
116.00
102.00
112.00
116.00
146.00
134.00
106.00
110.20
(6.43)
Hardness,
(mg/L as
CaCOj)
118.00
155.00
185.00
150.00
168.00
115.00
160.00
183.00
154
(9.32)
160.00
160.00
150.00
160.00
160.00
150.00
150.00
180.00
160.00
160.00
159.00
(2.77)
D.O. ,
fma/L)
7.90
3.50
4.00
4.00
3.6
3.30
1.80
3.20
3.91
(0.62)
4.40
8.20
4.90
3.40
4.00
3.20
3.00
3.00
3.00
3.20
4.03
(0.51)
Conductivity ,
(umhos/cm)
259.46
335.14
432.43
362.16
486.49
383.78
443.24
951.35
456.76
(74.91)
464.86
400.00
347.03
367.57
335.14
367.57
378.38
410.81
421.62
378.38
387.14
(12.10)
Unionized
ammonia,
fma/L)
ND
0.12
0.09
0.24
0.27
0.37
0.09
0.59
0.25
(0.07)
1.37
0.03
0.02
0.02
0.02
0.02
0.01
0.02
0.02
0.01
0.15
(0.14)
Chloride,
-------
Table 2.3 continued.
AOC and
sample
Saginaw River
SR-01-07
SR-01-09
SR-01-10
Mean
Saqinaw River
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Mean
PH
Alkalinity,
(mg/L as
CaCCM
( first survey >
7.52
7.54
7.71
7.48
(0.07)
164.00
146.00
106.00
138.75
(17.59)
Hardness,
(mg/L as
CaCO})
196.00
192.00
176.00
183.50
(16.08)
D.O. ,
-------
Table 2.4 Mean and standard error of the mean (in parentheses) water qualtiy
during 48-h acute toxicity tests with Daphnia maana exposed to
elutriates prepared from sediments collected from each Area of
Concern (AOC).
AOC and
sample
DH
Indiana Harbor
Control
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Buffalo River
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Saainaw River
Control
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
8.
(first
7.
8.
8.
8.
7.
8.
8.
8.
18
26
31
31
33
11
30
26
64
39
24
31
32
14
23
29
17
25
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
.05)
.01)
.06)
.01)
.05)
.08)
.02)
.03)
.25)
.0)
.17)
.12)
.13)
.14)
.12)
.10)
.16)
.13)
D.O.,
(ma/Li
8.
8.
8.
8.
8.
8.
8.
8.
9.
9.
9.
9.
9.
9.
9.
9.
9.
9.
45
20
35
13
18
05
50
45
33
85
90
70
70
10
63
60
43
50
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
03)
19)
09)
19)
13)
41)
07)
17)
12)
25)
37)
24)
17)
11)
22)
23)
17)
19)
Conductivity,
(ianhos/cm\
276
330
369
332
374
363
391
567
356
313
329
352
332
353
324
338
334
320
.75
.81
.19
.16
.87
.52
.08
.57
.76
.51
.73
.70
.43
.01
.32
.74
.23
.00
(13.25)
(13.26)
(29.52)
(14.56)
(29.40)
(18.31)
(23.71)
(113.96)
(36.66)
(34.47)
(41.05)
(31.78)
(31.94)
(23.82)
(5.58)
(17.30)
(16.61)
(7.23)
survey)
98
27
36
34
91
24
17
15
(0
(0
.25)
.22)
(0.24)
(0
(0
(0
(0
(0
.19)
.22)
.17)
.23)
.21)
9.
9.
9.
9.
9.
9.
9.
9.
05
08
10
17
05
43
30
48
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
40)
19)
28)
35)
29)
49)
45)
51)
294
403
481
400
339
356
372
334
.87
.51
.62
.27
.19
.21
.16
.05
(19.02)
(33.23)
(76.94)
(45.39)
(20.39)
(62.33)
(29.76)
(11.09)
-------
Table 2.4 continued.
AOC and
sample
PH
D.O.,
(ma/L)
Conductivity,
(umhos/cm)
Saainaw River (third survey)
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
8
8
8
8
8
8
8
8
8
8
8
8
8
7
8
.11
.08
.08
.11
.08
.06
.06
.00
.14
.11
.08
.01
.16
.87
.11
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
.09)
.11)
.12)
.11)
•14)
.14)
.15)
.11)
.08)
.11)
.14)
.07)
.07)
.16)
.07)
8.22
8.22
8.14
8.10
8.24
8.16
8.26
7.95
9.33
8.90
8.58
8.44
9.08
7.30
8.83
(0
(0
(0
(0
(0
(0
(0
(1
(0
(0
(0
(0
(1
(0
(0
.55)
.52)
.57)
.48)
.62)
.61)
.59)
.02)
.87)
.91)
.72)
.86)
.04)
.96)
.99)
321.
355.
352.
327.
343.
352.
386.
420.
377.
340.
409.
341.
380.
450.
400.
30
68
43
57
79
43
60
54
57
54
46
19
54
00
54
(9.
(22
(20
(13
(20
(22
(35
(60
(35
(38
(45
(31
(31
(57
(28
16)
.31)
.33)
.63)
.41)
.09)
.91)
.35)
.63)
.27)
.78)
.00)
.34)
.39)
.06)
-------
Table 2.5. Concentrations of total metals in elutriates prepared from Indiana Harbor sediment samples.
Metal concentration (jtg/L)
Sample
Control
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Ag As
<0.03 <1.8
<0.03 15
<0.03 5.5
0.21 9.8
0.26 10.2
1.3 152
<0.03 24.8
0.45 13.0
Cd
0.05
3.3
1
2.9
2.1
69
5.8
2.6
Cr
1.1
61
37
73
86
4700
196
73
Cu
2.6
65
28
42
36
1329
70
23
Hg
0.0011
0.0383
0.0233
0.0345
0.0772
0.6670
0.1203
0.0046
Ni
2.5
23
8.8
18
15
650
46
8.8
Pb
12
489
293.9
391.9
343.9
11756
2449
637
Zn
6.9
517
183
535
348
10870
814
248
-------
Table 2.6. Concentration of totals metals in Indiana Harbor sediment porewater samples. The ranges for duplicate samples
are in parentheses.
Metal concentration (ua/L)
Sample
IH-01-03
IH-01-04
IH-01-06
IH-01-07
Ad
<0.05
<0.05
<0.05
-------
Table 2.7 Acute toxicity of Indiana Harbor, Buffalo River, and Saginaw River elutriates
to Daphnia tnaana and Photobacterxum phosphoreum(Microtox1*). The 48-h EC50"s
for daphnids and 5 and 15 min ECSO's for MicrotoxR (95% confidence intervals
in parentheses) are percent dilutions of full-strength elutriate samples.
Survival of daphnids in dilution water controls ranged from 80 to 100%.
EC50 and 95% confidence interval
Sample
Indiana Harbor
Control
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Buffalo River
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Saainaw River
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
Daphnia tnaana
(48-h)
>100
1.0 (0.008 - 9.6)
70.7 (25 - 100)
25.0 (18 - 100)
>100
0.7 (0.001 - 3.1)
5.4 (2.5 - 11.0)
39.4 (30.2 - 53.0)
>100
>100
76.2 (50 - 100)
>100
>100
0.11 (0.001 - 12.5)
ND
<100
>100
>100
(first survey)
>100
>100
>100
>12.5 - 100
>12.5 - 100
>100
>100
Microtox"
(5-min)
>100
33.6 (29.
58.7 (27.
48.4 (38.
9.0 (8.3
37.2 (27.
12.1 (11.
12.9 (12.
33.4 (23.
>100a
40.8 (38.
>100a
63.5 (37.
19.3 (17.
55.5 (41.
>100a
>100a
>100a
>100a
>100a
>100a
14.1 (12.
>100a
>100a
>100a
0 - 39.0)
6 - 124.8)
2 - 61.2)
- 9.8)
3 - 50.8)
5 - 12.7)
2 - 13.7)
6 - 47.4)
5 - 43.3)
3 - 108.3)
1 - 21.8)
7 - 73.8)
0 - 16.6)
Microtox"
(15-min)
>100
20.3 (18.7
31.5 (27.1
27.3 (25.4
4.3 (3.2 -
34.4 (23.6
6.0 (5.4 -
6.8 (5.8 -
40.5 (25.4
>100a
44.2 (32.8
>100a
65.8 (52.5
6.7 (6.0 -
46 -100
>100a
>100a
>100a
>100a
>100a
>100a
13.0 (9.5
>100a
>100a
>100a
- 21.9)
-36.6)
- 29.3)
5.8)
- 50.2)
6.7)
7.9)
- 64.4)
- 59.5)
- 82.4)
7.5)
-17.7)
-------
Table 2.7 continued
EC50 and 95% confidence interval
Sample
Saainaw River
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05 02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Daphnia
f48-h>
(third survey >
>100
>100
>100
>100
>100
>100
>100
MicrotoxR
(5-min»
>100a
>100a
>100a
>100a
>100a
>100a
>100a
43.1 (35.0 - 53.0)a
>100a
>100a
88.1 (55.6 - 139. 5)a
>100a
76.5 (44.7 - 131. 0)a
30.5 (19.8 - 47.0)a
71.3 (54.3 - 93.5)a
MicrotoxR
(15-min)
>100a
>1008
>100a
>100a
>100a
>100a
>100a
34.0 (26.7 - 43.4)a
>100a
>100a
>100a
53.7 (42.3 - 68.3)a
>100a
31.1 (22.4 - 43.1)a
95.7 (64.0 - 143. l)a
ND = Not determined
a 100% elutriates testesd
-------
Table 2.8 Concentrations of total metals in elutriates prepared from Buffalo River sediment samples,
Metal concentration (/tg/L)
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Ag
<0.03
<0.03
0.33
<0.03
0.12
0.04
0.04
<0.03
<0.03
<0.03
As
27
0.7
15
8
8.4
17
12
12
9.8
2.6
Cd
1.5
<0.15
2.2
0.83
2.9
2
1.2
0.61
0.44
<0.15
Cr
22
1.4
44
10
24
31
10
2.3
1.3
0.9
Cu
14
3.2
34
16
30
60
17
11
7.4
4.8
Hg
0.005
0.004
0.01
0.004
0.009
0.10
0.007
0.006
0.003
0.002
Ni
6.6
<5
5.5
<5
7.7
11
9.9
<5
<5
<5
Pb
45
8.5
115
40
156
238
62
25
23
13
Se
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
<0.25
Zn
122
22
150
60
161
306
67
28
28
17
-------
Table 2.9 Concentrations of total metals in Buffalo River sediment porewater samples. The ranges for duplicate samples
are in parentheses.
Metal concentration (/xg/L)
Sample
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Ag
<0.05
<0.05
<0.052
<0.05
<0.052
As
14.5
(1.2-
2.3)
1.7
<1.0
1.7
Cd
0.1
<0.07
<0.07
<0.07
<0.07
Cr
2.5
0.7
0.3
0.2
0.27
Cu
3.3
(1.3-
2.6)
1.65
1.65
2.3
Hg
0.1
(+0.003)
(0.0098-
0.0096)
0.004
0.002
0.002
Ni
<4.0
<4.0
<4.0
<4.0
<4.0
Pb
5.7
(1.6-
2.0)
0.8
<0.7
<0.7
Zn
0.9
<0.93
<0.93
<0.93
<0.93
-------
Table 2.10. Concentrations of total metals in elutriates from sediment samples collected during the first survey of
Saginaw River.
Metal concentration ((Jig/L)
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
Ag
<0.09
<0.09
<0.09
<0.09
<0.09
<0.09
<0.09
As
5.9
8.6
5.6
4.6
3.4
5.3
2.2
Cd
0.17
0.11
0.35
7.4
0.24
0.11
<0.06
Cr
3.1
3.0
4.8
253.0
2.6
1.2
1.2
Cu
7.6
5.6
10.4
98.0
7.2
3.6
<1.2
Hg
0.0057
1.00
0.003
0.005
0.002
0.002
0.008
Ni
<5.0
<5.0
<5.0
124.0
<5.0
<5.0
<5.0
Pb
11.2
11.8
15.1
39.2
11.2
5.0
3.4
Zn
28.0
28.0
50.0
139.0
33.0
28.0
17.0
-------
Table 2.11. Concentrations of total metals in porewater from sediments collected during the first survey of Saginaw River.
Metal concentration (jtg/L)
Sample
SR-01-03
SR-01-06
SR-01-10
Ag As
<0.05 <1.0
<0.05 <1.0
<0.05 1.7
Cd
<0.07
0.84
<0.066
Cr
0.8
24.4
0.33
Cu
<0.57
19.8
2.6
Hg
0.0034
0.006
0.002
Ni
<4.0
97.8
<4.0
Pb
<0.7
36.2
0.81
Zn
<0.93
19.0
<0.93
-------
Table 2.12 Concentrations of total metals in elutriates prepared from sediments collected during the third survey of
Saginaw River. The ranges for duplicates are in parentheses.
Metal concentration (/*g/L)
Sample
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Ag
<0.085
<0.085
<0.085
0.096
<0.085
0.12
0.12
1.09
(0.15-
0.24)
0.54
0.29
0.73
0.07
1.36
0.13
As
1.7
2.6
2.6
(2.6-2.9)
2.3
3.2
5.2
66.2
9.0
42.1
63.3
7.6
5.8
4.1
6.7
Cd
<0.18
<0.18
0.29
0.34
0.52
0.52
0.43
8.53
(0.16-
(0.19)
4.51
1.01
6.11
0.069
14.1
0.44
Cr
2.85
1.22
3.52
2.98
4.34
6.37
3.80
334.0
(9.2-
9.5)
83.0
34.0
334.0
3.1
597.0
14.6
Cu
4.5
3.00
4.51
4.88
6.01
9.39
10.5
122.0
(9.2-
10.0)
35.0
16.0
61.0
12.0
107.0
17.0
Hg
0.0004
0.0004
0.001
0.001
0.0009
0.002
0.001
0.01
(0.0021-
0.0024)
0.003
0.005
0.005
0.003
0.006
0.005
Ni
<4.3
4.9
<4.3
(4.9-7.4)
<4.3
6.1
4.9
16.0
<4.3
13.5
6.1
95.8
<4.3
209.0
8.6
Pb
9.7
5.5
9.7
(7.6-8.9)
18.6
25.4
32.2
166.0
12.3
52.9
35.5
49.9
11.4
133.0
17.3
Zn
9.4
<7.2
31.0
31.0
53.0
127.0
148.0
148.0
9.4
75.0
39.0
16.0
9.4
302.0
24.0
-------
Table 2.13 Concentrations of total metals in porewater from sediment samples collected during the third survey of Saginaw
River. The ranges for duplicate samples are in parentheses.
Metal concentration (/tg/L)
Sample
SR-03-01
SR-03-02
SR-03-06
SR-03-08
Ag
<0.5
<0.5
<0.5
<0. 5
As
1.4
0.8
1.6
(2.3-
3.9)
Cd
<0.2
<0.2
<0.2
<0.2
Cr
0.7
0.9
1.9
(2.5-
2.8)
Cu
2.7
1.1
3.0
(3.0-
4.3)
Hg
0.002
0.0031
0.003
(0.003-
0.006)
Ni
3.15
3.15
4.90
4.90
Pb
3.5
3.5
3.5
(4.1-
6.4)
Zn
1.3
2.4
10.7
(2.3-
3.3)
-------
Table 2.14 Acute toxicity of elutriates prepared from stored sediments from Indiana Harbor, Buffalo River, and Saginaw
River to Photobacterium phosphoreum in MicrotoxR tests. The 5- and 15-min ECBO's (95% confidence intervals
in parentheses) are percent dilutions of full-strength elutriate samples. The El elv
from whole sediment samples that were stored up to 7 d; E2's from whole sediment samj
and E3's from whole sediment samples stored 21 to 28 d. "Days Storage" is the number
collection at AOC and day of MicrotoxR test.
Sample
Elutriate Days of Percent samp!
number storage dilution tet
Le Microtox" ECSO's and 95% confidence intervals
3ted 5-min
15-min
Indiana Harbor
IH-01-03
IH-01-03
IH-01-03
IH-01-04
IH-01-04
IH-01-04
IH-01-06
IH-01-06
IH-01-06
IH-01-07
IH-01-07
IH-01-07
Buffalo River
BR-01-01
BR-01-01
BR-01-03
BR-01-03
BR-01-07
BR-01-07
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
El
E2
El
E2
6
21
28
6
21
28
7
21
28
7
21
28
5
20
5
20
7
20
45.
45.
45.
45.
45.
45.
45.
45.
45.
45.
45.
11.
45.
45.
45
100
45
100
0
0
0
0
0
0
0
0
0
0
0
2
0
0
.0
.0
.0
.0
33
31
32
58
61
51
9
10
9
37
1
3
33
42
40
.6
.0
.7
.7
.8
.5
.0
.1
.0
.2
.3
.8
.4
.0
.8
>100
55
70
.5
.7
(29.0 -
(24.5 -
(28.0 -
(27.6 -
(22.8 -
(10.4 -
(8.3 - 9
(9.4 -10
(8.7 -
(27.3 -
(0.7 - 2
(3.5 - 4
(23.6 -
(32.6 -
(38.5 -
(NC)
(41.7 -
(48.7 -
39.0)
39.1)
38.4)
124.8)
167.9)
254.7)
.8)
•7)
9.2)
50.8)
•5)
•3)
47.4)
54.0)
43.3)
73.8)
102.5)
20.3
14.3
18.8
31.5
34.1
33.5
4.3
5.0
4.7
34.4
0.6
3.3
40.5
49.6
44.2
>45
91.8
(18.7 -
(13.7 -
(17.8 -
(27.1 -
(28.0 -
(25.0
(3.2 -
(4.3 -
(3.7 -
(23.6 -
(0.2 -
(2.8 -
(25.4 -
(44.0 -
(32.8 -
>100
(NC)
(28.3 -
21.9)
15.0)
20.0)
36.6)
41.6)
45.0)
5.8)
5.6)
6.0)
50.2)
1.5)
3.8)
64.4)
55.8)
59.5)
(NC)
298.0)
-------
Sample
Buffalo River
BR-01-08
BR-01-08
BR-01-09
BR-01-09
Saqinaw River
SR-01-03
SR-01-03
SR-01-03
SR-01-06
SR-01-06
SR-01-06
SR-01-10
SR-01-10
SR-01-10
Saqinaw River,
SR-03-01
SR-03-03
SR-03-03
SR-03-02
SR-03-02
SR-03-02
SR-03-05
SR-03-05
SR-03-05
SR-03-06
SR-03-06
SR-03-06
Elutriate Days of
number storage
El
E2
El
E2
( first
El
E2
E3
El
E2
E3
El
E2
£3
(third
El
E2
E3
El
E2
E3
El
E2
E3
El
E2
E3
8
20
8
20
survey)
5
18
26
7
18
26
6
18
26
survey )
5
18
21
5
18
21
5
18
21
6
18
21
Percent sample
dilution tested
100.0
100.0
100.0
100.0
100.0
100.0
100.0
45.0
45.0
45.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
Microtox" ECBO's and
5-min
>100
>100
>100
>100
>100
>100
>100
14.1 (12.0 - 16.6)
17.8 (16.5 - 19.2)
20.7 (20.0 - 21.4)
>100
100
>100
>100
92.2 (61. - 14.)
11.9 (34.6 - 355.1)
>100
>100
>100
>100
>100
>100
>100
>100
>100
95% confidence
1 5-min
>100
>100
>100
>100
>100
>100
>100
13.0 (9.5
14.8 (12.9
12.0 (6.0
>100
>100
>100
>100
>100
>100)
>100
>100
>100
>100
>100
>100
>100
>100
>100
intervals
-17.7)
- 17.0)
- 24.0)
-------
Table 2.14 continued.
Sample
Saginaw River.
SR-03-08
SR-03-08
SR-03-08
SR-03-16
SR-03-16
SR-03-16
SR-03-24
SR-03-24
SR-03-24
Elutriate
number
Days of
storage
Percent sample
dilution tested
Microtox" ECSO's
5-min
and 95% confidence
1 5-min
intervals
(third survey)
El
E2
E3
El
E2
E3
El
E2
E3
6
18
25
6
18
25
6
18
25
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
100.0
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
>100
92.3 (63.1
>100
>100
>100
>100
- 135.1)
-------
CHAPTER 3; ELUTRIATE TOXICITY TESTS; SELENASTRUM CAPRICORNUTUM
Hall, N.E., T.W. La Point, P.R. Heine and J.F. Fairchild
INTRODUCTION
Algae serve a critical functional role in aquatic ecosystems. They
convert solar energy and carbon dioxide to organic matter or algal biomass.
Algal biomass serves as food for herbivorous and omnivorous animals, from
Daphnia to paddlefish. Thus, algae are a primary base for the transfer of
energy to consumer levels of trophic food webs. Algae also serve numerous
other critical ecological functions. They cycle chemical elements through the
ecosystem by incorporating and transferring dissolved nutrients, vitamins,
metals, and other elements to other ecosystem components. If not consumed,
algae die and are decomposed by bacteria, thereby transforming, solubilizing
and recycling chemicals to be used again as nutrients by algae and other
organisms. The flow of energy and materials through the ecosystem is affected
by phytoplankton population dynamics, which are in turn influenced by
temperature, solar radiation, nutrient concentration and animal grazing.
Anthropogenic disturbance, such as the introduction of toxicants, can alter
the structure and function of the phytoplankton community (Amblard et al.
1990, Bokn 1990, Havens and Heath 1990, Molander et al. 1990, Rai et al.
1990). Such changes can ul :imately affect the entire aquatic ecosyttem.
Algae were used in the present study to evaluate the toxicity of the
Great Lakes sediments, through testing of elutriates of sediment samples.
Elutriates and their use in sediment testing are discussed in Chapter 2.
Currently, there is no method for exposing algae directly to whole sediments,
an elutriate most closely simulates the most likely exposure conditions for
natural algal populations. A single-species laboratory toxicity test was
selected that was rapid and had small volume requirements. The species
selected for testing was Selenastrum capricornutum, recommended in standard
methods (EPA 1985, IJC 1988, Clesceri et al. 1989, ASTM 1990). The method is
derived from the U.S. Environmental Protection Agency's standard 96-h algal
assay procedure (AAP) (Miller et al. 1978). The AAP was originally designed
to measure eutrophication. However, it has been recommended for use in
testing the toxicity of complex effluents (Porcella 1983) and has been widely
used to test single chemicals (Elnabarawy and Welter 1984, Whitton 1984,
Greene et al. 1988). The AAP has a 96-h duration, cell-counts are the test
endpoint and flasks are the test vessels; thus labor, time, space and
equipment requirements may be prohibitive for screening large numbers of
samples. The AAP has been converted into a 24-h photosynthesis test with
smaller sample volume requirements (Ross et al. 1988, Sloterdijk et al. 1989).
Algal cells are exposed to an elutriate supplemented with nutrients and
radioactive inorganic carbon. Toxicity is measured in terms of relative
radioactive carbon uptake compared to controls. Our tests are modifications
of their 24-h radioactive carbon assimilation (photosynthesis) tests.
The objective of this chapter is to identify AOC samples from the Great
Lakes toxic to algae. Toxicity is defined for the algal assay as negatively
affecting carbon assimilation of algal test subpopulations. The short
generational time and meager volume requirements of a microscopic algae allows
testing of subcultures comprised of thousands of individuals and production of
one or more generations within a 24-h period. Algal test responses are
population responses incorporating effects upon individual cells' metabolism,
growth, mortality and reproduction. It is assumed that if elutriate has a
toxic effect on assay algal populations, natural algal communities in contact
with sediment at the sampling site may also experience a negative effect.
Effects may range from slower growth rates to lethality. Either may have
profound effects on the ability of a species to compete for resources and
could result in changes in algal communities, balance with bacterial
competitors, phytoplankton consumers, and creating a ripple of effect
throughout the ecosystem.
3-1
-------
MATERIALS AND METHODS
METHOD DEVELOPMENT
The original objective of our study was to evaluate the toxicity of
elutriate samples prepared from all of the sediment samples listed in table
2.1. Initially, elutriates from all of the sediments except the third survey
of Saginaw River samples were tested using a method similar to that described
by Ross et al. (1988).
Elutriates were filtered through 0.7 pm Whatman GF-F glass fiber filters
to remove indigenous algae. Exposure volume was 11.2 mL, placed within a 16-
mL capped borosilicate glass tube. Nutrients were added in 1 mL of medium
concentrate to produce the same nutrient concentration as the culture medium
for the control treatment (Table 3.1). Algal exposure occurred at a density
of 190,000 cells/mL, for 24 h at 21 + 2° C with 400 to 500 ftc (approximately
40 to 50 /jE/m2-sec) of light. At 20 h, sodium MC-bicarbonate was added to
produce a concentration of 1 to 1.15 /jCi/mL. At 24 h, algal Mcarbon
assimilation was halted by adding HC1 and air bubbling for 5 min to remove
unassimilated 14carbon. The amount of radioactive carbon assimilated was
measured in a beta liquid scintillation counter.
Results from these exposures indicated the test required modification to
determine elutriate toxicity. The high nutrient concentrations likely to
occur in the elutriates violated two assumptions of the algal test design:
equal concentrations among treatments of (1) non-radioactive inorganic carbon
(Kusk and Nyholm 1991) and (2) nutrients limiting growth.
Non-radioactive inorganic carbon concentrations of elutriates (estimated
from pH and alkalinity), were about 10 times higher than the concentration in
the algal medium. Therefore radioactive inorganic carbon was diluted with
non-radioactive inorganic carbon e.g., radioactive carbon assimilation
decreased with increased elutriate concentration. The method was revised to
include measurement of non-radioactive inorganic carbon in test solutions when
radioactive carbon was added, at 20 h. A carbon dioxide ion selective
electrode (Fisher model 13-620-506/507) was used to determine total inorganic
non-radioactive carbon upon an acidified 20-tnL sub-sample from each test
replicate. Test volume was increased to 24.4 mL and 25-mL capped borosilicate
glass tubes used as test chambers. The 4-mL portion remaining after sub-
sampling received radioactive carbon. Algal response was measured as total
inorganic carbon assimilated, calculated from measurements of radioactive
carbon assimilated and total inorganic non-radioactive carbon available,
rather than simply radioactive carbon assimilated.
The second problem became apparent after the variable non-radioactive
inorganic carbon concentration problem was resolved. Elutriates generally
stimulated algal carbon assimilation. Reducing algal cell density, increasing
medium concentration, or adding sodium nitrate, potassium phosphate dibasic or
sodium EDTA, singly, or in combination, did not increase algal carbon
assimilation in the control. Addition of sodium bicarbonate (60 mg/L) did
enhance growth of algae in the control, but elutriates were still stimulatory.
The nutrient concentration problem was not completely resolved. Only the
presence or absence of toxicity could be determined.
Based on the previous findings, the original method was revised. The
revised method will be described fully under the heading Toxicity Testing.
EFFECTS OF SAMPLE STORAGE
Whole sediments were stored for 12 to 16 months at 4" C before we could
re-test samples with the revised method. Because toxicity may change with
storage (Burton 1991), only a few samples were re-tested.
Storage effects were determined by preparing and testing elutriates from
stored whole sediments using the original algal method of initial studies.
Storage effects tests using the original algal method were conducted
concurrent with toxicity re-testing using the revised method. Although the
3-2
-------
toxicity of elutriates using the initial algal method could not be
interpreted, any change in the response pattern for a particular treatment
over time could indicate a change in toxicity if carbon concentrations were
unchanged. If changes in inorganic non-radioactive (I2carbon) concentrations
occurred during storage, the ratio of I4carbon/I2carbon would also change upon
testing and affect carbon assimilation. Therefore Microtox was also
performed as a second method to detect changes in toxicity with storage. The
Microtox test was selected for detection of long-term storage effects for the
same reasons it was used to evaluate weekly changes in sediment toxicity
described in Chapters 2 and 4. Methods for Microtox testing are described in
Chapter 2.
Both algal and Microtox methods indicated that storage of samples for
approximately a year would not affect the classification of elutriates as
toxic or non-toxic with the revised algal method. Toxicity to algae of these
elutriates of stored samples, using the revised algal method, is reported in
the remainder of this chapter.
SAMPLES TESTED USING THE REVISED METHOD
Elutriates prepared from sediment samples from three Great Lakes Areas
Of Concern (AOC) were re-tested: Indiana Harbor, IN (IH-01-07); Buffalo
River, NY (BR-01-03 and BR-01-08); Saginaw River and Bay, MI (SR-01-06 and SR-
01-10).
In addition to testing sediments from the AOCs, we tested a soil
elutriate (Missouri River sediment) from Florrissant, MO for comparison as a
control sediment from an uncontaminated site (Ingersoll and Nelson 1990).
Sediment collection, handling, and storage as well as elutriate
preparation procedures are described in Chapter 2.
ALGAL CULTURE
Selenastrum capricornutum Printz was cultured before tests under axenic
conditions in borosilicate glass flasks, with a 1:5 (v/v) medium:air ratio.
The culture medium was AAP medium but with EDTA reduced by two-thirds (Table
3.1). Culture flasks were continuously agitated at 50 to 100 rpm on a rotary
shaker table under constant light (300-350 ftc, approximately 30-35 ^E/m2-
sec).
An inoculum was prepared for each study from fresh culture stocks during
the exponential growth phase. We quantified stock culture cell density,
concentrated cells by centrifugation, rinsed cells with fresh medium, and then
diluted the algae in culture medium to an inoculum density of 10.5 x 106
cells/mL.
TOXICITY TESTING
Algal tests were conducted over a 24-h exposure period. Radiolabeled
carbon was added after 20 h of exposure. At 24 h (4 h incubation), the test
was ended. Photosynthesis was estimated as the amount of total carbon
assimilated by S. capricornutum. Nutrients (Table 3.1) were supplied equally
to all treatments.
The tests were conducted under sterile conditions. Bacteria, as well as
spuriousT(algae, were removed from elutriates samples with 0.45 jum Gelman
Metricel GN-6 cellulose acetate filters and glass fiber prefilters. All
dilution water and glassware were autoclaved. Minimization of bacteria
prevented continual addition of an unknown and varying amount of carbon
dioxide from bacterial decomposition of organic material.
Five serial dilutions were tested: 1%, 10%, 25%, 50%, and 94% elutriate.
Deionized water (<1 megohms/cm) was used as dilution water and as the negative
control (0% elutriate) treatment. Exposure chambers were 25-mL capped
borosilicate glass tubes. Exposure volume was 24.4 mL.
Six percent of each treatment was composed of medium and algal
3-3
-------
additions. Nutrients were added in 1 mL of concentrated medium to all control
and treatment tubes before addition of algae. Dilution of the medium
concentrate in the control treatment produced the same nutrient concentration,
except for sodium bicarbonate, as the culture medium, and at least the culture
medium concentration for elutriates with intrinsic nutrients. Sodium
bicarbonate concentration was increased so that dilution of the medium
concentrate in treatments produced a four-fold increase in carbon from the
culture medium. Algal inoculum was added to produce an exposure density of
190,000 cells/mL.
Total exposure time was 24 h at 21 + 2° C and 400 to 500 ftc
(approximately 40 to 50 pE/m2-sec) of light. After 20 h of exposure, total
inorganic carbon was determined before treatments were spiked and incubated
with carbon. Total dissolved carbon dioxide (equivalent to total inorganic
carbon) of acidified sub-samples removed before spiking, was measured with a
carbon dioxide ion selective electrode (Fisher model 13-620-506/507). After
sub-sampling, all treatments were spiked with 1 to 1.15 pCi of sodium '"c-
bicarbonate per mL of test volume. Incubation period was 4 h. At the end of
the incubation, unmetabolized 14carbon was removed by acidifying treatments
with concentrated HC1 and bubbling for 5 min with air. Scintillation cocktail
(Safety Solve") was added and radioactive carbon assimilation was measured in
a Beckman LS 3801 Beta Liquid Scintillation Counter. Treatment responses were
total inorganic carbon assimilation values, calculated using Vollenweider's
(1974) equation:
Carbon assimilated = carbon available * carbon assimilated / '"carbon
available.
To compare test results of different batches of algae (different dates),
responses were standardized as "percent of control" = (Treatment response /
average control response) * 100%. (A treatment with a response of 100% would
indicate no difference from controls.)
Blanks (additional control and 94% dilution treatment tubes which
received no algal inoculum) were included to quantify exposure fluorescence
due to factors other than S. capricornutum '"carbon assimilation.
WATER QUALITY CHARACTERIZATION
Alkalinity, pH, unionized ammonia, total chloride, turbidity, dissolved
oxygen, hardness and conductivity were measured on 0.45pm filtered elutriates
and control (dilution) water using the same methods described in Chapter 2.
Additionally, inorganic carbon was calculated from pH and alkalinity data
(Stumm and Morgan 1981).
PHYSICAL AND CHEMICAL CHARACTERIZATION OF SAMPLES
No chemical analysis was performed on the samples tested with the
revised algal method, filtered elutriates prepared from stored sediments.
Unfiltered elutriates, prepared shortly after sediment arrival, were analyzed
for inorganic contaminants (Chapter 2) and whole sediments were analyzed for
organic and inorganic contaminants (Chapter 4).
QUALITY CONTROL AND QUALITY ASSURANCE
Measures of quality control are reported in Table 3.2. The algal
toxicity test was performed with a reference toxicant, copper sulfate.
Negative (0% elutriate) control assimilation was determined by comparing
control assimilation to blank treatments which did not receive £.
capricornutum. Control assimilation was considered acceptable if it was
greater than blank assimilation: The actual amount of assimilation was not
expected to be consistent between tests because it is a function of several
factors besides culture health including age of the algal cells, the specific
activity of the '"carbon, and the length of time between reading and
preparation of scintillation samples. Unassimilated carbon removal
3-4
-------
efficiency was determined as:
(the average I4carbon spike (dpm) - ucarbon remaining in blanks after
bubbling (dpm)) / the average I4carbon spike (dpm).
Removal efficiency, while a function primarily of sample acidification
and bubbling, is also influenced by sample chemistry and the presence of
bacteria or other microorganisms. The carbon dioxide ion selective electrode
was calibrated to dilutions of a commercially prepared sodium bicarbonate
standard.
3-5
-------
RESULTS
WATER QUALITY CHARACTERIZATION
Water quality results for the 0.45 /jm filtered elutriates prepared from
stored sediments are presented in Table 3.4. Alkalinity, inorganic carbon,
hardness, conductivity, unionized ammonia, total chloride, and turbidity
varied most between AOC elutriate samples and control water (deionized water
with nutrients added). The control sediment was similar to the control water
for unionized ammonia, alkalinity, inorganic carbon and total chloride, but
more similar to AOC elutriate samples in terms of hardness, conductivity and
turbidity.
EXPOSURES
All of the elutriates of the stored samples were stimulatory i.e.,
produced greater algal carbon assimilation than the negative control (0%
elutriate), at one or more dilution concentrations (Figure 3.1). A positive
relationship between carbon assimilation and elutriate concentration was
evident for the control sediment and SR-01-10. Inhibition, i.e. mean response
less than the mean negative control response, occurred only in the 1%
elutriate concentrations of BR-01-03 and SR-01-06, but IH-01-07 and BR-01-08
did exhibit a decrease in stimulation at some of the higher concentrations
which may indicate toxicity.
CHEMICAL ANALYSIS
Results of whole sediment chemical analyses for selected organic and
inorganic compounds and metal chemistry for elutriates are reported in
chapters 4 and 2, respectively. Elutriate metal chemistry from chapter 2 is
presented in Table 3.3 for the subset of sites re-tested with the algal assay;
no chemical analyses were performed on the filtered elutriates prepared from
stored sediments.
Aging of the sediment and filtration of the elutriate propably altered
metal concentrations in the elutriates tested with the algal assay from the
elutriates described in chapter 2. The chemistry data is best used as an
indicator of relative site potential for toxicity. Assuming any contaminant
loss to be proportional to concentration regardless of the sample, the
elutriate prepared from IH-01-07 was the most contaminated, and the elutriates
prepared from the control sediment and SR-01-10 sediment were least
contaminated with metals. Organic toxicants were probably also present, but
relative concentrations in algal elutriates are difficult to estimate from
only whole sediment data.
3-6
-------
DISCUSSION
It was possible to classify sediments as toxic or non-toxic with the
algal assay, by comparing responses within an elutriate dilution series. A
toxic sediment is defined to be a sediment, that if suspended, is likely to
have adverse effects on natural algal populations. Effects may be sublethal,
such as reduced carbon assimilation rates, or lethal, to individual algal
cells, but either may ultimately result in algal population declines and / or
changes in algal community structure.
The algal stimulation produced by all the elutriates necessitated the
different approach to data interpretation from the planned comparison to a
negative control. Sediment elutriates may have contained both growth
promoting and inhibiting elements relative to the AAP medium. For a given
treatment, the response observed may have been the difference between
stimulation and inhibition relative to the AAP medium.
Stimulation also made it impossible to determine effective
concentrations (Nyholm and Kallqvist 1989) for elutriates. Thus, toxic
sediments could not be ranked or categorized by severity of toxicity as in
Chapter 2.
Test materials of a complex chemical nature may produce algal
stimulation relative to a negative control of AAP medium. Thomas et al.
(1986) reported stimulation from more than half the elutriate samples tested,
but none from identified toxicants tested as single chemicals. Stimulation is
reported in response to resuspended sediment (Ahlf et al. 1989), industrial
wastes (Walsh et al. 1982), textile mill effluents (Walsh et al. 1980) as well
as elutriates (Thomas et al. 1986). The phenomenon is not specific to
photosynthetic carbon assimilation, but is reported from growth (cell
enumeration) toxicity tests (Walsh et al. 1980, Walsh et al. 1982, Thomas et
al. 1986) and a test measuring chlorophyll a (Ahlf et al. 1989). Also, algal
stimulation does not appear to be a initial compensatory response to elutriate
toxicity in our test, for the following reasons: (1) the control sediment was
stimulatory and (2) the magnitude (about 200 to 300% of control) of
stimulation from the samples.
Elutriates and other complex mixtures, through perhaps either greater
concentrations and variety of nutrients or in creating more growth-conducive
chemical environments with pH, pH buffering, redox potential, etc., encourage
greater algal growth than the AAP medium alone can support. Water quality
measurements (Table 3.4) illustrate the disparities between the AAP medium and
the elutriates, especially between alkalinity, inorganic carbon concentration,
hardness, and conductivity. The elutriates have the greater ion
concentrations, and perhaps nutritive concentrations as well.
In the absence of toxicants, growth increases with elutriate
concentration. Increasing elutriate concentration of the control sediment
increased stimulation (Figure 3.1). A resuspended sediment produced
chlorophyll a stimulation of Ankistrodesmus bibraianus in an assay using a
chamber device technique, while an elutriate, a more dilute preparation of the
same sediment, was not stimulatory in a growth assay (Ahlf et al. 1989).
If potential growth was equally limited in all treatments, i.e., the
normal test conditions for single chemical testing, observed when we tested
copper sulfate, treatment responses would be less than or equal to the
negative control. The null response curve for the sample dilution series
would be a slopeless line equivalent to the negative control response ("no
stimulation, no toxicity", Figure 3.2). Toxicity would be indicated by
decreased carbon assimilation relative to the negative control as in "no
stimulation, constant toxicity" and "no stimulation, increasing toxicity",
Figure 3.2. Results could be expressed as the effective concentration which
reduced growth (assimilation) potential by 50% (i.e., EC50).
Great Lakes samples were labeled toxic or non-toxic using the following
procedure: Results were interpreted without relation to negative controls,
because each treatment has a different unknown potential for growth in the
absence of toxicants (dependent upon the concentration of elutriate and its
concentration of nutritive elements). The AAP medium was intended to function
3-7
-------
as a negative control, but instead served only as a means of standardizing
algal responses for different test dates.
Without a negative control, the null response for a sample dilution
series was an unknown. Sample toxicity can be indicated by deviation of the
dilution-series response curve ("stimulation, increasing toxicity", Figure
3.2) from the shape of a hypothetical non-toxic sample's response curve
("stimulation, no toxicity", Figure 3.2), however. This null response curve
would be a function of the increasing concentrations of limiting sample
nutrients. It would have a positive slope, although a plateau might be
reached, and the 0% elutriate control response would be the lowest value. An
actual negative control curve could not be constructed for a given sample
dilution series because points are functions of unknown nutrient
concentrations.
Constant toxicity across dilutions would have a response curve
indistinguishable from the null (no toxicity) response curve, and therefore
may not be recognized ("stimulation, constant toxicity", Figure 3.2).
However, increasing toxicity would be indicated by a decrease in stimulation
("stimulation, increasing toxicity", Figure 3.2). Thus, samples could be
classified as toxic, or with less certainty, non-toxic.
SEDIMENT TOXICITY
The control sediment and SR-01-10 produced increasing stimulation with
increasing concentration, as expected for a non-toxic sample (Figure 3.1).
There was no evidence of toxicity for either sample at any dilution.
The IH-01-07 and BR-01-08 elutriates were classified as toxic (Figure
3.1). After exhibiting increasing stimulation, a decline occurred at the
upper dilutions.
The BR-01-03 elutriate may have been toxic. Algal response was similar
to the control sediment and SR-01-10, but the response was lower than the
negative control at the lowest dilution (Figure 3.1).
The SR-01-06 elutriate was classified as toxic but with less certainty
than the IH-01-07 and BR-01-08 samples. The response curve vacillated between
dilutions unlike other elutriates, and although stimulation tended to increase
with increasing concentration, carbon assimilation was less than the negative
control at the lowest dilution.
CHEMISTRY
The elutriate with the highest metal concentrations (IH-01-07) was also
most certainly classified as toxic with the previous approach, although algae
could be responding to contaminants other than those analyzed.
Chemical analyses and algal toxicity tests were performed upon samples
from the same sediments, but they were not performed upon the same samples.
The processes of storage, elutrification and filtration affect the
concentrations of toxicants in the algal test samples. Storage may decrease
or increase sediment toxicity as some toxicants volatilize, and others degrade
chemically or microbially to lesser or more toxic forms. Elutrification,
discussed in chapter 2, may extract water soluble toxicants from the sediment,
while toxicants organically bound to particulate matter may be lost when the
elutriate is poured off. Filtration may further reduce contaminant levels by
removing more particulate matter (Burton 1991). It is impossible to say what
chemical concentrations were present in the elutriates tested in the algal
assay, and therefore to attempt to correlate chemistry and algal toxicity.
3-8
-------
SUMMARY AND RECOMMENDATIONS
Elutriates prepared from selected stored sediment samples were tested
with an algal carbon assimilation method developed for use with materials of
varying carbon concentrations. IH-01-07, BR-01-08, BR-01-03, and SR-01-06
were classified as toxic to algae. The control sediment and SR-01-10 were
non-toxic. It was not possible to correlate sample chemistry and algal
toxicity, but sites identified as toxic did have higher concentrations of
suspected toxicants than the sites designated non-toxic. These results
suggest sites to target for clean up exist at all 3 AOC, but that SR-01-10 may
serve as a non-toxic reference site for algae.
All elutriates were stimulatory relative to the negative control, AAP
medium. Stimulation complicated interpretation of results and prevented
calculation of effective concentrations. The AAP medium did not suffice as a
negative control and nutrient supplement for elutriates. It did not provide
unlimited algal nutrients or other unknown growth-limiting factors in all
treatments of the toxicity test. Samples were classified as toxic or non-
toxic based upon dilution series response curves, but effect concentrations
(i.e., ECSOs) could not be calculated.
The sensitivity demonstrated by S. capricornutum to sediment
contaminants warrants the development of an algal medium that can be used to
evaluate toxicity, without the complication of stimulation. With the right
medium, this algal test could be a consistent, sensitive, rapid, reproducible,
small-volume, means of screening and ranking environmental samples for
toxicity.
There are several approaches that could be taken to find a medium
appropriate to toxicity testing of environmental samples. Further
experimentation could be done on appropriate algal density, medium
concentration, and concentrations of medium nutrients individually or in
combination; effects of aerating chambers or modifying the AAP medium by
adding a pH buffer or a soil extract or other organic materials (Berman et al.
1991); or another published medium could be substituted. Bold's and OECD
media produce significantly greater algal growth than the AAP medium
(Millington et al. 1988).
The ideal medium would have an excess of nutrients such that nutrients
inherent to test materials would not stimulate growth. However, the ideal
medium would not have nutrient concentrations in excess of what is necessary
to provide unlimited growth. Algal nutrients may affect the toxicity of other
materials (Hornstrum 1990) or become toxicants with increasing concentrations
(Malone et al. 1975). Finally, an ideal medium would be defined, i.e., one in
which all materials and their concentrations are identified, for comparability
of results between laboratories.
3-9
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Ahlf, W., W. Calmano, J. Erhard and U. Forstner. 1989. Comparison of five
bioassay techniques for assessing sediment-bound contaminants. Hydrobiologia
188/189:285-289.
Amblard, C., P. Couture and G. Bourdier. 1990. Effects of a pulp and paper
mill effluent on the structure and metabolism of periphytic algae in
experimental streams. Aquat. Toxicol. 18:137-162.
American Society for Testing and Materials. 1990. D 3978-80: Standard practice
for algal growth potential testing with Selenastrum capricornutum. In ASTM,
Annual Book of Standards, Vol. 11.04. ASTM, Philadelphia, PA, pp.19-23.
Herman, T., S. Chava, B. Kaplan and D. Wynne. 1991. Dissolved organic
substrates as phosphorus and nitrogen sources for axenic batch cultures of
freshwater green algae. Phycologia 30:339-345.
Bokn, T. 1990. Effects of acid wastes from titanium dioxide production on
biomass and species richness of benthic algae. Hydrobiologia 204/205:197-203.
Burton, G.A. Jr. 1991. Assessing the toxicity of freshwater sediments.
Environ. Toxicol. Chem. 10:1585-1627.
Clesceri, L.S., A.E. Greenberg and R.R. Trussell (eds.). 1989. Standard
Methods for the Examination of Water and Wastewater. 17th ed. American Public
Health Association, Washington, D.C. pp. 8-42 to 8-52.
Couture, P., R. Van Coillie, P.G.C. Campbell and C. Thellen. 1981. Le
phytoplancton, un reactif biologique sensible pour detecter rapidement la
presence de toxiques. INSERM (Inst. Nat'l. Sante Rech. Med.) Colloq. 106:255-
272.
Elnabarawy, M.T. and A.N. Welter. 1984. Utilization of algal cultures and
assays by industry. In L.E. Shubert, ed., Algae as Ecological Indicators.
Academic Press, New York, NY. pp. 317-328.
Environmental Protection Agency. 1985. Subpart B—aquatic guidelines, section
797.1050, algal acute toxicity test. Federal Register 50(188):39321-39331.
Greene, J.C., W.E. Miller, M. Debacon, M.A. Long and C.L. Bartels. 1988. Use
of Selenastrum capricornutum to assess the toxicity potential of surface and
ground water contamination caused by chromium waste. Environ. Toxicol. Chem.
7:35-39.
Havens, K.E. and R.T. Heath. 1990. Phytoplankton succession during
acidification with and without increasing aluminum levels. Environ. Pollut.
68:129-145.
Herman, D.C., W.E. Inniss and C.I. Mayfield. 1990. Impact of volatile aromatic
hydrocarbons, alone and in combination, on growth of the freshwater alga
Selenastrum capricornutum. Aquat. Toxicol. 18:87-100.
Hornstrom, E. 1990. Toxicity test with algae—a discussion of the batch
method. Ecotoxicol. Environ. Saf. 20: 343-353.
International Joint Commission Great Lakes Water Quality Board. 1987. Report
on Great Lakes water quality, Appendix A: Progress in developing remedial
action plans for areas of concern in the Great Lakes basin. Technical Report.
3-10
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Ingersoll, C.G. and M.K. Nelson. 1990. Testing sediment toxicity with Hvalella
azteca (Amphipoda) and Chironomus riparius (Diptera). In W.G. Landis and W.H.
van der Schalie, eds., Aquatic Toxicology and Risk Assessment. Vol. 13, ASTM
STP 1096. American Society for Testing and Materials, Philadelphia, PA, pp.
93-109.
Kusk, K.O. and N. Nyholm. 1991. Evaluation of a phytoplankton toxicity test
for water pollution assessment and control. Arch. Environ. Contam. Toxicol.
20:375-379.
Malone, R.F., K.A. Voos, W.J. Grenney and J.H. Reynolds. 1975. The effects of
media modifications upon Selenastrum capricornutum in batch cultures.
Proceedings. Biostimulation and Nutrient Assessment Workshop, Logan, UT, Sept.
10-12, 1975, EPA-660/3-75-034. pp. 267-292.
Miller, W.E., J.C. Greene and T. Shiroyama. 1978. The Selenastrum
capricornutum Printz algal assay bottle test. EPA-600/9-78-018. U.S.
Environmental Protection Agency, Corvallis, OR.
Millington, L.A., K.H. Goulding and N. Adams. 1988. The influence of growth
medium composition on the toxicity of chemicals to algae. Water Res.
22(12):1593-1597.
Molander, S., H. Blanck and M. Soderstrom. 1990. Toxicity assessment by
pollution-induced community tolerance (PICT), and identification of
metabolites in periphyton communities after exposure to 4,5,6-
trichloroguaiacol. Aquat. Toxicol. 18:115-136.
Nyholm, N. and T. Kallqvist. 1989. Methods for growth inhibition toxicity
tests with freshwater algae. Environ. Toxicol. Chem. 8:689-703.
Porcella, D.B. 1983. Protocol for bioassessment of hazardous waste sites. EPA-
600/2-83-054.
Rai, L.C., A.K. Singh and N. Mallick. 1990. Employment of CEPEX enclosures for
monitoring toxicity of Hg and Zn on in situ structural and functional
characteristics of algal communities of River Ganga in Varanasi, India.
Ecotoxicol. Environ. Saf. 20:211-221.
Ross, P., V. Jarry and H. Sloterdijk. 1988. A rapid bioassay using the green
alga Selenastrum capricornutum to screen for toxicity in St. Lawrence River
sediment elutriates. In J. Cairns, Jr. and J.R. Pratt, eds., Functional
Testing of Aquatic Biota for Estimating Hazards of Chemicals, ASTM STP 988.
American Society for Testing Materials, Philadelphia, PA, pp. 68-73.
Sloterdijk, H., L. Champoux, V. Jarry, Y. Couillard and P. Ross. 1989.
Bioassay responses of micro-organisms to sediment elutriates from the St.
Lawrence River (Lake St. Louis). Hydrobiologia 188/189:317-335.
Stumm, W. and J.J. Morgan. 1981. Aquatic Chemistry. 2nd ed. John Wiley & Sons,
New York, NY.
Thomas, J.M., J.R. Skalski, J.F. Cline, M.C. McShane, J.C. Simpson, W.E.
Miller, S.A. Peterson, C.A. Callahan and J.C. Greene. 1986. Characterization
of chemical waste site contamination and determination of its extent using
bioassays. Environ. Toxicol. Chem. 5:487-501.
Vollenweider, R.A. 1974. Chemical and physico-chemical procedures directly
involved in primary production measurements. In R.A. Vollenweider, ed., IBP
Handbook No. 12; A Manual on Methods for Measuring Primary Production in
Aquatic Environments. 2nd ed. Blackwell Scientific Publications, Oxford,
England, pp. 53-58.
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Walsh, G.E., K.M. Duke and R.B. Foster. 1982. Algae and crustaceans as
indicators of bioactivity of industrial wastes. Water Res. 16:879-883.
Walsh, G.E., L.H. Bahner and W.B. Horning. 1980. Toxicity of textile mill
effluents to freshwater and estuarine algae, crustaceans and fishes. Environ.
Pollut. (Series A) 21:169-179.
Whitton, B.A. 1984. Algae as monitors of heavy metals in freshwaters. In L.E.
Shubert, ed., Algae as Ecological Indicators. Academic Press, New York, NY,
pp. 257-280.
3-12
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LIST OF FIGURES IN CHAPTER 3
Figure 3.1 S. capricornutum responses to elutriate sample doses (Mean and
standard deviation).
Figure 3.2 Hypothetical S. capricornutum dose-response curves for stimulatory
and nonstimulatory samples.
3-13
-------
Percent of Control Response
c
CD
UJ
m
i—*
S'
CD
a a s as g a
-------
Stimulation
No Stimulation
NoToxicity
Constant Toxicity
Increasing Toxicity
CD
CO
C
o
Q.
CO
0
tr
o
o
CD
o
CD
Q_
100%
Figure 3.2
Low
Elutriate (%)
High
-------
Table 3.1. Algal culture medium (after Miller et al. 1978)
Compound
Concentration
NaNO,
MgCl26H20
CaCl22H20
MgSO47H2O
K2HPO4
NaHC03
HjBOj
MnCl24H2O
ZnCl2
COC126H2O
CuCl22H2O
FeCl36H20
Na2EDTA'2H2O
25.500 mg/L
12.160
4.410
14.70
1.044
15.OO1
185.52 pg/L
415.38
3.27
1.428
0.012
7.26
160.00
100.O2
7.5'
pH
C : N : P
30 : 50 : 1 mol.'
Parameter altered for the revised algal test: NaHCO3 = 60.000 mg/L, pH =
8.1, C : N: P = 119 : 50 : 1 mol.
Concentration reduced from the Algal Assay Procedure medium concentration of
300.0 Aig/L.
-------
Table 3.2. Measures of quality assurance for algal assays.
Assay Cu2" EC50 (p/g/L) Control Response1 '4CO2(free) Removal Efficiency
(Control / Blank 14C (%)
Assimilation)
IH-01-07, CONTROL2,
BR-01-08, SR-01-10
BR-01-03, SR-01-06
Couture et al. 1981
NM
10 (3-18)3
4 < EC50 < 8
104
58
NA
99.
99.
NA
97-99.98
94-99.97
i j
water.
Elutriate prepared Dec. 1990, from stored, dried soil of control sediment.
3Mean and 95% confidence interval.
NM Not measured.
NA Information not available.
-------
Table 3.3. Concentrations of metals in non-filtered elutriates prepared within 2 weeks of sample receipt.
Note, algal assays were performed on filtered elutriates prepared from the same samples after >
1 year storage.
Metal concentration (pg/L)
Sample
CONTROL
SEDIMENT
IH-01-07
BR-01-03
BR-01-08
SR-01-06
SR-01-10
Ag
<0.
1.
0.
<0.
<0.
<0.
03
3
33
03
09
09
As
-------
Table 3.4. Water quality of elutriates tested with the algal assay.
Sample
CONTROL SEDIMENT2
IH-01-07
BR-01-03
BR-01-08
SR-01-06
SR-01-10
0% ELUTRIATE CONTROL
i
PH
7.5
8.2
8.0
8.1
8.1
8.0
8.1
Alkalinity
(mg/L as
CaC03)
40
124
88
152
NM3
124
36
Hardness
(mg/L as
CaC03)
116
168
132
188
172
180
16
D.O.
(mg/L)
9.7
10.8
9.2
10.5
8.6
9.8
9.5
Conduct ivity
(pmhos/cm)
238
378
300
378
372
322
81
Unionized
Ammonia
(mg/L)
0.002
0.869
0.205
0.463
0.169
0.100
0.0054
Total
Chloride
(mg/L)
6.9
11.3
10.1
11.2
15.3
15.4
7.3
Turbidity
(NTU)
2.6
8.1
0.4
1.3
2.4
1.6
0.2
Inorganic
Carbon
(nmol/L)
850
2500
1800
3100
NM3
2500
720
Inorganic carbon is estimated from pH and alkalinity data (Stumm and Morgan 1981).
Elutriate prepared from stored dried soil 1990.
Values not measured.
Values below detectable limits.
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CHAPTER 4; WHOLE SEDIMENT TOXICITY TESTS
Nelson, M.K., J.J. Coyle, L.B. King, N.E. Kemble, E.A. Crecelius and I.E.
Greer
INTRODUCTION
Contaminated sediments are a persistent source of toxic materials to
aquatic resources because of their ability to accumulate contaminants in
concentrations frequently greater than that of the overlying water (Medine and
McCutcheon 1989), and as a result, can have significant effects on biota and
the overlying water quality. During re-suspension and redistribution, and
accumulation in depositional areas, contaminated sediment may cause
deleterious effects to the benthos and food chain organisms. Because
indigenous benthic organisms are continuously exposed to complex mixtures of
contaminants associated with sediment, they may exhibit lethal and sublethal
effects such as, mortality, diminished growth, delayed maturation, and
reproductive impairment.
The interaction of complex mixtures of sediment-associated contaminants
can be integrated by using whole sediment toxicity tests (Swartz et al.,
1979). Whole sediment laboratory toxicity tests can measure the effect of all
bioavailable contaminants, where bioavailable is defined as the fraction of
the total contaminant in the interstitial water and on the sediment particles
that is available for biological processes (Landrum and Robbins 1990). If the
test organism's critical life stages are used, ecotoxicologically relevant
information can be obtained (Samoiloff, 1989). Toxicity assessments using
aquatic invertebrates provide valuable information on the relative degree of
aquatic resource degradation and a direct measure of contaminant
bioavailability (Swartz et al. 1982, Nebeker et al. 1984, Breteler et al.
1989, Scott and Redmond 1989, Ingersoll and Nelson 1990). Test organisms
exposed to field-collected sediments integrate interactions among complex
mixtures of contaminants that may be present at the sediment collection site.
TOXICITY TESTS
The objective of the whole sediment laboratory exposures was to evaluate
the biological effects of sediment contamination to benthic
macroinvertebrates. The laboratory experimental design was based on the
assumption that the behavior and effects of the sediment-associated
contaminants is similar to that in in situ sediments. Relative comparisons
between species sensitivities were made. Sediment contaminant concentrations,
and the toxicity responses observed, were compared to recommended threshold
concentrations used to identify sediment toxicity (Barrick et al. 1988; Long
and Morgan 1990; SMS 1991).
The present studies evaluated the potential toxicity of whole sediments
collected from three Areas of Concern (AOCs) within the Great Lakes: Indiana
Harbor, IN; the Buffalo River, NY; and the Saginaw River, MI. The studies
were conducted for the Assessment and Remediation of Contaminated Sediments
(ARCS) with the U.S. Environmental Protection Agency's (USEPA) Great Lakes
National Program Office (GLNPO). Three test species were exposed in flow-
through or static exposures, in partial life-cycle (10-d, 14-d and 28-d)
toxicity tests. The test organisms included: Hyalella azteca (Amphipoda) 14
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and 28-d flow-through exposures; Chironomus riparius (Diptera) 14-d flow-
through exposure; and Chironomus tentans (Diptera) 10-d static exposure. The
toxicological endpoints measured were survival and growth (body length) for
all test species, antennal segment number and sexual maturation for H. azteca.
These species have been recommended by the International Joint Commission
(1987) and ASTM (E 1383-90) for whole sediment toxicity evaluations.
Hvalella azteca (Amphipodal
The amphipod, H. azteca, is a common and widely distributed Talitrid
amphipod inhabiting permanent lakes, ponds, and streams, throughout most of
the American continent and the Caribbean (Bousfield 1958, DeMarch 1981, Pennak
1989). Its life cycle is divided into three stages: (1) immature, including
the first five instars; (2) juvenile, including instars 6 and 7; and (3)
adult, including the 8th instar and older (Cooper 1965, Pennak 1989). Adult
males are larger than females and have an enlarged second gnathopod propodus
(Giesler 1944). As a toxicity test organism, H. azteca has several desirable
characteristics: (I) it has a short generation time, (2) is easily collected
from natural sources or laboratory cultured, (3) is ecologically relevant, and
(4) is sensitive to contamination (Maciorowski 1975, Borgmann et al. 1989).
For assessment and management of contaminated sediments, H. azteca has been
used in sediment toxicity testing, both under laboratory and field conditions
(Landrum and Scavia 1983, Cairns et al. 1984, Nebeker et al. 1984, 1986,
Nebeker and Miller 1988, Burton et al. 1989a, Borgmann and Munawar 1989,
Ingersoll and Nelson 1990).
Chironomus riparius and Chironomus tentans (Diptera)
Chironomus tentans and C. riparius (midge larvae) live most of their
life cycle as burrowing aquatic insects. They are frequently found in large
numbers in North American lakes and ponds (ASTM E 1383-90). Owing to their
frequently high density, chironomids constitute a biomass of considerable
ecological importance as nutrient processors and as food organisms for higher
trophic levels. The life cycles of C. riparius and C. tentans can be divided
into three stages: (1) a larval stage (first four instars); (2) a pupal stage;
and (3) an adult stage. Under optimal conditions C. riparius larvae will
pupate in 15 to 21 days at 20°C and C. tentans larvae will pupate after 24 to
28 days at 20°C. Because midge larvae are in direct contact with the sediment
and consume a wide variety of organic substances, they are sound biological
indicators of habitat quality and represent an important route of entry for
contaminants into food webs. Due to their short life span and ease of
laboratory culture, midges have been used frequently for sediment toxicity
testing (Adams 1985, Giesy et al. 1988, Pittinger et al. 1989, Ingersoll and
Nelson 1990).
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MATERIALS AND METHODS
SUMMARY OF SEDIMENT SAMPLES TESTED
The GLNPO ARCS Toxicity-Chemistry Work Group (Table 1.1) selected
stations for whole sediment toxicity evaluations. Stations were chosen based
on the expected presence of contaminants which would elicit both acute and
chronic effects. Of the 29 stations sampled between 1988 and 1990, from the
three AOC, 19 were chosen for whole sediment toxicity evaluation. The
following Great Lakes sediment samples were evaluated:
Indiana Harbor, IN, IH-O1-03, IH-O1-04, IH-O1-06 and IH-O1-07;
Buffalo River, NY, BR-01-01, BR-01-03, BR-01-07, BR-01-08 and BR- 01-09;
Saginaw River, MI (First survey), SR-01-03, SR-01-06 and SR-01-101;
Saginaw River, MI (Third survey), SR-03-01, SR-03-02, SR-03-05, SR-03-
06, SR-03-08, SR-03-16 and SR-03-24)2.
For additional information regarding locations, water depths, and sample
types, see Table 2.1. For descriptions of collection methods see Chapter 1
(Overview) this report.
Sediment Storage
Sediment sampling at all AOCs required more than one day. Reported
storage days are the number of days between the first date of sediment
collection at an AOC, and the date when sediments were put into test chambers.
Indiana Harbor—
Sediment samples were collected from Indiana Harbor between 8/9/89 and
8/10/89. Sediment from Stations IH-01-03, IH-01-04 and IH-01-06 were
collected on a/9/89, and from Station IH-01-07 on 8/10/89. All samples were
received at the National Fisheries Contaminant Research Center (NFCRC) on
8/11/89.
Elutriates were prepared on 8/15/89, 8/30/89 and 9/6/89 (see Chapter 2).
Sediments were homogenized and put into test chambers on the days indicated:
14 d H. azteca. 8/30/89 (21 days storage); C. riparius and C. tentans. 9/6/89
(28 days storage). The time between collection and placing sediment in test
chambers ranged from 21 to 28 days.
Buffalo River—
Sediment samples were collected from the Buffalo River between 10/6/89
and 10/8/89. Sediment was collected from the following stations on the days
indicated: BR-01-03, BR-01-04 — 10/6/89; BR-01-06, BR-01-07, BR-01-09 —
10/7/89; and BR-01-01, BR-01-08 — 10/8/89. Samples arrived at NFCRC on
10/10/89 at 8°C. Elutriates were prepared on 10/11/89 and 10/25/89 (see
Referred to as SR-01 when discussing stations, or Saginaw River (1)
when discussing AOCs.
2 Referred to as SR-03 when discussing stations, or Saginaw River (3)
when discussing AOCs.
4-3
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Chapter 2). Sediments were homogenized and put into test chambers on the days
indicated: C. riparius and 28-d H. azteca 10/23/89 (17 days storage); 14 d H.
azteca and C. tentans 10/25/89 (19 days storage). The time between collection
and placing sediment into test chambers ranged from 17 to 19 days.
Saginaw River (1)—
Sediment samples were collected between 12/01/89 and 12/02/89 from
Saginaw River (1). Sediment was collected from the following stations on the
days indicated: SR-01-10 — 12/01/89; and SR-01-03, SR-01-06 — 12/02/89.
Samples arrived at NFCRC on 12/5/89. Elutriates were prepared on 12/6/89,
12/19/89, and 12/27/89 (see Chapter 2). Sediment was homogenized and put into
test chambers on the days indicated: 28 d H. azteca, 12/19/89 (18 storage
days); C. riparius. 12/20/89 (19 storage days); 14 d H. azteca 12/27/89 (26
storage days). The time between collection and placing sediment into test
chambers ranged from 18 to 26 days.
Saginaw River (3)—
Sediment samples were collected between 6/21/90 and 6/23/90 from Saginaw
River (3). Sediment was collected at the stations on the following days: SR-
03-02, SR-03-05, SR-03-06 — 6/21/90; SR-03-08, SR-03-16, SR-03-24 — 6/22/90;
and SR-03-01 — 6/23/90. Samples arrived at NFCRC on 6/26/90 at temperatures
between 20°C to 21°C. Elutriates were prepared on 6/26/90, 07/09/90, 07/12/90
and 07/16/90. Sediment was homogenized and put into test chambers on the days
indicated: 28 d H. azteca 07/09/90 (18 days storage), C. riparius 7/12/90 (21
days storage), 14 d H. azteca, 7/16/90 (25 days storage), and 7/18/90 C.
tentans (27 days storage). The time between collection and placing sediment
into test chambers ranged from 18 to 27 days.
EXPOSURE PROCEDURES
Whole sediment tests conducted include the flow-through tests for
amphipods in 14- and 28-d exposures and C. riparius in a 14-d exposure
following procedures described in Ingersoll and Nelson (1990). The midge, C.
tentans. was tested in 10-d static exposures following procedures described in
Giesy et al. (1988). The effects of contaminants on survival and body length
were measured for all test species. In addition, effects on sexual
maturation, antennal segment number and increase in body length were measured
for amphipods. A fine silt- and clay-particle size soil was used for the
control sediment in all studies (Adams et al. 1985, Ingersoll and Nelson
1990).
Test animals were cultured at 20°C (~2°C) with a photoperiod of 16:8 h
light:darkness, and a light intensity of about 25 to 50 fc. Amphipods were
mass cultured in 80-L glass aquaria containing 50 L of NFCRC water (pH 7.8,
hardness 286 mg/L as CaCO3, alkalinity 258 mg/L as CaCO3, chloride 22 mg/L,
and conductivity 600 umhos/cm) with about three water additions daily, and
gentle aeration with air stones. Amphipods were cultured on hard maple leaves
(Acer spp.) that had been soaked in NFCRC water for about 30 d and then rinsed
with well water to remove any tannic acid. Also, ground Tetra" Standard Mix
Fish Food was fed weekly ad libitum (Ingersoll and Nelson 1990). Amphipods
were collected for testing by placing a part of the leaf substrate in a U.S.
4-4
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Standard stainless steel sieve stack (#25, #40, and #60) and rinsing with
culture water. Amphipods, about second instar, stopped by the #60 sieve (<2
mm), were used for testing.
Both of the midges were mass cultured in polyethylene chambers (30 x 30
x 30 cm) that were covered with nylon screen and contained 3 L of well water.
Chironomus riparius larvae were fed a combination of Cerophyl", green algae
Selenastrum capricornutum, and Hartz dog treats (Ingersoll and Nelson 1990).
The C. tentans chambers contained a shredded paper towel substrate, and larvae
were fed ground Tetrafin" in a well water suspension (Giesy et al. 1988).
Chironomus riparius were isolated for testing by placing fresh egg cases in
individual 200-mL crystallization dishes containing about 100 mL of culture
water at 20°C. Midge larvae began hatching after two to three days and were
collected for testing within 24 h of hatching. Chironomus tentans were
isolated for testing by placing fresh egg cases in glass baking dishes
containing culture water at 20°C. Larvae used for testing hatched after two
to three days and were maintained for 10 to 12 days (second instar), with
gentle aeration, and a diet of S. capricornutum and ground TetrafinR.
The reconstituted water used in the sediment tests is described in
Chapter 2 (pH 8.0, hardness 134 mg/L as CaC03, alkalinity 60 to 65 mg/L as
CaC03, sulfate 72 mg/L). Test animals were acclimated to the overlying water
before tests started on Day 0, by placing them at 2-h intervals into 50:50 and
25:75 mixtures of culture water to overlying water (Ingersoll and Nelson,
1990). At the start of each test (Day 0) a sub-sample of about 20 animals
were preserved with a sugar-formalin mixture (Ingersoll and Nelson 1990) and
archived. Test animals were also preserved and archived at the end of each
study. Animals were tested at 16:8 h light:darkness photoperiod and light
intensity of 25-50 fc (Ingersoll and Nelson, 1990, ASTM E 1383-90).
Flow-through Tests
The day before tests started (Day -1) sediment was homogenized, then
sediment and overlying water were added to each of four replicate test
chambers (Ingersoll and Nelson, 1990). Test chambers for amphipods and C.
riparius exposures contained 200 mL homogenized whole sediment and 800 mL
reconstituted overlying water (Ingersoll and Nelson 1990). Gentle aeration of
the overlying water was started, cover glasses placed on the test chambers,
and test systems equilibrated overnight in a 20°C water bath. Tests were
started on Day 0, using methods described in Ingersoll and Nelson (1990), with
the addition of 20 amphipods or 50 C. riparius larvae into each test chamber.
Larvae of C. riparius and amphipods were acclimated to the overlying water,
then counted into 30 mL beakers containing overlying water. Animals were
released into the test chambers and any animals on the surface were gently
submersed or replaced. Modified Mount and Brungs proportional flow-through
diluters delivered overlying water (Mount and Brungs 1967). To allow the
larvae to settle into the sediment (Pittinger et al. 1989), water delivery
(1.25 volume additions test chamber~1*day~1) was started on Day 1 of the C.
riparius test, and on Day 0 in the amphipod tests (Ingersoll and Nelson
1990). Cover glasses and airlines were removed from the test chambers before
the water flow started. Test chambers were observed daily for the presence of
animals in the water column or on the sediment surface. Water condition
(e.g., surface film, cloudiness) was recorded, and sediment surface was
4-5
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observed for the presence of mold, unconsumed food particles, or signs of
burrowing behavior.
Amphipods were fed Purina" Rabbit Pellets in a water suspension
(Ingersoll and Nelson 1990) three times weekly at 6 mg for the first 7 d, then
12 mg to test end. Chironomus riparius larvae were fed a combination of
Cerophyl" (10 mg daily for the first 6 d), S. capricornutum (6 X 107 cells
daily), and HartzR dog treats (10 mg daily) (Ingersoll and Nelson 1990). When
excessive mold was present on the sediment surface of any replicate, food was
withheld for the entire treatment.
At test end, sediments were sieved to retrieve surviving test organisms
using methods outlined in Ingersoll and Nelson (1990). Remaining debris and
sediment from the collecting pans was preserved and later inspected under a
low power binocular microscope to recover any small animals (particularly
amphipods) which were not seen at test end.
Preserved amphipods were measured using a ZeissR Interactive Digital
Analysis System in combination with a Zeiss SV8 stereomicroscope at a
magnification of 25 X (Ingersoll and Nelson 1990). Two measurements of
amphipod body length (-0.1 mm), along the curve of the dorsal surface from the
base of the first antenna to the tip of the third uropod, were recorded, and
antennal segments were counted. To determine sexual maturation, amphipods
were sexed by the appearance of an enlarged propodus on the second gnathopod
for males, and the presence of eggs in the brood pouch or lack of an enlarged
propodus for females (Nelson and Brunson 1992). Archived amphipods from Day 0
were measured for comparisons to amphipods at the end of the 14- and 28-d
exposures. Chironomus riparius larvae were photographed on a 1 cm gridded
background and C. riparius lengths were digitized from enlarged prints (20 x
30 cm) with a Houston Instrument True-grid" 1017 digitizing board. Up to 20
midge per replicate were measured from the anterior of the labrum to the
posterior of the last abdominal segment (Smock, 1980).
Static Tests
Sediments collected from Indiana Harbor, Buffalo River and Saginaw River
(3) were tested with C. tentans using static, 10-d whole sediment toxicity
test methods, described by Giesy et al. (1988), with some modifications. Due
to diminished reproduction in C. tentans cultures, test organisms were not
available to conduct whole sediment tests on sediment samples collected from
Saginaw River (1). Additionally, only 10 individuals per station were tested
in Saginaw River (3) due to a larger number of samples for this survey and
limited space in the water bath.
Two days before the start of the test, 7.5 g of homogenized sediment
from each station was placed in 15 50 mL polypropylene conical centrifuge
tubes, or 10 tubes for Saginaw River (3) exposures. Forty mL of overlying
water was added to each test chamber. Sediment and water were mixed by
capping chambers and inverting five times in accordance with Giesy et al.
(1988). Test chambers containing sediment from the same station were inserted
in a styrofoam holder and placed in a vented, temperature controlled (23-l°C)
waterbath. One day before introduction of test organisms, caps were removed
and gentle aeration (about 3 bubbles*s"1) were delivered to the chambers.
Tests were started by placing one second instar C. tentans larva in each test
chamber.
4-6
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Biological observations (e.g., organism behavior) and observations of
sediment and water conditions (e.g., presence of mold, water clarity) were
performed daily on each test chamber. TetraFinR (6 mg/day) was added to each
chamber in an aqueous suspension (Giesy et al. 1988). If mold was observed on
the sediment surface or if the overlying water appeared cloudy, feeding was
withheld until conditions cleared or the test ended. Water lost to
evaporation was replaced as needed with temperature acclimated deionized water
(Giesy et al. 1988).
At the end of the 10-d exposure, sediment was passed through a U.S.
Standard #50 sieve to recover larvae. Surviving C. tentans larvae from test
chambers for each station were counted and combined into a single glass vial,
preserved and archived. Growth (dry weight) was determined gravimetrically
for midge exposed to Buffalo River sediments. Larvae were blotted on
absorbent paper to remove excess water, then individuals were placed on pre-
weighed pans and dried at 80°C. Chironomus tentans exposed to Indiana Harbor
and Saginaw River (3) sediments were measured from photographs using methods
described above for C. riparius.
To determine if the MicrotoxR test response to elutriates prepared from
stored sediment samples had changed over time, Microtox" tests were conducted
for all sediments used in whole sediment tests. Before each whole sediment
test, stored sediment samples were homogenized, elutriates prepared, then
tested for Microtox" toxicity. Elutriates prepared from the stored sediments
were tested with the same methods described in Chapter 2 with one exception:
elutriates prepared for the Microtox" and Daphnia maqna tests were stored for
about 1 week at 4°C, in the dark, whereas elutriates prepared from stored
sediments were tested within 24 h.
WATER QUALITY CHARACTERIZATION
The reconstituted overlying water described in Chapter 2 was used as the
overlying water in the whole sediment exposures (pH 8.0, hardness 134 mg/L as
CaC03, alkalinity 60 to 65 mg/L as CaC03, sulfate 72 mg/L). The methods used
for measuring water quality characteristics were the same as described in
Chapter 2.
Whole sediment exposures began on separate days because of the time
required to set up individual tests. Because of the staggered test set-up,
water quality assessment efforts were combined. Before starting water flow on
Day 0 of the first flow-through exposure, 50 mL of overlying water was removed
from each of the four replicates per treatment and pooled. The pH, dissolved
oxygen, conductivity, water hardness, alkalinity, turbidity, ammonia, and
chloride were measured and the values were used as Day 0 water quality
determinations for the remaining whole sediment tests within an AOC.
In the 14-d exposures with amphipods and C. riparius. the pH, dissolved
oxygen, and conductivity were determined on Day 7. On Day 13, pH, dissolved
oxygen, conductivity, total water hardness, alkalinity, turbidity, ammonia,
and chloride were measured. In the amphipod 28-d exposures, pH, dissolved
oxygen, and conductivity were measured on Days 7, 14, and 21; pH, dissolved
oxygen, conductivity, total water hardness, alkalinity, turbidity, ammonia,
and chloride were measured on Day 27. In the C. tentans exposure, pH,
dissolved oxygen, conductivity, total water hardness, alkalinity, turbidity,
ammonia, and chloride were measured on Day 10; no Day 0 water quality
determinations were conducted. For the Day 10 water quality determinations,
4-7
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about 40 mL of exposure water per chamber (within each treatment) was
collected and pooled.
STATISTICAL ANALYSIS
Survival (except C. tentans), body length (H. azteca. C. riparius, C.
tentans), body weight (C. tentans), increase in length, antennal segment
number and sexual maturation (H. azteca) were tested for differences among
means. Before testing for differences, survival, increase in length, and
sexual maturation data were arcsine square root transformed. Variances were
compared using the general linear model (GLM) one way analysis of variance
(ANOVA) least squares means Type III test at alpha = 0.05 (SAS 1988).
Homogeneity of variance was tested using an F test, at alpha =0.05, in the
GLM and ANOVA procedures. Survival data for C. tentans was analyzed using a
categorical model procedure (SAS 1988) that performed a statistical analysis
of multivariate categorical data described by Koch et al. (1977). Significant
differences for sediment treatments are presented as differences compared to
control, and not differences between stations or AOCs. Sediment samples were
considered toxic if any test organism response was significantly different
from the control treatment.
SEDIMENT PHYSICAL AND CHEMICAL CHARACTERIZATIONS
Sediment physical characterization included (1) percentage solids, and
(2) particle size. Sediment chemical characterization included (1) metals,
(2) organometals, (3) acid volatile sulfide (AVS) and simultaneously extracted
metals (SEM), (4) chlorinated pesticides, (5) polychlorinated biphenyls
(PCBs), (6) polychlorinated dibenzo-dioxins (PCDDs) and -furans (PCDFs), (7)
select polynuclear aromatic hydrocarbons (PAHs), and (8) total organic carbon
(TOG). All chemicals analyzed are listed in Table 4.1. ^
All chemical analyses on sediment samples were provided by Battelle
Laboratory, Sequim, WA. The samples for chemistry were collected by personnel
of the Large Lakes Research Station, Grosse Isle, MI, and shipped by overnight
courier to Battelle. In most cases, one 2-L bottle of sample from each
station was collected and shipped for sediment chemical analyses. The samples
were then placed in an automated, continuously monitored cold storage room
until they were analyzed. For analyses, the samples were sub-sampled as
follows:
1. 50 g for metals, percentage solids, and TOC analyses,
2. 250 g for analysis of PAH,
3. 50 g for tributyltin analysis,
4. 20 g for AVS and 20 g for methylmercury analyses, and
5. 100 g for Ames (Chapter 6) and Mutatox (Chapter 7) assays.
The percentage solids in each sediment sample was estimated by freeze
drying the sample and then comparing wet and dry weights. Freeze drying
provided a fine, powdery sample that could be more uniformly homogenized. The
TOC in samples was determined with a Leco Model WR-12 carbon determinator.
Samples were pre-treated with concentrated HC1 to remove inorganic carbon.
Then the samples were burned at 800°C in an oxygen atmosphere connected to a
boat inlet which transferred the evolved CO2 directly into an organic carbon
analyzer. Particle size was determined with a Gilson Model WV-2 wet siever.
4-8
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using U.S. Standard #18 (1 mm), 60 (250 /«n) , 230 (63 pm) and 400 (38
sieves.
Acid volatile sulfides were determined with the method of Cutter and
Oattes (1987). In this method, hydrogen sulfide is selectively generated,
cryogenically trapped, separated by gas chromatography, and detected by
photoionization. The SEM are determined by analyzing the 1NHC1 solution
produced during the AVS determination. The 1NHC1 is filtered through a 0.3/tm
pore filter and analyzed by graphite furnace atomic absorption. Because
sediments samples were homogenized in the field and exposed to air, they may
not be representative of ambient field conditions, but are representative of
the sediment samples used in the toxicity tests which were homogenized and
stored in a similar manner. Sub-samples for AVS analyses should be isolated
before open air homogenization.
Simultaneously extracte'd metal concentrations were not determined for
Saginaw River (3), but were estimated using the vigorously extracted metal
concentrations (/ig/dry g). Saginaw River (3) vigorously extracted, individual
metal concentrations were converted to molar concentrations (/xM/dry g). To
obtain the estimates of the SEM concentration, the Saginaw River (3) metal
concentrations (/tM/dry g) were multiplied by the average Saginaw River (1) SEM
concentration for each metal.
The sediment samples were analyzed for total metals concentrations by
use of the USEPA method 200.4 (USEPA 1990). These techniques are not intended
to measure biologically significant metals. The samples were completely
dissolved by digestion with nitric, perchloric and hydrofluoric acids in
Teflon" pressure vessels and then analyzed by use of cold vapor atomic
absorption, or graphite furnace atomic absorption. For crustal elements that
are difficult to dissolve with strong acids a portion of the freeze-dried
samples was ball-milled to about 120 mesh, pelletized, and analyzed with x-ray
fluorescence (Nielson and Sanders 1983).
In methylmercury analyses, the homogenized samples were digested in 10
mL of a 25 percentage solution of potassium hydroxide in methanol at 60°C for
two to four hours. Samples were allowed to cool for 24 h and an additional 10
mL of methanol was added and mixed well by shaking. Before analysis
undissolved solids were allowed to completely settle. The samples were
analyzed with a cold vapor atomic fluorescence technique (Bloom 1989). The
technique is based on the emission of 254 nm radiation by exiting mercury
atoms in an inert gas stream. An ethylating agent, sodium tetraethylborate,
was added to the sample digestate to form a volatile methylethylmercury
derivative. The derivative was then purged onto graphite carbon traps for
pre-concentration and removal of interferences. Then the sample was subjected
to cryogenic chromatography and pyrolytic degradation to elemental mercury,
which was quantified with a cold vapor atomic fluorescence detector.
In analyses for organotins, samples were extracted with 0.2% tropolone
in methylene chloride, then filtered through glass wool. The filtrates were
derivitized with 1 mL hexyl magnesium bromide, a Grignard's reagent, and
cleaned-up with a Florisil column. Organotin concentrations were measured
with a Hewlett Packard Model 5890 gas chromatograph equipped with a flame
photometric detector.
Three groups of organic chemicals were measured for each sediment
sample: (1) PAH, (2) PCB and chlorinated pesticides, and (3) PCDDs and PCDFs.
The analytical procedure for each chemical group included solvent extraction,
extract purification with column chromatography, and chemical quantification
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with capillary column gas chromatography. In the analyses for pesticides and
PCBs, Aldrin, B-BHC, Gamma Chlordane, 4,4,-ODD, 4,4,-DDE, 4,4,-DDT, Dieldrin,
Endosulfan I, Endosulfan II, Endrin, Endrin aldehyde, Endrin ketone,
Heptachlor epoxide, Aroclor 1242 and 1254 were detected in some samples, but
either a less than 25% difference between two gas chromatography columns for
detected concentrations was observed, or the analyses were conducted at
secondary sample dilution factors.
Polynuclear aromatic hydrocarbons in sediment samples were determined
according to the USEPA method 3550 (USEPA 1986a). Before extraction, three
isotopically labelled surrogate PAH compounds (DIO-fluorene, DIO-anthracene,
DIO-pyrene) were added to the samples. Then the samples were extracted with
methylene chloride in a Soxhlet extractor. Potential interferences by
pigments, lipids and other macromolecules were removed by use of the USEPA gel
permeation chromatography (GPC) method 3540 (USEPA 1986a). Then the extracts
were exchanged into hexane and analyzed with the USEPA Gas Chromatography/Mass
Spectrometry (GC/MS) method 8270 (USEPA 1986a).
Aroclors quantified were 1016, 1221, 1232, 1242, 1248, 1254 and 1260 for
sediments from Indiana Harbor, Buffalo River and Saginaw River (1); and 1242,
1254 and 1260 for Saginaw River (3). Aroclors were extracted from the
sediment samples according to the USEPA method 3550 (USEPA 1986a). The GC
surrogate compound dibutyl chlorendate (DBC) was added to the samples, and the
samples were subsequently extracted with methylene chloride using sonication.
Potential interferences by oily-type materials from highly contaminated
sediments, lipids, and other macromolecules were eliminated by use of GPC or
alumina column chromatography (USEPA 1986a, methods 3540 and 3610). Aroclors
were quantified by USEPA method 8080 (1986a) using a DB-5 fused silica
capillary column (0.25 mm diameter x 30 m) and a Hewlett-Packard 5890 gas
chromatography equipped with an electron capture detector (GC/ECD) and a
computer for data acquisition. A dual column analysis was always performed
simultaneously and results from both columns were accepted if they showed no
more than 50% variation.
The USEPA isotope dilution method 8290 (USEPA 1986a) was used to extract
and clean-up the sediment samples for analysis of PCDDs and PCDFs.
Isotopically labelled PCDDs and PCDFs were added to the samples before
extraction. The samples were extracted with benzene in a Soxhlet extractor
for 18 h. Then a three-step column chromatography procedure with acidified
silica gel, alumina, and AX-21 activated carbon on silica gel was used to
enrich the samples and remove interferences. Isotopically labelled 2,3,7,8,-
TCDD was added to the samples before the enrichment to determine the
efficiency of the method. Two internal standards were added to the samples
after sample enrichment to determine percentage recoveries. The PCDDs and
PCDFs were quantified with capillary column gas chromatography and high
resolution mass spectrometry. Concentrations were determined by selected-ion-
monitoring chromatography of groups of ion masses described in the USEPA
method 8290 (USEPA 1986a).
QUALITY CONTROL AND QUALITY ASSURANCE
The precision of the whole sediment toxicity tests were evaluated using
four replicate treatments, except for C. tentans tests, in which one midge
larva per replicate was used, for 10 to 15 replicates. The acceptability of
the toxicity tests was assessed by response of the test organisms to the
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control sediment for survival. Survival in the control sediment was within
acceptable ranges for all of the H. azteca tests, greater than 80% as
recommended by American Society for Testing and Materials (ASTM) (E 1383-90).
Midge control survival for ASTM test acceptability is 70% (E 1383-90), and was
met for all tests except the C. riparius Buffalo River sediment exposure
(60.5% survival, Table 4.18), and the C. tentans Saginaw River (3) sediment
exposure (60% survival, Table 4.35). The results from these two midge studies
should be interpreted with caution.
Test organism sensitivity was evaluated by exposing the organisms to
saline water in acute toxicity tests. These reference toxicant tests are
routinely conducted by NFCRC to monitor the sensitivity and condition of
laboratory test organisms. Reference toxicant tests were conducted with NFCRC
water and Instant OceanR sea salts (pH 8.0, salinity 33 g/L) . The 48-h LC50
for C. riparius was 9 g/L salinity and for C. tentans was 10 g/L salinity.
The 72-h LC50 for H. azteca was 17 g/L salinity.
Accuracy and precision of the chemical analyses were determined by
analysis of one blank, one matrix spike, one certified reference material, and
one sample in duplicate or triplicate for each set of 20 samples. Acceptable
recovery values ranged from 85 to 115% of the spike concentration for metals,
and from 70 to 130% of the spike concentration for organics and organometals.
Analytical values for reference materials were acceptable if they were within
20% of the certified ranges. The acceptable coefficient of variation for
duplicate or triplicate sample analyses was <20%.
During chemical analyses, three to five standards containing
concentrations that bracketed the expected range of concentrations in the
samples were used for daily instrument calibrations. In analyses of samples
for metals by atomic absorption spectrophotometry, these standards were
analyzed as matrix spikes, and the slopes from linear regression analyses were
used to estimate sample concentrations. The minimum acceptable r in the
regression analyses was 0.97. The standards for each sample set were analyzed
at the beginning and end of each analytical run. The analytical results were
accepted if the values for standards were within 90 to 110% of their certified
values. For some samples analyzed by atomic absorption, average response
factors, rather than linear regression, were used for instrument calibration.
The accuracy of this calibration method was checked by dividing each response
factor by the average response value. The calibration values were accepted if
they were within 5% of the average response value.
During chemical analyses, the method's detection limits (MDL) was
estimated according to procedures in the USEPA Federal Register (1984). The
MDLs were calculated as:
3ffn
where an is the standard deviation of the response factors of 3 or
4 calibration spikes or blanks (if applicable). Additional MDLs were
calculated as:
Student t0.oi,,,-i * °n
where "n is the standard deviation of the measured concentration of a standard
value within an order of magnitude of the absolute detection limit of the
instrument. If standard values were not available, 15 replicates of certified
reference materials were used to obtain °n, and to calculate non-matrix-
specific MDLs.
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RESULTS
EVALUATION OF SEDIMENT STORAGE USING MICROTOX TESTS
With few exceptions, the Microtox" test indicated that the toxicity of
sediment samples did not change during laboratory storage, for up to 26 d at
4°C. Indiana Harbor sediments were stored the longest over other AOCs, for 26
d. The remaining samples were stored between 15 and 22 days before the last
whole sediment tests were started. Only two of the 19 samples tested
exhibited different toxicity over time, see Table 2.14 in Chapter 2. After 14
days of storage, elutriates prepared from sample IH-01-07 (Indiana Harbor)
elicited increased toxicity, and elutriates prepared from sample BR-01-03
(Buffalo River) exhibited decreased toxicity. All other samples exhibited no
appreciable changes in toxicity as measured by Microtox toxicity.
INDIANA HARBOR
Physical and Chemical Characterization
Concentrations of TOC, total solids, and organometals in Indiana Harbor
sediment samples are listed in Table 4.2. Total organic carbon ranged from
8.8 to 12.2% in Stations IH-O1-05, IH-O1-06, IH-O1-07, IH-O1-08 and IH-O1-10
and in Stations IH-O1-03 and IH-O1-04 from 5.6 to 7.8%. Solids ranged from
19.6% in sediment from Station IH-O1-10 to 50.2% in sediment from Station IH-
O1-05. Organometals in Indiana Harbor sediment samples ranged from less than
0.1 ng/dry g for methylmercury to 1500 ng/dry g for tributyltin.
Metals in Indiana Harbor sediments ranged from 0.7 /Kj/dry g for Hg to
7960 /tig/dry g for Zn (Table 4.3). Concentrations of Cr, Cu, Fe, Mn, Pb, and
Zn were the highest, and the other metals ranged from less than 1.0 to 100
/ig/dry g (Table 4.3).
Acid volatile sulfides and eight SEM for Indiana Harbor stations were
analyzed (Table 4.4). Among the Indiana Harbor stations, Station IH-O1-07 was
the highest for extractable Cd, Cr, Fe, Mn, Ni, Pb and Zn (Table 4.4). The
AVS ranged from 16 /iM/dry g in Station IH-O1-04 to 71.4 jtM/dry g in Station
IH-O1-07, and all SEM/AVS ratios were less than one (Table 4.4).
The Indiana Harbor sediments consisted primarily of particles that were
less than 38 /wn in size (36.8% to 77.7%) with the remainder of the particles
ranging from .063 mm to 1.0 mm in size (Table 4.5). Particles greater than
1.0 mm in size comprised only a small portion (0.2% to 1.0%) of the sediments
from Indiana Harbor.
Sediments from Indiana Harbor contained PAH and semivolatile compound
concentrations that ranged from less than 25 ng/dry g for dimethylphthalate to
290,000 ng/dry g for 2-methylnaphthalene (Table 4.6). Station IH-01-07
contained fluorene, phenanthrene, anthracene, fluoranthene and pyrene (Table
4.6). Polychlorinated dibenzo-dioxins and -furans in the Indiana Harbor
samples ranged from 3.8 pg/dry g for 1,2,3,7,8-pentachlorodibenzofuran to
43,000 pg/dry g for octachlorodibenzodioxin (Table 4.7).
Concentrations of PCBs in the sediment from Indiana Harbor for Aroclor
1242 ranged from 3,000 (IH-01-03) to 43,000 ng/dry g (IH-01-07), and in
addition, Aroclor 1254 was detected in sediment from Station IH-01-04 (1,000
ng/dry g) (Table 4.8). Pesticides in the Indiana Harbor sediments were below
analytical detection limits (Table 4.9).
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Hvalella azteca Exposure
Overlying Water Quality Characterization—
In the Indiana Harbor whole sediment amphipod exposure, mean overlying
water dissolved oxygen, pH, and hardness were similar to in-flowing
reconstituted water described earlier. Unionized ammonia concentrations were
greater than 0.137 mg/L in all of the treatments, with the highest unionized
ammonia level (0.320 mg/L) in overlying water from Station IH-O1-06 (Table
4.10); unionized ammonia in the overlying water of control was 0.057 mg/L
(Table 4.10). Alkalinity in all of the treatments, including control, were1
about 1.5 times greater than the in-flowing reconstituted water (Table 4.10).
Dissolved oxygen was at acceptable levels (greater than 40% of saturation,
ASTM E 1383-90) in all treatments throughout the exposure. Chloride was
elevated 1.5 to two times in all treatments (except control) relative to the
reconstituted water (Table 4.10). Turbidity in all treatments was about five
to six times greater than control (Table 4.10).
Toxicity—
Amphipod survival was significantly reduced in 14-d exposures to whole
sediment samples from Stations IH-O1-03 and IH-O1-06, and no amphipods
survived in Stations IH-O1-04 and IH-O1-07 (Table 4.11). The sublethal
endpoints, number of antennal segments and percent sexually mature
(identifiable secondary sex characteristics) of surviving amphipods exposed to
Indiana Harbor sediments, were not significantly different after 14 days
(Table 4.11). Because Indiana Harbor sediments were lethal in 14-d exposures,
amphipod 28-d exposures were not conducted.
By Day 1, amphipods were observed floating on the water surface in all
Indiana Harbor stations (except the control) within the first 24 h of the 14-d
exposure, most notably in Station IH-O1-07. An oily film was present on the
overlying water surface in all stations, especially heavy in IH-O1-07. Two
feedings were withheld from Station IH-O1-07 because of sediment surface mold.
Indigenous species seen during amphipod recovery at test end included
oligochaetes and ostracods.
Chironomus riparius Exposure
Overlying Water Quality Characterization—
During the Indiana Harbor C. riparius exposure, overlying water pH,
dissolved oxygen, and hardness were similar to in-flowing reconstituted water
described earlier. Unionized ammonia concentrations were greater than 0.140
mg/L in all of the treatments, with the highest unionized ammonia level,
(0.320 mg/L) in overlying water from Station IH-O1-06 (Table 4.10). Unionized
ammonia in the control overlying water was 0.055 mg/L (Table 4.10).
Alkalinity in all of the treatments, including control, were about 1.5 times
greater than the in-flowing reconstituted water (Table 4.10). Dissolved
oxygen was at acceptable levels (greater than 40% of saturation, ASTM E 1383-
90) in all treatments throughout the exposure. Chloride was 1.5 to two times
greater in all of the treatments (except control) relative to the
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reconstituted water (Table 4.10). Turbidity in the treatments was about two
times greater than control (Table 4.10).
Toxicity—
All of the Indiana Harbor sediments were toxic to C. riparius (Table
4.11). Whereas 75% of the larvae survived in the control, survival in
sediment from Indiana Harbor was 0 to 2.0% (Table 4.11). Body length was not
measured for the limited number of survivors.
An oily film developed on the overlying water surface of all treatments.
Controls were fed algae and HartzR dog treats Cerophyl" throughout the study.
All test chambers containing sediment from Indiana Harbor stations were fed
only algae between Day 5 and the end of the test, because of surface sediment
mold. Oligochaetes, fingernail clams, and snails were found in the Indiana
Harbor sediments at the end of the study.
Chironomus tentans Exposure
Overlying Water Quality Characterization—
In the C. tentans whole sediment exposure, unionized ammonia
concentrations were greater than 0.292 mg/L in all of the treatments, with the
highest unionized ammonia level, (1.110 mg/L) in overlying water from Station
IH-O1-07 (Table 4.10). Alkalinity in all of the treatments, including
control, was about 1.5 to two times greater than the reconstituted water
(Table 4.10). Dissolved oxygen was at acceptable levels (greater than 40% of
saturation, ASTM E 1383-90) in all treatments throughout the exposure.
Chloride was 1.5 to two times greater in all of the treatments (including
control) relative to the reconstituted water (Table 4.10). Turbidity in all
treatments was about two to four times greater than control (Table 4.10).
Toxicity—
All C. tentans exposed to sediment from Stations IH-O1-06 and IH-O1-07
died within 10 days. Survival and growth (body length) of C. tentans larvae
exposed to sediment from Stations IH-O1-03 and IH-O1-04 were significantly
reduced compared to the control sediment (Table 4.11).
Black sediment surface mold required withholding food for two days in
the control sediment, one day in Stations IH-O1-G3 and IH-O1-04, and three
days in Stations IH-O1-06 and IH-O1-07. An oil sheen was observed on the
overlying water surface of the exposure chambers containing sediment from
Stations IH-O1-03, IH-O1-04 and IH-O1-07. Indigenous species were not
observed at the end of the C. tentana exposure.
BUFFALO RIVER
Physical and Chemical Characterization
Concentrations of TOC, total solids, and organometals in Buffalo River
sediments are listed in Table 4.12. With the exception of Station BR-01-01
that contained about 9.0% TOC, concentrations of TOC in the Buffalo River
sediments were similar (< 2.0%). Solids ranged from 30% in sediment from
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Station BR-01-01 to 74% in sediment from Station BR-01-02. Organometal
concentrations in the Buffalo River samples ranged from less than 0.05 ng/dry
g for methylmercury to 36 ng/dry g for tributyltin.
Metals in sediment from Station BR-01-01 were elevated over sediments
from all other Buffalo River sediment (Table 4.13). The Buffalo River samples
contained a broad range of metal concentrations (Table 4.13): Cr, Cu, Mn, Pb,
and Zn ranged from less than 1.0 /*g/dry g to 1386 fig/dry g; all other metals
were < 57 /tg/dry g. Concentrations of AVS and eight SEM for Buffalo River
stations were analyzed (Table 4.4). The AVS in sediments from Buffalo River
stations ranged from 1.0 /iM/dry g (BR-01-02) to 161 jtM/dry g (BR-01-01). All
SEM/AVS ratios were less than one (Table 4.4).
The distribution of sizes for sediment particles in the Buffalo River
sediments are shown in Table 4.14. With the exception of Station BR-01-02,
the Buffalo River sediments consisted primarily of particles that were less
0.038 mm in size (43.7% to 92.0%); Station BR-01-02 consisted mainly of
particles that ranged from 0.063 to 0.25 mm (58.5%) and from 0.25 to 1.0 mm
(38.5%) (Table 4.14). Particles greater than 1.0 mm in size comprised only a
small portion (0.1% to 14.0%) of the Buffalo River sediments (Table 4.14).
Concentrations of PAHs in Buffalo River sediments ranged from less than
18 ng/dry g for dichlorobenzene to 59000 ng/dry g for bis-(2-ethylhexyl)
phthalate (Table 4.15). Polychlorinated dibenzo-dioxins and -furans in the
Buffalo River samples ranged from less than the analytical detection limit up
to 12,000 pg/dry g for octachlorodibenzodioxin (Table 4.16). Aroclors and
pesticides in all samples from Buffalo River were below analytical detection
limits (Tables 4.8 and 4.9).
Station BR-01-01, located nearest to the end of the shipping canal, had
the most distinctive physical and chemical characteristics compared to the
other sediments collected at the Buffalo River study area. Sediment from
Station BR-01-01 had the lowest dry weight and a TOC value more than four
times higher than other stations. In addition, Station BR-01-01 had the
highest measured values for AVS (Table 4.4), methylmercury, tri-, di-, and
monobutyltin (Table 4.12), nine out of 12 metals (Table 4.14); 13 out of 20
semi-volatile organic compounds (Table 4.15); and 22 out of 25 PCDDs and PCDFs
(Table 4.16). Sediments from Station BR-01-07 and BR-01-08 had the lowest TOC
values of any Buffalo River sediments tested, and also had intermediate to
lower levels of AVS, methylmercury, and tri-, di- and monobutyltin, compared
to other sediments tested.
Hyalella azteca Exposures
Overlying Water Quality Characterization—
In the Buffalo River amphipod 14- and 28-d whole sediment exposures,
overlying water dissolved oxygen and pH were similar to in-flowing
reconstituted water described earlier. Unionized ammonia concentrations were
greater than 0.052 mg/L in all of the treatments, with the highest unionized
ammonia level, (0.117 mg/L) in overlying water from Station BR-01-09 (28-d
exposure) (Table 4.17). Unionized ammonia in control overlying water was
0.004 mg/L (Table 4.17). Alkalinity, in all of the treatments, including
control, was about 1.5 times greater (mean alkalinity for Station BR-01-09 was
three times greater) than the in-flowing reconstituted water (Table 4.17).
Overlying water hardness was elevated about 1.5 times for sediment from
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Station BR-01-09 over the reconstituted water (Table 4.17). Dissolved oxygen
was at acceptable levels (greater than 40% of saturation, ASTM E 1383-90) in
all treatments throughout the exposure. Chloride was 1.5 to two times greater
in all of the treatments (including control), relative to the reconstituted
water (Table 4.17). Turbidity in all treatments (except BR-01-07) of the 14-d
Hvalella azteca exposure was elevated about four to eight times relative to
control; Station BR-01-07 in the 14-d exposure was elevated about 21 times;
and Station BR-01-07 of the 28-d exposure, was elevated about four times
(Table 4.17).
Toxicity—
Amphipod 14-d survival in sediments from Station BR-01-03 was
significantly reduced. Additionally, amphipod 28-d survival was significantly
reduced in sediments from Stations BR-01-01, BR-01-03, BR-01-07 and BR-01-09
(Table 4.18). Amphipods exposed to sediments from Station BR-01-03 exhibited
the lowest survival for both 14-d (63.8%) and 28-d (75.0%) exposures. Body
length for surviving 14-d H. azteca was significantly reduced in exposures
from all stations, but body length was significantly reduced in 28 days only
in sediments from Stations BR-01-03 and BR-01-07 (Table 4.18). Similarly,
amphipods exposed to sediments from BR-01-03 and BR-01-07 in the 28-d exposure
exhibited significant reduction for increase in length (Figure 4.1). The
number of antennal segments in the 14-d control amphipods was significantly
reduced for all stations, except Station BR-01-09, but was the same between
Stations BR-01-01 and BR-01-08, and Stations BR-01-03 and BR-01-07. Amphipods
in the 28-d sediment exposure exhibited no significant differences for number
of antennal segments and had similar number of segments to the 14-d amphipods
(Table 4.18). Sexually mature amphipods for the 14-d exposure was
significantly different (6.1%) in sediments from all stations except BR-01-03
(4.2%), however, no significant reduction in percent sexually mature was
evident at the end of the 28-d exposure (Table 4.18).
Except for the control sediment, a film was present on the overlying
water surface in all Buffalo River sediments, in both the 14- and 28-d
exposures. In the 28-d exposure, at least one feeding was withheld from all
stations, except for BR-01-08, because of sediment surface mold. No amphipods
were observed during the 14- or 28-d exposures. Indigenous species observed
at the end of the 14- and 28-d tests included oligochaetes, ostracods, and one
leach in station BR1-07 of the 14-d test.
Chironomus riparius Exposure
Overlying Water Quality Characterization—
In the Buffalo River C. riparius whole sediment exposure, overlying
water dissolved oxygen and pH were similar to in-flowing reconstituted water
described earlier. Unionized ammonia concentrations were greater than 0.051
mg/L in all treatments, with the highest unionized ammonia level (0.118 mg/L)
in overlying water from Station BR-01-09 (Table 4.17). Unionized ammonia in
control overlying water was 0.004 mg/L (Table 4.17). Alkalinity, in all
treatments, including control, were about 1.5 times greater (about three times
greater in Station BR-01-09) than the reconstituted water (Table 4.17). Water
hardness was elevated about 1.5 times in sediment from Station BR-01-09 over
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the reconstituted water (Table 4.17). Dissolved oxygen was at acceptable
levels (greater than 40% of saturation, ASTM E 1383-90) in all treatments
throughout the exposure. Chloride was 1.5 to two times greater in all of the
treatments relative to the reconstituted water (Table 4.17). Turbidity in all
treatments was elevated about seven to 15 times relative to control (Table
4.17).
Toxicity—
Control survival (61%) was not within the limits of test acceptability
(70%) (ASTM E 1383-90) so C. riparius Buffalo River data should be interpreted
with caution. Survival of C. riparius was significantly reduced in sediments
from Stations BR-01-07 or BR-01-08 (Table 4.18). Survival did not differ
significantly in sediments from Stations BR-01-01, BR-01-03 and BR-01-09
(Table 4.18). Only C. riparius larvae exposed to Station BR-01-03 sediment
were significantly smaller than larvae exposed to the control and the other
sediments, except Station BR-01-08. Body length did not differ significantly
for C. riparius larvae exposed to the control or sediments from Stations BR-
01-01, BR-01-07, BR-01-08, and BR-01-09 (Table 4.18).
By Day 7 all test sediments exhibited surface mold which temporarily
required withholding Hartz dog treats. Beginning at Day 8, and continuing
until the end of the exposures, the amount of Hartz dog treats added to the
test chambers was decreased by 50%, to minimize development of sediment
surface mold. At the end of the exposures, snails, oligochaetes and several
unidentified midge larvae were found in the Buffalo River sediments.
Chironomus tentans Exposure
Overlying Water Quality Characterization—
In the Buffalo River C. tentans whole sediment exposure, overlying water
dissolved oxygen, and pH were similar to in-flowing reconstituted water
described earlier. Unionized ammonia concentrations were greater than 0.066
mg/L in all treatments, with the highest unionized ammonia level, (0.559 mg/L)
in overlying water from Station BR-01-01 (Table 4.17). Unionized ammonia in
the control overlying water was 0.547 mg/L (Table 4.17). Alkalinity, in all
of the treatments, including control, were about 1.5 times greater (about
three times greater in sediment from Station BR-01-09) relative to the
reconstituted water (Table 4.17). Overlying water hardness was elevated about
1.5 times for all treatments, except sediments from Station BR-01-03 over the
reconstituted water (Table 4.17). Dissolved oxygen was at acceptable levels
(greater than 40% of saturation, ASTM E 1383-90) in all treatments throughout
the exposure. Chloride was three to seven times greater in all of the
treatments relative to the reconstituted water (Table 4.17).
Toxicity—
Survival of C. tentans was not significantly reduced in the Buffalo
River test (Table 4.18). While larval dry weight was reduced in sediments
from Stations BR-01-07 and BR-01-08, no significant effect was observed in
sediments from Stations BR-01-01, BR-01-03 or BR-01-09 (Table 4.18).
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Sediment surface mold required withholding food for five days for the
control sediments and sediment from Station BR-01-01; six days for BR-01-03;
seven days for sediments from Stations BR-01-08 and BR-01-09; and eight days
for sediment from Station BR-01-07. Indigenous species were not observed when
test organisms were recovered at the end of the C. tentans exposure.
SAGINAW RIVER (1)
Physical and Chemical Characterization
Concentrations of TOC, total solids, and organometals in sediments from
the Saginaw River (1) are listed in Table 4.19. With the exception of
sediments from Station SR-01-10 that contained less than 1.0% TOC, TOC in
Saginaw River were similar and ranged from 2.0 to 3.8%. Solids ranged from
33.0% in sediment from Station SR-01-07 to 64.0% in sediment from Station SR-
01-10. Sediments contained low (ng/dry g) concentrations of organometals.
Saginaw River (1) sediments contained a broad range of metal
concentrations: Cr, Cu, Mn, Pb, and Zn ranged from 16.0 /tg/dry g for Cu in
Station SR-01-10 to 819 ng/dry g for Mn in Station SR-01-03 (Table 4.20).
With exception of Fe, that ranged from 1.2 to 3.2% in Saginaw River (1)
sediments, all other metals were < 157 ng/dry g (Table 4.20).
Concentrations of AVS and eight SEM for Saginaw River (1) sediments were
analyzed (Table 4.21). The AVS in Saginaw River (1) sediments ranged from 1.5
^iM/dry g in Station SR-01-10 to 15.5 /tM/dry g in Station SR-01-06. All
SEM/AVS ratios were less than one (Table 4.21).
Sediment samples from Saginaw River (1) consisted primarily of particles
that were less than 0.038 mm in size (19.4% to 79.3%), and particles that
ranged from 0.063 mm to 0.25 mm (10.4% to 45.9%). Particles greater than 1.0
mm in size comprised only 0.1% to 5.4% of the samples (Table 4.22).
Concentrations of PAHs and other semivolatile compounds in Saginaw River
(1) sediments ranged from less than the analytical detection limit of 4.0
ng/dry g for dichlorobenzene up to 13000 ng/dry g for bis(2-ethylhexyl)
phthalate (Table 4.23). Saginaw River (1) sediments contained PCDDs and PCDFs
that ranged from 1.7 pg/dry g for 1,2,3,4,7,8-hexachlorodibenzodioxin in
Station SR-01-10 to 22,000 pg/dry g for total tetrachlorodibenzofuran (TCDD)
in Station SR-01-04 (Table 4.24). Levels of PCBs measured in sediment ranged
from 300 ng/dry g (Arochlor 1254) to 60,000 ng/dry g (Aroclor 1232) in
sediment from Station SR-01-06 (Table 4.8). Pesticides in sediments from
Saginaw River (1) were below analytical detection limits (Table 4.9).
Hvalella azteca Exposures
Overlying Water Quality Characterization—
Amphipod 14- and 28-d exposures to Saginaw River (1) sediments,
overlying water dissolved oxygen, hardness, and pH were similar to in-flowing
reconstituted water described earlier. Unionized ammonia concentrations were
greater than 0.021 mg/L in all of the treatments, with the highest unionized
ammonia level, (0.064 mg/L) in overlying water from Station SR-01-06 for both
the 14- and 28-d exposures (Table 4.25). Unionized ammonia in the control
overlying water were 0.003 mg/L (14-d exposure) and 0.004 mg/L (28-d exposure)
(Table 4.25). Alkalinity, in all of the treatments except control, was
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elevated about two times compared to the reconstituted water (Table 4.25).
Alkalinity for the control overlying water was 48 mg/L in the 14-d exposure,
and 66 mg/L in the 28-d exposure (Table 4.25). Dissolved oxygen was at
acceptable levels (greater than 40% of saturation, ASTM E 1383-90) in all
treatments throughout the exposure. Chloride, in all of the treatments, was
elevated two times compared to the reconstituted water (Table 4.25).
Turbidity in all treatments was elevated about four to 10 times relative to
control (Table 4.25).
Toxicity—
Amphipod 14-d and 28-d survival in sediments from Station SR-01-06 was
significantly reduced. Survival of amphipods ranged from 92.5% in sediments
from Station SR-01-03 to 10.0% in sediments from Station SR-01-06 (Table
4.26). No significant differences were detected in the 14-d exposure
sublethal responses (body length, number of antennal segments and percent
mature), but was significantly reduced in sediment exposures from Station SR-
01-06 and Station SR-01-03 in the 28-d exposure (Table 4.26). Increase in 28-
d amphipod body length was significantly reduced in exposures from Stations
SR-01-03 and SR-01-06 sediments (Figure 4.1).
In the 28-d exposure, gravid females or amplexed pairs were observed in
the control and in Station SR-01-10, however no young were isolated at the end
of the test. By Day 1, amphipods were observed floating on the water surface
in Station SR-01-06 of the 28-d exposure. In the 14-d exposure, at least one
feeding was withheld from the control and Station SR-01-06 because of mold,
and a build-up of food on the sediment surface. Indigenous species observed
at the end of the 14- and 28-d tests included oligochaetes, ostracods, and
cyclops. In sediment from Station SR-01-06, only one oligochaete was
observed.
Chironomus riparius Exposure
Overlying Water Quality Characterization—
In the Saginaw River (1) C. riparius exposure, overlying water dissolved
oxygen, hardness, and pH were similar to in-flowing reconstituted water
described earlier. Unionized ammonia concentrations were greater than 0.021
mg/L in all of the treatments, with the highest unionized ammonia level (0.063
mg/L) in overlying water from Station SR-01-06 (Table 4.25). The unionized
ammonia concentration for the control overlying water was 0.003 mg/L (Table
4.25). Alkalinity, in all of the treatments except control, were elevated
about two times compared to the in-flowing reconstituted water (Table 4.25).
Alkalinity for the control overlying water was 34 mg/L about 0.5 times less
than the reconstituted water (Table 4.25). Dissolved oxygen was at acceptable
levels (greater than 40% of saturation, ASTM E 1383-90) in all treatments
throughout the exposure. Chloride, in all of the treatments, was elevated two
times compared to the reconstituted water (Table 4.25). Turbidity in all
treatments was elevated about three to six times compared to control (Table
4.25).
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Toxicity—
Both survival and body length were significantly reduced among larvae
exposed to sediment from Station SR-01-06 (Table 4.26). Survival and body
length of C. riparius in sediment exposures from Stations SR-01-03 and SR-01-
10 did not differ significantly (Table 4.26).
An oily surface water film was observed in all test chambers beginning
at Day 5. By Day 7, mold on the sediment surface required withholding
CerophylR in all treatments. Surface sediment conditions in test chambers
containing Station SR-01-06 and control sediment were generally clearer than
sediments from Stations SR-01-03 and SR-01-10. Additions of Hartz" dog treats
were withheld from Stations SR-01-03 and SR-01-10 during the last four days of
the exposure because of sediment surface mold. At the end of the exposures,
oligochaetes were observed in the sediments from Saginaw River (1).
Chironomus tentans Exposure
No exposures with Chironomus tentans were conducted with samples
collected from the Saginaw River (1) because of problems with the cultures.
SAGINAW RIVER (3)
Physical and Chemical Characterization
Concentrations of TOC, total solids, and organometals in sediments from
Saginaw River (3) are listed in Table 4.27. With the exception of the
sediment grab sample from Station SR-03-06 that contained 0.2% TOC, TOC in the
Saginaw River (3) grab samples ranged from 1.0% to 4.0%. Total organic carbon
in core samples from Saginaw River (3) ranged from 0.6% in Station SR-03-06-02
to 5.0% in Station SR-03-02-X2. Solids ranged from 47.3% to 76.4% in Saginaw
River (3) sediment grab samples and from 49.2% to 76.3% in sediment core
samples. The Saginaw River (3) grab samples contained low concentrations of
organometals that ranged from less than 0.1 ng/dry g for methylmercury to 10.0
ng/dry g for tributyltin.
Generally, the sediments from Saginaw River (3) contained a broad range
of low (/tg/dry g) metal concentrations. In grab samples, Cr, Cu, Mn, Pb, and
Zn ranged from 16.9 ng/dry g for Pb in Station SR-03-06 to 549 /ig/dry g for Mn
in Station SR-03-24; all other metals in the grab samples were < 92.9 /xg/dry g
(Table 4.28).
AVS in Saginaw River (3) grab samples ranged from 1.2 /*M/dry g in
Station SR-03-06 to 8.9 fiM/dry g in Station SR-03-24 (Table 4.21). The
estimated SEM/AVS ratios were all less than one (Table 4.21).
Sediments from Saginaw River (3) consisted primarily of particles that
were less 0.038 mm in size (2.3% to 54.9%) and particles that ranged from
0.063 mm to 0.25 mm (22.7% to 77.7%) (Table 4.29). Particles greater than 1.0
mm in size comprised only 0.1% to 1.4% of the samples (Table 4.29).
Generally, PAHs in the sediments from Saginaw River (3) were below
analytical detection limits, but concentrations of some PAHs were detected
(Table 4.30). For example, pyrene ranged from less than the detection limit
up to 1800 ng/dry g in grab samples and up to 2700 ng/dry g in core samples,
and phenanthrene ranged from less than detection limits up to 1000 ng/dry g in
grab samples (Table 4.30).
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The samples from Saginaw River (3) contained PCDDs that ranged from less
than detection limits up to 6400 pg/dry g of octachlorodibenzodioxin in grab
sample SR-03-08, and up to 74000 pg/dry g of heptachlorodibenzofuran in core
sample SR-03-02-X2 (Table 4.31). Total dioxins ranged from 5.7 pg/dry g in
sediment from Station SR-03-06 to 230 pg/dry g in sediment from Station SR-03-
08; and total furans 230 pg/dry g in sediments from Station SR-03-01 (first
replicate) to 3100 pg/dry g in sediments from Station SR-03-06 (Table 4.31).
With the exception of Endrin ketone, which ranged from 2.0 to 380 ng/dry
g, pesticides in sediments from the Saginaw River (3) were below analytical
detection limits or present at low ng/dry g concentrations (Table 4.32). In
Saginaw River (3) sediment, Aroclor 1260 was generally below detection limits;
Aroclor 1254 ranged from less than detection limits up to 95 ng/dry g in grab
samples and up to 8100 ng/dry g in core samples; and Aroclor 1242 ranged from
less than detection limits up to 470 ng/dry g in grab samples and up to 79,000
ng/dry g in core samples (Table 4.33).
Hvalella azteca Exposures
Overlying Water Quality Characterization—
In the Saginaw River (3) amphipod 14- and 28-d whole sediment exposures,
overlying water dissolved oxygen, hardness, alkalinity, pH, and chloride were
similar to in-flowing reconstituted water described earlier. Unionized
ammonia concentrations were greater than 0.019 mg/L in all treatments, with
the highest unionized ammonia level (0.118 mg/L, 14-d exposure; and 0.101
mg/L; 28-d exposure) in overlying water from Station SR-03-24 (Table 4.34).
Unionized ammonia for the control overlying water was 0.005 mg/L (14-d
exposure) and 0.004 mg/L (28-d exposure) (Table 4.34). Dissolved oxygen was
at acceptable levels (greater than 40 percent of saturation, ASTM E 1383-90)
in all treatments throughout the exposure. Turbidity in all treatments,
except Station SR-03-01, was elevated about three to eight times compared to
control, but turbidity in SR-03-01 was similar to control (Table 4.34).
Toxicity—
Amphipod survival, body length and number of antennal segments were not
significantly reduced in 14-d or 28-d exposures, for all Saginaw River (3)
stations (Table 4.35). However, in the 28-d exposure, increase in amphipod
body length was reduced significantly in sediment exposures from Stations SR-
03-06 and SR-03-08 (Figure 4.1). Amphipods exposed to sediment from Station
SR-03-05 exhibited delayed sexual maturation in the 14-d exposure.
In both the 14- and 28-d exposures, mold was present on the sediment
surface in all treatments, except Station SR-03-01. A film was present on the
water surface of all treatments in the 28-d exposure, and in all treatments,
except the control and Station SR-03-01, of the 14-d exposure. One feeding
was withheld from the control treatment in the 28-d exposure due food build-up
on the sediment surface. Gravid females were observed in sediments from
Stations SR-03-16 and SR-03-24 at the end of the 14-d exposure, however, no
young were isolated. Indigenous species observed at the end of the 14- and
28-d tests were oligochaetes, most notably in Station SR-03-01.
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Chironomus riparius Exposure
Overlying Water Quality Characterization—
In the Saginaw River (3) whole sediment C. riparius exposure, overlying
water dissolved oxygen, hardness, alkalinity, pH, and chloride were similar to
in-flowing reconstituted water described earlier. Unionized ammonia
concentrations were greater than 0.015 mg/L in all of the treatments, with the
highest unionized ammonia level (0.101 mg/L) in overlying water from Station
SR-03-24 (Table 4.34). Unionized ammonia in the control overlying water was
0.004 mg/L (Table 4.34). Dissolved oxygen was at acceptable levels (greater
than 40% of saturation, ASTM E 1383-90) in all treatments throughout the
exposure. Turbidity in all treatments, except Station SR-03-01, was elevated
about three to five times compared to control, but turbidity in the sediment
exposure from Station SR-03-01 was similar to control.
Toxicity—
Survival of C. riparius larvae was not significantly different in
sediments for all stations from Saginaw River (3) (Table 4.35). Larval body
length exposed to sediments from all Saginaw River (3) stations was
significantly greater than that of C. riparius larvae exposed to the control
sediment. For C. riparius. body length ranged from 10.1 mm for the control to
12.5 mm for Stations SR-03-02 and SR-03-16 (Table 4.35).
All treatments were fed Cerophyl , Hartz dog treats and algae for the
first six days of the test. Beginning Day 7, additions of HartzR dog treats
were withheld from four of the seven Saginaw River (3) sediments. All feeding
was withheld on Day 10 and aeration started in all treatments, due to a
combination of low dissolved oxygen and the presence of black mold on the
surface of the sediments. Aeration was continued until the end of the test.
Surface mold disappeared after aeration started. Oligochaetes and snails were
found in the Saginaw River (3) sediments at the end of the exposures.
Chironomus tentans Exposure
Overlying Water Quality Characterization—
In the Saginaw River (3) C. tentans exposure, overlying water dissolved
oxygen, hardness, and pH, were similar to reconstituted water described
earlier. Unionized ammonia concentrations were greater than 0.074 mg/L in all
treatments, with the highest unionized ammonia level (0.886 mg/L) in overlying
water from Station SR-03-01 (Table 4.34). Unionized ammonia in the control
overlying water was 0.376 mg/L (Table 4.34). Dissolved oxygen was at
acceptable levels (greater than 40% of saturation, ASTM E 1383-90) in all
treatments throughout the exposure. Chloride, in all treatments except
Station SR-03-01, were elevated about two times compared to the reconstituted
water (Table 4.34). Turbidity in all treatments (except Station SR-03-01) was
elevated about three times compared to control, but turbidity in the sediment
exposure from Station SR-03-01 was similar to control.
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Toxicity—
Survival of C. riparius in the control (60%) was less than the
recommended limit for test acceptability (70%) (ASTM E 1383-90), therefore,
Saginaw River (3) whole sediment exposure results should be interpreted with
caution. Survival of C. tentans exposed to Saginaw River (3) stations ranged
from 0 to 80% (Table 4.35). Samples were grouped into two general classes,
those exhibiting survival greater than the control (SR-03-05, SR-03-08 and SR-
03-24) and those with survival less than the control (SR-03-01, SR-03-02, and
SR-03-06, and SR-03-16). No C. tentans survived in Station SR-03-06 and was
the only treatment significantly different from control (Table 4.36). Due to
difficulties controlling the air flow during these exposures, several chambers
in the study had increased overlying water evaporation. In three treatments:
control, SR-03-01 and SR-03-08, overlying water evaporated resulting in the
loss of test organisms.
Except in the control, mold developed by Day 5 on the surface of all
Saginaw River (3) sediments. Sediment surface mold required withholding food
for the last five test days in all Saginaw River (3) sediments. Indigenous
species were not observed at the end of the test.
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DISCUSSION
INDIANA HARBOR
Overlying Water Quality Characteristics
Overlying water quality for the amphipod and midge exposures differed
from the in-flowing reconstituted water in unionized ammonia, chloride,
alkalinity and turbidity. Chloride levels in the overlying water were 1.5 and
two times greater in all treatments compared to control. Culture water
chloride levels are about 20 mg/L (Kemble et al. 1992) indicating that the
two-fold chloride increase in the Indiana Harbor exposures would probably not
be directly toxic to amphipods or midges.
Unionized ammonia concentrations in the overlying water were > 0.137
mg/L which is above the 4-d unionized ammonia criterion of 0.035 mg/L at pH
8.0 and 20°C (USEPA 1986b). However, unionized ammonia concentrations of
about 1 mg/L were not toxic to either amphipods or C. tentans in 10-d water
only exposures (G. Ankley, USEPA, Duluth, MN, personal communication).
Unionized ammonia, in treatment IH-O1-07, of the 10-d C. tentans exposure, was
1.110 mg/L, high enough that it may have contributed to the observed toxicity.
However, amphipod and C. riparius exposures had complete mortality in the IH-
01-07 sediment exposure, despite having lower unionized ammonia. Unionized
ammonia levels in the overlying water of the C. tentans static exposure were
higher than unionized ammonia levels in the amphipod and C. riparius
treatments. This is most likely because the amphipods and C. riparius were
exposed in a flow-through system while the C. tentans exposure was in a static
system. In addition, higher test temperature in the C. tentans exposure (C.
tentans 23°C; H. azteca or C. riparius 20°C) may also have caused the increase
in unionized ammonia levels (Thurston et al. 1974).
Toxicitv Tests
The reductions in survival of amphipods, C. riparius or C. tentans.
exposed to whole sediments from Indiana Harbor, indicate that the quality of
sediments is not adequate to sustain populations of the test organisms or
organisms of equal or greater sensitivity. Higher trophic level organisms
which exploit midge larvae or amphipods as food items would likely be reduced
within the reaches of the Indiana Harbor sediments sampled for these
exposures, due to the probable absence of these benthic organisms.
Because all of the sediments from Indiana Harbor elicited severe effects
on test organism survival, spatial differences in sediment contamination were
difficult to identify. Observed effects on survival could not be attributed
to any specific contaminant measured in the Indiana Harbor sediments (Figures
4.2 and 4.3). The presence of concentrations of a broad range of contaminants
in the sediments provides additional evidence that poor sediment quality
exists in Indiana Harbor. However, because the sediments had a complex
mixture of contaminants, interactive toxicity (i.e., additivity, synergism, or
antagonism) (Marking 1985) may have occurred during the exposures.
Given the extreme spatial differences in sediment toxicity reported even
in small study areas (Stemmer et al. 1990), combined with the limited number
stations tested in the present study, it is difficult to determine if a
gradient of toxicity exists at Indiana Harbor. However, C. tentans test
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results suggest a trend. Samples collected from Indiana Harbor (Stations IH-
O1-03 and IH-O1-04) exhibited less toxicity compared to samples collected from
the main channel (Station IH-O1-06) or at the junction of the Lake George and
Grand Calumet Branch (Station IH-O1-07). Sediments from the two up-channel
locations had elevated metals and organic compound concentrations compared to
the other Indiana Harbor stations.
Sediment from Station IH-01-07, despite having an intermediate TOC
level, compared to other Indiana Harbor sediments, generally contained the
highest concentration of organic contaminants (Table 4.37). Station IH-O1-07
also had the highest measured metal concentrations (Tables 4.3 and 4.4). This
chemical profile suggests that Station IH-O1-07, located at the convergence of
the Grand Calumet Branch and the Lake George Branch, may be accumulating
contaminants from both systems.
Together, Stations IH-O1-06 and IH-O1-07 had the highest concentrations
of 16 out of 20 measured PAH. Of the 25 PCDDs and PCDFs measured, Stations
IH-O1-06 and IH-O1-07 accounted for 23 of the highest measurements.
Interestingly, Station IH-O1-03 (collected from Indiana Harbor proper) which
had the lowest toxicity to C. tentans, also contained the highest
concentrations of the toxic 2378 TCDD isomer (Helder 1981, Walker and Peterson
1991). These findings suggest that either 2378 TCDD is not toxic to C.
tentans larvae at these concentrations, or that it was not bioavailable under
the conditions tested.
BUFFALO RIVER
Overlying Water Quality Characteristics
Overlying water quality for the amphipod and midge Buffalo River whole
sediment exposures were similar to the in-flowing reconstituted water (except
unionized ammonia, and chloride). Culture water chloride levels are about 20
mg/L (Kemble et al. 1992) indicating that the two-fold chloride increase in
the Indiana Harbor exposures would probably not be directly toxic to amphipods
or midges. Survival of C. tentans was not adversely affected even with a
four-fold increase in chloride levels. Unionized ammonia concentrations in
the overlying water in all treatments, except control, were > 0.051 mg/L which
were above the 4-d unionized ammonia criterion of 0.035 mg/L at pH 8.0 and
20°C (USEPA 1986b). However, unionized ammonia concentrations of about 1 mg/L
were not toxic to either amphipods or C. tentans in 10-d water only exposures
(G. Ankley, USEPA, Duluth, MN, personal communication). Unionized ammonia
levels in the C. tentans exposure were higher than in the amphipod and C.
riparius treatments. This is most likely because the amphipods and C.
riparius were exposed in a flow-through system while the C. tentans exposure
was in a static system. However, higher test temperature in the C. tentans
exposure (C. tentans 23°C; H. azteca or C. riparius 20°C) may also have caused
the increase in unionized ammonia levels (Thurston et al. 1974).
Toxicitv Tests
Buffalo River sediment exposures were not consistently toxic to
amphipods or C. riparius. Amphipods identified toxicity (i.e., reduced
survival) in four of the five stations, whereas C. riparius identified two of
the five stations (Table 4.18). Amphipods were more sensitive than the midge
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in identifying toxicity in sediment from Buffalo River stations. In addition,
the results imply that differences in the spatial distribution of contaminant
loads existing in the Buffalo River stations may adversely influence survival
of H. azteca or C. riparius populations, or organisms of equal or greater
sensitivity. This implication is supported by the broad range in
concentrations of heavy metals, PAHs, and dioxins that were measured in
Buffalo River sediments (Tables 4.4, 4.12, 4.15 and 4.16). Differences in the
spatial distribution of contaminants can result from differences in
contaminant loading patterns, differences in contaminant cycling processes, or
contaminant transport (Burton 1991).
Of the five sediment samples tested, metal concentrations where higher
in stations closer the mouth of the river (Stations BR-01-01 and BR-01-02)
compared to stations sampled further up-stream (Stations BR-01-07, BR-01-08,
BR-01-09). Organic contaminants measured in the sediments did not follow the
trend of progressively higher concentrations near the mouth of the river as
seen with metals. While organic contaminants increased between Station BR-01-
09 and BR-01-07, in many cases concentrations of organic contaminants were
lower in Station BR-01-08 compared to Station BR-01-09. Growth effects were
not seen in sediments from Station BR-01-01 for midge or amphipods (28-d),
despite the higher levels of metal and organic contaminants found compared to
Station BR-01-08, in which amphipods (14-d) and C. tentans larvae showed
significant growth effects (Table 4.18). The high TOG found in sediment from
Station BR-01-01 may have minimized the availability of the PAHs, dioxins, and
furans and reduced their toxicity (Table 4.37).
The adverse effects of Buffalo River sediment on amphipod (14- and 28-d)
suggest that contaminants were bioavailable, but that a longer exposure (28-d)
was necessary to demonstrate effects on survival. In contrast though, the 28-
d exposure produced significant differences in growth (body length) at two
stations, while growth effects (body length) were observed at all stations in
the 14-d exposure (Table 4.18), which constituted a 50% reduction in number of
stations producing effects in 28 days. Percentage sexual maturation was
significantly different at four stations at 14-d, but by 28 days all amphipods
were sexually mature and not significantly different.
The amphipod Buffalo River whole sediment exposures demonstrate the need
to conduct both short- and long-term exposures to measure both lethal and
sublethal effects, as the information obtained is not always predictable from
one exposure or one endpoint. Disproportionate survival from the 14- to 28-d
exposure can be expected if the sediment-sorbed contaminants are slow to
accumulate in organisms (Landrum 1988), produce delayed effects at the site of
action (Landrum et al. 1985), or exhibit slow desorption kinetics (i.e.,
increased equilibration time between the contaminant and sediment) (Kenega and
Goring 1980, Karickhoff 1981, Landrum and Robbins 1990). For a better
understanding of the desorption kinetics of non-labile contaminants and its
relationship to bioavailability, rate limiting factors of sediment-sorbed
contaminants needs further investigation. But, if volatile contaminants are
lost from the test system or toxic compounds degrade, increased survival may
be expected in the 28-d exposures, in which case further investigation is
needed to assess the fate of the labile and degradable compounds in such
sediment test systems.
Effects of Buffalo River whole sediment on the survival and body length
of C. riparius were less severe than the effects elicited by Indiana Harbor
sediments; however, the Buffalo River results must be cautiously interpreted
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because survival of C. riparius (60.5%) in the control was less than the 70%
recommended by ASTM E1383-90 (1990). These effects of Buffalo River sediments
on C. riparius were consistent with overall lower measured concentrations of
heavy metals, PAHs, and dioxins in the Buffalo River sediments compared to
Indiana Harbor.
The Buffalo River sediments did not elicit uniform toxicity to C.
riparius. Survival of C. riparius from Stations BR-01-01, BR-01-03 and BR-01-
09, were similar to control. Sediments from Stations BR-01-07 and BR-01-08
elicited effects on survival similar to sediments from Stations BR-01-01 and
BR-01-03, but were significantly different from the control and the sediment
from Station BR-01-09 (Table 4.18). The only significant effect elicited by
exposure of C. tentans to Buffalo River sediments was the decreased weight of
organisms exposed to sediments from Station BR-01-07 and BR-01-08. Otherwise,
Buffalo River contaminated sediments were not toxic to C. tentans.
Exposure of C. tentans to sediment collected from Station BR-01-01 did
not result in significant decreases in survival or length. Two hypotheses may
be postulated to explain the lack of effects noted during the 10 day C.
tentans exposure to sediments from Station BR-01-01. The high TOG found in
sediment from Station BR-01-01 may have minimized the availability of the
PAHs, dioxins, and furans and reduced their toxicity (Table 4.37). The other
possibility is that the duration of the test (10-d) was insufficient for
contaminants present to produce toxic effects.
Withholding food to prevent additional growth of mold on the sediment
surface during the C. tentans exposures may have contributed to the decrease
in organism length in sediments from Stations BR-01-07 and BR-01-08. During
the 10 day exposures, food was withheld from chambers containing sediments
from Station BR-01-07 a total of eight days, and larvae exposed to sediments
from Station BR-01-08 were not fed a total of seven days. In comparison,
organisms exposed to sediment collected from Stations BR-01-01, BR-01-03 and
the control sediment were not fed on four to five days during the exposure
period. However, results of the C. tentans test conducted with sediment
collected from Station BR-01-09 with similar physical and chemical
characteristics contradicts the possibility of a feeding effect. Similar to
tests with sediment from Stations BR-01-07 and BR-01-08, the larvae exposed to
sediment from Station BR-01-09 were not fed for seven days but showed no
decrease in length by Day 10.
SAGINAW RIVER (1) and (3)
Overlying Water Quality Characteristics
Overlying water quality in the amphipod and midge exposures to Saginaw
River sediment were similar to the in-flowing reconstituted water (except
unionized ammonia for all exposures). Unionized ammonia concentrations for H.
azteca and C. riparius in the whole sediment overlying water were > 0.040 mg/L
in all treatments (except treatments SR-03-01, SR-03-06, and control), which
is above the 4-d unionized ammonia criterion of 0.035 mg/L at pH 8.0, and 20°C
(USEPA 1986b). Unionized ammonia concentrations in the 10-d C. tentans
exposures were > 0.074 mg/L, which is above the 4-d unionized ammonia
concentration of 0.035 mg/L at pH 8.0 and 20°C (USEPA 1986b). Unionized
ammonia levels in the C. tentans exposures were higher than unionized ammonia
levels in the amphipod and.C. riparius exposures. This is most likely because
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the amphipod and C. riparius exposures were in a flow-through system while the
C. tentans exposure was in a static system. In addition, higher temperature
in the C. tentans exposure (C. tentans 23°C; H. azteca or C. riparius 20°C)
may also have caused the increase in unionized ammonia levels (Thurston et al.
1974). However, unionized ammonia concentrations of about 1 mg/L were not
toxic to either amphipods and C. tentans in 10-d water only exposures (G.
Ankley, USEPA, Duluth, MN, personal communication).
Although dissolved oxygen in the 10-d C. tentans exposure was in the
acceptable range (greater than 40% saturation, ASTM E 1383-90), problems with
aeration were common (either no air flow or to much air flow to a tube). This
may have caused dissolved oxygen concentrations to drop below 40% saturation
during the exposure or caused supersaturation in some of the tubes. Rapid air
flow in one case (treatment SR-03-08) bubbled one of the exposure tubes dry.
Toxicitv Tests
Test results with whole sediment exposures from Saginaw River indicate
that sediment quality in Saginaw River may be more suitable for sustaining H.
azteca. C^ riparius and C. tentans populations than the sediment quality in
Indiana Harbor and Buffalo River. Only the sediment from Station SR-01-06 in
Saginaw River (1) elicited significant reductions in the survival and growth
for both amphipods and C. riparius. One sediment exposure from Saginaw River
(3) adversely effected growth (C. tentans. Station SR-03-06), and one sediment
effected amphipod sexual maturation (Tables 4.26 and 4.35). These results
were consistent with the generally higher measured concentrations of heavy
metals, methylmercury, PAHs, and dioxins in the sediment from Station SR-01-06
compared to the other sediments from Saginaw River. Also, these results
suggest a difference in the spatiotemporal distribution of toxicity and
contamination in Saginaw River sediments that may influence the distribution
and abundance of test organism populations as well as indigenous organisms.
Sediment collected from Station SR-03-06, which resulted in complete mortality
of exposed C. tentans. had elevated concentrations of PCBs and some tetra- and
penta-furans, compared to other stations.
Amphipod survival in Saginaw River (1) exposures was reduced
significantly in sediment from Station SR-01-06 in both 14- and 28-d tests.
Additionally, effects on body length, antennal segment number and sexual
maturation were observed at 28 days (Table 4.26). Body length at 28 days was
the most sensitive amphipod sublethal endpoint, since it detected toxicity for
all Saginaw River (1) sediments, whereas number of antennal segments and
sexual maturation could be predicted from reduced survival (Table 4.26).
Sediments from Saginaw River (3) were not lethal to amphipods at 14- or 28-d
exposures, and none of the sublethal endpoints added further sensitivity, with
the exception of one station (SR-03-05) at 14-d for percentage sexual
maturation (Table 4.35).
Samples collected from Saginaw River (1) were not evaluated using C.
tentans whole sediment tests. Larvae were not available at the time tests
were started.
The validity of the biological response of C. tentans exposed to
sediments collected from the Saginaw River (3) is uncertain. Control survival
was less than the test acceptability criteria at 70% (ASTM E 1383-90). Low
control survival may indicate that the cohort of test organisms used in the
exposures may have been unhealthy or subjected to some environmental or
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handling stress which decreased their vitality. In addition to low control
survival, technical problems with regulating air flow to the chambers resulted
in excessive evaporation of three chambers and the need for unusually high
water replacement in others. Lastly, due to an attempt to test all grab
samples, the number of replicates was reduced from 15 to 10. Exposing fewer
individuals enhanced the importance of mortalities in the controls.
COMPARISONS BETWEEN AREAS OF CONCERN AND SPECIES SENSITIVITIES
As mentioned earlier, a broad range of contaminant concentrations were
measured in the sediment samples from each AOC. But because of the potential
for these contaminated sediments to elicit toxicity as a result of contaminant
interactions, the responses observed in the whole sediment tests cannot be
attributed to any specific sediment-associated contaminant or component (i.e.,
particle size, TOC, ammonia). The survival of each species in the whole
sediment tests was inversely related to the sum of metal and PAH
concentrations in sediments from all stations across AOCs (Figures 4.2 and
4.3). The total metals, associated with high mortality, was less than 5000
ing/dry g for all test species. The total PAHs, associated with high
mortality, was about 50 fig/dry g for H_._ azteca and C. riparius. and about 200
ng/dry g for C. tentans (Figures 4.2 and 4.3).
Understanding the complex interactions of factors that control the
bioavailability of sediment-associated organic contaminants is fundamental to
evaluating the hazards of contaminated sediments with any degree of
certainty. Factors affecting the bioavailability of sediment-sorbed organic
compounds, and subsequently, the toxicity that these compounds may elicit
include: (1) the chemical characteristics of the contaminants, for example,
hydrophobicity, (2) sediment composition and characteristics, for example,
total organic carbon and particle size (Forstner 1990, Landrum and Robbins
1990, DiToro et al. 1991), and (3) the behavior and physiological
characteristics of the test organisms, for example, burrowing activity,
feeding, waste elimination (Riedel et al. 1987, Landrum and Robbins 1990). In
the present studies, many of the organic contaminants were hydrophobic, and as
such, their bioavailability may be regulated by sediment characteristics, such
as organic carbon or fine grain sediment particles. To evaluate organic
carbon interactions, concentrations of low and high molecular weight PAHs and
total PCBs, were normalized with percentage organic carbon (Table 4.37), then
compared with recommended threshold concentrations (Table 4.38).
Concentrations of low and high molecular weight PAHs and total PCBs, on a dry
weight basis, were also compared to recommended threshold concentrations
(Tables 4.37 and 4.38).
Concentrations of total PAHs (ng/dry g) and total PCBs (ng/dry g)
exceeded the Effects Range-Median (ER-M), estimates of the concentrations at
or above which effects were often detected at the 50th percentile (Long and
Morgan 1990), in all Indiana Harbor sediments (Table 4.38), whereas all
Buffalo River and Saginaw River (1) sediments only exceeded the PCB threshold,
and one sediment (Station BR-01-01) exceeded the total PAH threshold (Table
4.38). The reported total PAH Effects Range-Low (ER-L), an approximation of
the concentration at which adverse effects were first detected at the 10th
percentile, and ER-M, in Long and Morgan (1990), has a low subjective degree
of confidence, and has a moderate degree of confidence for total PCBs.
Compared to the Long and Morgan (1990) ER-M reported concentrations, the
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recommended threshold values reported by Barrick et al. (1988), based on
amphipod Apparent Effects Threshold (AET) concentrations (Table 4.38),
identified fewer sediments (Stations IH-01-06 and -07) as elevated for both
low and high molecular weight PAHs; and only one sediment (IH-01-03) was
identified as elevated for the high molecular weight PAHs (Tables 4.37 and
4.38). The amphipod AET (Barrick et al. 1988) identified all Indiana Harbor,
Buffalo River and Saginaw River (1) sediments as elevated for total PCBs,
similar to Long and Morgan (1990) (Tables 4.37 and 4.38).
Recommended threshold values from Puget Sound (SMS 1991), established by
AET and equilibrium partitioning methods, were contrasted with sediment
concentrations that were normalized for TOC (Table 4.38). Similar to amphipod
AET thresholds (Barrick et al. 1988) in Tables 4.37 and 4.38, the SMS (1991)
thresholds for PAHs, identified three Indiana Harbor sediments as elevated
(IH-01-03, -06, and -07). Similar to the AET thresholds reported in Barrick
et al. (1988), concentrations of total PCBs exceeded the SMS (1991)
recommended threshold concentrations in all Indiana Harbor, Buffalo River and
Saginaw River (1) sediments, but the SMS (1991) also identified Saginaw River
(3) sediments (SR-03-05, -06, and -08) as elevated for PCBs (Tables 4.37 and
4.38).
Single-compound tests with different TOC concentrations are providing
evidence that TOC and contaminant bioavailability are related. Measurement of
contaminant concentrations in the sediment pore water (DiToro et al. 1991),
rather than in the bulk sediment as in the present study, may better reflect
contaminant concentrations that the benthic test organisms encounter. This
does not imply that pore water or bulk sediment contaminant concentrations
normalized to TOC, are the only, or primary routes of exposure. All exposure
pathways are equally capable of contributing to the chemical activity of a
compound in an equilibrium analysis (DiToro et al. 1991). Further
investigations are needed to evaluate the influence of TOC on organic
contaminant bioavailability in sediments containing complex mixtures of
contaminants. Further, the relation between recommended threshold
concentrations for organic contaminants normalized to TOC or those evaluated
on a dry weight basis needs to be established.
Metal concentrations (jug/dry g) measured in the sediments tested in the
present study (Tables 4.3, 4.13, 4.20 and 4.28) were compared to recommended
threshold concentrations (Table 4.39). Metal concentrations (pig/dry g) that
exceeded recommended threshold concentrations (not AVS normalized), reported
for Puget Sound (SMS 1991), also exceeded the other reported thresholds, with
the exception of the ER-M value (Long and Morgan 1990) for As and Cd (Table
4.39). All the Indiana Harbor sediments contained at least three metals at
concentrations that exceeded the SMS (1991) and the Long and Morgan (1990)
thresholds, and sediments from Stations BR-01-01, SR-01-06 and SR-03-16
contained two metals that exceeded (Table 4.39). Although metal
concentrations (Tables 4.3, 4.13, 4.20 and 4.28) exceeded recommended
thresholds (Table 4.39), no relation between AVS and SEM were apparent. All
SEM/AVS ratios were less than one (Tables 4.4 and 4.21), indicating that the
divalent SEM may not have been bioavailable (DiToro et al. 1990, Carlson et
al. 1991). Measuring metals in sediments that are simultaneously released
during the AVS extraction is a more reasonable extraction procedure rather
than the strong acid total metal extraction procedure, because it provides a
better estimate of bioavailable metal concentrations (DiToro et al. 1990).
Additionally, the metal concentrations in the sediment can be normalized to
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AVS (DiToro et al. 1990). However, additional studies are needed with other
metals and complex mixtures of metals to further delineate the influence of
SEM/AVS on metal bioavailability in sediments (Carlson et al. 1991).
Sediment contaminants that exceed recommended threshold concentrations
are predicted to have an adverse effect on aquatic resources. Comparisons of
recommended threshold concentrations and test organism responses (i.e.,
significant reductions in survival or growth, Tables 4.11, 4.18, 4.26 and
4.35) are listed in Table 4.40. Three situations were observed in comparing
the recommended threshold concentrations to observed responses of the test
organisms. The first situation was designated as a positive hit ( + ) where
sediment concentrations exceeded recommended thresholds and toxicity occurred,
or sediment concentrations did not exceed thresholds and no toxicity occurred
(true positive and negative). The second situation was designated as a
negative hit (-) where sediment concentrations did not exceed thresholds and
toxicity occurred (false negative). The third situation was designated as a
zero hit (0) where sediment concentrations did exceed thresholds and no
toxicity occurred (false positive). The reported recommended thresholds were
ranked according to increasing ability to predict, or not to predict, the test
organism responses in the present study.
For positive hits (+) (thresholds were predictive):
Metals — SMS (1991) > Long and Morgan (1990);
PAHs — Long and Morgan (1990) > SMS (1991) = Barrick et al. (1988);
PCBs — Barrick et al. (1988) > SMS (1991) > Long and Morgan (1990).
For negative hits (-) (thresholds not predictive, false negatives):
Metals — Long and Morgan (1990) > SMS (1991);
PAHs — SMS (1991) = Barrick et al. (1988) > Long and Morgan (1990);
PCBs — Long and Morgan (1990) = Barrick et al. (1988) > SMS (1991).
For zero hits (0) (thresholds not predictive, false positives):
Metals — SMS (1991) = Long and Morgan (1990);
PAHs — SMS (1991) = Long and Morgan (1990) = Barrick et al. (1988);
PCBs — SMS (1991) = Long and Morgan (1990) > Barrick et al. (1988).
The rankings should be interpreted with caution. For example, SMS (1991)
predicted toxicity at Station SR-03-06 in total PCB concentration, but did not
predict toxicity for metals or PAHs. Although a negative hit indicated that a
threshold concentration for metals or PAHs was not exceeded, the total PCB
threshold concentration predicted the toxicity. To evaluate the negative hits
(false negatives) of the recommended threshold concentrations all chemical
classes need to be assessed.
In comparing recommended thresholds and observed toxicity, the
occurrence of false positives (sediments that exceeded contaminant recommended
thresholds but toxicity was not observed) is important for assessing the
predictive value of the threshold concentrations. Four stations in Saginaw
River (3), 57% of the stations, are characterized by this situation (Stations
SR-03-05, SR-03-08, SR-03-16 and SR-03-24) (Table 4.40). The SMS (1991) and
Long and Morgan (1990) recommended threshold concentrations for metals and
PCBs were not predictive. It appears that problems are associated with
defining the toxicity of individual contaminants from data on complex mixtures
of contaminants as observed in the above comparison of recommended thresholds.
Similar findings have been reported by Jenkins and Gillam (1991). The
occurrence of false positives indicates the need for further development of
the threshold concentration approach in sediment hazard assessments, as the
current thresholds have varying degrees of uncertainty associated with them.
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Although the sediments contained complex mixtures of contaminants that
may have been complexed by either TOC (e.g., hydrophobic organics) or AVS
(e.g., SEM), sediment particles may have also played an important role in the
sorption process which influences contaminant bioavailability. Normalization
to TOC may account for the extent of sorption to sediments, but fine sediment
particles (e.g., clays) may also contribute significantly to sorption and
reduce contaminant bioavailability (Landrum and Robbins 1990). If a large
proportion of the sediment particles are fines (less than 63 /im) ,
normalization to organic carbon alone, to estimate bioavailable organics, may
be misleading (Landrum and Robbins 1990). For example, all reduced survival
responses compared to control, for at least one species tested in the present
study, occurred in sediments (except sediment from Station SR-01-06)
containing greater than 90% fine sediment particles (Table 4.37). These
results suggest that a relationship may occur between particle size and
contaminant availability. However, further evaluation of the organic
contaminant data for these studies is needed to define relations between
percentage fine sediment particles, or TOC, to test organism responses. Both
acute and chronic endpoints should be evaluated in relation to contaminant
concentrations.
Sediment tests, and test organisms selected to predict effects on
indigenous biota, should be sensitive and able to identify affected areas,
such that subsequent remedial actions are directed accurately to the areas in
need (Becker et. al 1990). Although the amphipod exposures always had high
control survival, greater than or equal to the ASTM test acceptability at 80%
(ASTM E 1383-90), no one endpoint (survival, growth, sexual maturation), or
duration (14- or 28-day), consistently identified toxicity. Combining all
AOCs for amphipod endpoint contrasts between 14- and 28-d exposures, 28-d
survival identified 17% more toxic sediments than 14-d exposures, with only an
11% agreement on toxic sediments, and 72% agreement on non-toxic sediments
(Figure 4.4). When 14- to 28-d growth (body length) were contrasted they
matched closely in ability to identify toxic sediments, with a 1% difference,
and had the lowest percentage of non-toxic sediments (55%) to the other
amphipod endpoints (Figure 4.4). Although 28-d exposure survival may exceed
the 14-d survival (Tables 4.18, 4.26 and 4.35), 28-d exposures will identify
more toxic sediments than 14-d exposures, for both survival or growth (Figure
4.4). Although comparisons across AOCs for amphipods, indicated that growth
(body length) was a sound indicator of toxicity, growth did not consistently
estimate toxicity at all AOCs, when compared to the other sublethal endpoints
(increase in length, number of antennal segments, and sexual maturation).
Because of the inconsistencies observed in the ability of amphipod sublethal
endpoints to identify toxicity, several endpoints should be assessed in the
amphipod tests for a thorough toxicity evaluation.
Amphipod antennal segment number and body length were not predictors of
sexual maturation. For example, almost 100% of the amphipods exposed to
Buffalo River sediments were mature at 28 days and had about 14 antennal
segments, yet almost 4% were mature at 14 days which also had 14 antennal
segments (Table 4.18). The number of amphipod antennal segments at 28 days is
lower for Buffalo River exposures than the other AOCs, and is also lower than
values Nelson and Brunson (1992) reported for amphipods at the same age (about
20 segments at six weeks old). In addition, using Saginaw River (1) 28-d
amphipod responses, Nelson and Brunson (1992) observed that the relationship
between body length and antennal segment number was not a predictor of sexual
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maturation, or age, by instar. Nelson and Brunson (1992) also observed that
overlapping ranges for body length and antennal segment number occur because
of individual variation, density dependent factors (i.e., competitive
interactions), and density independent factors (i.e., temperature).
Given the population mean (p) and the population variance (a2) for
endpoints such as antennal segment number or sexual maturation (Tables 4.11,
4.18, 4.26 and 4.35) future studies can be designed using the following
formula:
16 * a2/(P * /t)
where: H. azteca 28-d (33%) > H. azteca 14-d (32%) >
C. tentans 10-d (31%);
Growth —
H. azteca 28-d (33%) > C. tentans 10-d (31%) > H. azteca 14-d (29%) > C.
riparius 14-d (13%);
Growth and Survival —
H. azteca 28-d (66%) > C. tentans 10-d (62%) > H. azteca 14-d (61%) > C.
riparius 14-d (50%).
The number of stations tested and the number of endpoints measured varied
between species. For example, C. tentans percentage hits for survival (31%)
was assessed in 84% of the stations, whereas C. riparius percentage hits for
survival (37%) was assessed at 100% of the stations (Table 4.41). The
proportions of hits and no hits to the number of stations influences the final
percentage determination. Caution must be taken with these comparisons as
both C. riparius (Buffalo River) and C. tentans (Saginaw River (3)) exposures
did not meet ASTM test acceptability criteria as stated earlier. In addition,
to fully evaluate species sensitivity exposure procedures should be identical
(e.g., temperature).
Amphipod survival in exposures to whole sediments from Waukegan Harbor
was more sensitive in identifying toxicity than C. riparius adult emergence
(Ingersoll and Nelson 1990), but amphipod 48 h survival to the same Waukegan
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sediment tested at another laboratory was not as sensitive as Ceriodaphnia
dubia survival (Burton et al. 1989). Two day amphipod exposures were
inadequate to identify toxicity with the Waukegan sediments, whereas, the 10-
and 29-d amphipod sediment exposures produced lethal and sublethal effects
(Ingersoll and Nelson 1990).
Growth of C. tentans was identified as the most sensitive endpoint in
studies conducted with Detroit River whole sediments over that of mortality
for Daphnia maqna or Hexaqenia limbata (Giesy et al. 1988). In another
Detroit River whole sediment evaluation, C. tentans was less sensitive than D.
maqna (Giesy et al. 1990). An important distinction between the C. tentans
exposures and the whole sediment tests with C. riparius and amphipods is that
the former tests were conducted with single test organisms in individual test
chambers. This configuration eliminates the potential of density dependent
effects, primarily growth, by preventing competition for common resources
(i.e., food), within a test container, and makes retrieving the organism
somewhat easier at the end of the test.
CONCLUSIONS AND RECOMMENDATIONS
Changes in the physicochemical and biological status of the sediment
samples, similar to those described by Patrick et al. (1977) (i.e., sediment
redox state, molecular diffusion) and Riedel et al. (1987) (i.e.,
bioturbation), may have occurred during sampling, transportation, and testing
(Burton 1991). Test organism responses from laboratory whole sediment
exposures may not reflect in situ responses because of the physicochemical and
biological changes that occur during sediment manipulation. But laboratory
exposures are useful tools to assess the bioavailability of complex mixtures
of contaminants under controlled conditions. Responses of the test organisms
exposed to sediments from each AOC, and measured concentrations of
contaminants in the sediments tested, provided strong evidence that a gradient
in environmental toxicity exists across the AOCs. However, specific
contaminants causing the effects could not be identified. Sediments from
Indiana Harbor were generally more toxic to the test organisms than sediments
from Buffalo River which were generally more toxic than the sediments from
Saginaw River.
The contaminant exposure that benthic infaunal organisms encounter in
the environment often involves a prolonged exposure with slow contaminant
accumulation, that often results in sublethal responses, not acute toxicity
(Chapman 1989). Chronic sediment toxicity tests are needed to better define
(1) the nature of time-integrated contaminant uptake in complex chemical
mixtures; (2) sublethal responses using several sensitive benthic organisms,
covering phylogenetically diverse groups; and (3) the ability to estimate
toxicity accurately (identifying toxicity without false predictions) and
efficiently (consistently identifying toxicity). A better understanding of
the importance of sublethal responses is needed. Although interpreting
sublethal responses is not as clear as survival data, with continued
development and guidance, sublethal endpoints will provide additional toxicity
information necessary to make sound sediment quality decisions. Additionally,
sublethal responses need to be compared to benthic community effects and
sediment chemistry to see if the sublethal endpoints are predictive (Chapman
1989).
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A comprehensive approach to evaluating sediment quality using sublethal
test methods is to use a tiered testing approach (Burton et al. 1989a,b,
Becker et al. 1990, USEPA/USACE 1991, Burton 1991) and several types of
toxicity tests, with various species and durations (Burton et al. 1989a,b,
Ingersoll and Nelson 1990, Burton 1991), over that of a single species, single
duration, and survival as the only toxicity endpoint.
Because of biological, physical, and chemical relationships with
sediment associated contaminants, as well as unknown relationships, there is
no single whole sediment toxicity test ideal for all evaluations. It is
essential to use multiple toxicity tests as demonstrated by the varied
responses of the test species exposed to contaminated sediments in the present
study. The use of multiple toxicity tests is needed to accurately
(discriminating between toxic and non-toxic areas) and efficiently estimate
sediment toxicity. It is necessary to better understand what the test
responses mean biologically, ecologically and economically. Moreover, of
equal importance is the necessity that laboratory sediment toxicity test
results be field-verified (i.e., in situ exposures) and comparisons made to
benthic community responses, to establish the ability of laboratory exposures
to predict biological effects in the field.
The following recommendations are a summary of the future research needs
identified as important issues relevant to whole sediment toxicity testing and
interpretation of test results. Many of these issues are central to the
understanding of contaminant bioavailability and the relevance of sediment
studies with field-collected sediments.
0 The use of multiple toxicity tests is needed to accurately and
efficiently estimate sediment toxicity.
0 Chronic sediment toxicity tests need further development to better
define the nature of time-integrated contaminant uptake in complex
chemical mixtures, and sublethal responses, using sensitive benthic
organisms, covering phylogenetically diverse groups.
0 Sublethal responses need to be compared to benthic community effects and
sediment chemistry to see if the sublethal endpoints are predictive.
0 Laboratory sediment toxicity test results need to be field-verified
(i.e., in situ exposures) and comparisons made to benthic community
responses, to establish the ability of laboratory exposures to predict
biological effects in the field.
0 Additional studies are needed with other metals and complex mixtures of
metals to further determine SEM/AVS relationships and its influence on
bioavailability.
0 Rate limiting factors of sediment-sorbed contaminants need further
investigation for a better understanding of the desorption kinetics of
non-labile contaminants, and the relationship to bioavailability.
0 Further investigation is needed to evaluate the influence of TOC and
organic contaminant bioavailability in complex mixtures, and the
relationship to recommended threshold values, at varying TOC levels, or
those evaluated on a dry weight basis.
0 Further investigation of organic contaminants is needed to define
relationships between percentage fines or TOC to test organism
responses, including acute and chronic endpoints, in relation to
contaminant concentrations.
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SUMMARY
Responses of the test organisms exposed to sediments from each AOC, and
measured concentrations of contaminants in these sediments, provided strong
evidence that a gradient in environmental toxicity exists across the AOCs.
However, specific contaminants causing the effects could not be identified.
Sediments from Indiana Harbor were generally more toxic to the test organisms
than sediments from Buffalo River which were generally more toxic than the
sediments from Saginaw River.
Comparisons of survival and growth for test species relative sensitivity
expressed as percentage of hits (ability to identify toxicity) include:
Survival --
C. riparius 14-d > H. azteca 28-d > H. azteca 14-d > C. tentans 10-d;
Growth —
H. azteca 28-d > C. tentans 10-d > H. azteca 14-d > C. riparius 14-d;
Growth and Survival —
H. azteca 28-d > C. tentans 10-d > H. azteca 14-d > C. riparius 14-d.
INDIANA HARBOR
Indiana Harbor sediments were toxic to all of the test species. The
sediments caused significant reduction in survival or body length, but all
test species did not identify toxicity with equal sensitivity for the Indiana
Harbor sediment exposures. Survival of C. tentans was greater than survival
of amphipods or C. riparius. The acute toxicity observed, however, did verify
extensive chemical contamination. Overall, C. riparius and H. azteca were the
most sensitive species in the Indiana Harbor sediment exposures.
BUFFALO RIVER
Buffalo River sediments significantly reduced amphipod survival in the
28-d exposure at 80% of the stations. Survival of C. riparius was
significantly reduced in the 14-d exposure in two stations (40%), but survival
was not significantly reduced for C. tentans for any stations in the 10-d
exposure. Amphipod 14-d growth identified 100% of the Buffalo River sediments
as toxic, and significant differences from control for antennal segment number
or sexual maturation was predicted by amphipod reductions in body length at
80% of the stations. Growth of C. riparius identified 20% of the Buffalo
River sediments as toxic, and growth of C. tentans identified 40% of the
stations as toxic. Overall, H. azteca was the most sensitive species in the
Buffalo River sediment exposures.
SAGINAW RIVER (1)
Saginaw River (1) sediment from Station SR-01-06 was toxic to amphipods
and C. riparius. with significant reductions in survival and growth. The
amphipod 28-d growth response identified 100% of the Saginaw River (1)
stations toxic, whereas C. riparius growth identified only 33% of the
stations. Overall, H. azteca was the most sensitive indicator of toxicity in
the Saginaw River (1) sediments tested.
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SAGINAW RIVER (3)
Saginaw River (3) sediments were not toxic to the species tested in the
whole sediment exposures. Survival at any stations, with the exception of C.
tentans (Station SR-03-06), was not significantly different from control.
Growth for amphipods and C. tentans was not significantly reduced compared to
control. Contaminated sediment from Station SR-03-05 exposed to H. azteca
delayed sexual maturation in the 14-d exposure, but not in the 28-d exposure.
Results from the toxicity tests for all test species indicate an equal
sensitivity to Saginaw River (3) sediments, except Station SR-03-06.
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4-43
-------
LIST OF FIGURES IN CHAPTER 4
4.1 Amphipod percentage increase in body length for the 28-d exposures,
significant differences from control denoted with an *.
4.2 Relation of summed total metal concentrations to the survival of each
species tested in sediments across all Areas of Concern.
4.3 Relation of summed total PAH concentrations to the survival of each
species tested, in sediments across all Areas of Concern.
4.4 Comparison of H. azteca test sensitivity. The percentage hits are based
on the number of stations that elicited significant reductions in
survival, body length or sexual maturation.
4-44
-------
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72.0%
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22.0%
6.0%
72.0%
HA-28
Survival / Sexual Maturation
76.8%
77.8%
70.9%
54.5%
HA-28 Growth / HA-14 Growth
Figure 4.4
response / response
no response I no response
+ /-
response / no response
no response / response
-------
Table 4.1 Chemical names for compounds analyzed in the sediments from
stations tested in laboratory whole sediment toxicity tests.
CHEMICAL CODES
CHEMICAL NAME
2378-TCDF
Total TCDF
2378 TCDD
Total TCDD
12378-PeCDF
23478-PeCDF
Total PeCDF
12378-PeCDD
Total PeCDD
123478-HxCDF
123678-HxCDF
123789-HxCDF
234678-HxCDF
Total HxCDF
123478-HxCDD
123678-HxCDD
123789-HxCDD
Total HxCDD
1234678-HpCDF
1234789-HpCDF
Total HpCDF
1234678-HpCDD
Total HpCDD
OCDF
OCDD
Ag
As
Cd
Cr
Cu
Fe
Hg
Mn
Ni
Pb
Se
Zn
TOC
AVS
SOLIDS
Methyl-mercury
TBT
DBT
MBT
1,4 DCB
2,3,7, 8-Tetrachlorodibenzof uran
Total Tetrachlorodibenzofuran
2,3,7, 8-Tetrachlorodibenzodioxin
Total Tetrachlorodibenzodioxin
1,2,3,7, 8-Pentachlorodibenzof uran
2,3,4,7, 8-Pentachlorodibenzof uran
Total Pentachlorodibenzofuran
1,2,3,7, 8-Pentachlorodibenzodioxin
Total Pentachlorodibenzodioxin
1,2,3,4,7, 8-Hexachlorodibenzof uran
1,2,3,6,7, 8-Hexachlorodibenzof uran
1,2,3,7,8, 9-Hexachlorodibenzof uran
2,3,4,6,7, 8-Hexachlorodibenzof uran
Total Hexachlorodibenzofuran
1,2,3,4,7, 8-Hexachlorodibenzodioxin
1,2,3,6,7, 8-Hexachlorodibenzodioxin
1,2,3,7,8, 9-Hexachlorodibenzodioxin
Total Hexachlorodibenzodioxin
1,2,3,4,6,7, 8-Heptachlorodibenzof uran
1,2,3,4,7,8, 9-Heptachlorodibenzof uran
Total Heptachlorodibenzofuran
1,2,3,4,6,7, 8-Heptachlorodibenzodioxin
Total Heptachlorodibenzodioxin
Octachlorodibenzofuran
Octachlorodibenzodioxin
Silver
Arsenic
Cadmium
Chromium
Copper
Iron
Mercury
Manganese
Nickel
Lead
Selenium
Zinc
Total Organic Carbon
Acid Volatile Sulfides
Total Solids
Methyl-mercury
Tributylin
Dibutylin
Monobutylin
1 , 4-Dichlorobenzene
-------
Table 4.1 (Continued).
Naph
2-MNaph
Acnaph
DM PH
DBF
Fluore
Phen
Anth
Fluora
Pyrene
BBPh
BaAnth
BisPh
Chrys
DnOPh
BbFluor
BkFluor
BaPyr
IndPyr
BghiPer
PCB 1016
PCB 1221
PCB 1232
PCB 1242
PCB 1248
PCB 1254
PCB 1260
Aldrin
A-BHC
B-BHC
D-BHC
Chlordane
4, 4, ODD
4, 4, DDE
4, 4, DDT
Dieldrin
Endosulfan I
Endosulfan II
Endosulfan
sulf ate
Endrin
Endrin Aldehyde
Endrin Ketone
Heptachlor
Heptachlor
epoxide
Lindane (G-BHC)
Toxaphene
Methoxychlor
Naphthalene
2-Methylnaphthalene
Acenapthene
Dimethyl Phthalate
Dibenzofuran
Fluorene
Phenanthrene
Anthracene
Fluoranthene
Pyrene
Butyl Benzyl Phthalate
Benzo ( a ) anthracene
Bis ( 2-Ethylhexyl ) Phthalate
Chrysene
Di-n-octylphalate
Benzo ( b ) Fluoranthene
Benzo ( k) Fluoranthene
Benzo ( a ) Pyrene
Indeno ( 1, 2, 3-cd) Pyrene
Benzo (g,h, i)perylene
PCB 1016
PCB 1221
PCB 1232
PCB 1242
PCB 1248
PCB 1254
PCB 1260
Aldrin (pesticide)
Hexachlorocyclohexane-Alpha
Hexachlorocyclohexane-Beta
Hexachlorocyclohexane-Delta
Chlordane
Dieldrin
Alpha Endosulfan
Beta Endosulfan
Endosulfan sulphate
Endrin
Endrin Aldehyde
Endrin Ketone
Heptachlor
Heptachlor epoxide
Hexachlorocyclohexane-Gamma
Toxaphene
Methoxychlor
-------
Table 4.2 Concentrations of total organic carbon, percent solids, and organometals in whole sediment
samples from Indiana Harbor, IN. Ranges for samples analyzed in replicate are shown in parentheses. See
Table 4.1 for an explanation of chemical abbreviations.
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
TOC
(%)
7.7
5.6
11.1
11.6
8.8
10.4
12.3
Solids
(%)
40.8
44.8
50.2
29.8
46.7
33.0
19.6
Methyl-
mercury
(ng/dry g)
<0.1
2.0
<0.1
<0.1
0.5
1.4
<0.1
TBT
(ng/dry g)
240
110
300
1500
(14-
19)
370
530
DBT
(ng/dry g)
47
32
58
370
(11-
19)
110
160
MBT
(ng/dry g)
7.4
7.2
17.0
39.0
12.0
28.0)
12.0
26.0
< Indicates that compound was not detected at detection limits shown.
-------
Table 4.3 Concentrations of metals (jig/dry g) in whole sediment samples from Indiana Harbor, IN. See Table
4.1 for an explanation of chemical abbreviations.
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Ag
0.2
0.04
0.02
6.0
7.1
4.7
5.2
As
60
32
45
52
93
56
63
Cd
9.1
5.2
10.4
11.7
24.2
12.4
18.4
Cr
572
407
580
1132
2610
780
1412
Cu
226
182
219
379
287
284
354
Metal
Fe(%)
19.7
14.4
23.4
7.9
28.8
12.1
21.4
(fig/dry g)
Hg
0.9
0.7
0.9
1.9
2.1
1.8
1.8
Mn
2420
1970
2740
2410
3280
1674
2450
Ni
50
50
<50
103
<58
95
88
Pb
589
396
415
878
1354
1223
791
Se
2.6
2.3
2.0
3.8
3.1
3.9
3.3
Zn
3250
2250
2290
4460
7960
3540
4080
< Indicates compound not detected at detection limit shown.
-------
Table 4.4 Simultaneously extracted metals (/iM/dry g) and the acid volatile sulfides (/an/dry g) in Indiana Harbor, IN, and
Buffalo River, NY whole sediment samples. See Table 4.1 for an explanation of chemical abbreviations.
AVS
Cd
Cr
Cu
Fe
Mn
Ni
Pb
Zn
SUM SUM
Cd,Cr,Cu SEM/AVS
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06 REP1
BR-01-06 REP2
BR-01-07
BR-01-08
BR-01-09
BR-01-10
(/iM/dry g)
33.5
15.7
21.8
52.6
71.4
54.1
31.7
161.0
1.3
5.8
4.2
16.9
10.7
10.4
5.7
2.6
5.1
13.5
0.013
0.007
0.008
0.012
0.032
0.011
0.008
0.012
0.001
0.003
0.002
0.004
0.003
0.004
0.005
0.002
0.005
0.004
0.790
0.428
0.796
1.160
4.994
0.784
0.740
0.288
0.007
0.064
0.015
0.043
0.039
0.034
0.024
0.013
0.025
0.009
0.280
0.247
0.212
0.110
0.267
0.023
0.059
0.062
0.002
0.069
0.054
0.050
0.079
0.080
0.047
0.035
0.029
0.032
152
95
96
104
283
78
83
64
1
22
21
21
23
21
21
13
16
17
3.41
2.74
2.91
2.35
4.61
1.61
1.59
2.40
0.04
1.16
1.56
1.01
1.20
1.06
1.42
1.09
1.35
1.04
0.083
0.058
0.069
0.114
0.154
0.041
0.106
0.111
0.013
0.030
0.002
0.024
0.022
0.020
0.043
0.023
0.044
0.015
0.494
0.289
0.345
0.515
1.255
0.431
0.285
0.189
0.011
0.067
0.045
0.074
0.104
0.097
0.036
0.026
0.037
0.030
Ni,Pb,Zn
1.740
1.008
1.171
1.564
4.552
1.131
0.878
0.419
0.009
0.102
0.068
0.131
0.164
0.151
0.053
0.072
0.039
0.045
3.40
2.04
2.60
3.47
11.25
2.42
2.08
1.08
0.05
0.33
0.19
0.32
0.41
0.38
0.20
0.17
0.18
0.14
0.010
0.130
0.119
0.066
0.158
0.045
0.066
0.007
0.038
0.058
0.045
0.019
0.038
0.034
0.036
0.067
0.035
0.010
-------
Table 4.5 Particle size distribution (mm), on a dry weight basis, in whole
sediment samples from Indiana Harbor, IN.
Particle
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
0.25
>1.0 - 1.0
1.0 2.0
0.2 2.2
4.4 10.5
1.7 2.0
0.9 3.2
0.9 2.3
0.4 1.1
0.
14
26
40
12
11
16
13
Size (mm)
063
0.25
.4
.1
.6
.3
.5
.4
.1
0.038
- 0.063
5.5
6.3
6.2
6.1
4.8
6.5
9.9
<0.038
72.3
63.6
36.8
77.7
75.5
70.5
72.1
-------
Table 4.6 Concentrations of polynuclear aromatic and other semivolatile compounds (ng/dry g) in whole sediment samples from
Indiana Harbor, IN. Ranges for samples analyzed in replicate are shown in parentheses. See Table 4.1 for an explanation of
chemical abbreviations.
Polynuclear Aromatic Hydrocarbon (ng/dry g)
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
1-4
DCS
82
31
45
380
125
(110-
140}
160
930
BBPh
<160
<100
<58
16000
<122
(<85-
<160)
<240
<220
Naph
7300
3600
6300
8500
4550
(4100-)
5000)
6100
24000
BaAnth
7300
4200
5800
16000
32000
(25000-
39000)
30000
6900
2-M
Naph
2000
930
1200
5400
21015
(<31-
42000)
20000
4200
BisPh
10000
4700
3800
290000
7150
(5900-
8400)
18000
15000
DMPh
<68
<43
<25
<95
<52
(<37-
<68)
<110
<95
Chrys
8600
5200
7200
26000
31500
(24000-
39000)
33000
9400
DBF
2300
920
1200
5700
26530
(<61-
53000)
6900
2400
DnOPh
1900
4100
430
37000
<130
(<90-
<170)
2600
<240
Fluore
2400
790
1400
3200
30530
(<61-
61000)
12000
3200
BbFluor
7800
5600
6300
24000
20500
(19000-
22000)
26000
8900
Phen
7000
3400
5100
9900
151500
(33000-
270000)
79000
13000
BkFluor
10000
4200
5100
23000
13500
(12000-
15000)
21000
9700
Anth
2400
1400
2200
3400
215000
(130000-
300000)
26000
3500
BaPyr
10000
7000
5700
25000
31000
(21000-
41000)
29000
9200
Fluora
8600
4800
7200
14000
80000
(40000-
120000)
56000
9600
IndPyr
7300
5300
6600
2000
10100
(9200-
11000)
19000
5800
Pyrene
16000
5500
10000
40000
46500
(38000-
55000)
43000
16000
BghiPer
9600
6300
8800
28000
17500
(14000
21000)
31000
7600
< Indicates that compound was not detected at detection limit shown.
-------
Table 4.7 Concentrations of dioxins and furans (pg/dry g) in whole sediment samples from Indiana Harbor, IN. Ranges
for samples analyzed in replicate are shown in parentheses. See Table 4.1 for an explanation of chemical abbreviations.
Polychlorinated-dibenzo-dioxins or
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
2378-
TCDF
290
27
11
600
610
(480-
740)
320
310
Total
TCDF
860
400
170
3700
3450
(2400-
4500)
2200
170O
2378-
TCDD
130
ND
ND
<59
73.5
(<37-
<39
<18
Total
TCDD
190
37
32
490
195
(160-
230)
230
110
12378-
PeCDF
27
12
3.8
56
104
(28-
<180)
27
30
23478-
PeCDF
29
21
7.8
120
106
(82-
130)
89
68
Polychlorinated-dibenzo- furans
Total
PeCDF
340
190
76
1300
1350
(1300-
1400)
680
720
12378-
PeCDD
<52
ND
ND
42
80
(<76-
84)
29
20
Total
PeCDD
ND
22
35
510
1900
140
66
123-
478-
HxCDF
41
16
15
130
225
(210-
240)
95
86
(pg/dry g)
123-
678-
HxCDF
<66
12
6.8
76
98
(86-
110)
<45
43
123- 234-
789- 678-
HxCDF HxCDF
32 <5.6
10 ND
5.2 ND
55 13
77.5 85.5
(56- (<31-
<99) <140))
32 <13
30 <18
-------
Table 4.7 (Continued).
Polychlorinated-dibenzo-dioxins or Polychlorinated-dibenzo-furans (pg/dry g)
Sample
IH-01-03
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Total
HxCDF
700
250
220
1900
3600
(3500-
3700)
2100
920
123-
478-
HxCDD
53
13
17
130
390
(220-
560)
<47
32
123-
678-
HxCDD
73
23
31
210
420
(360-
480)
230
99
123-
789-
HxCDD
97
14
19
380
660
(520-
<800)
290
260
Total
HxCDD
950
350
420
2500
6800
(4600-
9000)
2600
1700
1234-
678-
HpCDF
<38
180
220
1600
3150
(3000-
3300)
340
810
1234-
789-
HpCDF
660
ND
8.8
81
96
(72-
120)
700
36
Total
HoCDF
660
380
510
4200
7700
(6600-
8800)
8200
1500
1234-
678-
HpCDD
1400
410
580
5100
10750
(6500-
15000)
4700
1600
Total
HDCDD
3300
980
1200
9300
23000
(15000-
31000)
5300
3100
OCDF
1600
180
250
6900
17300
(2600-
32000)
12000
2500
OCDD
6700
2300
2900
43000
43500
(41000-
46000)
25000
12000
< Indicates that compound was not detected at detection limit.
ND = Not detected.
-------
Table 4.8 Polychlorinated biphenyls (ng/dry g) in whole sediment samples from Indiana Harbor, IN, and the
Buffalo River, NY. See Table 4.1 for an explanation of chemical abbreviations.
Sample
IH-01-03
IH-01-04
IH-01-06
IH-01-07
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
SR-01-03
SR-01-06
SR-01-10
Polychlorinated Biphenyl
PCB 1016
<360
<360
<500
<410
<510
<300
<360
<340
<360
<340
<300
<260
PCB 1221
<360
<360
<500
<410
<510
<300
<360
<340
<360
<340
<300
<260
PCB 1232
<360
<360
<500
<410
<510
<300
<360
<340
<360
<340
60000
<260
PCB 1242
10000
3000
24000
43000
<510
<300
<360
<340
<360
<340
<300
440
(ng/dry g)
PCB 1248
<360
<360
<500
<410
<510
<300
<360
<340
<360
<340
7900
2300
PCB 1254
<360
1000
<500
<410
<510
<3400
<360
<340
<360
<340
300
<260
PCB 1260
<360
<360
<500
<410
<510
<300
<360
<340
<360
<340
<300
<260
< Indicates that compound was not detected at detection limit shown.
-------
Table 4.9 Pesticide ranges of analytical detection limits (ng/g) measured in
whole sediment samples from Indiana Harbor, IN, Buffalo River, NY, and Saginaw
River, MI. See Table 4.1 for an explanation of chemical abbreviations.
Area of Concern
Pesticide
(ng/dry g)
Aldrin
A-BHC
B-BHC
C-BHC
Gamma cholordane
Alpha Chlordane
4, 4, ODD
4,4, DDE
4, 4, DDT
Deildrin
Endoaulfan I
Endosulfan II
Endosulfan
sulfate
Endrin
Endrin aldehyde
Endrin ketone
Heptachlor
Heptachlor
epoxide
Lindane (G-BHC)
Toxaphene
Methoxychlor
Indiana
Harbor
NA
34-69
34-69
34-69
> 36
34-69
34-69
> 36
34-69
34-69
34-69
34-69
34-69
34-69
34-69
34-69
34-69
> 36
34-69
340-500
170-340
Buffalo
River
24-51
24-51
24-39
24-51
24-51
24-51
24-39
24-51
24-39
24-39
24-51
24-51
24-51
24-51
24-51
24-51
24-51
24-51
24-51
140-510
120-250
Saginaw
River (1)
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
26-43
260-430
130-220
Saginaw
River (3)
2.5-460
2.5-360
2.8-380
2.5-380
2.5-360
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-380
2.5-500
2.5-380
2.5-3800
13-1600
NA indicates not applicable.
-------
Table 4.10 Mean (standard error of the mean in parentheses) measured overlying water quality for Indiana Harbor, IN whole
sediment toxicity tests.
Sample
Alkalinity
(mg/L as
pH
Hardness
(mg/L as
CaCOj)
D.O.
(mg/L)
Unionized
Conductivity Ammonia Chloride Turbidity
(^mhos/cm) (mg/L) (mg/L) (NTU)
Hvalella azteca - 14-day
Control
IH-01-03
IH-01-04
IH-01-06
IH-01-07
8.5(0.49)
8.1(0.09)
8.1(0.12)
8.1(0.09)
8.0(0.09)
82(18.0)
82(06.0)
98(18.0)
90(14.0)
80(08.0)
142(14.0)
156(04.0)
146(10.0)
160(08.0)
156(04.0)
8.7(0.29)
7.9(0.48)
8.0(0.55)
7.9(0.44)
7.8(0.40)
263(07.2)
299(23.6)
323(24.7)
316(31.2)
317(23.3)
0.057(0.05)
0.137(0.13)
0.280(0.28)
0.320(0.31)
0.297(0.27)
8.6(0.03)
10.4(1.09)
12.7(0.95)
14.8(2.82)
12.1(1.68)
7.8(05.55)
35.8(34.20)
39.2(37.71)
44.2(42.80)
31.6(30.18)
Chironomus riparius - 14-day
Control
IH-01-03
IH-01-04
IH-01-06
IH-01-07
8.3(0.60)
8.1(0.10)
8.1(0.12)
8.0(0.11)
8.0(0.09)
84(16.0)
82(06.0)
96(20.0)
90(14.0)
78(10.0)
136(08.0)
150(02.0)
146(10.0)
158(10.0)
152(08.0)
8.2(0.53)
8.2(0.38)
8.2(0.45)
8.2(0.33)
7.9(0.35)
270(10.8)
312(17.0)
322(23.0)
323(28.0)
311(17.7)
0.055(0.05)
0.140(0.13)
0.280(0.28)
0.325(0.31)
0.298(0.27)
8.6(0.02)
10.5(1.02)
12.1(1.59)
14.3(3.27)
13.1(0.67)
16.7(03.35)
35.6(34.46)
39.2(37.75)
44.1(42.93)
31.5(30.31)
Chironomus tentans - 10-day
Control
IH-01-03
IH-01-04
IH-01-06
IH-01-07
8.7(0.73)
8.3(0.04)
8.3(0.11)
8.2(0.09)
8.2(0.08)
92(08.0)
116(28.0)
102(14.0)
110(06.0)
104(16.0)
130(02.0)
182(30.0)
168(32.0)
174 (06.0)
166 (06.0)
7.6(1.60)
7.7(1.20)
8.2(0.95)
7.5(1.35)
7.9(0.70)
296(59.3)
402(71.7)
380(29.9)
412(51.6)
397(67.0)
0.292(0.17)
0.565(0.24)
0.364(0.31)
0.709(0.07)
1.110(0.42)
10.9(2.28)
13.5(1.94)
17.8(4.13)
20.3(2.75)
16.5(2.78)
21.2(07.90)
40.9(29.10)
84.2(07.30)
74.3(12.75)
40.2(21.65)
0.057(0.05)
0.137(0.13)
0.280(0.28)
0.320(0.31)
0.297(0.27)
0.055(0.05)
0.140(0.13)
0.280(0.28)
0.325(0.31)
0.298(0.27)
0.292(0.17)
0.565(0.24)
0.364(0.31)
0.709(0.07)
1.110(0.42)
8.6(0.03)
10.4(1.09)
12.7(0.95)
14.8(2.82)
12.1(1.68)
8.6(0.02)
10.5(1.02)
12.1(1.59)
14.3(3.27)
13.1(0.67)
10.9(2.28)
13.5(1.94)
17.8(4.13)
20.3(2.75)
16.5(2.78)
-------
Table 4.11 Responses of Hvalella azteca. Chironomus riparius. and Chironomus
tentana exposed to whole sediment samples from Indiana Harbor, IN. Mean
values (standard errors in parentheses) within columns and tests with common
letters are not significantly different (p < 0.05).
Test
Survival
Total
Body
Length(mm)
Antennal
Segment(#)
Mature
(*)
Start
End
Hvalella azteca
Control
IH-01-03
IH-01-04
IH-01-06
IH-01-07
14-day
92.
1.
0.
1.
0.
5(
3(
0
3(
0
.06) a
.06)b
.06)b
2.1 2
2
-
1
-
.5(0
.4(1
-
.7(1
-
•2)
.8)
.8)
16.
17.
—
15.
—
3(0.
0(4.
0(4.
5)
6)
6)
1.2(.06)
0.0
—
0.0
—
Chironomus rjparius 14-day
Control
IH-01-03
IH-01-04
IH-01-06
IH-01-07
75.0(3.5)a
2.0(0.8)b
0.0
0.0
12.9(0.3)
Chironomus tentans 10-day
Control
IH-01-03
IH-01-04
IH-01-06
IH-01-07
100.Oa
46.7b
26.7b
0.0
0.0
6.4
18.7(0.9)a
10.3(1.4)b
9.8(1.8)b
-------
Table 4.12 Concentrations of total organic carbon, percentage solids, and organometals in whole sediment
samples from Buffalo River, New York. Ranges for samples analyzed in replicate are shown in parentheses.
See Table 4.1 for an explanation of chemical abbreviations.
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
TOC
(%)
8.9
0.25
2.0
1.8
1.7
(2.1-2.2)
1.7
1.7
2.2
1.9
Solids
(%)
30.0
73.9
50.5
46.8
52.4
(47.7-49.1)
44.1
45.9
41.7
42.8
Methyl-
mercury
(ng/dry g)
1.6
<0.05
<0.05
<0.05
<0.05
(2.1-3.4)
<0.05
<0.05
<0.05
<0.05
TBT
(ng/dry g)
26.0
<0.5
14.0
3.1
4.6
(3.3-3.4)
2.5
1.7
1.3
1.3
DBT
(ng/dry g)
36.0
0.7
4.6
2.6
2.5
(2.5-4.6)
<0.9
0.9
<0.9
<1.0
MBT
(ng/dry g)
15.0
<0.5
<0.8
<0.8
<0.7
(1.1-1.7)
<0.9
<0.8
<0.9
<1.0
< Indicates that compound was not detected at detection limits shown.
-------
Table 4.13 Concentrations of metals (/tg/dry g) in whole sediment samples from Buffalo River, NY. Ranges for samples
analyzed in replicate are shown in parentheses. See Table 4.1 for an explanation of chemical abbreviations.
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Ag
0.5
<0.03
0.4
0.2
0.2
(0.21-
0.24)
0.1
0.1
0.1
0.1
As
34.0
<1.4
13.0
12.0
<4.5
(12.0-
13.0)
12.0
12.0
11.0
8.2
Cd
4.0
0.04
1.4
1.0
1.6
1.2
0.9
0.7
0.7
0.6
Cr
312
<13
113
77
100
(log-
ISO)
92
70
56
46
Cu
148.0
8.2
67.0
50.0
60.0
(90.0-
93.0)
49.0
46.0
41.0
35.0
%Fe
5.5
0.3
4.4
4.2
5.4
4.2
4.1
3.7
3.4
3.0
Metal
Hg
1.9
0.02
0.6
0.2
0.3
(1.6-
1.8)
0.2
0.1
0.1
0.1
(ua/drv o)
Mn
1386
40
685
789
673
(628-
630)
726
731
726
556
Ni
57.0
5.2
45.0
50.0
47.0
(46.0-
(52.0)
44.0
43.0
40.0
34.0
Pb
286
28
107
67
314
(143-
(151)
70
51
49
43
Se
3.8
<0.5
<0.9
<0.8
<1.0
(0.8-
0.9)
<0.9
<0.9
<0.8
<0.8
Zn
900
32
286
224
371
(387
389)
195
166
159
142
< Indicates that compound was not detected at detection limit shown.
-------
Table 4.14 Particle size distribution (mm), on a dry weight basis, in whole
sediment samples from Buffalo River, NY.
Particle Size (mm)
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
>1.0
11.7
0.3
2.8
0.2
14.3
0.5
0.1
0.3
0.5
1.6
0.25
- 1.0
11.7
38.3
4.1
0.6
14.1
0.7
0.7
0.8
2.0
10.3
0.063
- 0.25
30.5
58.5
30.5
6.9
14.6
4.7
6.3
9.2
18.9
23.3
0.038
- 0.063
7.6
0.2
7.0
3.7
5.2
5.4
5.1
9.5
10.0
7.9
<0.038
43.7
0.9
54.0
92.0
51.5
89.1
86.4
74.3
67.0
55.8
-------
Table 4.15 Concentrations of polynuclear aromatic and other semivolatile compounds (ng/dry g) in whole sediment samples
from Buffalo River, NY. See Table 4.1 for an explanation of chemical abbreviations.
Polvnuclear Aromatic Hydrocarbons (na/drv a I
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
1,4
DCB
730
<18
810
58
380
590
68
54
<26
<36
2-M
Naph
20000
<29
470
<67
180
790
140
59
<42
<57
Naph
2400
<29
230
<67
190
2100
83
<47
45
<57
DMPh
<86
<34
<73
<79
<63
<48
<45
<56
<50
<68
DBF
1600
<30
120
<71
140
1200
63
<50
<45
<61
Fluore
1800
<30
400
<71
380
3400
140
<50
46
<61
Phen
6100
<36
1400
580
2700
10000
680
460
540
520
Anth
1700
<34
1100
170
640
4300
240
120
100
99
Fluora
7500
<55
1900
1200
2700
5100
990
760
1200
840
Pyrene
6100
<68
2100
880
2500
6700
1100
690
750
910
-------
Table 4.15 (Continued).
Polvnuclear Aromatic Hydrocarbons (ncr/drv a) _ ___
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
BBPh
15000
<79
<170
<180
1500
6100
2600
<130
210
1500
BaAnth
3500
<21
680
460
870
1800
470
260
310
330
BisPh
39000
880
7000
3100
8800
41000
14000
2400
2700
59000
Chrys
4000
<27
1100
610
1200
2600
690
440
470
480
DnOPh
38000
<84
<180
<200
6300
24000
7800
210
560
1300
BbFluor
7000
<30
1000
640
1400
1500
670
550
610
770
BkFluor
9500
<41
910
650
1200
1500
600
370
460
430
BaPyr
5800
<27
470
600
1200
1300
620
350
440
460
IndPyr
3800
<45
520
<100
640
990
250
78
220
160
BghiPer
3800
<55
620
<130
460
1100
260
<91
240
170
< Indicates that compound was not detected at detection limit shown.
-------
Table 4.16 Concentrations of dioxins and furans (pg/dry g) in whole sediment samples from Buffalo River,
NY. Ranges for samples analyzed in replicate are shown in parentheses. See Table 4.1 for an explanation of
chemical abbreviations.
Polychlorinated-dibenzo-dioxins or Polychlorinated-dibenzo-furans (pg/dry g)
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
2378-
TCDF
7.6
<2.2
4.4
<2.2
<3.7
5.2
4.7
<2.4
<2.0
<2.2
Total
TCDF
51.0
ND
12.0
1.5
ND
37.0
10.0
ND
ND
ND
2378- Total
TCDD TCDD
<2.0 12.0
<2.3 ND
<1.0 ND
<1.7 ND
<1.7 ND
<2 . 1 ND
<1.3 ND
<1.3 ND
<1.9 ND
<0.9 ND
12378-
PeCDF
1.7
<0.8
1.0
<1.3
<0.7
<0.7
2.7
<0.6
<0.7
<0.7
23478- Total
PeCDF PeCDF
2.8 52.0
<0.6 ND
<1.1 33.0
<2.2 19.0
<1.5 16.0
<1.0 83.0
<1.1 36.0
<1.6 9.7
<1.5 3.1
<1.3 3.5
12378- Total
PeCDD PeCDD
<3.3 3.6
<0.9 ND
<1.3 ND
<0.9 5.1
<1.4 ND
<1.5 ND
<0 . 9 ND
<0.6 ND
<0.7 ND
<3.5 7.4
123- 123- 123-
478- 678- 789-
HxCDF HxCDF HxCDF
3.8 <2.5 4.2
<0.7 <0.6 <0.7
1.6 <0.8 <2.3
<7.1 <1.0 <2.5
<1.5 <0.8 <2.2
<2.4 <1.1 <1.2
11.0 3.6 <1.5
<2.7 <1.5 <1.4
<1.5 <0.6 <1.1
<1.2 <0.7 <1.6
234-
678-
HxCDF
2.8
<1.0
<1.1
<1.1
<1.1
10.0
<2.3
<0.9
<1.0
<0.9
-------
Table 4.16 (Continued).
Polychlorinated-dibenzo-dioxins or Polychlorinated-dibenzo-furans (pg/dry g)
Sample
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
Total
HxCDF
67.0
ND
24.0
26.0
12.0
110.0
54.0
5.6
8.7
13.0
123- 123- 123-
478- 678- 789- Total
HxCDD HxCDD HxCDD HxCDD
9.2 36.0 19.0 190.0
<1.1 <0.9 <1.0 ND
<1.2 <3.2 <2.1 14.0
<1.7 <2.4 <1.7 ND
<1.1 <1.8 <1.8 8.0
<1.4 <3.5 <1.8 14.0
<2.0 <1.5 <1.4 ND
<0.8 <1.4 <0.8 ND
<0.8 <1.1 <0.5 5.0
<0.9 <1.2 <1.4 4.0
1234-
678-
HpCDF
150.0
<1.8
13.0
11.0
12.0
23.0
15.0
7.8
2.8
5.2
1234-
789- Total
HpCDF HpCDF
9.1 640.0
<1.1 3.8
<1.9 42.0
<4.2 32.0
<1.7 34.0
<3.8 61.0
<3.1 46.0
<0.95 22.0
<0.7 15.0
<0.8 27.0
1234-
678-
HpCDD
1200.0
6.5
64.0
43.0
52.0
54.0
39.0
28.0
33.0
36.0
Total
HpCDD
2000.0
12.0
120.0
74.0
93.0
100.0
69.0
50.0
59.0
61.0
OCDF
780.0
<3.9
39.0
26.0
32.0
42.0
34.0
21.0
20.0
20.0
OCDD
12000.0
53.0
560.0
340.0
400.0
400.0
290.0
250.0
250.0
260.0
< Indicates that compound was not detected at detection limit shown.
ND = Not detected.
-------
Table 4.17 Mean (standard error of the mean in parentheses) measured overlying water quality for Buffalo River, NY whole
sediment toxicity tests.
Sample
PH
Alkalinity Hardness
(mg/L as (mg/L as
CaCOr ) CaCOi )
Hvalella azteca - 14-day
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
7.9 (0.09)
8.3 (0.06)
8.1 (0.06)
8.1 (0.07)
8.1 (0.05)
8.1 (0.11)
90 (6.0)
87 (1.0)
87 (1.0)
93 (7.0)
113(27.0)
180(92.0)
160
165
166
170
175
220
(0.0)
(5.0)
(4.0)
(10.0)
(15.0)
(60.0)
D.O. Conductivity
(mq/L) (umhos/cm)
8.4
7.5
7.7
7.6
7.7
7.3
(0.24)
(0.07)
(0.33)
(0.22)
(0.38)
(0.35)
283
329
307
319
332
408
(17.3)
(18.3)
(8.8)
(8.5)
(14.3)
(93.2)
Unionized
Ammonia
(ma/L)
0.004
0.063
0.052
0.074
0.069
0.116
(0.00)
(0.06)
(0.05)
(0.07)
(0.06)
(0.11)
Chloride
(ma/L)
12.1 (0.25)
21.3 (10.05)
12.1 (1.10)
12.1 (1.85)
11.9 (2.10)
19.7 (8.95)
Turbidity
(NTU)
1.2 (0.69)
13.4 (12.90)
10.2 (9.65)
42.7 (42.27)
16.1 (15.45)
8.0 (7.52)
Hvalella azteca - 28-day
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Chironomus
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Chironomus
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
7.9 (0.06)
8.2 (0.09)
8.1 (0.06)
8.2 (0.07)
8.2 (0.08)
8.2 (0.09)
riparius -
8.0 (0.14)
8.1 (0.14)
8.0 (0.03)
8.1 (0.05)
8.1 (0.05)
8.0 (0.05)
tentans -
8.1 (0.32)
8.4 (0.05)
8.3 (0.22)
8.3 (0.09)
8.3 (0.09)
8.2 (0.13)
82 (2.0)
82 (6.0)
88 (0.0)
94 (6.0)
116(24.0)
184(88.0)
14-day
82 (2.0)
85 (3.5)
86 (2.0)
92 (8.0)
112(28.0)
182(90.0)
10-day
156(72.0)
156(68.0)
114(26.0)
130(30.0)
150(10.0)
208(64.0)
162
163
169
170
179
228
158
167
161
168
173
216
200
237
187
200
205
240
(2.0)
(7.0)
(1.0)
(10.0)
(11.0)
(52.0)
(2.0)
(3-0)
(9.0)
(12.0)
(17.0)
(64.0)
(40.0)
(67.0)
(17.0)
(20.0)
(15.0)
(40.0)
8.5
7.8
7.9
7.9
7.9
7.6
7.9
6.8
7.7
7.5
7.7
7.3
7.9
7.4
7.9
7.1
7.4
7.1
(0.14)
(0.13)
(0.17)
(0.12)
(0.18)
(0.16)
(0.52)
(0.50)
(0.47)
(0.42)
(0.54)
(0.55)
(1.05)
(0.20)
(0.55)
(0.90)
(1.10)
(0.90)
281
324
318
319
329
382
284
331
313
317
329
408
418
581
375
412
424
490
(15.6)
(10.5)
(3.9)
(4.9)
(8.0)
(53.4)
(22.0)
(21.0)
(10.5)
(13.0)
(17.0)
(93.3)
(180.4)
(233.0)
(68.0)
(92.8)
(80.9)
(77.3)
0.004
0.065
0.052
0.075
0.072
0.117
0.004
0.065
0.051
0.074
0.069
0.118
0.547
0.559
0.066
0.095
0.119
0.232
(0.00)
(0.05)
(0.05)
(0.07)
(0.06)
(0.11)
(0.00)
(0.05)
(0.05)
(0.07)
(0.06)
(0.11)
(0.54)
(0.42)
(0.05)
(0.08)
(0.04)
(0.04)
11.9 (0.45)
21.2 (10.15)
11.7 (1.55)
12.3 (1.60)
12.6 (1.40)
19.9 (8.70)
11.2 (1.10)
20.6 (10.70)
12.0 (1.20)
10.9 (3.00)
11.2 (2.85)
18.9 (9.75)
26.3 (13.95)
53.4 (22.10)
23.4 (10.20)
19.8 (5.85)
25.3 (11.30)
32.1 (3.50)
10.2 (8.30)
17.7 (8.65)
10.5 (9.26)
43.4 (41.60)
16.6 (14.90)
8.6 (6.90)
2.9 (1.07)
21.9 (4.40)
12.4 (7.45)
46.1 (38.88)
17.7 (13.80)
13.7 (1.80)
22.3 (3.80)
23.2 (3.15)
23.3 (3.45)
55.5 (29.50)
31.5 (0.00)
45.8 (30.25}
-------
Table 4.18 Responses of Hyalella azteca, Chironomus riparius, and Chironomua tentans exposed to whole
sediment samples from Buffalo River, NY. Mean values (standard error in parentheses) within columns and
tests with common letters are not significantly different (p < 0.05).
Total Body
Survival (%) Lenath(mm)
Test
Hvalella azteca 14-day
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Hvalella azteca 28-day
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Chironomus riparius 14-day
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Chironomus tentans 10-day
Control
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
Start
82.5(.07)ab 1.3
82.5(.07)ab
63.8(.07)c
71.3(.07)bc
93.8(.07)a
68.8(.07)bc
95.0(.05)a 1.1
85.0(.05)bc
75.0(.05)c
76.3(.05)c
92.5(.05)ab
83.8(.05)bc
60.5(11.0)a
41.5(11.8)ab
38.0(07.0)ab
29.0(06.0)b
23.0(07.3)b
63.5(05.4)a
Wt. (ma)
86. 7a 0.1
93. 3a
93. 3a
93. 3a
93. 3a
100. Oa
End
2.1( .04) a
1.8 (.04) be
1.8( .05) be
1.8 (.04) be
1.8(.04)b 14.
1.9(.04)c
2.7(.09)b
2.6(.09)b
2.3(0.1)a
2.3(0.1)a
2.7(.09)b
2 . 5 ( . 09 ) ab
11.7(0.4)a
12.5(0.5)a
10.2(0.4)b
11.7(0.5)a
11.4(0.7)ab
12.7(0.4)a
Wt. (mq)
3.6(0.3)b
3.2(0.3)b
3 . 1 ( 0 . 3 ) ab
2.5(0.3)a
2.6(0.3)a
3.2(0.2)b
Antennal
Seoments (#) Mature (%)
14.7(.09)c
14.3(.08)a
6.1(.05)a
0.0(.05)b
14.1(0.1)b 4.2(.05)ab
13.8(0.1)b
l(.09)a
14.4(0.1)c
14.7(0.2)
14.2(0.2)
14.3(0.2)
14.1(0.2)
14.0(0.2)
14.5(0.2)
0.0(.05)b
1.4(.05)b
0.0(.05)b
97.4(.04)
100.0 (.04)
98.4(.04)
100.0(.04)
100.0(.04)
100.0(.05)
a
a
a
a
a
a
-------
Table 4.19 Concentrations of total organic carbon, percentage solids, and organometals in whole sediment
samples from Saginaw River, MI first survey. Ranges for samples analyzed in replicate are shown in
parentheses. See Table 4.1 for an explanation of chemical abbreviations.
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
TOG
(%)
(3.1-
3.2)
3.0
2.6
2.1
3.0
3.8
1.0
Solids
(%)
(36-
37)
36
44
56
33
42
64
Methyl- TBT DBT MBT
mercury (ng/dry g) (ng/dry g) (ng/dry g)
(ng/dry g)
0.1 (19.0-
20.0)
<0.1 15.0
<0.1 14.0
<0.1 6.9
<0.1 12.0
<0.1 8.3
<0.1 2.2
(11.0-
14.0)
21.0
7.4
10.0
9.0
5.8
2.3
<1.1
<1.0
<0.9
1.2
<1.1
<0.8
<0.6
< Indicates that compound was not detected at detection limit shown.
-------
Table 4.20 Concentrations of metals (/*g/dry g) in whole sediment samples from Saginaw River, MI first survey. The ranges
for samples analyzed in replicate are noted in parentheses. See Table 4.1 for an explanation of chemical abbreviations.
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
Ag
0.6
0.6
0.5
1.5
0.6
0.2
0.1
As
(13.0-
14.0)
13.1
10.9
3.6
16.0
9.1
5.1
Cd
(0.9-
1.0)
0.9
1.0
10.0
1.0
0.6
0.2
Cr
(74-
84)
90
73
319
84
46
40
Cu
(46.0-
49.0)
49.0
45.0
187.0
51.0
31.0
16.0
Metal
Fe(%)
2.8
3.1
2.6
1.5
3.2
1.8
1.2
(ua/drv
Hg
(0.1-
0.2)
0.2
0.1
0.3
0.2
0.1
0.05
a)
Mn
(671-
672)
819
661
334
817
374
305
•
Ni
(36-
38)
37
35
157
43
28
15
Pb
(56-
58)
55
511
86
58
39
19
Se
(<0.2-
0.5)
0.3
0.4
0.3
<0.2
0.5
0.5
Zn
(386-
389)
352
326
381
372
319
99
< Indicates that compound was not detected at detection limit shown.
-------
Table 4.21 Simultaneously extracted metals (/tM/dry g) and the simultaneously extracted metals/acid volatile sulfide ratios
for Saginaw River, MI first survey sediments. Also included are the Saginaw River, MI third survey sediment estimated SEM
and SEM/AVS ratios. See Table 4.1 for an explanation of chemical abbreviations.
Sample
SR-01-02 REP1
SR-01-02 REP2
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
AVS
(/iM/dry
4.9
4.4
6.0
5.6
15.5
3.6
5.8
1.5
3.3
3.7
1.2
5.8
3.2
8.9
5.0
4.9
7.4
10.0
10.9
24.6
36.8
8.4
Cd
g)
0.002
0.004
0.002
0.002
0.012
0.001
0.002
0.001
0.001
0.002
0.001
0.005
0.008
0.003
0.015
0.001
0.013
0.006
0.018
0.001
0.047
0.003
Cr
0.033
0.027
0.035
0.032
0.417
0.014
0.014
0.008
0.011
0.015
0.015
0.042
0.026
0.026
0.130
0.013
0.068
0.071
0.113
0.016
0.305
0.038
Cu
0.064
0.061
0.050
0.042
0.018
0.030
0.029
0.014
0.021
0.029
0.016
0.048
0.046
0.044
0.133
0.015
0.076
0.074
0.126
0.023
0.331
0.037
Fe
(/iM/dry
19 1
20 1
26 1
20 1
15 0
14 1
14 0
8 0
0
0
0
0
0
1
1
0
0
1
0
0
1
0
Mn
9)
.39
.44
.98
.51
.51
.07
.74
.52
.57
.73
.19
.77
.95
.06
.12
.48
.89
.10
.56
.30
.17
.37
Ni
0.021
0.035
0.019
0.016
0.223
0.012
0.017
0.009
0.010
0.019
0.006
0.026
0.021
0.020
0.027
0.008
0.036
0.030
0.076
0.008
0.214
0.020
Pb
0.036
0.039
0.044
0.031
0.050
0.024
0.029
0.011
0.019
0.021
0.011
0.043
0.036
0.042
0.069
0.011
0.044
0.057
0.047
0.015
0.105
0.025
Zn
0.186
0.192
0.185
0.165
0.195
0.107
0.181
0.055
0.082
0.108
0.028
0.170
0.180
0.266
0.110
0.023
0.108
0.097
0.146
0.030
0.350
0.055
Sum SEM
Cd,Cr,Cu
Ni,Pb,Zn
0.34
0.36
0.34
0.29
0.92
0.18
0.27
0.09
0.14
0.20
0.08
0.33
0.32
0.40
0.48
0.07
0.35
0.33
0.52
0.10
1.36
0.18
Sum
SEM/AVS
0.069
0.082
0.056
0.052
0.059
0.051
0.047
0.065
0.043
0.054
0.067
0.057
0.100
0.045
0.096
0.015
0.047
0.033
0.048
0.004
0.037
0.021
-------
Table 4.22 Particle size distribution (mm), on a dry weight basis, in whole
sediment samples from Saginaw River, MI first survey.
Particle Size (mm)
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
0.25
>1.0 - 1.0
0.1 0.9
0.1 0.9
0.2 6.0
5.4 41.0
0.0 0.3
0.5 1.6
5.3 40.2
0.063
- 0.25
17.0
10.4
21.3
28.2
8.8
45.9
27.7
0.038
- 0.063
10.1
9.6
9.0
6.0
9.6
9.6
5.9
<0.038
71.4
79.3
55.6
19.8
79.3
39.0
19.4
-------
Table 4.23 Concentrations of polynuclear aromatic hydrocarbons and other semivolatile compounds (ng/dry g) in whole
sediment samples from Saginaw River, MI first survey. Ranges for samples analyzed in replicate are noted in parentheses.
See Table 4.1 for an explanation of chemical abbreviations.
Polynuclear Aromatic
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
1-4
DCB
(33-
47.0)
33
47
130
52
25
<4
BBPh
(<34-
<51)
1300
270
<25
240
<39
<18
Naph
(34-
46)
35
48
55
53
27
<6
BaAnth
(140-
180)
170
160
300
190
64
15
2-M
Naph
(35.0-
41)
41
37
63
48
17
<6
BisPh
(1700-
2400)
13000*
1900
4200
3800
840
170
DMPh
(76-
80)
99
110
68
77
21
16
Chrys
(290-
330)
400
300
500
390
120
24
DBF
(18-
22)
<16
20
38
<18
<15
<7
DnOPh
(<36-
54)
2200
76
430
<49
<42
<19
Hydrocarbon
Fluore
(27-
34)
38
<12
69
25
<15
<7
BbFluor
(150-
400)
220
130
310
320
61
<7
(ng/dry g)
Phen
(220-
310)
340
270
390
290
99
27
BkFluor
(200-
220)
280
300
400
220
61
<9
Anth
(42-
70)
66
54
38
30
19
<8
BaPyr
(180-
240)
280
260
210
310
<13
<6
Fluora Pyrene
(130-
190)
190
160
160
280
130
36
IndPyr
(110-
120)
220
210
160
200
<22
<10
(470-
670)
550
460
570
570
190
44
BghiPer
(160-
190)
310
270
290
220
<27
<13
* Indicates above linear range.
< Indicates compound not detected at detection limit shown.
-------
Table 4.24 Concentrations of dioxins and furan (pg/dry g) in whole sediment samples from Saginaw River, MI first survey.
The ranges for replicates are noted in parentheses. See Table 4.1 for an explanation of chemical abbreviations.
Polychlorinated-dibenzo-dioxins or Polychlorinated-dibenzo-furans
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
Sample
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
2378-
TCDF
430
410
12000
2100
500
300
110
Total
HxCDF
(750-
820)
680
2100
1500
950
430
210
Total
TCDF
1300
1100
22000
4900
1400
950
320
123-
478-
HxCDD
(5.6-
7.3)
10
6.9
12
8.9
3.3
1.7
2378-
TCDD
(14-
17)
11
13
38
12
9.3
5.9
123-
678-
HxCDD
(42-
69)
32
39
100
48
21
11
Total
TCDD
140
100
120
230
160
94
54
123-
789-
HxCDD
(17-
30)
9.5
16
26
20
11
5.6
12378-
PeCDF
(170-
220)
220
1800
630
210
130
34
Total
HxCDD
(360-
460)
270
340
620
360
200
98
23478-
PeCDF
(140-
160)
150
2500
540
200
120
29
1234-
678-
HpCDF
(1100-
1300)
770
1200
1900
1100
540
400
Total 12378- Total
PeCDF PeCDD PeCDD
(790- (13- (74-
820) 14) 86)
790 9.5 60
8700 15 73
2800 22 120
1000 17 110
650 7.3 50
170 4.2 15
1234- 1234-
789- Total 678-
HpCDF HpCDF HpCDD
(46- (2200-
57) 3000)
40 1600
63 2300
69 4000
85 1800
26 980
12 770
123-
478-
HxCDF
(160-
180)
190
880
400
270
110
41
(pg/dry
123-
678-
HxCDF
(37-
45)
35
200
76
61
28
7.4
g)
123- 234-
789- 678-
HxCDF HxCDF
(19- (5.5-
22) 11)
17 6
88 29
46 17
30 6.4
13 <3.6
4.1 <2.3
Total
HpCDD
(700-
1300)
530
640
1100
630
300
160
(1200-
2300)
980
1200
1900
1200
550
300
OCDF OCDD
(1400-
2800)
1100
1300
2700
1300
610
400
(5700-
14000)
4100
4900
9300
5000
2200
1300
-------
Table 4.25 Mean (standard error of the mean in parentheses) measured overlying water quality for Saginaw River, MI first
survey whole sediment toxicity tests.
Sample
Hvalella
Control
SR-01-03
SR-O1-06
SR-01-10
Hvalella
Control
SR-01-03
SR-01-06
SR-01-10
PH
azteca
8.0
8.0
8.0
8.0
azteca
7.9
8.1
7.9
8.0
14-day
(0.04)
(0.02)
(0.03)
(0.03)
28-day
(0.05)
(0.07)
(0.02)
(0.05)
Alkalinity,
(mg/L as
CaCC^)
48
117
92
96
66
120
94
96
(0.0)
(27.0)
(8.0)
(8.0)
(18.0)
(24.0)
(6.0)
(8.0)
Hardness,
(mg/L as
CaCOj)
154
186
162
172
152
188
160
172
(10.0)
(18.0)
(2.0)
(8.0)
(8.0)
(16.0)
(0.0)
(8.0)
D.O.
(mg/L)
8.3 (0.40)
7.5 (0.40)
7.6 (0.47)
7.9 (0.44)
8.3 (0.23)
7.2 (0.24)
7.5 (0.20)
7.6 (0.25)
Conduct ivity
(^mhos/cm)
295
349
323
326
298
341
320
323
(16.0)
(26.2)
(10.3)
(11.7)
(9.0)
(15.3)
(8.2)
(9.6)
Unionized
Ammonia
(mg/L)
0.003
0.058
0.064
0.021
0.004
0.059
0.064
0.021
(0.00)
(0.05)
(0.06)
(0.02)
(0.00)
(0.05)
(0.06)
(0.02)
Chloride
(mg/L)
9.0
12.2
11.0
15.3
10.0
12.4
11.0
15.6
(0.96)
(4.26)
(2.91)
(7.33)
(0.05)
(4.10)
(2.95)
(7.05)
Turbidity
(NTU)
2.6 (1.10)
8.9 (7.15)
16.3 (15.70
28.4 (26.60
2.9 (0.85)
10.6 (5.41)
16.4 (15. 5f
29.9 (25.1
Chironomus riparius - 14-day
Control
SR-01-03
SR-01-06
SR-01-10
7.8
7.8
7.7
7.8
(0.09)
(0.11)
<0.13)
(0.13)
34
122
90
98
(14.0)
(22.0)
(10.0)
(6.0)
148
186
160
170
(4.0)
(18.0)
(0.0)
(10.0)
7.9 (0.67)
6.6 (0.77)
6.9 (0.84)
7.0 (0.83)
299
357
310
333
(14.8)
(21.6)
(13.6)
(7.5)
0.003
0.057
0.063
0.021
(0.00)
(0.05)
(0.06)
(0.02)
11.6
12.4
10.4
15.5
(1-55)
(4.10)
(3.50)
(7.15)
7.1 (3.4C
21.0 (5.0C
17.0 (15. C
42.0 (13.C
-------
Table 4.26 Responses of Hvalella azteca and Chironomus rioarius exposed to whole sediment samples from
Saginaw River, MI first survey. Mean values (standard error in parentheses) within columns and 1
common letters are not significantly different (p < 0.05).
Test
Hyalella azteca 14-day
Control
SR-01-03
SR-01-06
SR-01-10
Hyalella azteca 28-day
Control
SR-01-03
SR-01-06
SR-01-10
Chironomus ritaarius 14-day
Control
SR-01-03
SR-01-06
SR-01-10
Survival m
80.0(0.1)a
92.5(0.1)a
10.0(0.1)b
81.3(0.1)a
97.5(0.06)a
93.8(0.06)a
10.0(0.06)b
97.5(0.06)a
77. 5( 0.5)a
81.0(05.1)a
14.5(03.3)b
70.0(13.0)3
Total Body
Lencrthfmm)
Start End
1.4 2.4(0.1)
2.3(0.9)
2.1(0.3)
2.4(0.1)
1.4 3.5(.07)a
3.2(.07)c
2.1(0.2)b
3.4(.07)c
11.3(0.3)a
12.0(0.3)a
5.9(0.5)b
11.5(0.3)a
Antennal
Seoments If}
15.2(0.3)
15.4(0.3)
15.0(0.9)
15.6(0.3)
20.2(0.3)a
19.8(0.3)a
16.2(0.9)b
20.0(0.3)a
Mature
4.9(.
11. 4(.
0.0(,
16. 2(.
98.8(.
100. 0(.
37. 5(,
100. 0(.
f%>
.11)
.16)
.16)
.11)
.19)a
,19)a
,19)b
.19) a
-------
Table 4.27 Concentrations of total organic carbon, percent solids, and organometals in whole sediment samples from
Saginaw River, MI third survey. The ranges for replications are noted in parentheses. See Table 4.1 for an explanation
of chemical abbreviations.
Sample
Grab samples
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core samples
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
TOC
m
(1.0-1.1)
1.3
1.4
0.2
3.6
2.9
4.0
5.0
1.5
3.1
2.9
1.9
0.6
4.1
0.8
Solids
(55.1-56.8)
55.2
61.3
76.4
47.3
52.1
48.7
53.0
67.5
59.0
53.8
64.4
73.5
49.2
76.3
Methylmercury TBT
(nq/q) (na/a)
(<0.1-<.2)
<0. 2
<0.1
<0. 1
<0. 1
<0.2
<0.2
<0.2
<0.2
<0. 1
<0. 2
<0.2
<0.2
<0.2
<0. 1
(<0.4-<1.0)
6.2
3.9
0.8
6.6
10.0
9.2
<0.6
<0.4
4.0
<0.9
3.9
(<0. 4-7.0)
33.0
0.7
DBT
(na/a)
(<0.4-3.3) (<0
3.0
2.7
0.7
3.7
6.1
5.6
3.1
<0.4
4.2
1.9
6.3
(<0.4-6.6)
25.7
<0.3
MBT
(na/a)
.4-<0.8)
1.4
0.8
0.4
0.7
1.8
0.9
2.8
<0.2
1.6
1.3
3.0
(<0.8-<3.1)
11.6
0.8
< Indicated that compound was not detected at detection limit shown.
-------
Table 4.28 Concentrations of metals (^g/dry g) in whole sediment samples from Saginaw River, MI third survey. The ranges
for replicates are noted in parentheses. See Table 4.1 for an explanation of chemical abbreviations.
Grab sample
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Core sample
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X2
SR-03-06-X3
Aa
(0.18-
0.19)
0.3
0.4
0.2
0.6
0.5
0.5
2.8
0.1
1.1
0.8
1.3
0.2
3.3
0.35
As
(6.4-
41.1)
25.0
16.6
15.2
12.0
92.9
14.4
217.0
24.1
60.2
59.5
14.5
21.5
40.6
70.1
Cd
(0.7-
0.8)
0.5
0.9
0.5
2.0
2.9
1.1
5.5
0.5
4.9
2.3
6.9
0.42
17.4
1.2
Cr
(42.0-
67.0)
24.7
34.0
34.8
95.0
58.0
59.0
292.0
29.6
154.0
161.0
255.0
35.3
687.0
86.0
Metal (fig/dry g)
Cu Ha
20.3-
24.0)
24.0
33.1
18.3
54.8
51.9
49.8
150.7
16.6
86.3
84.2
142.8
26.6
375.0
42.2
(0.09-
0.1)
0.1
0.1
0.04
0.1
0.1
0.2
0.6
0.1
0.3
0.6
0.3
0.2
0.7
0.3
Mn
(304-
340)
293
379
99
397
492
549
578
250
460
569
292
155
604
194
Ni
(18.5-
19.1)
15.3
28.5
8.3
37.9
30.9
29.3
40.2
11.7
53.3
43.5
111.4
11.9
316.0
30.2
Pb
(22.6- (<0
26.8)
29.8
34.2
16.9
68.7
58.0
67.7
110.9
17.4
70.5
91.0
75.8
24.0
168.2
39.8
Se
.8- (69
0.9)
<0.8
<0.8
<0.7
1.2
2.0
1.0
3.0
<0.8
<0.8
1.7
<0.8
<0.7
<0.9
<0.76
Zn
.8-
81.6)
166.3
219.0
56.3
347.0
367.0
541.0
224.0
46.1
219.0
197.0
298.0
2.0
714.0
111.6
< Indicates compound not detected at detection limit shown.
-------
Table 4.29 Particle size distribution (mm), on a dry weight basis, in whole
sediment samples from Saginaw River, MI third survey. Ranges for samples
analyzed in replicate are shown in parentheses.
Particle Size (mm)
Sample
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
>1.0
0.1
0.6
(0.3-
0.5)
0.3
0.4
0.6
1.4
0.25
- 1.0
0.9
2.7
(9.3-
9.4)
40.0
2.7
5.5
5.3
0.063
- 0.25
56.5
77.7
(60.6-
63.1)
58.2
52.3
41.1
22.7
0.038
- 0.063
13.2
6.1
(9.9-
9.9)
1.5
13.6
12.9
14.3
<0.038
29.2
12.7
(17.8-
15.0)
2.3
29.9
38.8
54.9
-------
Table 4.30 Concentrations of polynuclear aromatic and other semivolatile compounds (ng/dry g) in whole sediment samples
from Saginaw River, MI third survey. The ranges for replicates are noted in parentheses. See Table 4.1 for an explanation
of chemical abbreviations.
Polvnuclear Aromatic
Sample
Grab samples
SR-03-24
SR-03-06
SR-03-05
SR-03-02
SR-03-16
SR-03-08
SR-03-01
Core samples
SR-03-05-01
SR-03-06-X2
SR-03-02-X3
SR-03-06-X3
SR-03-05-02
SR-03-02-X2
SR-03-06-02
SR-03-06-01
1,2
DCS
<290
<160
<200
<250
<230
<250
(<250-
270)
320
<290
<190
<170
<310
680
<170
<170
1,3
DCS
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
<170
<310
1100
<170
<170
1,4,
DCB
290
<160
<200
<250
<230
<250
(<250-
270)
450
340
<190
<170
<210
1300
<170
<170
4-
Methph
300
<160
<200
370
320
450
(410-
870)
<250
1000
<190
<170
<310
<290
<170
<170
Naph
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
<170
<310
<290
<170
<170
2-M
Naph
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
170
<310
<410
<170
<170
DMPh
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
<170
<310
<290
<170
<170
Ace-
Naph
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
<170
<420
<290
<170
<170
Hydrocarbons
DBF
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
<170
<310
<290
<170
<170
Fluore
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
<170
<310
300
<170
<170
(na/drv
Phen
1000
<160
720
510
460
580
(<250-
270)
950
1300
<190
440
1100
3300
350
1200
a)
Anth
<290
<160
<200
<250
<230
<250
(<250-
270)
<250
<290
<190
<170
<310
800
<170
210
Di-
n-B
290
<160
<200
270
<230
<250
(<250-
270)
<250
<290
<190
<170
<310
<290
<170
<170
Fluora
1200
<160
630
500
420
580
(<250-
270)
610
1000
<190
370
490
1400
250
870
Pyren
1800
<160
880
710
750
1000
(<250-
270)
2000
2700
260
880
2100
6600
850
2000
-------
Table 4.30 (Continued).
Polvnuclear Aromatic Hydrocarbons
-------
Table 4.31 Concentrations of dioxins and furans (pg/dry g) in whole sediment samples from Saginaw River, MI third survey.
Ranges for samples analyzed in replicate are noted in parentheses. See Table 4.1 for an explanation of chemical
abbreviations.
Polvchlorinated-dibenzo-dioxins or Polvchlorinated-dibenzo-furans {pa/dry a)
Sample
Grab samples
SR-03-24
SR-03-06
SR-03-05
SR-03-02
SR-03-16
SR-03-08
SR-03-01
Core samples
SR3-05-01
SR3-06-X2
SR3-02-X3
SR3-06-X3
SR3-05-02
SR3-02-X2
SR3-06-02
SR3-06-01
2378-
TCDF
290
1600
710
1700
240
580
(99-
110)
1300
6300
1500
490
1100
2700
870
8600
Total
TCDF
1100
3100
1600
3400
460
1700
(230-
330)
3400
15000
3300
960
3800
9600
1600
20000
2378-
TCDD
8.0
1.3
11.0
5.2
6.2
32.0
(4-
4.5)
16.0
110.0
ND
3.6
ND
19
ND
42
Total
TCDD
64.0
5.7
50
38
17
230
(18-
26)
140
740
1.2
9.3
14
400
2.2
320
12378-
PeCDF
120
240
260
480
110
250
(36-
50)
550
1900
1500
180
520
850
340
2800
23478-
PeCDF
110
250
190
690
72
160
(23-
32)
400
1500
670
130
330
850
210
2100
Total
PeCDF
560
850
940
2500
280
940
(120-
220)
2700
7800
3800
640
2000
8100
1100
11000
12378-
PeCDD
ND
1.3
15
ND
ND
20
(3.5-
4.4)
ND
ND
ND
ND
ND
56
ND
28
Total
PeCDD
31
3.8
24
11
ND
150
(20-
27)
400
310
ND
26
ND
990
ND
120
123-
478-
HxCDF
200
95
230
350
120
290
(52-
73)
1200
1200
1200
220
ND
2700
260
1800
123-
678-
HxCDF
41
24
43
60
25
82
(24
28)
280
220
210
42
130
720
52
1600
123-
789-
HxCDF
19.0
8.9
15.0
16.0
7.0
22.0
(6.8-
8.2)
73.0
88.0
46.0
14
36
190
11
110
234-
678-
HxCDF
5.8
3.8
7.6
4.6
2.1
111.0
(1-
1.9)
21
21
23
ND
ND
98
2.9
35
-------
Table 4.31 (Continued).
Polvchlorinated-dibenzo-dioxins or Polvchlorinated-dibenzo-furans (pa/drv a)
Sample
Grab samples
SR-03-24
SR-03-06
SR-03-05
SR-03-02
SR-03-16
SR-03-08
SR-03-01
Core samples
SR3-05-01
SR3-06-X2
SR3-02-X3
SR3-06-X3
SR3-05-02
SR3-02-X2
SR3-06-02
SR3-06-01
Total
HxCDF
610
200
490
660
340
1200
(230-
370)
5900
4600
1900
510
880
22000
440
5600
123-
478-
HxCDD
4.1
1.1
6.2
ND
ND
9.5
(1.9-
2.8)
30
19
ND
1.7
ND
95
ND
ND
123-
678-
HxCDD
ND
3.4
ND
9.3
18
60
(11-
13)
260
ND
ND
18
ND
ND
ND
67
123-
789-
15
2.6
15
3.9
13
27
(6.1-
6.7)
98
66
ND
6.6
ND
210
ND
22
Total
HxCDD
190
33
280
87
160
460
(65-
150)
1500
1200
22
120
97
3300
3.7
330
1234-
678-
HPCDF
210
49
520
280
540
930
(440-
450)
9400
4300
230
550
980
39000
110
2400
1234-
789-
HPCDF
68
4.5
44
33
21
61
(11-
13)
450
240
64
37
79
1100
15
110
Total
HpCDF
770
120
1700
570
1300
2100
(770-
810)
18000
10000
450
1100
1700
74000
180
5000
1234-
678-
HpCDD
310
40
700
180
430
700
(210-
220)
2500
3800
50
200
34
12000
9.5
1400
Total
HpCDD
550
74
1100
310
790
1300
(390-
420)
4400
6400
86
350
34
22000
16
2500
OCDF
850
89
1600
530
940
1800
(540-
730)
12000
10000
290
790
1200
56000
140
4300
OCDD
2600
340
5100
1800
4100
6400
(2200-
2500)
25000
34000
550
1900
180
12000
89
14000
ND = Not detected.
-------
Table 4.32 Concentrations of pesticides (/ig/g) in the Saginaw River, MI third survey whole sediment samples. Ranges for
samples analyzed in replicate are shown in parentheses. See Table 4.1 for an explanation of chemical abbreviations.
Pesticide
Aldrin
Sample
SR-03-01 (3.8-
4.6)
SR-03-02 8.0
SR-03-05 9.8
SR-03-06 <2.5
SR-03-08 <3.8
SR-03-16 <3.2
SR-03-24 <3.9
SR-03-02-X2 <360.0
SR-03-02-X3 <2.9
SR-03-05-01 (<3.9-
<320.0)
SR-03-06 (2.7-
460.0)
SR-03-06-X2 1400.0
SR-03-06-X3 <260.0
A-BHC
<3.4)
<3.3
<2.8
<2.5
<3.8
<3.2
<3.9
<360.0
<2.9
<320.0)
<270.0)
880.0
<260.0
B-BHC
<3.4)
6.4
<2.8
3.0
<3.8
9.0
<3.9
<360.0
7.0
<320.0)
<270.0)
<380.0
<260.0
C-BHC
<3.4)
<3.3
<2.8
<2.5
<3.8
<3.2
<3.9
<360.0
<2.9
<320.0)
<270.0)
500.0
<260.0
Gamma
Chlor-
dane
(<3.3-
5.7)
5.9
<2.8
<2.5
9.0
<3.2
7.0
<360.0
5.3
<320.0)
<270.0)
<380.0
<260.0
Alpha
Chlor-
dane
(<3.3-
<3.4)
<3.3
<2.8
<2.5
<3.8
<3.2
<3.9
<360.0
<2.9
<320.0)
<270.0)
<380.0
<260.0
4,4, DDD
7.6)
8.4
<2.8
<2.5
<3.8
<3.2
13.0
<360.0
<2.9
(6.4-
<320.0)
<270.0)
<380.0
<260.0
4,4, DDE
3.9)
5.6
<2.8
<2.5
11.0
<3.2
<3.9
<360.0
<2.9
<320.0)
<270.0)
<380.0
<260.0
4, 4, DDT
13.0)
<3.3
<2.8
<2.5
<3.8
<3.2
<3.9
<360.0
<2.9
<320.0)
<270.0)
<380.0
<260.0
Dieldrin
(3.3-
<3.4)
<3.3
<2.8
<2.5
<3.8
<3.2
7.7
<360.0
<2.9
<320.0)
<270.0)
<380.0
<260.0
Endo-
sulfan I
(<3.3-
<3.4)
<3.3
<2.8
<2.5
12.0
<3.2
6.0
<360.0
<2.9
<320.0)
<270.0)
<380.0
<260.0
Endo-
sulfan II
(<3.<
9.4)
<3.3
7.7
<2.5
11.0
5.2
<3.9
<360
<2.9
<320
<270
<380
<260
.0
.0)
.0)
.0
.0
-------
Table 4.32 (Continued).
Pesticide
Sample
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
SR-03-02-X2
SR-03-02-X3
SR-03-05-01
SR-03-05-02
SR-03-06-01
SR-03-06-02
SR-03-06-X3
Endosulfan Endrin
Sulfate
<3.4)
<3.3
<320.0
<2.5
<3,8
<3.2
<3.9
<360.0
<2.9
<320.0
<3.9
<270.0
<2.5
<260.0
<3.4)
<3.3
<320.0
<2.5
<3.8
<3.2
4.1
<360.0
<2.9
<320.0
4.5
<270.0
<2.5
<260.0
Endrin Hepta-
Aldehyde Chlor
17.0)
<3.3
<320.0
<2.5
<3.8
<3.2
<3.9
<360.0
<2.9
<320.0
<3.9
<270.0
<2.5
<260.0
<3.4)
<3.3
<320.0
<2.5
<3.8
<3.2
<3.9
<360.0
<2.9
<320.0
<3.9
<270.0
<2.5
<260.0
Heptachlor Lindane
Epoxide (G-BHC)
11.0)
<3.3
<320.0
<2.5
17.0
40.0
15.0
<360.0
5.3
<320.0
<3.9
<270.0
<2.5
<260.0
<3.4)
<33.0
<320.0
<2.5
17.0
<3.2
<3.9
<360.0
<2.9
<320.0
<3.9
<270.0
<2.5
<260.0
Toxaphene
(<33.0-
<34.0)
<33.0
<3200.0
<2.5
<38.0
<3.2
39.0
<3600.0
<29.0
<3200.0
<39.0
<2700.0
<25.0
<2600.0
Methoxy-
chlor
(<17.0)
<16.0
<1600.0
<25.0
<19.0
32.0
<19.0
<1800.0
<15.0
<1600.0
<19.0
<1350.0
<13.0
<1300.0
Endrin
Ketone
(<3.3
<3.4)
<3.3
<320.
12.0
<3.8
16.0
<3.9
<360.
<2.9
<320.
<3.9
<270.
<2.5
<260.
-
0
0
0
0
0
< Indicated that compound was not detected at detection limit shown.
-------
Table 4.33 Polychlorinated biphenyl concentrations and analytical detection
limits (ng/dry g) for whole sediments from the Saginaw River, MI third
survey. See Table 4.1 for an explanation of chemical abbreviations.
Grab Sample
SR-03-24
SR-03-06
SR-03-05
SR-03-02
SR-03-16
SR-03-08
SR-03-01
Core Sample
SR-03-05-01
SR-03-06-X2
SR-03-02-X3
SR-03-06-X3
SR-03-05-02
SR-03-02-X2
SR-03-06-02
SR-03-06-01
Polvchlorinated
1242
381
95
370
240
470
21
210
(721-260)
3700
79000
291
4300
62
410
96
28000
Biphenvl (ncr/dry
1254
381
251
120
95
871
94
341
(331-341)
32001
8100
291
26001
100
351
251
27001
a)
1260
381
251
291
321
321
381
341
(331-341)
32001
38001
291
26001
381
361
251
27001
Indicates that compound was not detected at detection limit shown.
-------
Table 4.34 Mean (standard error of the mean in parentheses) measured overlying water quality for Saginaw River, MI third
survey whole sediment toxicity tests.
Samole
DH
Alkalinity
(mg/L as
CaCO,!
Hardness
(mg/L as
CaCOji
D.O.
(ma/L>
Conduct ivity
(umhos/cm)
Unionized
Ammonia
(ma/L)
Chloride
(mq/L)
Turbidity
(NTtn
Hvalella azteca - 14-day
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Hvalella
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
7.8 (0.05)
8.1 (0.05)
8.0 (0.06)
8.1 (0.04)
8.1 (0.04)
8.0 (0.06)
8.0 (0.05)
8.0 (0.10)
azteca - 28-day
7.8 (0.07)
8.1 (0.04)
8.0 (0.05)
8.0 (0.06)
8.0 (0.04)
8.0 (0.05)
8.0 (0.06)
7.9 (0.06)
54
76
82
82
76
84
82
88
62
76
84
81
74
83
84
86
(2.0)
(0.0)
(10.0)
(8.0)
(0.0)
(8.0)
(10.0)
(12.0)
(6.0)
(0.0)
(8.0)
(9.0)
(2.0)
(9.0)
(8.0)
(14.0)
131 (1
149 (3
149 (5
148 (4
151 (3
152 (4
150 (6
156 (8
139 (9
145 (1
153 (1
152 (0
153 (1
150 (6
154 (2
158 (6
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
8
8
7
7
7
7
7
7
8
7
7
7
7
7
7
7
.2
.0
.4
.7
.7
.5
.5
.3
.2
.7
.6
.6
.9
.6
.4
.3
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
(0
.09)
.20)
.13)
.17)
.06)
.19)
.24)
.47)
.24)
.19)
.22)
.23)
.21)
.26)
.33)
(0.28)
283
293
315
305
315
319
323
341
286
292
308
304
303
307
310
308
(9.0)
(6.2)
(16.1)
(7.9)
(15.5)
(15.3)
(14.8)
(26.6)
(7.3)
(4.3)
(9.2)
(9.2)
(7.9)
(9.4)
(9.5)
(19.0)
Chironomus riparius - 14-day
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
7.9 (0.16)
8.0 (0.13)
7.9 (0.22)
7.9 (0.21)
8.0 (0.12)
7.9 (0.19)
8.0 (0.16)
7.9 (0.18)
60
76
77
83
76
86
88
92
(4.0)
(0.0)
(15.0)
(7.0)
(0.0)
(6.0)
(4.0)
(8.0)
141(11
149(03
149(05
152(00
153(01
154(02
158(02
160(04
.0)
.0)
.0)
.0)
.0)
.0)
.0)
.0)
7.
7.
7.
7.
7.
7.
7.
7.
9
6
0
1
5
2
3
2
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
50)
42)
96)
89)
61)
80)
62)
75)
287
285
309
306
307
311
310
329
(11.2)
(1.1)
(15.9)
(14.4)
(9.0)
(12.0)
(15.3)
(21.9)
0.
0.
0.
0.
0.
0.
0.
0.
0.
0.
0.
0.
005
024
049
049
030
047
061
118
004
019
047
049
0.026
0.
0.
0.
0.
0.
0.
0.
0.
0.
0.
040
062
101
004
015
047
049
026
040
063
(0.
(0.
(0.
(0.
00)
01)
03)
04)
(0.02)
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
04)
05)
11)
00)
01)
04)
05)
02)
04)
06)
10)
00)
01)
04)
05)
02)
04)
06)
7
6
8
8
8
8
8
10
6
7
8
8
8
8
8
10
7
7
8
8
8
8
8
.1
.7
.7
.5
.5
.0
.4
.2
.8
.5
.6
.7
.8
.2
.4
.7
.1
.3
.8
.9
.9
.5
.5
(0.
(0.
(2.
(2.
(2.
(2.
(2.
(4.
(0.
(0.
(2.
(2.
(2.
(2.
(1.
(4.
(0.
(0.
(2.
(2.
(2.
(1.
(1.
04)
89)
32)
71)
74)
29)
02)
58)
33)
12)
42)
52)
44)
10)
96)
11)
05)
32)
23)
29)
27)
77)
89)
8.
8.
27.
34.
33.
60.
27.
48.
8.
8.
27.
34.
32.
60.
27.
48.
12.1
11.1
33.5
40.0
37.8
69.5
36.0
9
4
6
7
0
5
0
1
8
5
5
5
9
9
7
5
(8.10)
(7.65)
(26.40)
(33.35)
(32.00)
(59.50)
(26.00)
(46.95)
(8.20)
(7.50)
(26.56)
(33.52)
(32.09)
(59.10)
(25.30)
(46.55)
(4.90)
(4.90)
(20.50)
(28.00)
(27.25)
(50.50)
(17.00)
0.101 (0.10) 10.7 (4.14) 57.8 (37.25)
-------
TABLE 4.34 (Continued).
Sample r»H
Alkalinity
(mg/L as
CaCO,!
Hardness
(mg/L as
CaCO,!
D.O.
fma/Ll
Conductivity
fumhos/cm)
Unionized
Ammonia
(mo/Li
Chloride
fma/L)
Turbidity
(NTU)
Chironomus tentans - 10-day
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
8.3
8.4
8.4
8.6
8.5
8.3
8.5
8.4
(0.17)
(0.23)
(0.25)
(0.42)
(0.30)
(0.23)
(0.33)
(0.22)
112
150
114
99
134
134
110
108
(56.0)
(74.0)
(22.0)
(9.0)
(58.0)
(42.0)
(18.0)
(8.0)
153(23,
169(23.
163(09.
166(14.
185(31.
180(24.
172(16.
178(14.
.0)
0)
0)
0)
0)
0)
0)
0)
6.1
6.6
7.1
8.0
7.8
7.2
7.9
7.6
(2.
(1.
(1.
(0.
(0.
(0.
(0.
(0.
50)
50)
05)
15)
55)
88)
35)
63)
326
348
352
351
370
368
374
389
(68.0)
(74.7)
(27.3)
(30.9)
(60.8)
(47.9)
(49.0)
(33.5)
0.
0.
0.
0.
0.
0.
0.
0.
376
886
361
074
846
219
248
188
(0.
(0.
(0.
(0.
(0.
(0.
(0.
(0.
37)
85)
25)
04)
79)
13)
10)
06)
9.
9.
12.
12.
12.
13.
14.
18.
3
4
3
8
1
7
2
1
(2
(1
(1
(1
(0
(3
(3
(3
.18)
.86)
.25)
.55)
.90)
.40)
.75)
.30)
20.
14.
38.
58.
36.
76.
35.
64.
8(3.75)
3
0
5
0
5
5
0
(1.
(16
(9.
(29
(43
(17
(31
75)
.00)
50)
.00)
.50)
.50)
.00)
-------
Table 4.35 Responses of Hvalella azteca. Chironomus riparius. and Chironomus tentans exposed to whole
sediment samples from Saginaw
columns and tests with common
Test
Hvalella azteca - 14 day
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Hvalella azteca - 28 day
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
C. riparius - 14 day
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
River, MI third
letters are not
Survival (%)
85.0(0.1)
98.8(0.1)
90.0(0.1)
88.8(0.1)
95.0(0.1)
93.8(0.1)
95.0(0.1)
96.3(0.1)
96.3(.09)
93. 8 (.09)
100.0(.09)
92. 5 (.09)
96.3(.09)
89. 5 (.09)
92. 5 (.09)
98.8(.09)
91.5(3.3)
88.5(7.6)
92.0(4.8)
96.0(2.8)
94.5(3.8)
87.0(3.4)
94.0(3.5)
93.5(2.2)
survey. Mean values (standard error
significantly different
Total
Body
Length (mm)
Start End
1.2 1.9(.06)ab
2.0(.05)b
2.0(.06)ab
1.8(.06)a
1.8(.05)a
1.9(.05)a
2.0(.05)ab
2.0(.05)b
1.3 3.9(0.1)ab
3.7(0.1)a
3.9(0.1)ab
3.7(0.1)a
3.6(0.1)a
3.6(0.1)a
4.1(0.1)b
4.0(0.1)b
10.1(0.2)e
11.0(0.2)d
12 . 5 ( 0 . 2 ) ab
11.8(0.2)c
11.7(0.2)c
11.9(0.2)bc
12.5(0.2)a
11.9(0.2)c
(p < 0.05).
Antennal
Seoment It)
15.0(0.1)
15.2(0.1)
15.0(0.1)
14.9(0.1)
14.9(0.1)
15.0(0.1)
15.3(0.1)
15.3(0.1)
19.6(0.3)abc
20.0(0.2)ac
20.2(0.3)ac
19.9(0.2)ac
19.5(0.2)bc
18.9(0.3)b
20.2(0.3)ac
20.2(0.2)a
in parentheses)
Mature m
12.6(.08)ab
15.9(.08)ab
6.5(.08)bc
2.8(.08)c
9 . 4 ( . 08 ) abc
7.8(.08)bc
20.7(.08)a
18.2(.08)ab
74. 9 (.04)
83.8(.04)
75.0 (.04)
83.1(.04)
72.1 (.04)
71.5(.04)
71.6(.04)
82.3(.04)
-------
Table 4.35 (Continued).
Test
Survival I % )
Total
Body
Lenoth (nun)
Start End
C. tentans - 10 day
Control
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
60.0*
40.0*
40.0*
80.0*
0.0*
70.0*
30.0*
80.0*
5.6 14.6(1.3)
12.6(1.6)
15.4(1.6)
16.2(1.2)
—
16.8(1.2)
16.2(1.9)
15.3(1.2)
* See Table 4.36 for categorical model statistics.
-------
Table 4.36 Categorical comparison matrix of Chironomus tentans survival data for Saginaw River, MI third
survey. Values above the diagonal separator are probability values and below the separator are the Chi
square values. Probability values with * indicate that significant differences exist in the comparison
between sites (p< 0.05).
Saginaw River Third Survey Stations
01
02
05
06
08
16
24
Control
01
****
****
0.00
3.08
2.67
1.76
0.22
3.08
0.79
02
1.00
****
****
3.08
2.67
1.76
0.22
3.08
0.79
05
0.079
0.079
****
****
7.05
0.26
4.53
0.00
0.92
06
0.102
0.102
0.008*
****
****
5.73
1.79
7.05
4.60
08
0.185
0.185
0.6075
0.017*
****
****
3.02
0.26
0.22
16
0.640
0.640
0.033*
0.181
0.82
****
****
4.53
1.76
24
0.079
0.079
1.00
0.008*
0.607
0.033*
****
****
0.92
Control
0.374
0.374
0.336
0.032*
0.640
0.185
0.336
****
-------
Table 4.38 Recommended threshold concentrations (ng/dry g) for polynuclear aromatic
hydrocarbons and total polychlorinated biphenyls. See Table 4.1 for an explanation of
chemical abbreviations.
Compound
Recommended Threshold
Concentrations
Barrick
et al.
(1988)
(ng/dry g)
USEPA
(1988)
(ng/dry g)
SMS
(1991)
(ng/dry g OC)
Long and
Morgan
(1990)*
(ng/dry g)
Low Molecular Weight PAH
High
Total
Total
Naph
Acnaph
Fluore
Phen
Anth
Molecular Weight PAH
Fluora
Pyrene
BaAnth
Chrys
BbkFlour**
BaPyr
IndPyr
BghiPer
PAH
PCBs
24,
2,
2,
3,
6,
13,
69,
30,
16,
5,
9,
7,
3,
1,
1,
-
1,
000
400
000
600
900
000
000
000
000
100
200
800
000
800
400
—
900
370
99
16
23
1,000 100
220
960
19,000 160
13,000 1,000
13,000 110
110
230
11000 99
33
31
1,330
420 12
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
,000
340-2,100
150-650
35-640
255-1,380
85-960
600-3,600
350-2,200
230-1,600
400-2,800
400-2,500
4,000-35,000
50-400
* The values for each compound indicates
and Effects Range-Median (ER-M).
the range between the Effects Range-Low (ER-L)
** BbFlour and BkFlour are combined and listed as BbkFlour.
-------
Table 4.39 Recommended metals threshold concentrations (/ig/dry g) for sediment.
explanation of chemical abbreviations.
See Table 4.1 for an
Metals
USEPA/
USAGE
(1977)
(^g/dry g)
As 3-8
Cd >6
Cr 25-75
Cu 26-50
Hg 1
Ni 20-50
Pb 40-60
Zn 90-200
Recommended
MOE
(1978)
(jig/dry g)
8.0
1.0
25.0
25.0
0.3
25.0
50.0
100.0
Threshold Concentrations
IEPA
(1984)
(Aig/dry g)
11.0
1.0
23.0
60.0
0.01
38.0
100.0
IJC
(1988)
(tig/dry g)
1.1
0.6
37.1
21.0
0.03
32.8
27.5
120.0
SMS
(1991)
(itg/dry g OC)
57
5.1
260
390
450
410
Long and
Morgan
(1990)*
(jig/dry g)
33-85
5-9
80-145
70-390
0.15-1.3
30-50
35-110
120-270
* The values for each metal indicates the range between the Effects Range-Low (ER-L) and Effects Range-
Median (ER-M).
-------
Table 4.37 Percentage fine particle size (<63 pm) for sediments from all AOCs tested in whole sediment exposures.
Concentrations of low and high molecular weight polynuclear aromatic hydrocarbons (ng/dry g), and total polychlorinated
biphenyls (ng/dry g), and concentrations normalized with percentage total organic carbon for low and high molecular weight
polynuclear aromatic hydrocarbons (fig/dry g OC), and total polychlorinated biphenyls (ng/dry g OC). See Table 4.1 for an
explanation of chemical abbreviations.
Station
Particle
Size (%)
Fines
(<63 ^im)
TOC
(%)
PAHs
(ng/dry g)
Molecular Weight
Low High Sum
PAH /TOC
(ng/dry g OC)
Molecular Weight
Low High
Sum of
PAH/TOC
PCBs
Total
(ng/dry g)
PCB/TOC
(ng/dry g OC)
IH-01-03*
IH-01-04*
IH-01-06*
IH-01-07*
BR-01-01*
BR-01-03*
BR-01-07*
BR-01-08
BR-01-09*
SR-01-03
SR-01-06*
SR-01-10
SR-03-01
SR-03-02
SR-03-05
SR-03-06*
SR-03-08
SR-03-16
SR-03-24
92.2
96.0
96.1
91.8
81.8
91.5
97.8
93.0
95.9
99.3
54.0
53.0
98.9
96.5
88.2
62.0
95.8
92.3
91.9
7.7
5.6
11.6
8.8
8.9
2.0
1.7
1.7
2.2
3.0
2.1
1.0
1.0
1.3
1.4
0.2
3.6
2.9
4.0
19100
9190
30400
401580
12000
3940
1143
580
731
479
552
27
1040
1260
1320
640
1330
1150
1870
* Survival for at least one test species
acceptability criteria were not included
85200
52800
198000
292600
51000
9300
19650
3498
4700
2620
2900
119
2250
2960
3300
1400
3750
2840
6200
1043000
61990
228400
694180
63000
13240
20793
4078
5431
3099
3452
146
3290
4220
4620
2040
5080 v
3990
8070
248.05
164.11
262.07
4563.41
134.83
197.00
67.24
34.12
33.23
15.97
26.29
2.70
104.00
96.92
94.29
320.00
j 36.94
39.66
46.75
was significantly reduced from
(C. riparius 14-d Buffalo River
1106.49
942.86
1706.90
3211.36
573.03
465.00
1155.88
205.76
213.64
87.33
138.10
11.90
225.00
227.69
235.71
700.00
104.17
97.93
155.00
1354.55
1106.96
1968.97
7774.77
707.87
662.00
1223.12
239.88
246.86
103.30
164.38
14.60
329.00
324.62
330.00
1020.00
141.11
137.59
201.75
21160oo
17960oo
27000oo
45460oo
3570oo
5200oo
2520oo
2380oo
2520oo
2380oo
69400oo
4040oo
1140
1450
5190
3670
5890
1530
2780
control. Tests that did not meet ASTM
and C. tentans 10-d Saginaw River (3).
274.81
320.71
232.76
516.59
40.11
260.00
148.24
140.00
114.55
79.33
3304.76
404.00
11.40
11.15
37.07
183.50
16.36
5.28
6.95
test
oo PCBs analyzed: Aroclor 1016, 1221, 1232, 1242, 1248, 1254 and 1260.
0 PCBs analyzed: Aroclor 1242, 1254 and 1260.
-------
Table 4.40 Comparisons of recommended threshold concentrations and test organism
responses from whole sediment exposures. A hit was considered positive (+) if threshold
concentrations were predictive (sediment concentrations exceeded thresholds and toxicity
occurred, or sediment concentrations did not exceed thresholds and no toxicity occurred.
A hit was considered negative (-) if sediment concentrations did not exceed thresholds and
toxicity occurred. A hit was considered a zero (0) if sediment concentrations did exceed
thresholds and no toxicity occurred. Percentages represent proportion of total hits to
the number of AOC stations tested.
Station
SMS
(1991)
Metals
PAHs
PCBs
Long and Morgan
(1990)O
Metals
PAHs
PBCs
Barrick et al.
(1988)
PAHs
PCBs
IH-01
IH-01
IH-01
IH-01
BR-01
BR-01
BR-01
BR-01
BR-01
SR-01
SR-01
SR-01
SR-03
SR-03
SR-03
SR-03
SR-03
SR-03
SR-03
03*
04*
06*
07*
01*
03*
07*
08*
09*
03*
06*
10*
01
02
05
06*
08
16
24
0
0
+
+
+
0
0
Total + (%)
52.6
47.4
89.5
47.4
57.9
84.2
47.4
94.7
Total -
Total 0
(%)
(%)
36
10
.9
.5
52.6
0
0
10.5
42.1
10.5
42.1
0
5.3
10.5
52.6
0
5.
0
3
O Indicates concentrations exceeded the Effects Range-Median.
* Survival or growth for at least one test species was significantly reduced from
control.
-------
Table 4.41 Comparison of test species sensitivity across Areas of Concern. The
percentage hits are based o the number of contaminated sediments that elicited significant
reductions in survival or body length compared to control. A positive indicates a hit and
a negative indicates no hit.
Station
Survival
HA14*
HA28A
CR1400
CT10'
Growth
HA14*
HA28A
CR1400
CT10'
IH-01-03
IH-01-04
IH-01-06
IH-01-07
BR-01-01
BR-01-03
BR-01-07
BR-01-08
BR-01-09
SR-01-03
SR-01-06
SR-01-10
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Hits
Stations
+
+
-f
+
32
100
NA
NA
NA
NA
33
79
+
+
+
+
37
100
+
+
+
+
NA
NA
NA
31
84
NA
NA
29
89
NA
NA
NA
NA
+
+
+
33
79
NA
NA
NA
NA
13
79
* H. azteca 14 day exposure.
* H. azteca 28 day exposure.
°° C. ripariua 14 day exposure.
' C. tentans 10 day exposure.
NA Indicates not applicable.
+
+
NA
NA
NA
NA
NA
NA
31
68
HA14*
HA28A
CR14"
CT10'
HA14* 1 HA28~
CR14°°
CT10'
-------
Appendix 4.1 Concentrations of metals (/ig/dry g) in control sediment. See
Table 4.1 for an explanation of chemical abbreviations.
Metal (/ig/dry g)
Ag As Cd Cr Cu Hg Mn Ni Pb Se Zn
0.117 8.9B 0.18 52B 19B 0.022 675B 19.4B 19.6B <0.16 52.5B
B Indicates that data was not available for procedural blank.
< Indicates compound not detected at detection limit shown.
-------
Appendix 4.2 Concentrations of polynuclear aromatic and other semivolatile compounds (ng/dry g) for control
sediment. Ranges for samples analyzed in replicate are shown in parentheses. See Table 4.1 for an
explanation of chemical abbreviations.*
Polynuclear Aromatic Hydrocarbon (ng/dry g)
1-4
DCB
NA
DThiop
<5.24
Chrys
4.46
(4.19-4.72)
Naph Acenaphl
B <5.77
Phen Anth
B <4.77
DnOPh BbFlour
NA 8.52
(8.05-8.99)
Acenaph
<10.26
Fluora
8.495
(7.25-9.74)
BkFlour
<3.23
2-M
Naph
NA
Pyrene
B
BaPyr
<4.04
DMPh
NA
BBPh
NA
IndPyr
<2.54
DBF Fluore
NA <7.37
BaAnth BisPh
<3.76 NA
D(a,h)Anth BghiPer
<3.22 <2.85
* Other organic chemical analyses (Aroclor 1242, 1248, 1254, and 1260; and pesticides) were conducted for
the control sediment. All pesticides and Aroclors were not detected at or above the detection limits
used for the contaminated sediments from all AOCs. Total organic carbon was 1.02 percent.
NA Indicates that data is not available.
B Indicates that analyte was detected in the method blank associated with sample.
< Indicates that compound was not detected at detection limit shown.
-------
CHAPTER 5; BENTHIC COMMUNITY STRUCTURE EVALUATIONS
Canfield, T.J., T.W. La Point, M.C. Swift1, G.A. Burton2, J.A. Fairchild,
and N.E. Kemble.
1 University of Minnesota, Monticello, MN.
2 Wright State University, Dayton, OH.
INTRODUCTION
Contaminated sediments are a major source of pollution in the United
States and represent a continual threat to all levels of the aquatic ecosystem
(Sorensen et al. 1977, Landrum and Robbins 1990). Sediments act as a
repository for a whole array of organic and inorganic contaminants, and can
accumulate the contaminants to extremely high concentrations (Shimp et al.
1971, Oschwald 1972, Medine and McCutcheon 1989). Benthic macroinvertebrates
are integrally tied to the sediment and therefore are continuously exposed to
contaminants in sediments.
The entire aquatic ecosystem is linked together by virtue of
interdependent trophic levels. Therefore it is no longer adequate to study
single components of the ecosystem when making assessments of sediment
toxicity (Burton 1991). Complete ecological assessments of sediment toxicity
require the use of resident biota as indicators of sediment quality. For the
assessment to be successful, closely integrated biological, chemical, and
physical data are required. Sediments integrate historical water quality,
hence, the spatial and temporal distribution of resident organisms can reflect
the degree to which chemicals in the sediments are toxic. Field surveys of
invertebrates provide an essential component of biological assessments of
toxicity associated with contaminated sediments and have several advantages:
(1) indigenous benthic organisms complete all or most of their life cycles in
the aquatic environment and serve as continuous monitors of sediment quality,
(2) benthic invertebrates are relatively sedentary and are representative of
local conditions, (3) macroinvertebrates are relatively easy to collect and
are abundant and ubiquitous across a broad array of sediment types, (4) a
field assessment of natural populations can be used to screen potential
sediment contamination, (if a spatial gradient of contamination exists,
extrapolations of inter-species sensitivity, water quality differences, or
chemical interaction [additivity, antagonism, or synergism] are not
necessary), and (5) results of an assessment of indigenous populations are
biologically interpretable, which should quantify resource damage in a manner
more easily understood by managers, regulators and the general public (Cook
1976, Pratt and Coler 1976, Davis and Lathrop 1989).
This chapter describes the results of quantitative invertebrate surveys
taken simultaneously with sediment collection from stations within the Areas
of Concern (AOC) sampled for the GLNPO ARCS project. The objective of the
survey is to describe species distributions and abundances across 3 AOCs:
Indiana Harbor, Buffalo River, and Saginaw River. An attempt will be made to
relate the observed species abundance and community structure to sediment
metal concentrations and other physical and chemical parameters. This
information, when analyzed in the context of measures of sediment toxicity
will provide a more complete representation of the effects of in-situ
contaminants on the benthic invertebrate communities.
5-1
-------
MATERIALS AND METHODS
The GLNPO ARCS Toxicity-Chemistry Work Group (Table 1.1) selected
stations for whole sediment toxicity and benthic macroinvertebrate assessments
based on the expected presence of contaminants which could be deleterious to
resident aquatic biota. Three areas of concern (AOC), Buffalo river (BR-01),
Indiana Harbor (IH-01), and Saginaw River (SR-01) were sampled once in 1989;
Saginaw River (SR-03) was sampled a second time in June 1990 (See Table 2.1
for a complete listing of the stations sampled). Seven stations were sampled
in Indian Harbor and Saginaw River (first and third survey) while 10 stations
were sampled in Buffalo River. A standard ponar dredge (529 cm2 area; 4.69 L
volume) was used to collect benthic invertebrates and bulk sediments at each
station.
Sediments were analyzed for a variety of chemical and physical
characteristics. Sediment chemical characteristics included (1) total metals,
(2) organometals, (3) simultaneously extracted metals (SEM) and acid volatile
sulfide (AVS), (4) chlorinated pesticides, (5) polychlorinated biphenyls
(PCB), (6) select polynuclear aromatic hydrocarbons (PAH), and (8)
polychlorinated dioxins and furans. Sediment physical characteristics
included (1) total organic carbon, (2) inorganic carbon, and (3) sediment
particle size. For additional information on chemical and physical
characteristics of the sediments, see chapter 4 of this document.
A total of 155 benthic grab samples (about 5 grabs/station) were
collected from all AOCs. Artificial substrates were put at 5 stations in
Buffalo River, 4 stations at Indiana Harbor and 6 stations at Saginaw River.
All 155 benthic samples were collected before sediment samples for chemistry
and toxicity testing in an effort to minimize potential disturbance of the
sediments and associated invertebrates. The 5 replicate Ponar samples were
taken at each station within a 100 m2 area. Each collected sample was sieved
through a 500 jun brass screen using station water for rinsing. Material
retained by the sieve was rinsed into 500-mL Ball glass jars and preserved
with 10% buffered formalin. All jars were filled to the top with formalin to
prevent damage to the benthic invertebrates during transport. Before
shipment, jars were placed into a cardboard box with dividers to protect
individual samples. Each box was triple bagged with 114 liter plastic sacks
and put into a larger box and surrounded by styrofoam packing. Each container
was labeled with an 11 digit number describing designation codes for
collection site, survey number, transect number, station number, sample type,
replicate, and sample fraction. The 11 digit sample number was recorded in
the field log book, on the sample container and in the sample container on a
piece of paper with a soft lead (#2) pencil.
Samples were received at the National Fisheries Contaminant Research
Center (NFCRC) in Columbia, MO, and immediately inspected for shipping damage.
Samples broken in shipping were recorded and discarded. Before sorting,
samples were rinsed thoroughly with tap water to remove formalin and excess
silt or mud. The "clean" samples were drained of excess water, returned to
the original jars, filled with 95% ethanol and allowed to soak for at least 24
h to facilitate extraction of any volatiles. After the 24 h extraction
period, each sample was rinsed again with tap water to remove the ethanol and
volatiles. The samples were placed into a 4-L wide-mouth jar, agitated with
tap water to float the invertebrates and lighter detrital material while
leaving the snails, clams and heavier material in the bottom of the jar.
5-2
-------
Aliquots of the sample were sequentially removed from the jar to sort the
benthic invertebrates. This continued until the entire sample had been
sorted. Approximate sorting time ranged from 3 to 20 h/sample.
A 4X or greater binocular dissecting microscope was used to sort the
samples. Organisms were sorted and enumerated into the following orders or
families: Oligochaeta, Chironomidae, Bivalvia, Gastropoda, Ephemeroptera,
Odonata, Plecoptera, Hemiptera, Megaloptera, Tricoptera, Coleoptera, Diptera,
Hirudinea, and Amphipoda. These samples were used to estimate
macroinvertebrate abundance (number/m ), species composition, and taxa
richness. Taxonomic identification was made to the lowest level for each
group by NFCRC personnel using published taxonomic keys (Wiederholm 1983;
Merritt and Cummins 1984; Pennak 1989; Thorp and Covich 1991). Oligochaeta
were identified to genus and species (when possible) by Mark Wetzel at
Invertaxon, Urbana, IL. Mollusca were identified to genus and species by Jeff
Garner at the Aquatic Resources Center, Franklin, TN. Chironomidae samples
were identified to genus by NFCRC personnel and confirmations of
identifications were made by Dr. Leonard Ferrington, University of Kansas,
Lawrence, KS.
Chironomidae were examined for deformities in mouthpart structures.
These deformities consisted of various types of asymmetry, missing teeth,
extra teeth, fusion among various teeth, and labial separation described by
several investigators (Saether 1970; Hamilton and Saether 1971; Hare and
Carter 1976; Warwick et al 1987; Warwick 1989). Individual Chironomidae were
mounted on slides and examined for deformities in the mentum and ligula
(Tanypodinae). Occurrence of deformities was expressed as a proportion of the
total number of Chironomidae at each station.
Artificial substrates were constructed from a 3M™ synthetic mesh and
stainless steel wire rotisserie chicken baskets (Stauffer et al. 1976). Each
substrate consisted of 5 pieces of mesh (20X20cm) folded in half and placed
beside each other in a chicken basket. The rotisserie baskets were 26 cm in
length, 17 cm in diameter and 53.34 cm in circumference (Figure 5.1). The
baskets were wired shut and three baskets were wired to a cinder block at each
sampling station. The baskets were on six foot wires and sat horizontally on
the bottom near the cinderblock. A wire was attached to the cinderblock and
the other end was connected to a recognizable land mark on shore to facilitate
retrieval of the artificial substrates.
Artificial substrates were put at 5 stations in Buffalo River (October
1989), 4 stations in Indiana Harbor (August 1991), and 6 stations at Saginaw
River (survey 3, June 1990). The artificial substrates were incubated at
stations for 30 d. The substrates were lifted to the surface and then placed
into plastic dishpans. The mesh substrate material was removed and placed in
4-L wide-mouth jars and preserved in 4% buffered formalin. Water and mud
remaining in the dishpans was poured through a 250 /un mesh sieve and anything
retained on the sieve was preserved with the synthetic mesh.
Samples were rinsed in the laboratory through a 250 pm mesh sieve and
then each piece of mesh was unraveled under water in a dishpan. Sediment and
organisms were retained in the water, sieved (250 fm mesh) and stored in 70%
ethanol; the mesh was discarded. The entire preserved sample was placed in a
pan for sorting at 12X magnification. A 100 organism sub-sample was removed
from the sample; an attempt was made to sub-sample organisms in proportion to
their abundance in the sample. At Indiana Harbor Station 3, each sample
contained thousands of tiny (<500 urn), just-settled zebra mussels fDreissena
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sp.). A 100 organism sub-sample of invertebrates other than zebra mussels was
picked to determine other organisms present at Station 3 of Indiana Harbor.
At all stations, organisms too rare to be included in the 100 animal sub-
samples were qualitatively recorded.
Statistical analysis was performed with the Statistical Analysis System
(SAS) computer package for personnel computers (Statistical Analysis System
1988). Relations among the benthic invertebrate abundance within and among
areas of concern were analyzed with a nested analysis of variance (ANOVA)
(Snedecor and Cochran 1980). Comparisons between benthic invertebrate
abundance and physical and chemical data were compared with correlation
analysis and multivariate regression statistics. If not reported, statements
of statistical significance imply p< 0.05.
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RESULTS
BENTHIC INVERTEBRATE ABUNDANCE AND COMMUNITY STRUCTURE IN GRAB SAMPLES
Indiana Harbor
Samples were collected at Indiana Harbor in August, 1989. Benthic
invertebrate abundance (number/m2) for the entire community (Table 5.1) at
each station ranged from 609/m2 at IH-01-07 to a maximum of 493,917/m2 at IH-
01-10. Except for IH-01-10, total abundance values were less than 7,000/m2,
with abundance values at 4 of the 7 stations less than 4,000/m2 (Table 5.1).
Within the benthic invertebrate community the Oligochaeta were numerically
dominant at all stations sampled, bivalves comprised the majority of the
remaining community in 3 of the 7 stations. Oligochaeta abundance accounted
for 91% to 100% of the community across all stations sampled, with the
remainder of the benthic community abundance coming from the Bivalvia,
Hirudinea and Chironomidae (Table 5.2).
Oligochaeta Abundance—
Oligochaeta abundance ranged from 552/m2 at IH-01-07 to 493,887/m2 at
IH-01-10 (Table 5.1). As with total abundance, Oligochaeta abundance values
were all less than 6,000/m2 with the exception of IH-01-10 and abundance was
less than 3,000/m2 at 40% of the stations (Table 5.1). The Oligochaeta
community was made up entirely of the family Tubificidae (Table 5.3). The
Oligochaeta were comprised of 1 family, 5 genera and 8 species.
Limnodrilus sp. was the most common genera occurring at all 7 stations
sampled. Quistadrilus sp. was present at 6 stations with Aulodrilus sp. and
Ilyodrilus sp. present at 2 stations each. Tubifex sp. was present at only 1
station. The Limnodrilus sp. were represented by 3 species (L. cervix,
L. hof fmeisteri. L.. udekemianus). Limnodrilus hof fmeisteri occurred at all 7
stations, with L. cervix occurring at 5 stations and L. udekemianus occurring
at 2 stations. The genera Aulodrilus sp. was represented by 2 species, A.
limnobius and A. pluriseta. Aulodrilus pluriseta occurred at two stations and
A. limnobius occurred at 1 station. The remaining genera were represented by
1 species each (Ilyodrilus templetoni. Quistadrilus multisetosus).
Chironomidae Abundance—
Chironomidae abundance ranged from none in samples from several stations
to 8/m2 at IH-01-10 (Table 5.1). The Chironomidae community was comprised of
1 subfamily (Orthocladinae) and 1 genera (Cricotopus sp.) (Table 5.4). Only 2
individuals were found among all the stations sampled at Indiana Harbor.
Mollusca Abundance—
The Mollusca were represented by 1 class (Bivalvia), 1 family, 3 genera,
and at least 3 species (Table 5.5). Mean Bivalvia abundance ranged from none
in samples from several stations to 8/m2 at IH-01-03 and IH-01-07 (Table 5.1).
Muaculium sp. and Pisidium sp. were the most frequently observed genera each
occurring at 2 of 10 stations. Sphaerium sp. occurred only at IH-01-07. The
abundance of these genera was extremely low when they were present.
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Buffalo River
Samples were collected at Buffalo River in October 1989. Benthic
invertebrate abundance (number/m2) for the entire community (Table 5.1) at
each station ranged from a low of 2,294/m2 at BR-01-09 to a high of 19,418/m2
at BR-01-06. Total abundance values at all stations were less than 20,000/m2,
with abundance values at 7 of the 10 stations less than 10,000/m2 (Table 5.1).
Within the benthic invertebrate community the Oligochaeta were numerically
dominant at all stations sampled, Chironomidae comprised the majority of the
remaining community in 7 of the 10 stations. Combined Oligochaeta and
Chironomidae abundance accounted for 92% of the community at BR-01-04 to 99%
of the total benthic invertebrate community at several stations, with the
remainder of the benthic community abundance coming from the Gastropoda,
Bivalvia and Hirudinea (Table 5.6).
Oligochaeta Abundance—
Oligochaeta abundance ranged from 1,984/m2 at BR-01-09 to 19,157/m2 at
BR-01-06. As with total abundance, Oligochaeta abundance values were all less
than 20,000/m2 with abundance less than 9,000/m2 at 70% of the stations (Table
5.1). The Oligochaeta community was dominated by the family Tubificidae to
almost entire exclusion of other families (Table 5.7). The Oligochaeta were
comprised of 2 families (Naididae, Tubificidae), 5 genera and 7 species.
The Naididae were represented at 1 station BR-01-01 by 1 species (Dero
diqitata). The Tubificidae were present at all stations. Limnodrilus sp. was
the most common genera occurring at all 10 stations followed by Quistadrilus
sp. at 6 stations and Dero sp., Aulodrilus sp. and Tubifex sp. occurring at 1
station each. The Limnodrilus sp. were represented by 3 species. Limnodrilua
hoffmeisteri was found at 9 stations, L. cervix was found at 5 stations and L.
claperedianus at 1 station. The remaining genera were represented by 1
species each (Aulodrilus pioueti, Quistadrilus multisetosus. Tubifex) .
Chironomidae Abundance—
Chironomidae abundance ranged from 11/m2 at BR-01-05 to 2,771/m2 at BR-
01-10 (Table 5.1). Except for BR-01-10, all Chironomidae abundance was less
than 500/m2. The Chironomidae community was comprised of 3 subfamilies
(Chironomini, Tanipodinae, Tanytarsini) and 11 genera (Table 5.8).
The Chironomini were represented by 7 genera and were present at all
stations except BR-01-05 and BR-01-07 (Table 5.8). Cryptochironomus sp. was
the most frequently observed genera and occurred at 8 of 10 stations sampled.
Chironomus sp. and Cladopelma sp. were found at 4 stations, Dicrotendipes sp.
and Polvpedilum sp. occurred at 2 stations, and Microchiromomus sp. and
Glyptotendipes sp. each occurred at 1 station.
The Tanypodinae were represented by 3 genera and occurred at all
stations sampled. Procladius sp. was found at all stations. Coelotanvpus sp.
occurred at 3 of 10 stations with Tanvpus sp. occurring at 1 station.
The Tanytarsini were found at only 1 station, BR-01-10. The Tanytarsini
were represented by the genus Tanvtarsus.
Station BR-01-10 had the highest taxa richness with 3 families and 8
genera (Table 5.8). BR-01-01 and BR-01-09 had the next highest taxa richness
5-6
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with 2 families and 7 genera followed by BR-01-09 with 2 families and 6 genera
represented. The taxa richness of the remaining stations was 4 or less.
Mollusca Abundance—
The Mollusca were represented by 2 classes (Gastropoda and Bivalvia), 6
families, 10 genus and a minimum of 11 species (Table 5.9). Mean Gastropoda
abundance ranged from 0 at several stations to 378/m at BR-01-04. Mean
Bivalvia abundance ranged from 0 at BR-01-10 to 217/m2 at BR-01-03. Two of
the Gastropoda in the Buffalo River samples, Valvata lewisi (Currier, 1968)
and Cincinnatia cincinnatiensis (Anthony, 1849) are reported uncommon in New
York state (Eileen Jokinen, University of Connecticut, Storrs, CT, personal
communication).
The Gastropoda were represented by 4 families, 4 genera and 5 species
(Table 5.9). The Gastropoda were present at 6 of 10 stations sampled.
Valvata lewisi was the most frequently observed species occurring at 5 of 10
stations. Cincinnatia cincinnatiensis was the next most abundant species
occurring at 3 of 10 stations sampled. Bithynia tentaculata and Laevapex
fucus occurred at 2 stations each with Valvata tricarinata occurring at only 1
station. The abundance of these three species was low.
Station BR-01-04 had the highest taxa richness with 4 families, 4
genera, and 4 species represented. BR-01-03 had the next highest taxa
richness with 3 families, 3 genera, and 4 species present. Samples from
stations BR-01-02, BR-01-05, BR-01-08, BR-01-09, and BR-01-10 did not contain
Gastropoda.
The Bivalvia were represented by 2 families, 6 genera and at least 7
species (Table 5.9). The Bivalvia were present at 8 of 10 stations sampled.
Musculium sp. and Pisidium sp. were the most frequently observed taxa
occurring at 7 of 10 stations sampled. Sphaerium sp. was the next most
frequently observed species occurring at 6 of 10 stations. The Sphaeridae
were the most dominant family at any of the stations.
The Unionidae were represented by 2 genera and 3 species (Table 5.9).
The Unionidae were found at 4 stations. The abundance of the Unionidae were
relatively small compared to the Sphaeridae.
Saqinaw River (first survey)
Samples were collected from Saginaw River (first survey) in December,
1989. Benthic invertebrate abundance (number/m ) for the entire community
(Table 5.1) at each station ranged from 888/m at SR-01-10 to a maximum of
7,129/m at SR-01-03. Total abundance values at all stations were less than
8,000/m . Abundance values were evenly distributed across the stations.
Within the benthic invertebrate community, the Oligochaeta were numerically
dominant at all stations sampled, Chironomidae comprised the majority of the
remaining community at 6 of the 7 stations. Combined Oligochaeta and
Chironomidae abundance accounted for 99% to 100% of the total benthic
invertebrate community at all stations, with the remainder of the benthic
community abundance coming from the Bivalvia, Diptera, Coleoptera and
Tricoptera (Table 5.10).
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Oligochaeta Abundance—
Oligochaeta abundance ranged from 813/m2 at SR-01-10 to 6,974/m2 at SR-
01-03. Oligochaeta abundance values were all less than 7,000/m2 (Table 5.1).
The Oligochaeta community was dominated by the family Tubificidae to almost
entire exclusion of other families (Table 5.11). The Oligochaeta were
comprised of 2 families, 5 genera, and 8 species.
The Naididae were represented at 1 station SR-01-09 by 1 species (Dero
digitata). The Tubificidae were present at all stations. Limnodrilus sp. was
the most common genera occurring at all 7 stations, followed by Ilyodrilus sp.
at 5 stations, Quistadrilus sp. at 2 stations and Tubifex sp. at 1 station.
The Limnodrilus sp. were represented by 4 species (L. cervix, L.
claparedianus. L. hoffmeisteri. L. maumeensis). Limnodrilus hoffmeisteri and
L. cervix were found at all 7 stations, with L. maumeensis occurring at 2
stations and L_. c 1 aparedianus at 1 station. Ilyodrilus templetoni occurred at
5 of 7 stations with Quistadrilus multisetosus at 2 stations and Tubifex
tubifex 1 station.
Chironomidae Abundance—
Chironomidae abundance ranged from 11/m at SR-01-06 to 363/m at SR-01-
09 (Table 5.1). The Chironomidae community was comprised of 3 subfamilies
(Chironominae, Tanipodinae, Orthocladinae) and 6 genera (Table 5.12).
The Chironomini were represented by 4 genera and were present at 5 of 7
stations. Crvptochironomus sp. was the most frequently occurring genera and
was present at 3 of 7 stations. Of the remaining genera only the Chironomus
sp. occurred at more than 1 station (SR-01-02, SR-01-09).
The Tanypodinae were represented by 1 genera (Procladius sp.) and
occurred at all stations except SR-01-06. Procladius sp. was the most
abundant genera at all stations it was in occurrence.
The Orthocladinae were represented by the genera Cricotopus sp. and
occurred only at SR-01-02. Cricotopus sp. abundance only made up a small
proportion of the abundance at SR-01-02.
Station SR-01-02 had the highest taxa richness with 3 families and 4
genera represented (Table 5.12). Station SR-01-09 had the next highest taxa
richness with 2 families and 3 genera represented, while SR-01-07 and SR-01-10
had only 2 families and 2 genera represented. The remaining stations had only
1 family and 1 genus present. The abundance of the genera at SR-01-10 was low
compared to most stations in Saginaw River.
Mollusca Abundance—
The Mollusca were represented by 1 class (Bivalvia), 1 family, 3 genus
and at least 3 species (Table 5.13). Mean Bivalvia abundance ranged from 0 at
several stations to 16/m at SR-01-06 (Table 5.1). Sphaerium sp. was the most
frequently observed genera each occurring at 2 of 10 stations. Musculium sp.
and Pisidium sp. occurred only at SR-01-06. The abundance of all these genera
was extremely low.
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Saainaw River (third survey)
Samples were collected at Saginaw River (third survey) in June, 1990.
Benthic invertebrate abundance (number/m2) for the entire community at each
station ranged from 321/m2 at SR-03-06 to a maximum of 7,152/m at SR-03-01
(Table 5.1). Total abundance values at all stations were less than 8,000/m2,
with abundance values at 3 of the 7 stations less than 2,000/m2. Within the
benthic invertebrate community the Oligochaeta were numerically dominant at
all stations sampled, Chironomidae comprised the majority of the remaining
community in all 7 stations. Combined Oligochaeta and Chironomidae abundance
accounted for 99% of the community at several stations to 100% of the total
benthic invertebrate community at SR-03-01 and SR-03-06, with the remainder of
the benthic community abundance coming from the Bivalvia, Diptera and
Coleoptera (Table 5.14).
Oligochaeta Abundance—
Oligochaeta abundance ranged from 302/m2 at SR-03-06 to 4,944/m at SR-
03-01. Oligochaeta abundance values were all less than 5,000/m with
abundance less than 1,000/m2 at 40% of the stations (Table 5.1). The
Oligochaeta community was dominated by the family Tubificidae to almost entire
exclusion of other families (Table 5.15). The Oligochaeta were comprised of 2
families, 5 genera and 7 species.
The Naididae were represented at 2 stations (SR-03-05 and SR-03-16).
Station SR-03-05 had 1 species (Dero dicritata). while the Naidid at SR-03-16
was unidentifiable. The Tubificidae were present at all stations.
Limnodrilus sp. was the most common genera occurring at all 7 stations,
followed by Ilvodrilus sp. at 4 stations, Aulodrilus sp. at 2 stations, and
Quistadrilus sp. and Tubifex sp. at 1 station. The Limnodrilus sp. were
represented by 3 species (L. cervix. L. hoffmeisteri, L. maumeensis) and were
found at all 7 stations. Ilvodrilus templetoni occurred at 4 of 7 stations,
Aulodrilus pluriseta occurred at 2 of 7 stations with Quistadrilus
multisetosus occurring at 1 station.
Chironomidae Abundance—
Chironomidae abundance ranged from 15/m2 at SR-03-06 to 2,204/m2 at SR-
03-01 (Table 5.1). The Chironomidae community was comprised of 3 subfamilies
(Chironominae, Tanipodinae, Tanytarsini) and 7 genera (Table 5.16).
The Chironomini were represented by 3 genera and occurred at all
stations (Table 5.16). Chironomus sp. was the most frequently observed genus
and occurred at all 7 stations. Cryptochironomus sp. was found at 5 of 7
stations and Microchironomus sp. occurred at 2 stations.
The Tanypodinae were represented by 3 genera and occurred at all
stations sampled. Procladius sp. was found at all stations while Tanvpus sp.
occurred in low abundance at SR-03-08 and SR-03-24 and Coelotanypus sp.
occurred only at SR-03-08.
The Tanytarsini were represented by Tanvtarsus sp. and occurred only at
SR-03-08. Tanvtarsus sp. constituted a relatively small proportion (<5%) of
the Chironomidae community at SR-03-08.
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Station SR-03-08 had the highest taxa richness with 3 families and 7
genera represented. Stations SR-03-16 and SR-03-24 had the next highest taxa
richness with 2 families and 4 genera present.
Mollusca Abundance—
The Mollusca were represented by 1 class (Bivalvia), 1 family, 1 genus
and 1 species (5.17). Mean Bivalvia abundance ranged from none in samples
from several stations to 11/m at SR-03-05. Musculium sp. (Sphaeridae) was
the only genus of Bivalvia collected at any of the stations. Musculium sp.
was collected at 3 of 7 stations and abundances were extremely low.
COMPARISON AMONG AOCs
Mean total abundance values ranged from 2,783/m in Saginaw River
(survey 3) to 9,323/m2 at Buffalo River (Table 5.18). Median values for total
benthic invertebrate abundance ranged from 2003/m2 in Indiana Harbor (station
10 deleted) to 8,108/m in Buffalo River samples. Indiana Harbor station 10
was deleted from some comparisons because inclusion of IH-01-10 drastically
skewed the data from Indiana Harbor and was not comparable to the other AOCs.
Deleting station IH-01-10 from certain comparisons was justified because
station IH-01-10 was on the upstream side of a low clearance bridge and
therefore did not receive the heavy disturbance caused by boat traffic common
at all other stations. Total invertebrate abundance was in general lowest at
Indiana Harbor and highest at Buffalo River.
Comparisons between the three AOCs show that overall mean Oligochaeta
abundance ranged from 2,429/m2 at Saginaw River (survey 3) to 8,726/m2 at
Buffalo River (Table 5.18). Median values for Oligochaeta abundance ranged
from 1,994/m2 at Indiana Harbor (station 10 deleted) to 7,333/m2 at Buffalo
River again indicating that Oligochaeta abundance was generally lowest at
Indiana Harbor and highest at Buffalo River.
The Oligochaeta community across all areas of concern was comprised of 2
families, 6 genera, and 12 species. All AOCs except Indiana Harbor had
representatives of the Naididae. Buffalo River, Indiana Harbor, and Saginaw
River (survey 1) had representatives of the genus Tubifex sp. present, while
Tubifex sp. was entirely absent from the Saginaw River (survey 3) samples.
All AOCs had similar numbers of genera and species present in the samples.
Overall mean Chironomidae abundance ranged from none in samples from
Indiana Harbor (station 10 deleted) to 465/m2 in Saginaw River (survey 3).
Chironomidae abundance was similar between Buffalo River and Saginaw River
(survey 3), but were not found in Indiana Harbor (station 10 deleted) sediment
samples. In Saginaw River Chironomidae abundance was higher during survey 3
(June 1990) compared to survey 1 (December 1989) (Table 5.18).
The Chironomidae community across all AOCs was comprised of 4
subfamilies (Chironominae, Tanipodinae, Tanytarsini, Orthocladinae) and 12
genera. Buffalo River had the highest taxa richness with 10 of the eleven
genera present. Saginaw River had 7 genera of Chironomidae in the third
survey and 6 genera in the first survey. Three genera were similar between
the two Saginaw river surveys, with Saginaw River having 4 genera in the third
survey which were not present in the first survey. Indiana Harbor had 2
Chironomidae (Cricotopus sp.) present at Station 10. There were no
Chironomidae at any of the other Indiana Harbor stations.
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The Mollusca were represented by 2 classes (Gastropoda and Bivalvia), 6
families, 9 genera, and at least 11 species. Buffalo River contained the
highest taxa richness with all the family, genera and species represented.
Indiana Harbor and Saginaw River (survey 1) had a relatively depauperate
community with only 1 family (Sphaeridae) and 3 genera found in these areas.
Saginaw River (third survey) had only 1 family (Sphaeridae) and 1 genus
present.
Comparisons of mean abundance between stations within an AOC
demonstrated no consistent trends (Table 5.19). Buffalo River and Indiana
Harbor had the most consistent pattern in the Oligochaeta and total abundance.
Oligochaeta and total abundance at Buffalo River tended to be highest in the
middle numbered stations. Oligochaeta and total abundance at Indiana Harbor
(station 10 excluded) tended to be higher in the low numbered stations and
lowest in the higher numbered stations. Discrepancies in the pattern occurred
with the 2 upper stations and the lowest station at Buffalo River and station
10 at Indiana Harbor. There was no recognizable pattern with the Saginaw
River stations. Chironomidae abundance demonstrated no pattern with respect
to stations at any of the AOCs.
BENTHOS-CHEMISTRY COMPARISONS IN GRAB SAMPLES
Comparisons between measured concentrations of metals and
macroinvertebrate abundance estimates using correlation analysis indicated
that no single metal explained a significant amount of the variation
associated with macroinvertebrate abundance at p< 0.05. This result was the
same regardless of whether the data was compared across AOCs or across
stations within an AOC.
Comparisons between macroinvertebrate mean total abundance and total
simultaneously extracted metals (SEM, weak acid extracts of Cd, Cr, Cu, Ni,
Pb, and Zn), total polyaromatic Hydrocarbons (PAH, normalized to total organic
carbon) and total Polychlorinated Biphenyls (PCB, normalized to total organic
carbon) demonstrated a consistent pattern of decreasing abundance with
increasing contaminant concentration (Figures 5.2 to 5.4).
Relations between SEM and mean Oligochaeta abundance demonstrated a
downward curvilinear plot (Figure 5.5). Below I /unol/dry g Oligochaeta
abundance seems to be independent of SEM concentration. Between 1 /unol/dry g
and 4 /unol/dry g Oligochaeta abundance tended to increase and low Oligochaeta
abundance was associated with SEM concentrations above 4 /unol/dry g.
Similar relations are observed between the SEM and mean Chironomidae,
mean Bivalvia and mean Gastropoda abundance (Figure 5.6 to 5.8). The
threshold was about 1 /unol/dry g.
Comparisons of total PAH's and mean Oligochaeta abundance demonstrated
at concentrations below 2,000 /xg/dry g Oligochaeta abundance seems to be
independent of PAH concentration (Figure 5.9) Above 2,000 /tg/dry g PAH
concentration was associated with low Oligochaeta abundance. Similar
relations were observed between total PAH's and mean Chironomidae and mean
Bivalvia abundance (Figure 5.10 and 5.11). Chironomidae abundance tended to
be reduced at PAH concentrations around 500 /*g/dry g. Bivalvia abundance
tended to be lower above 2,000 /tg/dry g, similar to the response of the
Oligochaeta. There was insufficient data to make any statement concerning the
response of Gastropoda abundance to PAH levels (Figure 5.12).
5-11
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Comparisons of total PCB and mean Oligochaeta abundance demonstrates a
downward curvilinear plot with Oligochaeta abundance being independent of PCB
concentration below 500 /*g/dry g (Figure 5.13). Above 500 jig/dry g PCB,
Oligochaeta abundance tended to be lower. Similar relations were observed
between total PCB concentration and mean Chironomidae and mean Bivalvia
abundance (Figures 5.14 and 5.15). Chironomidae and Bivalvia abundance tended
to be more variable at concentrations of PCB's below 250 fig/dry g.
Concentrations above 500 /tg/dry g were associated with decreased Chironomidae
and Bivalvia abundance. There was insufficient data to make any statements
about Gastropoda/PCB relations (Figure 5.16).
DEFORMITIES IN CHIRONOMIDAE FROM GRAB SAMPLES
Chironomidae were examined for deformities in the mentum and ligula
(Prodadius sp.). Deformities in the Chironomidae larvae consisted of various
types of asymmetry, missing teeth, extra teeth and fusion among various teeth.
Mouth part deformities in the Chironomidae community ranged from a low of 0%
at 2 stations BR-01-05 and SR-01-06 to a maximum of 100% at IH-01-10 (Table
5.20). On average the lowest percentage of deformities was from Buffalo River
samples (7.0 %), followed by Saginaw River first survey (12.7%), Saginaw River
third survey (16.7) and Indiana Harbor (100%). Deformities were present in 19
out of 42 replicate at Buffalo River, 17 out of 29 at Saginaw River survey 1,
23 out of 30 at Saginaw River survey 3 and 2 for 2 at Indiana Harbor.
ARTIFICIAL SUBSTRATES VS. GRAB SAMPLES
The ponar grab samples and the artificial substrates collected different
taxa at all the AOCs. The ponar grab samples collected an infaunal community
comprised primarily of Oligochaeta and Chironomidae (Table 5.1). Gastropoda,
Bivalvia, Hirudinea and rare occurrences of other aquatic insect larvae made
up a very small percentage of the grab samples at all the stations.
Artificial substrates collected a much different fauna than the grab
samples (Figure 5.17 to 5.19). The artificial substrate samples primarily
collected the epifaunal community. Artificial substrates in Indiana Harbor
station 3 were almost entirely Dreissena sp. (zebra mussels) (Table 5.17).
Along with the Dreissena sp., Station 3 contained largely Turbellaria,
Oligochaeta, and Amphipoda. The community at Station 3 also included
Cordvlophora sp., Hirudinea, Decapoda, and Cyclopoid and Harpacticoid
copepods. The number of Dreissena sp. decreased substantially at Indiana
Harbor Station 4. Turbellaria and Oligochaeta made up the rest of the
community. Throughout the Indiana Harbor samples, occasional Zygoptera
nymphs, Hirudinea and Bivalvia were also present. At Station 6 there were
relatively few animals, but what was there was predominantly Oligochaeta.
The Buffalo River stations were dominated by Amphipoda and Isopoda with
the remainder of the community comprised of Oligochaeta, Chironomidae, and
Turbellaria (Table 5.18). Buffalo River station 8 had large numbers of
Turbellaria (Flatworms). Oligochaeta abundance collected with the artificial
substrates at the Buffalo River stations was always low.
The macroinvertebrate community in Saginaw River was dominated by
Amphipoda (Garomarus sp.), Chironomidae and Coelenterata (Hvdra sp.) (Table
5.19). Amphipoda were most abundant at Saginaw River Station 2; Chironomidae
5-12
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at stations 6, 8, and 16; Hydra sp. at Station 5; and Station 24 had
relatively even abundances of Amphipoda and Chironoenidae.
The density of animals in the artificial substrates was an order of
magnitude greater than the density of animals in the grab samples at Saginaw
River (Figure 5.20). Similar data was not available to compare the results at
Buffalo River and Indiana Harbor.
VARIATION IN BENTHIC INVERTEBRATE SAMPLING USING PONAR GRABS
In this study variation in the estimate of benthic invertebrate
abundance in grab samples was partitioned into three separate units: (1) AOC
variability defined as the difference in benthic invertebrate estimates
between AOCa; (2) station variability, defined as station to station
differences in benthic estimates in a particular AOC; and (3) sampling
variability defined as differences between benthic invertebrate abundance
estimates between individual replicates at each station.
Estimates of variance components (Nested Analysis of Variance; [ANOVA],
Snedecor and Cochran 1982) identified that combined station and sampling
variability accounted for 74% to 99% of the explained variability in the
estimates for all major groups (Table 5.21). Variability associated with AOC
ranged from 1% to 26% for all major taxa.
Station variability explained almost 50% of the variability in the
estimates of Oligochaeta abundance (Table 5.21). Variability associated with
AOC and sampling each accounted for 25% of the remaining variability in the
estimates of Oligochaeta abundance.
Sampling variability accounted for 62% of the variability in the
estimates of Chironomidae abundance. Variability associated with station to
station differences accounted for 37% of the associated variability.
Variability associated with AOC was extremely low, accounting for only 1% of
the total associated variability in the estimates of Chironomidae abundance.
Variability associated with Bivalvia and Gastropoda estimates
demonstrated contrasting results. Station variability accounted for 25% of
the variability in the estimates of Bivalvia abundance, but accounted for 59%
of the variability in the estimates of Gastropoda abundance (Table 5.21).
Sampling variability demonstrated the opposite, accounting for 55% of the
variability in estimates off Bivalvia abundance while accounting for only 37%
of the variability in the estimates of Gastropoda abundance. Variability
associated with AOC was 19% for estimates of Bivalvia abundance and only 4%
for estimates of Gastropoda abundance.
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DISCUSSION
BENTHIC INVERTEBRATE ABUNDANCE AND COMMUNITY STRUCTURE IN GRAB SAMPLES
The abundance estimates of benthic invertebrates in this study are most
likely conservative because of the sampling device used to collect the
invertebrates (Resh 1979), and the size of the mesh used to sieve the
invertebrates (Brinkhurst 1974, Resh 1979, Heushelle 1982). Ponar samplers
are heavy by nature in order to penetrate the sediment surface more evenly and
efficiently. Problems arise as the sampler nears the sediment. A shock wave
of displaced water impacts the sediment just before the sampler, many times
causing small organisms and surface dwellers to be pushed out of the way of
the sampler (Flannagan 1970, Howmiller 1971, Howmiller and Beeton 1971,
Millbrink and Wiederholm 1973). The effect of this type of disturbance is not
easily quantified and was not quantified in this study. However, any bias
caused by this type of sampler should be consistent across stations and AOC's.
All samples were sieved through a 500 /an brass screen upon collection in
the field. Although this size screen is good at separating the benthic
invertebrates from the sediments, many of the smaller organisms such as the
Naidids pass through with the sediments (Brinkhurst 1974, Resh 1979, Heushelle
1982). A more appropriate method of sieving may be to sieve samples through a
500 /on mesh screen followed by a 250 /m screen to collect organisms that pass
through a 500 /im mesh screen (Burt et al. 1991, Dr. Ralph Brinkhurst, Aquatic
Resources Center, Franklin TN, personnel communication).
COMPARISON AMONG AOCs
Benthic Invertebrate Abundance and Species Composition
Comparisons among the three AOCs indicated that Indiana Harbor was the
least suitable to the benthic invertebrates and Buffalo River was the most
suitable. This was based on overall abundances (BR>SR>IH) and total number of
genera and species (BR=33>SR=20>IH=14) present at each AOC. Buffalo River had
many genera that were not present or were present in low numbers in Indiana
Harbor and Saginaw River samples. Varying degrees of contamination at these
AOCs could influence abundance and species composition, but differences in
substrate quality, and amount and quality of food sources can not be
discounted. There was no relation that could be attributed to any single
contaminant or physical variable at any of the stations.
Indiana Harbor
Indiana Harbor was a very depauperate benthic invertebrate community.
Except for the 2 Chironomidae collected at IH-01-10, no other aquatic insect
orders were present at the stations in Indiana Harbor. The Bivalvia were rare
occurrences at three stations (IH-01-03, IH-01-04, IH-01-07). The genera of
Bivalvia present have been indicated as being somewhat tolerant of organic
contamination (Carr and Hiltunen 1965, Fuller 1974, Bode 1988).
The invertebrate community, dominated by the Oligochaeta family
Tubificidae, is indicative of a benthic invertebrate community subjected to
heavy organic pollution (Brinkhurst et al. 1972, Brinkhurst and Cook 1974,
Cook and Johnson 1974, Burt et al. 1991). All of the Tubificidae genera
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present in Indiana Harbor are known to be very tolerant of organic pollutants
(Kennedy 1965, Brinkhurst et al 1972). Limnodrilus hoffmeisteri. one of the
most pollution tolerant Oligochaeta species, was the most abundant species at
all stations sampled.
Oligochaeta abundance was lowest at stations IH-01-07 (junction of Lake
George Branch and Grand Calumet Branch) and IH-01-06 (main channel), and
highest at station IH-01-10. Sediments from station IH-01-07 generally had
the highest concentrations of metals (Tables 4.3 and 4.4) and organic
contaminants (Table 4.37) with station IH-01-06 having the next highest
concentration of metals and organic contaminants. This would tend to indicate
that the combined input of metals and organic contaminants from both the Grand
Calumet Branch and the Lake George Branch is having a pronounced effect on the
Oligochaeta communities at these stations. Samples from these stations were
also among the most toxic in laboratory exposures (see Chapters 2 and 4).
High concentrations of metals may reduce Oligochaeta abundance by reducing the
bacteria on which they feed.
Oligochaeta abundance was extremely high at station IH-01-10,
approaching 1,000,000/m2 in individual grab samples. Although metals
concentrations are the second lowest of any stations sampled, the most
pronounced reason for the higher abundances may be the high density of aquatic
vegetation present at IH-01-10. Large amounts of vegetation were most likely
present because station IH-01-10 was on the upstream side of a low clearance
bridge, which tended to minimize the amount of disturbance caused by boat
traffic (Joe Rathbun, ASCI Corporation, Grosse lie, MI, personal
communication). This vegetation most likely provided a habitat which
collected decaying plant material resulting in a considerable amount of
bacterial activity.
Many of the grab samples had large remnants of Gastropoda and Bivalvia
shells. The stations sampled were depositional areas and the shells may have
washed in from upstream, but the possibility that resident Mollusca
communities at these locations have died off can not be excluded with the data
in this study.
Buffalo River
Benthic invertebrate communities samples from the Buffalo River
exhibited a wide range of abundance across all stations. Although the
Oligochaeta were the most abundant taxa, several stations had significant
number of representatives from the Chironomidae, Bivalvia, and Gastropoda. On
rare occasions, representatives from the orders Ephemeroptera, Odonata,
Hemiptera, Tricoptera, Coleoptera, Diptera (other than Chironomidae),
Hirudinea, and Amphipoda were present.
The Oligochaeta community was dominated by the family Tubificidae, with
species which are generally considered tolerant of organic and metal pollution
(Kennedy 1965, Brinkhurst et al 1972, Burt et al. 1991). Limnodrilus
hof^meisterjj. was the most abundant Oligochaeta at all stations except for BR-
01-01 and BR-01-10. The reasons for this are not clear, but it may be a
result of food preference or habitat preference not being met at these
stations (Verdonschot 1989).
The Chironomidae community at Buffalo River consisted primarily of
tolerant genera (Hilsenhoff 1982, 1987, Bode 1988). The exception to this
generalization was Tanvtarsus sp. at BR-01-10 which reportedly prefers less
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organically enriched environments (Krieger 1984). Procladius sp., Chironomus
sp. and Crvptochironomua sp. are generally considered to be the most
frequently encountered genera in heavily polluted environments (Cook and
Johnson 1974, Krieger 1984). These three genera were generally the most
frequently sampled genera at Buffalo River.
Several genera and species of Bivalvia and Gastropoda were present in
Buffalo River. Two genera, Musculium sp. and Pisidium sp., have been reported
from systems receiving organic pollution (Carr and Hiltunen 1965, Fuller
1974). Species of the genus Sphaerium may be somewhat tolerant to organic
contaminants by virtue of being in the family Sphaeridae which reportedly is
tolerant of organic pollution (Bode 1988, Plafkin et al. 1989).
Representatives from the family Unionidae were collected only at 4 of 10
stations and their abundances were always low. This could be an indication
that these organisms are less tolerant of contaminants than some of the
Sphaeridae, however members in this family tend to be large so the ponar grabs
may not efficiently collect the Unionidae.
Representatives from the genera Valvata sp. and Bithvnia sp. have been
reported to be somewhat tolerant of organic pollution (Carr and Hiltunen 1965,
Krieger 1984). Even so the occurrence of these genera was limited to 6 of 10
stations and abundances of these genera were usually low at most stations.
The lack of abundance may be due to toxic effects of metals and organic
contaminants, insufficient grazing material, lack of suitable habitat , or a
combination of all the above. Interestingly, 2 species of Gastropoda, Valvata
lewisi and Cincinnatia cincinnatiensis which are reportedly uncommon in New
York State (Eileen Jokinen, University of Connecticut, Storrs, CT, personal
communication) seems to be the most abundant Gastropoda species at the Buffalo
River stations. Other reported occurrences for these species in New York are
in Oneida Lake, Seneca Lake, and a few tributaries in the Oswego drainage
basin. All of these areas are subject to high organic enrichment. This may
be indicative of a tolerance for organic contaminants which allows these
species to out compete other Gastropoda and survive in these polluted areas.
Concentrations of metals and organic contaminants were not consistently
related to patterns of invertebrate abundance. Station BR-01-04 and BR-01-10
had the lowest metal concentration and the highest number of taxonomic orders
(8) of the Buffalo River stations. Concentrations of contaminants may have
had an inconsistent effect on the benthic invertebrate community because of
differences in spatial distribution of contaminants or that these contaminants
were not readily bioavailable to the organisms (Burton 1991).
Saqinaw River (first and third survey)
The benthic invertebrate communities sampled from the Saginaw River
exhibited a fairly narrow range of abundance. As with Indiana Harbor and
Buffalo River, the Oligochaeta were the most abundant taxa at all stations.
The Oligochaeta constituted a higher percentage of the invertebrate community
across stations during the first survey sampled in December 1989. Samples
from the third survey in June of 1990 had a higher percentage of the family
Chironomidae present.
The Oligochaeta were dominated by the pollution-tolerant Tubificidae
similar to Indiana Harbor and Buffalo River. Limnodrilus hoffmeisteri was more
dominant in the samples from the first survey compared to the third survey
where distributions were spread more evenly across several other species.
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These differences may be attributed to seasonal variability, spatial or
temporal variability, differing food sources, or a combination of all the
above.
The Chironomidae community at Saginaw River was predominantly pollution-
tolerant genera (Hilsenhoff 1982, 1987, Bode 1988). The exception was
Tanvtarsus sp. at SR-03-08 which was present but in very low abundance.
Procladius sp. and Chironomus sp. were most frequently collected and are
reported tolerant of metal and organic pollution (Cook and Johnson 1974,
Krieger 1984).
The only Mollusca present at Saginaw River were those in the family
Sphaeridae. All three genera, Musculium sp., Pisidium sp. and Sphaerium sp.
were present in the first survey, but only Musculium sp. was present in the
third survey. The reasons for this are unclear. Perhaps these genera were
present in very low abundances during the third survey and were missed when
the ponar grabs were taken, or that these genera were not present at any of
the stations in the third survey due to loss of habitat or increased
contamination.
There were no families of Gastropoda present in Saginaw River. The
exact reason for this is not clear from the data, but may be the result of
contaminants in the area or the habitat may be insufficient to support this
group. Further investigation designed to identify contaminant, abiotic and
biotic factors controlling Gastropoda and overall invertebrate distributions
need to be conducted before any definitive conclusions can be made.
Benthos-Chemistry Comparisons
Comparisons between concentrations of Simultaneously extracted metals
(SEM), total PAHs and total PCBs all demonstrated a consistent pattern of
decreasing abundance with increasing contaminant concentration (Figures 5.2 to
5.4). Regardless of the contaminant examined there seemed to be a
concentration below which invertebrate abundance was independent of
contaminants. There also tended to be a concentration above which
invertebrate abundance was consistently reduced. This suggests a threshold
concentration of contamination below which invertebrate abundance is
controlled by other factors and above which the influence of the contaminant
is more pronounced. Even at low concentrations of contaminants the community
is still indicative an impacted environment.
The value of this threshold limit, regardless of contaminant examined,
seems to be consistently higher for the Oligochaeta community compared to the
other invertebrate orders. This is consistent with the findings that identify
Oligochaeta as some of the most tolerant aquatic invertebrates (Hilsenhoff
1982, 1987; Bode 1988).
DEFORMITIES IN CHIRONOMIDAE
Different genera of Chironomidae exhibit different levels of
susceptibility or tolerance to contaminants (Hamilton and Saether 1971; Hare
and Carter 1976; Warwick 1985, 1988; Wiederholm 1984). Some genera are quite
intolerant and low levels of contaminants eliminate these genera from
Chironomidae communities, while some genera such as Procladius sp., Chironomus
sp. and Cryptochironomus sp. are more tolerant and persistent (Warwick 1985,
Bode 1988). A relationship between increased contamination and the presence
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of Chironoroidae deformities has long been documented by many investigators
(Hamilton and Saether 1971, Warwick 1980, 1985, Tennessen and Gottfried 1983,
Cushman 1984, Wiederholm 1984). Some of the reported deformities include:
thickening of the exoskeleton, enlargement and darkening of the head capsule,
asymmetry in mouth parts, missing or fused lateral teeth, and antennal
deformities.
The mentum and ligula (Tanypodinae) of Chironomidae were examined for
deformities at all AOCs. The specimens had various mouthpart deformities
including missing lateral and central teeth, asymmetry in the mentum, badly
deformed and twisted lateral teeth on the mentum and missing teeth on the
ligula (Procladius sp.). None of the specimens examined exhibited antennal
deformities. The occurrence of deformities in this study tended to occur in
the Procladius sp. and Chironomus sp. genera. Even when other genera were
present they did not seem to display frequent mouth part deformities. The
reasons for this are not clear but may be due to these genera dying before
they can exhibit abnormalities.
The occurrence of deformities in Chironomidae reportedly is less than 1%
in non-impacted communities (Wiederholm 1984; Warwick et al. 1987). Several
investigators have suggested deformity frequency in the range of 5% to 25% or
greater is indicative of moderate to severe contamination (Wiederholm 1984,
Warwick et al. 1987). Given this criteria, occurrence of deformities in the
Chironomidae community at the three AOCs indicates the areas ranged from
moderately to heavily polluted. The stations that had Chironomidae without
deformities were also the areas that had few or no Chironomidae. Only 2
Chironomidae were collected from Indiana Harbor (station 10) and both had
deformities.
As with the abundance data, the frequency of deformities in Buffalo
River indicates less toxic conditions compared to either Indiana Harbor or
Saginaw River. It is interesting that the occurrence of deformities is
consistently high at the Saginaw River stations, which indicates that
contaminants were at a sufficient level impact the Chironomidae community at
most stations.
Although it would seem that the occurrence of these deformities may be
related to the degree of contamination in the sediments, the limited number of
samples made comparisons difficult. Frequency of deformities exhibited by
Chironomidae exposed in laboratory sediment toxicity tests (Chapter 4) need to
be compared to the frequency of deformities in the field-collected
Chironomidae. A more specific study designed to elucidate the relations
between particular contaminants and Chironomidae mouthpart deformities and
which would encompass a broad range of impacted and non-impacted areas would
be beneficial to future studies.
ARTIFICIAL SUBSTRATES VS. GRAB SAMPLES
Several investigators have examined the advantages and disadvantages in
using artificial substrates to examine the benthic invertebrate community
(Cairns 1982). The artificial substrates used in the present study were
modified from Stauffer et al. (1976). These substrates were very easy to work
with and the synthetic mesh was easily pulled apart thereby allowing easy
access to the invertebrates with a minimum of specimen damage.
The invertebrates from the artificial substrates were processed
following the methods outlined in the rapid bioassessment protocols (Plafkin
5-18
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et al. 1989). Invertebrates were enumerated into major taxonomic groups and
no further identification of these taxa was attempted. Therefore a rigorous
analysis of the data have yet to be attempted. The results of the data for
the artificial substrate samples compared to the ponar grab samples should be
used with caution.
The artificial substrates collected a benthic invertebrate community
which was considerably more diverse and abundant than the benthic grabs. This
indicates there were more invertebrates in the areas sampled than would be
predicted by the ponar grabs alone. Perhaps quality substrate is limited in
these areas. Invertebrates collected on the artificial substrates may not be
indicative of the true abundance of benthic invertebrates in these areas. The
artificial substrates may act as a focal point of colonization for surrounding
invertebrates inhabiting the sediments.
The differences observed between the ponar grabs and artificial
substrates collected presents a problem for benthic community assessments.
Estimates of the benthic community composition made from the ponar grabs would
indicate a severely impacted community based on the low diversity of taxa.
Considerably more invertebrates were at the station based on the results of
the artificial substrates samples. This becomes extremely important if
potential food chain bioaccumulation estimates are being considered. Since
the artificial substrates probably sample the epibenthic community more
readily available to vertebrates future studies may require the use of both
types of samplers to make estimates the benthic invertebrate community.
VARIATION IN BENTHIC INVERTEBRATE SAMPLING USING PONAR GRABS
In large scale ecological studies it is important to identify sources of
variability so that a meaningful interpretation of the data can be made
(Collins and Sprules 1983) and so future studies can be designed to address
major sources of variation. The results of the variance partitioning
indicates that station and sampling variability accounted for most of the
explained variability in the taxa estimates. It is not uncommon for station
to station variance to account for a considerable amount of the explained
variation between invertebrate abundance estimates (Lewis 1978, Threlkeld
1983). The differences associated between stations may be due to (1) some
stations being closer to contaminant effluent outfalls, (2) variation in
substrate composition and structure which would preclude colonization by the
invertebrates, (3) depth of the station which may prohibit invertebrates due
to wave action or shipping traffic, or (4) a combination of all of these
factors.
Variance associated with AOC was significant in the estimates of
Oligochaeta and Bivalvia abundance while minimal in the estimates of
Chironomidae and Gastropoda abundance. The benthic invertebrate abundances
between the areas of concern were quite similar. This is not an unexpected
considering all three of these areas have received substantial industrial and
municipal input and the bulk of the community consists of Oligochaeta. Future
studies would be enhanced if a relatively pristine reference station could be
identified and sampled.
The remaining variance was associated with sampling technique.
Invertebrate estimates between successive grabs could vary by as much as 68%
depending on the taxa. Since benthic invertebrate populations exhibit a
patchy distribution in the sediment, the result is not unusual (Elliott 1977).
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But for most studies with benthic invertebrates you would want this estimate
to be as low as possible (eg. additional replicates).
Partitioning of the variance into different components indicates that
future studies might provide better data if: (1) smaller size grabs were
taken, but with more grabs to try to minimize the influence of a patchy
benthic invertebrate distribution; and (2) more stations were sampled in a
given AOC in an attempt to obtain stations which would better sample across
the gradient of effluents from upstream to downstream. A main reason for
limiting the amount of grabs taken from an area of concern is the time and
cost it takes to process the samples once they are taken. By taking smaller
samples, but still taking the same area of sample, the processing time would
be shorter and the estimates between grabs should have a lower variance (Leigh
Frederickson, University of Missouri, Columbia, MO, personal communication).
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SUMMARY AND RECOMMENDATIONS
Oligochaeta and Chironomidae abundance constituted over 90% of the
benthic invertebrate community collected with the ponar grab at all stations
and Areas of Concern. This overwhelming dominance by these 2 taxonomic groups
is indicative of an impacted benthic invertebrate community. Buffalo River
had the largest number of genera and species (n=33) present, followed by
Saginaw River (n=20) and Indiana Harbor (n=14). Based on measures of taxa
dominance and taxa richness Indiana Harbor was the least suitable Area of
Concern to benthic invertebrates and Buffalo River was the most suitable.
Comparisons between concentrations of simultaneously extracted metals
(SEM), total PAH, and total PCB with measures of benthic invertebrate
abundance demonstrates a consistent pattern of decreasing invertebrate
abundance with increasing contamination. The data suggest a threshold
concentration below which abundance may be controlled by factors other than
contaminant concentration and above which the influence of the contaminant is
pronounced. However, direct cause and effect relations between invertebrate
abundance and any individual contaminant were not demonstrable with the data
at this time.
The occurrence of Chironomidae deformities was pronounced at all Areas
of Concern. Buffalo River consistently had the lowest occurrence of
deformities across all stations. The occurrence of Chironomidae deformities
was more prevalent at Saginaw River than Buffalo River. The 2 Chironomidae
collected from Indiana Harbor were both deformed. Overall, the percentages of
deformities at all the Areas of Concern would indicate that these area are
moderately to severely polluted.
The ponar grabs and artificial substrates collected tremendously
different numbers and taxa composition of aquatic invertebrates. While the
grab samples were predominantly Oligochaeta and Chironomidae, the artificial
substrates collected predominantly Amphipoda, Isopoda, Turbellaria, and
Dreissena sp.. Indications are the ponar grabs may be missing a considerable
component of the benthic invertebrate community. Incorporation of artificial
substrates may enhance the estimation of benthic invertebrate community
composition.
Results of variance partitioning indicates station and replicates
accounted for most of the explained variability in the estimation of
invertebrate abundance collected with the ponar grab. Variance associated
with Area of Concern was significant in the estimation of Oligochaeta and
Bivalvia abundance, while minimal in the estimation of the remaining taxa
abundance.
The following recommendations are concerns and future research
identified as important factors relevant to interpreting results of benthic
invertebrate surveys.
• Additional research is needed to evaluate specific contaminant, biotic
and abiotic factors controlling invertebrate abundance and community
structure in sediments.
• Sediment chemistry and physical variables need to be obtained from
splits of the samples from which the invertebrates are collected to
overcome problems associated with benthic invertebrate patchiness and
heterogenous contaminant deposition.
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Assessment of benthic communities should use both artificial substrates
and grab samples.
Frequency of deformities in Chironomidae exposed in laboratory toxicity
tests (Chapter 4) should be measured and compared to the frequencies of
deformities in the Chironomidae collected from the field.
With most of the variance in benthic community estimates associated with
station and replicates, future studies should sample more stations and
take more replicates using smaller grab samples.
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Kennedy, C.R. 1965. The distribution and habitat of Limnodrilus claparede and
its adaptive significance. Oikos 16: 26-28.
Kosalwat, P. and A.W. Knight. 1987. Chronic toxicity of copper to a partial
life cycle of the midge Chironomus decorus. Arch. Environ. Contam. Toxicol.
16: 283-290.
Krieger, K.A. 1984. Benthic macroinvertebrates as indicators of environmental
degradation in the southern nearshore zone of the central basin of Lake Erie.
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contaminants to benthic invertebrates. Chapter 8. In R. Baudo, J.P Giesy and
H. Muntau, eds., Sediments; Chemistry and Toxicitv of In-Place Pollutants.
Lewis Publishers, Inc. Ann Arbor, MI.
5-24
-------
Lewis, W.M., Jr. 1978. Comparison of temporal and spatial variation in the
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Medine, A.J. and S.C. McCutcheon 1989. Fate and transport of sediment-
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Plafkin, J.L., M.T. Barbour, K.D. Porter, S.K. Gross and R.M. Hughs. 1989.
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5-25
-------
Tennessen, K.J. and P.K. Gottfried 1983. Variation in structure of ligula of
Tanypodinae larvae (Diptera: Chironomidae). Ent. News. 94: 109-116.
Thorp, J.H. and A.P. Covich. 1991. eds. Ecology and classification of North
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larvae as measures of toxic stress in freshwater ecosystems: indexing antennal
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as biological indicators of toxic stress, p. 281-320. In M.S. Evans ed. Toxic
contaminants and ecosystem health; a Great Lakes focus. John Wiley and Sons,
New York, NY.
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(Diptera:: Chironomidae) and their biomonitoring potential. Can. J. Fish.
Aquat. Sci. 46: 1255-1270.
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incidence of deformities in Chironomus sp. from Port Hope Harbour, Lake
Ontario. J. Great Lakes Res. 13: 88-92.
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diagnoses, Part 1. Larvae". Entomologia Scandinavica Supplement No. 19.
Wiederholm, T. 1984. Incidence of deformed chironomid larvae (Diptera:
Chironomidae) in Swedish lakes. Hydrobiologia 109: 243-249.
5-26
-------
LIST OF FIGURES IN CHAPTER 5
Figure 5.1 Artificial substrate samplers used to collect aquatic
invertebrates at select stations from each Area of Concern
(AOC). The rotisserie baskets were 26 cm in length, 17 cm in
diameter, and 53 cm in circumference.
Figure 5.2 Relation of the summed total simultaneously extracted metals
(Cd, Cr, Cu, Ni, Pb, Zn) concentration (/unol/dry g) to mean
total invertebrate abundance (number/m2) across all Areas of
Concern.
Figure 5.3 Relation of the summed total PAH concentration (^ig/dry g OC)
to mean total invertebrate abundance (number/m2) across all
Areas of Concern.
Figure 5.4 Relation of the summed total PCB concentration (ng/dry g OC)
to mean total invertebrate abundance (number/m ) across all
Areas of Concern.
Figure 5.5 Relation of the summed total simultaneously extracted metals
(Cd, Cr, Cu, Ni, Pb, Zn) concentration (/unol/dry g) to mean
Oligochaeta abundance (number/m2) across all Areas of Concern.
Figure 5.6 Relation of the summed total simultaneously extracted metals
(Cd, Cr, Cu, Ni, Pb, Zn) concentration (/unol/dry g) to mean
Chironomidae abundance (number/m ) across all Areas of
Concern.
Figure 5.7 Relation of the summed total simultaneously extracted metals
(Cd, Cr, Cu, Ni, Pb, Zn) concentration (/xraol/dry g) to mean
Bivalvia abundance (number/m2) across all Areas of Concern.
Figure 5.8 Relation of the summed total simultaneously extracted metals
(Cd, Cr, Cu, Ni, Pb, Zn) concentration (/unol/dry g) to mean
Gastropoda abundance (number/m ) across all Areas of Concern.
Figure 5.9 Relation of the summed total PAH concentration (/ig/dry g OC)
to mean Oligochaeta abundance (number/m ) across all Areas of
Concern.
Figure 5.10
Relation of the summed total PAH concentration (fig/dry g OC)
to mean Chironomidae abundance (number/m ) across all Areas of
Concern.
Figure 5.11
Relation of the summed total PAH concentration (/*g/dry g OC)
to mean Bivalvia abundance (number/m ) across all Areas of
Concern.
Figure 5.12
Relation of the summed total PAH concentration (fig/dry g OC)
to mean Gastropoda abundance (number/m ) across all Areas of
Concern.
5-27
-------
Figure 5.13 Relation of the summed total PCB concentration (fig/dry g OC)
to mean Oligochaeta abundance (number/m2) across all Areas of
Concern.
Figure 5.14 Relation of the summed total PCB concentration (fig/dry g OC)
to mean Chironomidae abundance (number/m ) across all Areas of
Concern.
Figure 5.15 Relation of the summed total PCB concentration (/xg/dry g OC)
to mean 1
Concern.
Figure 5.16 Relation of the summed total PCB concentration (fig/dry g OC)
to mean <
Concern.
to mean Bivalvia abundance (number/m ) across all Areas of
to mean Gastropoda abundance (number/m ) across all Areas of
Figure 5.17 Percentage composition of the invertebrate taxa collected
using artificial substrates and a ponar grab at Indiana
Harbor, IN, August 1990.
Figure 5.18 •" Percentage composition of the invertebrate taxa collected
using artificial substrates and a ponar grab at Buffalo River,
NY, October 1989.
Figure 5.19 Percentage composition of the invertebrate taxa collected
using artificial substrates and a ponar grab at Saginaw River,
MI, June 1990.
Figure 5.20 Comparison of the total amounts of invertebrates collected
with artificial substrates and a ponar grab at Saginaw River,
MI, June 1990.
5-28
-------
Figure 5.1
River
-------
MEAN TOTAL ABUNDANCE
01
ro
o
>
m
C/5
m
O
m
o
m
53
c
(Q
ro
n
n1 n
n
o
n
D>
t>
v
CO
DO
CO =
03
I!
CO
(D
pi
N)
-------
MEAN TOTAL ABUNDANCE
3
\i^ i_
cn
o
8
* i o
0
§
cn
8
O
IV)
o
8
O
T)
I
a
(D O
08
o
00
o
o
o
o
o
o
-x-
n
n
CO
*?
£^
CO
CO — CD
o "P o ' n 7
cb 2 o
3
(Q
c
"1
m
. Ni^
ai
CO
-------
O
-a
O
00
3"
CQ
CQ
O
MEAN TOTAL ABUNDANCE
P .P1
o o
o o
Ol
o
ro
o
o
Ol
o
o
cn
ro
01
CO
o
o
o
CO
"01
o
o
-&-
D
o
n
o
o
C/5 C75
v 33
TV 1
O
CO
= CD
5 D?
(Q
C
CD
01
-------
Figure 5.5
20,000
LLJ
O
< 15,000
Q
Z
CD
ID
o
o
o
10,000
5,000
n
n
n
n
O
n
o
0
n
o
A
A
A
A
BR-01
n
IH-01
A
SR-01
o
SR-03
*
A
2 4 6 8 10
SIMULTANEOUSLY EXTRACTED METALS (umol/dry g)
12
-------
Figure 5.6
o,uuu
g 2,500
Z
O
| 2,000
111
| 1,500
o
0
jf 1,000
o
Z
•E 500
n
n
—
Dn
nH D
$& r> K AA A\
BR-01
n
IH-01
A
SR-01
o
SR-03
*
A
2 4 6 8 10
SIMULTANEOUSLY EXTRACTED METALS (umol/dry g)
12
-------
Figure 5.7
250
n
200
LU
o
I
§ 150
CD
BR-01
n
IH-01
A
SR-01
o
SR-03
100
CO
<
LU
D
50
n
0
O
AA
A
2 4 6 8 10
SIMULTANEOUSLY EXTRACTED METALS (umol/dry g)
12
-------
Figure 5.8
400
LU
O
5 300
O
z
D
QQ
O 200
Q_
O
CC
s
0
iS
100
D
D
A/V
BR-01
D
IH-01
A
SR-01
o
SR-03
*
0
2 4 6 8 10
SIMULTANEOUSLY EXTRACTED METALS (umol/dry g)
12
-------
Figure 5.9
20,000
UJ
O
< 15,000
Q
Z
CD
UJ
<
O
O
O
<
ill
10,000
5,000
0
D
O
D
A
* D
0
2,000
4,000 6,000
TOTAL PAH (ug/dry g OC)
A
BR-01
D
IH-01
A
SR-01
O
SR-03
*
8,000
10,000
-------
Figure 5.10
Q
Z
LJU
2,500
2,000
| 1,500
O
O
^ 1,000
O
Z
LJJ
500
D
0
A
2,000
4,000 6,000
TOTAL PAH (ug/dry g OC)
8,000
BR-01
D
IH-01
A
SR-01
o
SR-03
10,000
-------
o
o ©
(Q
(Q
O
O
O)
00
o
t>
D>
en
o
MEAN BIVALVIA ABUNDANCE
8 8
n
n
n
ro
8
•
CO
CO
n
CD
ro
u
(Q
i
U1
-------
I>
ro
So
(Q
(Q P
O Q
O Q
n
n
MEAN GASTROPODA ABUNDANCE
ro w
80
o
n
CO
DO
0) =
DD
O5 -*>
31
(Q
ro
-------
Figure 5.13
20,000
UJ
O
< 15,000
Q
Z
D
CD
O
O
g
4
O
LU
10,000
5,000
0
0
500
1,000 1,500 2,000 2,500
TOTAL PCB (ng/dry g OC)
BR-01
D
IH-01
A
SR-01
SR-03
*
3,000
3,500
-------
MEAN CHIRONOMIDAE ABUNDANCE
_L _k ro
o on o
cn
0
o
ro
01
CO
en
o
o
01
TJ
O
CD
CQ
o: ro
13 8
CQ O
O
o
ro
01
CO
01
D
o
0)
Y ^W
CO
C/5 = CO
^n ^ JH ^n
°g >6 °g
^^ M^.
(Q
C
Q
Ol
-------
Figure 5.15
250
LLJ
o
200
Q
§ 150
CO
> 100
CD
<
LLJ
50
D
D
D
-R
BR-01
D
IH-01
A
SR-01
SR-03
*
0 $^_»
0
500
1,000 1,500 2,000 2,500
TOTAL PCB (ng/dry g OC)
3,000
3,500
-------
O
01
o
O
o
2
TJ
O
DO
3"
(Q
ro
CO 8
o
o
N)
c
GO
n
MEAN GASTROPODA ABUNDANCE
-*• ro 03
o
o
I
n
C/)
0) —
03
03 -••
-n
(Q
I
pi
o>
-------
Percent Composition
8
8
o
Percent Composition
8
8
CO2
i—h
£
o
3 8
8
8
O
s
cr
CO
CD
0)
CD
00
CO
0)
I
CD
O"
o
III
I!
pi
«>i
-------
Percent Composition
o 8 S 8 8
8
.'
8
CO
5T
5:3
O
D
8
8
c
o
8
CO
i-*
0)
£+•
— • o
o •"
8
8
' 1 ' 1 •— r ' i
f f ( t r r frirrr c
cr
J2-
& z+
CD
' -I
.LUJJJJ //A/./,.^ (Q
Percent Composition
8 * 8 8 8
• i • i • i • i •
^ ?
3
? CO
D)
CD
0)
4 g
CO
5^^
S 0 • H •
HIH
i!l|!
CD
=t
0)
O
33
<"
3
(Q
i
en
_*
00
-------
Percent Composition
Percent Composition
0)
(Q
1
0
I!
«
S
01
Li
CO
-------
Abundance (thousands/nrP)
8
8 8 8
S3
O
en
CO 8
B
O
13 §
O)
Q W
O" co
o
3
cr
5 CO
£ 05
g-CQ
CO —•
9 u
c? JD
%Z
t3
8 <•
CD
3
(O
§
en
lo
o
-------
Table 5.1 Continued
AOC
SR-01-023
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
Oligochaeta
6418
6974
3621
1682
5954
1478
813
4944
3735
2861
302
820
3538
805
(761)
(242)
(515)
(432)
(365)
(204)
(630)
(863)
(648)
(244)
(108)
(240)
(253)
(203)
Chironomidae
238
121
45
11
87
363
61
2204
23
144
15
360
389
125
(77)
(22)
(18)
(5)
(14)
(79)
(55)
(372)
(9)
(35)
(7)
(105)
(45)
(36,
Bivalvia
0
4
8
16
0
0
0
0
8
11
0
0
4
0
(0)
(4)
(5)
(7)
(0)
(0)
(0)
(0)
(5)
(8)
(0)
(0)
(4)
(0)
Gastropoda
0
0
0
0
0
0
0
0
0
0
0
0
0
0
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
Total
6664
7129
3686
1709
6056
1890
888
7152
3780
3047
321
1157
3977
941
(822)
(239)
(511)
(437)
(377)
(287)
(693)
(1194)
(658)
(227)
(114)
(296)
(240)
(238)
'BR-Buffalo River
2IH-Indiana Harbor
3SR-Saginaw River
-------
Table 5.1 Mean abundance data (number/m2, standard error in parentheses) for major benthic
invertebrate groups. Values are averages (n=5) of the abundance values obtained for each
station within an AOC
Station
BR-01-011
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
IH-01-032
IH-01-04
IH-01-05
IH-01-06
IH-01-07
IH-01-08
IH-01-10
Oligochaeta
2714
7394
7059
8407
6403
19157
14496
16220
1984
3092
3765
5935
5198
1501
552
2903
493887
(1045)
(1389)
(1712)
(989)
(1920)
(2756)
(1838)
(2254)
(492)
(860)
(1618)
(2090)
(1742)
(915)
(272)
(1430)
(180680)
Chironomidae
197
110
151
386
11
114
26
181
287
2771
0
0
0
0
0
0
8
(60)
(44)
(46)
(37)
(8)
(44)
(11)
(19)
(86)
(1633)
(0)
(0)
(0)
(0)
(0)
(0)
(8)
Bivalvia
49
0
217
79
12
87
84
68
12
0
8
4
0
0
8
0
0
(40)
(0)
(103)
(42)
(8)
(32)
(22)
(13)
(8)
(0)
(8)
(4)
(0)
(0)
(8)
(0)
(0)
Gastropoda
27
0
72
378
4
57
23
4
0
0
0
0
0
0
0
0
0
(27)
(0)
(14)
(118)
(4)
(16)
(11)
(4)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
(0)
Total
3013
7530
7536
9461
6445
19418
14708
16473
2294
6067
3791
6025
5307
1501
609
2907
493917
(1190)
(1419)
(1787)
(1210)
(1927)
(2809)
(1839)
(2248)
(553)
(2423)
(1623)
(2081)
(1788)
(915)
(255)
(1435)
(180676)
-------
Table 5.2 Percentage contribution of each taxa to the total number
of tax«i collected from Indiana Harbor, IN, August 1989.
Taxa
Oligochaeta
Chironomidae
Bivalvia
Gastropoda
Ephemeroptera
Odonata
Pelcoptera
Hemiptera
Megaloptera
Trichoptera
Coleoptera
Diptera
Hirudinea
Amphipoda
03
99
0
<1
0
0
0
0
0
0
0
0
0
<1
0
04
98
0
<1
0
0
0
0
0
0
0
0
0
1
0
STATION
05 06
98
0
0
0
0
0
0
0
0
0
0
0
2
0
100
0
0
0
0
0
0
0
0
0
0
0
0
0
07
91
0
1
0
0
0
0
0
0
0
0
0
8
0
08
100
0
0
0
0
0
0
0
0
0
0
0
<1
0
10
100
<1
0
0
0
0
0
0
0
0
0
0
<1
0
-------
Table 5.3 Mean abundance values (number/m, n=5/station) for aquatic
Oligochaeta species collected from Indiana Harbor, IN, August
1989.
Taxa
Naididae
Dero diaitata
UN1
Tubif icidae
Aulodrilus pjqueti
Aulodrilus limnobius
Aulodrilus pluriseta
Ilyodrilus templetoni
Limnodrilus cervix
Limnodrilus cervix variant
Limnodrilus claparedianus
Limnodrilus clap.-cerv. complex
Limnodrilus hof fmeisteri
Limnodrilus maumeensis
Limnodrilus udekemianus
Limnodrilus sp.
Quistadrilus multisetoaus "1"
Quistadrilus multisetosus "m"
Tubifex tubifex
UIW/OCC2
UW/CC3
03
0
0
0
0
113
0
128
72
0
0
926
0
72
166
15
0
0
2191
30
STATION
04 05
0
0
0
77
0
0
0
101
0
0
1056
0
53
0
30
178
0
4445
0
0
0
0
0
0
0
0
0
0
0
790
0
0
0
0
858
0
3524
26
06
0
0
0
0
0
0
0
0
0
0
102
0
0
21
0
102
0
1246
30
07
0
0
0
0
0
15
0
6
0
0
238
0
0
10
0
6
0
258
15
08
0
0
0
0
0
0
0
38
0
0
1112
0
0
52
0
38
6
1646
0
10
0
0
0
0
0
1975
0
0
0
0
35560
0
0
0
0
0
0
419310
37535
unidentifiable naidid
unidentifiable, without capilliform chaetee (mostly Tubificidae)
unidentifiable, with capilliform chaetee (mostly Tubificidae)
-------
Table 5.4 Mean abundance values (number/m2, n=5/station) for Chironomidae
genera collected from Indiana Harbor, IN, August 1989.
STATION
Taxa 03 04 05 06 07 08 10
Tanypodinae
Coelotanvpus sp. ____---
Procladius sp. ____---
Tanvpus sp. -------
Tanytarsini
Tanvtarsus _______
Chironomini
Chironomus sp. _____--
Cladopelma sp. -------
Cryptochironomus sp. _____--
Dicrotendipes sp. -------
Glyptotendipes sp. ____---
Microchironomus sp. ____---
Polypedilum sp. -------
Orthocladinae
Cricotopus sp. _____-8
Total 0000008
Taxa Richness Q 0 0 Q 0 0 L_
-------
Table 5.5 Mean abundance values (numbers/m2, n=5/station) for Mollusca genera
and species collected from Indiana Harbor, IN, August 1989.
Taxa
03
STATION
04 05 06 07
08
10
Gastropoda
Valvatidae
Valvata lewisi
Valvata tricarinata
Bithyniidae
Bithvnia tentaculata
Ancylidae
Laevapex fucus
Hydrobiidae
Cincinnatia cincinnatiensis
Total
Taxa Richness
0
0
0
0
0
0
0
0
0
0
0
0
0
0
Bivalvia
Sphaeriidae
Musculium sp.
Pisidium sp.
Sphaerium sp.
Sp. unidentified
Unionidae
Anodonta imbecillis
Anodonta qrandis
Eliptio complanata
4
4
4
4
Total
Taxa Richness
8
2
o
0
0
0
8
2
0
0
0
0
-------
Table 5.6 Percentage contribution of each taxa to the total number of taxa
collected from Buffalo River, NY, October 1989.
Taxa
Oligochaeta
Chironomidae
Bivalvia
Gastropoda
Ephemeroptera
Odonata
Pelcoptera
Hemiptera
Megaloptera
Trichoptera
Coleoptera
Diptera
Hirudinea
Anvphipoda
01
90
7
2
<1
0
0
0
0
0
0
0
0
-------
Table 5.7 Mean abundance values (number/m, n=5/station) for aquatic Oligochaeta species collected
from Buffalo River, NY, October 1989.
Taxa
Naididae
Dero dioitata
UN1
Tub if icidae
Aulodrilus piqueti
Aulodrilus limnobius
Aulodrilus pluriseta
Ilvodrilus templetoni
Limnodrilus cervix
Limnodrilus cervix variant
Limnodrilus c 1 aparedianu s
Limnodrilus clap.-cerv. complex
Limnodrilus hof fmeisteri
Limnodrilus maumeensis
LimnodriluB udekemianus
Limnodrilus sp.
Quistadrilus multisetosus "1"
Quistadrilus multisetosus "m"
Tubifex tubifex
UIW/OCC2
UW/CC3
01
16
0
0
0
0
0
100
100
0
30
0
0
0
43
100
0
0
2249
73
02
0
0
30
0
0
0
0
0
0
0
370
0
0
111
30
0
0
6766
81
03
0
0
0
0
0
0
0
0
0
0
1884
0
0
494
219
28
0
4377
56
04
0
0
0
0
0
0
0
0
0
0
1224
0
0
588
261
34
0
5212
67
05
0
0
0
0
0
0
0
0
0
0
160
0
0
0
0
0
0
6172
70
STATION
06 07
0
0
0
0
0
0
0
77
0
0
1743
0
0
747
0
0
0
16590
0
0
0
0
0
0
0
0
58
0
0
2305
0
0
449
0
0
0
11698
0
08
0
0
0
0
0
0
0
0
0
0
1233
0
0
277
277
0
65
14322
130
09
0
0
0
0
0
0
0
54
0
0
125
0
0
26
10
20
0
1706
46
10
0
0
0
0
0
0
0
62
0
0
49
0
0
37
12
0
0
2829
99
unidentifiable naidid
Unidentifiable, without capilliform chaetee (mostly Tubificidae)
'unidentifiable, with capilliform chaetee (mostly Tubificidae)
-------
Tabie 5.8 Mean abundance values (number/m2, n=5/station) for Chironomidae genera collected from
Buffalo River, NY, October 1989.
Taxa
Tanypodinae
Coelotanypus sp.
Procladius sp.
Tanvpus sp.
Tanytarsini
Tanvtarsus
Chironomini
Chironomus sp.
Cladopelma sp.
Cryptochironomus sp.
Dicrotendipes sp.
Glyptotendipes sp.
Microchironomus sp.
Polvpedilum sp.
Orthocladinae
Cricotopus sp.
Total
Taxa Richness
STATION
01 02 03 04 05
- 28 26 -
45 8 71 325 11
— — — — —
-----
76 53 - - -
4 34 - - -
30 15 52 34
4
34
_
4
-----
197 116 151 386 11
74331
06 07 08 09
_ 4 -
110 26 174 57
_ — _ —
— — - -
- - - 64
- 15
4-4 140
_
- - - ' -
_ 4
- 8
— — — —
114 26 181 287
21 36
10
-
261
11
404
355
4
284
155
-
-
1297
—
2771
8
-------
Table 5.9 Mean abundance values (numbers/m2, n=5/station) for Mollusca genera and species collected
from Buffalo River, NY, October 1989.
Taxa
Gastropoda
Valvatidae
Valvata lewis!
Valvata tricar inata
Bithyniidae
Bithvnia tentaculata
Ancylidae
Laevapex fucus
Hydrobiidae
Cincinnatia cincinnatiensis
Total
Taxa Richness
Bivalvia
Sphaeriidae
Musculium sp.
Pisidium sp.
Sphaeriutn sp.
Sp. unidentified
Unionidae
Anodonta imbecillis
Anodonta arandis
Eliptio complanata
Total
Taxa Richness
01
STATION
01 02 03 04 05 06 07 08 09
23 - 5
- 5
4 -
- - 5
- - 57
27 0 72
20 4
11 - 70
30 - 109
- 33
— ™ ~
4 -
4 5
- — —
49 0 217
40 4
163 - 57 23 -
_ _ — — — —
23 - - - -
4
185 4
375 4 57 23 0 0
41 1 10 0
11 8 15 4 8 -
45 - 53 57 30 8
15 4 19 19 30
4
4 _ - - - 4
_ _ _ — — —
4 -----
79 12 87 84 68 12
523432
10
-
w
"
—m
0
0
—
—
—
-
—
^
0
0
-------
Table 5.10 Percentage contribution of each taxa to the total number
of taxa collected from the first survey of Saginaw
River, MI, December 1989.
Taxa
Oligochaeta
Chironomidae
Bivalvia
Gastropoda
Ephemeroptera
Odonata
Pelcoptera
Hemiptera
Megaloptera
Trichoptera
Coleoptera
Diptera
Hirudinea
Amphipoda
02
96
4
0
0
0
0
0
0
0
0
0
<1
0
0
03
98
2
<1
0
0
0
0
0
0
0
0
<1
0
0
STATION
04 06
98
1
<1
0
0
0
0
0
0
0
0
<1
0
0
98
1
1
0
0
0
0
0
0
0
0
0
0
0
07
98
1
0
0
0
0
0
0
0
0
0
<1
0
09
80
19
0
0
0
0
0
0
0
0
0
<1
0
10
92
7
0
0
0
0
0
0
0
<1
<1
<1
0
0
-------
Table 5.11 Mean abundance values (number/m , n=5/station) for aquatic
Oligochaeta species collected from the first survey of Saginaw
River, MI, December 1989.
Taxa
Naididae
Dero diaitata
UN1
Tubif icidae
Aulodrilus piquet i
Aulodrilus limnobius
Aulodrilus pluriseta
Ilvodrilus templetoni
Limnodrilus cervix
Limnodrilus cervix variant
Limnodrilus claparedianus
Limnodrilus clap.-cerv. complex
Limnodrilus hof fmeisteri
Limnodrilus maumeensis
Limnodrilus udekemianus
Limnodrilus sp.
Quistadrilus multisetosus "1"
Ouistadrilus multisetosus "m"
Tubifex tubifex
UIW/OCC2
UW/CC3
02
0
0
0
0
0
32
32
90
0
0
809
0
0
122
0
0
0
5295
32
STATION
03 04
0
0
0
0
0
98
265
70
0
0
1318
0
0
133
0
0
0
5056
35
0
0
0
0
0
0
174
109
0
0
391
22
0
221
22
0
0
2680
0
06
0
0
0
0
0
34
59
34
19
0
232
0
0
86
8
0
8
1189
8
07
0
0
0
0
0
30
101
30
0
0
536
0
0
286
0
0
0
4972
0
09
15
0
0
0
0
15
62
21
0
0
268
7
0
109
0
0
0
998
0
10
0
0
0
0
0
0
109
0
0
0
35
0
0
71
0
0
0
585
13
unidentifiable naidid
Unidentifiable, without capilliform chaetee (mostly Tubificidae)
unidentifiable, with capilliform chaetee (mostly Tubificidae)
-------
Table 5.12 Mean abundance values (number/m2, n=5/station) for Chironomidae
genera collected from the first survey of Saginaw River, MI,
December 1989.
Taxa
STATION
02 03 04 06 07 09 10
Tanypodinae
Coelotanypus sp.
Procladius sp.
Tanypus sp.
Tanytarsini
Tanvtarsus
Chironomini
Chironomus sp.
Cladopelma sp.
Crvptochironomus sp.
Dicrotendipes sp.
GIvptotendipes sp.
Microchironomus sp.
Polvpedilum sp.
Orthocladinae
Cricotopus sp.
Total
Taxa Richness
227 121 45
83 280 38
11
79
4 24
238
4
121
1
45
1
11
1
87
2
363
3
61
2
-------
Table 5.13 Mean abundance values (numbers/in , n=5/station) for Mollusca genera
and species collected from the first survey of Saginaw River, MI,
December 1989.
Taxa
02
03
STATION
04 06 07
09
10
Gastropoda
Valvatidae
Valvata lewisi
Valvata tricarinata
Bithyniidae
Bithvnia tentaculata
Ancylidae
Laevapex fucus
Hydrobiidae
Cincinnatia cincinnatiensis
Total
Taxa Richness
0
0
0
0
0
0
0
0
0
0
0
0
0
0
Bivalvia
Sphaeriidae
Musculium sp.
Pisidium sp.
Sphaerium sp.
Sp. unidentified
4
12
Unionidae
Anodonta imbecillis
Anodonta arandis
Eliptio complanata
Total
Taxa Richness
0
0
4
1
8
I
16
2
0
0
0
0
0
0
-------
Table 5.14 Percentage contribution of each taxa to the total number
of taxa collected from the third survey of Saginaw
River, MI, June 1990.
Taxa
Oligochaeta
Chironomidae
Bivalvia
Gastropoda
Ephemeroptera
Odonata
Pelcoptera
Hemiptera
Megaloptera
Trichoptera
Coleoptera
Diptera
Hirudinea
Amphipoda
01
69
31
0
0
0
0
0
0
0
0
0
0
0
0
02
99
<1
<1
0
0
0
0
0
0
0
<1
<1
0
0
STATION
05 06
94
5
<1
0
0
0
0
0
0
0
0
<1
0
0
95
5
0
0
0
0
0
0
0
0
0
0
0
o
08
69
30
0
0
0
0
0
0
0
0
0
<1
0
0
16
90
9
<1
0
0
0
0
0
0
0
0
<1
0
0
24
86
13
0
0
0
0
0
0
0
0
<1
0
0
0
-------
Table 5.15 Mean abundance values (number/m, n=5/station) for aquatic
Oligochaeta species collected from the third survey of Saginaw
River, MI, June 1990.
Taxa
Naididae
Dero diaitata
UN1
Tubif icidae
Aulodrilus piaueti
Aulodrilus limnobius
Aulodrilus pluriseta
Ilvodrilus tetnpletoni
Limnodrilus cervix
Limnodrilus cervix variant
Limnodrilus claparedianus
Limnodrilus clap.-cerv. complex
Limnodrilus hof fmeisteri
Limnodrilus maumeensis
Limnodrilus udekemianus
Limnodrilus sp.
Quistadrilus multisetosus "1"
Quistadrilus multisetosus "m"
Tubifex tubifex
UIW/OCC2
UW/CC3
— i . . — .
'
-------
Table 5.16 Mean abundance values (number/m2, n=5/station) for Chironomidae
genera collected from the third survey of Saginaw River, MI,
June 1990.
Taxa
Orthocladinae
Cricotopus sp.
Total
Taxa Richness
STATION
01 02 05 06 08 16
2204 23
2 3
144
2_
15
3
360
7
389
4_
24
Tanypodinae
Coelotanypus sp.
Procladius sp.
Tanypus sp.
Tanytarsini
Tanvtarsus
Chironomini
Chironomus sp.
Cladopelma sp.
Crvptochironomus sp.
Dicrotendipes sp.
Glyptotendipes sp.
Microchironomus sp.
Polypedilum sp.
- - 8
144 15 125 4 147 234
_ _ _ 4 -
- - - 8
2045 4 19 4 178 34
_ _ _ - - -
- 4 8 4 113
- - - - - -
_ _ _ _
- 11 8
_ _ _ - - -
-
87
8
~
26
—
4
—
-
-
-
125
4
-------
Table 5.17 Mean abundance values (numbers/m , n=5/station) for Mollusca genera
and species collected from the third survey of Saginaw River, MI,
June 1990.
Taxa
01
_Q2_
STATION
05 06 08 16 24
Gastropoda
Valvatidae
Valvata lewisi
Valvata tricarinata
Bithyniidae
Bithvnia tentaculata
Ancylidae
Laevapex fucus
Hydrobiidae
Cincinnatia cincinnatiensis
Total
Taxa Richness
0
0
0
0
0
0
0
0
0
0
0
0
0
0
Bivalvia
Sphaeriidae
Musculium sp.
Pisidium sp.
Sphaerium sp.
Sp. unidentified
Unionidae
Anodonta imbecillis
Anodonta arandis
Eliptio complanata
Total
Taxa Richness
11
0
0
8
1
11
1
0
0
0
0
4
1
0
0
-------
Table 5.18 Comparison of total invertebrate abundance (number/m ) values
for each area of concern (AOC).
Studv
Taxa
Abundance ( number )
Oligochaeta
Mean
Median
Range
Std. Dev.
Chironomidae
Mean
Median
Range
Std . Dev .
Total
Mean
Median
Range
Std. Dev.
BR-01
(n=49)
8726
7333
170-28047
6669
426
132
0-9148
1325
9323
8108
189-28558
6680
IH-01
-------
Table 5.19 Comparison of mean abundance (number/m2) values among stations within an AOC
using Duncans Multiple Range test. Values with the same letter within ans AOC
are not significantly different (P<0.05).
AOC
BR-01
BR-01
BR-01
BR-01
BR-01
BR-01
BR-01
BR-01
BR-01
BR-01
IH-01
IH-01
IH-01
IH-01
IH-01
IH-01
IH-01
Station
06
I
08
07
04
02
03
05
10
01
09
10
04
05
03
08
06
07
Oligochaeta
19157
16220
14496
8407
7394
7059
6403
3092
2714
1984
493887
5935
5198
3765
2903
1501
552
A
A
A
B
BC
BCD
BCD
CD
CD
D
A
B
BC
BC
BC
BC
C
Station
10
04
09
01
08
03
06
02
07
05
10
04
03
06
07
08
05
Chironomidae
2771
306
287
197
181
151
114
110
26
11
4
0
0
0
0
0
0
A
B
B
B
B
B
B
B
B
B
A
A
A
A
A
A
A
Station Total
06
08
07
04
03
02
05
10
01
09
10
04
05
03
08
06
07
19418
16447
14700
9461
7536
7530
6445
6067
3016
2294
493917
6025
5307
3791
2907
1501
609
A
A
AB
BC
BC
CD
CD
CD
D
D
A
B
BC
BC
BC
BC
C
-------
Table 5.19 Continued.
AOC
SR-01
SR-01
SR-01
SR-01
SR-01
SR-01
SR-01
SR-03
SR-03
SR-03
SR-03
SR-03
SR-03
SR-03
Station
03
02
07
04
06
09
10
01
02
16
05
08
24
06
Oligochaeta
6974
6418
5954
3621
1682
1478
813
4944
3735
3538
2862
820
805
302
A
A
A
B
C
C
C
A
AB
B
B
C
C
C
Station
09
02
03
07
10
04
06
01
16
08
05
24
02
06
Chironomidae
363
238
121
87
61
45
11
2204
389
360
144
125
23
15
A
AB
BC
C
C
C
C
A
B
B
B
B
B
B
Station
03
02
07
04
09
06
10
01
16
05
02
08
24
06
Total
7129 A
6664 A
6056 A
3686 B
1890 C
1705 C
888 C
7152 A
3977
3047
2888
1157
941
321
B
B
BC
CD
D
D
-------
Table 5.20 Comparison of Chironomidae deformities (%) for each
area of concern (AOC).
AOC/STATION
BR-01-01
BR-01-02
BR-01-03
BR-01-04
BR-01-05
BR-01-06
BR-01-07
BR-01-08
BR-01-09
BR-01-10
MEAN
IH-01-10
SR-01-02
SR-01-03
SR-01-04
SR-01-06
SR-01-07
SR-01-09
SR-01-10
MEAN
SR-03-01
SR-03-02
SR-03-05
SR-03-06
SR-03-08
SR-03-16
SR-03-24
n
52
29
32
102
3
30
7
48
76
210
10
2
63
32
12
3
23
96
6
7
334
6
38
4
94
103
33
OCCURRENCE OF
DEFORMITIES (%)
8
7
6
5
0
7
14
4
17
7
7
100
14
15
25
0
13
16
6
13
2
14
23
25
16
21
16
MEAN
17
Overall occurance among individual replicate grabs
Buffalo River (19/42)
Indiana Harbor (1/1)
Saginaw River 1 (17/29)
Saginaw River 3 (23/30)
45
100
59
77
-------
CHAPTER 6; AMES MUTAGENICITY ASSAY
Papoulias, D. and D.R. Buckler
INTRODUCTION
The presence of chemical contaminants in the Great Lakes is well-
documented. In 1980 the International Joint Commission (IJC) estimated that
about 2,500 chemicals were in common use in the Great Lakes Basin ecosystem
(International Joint Commission 1980). A comprehensive review of fish-tissue
residues prepared by the IJC's Great Lakes International Surveillance Plan
(Hesselberg and Seelye 1982) tentatively identified 476 organic compounds.
Most of these organic compounds are also present in Great Lakes sediments, and
extracts of these sediments have tested positive in mutagenicity assays (Allen
et al. 1983, Metcalfe et al. 1990, Maccubbin and Ersing 1991). Baumann (1984)
and Baumann and Whittle (1988) suggested a link between high levels of
industrial contaminants and frequency of tumors in fish populations. Recent
study of contaminated sediments from tributaries of the Great Lakes using
Medaka (Orvzias latipesl supports the hypothesis that chemical contaminants in
lake sediment are associated with neoplasms in fishes near contaminated sites
(Fabacher et al. 1991).
Short-term bioassays are a fundamental component of tier testing to
assess the genotoxicity (DNA-damaging properties) of chemical contaminants
(Barfknecht and Naismith 1984, Lovell 1989, International Joint Commission
1988). The bioassays are typically performed in vitro over a period of hours
to weeks and encompass a wide range of genetic endpoints. In general, short-
term assays identify specific genotoxic contaminants or those in complex
mixtures of contaminants, provide baseline data for monitoring changes in
environmental conditions, and predict potential long range genotoxic health
effects (Epler 1980). Genotoxicity assays encompass a broad range of tests
that detect damage at the genetic level, while the mutagenicity end-point
specifically identifies those changes to the DNA that are heritable.
Mutagenicity assays are relatively rapid and inexpensive and are especially
useful for establishing priorities for more definitive chemical analysis and
validation testing with whole animals (Ashby 1988, Ashby and Tennant 1988;
also see Benigni 1990).
In this investigation, part of a series of extensive sediment studies,
the Ames/Salmonella Test (Maron and Ames 1983) was used to determine the
mutagenicity of organic chemical extracts of contaminated sediments from three
priority areas: Saginaw River, Indiana Harbor, and Buffalo River.
6-1
-------
MATERIALS AND METHODS
SUMMARY OF SAMPLES TESTED
Sediment samples were collected at 29 stations from 1988 to 1990 from
the three AOCs. At each AOC, splits of the collected sediment were used to
prepare blind replicate samples. For additional details on stations and
sample collection, see the section in Chapter 1 dealing with Sediment
Collection, Handling, and Storage and the LLRS 1991 Final Report.
Indiana Harbor
Grab samples from each of 7 stations from the first survey of Indiana
Harbor were assayed. The stations included: 3, 4, 5, 6, 7, 8, and 10. A
replicate sample was prepared from Station 7. See Table 2.1 for complete
station identification numbers.
Buffalo River
Grab samples from each of 10 stations from the first survey of the
Buffalo River were assayed. The stations included: 1, 2, 3, 4, 5, 6, 7, 8,
9, and 10. A replicate sample was prepared from Station 6. See Table 2.1 for
complete station identification numbers.
Saoinaw River
Saginaw River was sampled twice. Grab samples from each of 7 stations
from the first survey of Saginaw River were assayed. The stations included:
2, 3, 4, 6, 7, 9, and 10. A replicate sample was prepared from Station 2.
Grab samples from each of 7 stations from the third survey of Saginaw River
were assayed. Sampling of two of the initial stations was repeated. The
stations included: 1, 2, 5, 6, 8, 16, and 24. Two replicate samples of
Station 1 were included. In addition, various depths of core samples from
Stations 2, 5, and & collected during the third survey were assayed. See
Table 2.1 for complete station identification numbers.
SEDIMENT PHYSICAL AND CHEMICAL CHARACTERIZATIONS
Sediment physical characterization included (1) organic carbon, (2)
inorganic carbon, (3) percentage water, and (4) particle size. Sediment
chemical characterization included (1) metals (Ag, As, Cd, Cr, Cu, Hg, Mn, Ni,
Pb, Se, Zn), (2) organometals (butyltins and methyl mercury), (3) acid
volatile sulfide (AVS) and simultaneously extracted metals (SEM), (4)
chlorinated pesticides, (5) PCB congeners, (6) polychlorinated dioxins and
furans, and (7) select PAHs. For additional details on physical and chemical
characterizations, see Chapters 2 and 3.
EXPOSURE PROCEDURES
Disposable gloves, tyvek suits, sleeve protectors, and safety glasses
were used for handling potentially genotoxic substances. All toxicological
6-2
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transfers were conducted in a laminar flow hood under yellow light. Toxicant
disposal was pursuant to National Fisheries Contaminant Research Center
(NFCRC) Safety Plan.
Methylene chloride extracts of test sediments and a control sediment
were prepared by Battelle Northwest as follows: sub-samples of about 100 to
200 g wet sediment were dried with sodium sulfate, triple-extracted with
methylene chloride, and concentrated by roto-evaporation to 11 mL (Eric
Crecilius, Battelle, Sequim, WA, personal communication). Upon receipt at
NFCRC, extracts were stored in amber glass vials with wax septa caps at 4'C
until processed.
Processing of methylene chloride extracts included gel permeation
chromatography (GPC) followed by transfer into dimethyl sulfoxide (DMSO).
This chromatographic system involves a mixture of size exclusion and
adsorption to remove interfering materials from the sediment extracts. A
chromatographic column was packed with 60 g of Bio-beads" SX-3 (200 to 400
mesh) and was wetted with 100 mL of a 50:50 (v:v) mixture of methylene
chloride and cyclopentane. Before application of the methylene chloride
sediment extracts, the GPC was calibrated with a standard solution containing
di(2-ethylhexyl) phthalate (DEHP), Dacthal, and pyrene. The potential
interferents (alkanes, phthalate esters, fatty acids and their esters,
biogenic pigments, and tannic acids) elute earlier than do the standards.
Analytes that are traditionally targeted for mutation assays, such as PCBs,
most organochlorine pesticides, phenols, chlorinated dioxins and furans, and
neutral and nitrogen-containing PAHs, elute from the gel later than the
standards.
Samples collected from GPC were concentrated by roto-evaporation to
about 5 mL. Five mL of DMSO was then added and the remaining methylene
chloride:cyclopentane was evaporated under a gentle stream of nitrogen.
Procedures were conducted under yellow lights to reduce possible photo-
degradation. Resulting extracts, representing 5 or 10 g-equivalents dry
sediment/mL extract, were kept in the dark at room temperature in 8-mL amber
glass vials with teflon cap liners.
The Ames assay procedure generally followed Maron and Ames (1983) for
the plate incorporation test with preincubation. Briefly, 100 /tL of cultured
test strain was combined with 500 /iL of either phosphate buffer or activating
S9 enzyme mixture. To this, 100 j«L of extract or DMSO solvent was added and
the entire mixture was incubated at 37*C for 20 to 30 min in a dry block
heater. After incubation, 2 mL of top agar containing trace histidine and
biotin was added, and the mixture was poured onto agar plates. Plates were
incubated in the dark at 37"C and the resulting colonies were counted at 72 h.
Plates with fewer than 100 colonies were counted manually; all other plates
were mechanically enumerated with an electric counter as described by Johnson
(1990).
The experimental design consisted of 3 replicates of 3 nominal
concentrations of sediment extract: 5 or 10, 50 or 100 and 500 or 1000 mg-
eguivalents dry sediment/plate; and 3 concentrations of S9: 0, 1, 3 mg-
equivalents protein/plate; tested with 4 strains of Salmonella tvphimurium,
TA97a, TA98, TA100 and TA102 (Table 6.1). Preparation of reagents and
solutions used in the Ames assay was as described by Maron and Ames (1983)
except that the 30% S9 mixture (3 mg/plate) contained twice the normal amount
of NADP co-factor with a corresponding decrease in water (Gatehouse 1987). A
positive mutagenic response was indicated when the number of revertants on
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test plates was greater than or equal to 2 times the number of colonies on the
DMSO solvent control plate.
In order to discriminate between prototroph (revertant) and auxotroph
(non-revertant) colonies, an additional step was included to confirm histidine
independence. Plates indicating potential mutagenicity (e.g., > 2 times their
respective controls) were evaluated by lightly stamping the colonies with a
velvet-covered disk and then transferring the colonies to new minimal glucose
plates containing biotin but no histidine. If the pattern and number of
transferred colonies resembled that on the original plate, the response was
scored as mutagenic. When relatively few colonies transferred, the original
plates contained primarily auxotrophs resulting from cytotoxicity.
STATISTICAL ANALYSES
Means and standard deviations of the number of colonies/plate were
calculated from the three replicate plates. Test results were summarized
based on three categories: no effect, cytotoxic, and mutagenic. Plates were
scored as "Mutagenic" if mean revertants on extract exposed plates were
greater or equal to two times the mean number of spontaneous revertants on
solvent control plates. Plates were scored as "Cytotoxic" if mean revertant
values on extract exposed plates were less than half the mean spontaneous
revertants on solvent control plates, or if cytotoxicity was detected through
the confirmation step. "No-effect" describes those plates that were neither
identified as cytotoxic nor mutagenic. Means and standard deviations are
provided in Appendices 6.1 to 6.13.
QUALITY CONTROL AND QUALITY ASSURANCE
Precision in the Ames Salmonella microsome test was assessed through
replication (triplicate minimum), inclusion of splits of sediment samples
which were run blind, and by making three counts per plate with an electronic
colony counter.
The integrity and sensitivity of the test organisms used in the Ames
test was assessed by confirmation of genotypes, measurement of spontaneous
reversion frequency, and confirmation of reversion properties of the tester
strains using the known mutagens 2,4,7-trinitro-9-fluorenone; 2-aminofluorene;
methyl methanesuIfonate; and benzo[a]pyrene (Maron and Ames 1983).
Additional quality control included determination of S9 protein content,
(-20 mg/ml; Bradford 1976) and EROD activity (-7.05 nmol/mg/min; Pohl and
Fouts 1980). A most-probable-number test was used to estimate population
densities of Salmonella cell cultures. Procedural blanks (no sediment) from
the extraction process were prepared and control samples (extracts from a fine
silt- and clay-particle size sediment obtained from local undisturbed
agricultural soil (Ingersoll and Nelson 1990) were prepared and assayed along
with AOC sediments. See also the Quality Assurance Project Plan for
"Assessment and Remediation of Contaminated Sediments (ARCS) Assistance",
National Fisheries Contaminant Research Center.
6-4
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RESULTS
Mutagenicity was detected in both grab and core samples from every
station tested. Of the four strains, only TA98 identified 100% of the
stations as mutagenic.
INDIANA HARBOR
Without Metabolic Activation
Mutagenicity was detected at Station 7 with strain TA98 (Table 6.2,
Appendix 6.1). No effect of the extracts was observed at the lowest dose (5
mg-equiv sediment) at all stations except Stations 8 and 10 with strains TA98
and TA100. Cytotoxicity tended to increase with increasing dose of sediment
extract and all but Station 10 samples were cytotoxic at the highest dose.
With Metabolic Activation
Mutagenicity was detected at each Indiana Harbor station with strains
TA98 and TA100 at one or more concentrations of sediment extract or S9 (Table
6.3, Appendix 6.2). Strain TA97a detected mutagenicity at every station
except 5 and 7, and strain TA102 detected mutagenicity at every station except
Station 10.
BUFFALO RIVER
Without Metabolic Activation
A no-effect response was obtained for all Buffalo River stations (except
Stations 4 and 6) at the lowest concentration of sediment extract, but all
sample extracts were cytotoxic at the highest concentration (Table 6.4,
Appendix 6.3). Only Station 3, with strain TA100 at 100 mg-equiv sediment
indicated mutagenicity.
With Metabolic Activation
All stations tested with strains TA97a, TA100 and TA102 responded with
no effect at the lowest concentration of sediment extract at both
concentrations of S9 tested (Table 6.5, Appendix 6.4). No mutagenicity was
detected with strain TA102, while either strain TA97a or TA100 indicated
mutagenicity at every station except Station 3. All stations were mutagenic
with strain TA98 generally at more than one concentration of sediment extract
and S9.
SAGINAW RIVER
Without Metabolic Activation
In the first survey samples from Saginaw River, Station 2 was mutagenic
with strain TA98 (Table 6.6, Appendix 6.5). Except for Station 2, samples
from all stations indicated a no-effect response for all strains at the lowest
6-5
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extract concentration and a cytotoxic response at the highest concentration.
Overall, results from the third survey of Saginaw River were similar (Table
6.7, Appendix 6.6). Station 2, although mutagenic in the first set of samples
was not mutagenic in the set of samples from the third survey. A positive
mutagenic response was observed for Station 24 with strain TA100.
No-effect and cytotoxicity patterns were generally the same for the core
samples as for surficial grab samples from the third survey collection (Table
6.8, Appendix 6.7). Mutagenicity was detected only in one section of core
from Station 2 with strain TA100.
With Metabolic Activation
Assay results for all stations from the first survey samples were
similar with no mutagenicity evident for strains TA97a or TA102 (Table 6.9,
Appendix 6.8). With strain TA100 this pattern varied only at Station 6 which
was mutagenic. In contrast, with strain TA98 mutagenicity was detected at
each station. Station 6 with strain TA100 was again mutagenic in grab samples
from the third survey. Mutagenicity was detected by TA97a at Station 8 in
this survey, but not in the first (Table 6.10, Appendix 6.9). Similar to the
first survey results, no stations from the third survey were mutagenic with
strain TA102. Mutagenicity was not observed at Station 1 with TA98, however
as in the first survey results, mutagenicity was detected at all other
stations with this strain.
Core samples tested with S9 were mutagenic for all strains but TA102 for
at least one section of the core (Table 6.11, Appendix 6.10).
QUALITY CONTROL AND QUALITY ASSURANCE
At the high dose, the control sediment was cytotoxic to most strains
with and without metabolic activation (Tables 6.12 and 6.13, Appendices 6.11
and 6.12). Additionally, there were indications of cytotoxicity and
mutagenicity at lower doses. Only strain TA98 detected the mutagenicity. The
procedural blank (no sediment) was cytotoxic at the highest dose (100 /*L of
undiluted sample per plate) with no S9. Additions of S9 eliminated the
cytotoxic response; mutagenicity was not observed. Results for blind
replicate split samples were similar to their complimentary original samples
except for the second samples from Saginaw River which were more cytotoxic
than the original samples (Tables 6.14 and 6.15, Appendices 6.11 and 6.12).
In each test series, the results of the diagnostic tests confirmed
strain genotype and sensitivity (Table 6.16, Appendix 6.13; Maron and Ames
1983). Bacterial culture densities and spontaneous reversion frequencies were
within acceptable limits (Maron and Ames 1983). Acceptable reversion
frequencies were obtained with each combination of strain and positive mutagen
(Maron and Ames 1983). See also the Data Verification Report for Assessment
and Remediation of Contaminated Sediment Program, Report Number 4,
Environmental Monitoring Systems Laboratory, Office of Research and
Development, U.S.E.P.A.
6-6
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DISCUSSION
Sediments are sinks for many anthropogenic contaminants. As such,
organic extracts of contaminated sediments often contain a wide variety of
chemicals which makes mutagenicity testing of sediments complex. The
individual components of chemical extracts of environmental samples can be
both cytotoxic and mutagenic. A predominance of cytotoxic compounds and
synergistic interactions among individual compounds may mask the expression of
a mutagenic response (Epler 1980, Carver et al. 1985). Consequently, a test
may not detect mutagenicity. Class fractionation of samples before testing
can reduce problems of cytotoxicity and aid in the identification of mutagenic
components (Alfheim et al. 1984). However, individual components of the
sample may be modified or lost and synergistic effects may be altered (Epler
1980). Fractionation may also significantly increase the number of samples to
be tested.
Due to the cytotoxicity of methylene chloride to the bacterial test
strains, the crude sediment extracts could be used only after transfer to an
Ames compatible solvent. Initially, a direct transfer to DMSO and other Ames
solvents, resulted in separation of samples into two distinct phases- a clear
or slightly amber-colored solution with a thicker dark-colored solution on the
surface. Since these samples were not homogenous and components could be
trapped in the upper layer, the crude methylene chloride extracts were
pretreated with gel permeation chromatography (GPC). Gel permeation
chromatography is a size exclusion process that removes large biogenic
molecules (e.g., lipids, tannins) which can interfere with the complete
transfer of material from crude extracts to DMSO.
Optimizing the transfer of complex mixtures containing mutagenic
components into DMSO does not ensure a positive response with the Ames assay.
Extracts of sediments collected from polluted areas are often quite cytotoxic
(Metcalf 1990, Fabacher et al. 1988) and excessive cytotoxicity can prevent
the expression of mutagenicity (Epler 1980). Dilution can remove
cytotoxicity, but with a concomitant reduction of potentially mutagenic
compounds. Dilution is particularly ineffective when the sample is highly
cytotoxic and only slightly mutagenic (Maron and Ames 1983).
Cellular microsomal and cytosolic enzymes catalyze the biotransformation
of a variety of chemicals. Biotransformation may result in polar metabolites
that are biologically inactive and readily excretable, or may produce
metabolites that are reactive intermediates that can damage DNA. Exogenous S9
enzyme preparations are routinely used in the Ames assay to more closely
approximate the chemical metabolizing abilities of eucaryotes. Maron and Ames
(1983) suggest rat liver as the source of S9, but other species (Mueller et
al. 1980, Johnson 1990, Blevins 1991) may provide better results depending on
the compounds to be tested. The concentration of S9 must also be considered.
Again, Maron and Ames (1983) recommends a 4 and 10% S9 mixture with the caveat
that it may be necessary to define the optimal concentration for a particular
sample.
Coupling extract dilution with S9 and optimizing the S9 concentration,
allowed detection of mutagenicity at more stations than would have been
possible had the samples only been diluted to non-cytotoxic concentrations
then assayed with the standard concentrations of S9. Extracts of sediment
samples from every station for the three AOCs were both cytotoxic and
6-7
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mutagenic to the bacteria. At 7 of 29 stations mutagenicity could not be
identified at dilutions sufficient to eliminate cytotoxicity. Diluting to
non-cytotoxic concentrations likely co-diluted mutagenic components. At these
stations, cytotoxicity reduction by use of S9 enzymes was required to observe
mutagenicity. Moreover, for 3 of the 29 stations, an increase in S9
concentration from 1 to 3 mg-equiv. protein per plate was necessary for
detection of mutagenicity. This procedure^of combining extract dilution with
additions of increasing concentrations of S9, identified mutagenicity in the
complex contaminant mixtures from each station in the AOCs.
Use of S9 with single compounds allows distinction of classes of
mutagens that are direct acting from compounds requiring metabolic activation.
It is more difficult to identify these classes of mutagens with complex
mixtures (Ball et al. 1990, Pederson and Siak 1981). For the purpose of this
study, mutagens were identified as direct-acting when a positive response was
observed in the absence of S9. Promutagens were identified when addition of
S9 to non-cytotoxic dilutions, or addition of S9 beyond that necessary for
elimination of cytotoxicity, resulted in a positive response. Mutagenicity in
2 of the samples could be attributed to promutagens alone. Two other samples
contained both promutagens and direct acting mutagens. At the remaining 25
stations, cytotoxicity was not sufficiently reduced through dilution or
addition of S9 to completely characterize the classes of mutagens in the
sample. For example, the positive response from Indiana Harbor Station 6 with
strain TA98 at 5 mg-equiv. sediment, can be attributed to promutagens since
there was no observed effect except with metabolic activation (Tables 6.2 and
6.3). If the sample contains direct-acting mutagens, they have been diluted
out at this concentration. At 50 mg-equiv. sediment the sample is cytotoxic
and upon addition of S9 we observe a mutagenic response. This positive
response may be due to reduction of cytotoxicity alone (allowing the
expression of direct-acting mutagens), reduction of cytotoxicity coupled with
activation of promutagens, or a combination of both.
Two strains detected mutagenicity more frequently. Strain TA98 with S9
detected mutagenicity at every station. Strain TA98 without S9 detected
mutagenicity at 3 stations. Of the other 3 strains, positive responses were
most commonly observed with TA100. In a comparison by Zeiger et al. (1985),
strains TA98 and TA100 positively identified 89% of the mutagenic chemicals in
the NTP (National Toxicity Program) database. While strain TA98 detected
mutagenicity at no and low concentrations of S9, the other strains generally
required high concentrations of S9 for a mutagenic response.
Chemical analysis (Table 6.17; also see Chapter 4) of whole sediment
from the three AOCs and the control sediment indicates that as a class, PAHs
comprise the greatest percent by weight of the total identified organics.
High molecular weight PAHs were more abundant than low molecular weight PAHs.
Of the classes of compounds tested, PAHs contain the greatest number of
compounds known to be mutagenic in the Ames assay. Within this class, the
high molecular weight PAHs are more likely to cause mutation in the Ames
assay. It should be noted, however, that many of the other sediment
contaminants identified are reported to cause DNA damage in other organisms.
Mutagenicity detected in samples from the AOCs could be primarily
attributable to the PAH compounds. Other investigations have identified
sediment PAHs as the most likely contaminant causing a positive Ames result
(Maccubbin and Ersing 1991, Abe et al. 1989, Grifoll et al. 1988, Odense et
al. 1988, Oishi and Takahashi 1987, Sato et al. 1983, Waldron and White 1989).
6-8
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Crude extracts of freshwater sediments from Hamilton Harbor, Lake Ontario,
Canada and marine sediments from Long Island Sound, USA. contained
concentrations of high molecular weight PAHs similar to those in the present
study and were also mutagenic while control samples were not (Metcalfe et al.
1990, Gardner et al. 1987). Furthermore, PAH contaminated sediment reportedly
caused induction of neoplasia in oyster and flounder (Gardner et al. 1991).
A ranking was develped to describe the relative cytotoxicity and
mutagenicity among stations and AOCs. A no-effect response at all three doses
was given a value of 1; no-effect for the two lowest doses with mutagenicity
or cytotoxicity only at the highest dose was given a value of 2; no-effect at
only the lowest of the three doses received a value of 3; and mutagenicity or
cytotoxicity at all three doses received a, value of 4. The values from each
of the four strains tested for each sample were summed to obtain a score from
4 (low) to 16 (high) for each station. Figures 6.1 and 6.2 compare the
cytotoxicity and mutagenicity of the stations. For each AOC, the procedural
blank (no sediment) had a lower score than any of the stations. For Indiana
Harbor Station 6 had the highest mutagenicity score and Stations 5, 8, and 10
had the lowest mutagenicity scores. Stations 4 and 6 had the highest scores
and Station 3 the lowest for Buffalo River. Saginaw River Stations 1 and 4
received the high and low scores, respectively. No distinct relationship
between cytotoxicity and mutagenicity scores is apparent, although the Saginaw
station with the highest mutagenicity score (Station 1) also has the highest
cytotoxicity score and Station 4 scored lowest for both cytotoxicity and
mutagenicity. The range of mutagenicity and cytotoxicity scores and the modes
for all stations for each AOC are depicted in Figures 6.3 and 6.4.
Mutagenicity scores for Indiana Harbor were distributed over a narrower range
than those for Buffalo and Saginaw Rivers. In general, Indiana Harbor
stations scored at the upper end of the range of all stations combined while
Buffalo and Saginaw River stations scored in the middle to lower end of the
range.
The information provided through the use of these station ranks needs to
be evaluated with caution. The control sediment is considerably lower in
potential types and quantities of mutagenic compounds than the lowest ranking
station from the AOCs (Table 6.17). Yet the control sediment out-ranks 7 of
the 29 stations for mutagenicity. A test with such a high degree of
sensitivity is probably most useful for detecting presence or absence of
mutagens and its utility for more detailed discrimination is questionable.
The ultimate goal of mutagenicity testing is to identify the carcinogenic and
intergenerational effects a compound may have on organisms of concern.
Mutagenicity is only one of many genotoxicity end-points that can contribute
information towards this goal. Furthermore, detection of a mutation in the
Salmonella organisms in the Ames assay is dependent upon optimization of the
test parameters. For these reasons, it is generally accepted that a tiered
approach using a variety of tests and end-points is most valuable for
detecting potential gene and chromosome damage (Heinze and Poulsen 1983).
These studies demonstrate the utility of the Ames assay for screening
crude extracts of complex mixtures. By varying S9 concentration and extract
dilution, cytotoxicity was controlled so that mutations could be observed.
However, due to the assay's sensitivity, it is not able to discriminate among
discrete mutagenic responses. Fractionation of samples before testing may
allow a greater degree of discrimination and aid in identifying those
compounds which are causing the observed mutagenicity. Additionally, the Ames
6-9
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test as applied in this study does not address the issue of bioavailability of
mutagenic compounds in sediments. Development of methods to test elutriates
and pore waters and studies utilizing passive membrane bag samplers may be
useful for this purpose.
6-10
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Oishi, S. and O. Takahashi. 1987. Mutagenicity of Tama River sediments. Bull.
Environ. Contain. Toxicol. 39:696-700.
Pederson, T.C. and J-S. Siak. 1981. The activation of mutagens in diesel
particle extract with rat liver S9 enzymes. Journal of Applied Toxicology
l(2):61-66.
Pohl, R.J. and J.R. Fouts. 1980. A rapid method for assaying the metabolism
of 7-Ethoxyresorufin by microsomal subcellular fractions. Analytical
biochemistry 107:150-155.
Sato, T., T. Momma, Y. Ose, T. Ishikawa and K. Kato. 1983. Mutagenicity of
Nagara River Sediment. Mutation Research 118:257-267.
Schoeny, R. 1982. Mutagenicity testing of chlorinated biphenyls and
chlorinated dibenzofurans. Mutation Research 10:45-56.
Waldron, M.C., and A.R. White. 1989. Non-volatile chemical mutagens in
sediments of the Kanawha River, West Virginia. Ohio J. Sci 89(5):176-180.
6-13
-------
Zeiger, E., K.J. Risko, and B.H. Margolin. 1985. Strategies to reduce the cost
of mutagenicity screening with the Salmonella Assay. Environmental Mutagenesis
7:901-911.
6-14
-------
LIST OF FIGURES IN CHAPTER 6
6.1 Toxicity Scores for AOC Stations.
6.2 Mutagenicity Scores for AOC Stations.
6.3 Summary of Toxicity Score Results for AOCs.
6.4 Summary of Mutagenicity Score Results for AOCs;
-------
16
Toxicity
Indiana Harbor
Figure 6.1
8
CO
8H
.'A
06
B 03 05 07 04
Stations
10
08
16
Buffalo River
12-
£ *
8
CO
8-
Z rx/
B 08 09 01 02 03 10 05 07 04 06
Stations
16
Saginaw River
^2•\
O it
o
o
CO
8H
B 04 05 09 24 02 03 06 07 10 16 08 01
Stations
-------
16
a)12"*
o
o
co *
8-
v//\
B
08
Mutagenicity
Indiana Harbor
05
10 04
Stations
03
Figure 6.2
07
06
16
Buffalo River
12-
£
8
co *
B 03 01 08
05 10 02
Stations
09 07 04 06
16
Saginaw River
£
8
12-
*
8-
/
B 04 08 09 24 02 03 16
Stations
07 10 05 06 01
-------
Figure 6.3
Toxicity
ID
12 -
*
-------
Score
co
ro
i
o>
o 2
o 3
c
rt
0)
vQ
(D
3
H-
O
H-
ft
CO
3
C
r-(
(D
CT>
-------
Table 6.1 Ames Assay test strain characteristics (Maron and
Ames 1983).
Strain
TA97a
TA98
TA100
TA102
Mutation
rfa1 uvrB2 HIS3
+ - hisD6610
+ - hisD3052
+ - hisG46
+ + hisG428
Specificity
frameshift
frameshift
base-pair
excision
repair
Spontaneous
Revertants
90-180
30-50
120-200
240-320
1 causes partial loss of the bacteria's lipopolysaccharide
barrier
2 causes deletion of a gene coding for the DNA excision
repair system
3 mutation at the histidine operon
-------
Table 6.2 Results of Ames assay (n=3) for 4 strains, 3 doses of extract,
and no S9 for 7 sites in Indiana Harbor. "T" = toxicity,
"NE" = no-effect, and "M" = mutagenicity.
Strain
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA100
TA100
TA100
TA102
TA102
TA102
Dose
(mg-equiv.
sediment )
5
50
500
5
50
500
5
50
500
5
50
500
3
X
NE
NE
T
NE
NE
T
NE
NE
T
NE
NE
T
4
X
NE
T
T
NE
T
T
NE
T
T
NE
T
T
X
NE
T
T
NE
T
T
NE
NE
T
NE
NE
T
X
NE
T
T
NE
T
T
NE
T
T
NE
T
T
7
X
NE
NE
T
NE
M
T
NE
T
T
NE
T
T
8
X
NE
T
T
It
T
*
T
X
T
NE
T
T
NE
T
T
X
NE
T
T
T
T
T
T
T
T
NE
NE
NE
only 2 replicates
-------
Table 6.3 Results of Ames assay (n=3) for 4 strains, 3 doses of extract, and 2
concentrations of S9 for 7 stations in Indiana Harbor. "T" = toxicity,
"NE" = no-effect, and "M" = mutagenicity.
Station
Strain
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA98
TA98
TA98
TA100
TA100
TA100
TA100
TA100
TA100
TA102
TA102
TA102
TA102
TA102
TA102
Dose
(mg-equiv.
sediment)
5
5
50
50
500
500
5
5
50
50
500
500
5
5
50
50
500
500
5
5
50
50
500
500
S9
(mg-
equiv.
protein)
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
3
X
NE
NE
M
NE
M
NE
NE
NE
M
M
NE
M
NE
NE
M
M
NE
M
NE
M
NE
M
NE
NE
4
X
NE
NE
NE
NE
NE
M
NE
NE
NE
M
T
T
NE
NE
NE
M
T
T
NE
NE
NE
M
T
M
5
X
NE
NE
NE
NE
NE
NE
NE
NE
M
M
NE
M
NE
NE
M
M
NE
NE
NE
NE
M
M
M
M
6
X
NE
NE
M
NE
M
NE
M
NE
M
M
NE
M
NE
NE
M
M
M
M
M
M
M
M
T
M
7
X
NE
NE
NE
NE
T
T
NE
NE
NE
M
T
T
NE
NE
NE
M
T
T
NE
M
NE
M
T
T
8
X
NE
NE
NE
NE
M
NE
K
M
It
M
K
NE.
M"
K
NE
K
NE
NE
NE
NE
NE
M
M
NE
M
NE
NE
NE
NE
10
X
NE
NE
M
NE
M
NE
NE
NE
M
M
T
T
NE
NE
NE
M
NE
NE
NE
NE
T
T
T
T
only 2 replicates
-------
Table 6.4 Results of Ames assay (n=3) for 4 strains, 3 doses of extract, and no
S9 for 10 stations in Buffalo River. "T" = toxicity, "NE" = no-effect, and
"M" = mutagenicity.
Dose
Strain (mg-equiv
sediment)
TA 97a
TA 97a
TA 97a
TA 98
TA 98
TA 98
TA 100
TA 100
TA 100
TA 102
TA 102
TA 102
10
100
1000
10
100
1000
10
100
1000
10
100
1000
X
NE
T
T
NE
T
T
NE
NE
T
NE
NE
T
X
NE
T
T
NE
T
T
NE
NE
T
NE
NE
T
X
NE
T
T
NE
T
T
NE
M
T
NE
NE
T
X
NE
II
T
T
NE
T
T
T
T
T
NE
T
T
5
X
NE
T
T
NE
T
T
NE
T.
T*
NE
T
T
X
NE
T
T
NE
T
T
T
T
T
NE
T
T
7
X
NE
T
T
NE
T
T
NE
T
T
NE
T
T
X
NE
NE
T
NE
NE
T
NE
NE
T
NE
NE
T
9
X
NE
NE
T
NE
T
T
NE
NE
T
NE
NE
T
10
X
NE
T
T
NE
T
T
NE
T
T
NE
NE
T
only 2 replicates
-------
Table 6.5 Results of Ames assay (n=3) for 4 strains, 3 doses of extract, and 2
10 stations in Buffalo River. "T" - toxicity, "NE" - no-effect, and
concentrations of S9 for
"M" >• mutagenicity.
Strain
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA98
TA98
TA98
TA100
TA100
TA100
TA100
TA100
TA100
TA102
TA102
TA102
TA102
TA102
TA102
Dose
(mg-eguiv.
sediment)
10
10
100 .
100
1000
1000
10
10
100
100
1000
1000
10
10
100
100
1000
1000
10
10
100
100
1000
1000
S9
(mg-
eguiv.
orotein)
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
X
NE
NE
NE
NE
M
M
NE
NE
M
M
M
M
NE
NE
NE
NE
T
M
NE
NE
NE
NE
NE
NE
X
NE
NE
NE
M
T
NE
NE
NE
M
M
T
T
NE
NE
M
M
T
T
NE
NE
NE
NE
NE
NE
X
NE
NE
NE
NE
T
NE
NE
NE
M
M
T
M
NE
NE
NE
NE
T
T
NE
NE
NE
NE
NE
NE
X
NE
NE
M
M
T
T
M
NE
M
M
T
T
NE
NE
NE
M
T
T
NE
NE
NE
NE
T
T
X
NE
NE
NE
NE
NE
NE
M
NE'
M
M
T
T
NE
NE
NE
M
T
NE
NE
NE
NE
NE
NE
T
X
NE
NE
M
M
T
T
M
NE
M
M
T
T
NE
NE
NE
M
T
NE
NE
NE
NE
NE
T
T
X
NE
NE
M
NE
T
T
M
M
M
M
T
T
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
8
X
NE
NE
M
NE
T
M
NE
NE
M
M
T
M
NE
NE
NE
NE
NE
M
NE
NE
NE
NE
T
NE
X
NE
NE
M
NE
T
T
NE
T
M
M
T
T
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
1 only 2 replicates
-------
Table 6.6 Results of Ames assay (n=3) for 4 strains, 3 doses of extract,
and no S9 for 7 stations from the first survey (grab) of Saginaw River.
"T" = toxicity, "NE" = no-effect, "M" = mutagenicity.
Station
Strain
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA100
TA100
TA100
TA102
TA102
TA102
Dose
(mg-equiv.
sediment )
10
100
1000
10
100
1000
10
100
1000
10
100
1000
X
NE
NE
T
M
T
T
NE
T
T
NE
NE
T
X
NE
T
T
NE
NE
T
NE
T
T
NE
NE
T
4
X
NE
NE
T
NE
T
T
NE
NE
T
NE
NE
T
6
X
NE
T
T
NE
T
T
NE
NE
T
NE
NE
T
7
X
NE
NE
T
NE
T
T
NE
T
T
NE
NE
T
9
X
NE
NE
T
NE
NE
T
NE
T
T
NE
NE
T
10
X
NE
NE
T
NE
T
T
NE
T
T
NE
NE
T
-------
Table 6.7
Results of Ames assay (n=3) for 4 strains, 3 doses of extract,
and no S9 for 7 stations the third survey (grab) of Saginaw River.
"T" = toxicity, "NE" = no-effect, "M" = mutagenicity.
Station
Strain
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA100
TA100
TA100
TA102
TA102
TA102
Dose
(mg-equiv.
sediment)
10
100
1000
10
100
1000
10
100
1000
10
100
1000
X
NE
T
T
NE
T
T
NE
T
T
NE
T
T
2
X
NE
T
T
NE
T
NE
NE
NE
T
NE
NE
T
5
X
NE
NE
T
NE
T
T
NE
NE
T
NE
NE
T
6
X
NE
NE
T
NE
NE
T
NE
NE
T
NE
NE
NE
8
X
T
T
T
NE
T
T
NE
T
T
NE
NE
T
16
X
NE
T
T
NE
T
T
NE
NE
T
NE
NE
T
24
X
NE
T
T
NE
T
T
NE
NE
M
NE
NE
T
-------
Table 6.8 Mean number of revertants (n=3) + St> for 4 strains, 3 doses of extract, and no S9 for
8 core samples from the third survey of Saginaw River. Toxicity determined by visual
inspection or replicate plating is denoted with a "T".
Strain
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA100
TA100
TA100
TA102
TA102
TA102
core
core
3
core
4.
core
5
core
6
core
Dose
(mg-equiv.
sediment)
10
100
1000
10
100
1000
10
100
100 0
10
100
1000
1
X
NE
NE
T
NE
T
T
NE
NE
T
NE
NE
T
52
X
NE
T
T
NE
T
T
NE
T
T
NE
T
T
1
X
NE
T
T
NE
T
T
NE
T
T
NE
T
T
62
X
NE
NE
T
NE
NE
T
NE
NE
T
NE
NE
T
23
X
NE
T
T
NE
T
T
NE
T
T
NE
NE
T
4
X
NE
NE
T
NE
NE
T
NE
NE
M
NE
NE
T
65
X
NE
T
T
NE
T
T
NE
T
T
NE
NE
T
66
X
NE
T
T
NE
T
T
NE
T
T
T
T
T
from 0-2 feet deep
from 2-4 feet deep
from 8-20
from 20 - 35
from 5-13
from 13 - 32
inches deep
inches deep
inches deep
inches deep
-------
Table 6.9
Results of Ames assay (n=3) for 4 strains, 3 doses of extract,
and 2 concentrations of S9 for 7 stations from first survey (grab)
of Saginaw River. "T" = toxicity, "NE" = no-effect, "M" = mutagenicity.
Station
Strain
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA98
TA98
TA98
TA100
TA100
TA100
TA100
TA100
TA100
Dose
(mg-equiv.
sediment)
10
10
100
100
1000
1000
10
10
100
100
1000
1000
10
10
100
100
1000
1000
S9
(mg-
equiv.
protein)
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
2
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
NE
NE
NE
NE
T
T
3
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
NE
NE
NE
NE
T
T
4
X
NE
NE
NE
NE
T
NE
NE
NE
NE
M
NE
M
NE
NE
NE
NE
NE
NE
6
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
NE
NE
NE
M
T
T
7
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
NE
NE
NE
NE
T
T
9
X
NE
NE
NE
NE
T
NE
NE
NE
M
M
T
M
NE
NE
NE
NE
NE
NE
10
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
NE
NE
NE
NE
T
T
-------
Table 6.9 Continued.
fi-ha-Mon
Strain
TA102
TA102
TA102
TA102
TA102
TA102
Dose
(mg-equiv.
sediment)
10
10
100
100
1000
1000
S9
(mg-
equiv.
croteirn
1
3
1
3
1
3
2
X
NE
NE
NE
NE
T
NE
3
X
NE
NE
NE
NE
T
T
4
X
NE
NE
NE
NE
T
NE
6
X
NE
NE
NE
NE
T
T
7
X
NE
NE
NE
NE
T
T
9
X
NE
NE
NE
NE
T
T
10
X
NE
NE
NE
NE
T
T
-------
Table 6.10 Results of Ames assay (n=3) for 4 strains, 3 doses of extract,
and 2 concentrations of S9 for 7 stations from third survey (grab)
Saginaw River. "T" = toxicity, "NE" = no-effect, "M" = mutagenicity.
Station
Strain
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA98
TA98
TA98
TA100
TA100
TA100
TA100
TA100
TA100
Dose
(mg-equiv.
sediment)
10
10
100
100
1000
1000
10
10
100
100
1000
1000
10
10
100
100
1000
1000
S9
(mg-
equiv.
protein)
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
X
NE
NE
T
T
T
T
NE
NE
T
T
T
T
NE
NE
T
T
T
T
2
X
T
NE
NE
NE
T
T
NE
NE
NE
NE
M
M
NE
NE
T
T
T
T
5
X
NE
NE
NE
NE
T
T
NE
M
M
M
T
T
NE
NE
NE
NE
T
T
6
X
NE
NE
NE
NE
M
NE
NE
NE
NE
NE
M
M
NE
NE
NE
NE
NE
NE
8
X
NE
NE
NE
NE
NE
NE
NE
M
NE
M
T
M
NE
NE
NE
NE
NE
NE
16
X
NE
NE
NE
NE
T
NE
NE
M
M
M
T
M
NE
NE
NE
NE
T
T
24
X
NE
NE
NE
NE
T
NE
NE
NE
M
NE
T
M
NE
NE
NE
NE
NE
M
-------
Table 6.11
Mean number of revertants (n=3) ± SD for 4 strains, 3 doses of extract,
and 2 concentrations of S9 for 8 core samples from third survey of Saginaw River.
Toxicity determined by visual inspection or replicated plating is denoted
with a "T".
Strain
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA98
TA98
TA98
Dose
(mg-equiv.
sediment)
10
10
100
100
1000
1000
10
10
100
100
1000
1000
S9
(mg-
equiv.
protein)
1
3
1
3
1
3
1
3
1
3
1
3
51
X
NE
NE
NE
M
T
NE
M
M
M
M
T
M
52
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
61
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
T
62
X
T
T
T
T
T
T
NE
NE
M
M
T
T
23
X
NE
NE
NE
NE
NE
NE
NE
NE
M
M
NE
M
24
X
NE
NE
NE
NE
M
M
NE
NE
NE
NE
M
M
6s
X
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
66
X
NE
NE
NE
M
T
T
NE
NE
M
M
T
T
-------
Table 6.11 Continued.
Station
Strain
Dose S9
(ma-eouiv. (ma-
51
52
61
62
23
4
c
66
sediment) equiv. _ _______
TA100
TA100
TA100
TA100
TA100
TA100
TA102
TA102
TA102
TA102
TA102
TA102
1 core
2 core
3 core
4 core
5 core
10
10
100
100
1000
1000
10
10
100
100
1000
1000
from 0 -
from 2 -
from 8 -
from 20
from 5 -
protein)
1
3
1
3
1
3
1
3
1
3
1
3
2 feet deep
4 feet deep
20 inches deep
X
NE
NE
NE
M
NE
M
NE
NE
NE
NE
T
NE
X
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
X
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
X
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
X
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
X
NE
NE
NE
NE
NE
M
NE
NE
NE
NE
NE
M
X
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
X
NE
NE
NE
M
T
T
NE
NE
NE
NE
T
T
-35 inches deep
13 inches deep
6 core from 13-32 inches deep
-------
Table 6.12
Results of Ames assay (n=3) for 4 strains, 3 doses of
extract, and no S9 for 4 Quality Assurance and Quality
Control samples. "T" = toxicity, "NE" = no-effect, and
"M" = mutagenicity.
Strain
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA100
TA100
TA100
TA102
TA102
TA102
Dose
(mg-equiv.
sediment)
10
100
1000
10
100
1000
10
100
1000
10
100
1000
BR-CON
X
NE
T
T
NE
T
T
NE
T
T
NE
NE
T
"~"'p^-*-
SR-CON
X
T
T
T
T
NE
T
NE
NE
T
NE
NE
T
IH-BLANKa
X
NE
NE
T
NE
NE
T
NE
NE
T
NE
NE
T
SR-BLANK3
X
NE
NE
T
NE
NE
T
NE
NE
NE
NE
NE
NE
Procedural blanks do not contain sediment.
-------
Table 6.13
Results of Ames assay (n=3) for 4 strains, 3 doses of extract,
and 2 concentrations of S9 for 4 Quality Assurance Quality
Control samples. "T" = toxicity, "NE" = no-effect, and
"M" = mutagenicity.
Sample
Strain
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA98
TA98
TA98
TA100
TA100
TA100
TA100
TA100
TA100
Dose
(mg-equiv.
sediment)
10
10
100
100
1000
1000
10
10
100
100
1000
1000
10
10
100
100
1000
1000
S9
(mg-
equiv.
protein)
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
BR-CON
X
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
NE
NE
NE
NE
T
T
SR-CON
X
NE
NE
NE
NE
NE
NE
NE
M
M
M
T
T
NE
NE
NE
NE
T
T
IH-BLANKa
X
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
T
NE
SR-BLANK8
X
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
-------
Table 6.13 Continued
Strain
TA102
TA102
TA102
TA102
TA102
TA102
Dose
(mg-equiv.
sediment)
10
10
100
100
1000
1000
S9
(mg-
equiv.
protein)
1
3
1
3
1
3
BR-CON
X
NE
NE
NE
NE
T
T
SR-CON
X
NE
NE
NE
NE
T
T
IH-BLANK
X
NE
NE
NE
NE
NE
NE
SR-BLANK
X
NE
NE
NE
NE
NE
NE
a Procedural blanks do not contain sediment,
-------
Table 6.14 Results of Ames assay (n=3) for 4 strains, 3 doses of
extract, and no S9 for replicate samples. "T" = toxicity,
"NE" = no-effect, and "M" = mutagenicity.
Dose
Strain (mg-equiv.
sediment )
TA97a 10*
TA97a 100b
TA97a 1000°
TA98 10*
TA98 100b
TA98 1000C
TA100 10*
TA100 100b
TA100 1000°
TA102 10a
TA102 100b
TA102 1000°
For sample IH-REP
For sample IH-REP
For sample IH-REP
Blind duplicate of
Blind duplicate of
Blind duplicate of
0 Blind duplicate of
IH-REPd
X
NE
NE
T
NE
T
T
T
T
T
NE
T
T
BR-REP"
X
NE
NE
T
NE
M
T
NE
T
T
NE
NE
T
dose, is 5 mg-equiv.
dose, is 50
dose, is 500
IH-7
BR-6
SR-2 (first
SR-1 (third
mg-equiv.
mg-equiv
survey )
survey)
SRl-REPf
X
NE
T
T
NE
T
T
NE
T
T
NE
NE
T
sediment
sediment
. sediment
SR2-REP10
X
T
T
T
NE
NE
T
NE
NE
T
NE
NE
T
SR2-REP20
X
NE
NE
T
NE
NE
T
NE
NE
T
NE
NE
T
-------
Table 6.15
Results of Ames assay (n=3) for 4 strains, 3 doses of
extract, and 2 concentrations of S9 for replicate samples.
"T" = toxicity, "NE" = no-effect, and "M" = mutagenicity.
Dose S9
Strain (ma-eauiv. ( ma-
IH-REPd BR-REP6 SRl-REPf SR2-REP19 SR2-REP29
sediment) equiv. _ _ _ _
protein) X
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
TA98
TA98
TA98
TA98
TA98
TA98
TA100
TA100
TA100
TA100
TA100
TA100
TA102
TA102
TA102
TA102
TA102
TA102
a For
b For
0 For
10°
10°
100b
100b
1000C
1000C
10a
10a
100b
100b
1000C
1000°
10B
10a
100b
100b
1000C
1000C
10a
10a
100b
100b
1000C
1000C
sample IH-REP,
sample H-REP,
sample IH-REP,
d Blind duplicate of
e Blind duplicate of
f Blind duplicate of
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
dose
dose
dose
IH-7
BR-6
SR-2
M
NE
M
M
T
T
M
M
M
M
T
T
NE
NE
M
M
T
T
NE
NE
NE
NE
T
NE
is 5 mg-equiv.
is 50 mg-equiv.
is 500 mg-equiv
(first survey)
X
NE
NE
NE
NE
T
T
NE
NE
M
M
T
M
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
T
T
sediment
sediment
. sediment
X
NE
NE
M
NE
T
T
NE
NE
M
M
T
T
NE
NE
NE
M
T
T
NE
NE
NE
NE
T
T
X
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
M
M
NE
NE
NE
NE
NE
NE
NE
NE
NE
T
NE
T
X
NE
NE
NE
NE
NE
NE
NE
NE
NE
M
T
T
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
NE
9 Blind duplicate of SR-1 (third survey)
-------
Table 6.16 Summary of results of Quality Assurance Quality Control tests for Ames Assay.
Strain
97a
98
100
102
Solvent
Control1
173 (80-325)
32 (9-65)
170 (84-283)
357 (166-531)
Positive
Control1-2
521 (216-996)
1828 (323-2421)
846 (436-1394)
1669 (661-2559)
Positive
Control1-3
926 (545-1496)
1641
551
465
(955-2353)
(335-990)
(295-662)
Density
Estimate1
6.3 X 109 (0.
5.2 X 10* (0.
3.0 X 109 (0.
8.8 X 109 (0.
(
I 1
1-16)
3-9.2)
3-5.4)
3-16)
Crystal
Fiolet Ampicillin Tetracvcline
NG* G5 NG
NG G NG
NG G NG
NG G G
Completeness6
100%
100%
100%
100%
1 Mean for all AOCs and range of means for individual AOCs (in parentheses). Test acceptable at + 2 S.D.
z Positive control is direct-acting.
1 Positive control requires activation.
* NG = no growth
s G = growth
6 7 samples from Indiana Harbor plus rep and blank samples; 10 samples from Buffalo River plus rep and control sediment samples;
13 grab samples and 7 core samples from Saginaw River plus rep, blank and control sediment samples.
-------
Tablt 6.17 SuHMry of chemistry data for grab samples from the AOCs and the control sediment with literature
reported results of Ames assay with individual chemicals.
Indiana
Harbor
range
Pesticides (ug/kg)
Aldrin
(Alpha) Hexachlorocyclohexane
(Beta) Hexachlorocyclohexane
(c) Hexachlorocyclohexane
(Gaana) Chlordane
(Alpha) Chlordane
4,4 000
4,4. DDE
4,4, DDT
Dieldrin
Endo Sulfan I
Endo Sulfan II
Endo Sulfan sulfate
Endrin
Endrin Aldehyde
Heptachlor
Heptachlor expoxide
Lindane
Toxaphene
Hethoxychlor
Endrin ketone
PAHs
LOM Molecular Wt PAHs (ug/kg)
1,4 Dfchlorbenzene
Napthalene
2-Nethylnaphthalane
Dimethyl pthalate
Dibanzofuran
Fluorene
Phananthrene
Anthracene
Buffalo
River
(n-10)
range
N.D.
N.D.
N.D. -
N.D.
N.D.
N.D.
N.D. -
N.D.
N.D. -
N.D. -
N.D.
N.D. -
N.D.
N.D.
N.D. •
N.D.
N.D. -
N.D.
N.D.
N.D.
N.D.
150
75
100
120
80
60
30
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
- 330
- 290
- 170
- 70
- 210
- 100
- 343
- 41
- 160
- 44
- 65
- 320
- 320
<36 -810
range
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
Reported
Ames assay
Mutagen-
icity Notes
1
1.2
1,2
1.2
- 1
1
1,3
- 1
- 1
1
3
3
3
- 1
3
3
2
3
+ 1, (a)
- 1, (a)
3
Citation
NIOSH (1992)
USPHS (1989)
USPHS (1989)
USPHS (1989)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
930 - 26500
790 - 30500
3400 - 151500
1400 - 215000
<4 - 130
<6 - 55
<6 - 63
N.D. - 110
<7 - 38
<7 -69
N.D. - 1000
<8 - 70
13.5
N.D. - 3
N.D.
N.D.
High Molecular Wt PAHs (ug/kg)
Fluoranthene <55 -
Pyrene <68 -
Butyl Benzle Phthalate <180
Benzo (a) anthracene <21
Bis (2-Ethylhexyl) phthalate 880
Chrvsene <27
Di-n-octypthalate <84
Benzo-b-fluoranthene <30 -
Benzo-k-fluoranthene
Benzo(a)pyrene <27 -
Indeno 123 CD Pyrene <100
Benzo (ghi) perlyene <130
Dioxins (ng/kg)
TCDD
PECOO
HXCDD
HPCDD
OCDD
Furans (ng/kg)
TCDF
PECDF
HXCDF
HPCOF
OCDF
PCB (ug/kg)
PCB 1016
PCB 1221
PCB 1232
PCB 1242
PCB 1248
PCB 1254
PCB 1269
7500
6700
- 15000
3500
59000
4000
38000
7000
9500
5800
- 3800
- 3800
4800 -
5500 -
<240 -
4200 -
3800 -
5200 -
<240 -
5600 -
4200 -
5700 -
5300 -
6300 -
80000
46500
16000
32000
290000
33000
37000
26000
23000
31000
22000
31000
N.D. - 1200
N.D. - 1800
<18 - 3000
N.D. - 690
170 - 13000
N.D. - 600
<19 - 2200
<7 -600
<9 - 400
<6 -440
<10 - 220
<13 - 310
7 - 10
29
N.D.
4 - 5
8 - 9
N.D.
N.D.- 2
2
2
N.D. - 12
N.D. - 7.4
N.D. - 190
12 - 2000
53 - 12000
32 - 230
N.D. - 950
350 - 8700
980 - 22000
2300 - 46000
6 - 230
N.D. - 150
33 - 460
74 - 2100
340 - 26800
N.D - 1.5
N.D. - 83
N.D. - 110
3-640
<3.9 - 780
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
170 - 3700
76 - 1350
220 - 3600
380 - 8200
250 - 17300
230 - 22000
120 - 8700
200 - 2100
120 - 4000
89 - 2800
1000
N.D.
N.D.
N.D.
N.D.
3000
N.D.
N.D.
N.D.
- 43000
- 4450
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
N.D.
- 60000
- 7900
- 300
N.D.
N.D.
N.D.
Notes: (a), Florissant data fro* NFCRC historical records not Battelle
1, demonstrated autagenicity in tests other than Ames Assay.
2, data inconclusive
3, Information not available
(a)
,3
(b)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
+ 1
- 1 (a)
-
+ 1
+ 1
+ 1
+
+ 1
+
+ 1
+ 1
2
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
NIOSH (1992)
Schoeny (1982)
Schoeny (1982)
Schoeny (1982)
-------
Appendix 6.1 Mean number of revertants (n=3) + SD for 4 strains, 3 doses of extract,
and no S9 for 7 stations in Indiana Harbor. Suspected toxicity is
denoted by a "*", confirmed toxicity and mutagenicity is denoted by a "T"
and "M", respectively.
Strain
spont.
TA97a
TA97a
TA97a
spont.
TA98
TA98
TA98
spont.
TA100
TA100
TA100
spont.
TA102
TA102
TA102
Dose
(mg-egiaiv.
sediment )
revertants
5
50
500
revertants
5
50
500
revertants
5
50
500
revertants
5
50
500
3
X SD
135 7
140 25
135 3
242*132
17 1
29 8
15 7
215 172
124 19
130 12
116 19
386*263
228 29
231 23
210 21
249*256
4
X
135
143
4
0
17
26
1
0
124
133
1
0
228
266
0
0
SD
7
2
5
-
1
8
2
-
19
26
2
-
29
24
-
—
5
X
135
135
171*
0
17
23
295*
0
124
137
102
45
228
254
183
0
sn
7
4
58
-
1
6
59
-
19
17
12
38
29
6
9
~
X
135
141
328*
0
17
22
458*
1
124
98
6
SD
7
11
117
-
1
2
26
2
19
8
641*218
58*
228
268
276*
1
58
29
24
52
1
X
135
134
126
0
17
19
109
0
124
127
7
SD
7
20
25
-
1
3
3
—
19
9
496*157
369*
228
266
312*
0
49
29
8
346
"
8
X SD
135 7
164 6
430* 91
463* 19
311 -
61 -
0
0
124 19
104 15
95* 9
685*162
228 29
225 41
152* 35
1129*221
10
X
91
65
42*
8
32
11*
0
0
212
SD
16
10
4
4
11
4
-
—
16
153*131
72*
99*
394
286
226
671
7
36
18
14
22
42
only 2 replicates
-------
Appendix 6.1 Mean number of revertants (n=3) + SD for 4 strains, 3 doses of extract,
and no S9 for 7 stations in Indiana Harbor. Suspected toxicity is
denoted by a "*", confirmed toxicity and mutagenicity is denoted by a "T"
and "M", respectively.
Station
Strain
spont.
TA97a
TA97a
TA97a
spont.
TA98
TA98
TA98
spont .
TA100
TA100
TA100
spont .
TA102
TA102
TA102
Dose
(mg-equiv.
sediment )
revertants
5
50
500
revertants
5
50
500
revertants
5
50
500
revertants
5
50
500
3
X SD
135 7
140 25
135 3
242*132
17 1
29 8
15 7
215 172
124 19
130 12
116 19
386*263
228 29
231 23
210 21
249*256
X
135
143
4
0
17
26
1
0
124
133
1
0
228
266
0
0
4
SD
7
2
5
-
1
8
2
-
19
26
2
-
29
24
-
—
5
X
135
135
171*
0
17
23
295*
0
124
137
102
45
228
254
183
0
SD
7
4
58
-
1
6
59
-
19
17
12
38
29
6
9
—
6
X
135
141
SD
7
11
328*117
0
17
22
458*
1
124
98
-
1
2
26
2
19
8
641*218
58*
228
268
276*
1
58
29
24
52
1
X
135
134
126
0
17
19
109
0
124
127
496*
369*
228
266
312*
0
7
SD
7
20
25
-
1
3
3
-
19
9
157
49
29
8
346
8
X
135
164
430*
463*
311
61
0
0
124
104
95*
685*
228
225
152*
SD
7
6
91
19
_
-
-
-
19
15
9
162
29
41
35
- 1129*221
10
X
91
65
42*
8
32
11*
0
0
212
SD
16
10
4
4
11
4
-
—
16
153*131
72*
99*
394
286
226
671
7
36
18
14
22
42
only 2 replicates
-------
Appendix 6.2 Continued
Station
Strain
spont.
spont.
TA100
TA100
TA100
TA100
TA100
TA100
spont.
spont.
TA102
TA102
TA102
TA102
TA102
TA102
Dose
(g equiv.
sediment )
revertants
revertants
5
5
50
50
500
500
revertants
revertants
5
5
50
50
500
500
S-9
(mg-
equiv.
protein J
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
3
X
108
108
204
153
218
237
178
303
239
178
292
366
409
563
262
409
SO
3
36
21
8
39
19
24
39
12
13
11
26
31
14
195
34
X
108
108
149
144
139
216
115
265
239
178
346
305
404
516M
14
1565
4
SO
3
36
15
23
4
40
41
346
12
13
11
15
23
29
13
106
X
108
108
197
163
295
308
140
188
239
178
336
286
671M
794M
811
426
5
SD
3
36
13
11
18
44
15
17
12
13
60
8
40
95
184
99
6
X
108
108
180
203
249
295
191
256
239
178
505
639M
625
877
66
411
SO
3
36
14
24
19
13
22
18
12
13
68
26
85
72
71
41
7
X SO
108 3
108 36
142 4
150 10
140 12
222 7
637*252
981 170
239 12
178 13
406 14
442 20
282 16
390M 25
674*317
407*177
X
108
108
199
169
121
204
235
255
239
178
356
367
349
313
249
203
8
so
3
36
22
5
10
29
42
76
12
13
13
37
48
14
32
57
10
X
169
168
272
288
245
397
191
188
400
373
446
467
509*
635*
376*
SO
5
9
27
14
55
17
39
15
50
39
13
13
59
43
21
416*153
only 2 replicates
-------
Appendix 6.2 Mean number of revertants (n=3) + SD for 4 strains, 3 doses of extract, and 2
concentrations of S9 for 7 stations in Indiana Harbor. Suspected toxicity is
denoted by a "*". Confirmed toxicity and mutagenicity is denoted by a "T" and
"M", respectively.
Station
Strain
spont.
spont.
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
spont.
spont .
TA98
TA98
TA98
TA98
TA98
TA98
Dose
(mg-equiv.
sediment )
revertants
revertants
5
5
50
50
500
500
revertants
revertants
5
5
50
50
500
500
S-9
(mg-
equiv.
protein)
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
3
X
135
203
194
284
298
279
298
290
27
42
47
48
136
105M
37
93M
4
SD
6
9
28
38
15
7
24
10
7
8
4
13
16
8
6
20
X
135
203
185
195
218
284
76
472
27
42
35
41M
41
119M
11
331T
SD
6
9
22
9
6
20
22
171
7
8
10
4
5
14
11
56
5
X
135
203
196
234
252
351
127
336
27
42
28
29
57
SD
6
9
17
19
35
30
23
13
7
8
6
5
9
200M127
21
103M
2
2
6
X
135
203
238
285
354
377
317
393
27
42
55M
71
72
145
46
178
SD
6
9
39
18
7
45
9
64
7
8
3
21
23
23
2
79
7
X SD
135 6
203 9
181 12
204 13
221 18
269 20
803* 82
557*252
27 7
42 8
36 5
35 2
26 4
HIM 3
703*232
453T255
8
X SD
135 6
203 9
262 38
217 18
206 14
286 35
349 20
392 30
"I
311
701
961
571
1431
241
291
10
X
124
212
232
285
270
361
274
285
39
44
56
76
104M
181
110*
117*
SD
26
41
21
24
12
40
11
24
6
6
8
17
9
8
13
16
-------
Appendix 6.3 Mean number of revertants (n=3) + SD for 4 strains, 3 doses of extract, and no
S9 for 10 stations in Buffalo River. Toxicity determined by visual inspection or
replicated plating is denoted with a "T".
Station
Dose
(mg-equiv.
sediment )
spoxrt.
reverts . :
10
1000
1000
spont .
reverts . :
10
100
1000
spont .
reverts. :
10
100
1000
spont.
reverts. :
10
100
1000
1
X SD
107 23
128 16
344* 95
183* 48
23 5
20 4
108T 63
75* 38
265 17
158 9
503 165
129 25
340 18
392 20
323 33
141* 15
2
X
107
134
238*
0
23
29
132*
0
265
240
219
17
340
353
320
SD
23
5
48
-
5
8
111
-
17
11
92
18
18
18
5
201*258
3
X SD
107 23
133 7
266*158
2 4
23 5
30 1
169*210
0
265 17
253 13
658 71
17 29
340 18
374 24
281 33
10 10
4 5
X SD X
STRAIN:
107 23 107
89 12 98
ft
17 18 10
0-11
STRAIN
23 5 23
20 3 18
62T 87 0
SD
6
X
SD
7
X SD
8
X SD
9
X
SD
10
X SD
TA97a
23
23
9
19
107
133
174*
0
23
13
24
-
106 13
210 25
2 3
292*153
106 13
182 3
204 17
0
106
152
127
0
13
5
2
-
106 13
139 2
109* 46
0
: TA98
5
5
23
16
5
3
101*174
58*100 115*200
STRAIN:
265 17 265
90 4 168
0
-
29 2
29 3
1. 1
170*127
29 2
24 2
36 6
0
29
27
2
8
242T172
0
-
29 2
23 4
207*355
95* 40
TA100
17
25
131*202 276*264
288* 90 261
STRAIN:
340 18 340
369 27 333
449*103 150
88*152 10
1
265
95
259
0
17
7
152
-
138 13
179 27
25 30
62* 36
138 13
138 10
85 19
1 1
138
130
158*
0
13
6
87
-
138 13
142 8
190*277
55* 96
TA102
18
12
34
11
340
283
153
40*
18
7
24
35
361 47
426 15
294*168
1 1
361 47
402 8
393 13
295*480
361
430
379
0
47
19
32
-
361 47
387 22
289 58
117*107
-------
ppendix 6.4 Mean nunber of revertant. (n-3) + SD for 4 .train., 3 doee. of extract, and 2 concentration, of S9 for 10
•tation. in Buffalo River. Saapected toxiclty i. denoted by a "*". Confirmed toxicity and mutagenicity ie
denoted by a "T* and "M-, respectively.
Do.e s9
Strain («g-equiv. (cog- i
•ediment) equiv.
Protein 1 1 SD
•pont
•pont
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
•pont.
•pont.
TA98
TA98
TA98
tA98
TA98
TA98
•pont.
•pont.
TA100
TA100
TA100
TA100
TA100
TA100
• r evert i t i
- revert* t 3
10 i
10 3
100 l
100 3
1000 1
1000 3
revert* i 1
revert «i 3
10 i
10 3
100 i
100 3
1000 1
1000 3
reverts i i
revert* i 3
10 1
10 3
100 1
100 3
1000 1
1000 3
1(4 18
182 10
182 5
182 15
297 33
306 16
547 77
425 131
28 3
48 3
43 8
52 6
184 18
157M 19
131H 9
194M 39
217 48
224 28
282 18
262 29
407 14
383 34
280T 6
460M 54
X SD
164 11
183 10
184 8
197 6
312 10
437 19
686*118
274 21
28 3
48 3
48 11
45 7
119M 6
167 14
847*162
326* 22
217 48
224 28
277 20
257 21
451M 71
S39M 16
490T270
414*296
-J-
164 18
182 10
174 13
196 15
259 5
273 10
720* 57
226 27
28 3
48 3
39 3
39 4
148M 9
141 7
453*313
243M 18
217 48
224 28
248 7
260 29
386 44
390 26
847T128
224* 19
-^
1(4 18
182 10
212 6
226 8
328 12
404 36
19* 27
523*452
28 3
48 3
S3 6
45 16
142M 17
216M 12
599*373
776*420
217 48
224 28
263 6
237 7
364 20
449M 39
260* 42
297 168
c
•
164
182
210
173
277
361
132
138
28
48
70
5l'
89
170M
26
77*
217
224
276
251
367
514M
100
117
18
10
10
21
11
44
9
10
3
3
9
3
5
3
64
48
28
19
6
64
13
19
14
-t-
164 18
182 10
213 16
211 15
330 12
415 17
337*151
129* 34
28 3
48 3
56M 6
40 8
215H 10
226 31
609* 194
457T 90
217 48
224 28
270 18
266 22
397 46
460H 7
786* 21
125 39
-*-
143 6
180 10
229 33
236 33
3U 43
344 5
146* 15
887*183
i9 8
33 4
73 12
67 6
136 16
130M 16
42* 58
600* 154
139 13
139 20
182 9
170 12
233 23
213 28
575*144
1067T224
_fi
143
180
206
236
336
314
201*
799
29
33
27
60
137
135M
6
10
14
41
31
29
48
22
8
4
12
34
19
15
630*410
341M
139
139
155
150
232
213
129
450M
31
13
20
7
11
17
7
26
82
-i-
143 6
180 10
190 10
244 22
365 5
289 7
453*281
201* 38
29 8
33 4
27 6
230*256
163 14
138M 9
710* 55
507*339
139 13
139 20
148 4
167 13
264 26
273M 19
302*417
828*465
X SD
143 6
180 10
148 13
154 14
238 82
217 10
434* 85
777*570
29 8
33 4
22 5
23 10
63 13
9SM 36
351 123
500*688
139 13
139 20
202 26
152 15
202 26
188 9
480T420
509*205
-------
Appendix 6.4 Continued
Strain
Doae
(ng-equiv
S9
_J
•
2
_1
c
T
t |r
_£
__2
_fi
__2
i
sediment) equiv. ________
•pont.
•pont.
TA102
TA102
TA102
TA102
TA102
TA102
revert* i
revert* i
10
10
100
100
1000
1000
orotein)
1
3
1
3
1
3
1
3
X
394
299
469
460
492
474
601
539
SD
23
35
15
30
62
18
36
72
X
394
299
404
439
506
569
477
337
SD
23
35
18
14
24
45
192
31
X
394
299
436
423
534
SUM
435
364
SD
23
35
32
22
9
7
54
46
X SD
394 23
299 35
418 26
430 28
509 26
591M 33
255* 39
124*102
X
394
299
398
365
522
472
241
137
SD
23
35
64
62
14
27
45
20
X
394
299
347
353
489
422M
891*
5
sn
23
35
92
25
24
38
58
5
X
426
468
475
513
537
609
91
SD
21
14
72
31
33
51
17
903*597
X
426
4C8
401
356
522
471
378*
642
SD
21
14
94
58
6
63
57
59
X
42<
468
442
492
540
550
SD
21
14
31
17
15
38
842*658
584*270
X SD
42C 21
4C8 14
396 26
530 61
528 34
536 44
14 10
604*390
only 2 replicates
-------
Appendix 6.5 Mean number of revertants (n=3) + SD for 4 strains, 3 doses of extract,
and no S9 for 7 stations from first survey of Saginaw River. Suspected
toxicity is denoted by a "*". Confirmed toxicity and mutagenicity are
denoted by a "T" and "M", respectively.
Station
Strain
spent.
TA97a
TA97a
TA97a
spent.
TA98
TA98
TA98
spont.
TA100
TA100
TA100
spont.
TA102
TA102
TA102
Dose
(mg-equiv.
sediment )
revertants
10
100
1000
revertants
10
100
1000
revertants
10
100
1000
revertants
10
100
1000
2
X
136
120
112
0
19
39
118*
0
139
146
81*
0
407
456
379
0
3
SD
15
21
6
-
3
7
29
-
12
13
5
-
9
10
6
-
X
136
154
121*
9
19
29
29
96*
139
143
SD
15
7
97
10
3
1
34
83
12
10
382*159
0
407
403
256
0
-
9
8
4
-
4
X
136
135
140
0
19
23
57T
0
139
133
81
0
407
396
417
0
SD
15
12
4
-
3
4
8
-
12
12
7
-
9
9
16
-
X
136
141
4
0
19
24
0
0
139
123
122
0
407
443
226
0
6
SD
15
32
7
-
3
2
-
-
12
11
74
-
9
29
7
-
7
X
136
132
101
0
19
29
51*
0
139
141
85*
0
407
460
365
0
SD
15
3
9
-
3
7
7
-
12
13
4
-
9
20
26
-
9
X SD
216 48
203 9
152 6
0
34 1
28 4
20 6
117*113
165 41
145 7
198* 72
0
450 23
375 15
401 48
237*217
10
X
136
114
101
0
19
26
364*
0
139
117
105*
0
407
370
358
15*
SD
15
10
8
—
3
4
54
-
12
7
40
—
9
10
17
26
-------
Appendix 6.6 Mean number of revertants (n=3) ± SD for 4 strains, 3 doses of extract,
and no S9 for 7 stations from the third survey (grab) of Saginaw River.
Suspected toxicity is denoted by a "*". Confirmed toxicity and
mutagenicity is denoted by a "T" and "M", respectively.
Station
Strain
spont .
TA97a
TA97a
TA97a
spont.
TA98
TA98
TA98
spont.
TA100
TA100
TA100
spont.
TA102
TA102
TA102
Dose
(mg-equiv.
sediment)
revertants
10
100
1000
revertants
10
100
1000
revertants
10
100
1000
revertants
10
100
1000
X
225
150
29
1
60
41
2
0
206
229
1
0
215
241
0
0
1
SD
82
15
30
1
7
16
3
-
18
13
1
-
10
70
—
-
2
X
225
127
69
0
60
41
27
68
206
239
209
664
215
176
182
0
SD
82
28
11
-
7
21
25
23
18
18
29
347
10
6
79
-
X
156
159
130
0
22
35
5
SD
30
12
13
6
X
216
162
222
SD
48
5
15
392*226
6
3
283*344
0
167
171
115
1
336
418
371
0
—
4
11
7
1
57
22
36
-
34
22
28
943T
165
124
167
535*
450
433
390
345
1
7
2
60
41
11
18
5
23
19
21
41
8
X
216
77
85
3
34
17
15
322*
165
116
78
166*
450
388
352
125*
SD
48
16
2
5
1
4
3
103
41
16
5
41
23
34
13
23
16
X
216
170
107*
27
34
28
SD
48
11
2
31
1
7
205*173
0
165
157
307
0
450
416
446
0
~
41
16
38
~
23
20
59
24
X
156
90
62
0
22
23
152*
0
167
185
154
746
336
178
175
SD
30
8
21
~
6
8
49
~
4
33
24
149
57
11
34
342*104
-------
Appendix 6.7 Mean number of revertants (n=3) + SD for 4 strains, 3 doses of extract, and no S9 for
8 core samples from the third survey of Saginaw River. Suspected toxicity is denoted
by a "*". Confirmed toxicity and mutagenicity is denoted by a "T" and "M",
respectively.
Strain
spont.
TA97a
TA97a
TA97a
spont.
TA98
TA98
TA98
spont.
TA100
TA100
TA100
spont.
TA102
TA102
TA102
i
Dose
(mg-equiv.
sediment )
reverrants
10
100
1000
reverrants
10
100
1000
reverrants
10
100
1000
revertants
10
100
1000
_5
X
156
221
156
0
22
42
106*
1
167
168
131
.1
1
SD
30
21
13
-
6
13
14
2
4
10
5
224*148
336
384
395
324
57
35
44
371
5
X
156
106
2
SD
30
18
675*267
0
22
41
95
0
167
176
926*
0
336
194
130
0
-
6
9
66
-
4
19
335
-
57
51
21
—
X
225
168
3
0
60
59
6
0
206
192
52
0
215
271
172
0
61
SD
82
18
5
-
7
10
4
-
18
21
79
-
10
12
158
—
6
X
156
94
89
0
22
26
31
149*
167
203
161
0
336
198
174
22
2
SD
30
17
14
-
6
2
14
74
4
8
16
-
57
19
25
38
_2
X
216
146
79
0
34
25
187*
0
165
197
282*
2
450
285
408
561*
3
SD
48
44
13
-
1
3
49
-
41
14
66
3
23
52
18
78
24
X SD
156 30
181 10
195 21
335*353
22 6
21 4
27 7
484*443
167 4
137 4
123 14
557M 53
336 57
346 74
435 14
1142* 7
X
225
167
3
0
60
31
0
0
206
215
4
0
215
242
141
0
65
SD
82
85
1
-
7
4
_
-
12
50
4
-
10
33
19
-
6°
X SD
156 30
108 32
494*342
0
22 6
23 3
1043*721
0
167 4
195 6
1115T127
360*124
336 57
164 9
80 11
0
core from 0-2 feet deep
core from 2-4 feet deep
core from 8-20 inches deep
core from 20-35 inches deep
core from 5-13 inches deep
core from 13 - 32 inches deep
-------
Appendix 6.8 Mean number of revertants (n=3) ± SD for 4 strains, 3 doses of extract,
and 2 concentrations of S9 for 7 stations from the first survey of Saginaw
River. Suspected toxicity is denoted by a "*". Confirmed toxicity and
mutagenicity are denoted by a "T" and "M", respectively.
Station
Dose
(mg-equiv. (
sediment) e
p
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
S9
2
:quiv. _
>rotein) X
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
161
240
203
248
293
278
SD
12
4
6
12
10
14
592*192
199*
26
28
41
55
80
86M
24*
70*
19
4
4
7
7
9
1
3
6
3
X SD
161 12
240 4
194 5
239 9
256 11
259 26
79*113
482*313
26 4
28 4
43 8
47 4
65M 7
68 10
231T231
73T 11
X
STRAIN
161
240
197
255
263
259
234*
319
STRAIN
26
28
45
40
50
76M
23
83M
4
SD
X
6
SD
7
X SD
X
9
SD
10
X SD
TA97a
12
4
5
10
5
18
95
8
TA98
4
4
5
5
3
2
6
17
161
240
184
213
294
347
332*
271*
26
28
39
46
84
123
300*
12
4
8
18
12
21
103
432
4
4
3
11
13
10
67
614*532
161 12
240 4
230 8
261 12
268 20
272 8
301* 81
471*394
26 4
28 4
36 5
51 6
71 7
76M 11
567T 99
392T466
183
238
210
181
293
284
198*
364
24
17
28
27
54M
44
34*
73M
8
68
37
45
7
22
58
15
1
8
10
7
7
8
3
14
161 12
240 4
147 7
208 12
167 16
175 30
223*180
498*320
26 4
28 4
32 8
39 5
65M 6
70 6
19* 8
621T502
-------
Appendix 6.8 Continued
Dose S9
(rag equiv. mg-
sediment) equiv,
prote:
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
2
•
Ln) X
152
158
188
191
252
234
SD
16
3
14
14
21
17
552T442
130* 45
469
501
438
485
563
533
166
467
42
28
36
57
16
29
93
62
X
152
158
179
183
242
273
3
4
SD X
16
3
19
10
15
20
208*170
364*373
469
501
480
481
448
467
42
28
25
18
92
35
6 6
555*274
STRAIN
152
158
182
177
273
243
100
253
STRAIN
469
501
507
484
525
498
1023T
520
SD
X
TA100
16 152
3 158
10
4
15
14
14
13
186
173
278
367
6
SD
16
3
7
15
12
16
350* 39
665*470
TA102
42 469
28 501
23
8
15
75
91
18
502
522
561
611M
378*
409*
42
28
17
26
32
17
147
124
7
X SD
152 16
158 3
178 25
164 14
214 18
193 16
412 102
711T126
469 42
501 28
482 9
546 15
542 19
547 29
101 156
588* 66
x
140
178
149
152
194
173
129
233
388
430
405
466
439
381
9
SD
8
55
15
13
8
20
34
41
16
66
27
50
11
38
299 20
262T402
10
X SD
152 16
158 3
140 5
149 7
221 4
220 15
436*232
725* 62
469 42
501 28
404 35
420 59
457 17
433 28
1186T774
1459*982
-------
Appendix 6.9 Mean number of revertants (n=3) + SD for 4 strains, 3 doses of extract,
and 2 concentrations of S9 for 7 stations from third survey (grab) of Saginaw
River. Suspected toxicity is denoted by a "*". Confirmed toxicity and
mutagenicity is denoted by a "T" and "M", respectively.
Station
Dose S9
(mg-equiv. (mg-
sediment) equiv.
orotei
1
In) X
SD
X
2
SD
5
X SD
6
X SD
X
8
SD
16
X SD
24
X
SD
STRAIN TA97a
spont. reverts.:
spent, reverts.:
10
10
100
100
1000
1000
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
238
271
222
192
204*
183
0
0
55
54
49
54
266*
159
0
0
61
19
39
19
50
43
_
-
7
5
11
12
93
82
-
238
271
102
155
274
327
6
225
55
54
47
54
73
51
116
207
61
19
13
11
51
38
2
330
7
5
9
13
1
17
24
23
165 27
133 25
198 13
224 28
241 25
265 11
18 11
878*321
STRAIN TA98
26 10
22 8
45 3
46M 8
56 10
54 11
328*545
865*526
183 8
251 68
155 34
189 71
236 20
256 17
482 10
361 8
24 7
17 8
27 4
19 2
42M 11
31 4
123M 21
143M 23
183
251
132
169
214
200
116
326
24
17
20
28
47M
42
53*
119M
8
68
18
10
24
2
8
23
4
8
1
8
3
4
30
9
183 8
251 68
212 19
219 29
287 35
235 43
777*413
289 19
24 7
17 5
30 6
46 17
64M 12
55 22
395*354
140M 37
165
133
148
144
273
216
27
25
3
24
5
41
367*217
228
26
22
30
11
86M
36
77
10
4
7
8
14
20
460*229
139M 23
-------
Appendix 6.9 Continued
Dose S9
(mg-equiv. (ing-
sediment) equiv.
proteii
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
1
1) X SD
207 12
236 35
253 27
214 15
331* 20
206* 32
1 1
2 2
X
207
236
226
244
106
108
498
721
2
sr>
12
35
63
11
166
186
194
13
x
5
SD
X
STRAIN TA100
163
155
162
175
220
213
85
32
13
6
6
4
9
119
1010*139
140
178
160
147
171
156
259
288
6
SD
8
55
1
13
14
3
26
52
X
140
178
138
172
151
149
95
229
8
SD
8
55
12
23
17
7
12
26
16
X
140
178
162
151
235
245
124*
178*
SD
8
55
9
13
5
6
34
60
24
X
163
155
178
167
298
267
174
349M
SD
32
13
21
26
2
15
2
81
STRAIN TA102
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
334 28
349 27
327 12
288 12
767* 98
536* 64
503T868
776 796
334
349
271
262
228M
233
434
79
28
27
36
48
33
98
9
66
287
310
508
561
495
575
143
26
59
47
52
40
54
212
1167*668
388
430
412
322
328
326
434
511
16
66
33
23
89
41
59
35
388
430
412
499
427
456
305
658M
16
66
10
28
11
62
13
11
388
430
420
552
467
462
16
66
8
57
17
31
546*109
378
87
287
310
222
257
307
401
258
26
59
41
33
38
67
36
621M183
-------
Appendix 6.10 Mean number of revertants (n=3) ± SD for 4 strains, 3 doses of extract, and 2
concentrations of S9 for 8 core samples from the third survey of Saginaw River.
Suspected toxicity is denoted by a "*". Confirmed toxicity and mutagenicity is denoted
by a "T" and "M", respectively.
Dose S9
(mg-equiv. (mg-
51
sediment) equiv. _
crotein) X SD
spent, reverts.:
spont. reverts.:
10
10
100
100
1000
1000
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
165 27
133 25
253 34
225 22
283 49
351 83
481*205
248 71
26 7
22 5
62 12
51M 7
76 17
106 47
209* 88
114M 5
52
X SD
165 27
133 25
139 15
164 30
188 7
180 74
916*498
528 251
26 7
22 5
34 4
33 15
109 11
44M 29
1731*425
1680*1285
61
X SD
STRAIN
238 61
271 19
261 43
273 58
218 35
378 35
149* 61
2
STRAIN
55 10
54 8
55 22
73 6
132 12
123 7
0
327T180
62
X
TA97a
165
133
83
30
209
178
242
2293
TA98
26
22
27
32
74M
44M
SD
27
25
43
30
35
24
140
872
7
8
2
11
13
16
813*1360
1564*1182
23
X
183
238
201
194
238
289
285
344
24
17
30
25
56M
59M
34
227M
sr>
8
68
9
14
20
23
28
58
4
8
2
2
8
8
8
21
2*
X
165
133
187
205
213
187
332
362
26
22
36
37
40
34
52
114M
sn
27
25
31
38
15
44
43
39
7
5
8
2
3
6
13
6
x
238
271
176
173
213
291
96*
1
55
54
80
53
104
75
1
6s
66
SD X SD
61
19
53
19
30
20
4
1
10
4
9
8
9
52
1
368*294
165 27
133 25
147 19
182 35
228 17
288 40
1321*269
2556*320
26 7
22 5
47 10
39 2
132 37
114M 26
1618*924
2383*241
-------
Appendix 6.10 Continued
Dose S9
(mg-equiv. (mg-
sediment) equiv.
orotei
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
51
Ln) X SD
163 32
155 13
196 5
186 7
302 20
329M 49
183 13
318 35
52
X SD
163 32
155 13
245 60
231 44
312M 21
248 48
1432*311
1931*405
6
X
1
SD
STRAIN
207
236
246
250
241
338
1
0
12
35
38
10
22
23
2
1
STRAIN
spont. reverts.:
spont. reverts.:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
287 26
310 59
390 94
487 44
478 86
441 96
670*236
512 201
287 26
310 59
216 9
156 21
343 107
121 68
1248*833
385* 44
334
349
402
407
296
381
6
28
27
29
46
10
91
8
232*260
Station
62
X SD
TA100
163 32
155 13
213 32
204 57
316 43 '
275M 37
1129*232
2085T158
TA102
287 26
310 59
262 17
269 50
287 14
361 82
234 30
1895*1003
23
X
140
178
194
126
220
227
146
314
388
430
418
412
560
563
367
SD
8
55
38
8
23
17
15
60
16
66
19
100
50
18
81
676M172
2
X
163
155
142
150
191
194
247
341M
287
310
471
574
539
117
530
728M
4
SD
32
13
11
21
6
34
71
39
26
59
36
24
77
56
20
31
65
X SD
207 12
236 35
227 36
213 4
227 30
435 12
904*124
16 9
334 28
349 27
319 24
308 38
249 13
280 38
541*584
562T254
66
X SD
163 32
155 13
223 33
167 12
230 14
380M 59
604*514
423* 65
287 26
310 59
260 9
328 22
279 19
328 14
1457*185
293 273
core from 0 -
core from 2 -
core from 8 -
2 feet deep
4 feet deep
20 inches deep
4 core from 20-35 inches deep
5 core from 5-13 inches deep
6 core from 13-32 inches deep
-------
Appendix 6.11.
Mean number of revertants (n=3) ± SD for 4 strains, 3 doses of extract, and no S9 for
7 Quality Assurance Quality Control samples. Suspected toxicity is denoted by a "*".
Confirmed toxicity and mutagenicity is denoted by a "T" and "M", respectively.
Station
Strain
spont.
TA97a
TA97a
TA97a
spont.
TA98
TA98
TA98
spont.
TA100
TA100
TA100
spont.
TA102
TA102
TA102
Dose
(mg-equiv.
sediment)
revertants
10a
100b
1000C
revertants
10
100
1000
revertants
10
100
1000
revertants
10
100
1000
IH-REPd
X SD
91 16
104 18
112 36
0
32 11
19 6
0
0
212 16
84* 27
90* 21
28* 22
394 18
296 11
172 22
0
BR-REP6 SRl-REPf SR2-REP19 SR2-REP29
X
106
140
75
0
29
24
82
0
138
143
sp
13
13
11
-
2
2
17
-
13
19
360*217
0
361
360
408
0
-
47
10
3
—
X SD
136 15
141 11
185*147
157*273
19 3
17 4
580*324
0
139 12
143 23
371*262
0
407 9
403 21
218 16
0
X
225
101
60
71
60
57
59
175T
206
236
164*
21
215
147
148
326
RD
82
13
7
24
7
9
9
92
18
39
17
18
10
24
52
300
X SD
216 48
204 10
191 16
640*130
34 1
30 7
23 4
453*354
165 41
136 9
115 7
313* 55
450 23
333 27
375 13
487*210
BR-CON
X
106
115
89*
0
29
26
8
0
138
132
RD
13
7
23
—
6
6
2
—
4
18
365*498
0
361
379
180*
0
1
57
15
10
"
SR-CON
X SD
216 48
93 14
102 1
14 1
34 7
14 5
32 1
155*233
165 41
146 3
161 10
24 42
450 23
367 50
383 21
136*162
IH-BLh SR-BLh
X
91
122
150
546*
32
27
31
SD X
16 225
21 117
8 99
47 102
11 60
8 40
2 56
248*250 236*
212
205
186
36
394
288
334
16 206
16 219
25 204
27 178
18 215
14 219
19 217
625*225 212
SD
82
24
17
11
7
15
8
87
18
18
14
12
10
20
30
15
For sample IH-REP, dose is 5 mg-equiv. sediment
b For sample IH-REP, dose is 50 mg-equiv. sediment
c For sample IH-REP, dose is 500 mg-equiv. sediment
d Blind duplicate of IH-7
e Blind duplicate of BR-6
f Blind duplicate of SR-2 (first survey)
9 Blind duplicate of SR-1 (third survey)
h Procedural blanks do not contain sediment
-------
Appendix 6.12 Mean number of revertants (n=3)+ SD for 4 strains, 3 doses of extract, and 2 concentrations
of SS for 7 Quality Assurance Quality Control samples. Suspected toxicity is denoted by a
"*". Confirmed toxicity and mutagenicity is denoted by a "T" and "M", respectively.
Strain
spont .
spont .
TA97a
TA97a
TA97a
TA97a
TA97a
TA97a
spont .
spont.
TA98
TA98
TA98
TA98
TA98
TA98
Dose S9
(mg-equiv. (mg-
sediment) equiv
Drotei
reverts:
reverts:
10"
10"
ioob
ioob
1000°
1000°
reverts:
reverts:
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
IH-REPd
,n) X
124
212
334
338
390
426
169*
172*
39
44
109
131
125
245
111*
128*
SD
26
41
14
36
3
19
61
24
6
6
4
2
12
24
24
30
BR-REP"
X
143
180
162
202
232
235
204*
293
29
33
36
31
66
75M
12*
130M
SD
18
10
18
2
5
4
54
20
3
3
10
1
8
11
3
38
SRl-REPf
X
161
240
246
241
382
446
63*
3
26
28
43
51
148
179
39*
SD
12
4
7
4
12
54
68
6
4
4
3
6
4
10
32
184*304
SR2-REP18
X
238
271
149
190
212
278
262
352
55
54
70
66
79
64
125
114M
SD
61
19
9
50
17
27
13
39
10
4
15
14
10
23
26
14
SR2-REP2g BR-CON
X
183
251
203
227
217
188
334
415
24
17
24
26
32
39
68
SD X SD
8 143 18
68 180 10
20 166 6
31 213 20
23 121 5
46 160 19
12 349*253
64 721*567
4 29 3
8 33 3
2 37 9
10 38 7
3 28 6
19 37 2
12 1039*152
88T 41 105* 76
SR-CON
X
183
251
146
184
167
276
144
186
24
17
31
35
67M
74
75*
SD
8
68
10
22
9
47
30
1
4
8
7
5
7
9
34
225*473
IH-
X
124
212
201
222
195
191
124
182
39
44
34
40
40
39
24
64
h
BL
SD
26
41
6
8
5
2
8
15
6
6
4
7
1
6
9
5
SR-BLh
X
238
271
191
207
161
174
255
186
55
54
72
51
66
61
68
67
SD
61
19
104
29
46
22
7
28
10
4
6
4
13
12
23
9
-------
Appendix 6.12 Continued
Strain
spont.
spent.
TA100
TA100
TA100
TA100
TA100
TA100
spont.
spont.
TA102
TA102
TA102
TA102
TA102
TA102
Dose S9
(mg-equiv. (mg-
sediment) equiv
protei
reverts:
reverts :
10
10
100
100
1000
1000
reverts :
reverts :
10
10
100
100
1000
1000
1
3
1
3
1
3
1
3
1
3
1
3
1
3
1
3
IH-REPd
n) X
169
168
333
305
356
519
143
160*
400
373
457
498
434
671
160
198
sn
5
9
22
47
18
60
30
14
50
39
30
30
24
71
24
27
BR-REP"
X
139
139
144
145
217
181
82*
202
426
468
388
436
454
481
310*
351*
sn
48
28
28
15
23
16
3
31
23
35
34
44
49
69
69
65
SRl-REPf
X
152
158
165
188
251
344
SD
16
3
6
8
28
49
214*110
72*
469
501
527
498
557
606
146*
7
64
42
28
9
24
40
23
20
7
SR2-REP10
X
207
236
217
202
242
217
317
353
334
349
219
185
246
146
178
227
SD
12
35
29
45
37
55
105
101
28
27
43
68
21
71
78
197
SR2-REP2
X
140
178
200
168
163
153
176
261
388
430
354
422
368
419
377
592
SD
8
55
41
19
6
11
17
17
16
66
60
101
23
102
20
164
0 BR-CON
X SD
139 48
139 28
152 0
149 13
86 17
101 23
221 181
418T372
426 23
468 35
504 21
458 33
255 8
262 32
1214*634
1382T991
SR-CON IH-BLh
X
140
178
163
153
262
229
SD X SD
8 169
55 168
10 167
14 159
18 194
25 161
322*191 55
168*
388
430
363
358*
391
394
456*
17 97
16 400
66 373
74 324
85 236
52 355
69 222
44 220
879T364 206
5
9
30
8
21
5
14
22
50
39
25
40
18
123
16
24
SR-BLh
X
207
236
237
201
259
263
321
321
334
349
349
312
361
264
363
232
SD
12
35
13
21
27
28
50
36
28
27
23
20
27
30
40
21
* For sample IH-REP, dose is 5 mg-equiv. sediment ° Blind duplicate of SR-1 (third survey)
b For sample IH-REP, dose is 50 mg-equiv. sediment Procedural blanks do not contain sediment
0 For sample IH-REP, dose is 500 mg-equiv. sediment
d Blind duplicate of IH-7
* Blind duplicate of BR-5
f
Blind duplicate of SR-2 (first survey)
-------
Appendix 6.13 Mean number of revertants (n=3) + SD for known direct-acting and promutagens for each
test run.
Site
S9
Postive (mg-
Mutagen ~ equiv
orote
spont. reverts.:
spont. reverts.:
TNF
2AF
0
1
0
1
IH-1
in)X
135
203
535
833
SD
7
9
91
36
IH-2
x
91
212
664
649
SD
16
41
162
147
BR-1
x
SD
STRAIN
107
182
462
717
23
10
32
30
STRAIN
spont. reverts.:
spont. reverts.:
TNF
2AF
0
1
0
1
17
42
1022
1487
1
8
72
165
32
44
1092
1647
11
6
64
31
23
48
1022
1599
5
3
77
24
STRAIN
spont. reverts.:
spont. reverts.:
MMS
BAP
0
1
0
1
124
108
606
177
19
36
13
5
212
168
477
538
16
9
39
14
265
224
474
477
17
28
27
11
STRAIN
spont. reverts.:
spont. reverts.:
MMS
2AF
0
1
0
1
228
178
1007
302
29
13
73
12
394
373
1010
516
18
39
50
18
340
299
695
473
18
35
BR-2
x
SD
SR1-1
Y
TA97a
106
180
345
857
TA98
29
33
786
1714
TA100
138
139
996
370
TA102
361
468
39 2197
34
430
13
10
5
17
8
4
29
123
13
20
68
32
47
14
87
35
136
240
577
918
19
28
1008
1877
139
158
1050
401
407
501
2393
650
SD
15
4
72
18
3
4
60
87
12
3
61
27
9
28
144
12
SR2-1
x
216
251
971
1393
34
17
2283
2211
165
178
747
629
450
430
2104
527
SD
48
68
36
128
1
8
195
201
41
55
25
62
23
66
59
83
SR2-2
x
156
133
247
1020
22
22
702
1643
167
155
1004
558
336
310
1702
460
SD
30
25
34
80
6
5
176
171
4
13
20
43
57
59
328
100
SR2-3
X SD
225 82
271 19
426 169
1177 278
60 7
54 4
1333 283
1745 188
206 18
236 35
1379 13
914 73
215 10
349 27
1755 280
381 56
-------
CHAPTER 7: MUTATOX GENOTOXICITY ASSAY
Johnson, B.T.
INTRODUCTION
The Great Lakes of North America form the largest continuous mass of
fresh water on earth. Lakes Michigan, Superior, Huron, Ontario, and Erie
contain over 20 percent of the world's fresh water (Wetzel 1975). During the
last two centuries, many cities with their supportive industries spread along
their shorelines and tapped the natural wealth of the Great Lakes Basin
(Wheeler and Kostbade 1990). The Lakes have become a conduit of world trade,
a supporter of large fisheries, a dependable source of potable water, a
playground for aquatic sports and recreational fisheries, and a repository for
domestic and industrial anthropogenic wastes.
The presence of chemical contaminants in the Great Lakes is well-
documented. A comprehensive review by Hesselberg and Seelye (1982) in
conjunction with the International Joint Commission's (IJC) Great Lakes
International Surveillance Plan tentatively identified 476 organic compounds
in fish—tissue residues. Of these compounds, 53 were halogenated, 29 were
isomers of PCBs, nine were chlorinated pesticides, five were polycyclic
aromatic hydrocarbons, over 150 were oxygen—containing compounds such as
phenols, esters, and carboxylic acids, and the remaining were aliphatic
hydrocarbons. A study of sediment extracts with Medaka (Oryzias latipes) from
tributaries of the Great Lakes (Fabacher et al. 1991) suggested that chemical
contaminants can cause neoplasms in wild fishes. In 1980, the IJC estimated
that about 2,500 chemicals were in common use in the Great Lakes Basin
ecosystem (IJC 1980). Recently, a joint U.S.-Canadian committee investigating
water quality concluded that humans inhabiting the Great Lakes Basin were
exposed to and accumulated more toxic materials (including xenobiotics and
inorganic pollutants) than anyone else on this continent (NRC 1985).
In 1987 the U.S. Congress under amendments to the Clean Water Act, in
Section 118(c)(3), authorized the Great Lakes National Program Office (GLNPO)
of the U.S. Environmental Protection Agency to implement a five-year study of
chemical pollutants in Great Lake sediments. Data from this study will assist
the Assessment and Remediation of Contaminated Sediment (ARCS) Project develop
Remedial Action and Lake-wide Management Plans for removal of toxic materials
from sediment. The present study, as part of this extensive sediment
investigation, was designed to determine the potential genotoxicity (DNA-
dainage) of chemical contaminants in sediments from three Areas of Concern
(AOC): Indiana Harbor in IN, Buffalo River in NY, and Saginaw River in MI with
two procaryotic bioassays the Ames Salmonella Microsome Mutagenicity Test
(Maron and Ames 1973) (Chapter 6) and the Mutatox Genotoxicity Assay (Johnson
1992). The Ames and the Mutatox tests both use the induced reversion of
bacterial mutants to detect DNA-damaging substances. Unlike the Ames test,
the transformation in the Mutatox assay may not always be mutagenic;
therefore, the generic term genotoxicity is used to describe this assay.
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MATERIALS AND METHODS
EXPERIMENTAL DESIGN
The objectives of this study were twofold: to validate the relative
sensitivity and specificity of the activated Mutatox Genotoxicity Assay
(Microbics Corp., Carlsbad, CA) using known progenotoxins and non-genotoxins
in complex mixtures; and to use this assay to determine the potential geno—
toxicity of industrially contaminated sediments from selected Great Lakes
Areas of Concern.
SAFETY PROCEDURES FOR GENOTOXICITY EXPERIMENTS
Disposable gloves, TyvekR suits, sleeve protectors, and safety glasses
were used for handling potentially genotoxic substances. All toxicological
transfers were conducted in a laminar flow hood under yellow light; treated
samples were incubated in an enclosed vented hood. Toxicant disposal was
pursuant to National Fisheries Contaminant Research Center (NFCRC) Safety Plan
1991.
CHEMICALS: PROGENOTOXINS, NON-GENOTOXINS, AND SOLVENTS
Progenotoxins 2-acetamidofluorene (2-AAF), 2-aminoanthracene (2-AA), and
pyrene (PY) were purchased from Sigma Chemical Co. (St. Louis, MO), and 2-
aminofluorene (2-AF), 2-aminonaphthalene (2-AN), benzo(a)pyrene (BaP) , and 3-
methylcholanthrene (3-MC) were obtained from Aldrich Chemical Co. (Milwaukee,
WI). (A progenotoxin is defined as a chemical substance that must be metaboli-
cally activated to become a DNA-damaging agent.)
Non-genotoxins carbofuran (2,3-dihydro-2,2-dimethyl-7-benzofuranyl
methyl carbamate), a carbamate insecticide; di-2-ethylhexyl phthalate, a
plasticizer; malathion (0,0-dimethyl S-(l,2-dicarbethoxyethyl) phosphorodi-
thioate), an organophosphate insecticide; simazine (2-chloro-4,6-bis(ethyl-
amino)-s-triazine), a triazine herbicide; and permethrin (3-(phenoxyphenyl)-
methyl(I)-cis, trans-3-(2,2-dichloroethenyl)-2,2-dimethyl cyclopropane
carboxylate), a synthetic pyrethroid insecticide, were obtained from the
chemical repository of NFCRC.
Solvents dimethyl sulfoxide (DMSO), spectrophotometric grade, and
acetone were purchased from Burdwick Jackson Laboratory, Inc. (Muskegon,
Michigan). All progenotoxins were dissolved in DMSO (unless indicated),
stored frozen, and thawed at room temperature before use. Progenotoxin and
non-genotoxin working solutions were prepared in acetone (acetone:DMSO, 10:1,
V/V) and stored at 5°C in amber bottles with Teflon® cap liners.
SEDIMENT SAMPLES
Sediment samples were collected from 1988 to 1990 at twenty-eight
stations in three industrial areas: Indiana Harbor, in IN, Buffalo River in
NY, and Saginaw River in MI (Figures 1.1, 1.2, and 1.3). One-hundred g
samples, either grab or core, were dried with anhydrous sodium sulfate,
extracted with methylene chloride, subjected to gel-permeation chromatography
cleanup, evaporated under nitrogen, apd brought to volume in DMSO (Chapter 6).
Polychlorinated biphenyls (PCBs), low and high molecular polyaromatic
7-2
-------
hydrocarbons (PAHs), dioxins, and furans were present in sediment extracts
from all priority stations (Chapter 4 for complete physical and chemical
characterization of sediments). Working solutions of sediment extracts were
prepared in acetone (acetone:DMSO, 10:1, V/V) and contained 1000 mg equivalent
(eq) sediment dry weight/mL.
EXOGENOUS METABOLIC ACTIVATION
Metabolic activation of samples was accomplished with hepatic S9 from
Aroclor-1254 induced rats (Microbiological Associates, Inc., Bethesda, MD).
The S9 protein content was determined by the biuret method (Layne 1957); S9
preparations were stored at -80°C.
MUTATOX ASSAY
The Mutatox Assay uses rat liver S9 for exogenous metabolic activation
of progenotoxins and a dark mutant strain of the luminescent bacterium
Photobacterium phosphoreum for detection of genotoxins. DNA-damaging sub-
stances are detected by measuring the ability of a test extract or specific
chemical to restore the luminescent state in the bacterial cells (Ulitzur et
al. 1980). The degree of light increase indicates the relative genotoxicity
of the sample.
Genotoxicity in single chemicals (Ulitzur et al. 1980; Weiser et al.
1981; Ulitzur and Weiser 1981; Ulitzur 1986; and Microbics Corp. unpublished
data), as well as in complex mixtures was detected by Photobacterium without
activation (Kwan et al. 1990) and with activation (Johnson 1992).
ACTIVATED MUTATOX ASSAY PROTOCOL
The Mutatox Assay System,R developed and packaged by Microbics Corp.,
Carlsbad, NM, consisted of reagents, freeze—dried bacteria, cuvettes, and an
analyzer.
Preparation
Mutatox Assay Medium® (MAM), containing essential nutrients, growth
factors, buffer, and cofactor (NADP—glucose—6—phosphate regenerating system;
NADP 7.70 g/L and glucose—6—phosphate 3.40 g/L), was reconstituted with
deionized water. A 100—mL aliquot of MAM was prepared in a 250—mL beaker
using a magnetic stirrer and adjusted to 23 ±1°C. The rat hepatic S9 was
thawed in a water bath at 23°C. An ampoule of freeze—dried luminescent
bacteria (Photobacterium phosphoreum, Microbics Corp., lot #1089) was hydrated
with 1.0 mL of MAM at 23°C.
MAM was immediately inoculated with the bacteria at a ratio of 100:1,
stirred a few seconds, and incubated for 1 h at 23°C. Finally, rat hepatic S9
(40 mg protein/mL) was introduced into the MAM mixture at 0.4 mg/mL, stirred
Cor 5 s and stirred again before dispensing it into sample cuvettes.
Test Procedure
Sediment extracts were tested at concentrations from 100 mg eq dry
weight/mL to 0.19 mg eq dry weight/mL. Each test sample was serially double
7-3
-------
diluted 9 times in the bacteria—S9—MAM mixture over a 100—fold dose range to
elicit a normal dose response reaction. Ten cuvettes were placed in a row in
a holder and numbered from 1 to 10; the process was repeated for each test
sample. Using a 10—mL motorized electronic pipette (Rainin Instrument Co.,
Woburn, MA) that picks up and dispenses the liquid through a disposable
polypropylene tip, 1.0 mL of the MAM mixture was dispensed into cuvette 1 and
0.5 mL into each cuvette 2 to 10. The test sample (0.1 mL) was added to
cuvette 1 and the sample was briefly vortexed. With a disposable 1.0-mL
pipette tip, 0.5 mL of the test sample was transferred from cuvette 1 to
cuvette 2 (1:1 v/v) and vortexed a few seconds; the same procedure was
repeated sequentially diluting the entire series. Finally, to maintain a
similar surface-to-volume ratio in all cuvettes, 0.5 mL of the diluent from
the last cuvette was discarded. Each test sample as well as control(s) was
prepared in a standard dilution series as described. The treated cuvettes
were covered with aluminum foil, preincubated in a water bath at 37°C for 30
min, and immediately placed in a dark incubator with vented hood at 23 +1°C
for 16 to 24 h. Aseptic techniques were not required because of the high
salinity of test reagents and the relatively short incubation period of the
test.
Test Endpoint
The response of the luminescent bacteria was determined by measuring the
light intensity of each cuvette with a Model 500 Analyzer (Microbics Corp.,
Carlsbad, NM). A light value of 100 or more and at least three times the
light intensity of the negative control was defined as a GENOTOXIC RESPONSE;
sensitivity limits were the maximum peak concentration (MFC) and the lowest
detected concentration (LDC) in each dilution series. The DOSE-RESPONSE
NUMBER was defined as the number of genotoxic responses recorded at different
concentrations per dilution series. A DILUTION SERIES with a dose-response
number of three or more was designated genotoxic; with a dose—response number
less than three, the series was designated suspect; and without a genotoxic
response, the series was designated negative. The TEST SAMPLE was designated
GENOTOXIC when at least three replicate series indicated a mean dose—response
number of three or more, SUSPECT when at least three replicate series
indicated a mean dose-response number less than three, or NEGATIVE when ac
least three replicate series contained no genotoxic response. Three replicate
dilution series were conducted on different days before each test sample was
finally identified. All final designations were made after the mean value of
at least 30 data points (e.g., three replicate dilution series) over 100X
concentration was determined.
ASSAY CONTROLS
Three controls — positive, negative, and procedural — were used in
each assay. The positive controls consisted of four progenotoxins 2-AA, 2-AF,
BaP, and PY (tested at 10-, 5-, 2.5-, 1.2-, and 0.6 pg/cuvette); these
controls both determined the relative daily sensitivity of the assay and
established an historical sensitivity baseline. An LDC of all positive
controls (2-AA, 2-AF, BaP, and PY) <1.2/ig/cuvette validated the sensitivity of
the assay each day. When an LDC of any of the four positive controls was >1.2
7-4
-------
/zg/cuvette, the assay's sensitivity was considered suspect, the results were
rejected, and this test—run was repeated. The negative control was the
carrier solvent; this control identified the spontaneous light emission of the
dark mutant strain. The procedural control was an organic extract from a fine
silt and clay particle sediment obtained from local undisturbed agricultural
soil (Ingersoll and Nelson 1990).
CYTOTOXICITY
The relative cellular toxicity of the test chemical (or complex organic
extract) was determined by a turbidity test. Bacterial cytotoxicity of the
sediment extract(s) and test chemical(s) was determined visually by comparing
turbidity of bacterial growth in treatment and control tubes after 16 h.
Phenol (1%) was used as a positive control.
No effort was made to determine the cytotoxicity of known genotoxins at
concentrations >50 yug/test sample because the ecological relevancy of experi-
mental high—dose effects is questionable. In addition, use of high concentra-
tions of known genotoxins creates unnecessary handling hazards and disposal
problems.
MUTATOX ASSAY WITH COMPLEX MIXTURES
The sensitivity and specificity of the activated Mutatox Assay with
complex mixtures were determined by measuring genotoxic activity of known
arylamine and PAH progenotoxins in sediment extracts (procedural control, see
Assay Controls above) with and without exogenous metabolic activation, in non—
genotoxic mixtures (model complex mixture), and in binary progenotoxic
mixtures.
Progenotoxic Sensitivity
The relative sensitivity of the Mutatox Assay to detect model progeno—
toxins (2-AA, 2-AF, 2-AAF, 2-AN, BaP, 3-MC, and PY at concentrations ranging
from 0.019 to 10 jjg/cuvette) in environmental sediment extracts (methylene
chloride) was determined by comparing procedural and negative controls (see
Assay Controls above) and performing the assay with and without exogenous
activation (rat S9). An additional procedural control with the model progeno—
toxins included heat inactivation of exogenous enzymes and omission of the
regeneration system cofactor; rat S9 was heated 1 min in boiling water, cooled
in crushed ice, brought to 23°C in a water bath, and then added to the
bacteria-MAM mixture with or without NADP.
Non-genotoxic Specificity
A model complex mixture of potential non—genotoxic environmental
pollutants (carbofuran, di-2-ethylhexyl phthalate, malathion, simazine, and
permethrin) and model progenotoxins (2-AA or BaP) was tested to ascertain the
specificity of the Mutatox Assay in detecting genotoxins when other chemical
contaminants were also present in the test sample. Ten micrograms of each
non—genotoxin were mixed in MAM reaction mixture with a magnetic stirrer for
several minutes before preparing the dilution series; 10 pg of either 2-AA or
7-5
-------
BaP were introduced and serially diluted as previously described (See Test
Procedure).
Progenotoxin Interactions
Progenotoxins 2-AA, 2-AF, BaP, and PY were used to determine possible
interfering interactions of progenotoxins and metabolites in Mutatox sensitiv-
ity. In paired permutations with each other, each progenotoxin was introduced
at 10 ng (20/jg total for the combination) into the standard dilution series as
described above; the total acetone:DMSO carrier solvent did not exceed 100
/^L/cuvette.
STATISTICAL ANALYSIS
Sample variability was measured by determining the standard deviation of
the mean. Data were tested by analysis of variance with a Statistical
Analysis System computer package (SAS Institute Inc. 1985). Mean differences
were measured by one-way analysis of variance. If the overall F value was
significant (p <0.05), the mean differences were ascertained by using Fisher's
least-significant-difference test.
7-6
-------
RESULTS AND DISCUSSION
ASSAY VALIDATION: COMPLEX MIXTURES
Sensitivity
The activated Mutatox Assay detected all seven model progenotoxins in
organic sediment extracts; each clearly demonstrated a dose-response with no
observed cytotoxicity. Each progenotoxin was identified as a genotoxin when
there were three or more responses in each dilution series (dose-response
number >3). The Maximum Peak Concentration (MPC) and Lowest Dectected
Concentration (LDC) were routinely found to be 5.0 /jg/cuvette and <0.6
/zg/cuvette for each progenotoxin (Table 7.1). In Figures 7.1 and 7.2, single
data sets of 2-AA and BaP are shown in histograms; note the MPC, LDC, and
dose-response number. The relative sensitivity of the activated Mutatox Assay
to these arylamine and PAH progenotoxins (2-AA, 2-AF, 2-AAF, 2-AN, BaP, 3-MC,
and PY) was established at <1 /ig/cuvette.
Binary mixtures of four progenotoxins (2-AA + 2-AF, 2-AA + BaP, 2-AA +
PY, 2-AF +BaP, 2-AF + PY, and BaP + PY) at concentrations of 10 to 0.6
/jg/cuvette showed no evidence of inhibitory interactions (datii not shown) .
Exogenous Activation
Metabolic activation of complex mixtures was necessary for expression of
bacterial genotoxicity; controls without S9 or with heat-killed S9 (60s in
boiling water) showed no genotoxic activity in the assay (Table 7.2).
Specificity
The model complex mixture of carbofuran, di-2-ethylhexyl phthalate,
malathion, simazine, and permethrin, representing five classes of potential
aquatic contaminants, showed no genotoxic response or cytotoxicity at test
doses of <10 jig/cuvette nor did the mixture interfere with the genotoxic
expression of known progenotoxins (Table 7.2).
ENVIRONMENTAL COMPLEX MIXTURES: GREAT LAKES SEDIMENTS
Thirty—eight extracted samples collected from 28 stations in three Great
Lakes AOCs were evaluated for genotoxicity with the activated Mutatox Assay.
Indiana Harbor
Grab samples from seven stations along the Indiana Harbor were collected
in August 1989 (Table 2.1). All samples were genotoxic with an average 5.5
(0.8) dose-response number/station (Figure 7.3). The MPC detected ranged from
50 to 12 rag eq. sediment/mL and the LDC ranged from 0.7 to 0.09 mg eq.
sediment/mL. In Figure 7.4, a data set of Station 6 is shown in histogram;
note the MPC and LDC as well as dose-response number. Cytotoxicity was not
observed in the turbidity tests; positive controls (2-AA, 2-AF, B(a)P and PY)
were within acceptable sensitivity limits.
7-7
-------
Buffalo River
Grab samples from ten stations along the Buffalo River were collected in
October 1989 (Table 2.1). All samples were genotoxic with an average 5.0
(0.8) dose-response number/station (Figure 7.5). MFC detected ranged from 12
to 3 mg eq. sediment/mL and the LDC ranged from 0.75 to 0.37 mg eq.
sediment/mL. No cytotoxicity was detected; positive controls were
acceptable.
Saginaw River
Twenty—one grab and core samples from eleven stations along the Saginaw
River were collected in December 1989 (first survey) and May 1990 (third
survey). Ten of eleven staations showed evidence of genotoxic substances
(Figure 7.6 to 7.8). Grab samples from Stations 2, 3, 5, 6, 16, and 24 were
designated genotoxic and samples from Stations 4, 7, 8, and 10 were suspect
(Figure 7.6 and 7.7). In the third survey of the Saginaw River, the grab
sample from Station 1 failed to elicit a genotoxic response (Figure 7.7). The
grab sample in this survey from Station 6 was negative (Figure 7.7), but the
core sample was genotoxic (Figure 7.8). The average dose-response
number/station for grab samples was 2.3 (1.5), about one-half the number for
the Indiana Harbor and Buffalo River samples. The MFC detected ranged from 25
to 6 mg eq. sediment/mL and the LDC ranged from 0.75 to 0.18 mg eq.
sediment/mL. Core samples collected from Stations 2, 5, and 6 during the
third survey varied in depth from about 30 cm to 120 cm (Figure 7.8); all were
genotoxic with an average dose—response number of 3.7 (Figure 7.8). This
small core sampling suggested deep genotoxin penetration in the Saginaw
sediment. No cytotoxicity was observed; positive controls were acceptable.
GENOTOXIN BIOAVAIIABILITY
Most toxicological bioassays, procaryotic or eucaryotic, single cell or
multicellular, lack an important ingredient: the element of in situ exposure
to the real world. The best test can only simulate. The genotoxins recovered
from Great Lakes sediments were mobilized with organic solvents, concentrated
from large soil samples, and dissolved in compatible solvents. Therefore, the
genotoxicity findings described in this manuscript must be prefaced with the
word potential, e.g., existing in possibility, not in actuality. The bio-
availability of genotoxins in freshwater sediments — how they move in pore
water, how they sorb onto sediment components, and how they move through the
food-chain — is poorly understood. This study, however, clearly showed that
genotoxins were present in sediment as chemical contaminants and that they
could possibly alter the expression of a cell's genome, an event that in most
instances would be deleterious if not lethal to the organism.
MUTATOX AND AMES COMPARISON
The Mutatox Genotoxicity Assay compared favorably with the well-known
and validated Salmonella Mutagenicity Test (Maron and Ames, 1983; Johnson
1992) (Table 7.3 and Chapter 6). The spectra of sensitivity with seven known
or suspected arylamines and polycyclic aromatic hydrocarbon progenotoxins (2-
AAF, 2-AA, 2-AF, 2-AN, B(a)P, 3-MC, and PY) indicated both qualitative and
7-8
-------
quantitative similarities; the lowest detectable genotoxic concentrations were
in the low micrograra range (< 1) for both bacterial assays (Johnson 1992).
The Mutatox Assay and the Ames Test both with exogenous metabolic activation
(rat hepatic S9) detected genotoxins in complex mixtures extracted from the
all sediments from Areas of Concern — Indiana Harbor, Buffalo River, and
Saginaw River. In parallel studies, Mutatox with 1% S9 and one tester strain
and Ames with 10 to 30% S9 and four tester strains, the two genotoxic tests
compared favorably detecting evidence of genotoxic substances in all AOCs with
a 96% (27/28) station agreement (Chapter 6). Interestingly, the Great Lake
sediment extracts were not cytotoxic to Photobacterium. but extremely toxic to
Salmonella tester strains (Chapter 6). In the Ames test, however,
confirmation tests were necessary to demonstrate reversion of the histidine-
deficient tester strains from auxotrophic to prototrophic. When monitoring
complex environmental mixtures, confirmation with the Ames Test can be tedious
because sample extracts can cause cytotoxicity with the release of cellular
nutrients.
7-9
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SUMMARY
Genotoxins in complex organic mixtures were successfully detected by
measuring the restored bioluminescence of a dark mutant strain of Photo-
bacterium. Validation experiments showed that Mutatox was a sensitive,
specific, and predictive genotoxicity test. In field studies, evidence of
genotoxic substances was found in organic sediment extracts from all three
AOCs: Indiana Harbor, Buffalo River, and Saginaw River. Within the three AOCs
sampled, 27 stations showed evidence of genotoxins; 23 of 28 stations (82%)
were designated genotoxic, four were suspect (14%), and one was negative (3%).
Slightly over 80% (31/38) of all grab and core samples were genotoxic, about
13% (5/38) were suspect, and 5% (2/38) were negative. The Mutatox assay and
Ames test compared favorably — with 96% (27/28) station agreement — in
detecting evidence of genotoxic substances in all three AOCs. The activated
Mutatox Assay seemed to be a good screening tool because of its simplicity,
short-term duration, and sensitivity to detect genotoxins in complex
environmental sediments from the Great Lakes.
7-10
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REFERENCES CITED IN CHAPTER 7
Fabacher, D.L., J.M. Besser, C.J. Schmitt, J.C. Harshbarger, P.M. Peterman,
and J.A. Lebo. 1991. Contaminated sediment from tributaries of the Great
Lakes: chemical characterization and carcinogenic effects in Medaka (Oryias
latipes). Arch. Environ. Contam. Toxicol. 21:17-34.
Hesselberg, R.J., and J.G. Seelye. 1982. Identification of organic compounds
in Great Lakes fishes by gas chromatography/mass spectrometry: 1977. Great
Lakes Fisheries Laboratory, USFWS-GLFL/AR-82-1. U.S. Fish and Wildlife
Service, Ann Arbor, MI, 49 p.
Ingersoll, C.G., and M. K. Nelson. 1990. Testing sediment toxicity with
Hyalella azteca (Amphipoda) and Chironomus riparius (Diptera). P. 93-109. In
W.G. Landis and W. H. van der Schalie (eds.), Aquatic Toxicology and Risk
Assessment: Thirteenth Volume, ASTM STP 1096. American Society for Testing and
Materials, Philadelphia.
International Joint Commission. 1980. Pollution in the Great Lakes Basin from
land use activities. International Joint Commission, Wirisor, Ontario, pp. 7-
14.
Johnson, B.T. 1992. An evaluation of a genotoxicity assay using liver S9 for
activation and luminescent bacteria for detection. Environ. Toxicol. Chem.
11:473-480.
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7-11
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7-12
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1200 -I
1000-
Light Value
100 _
Genotoxic
Response
5 2.5 1.2 0.6 0.3 0.15 0.07 0.03 0.01
Concentration [jig/cuvette]
Figure 7.1 Genotoxicity of 2-aminoanthracene determined with the activated
Mutatox Assay (Photobacterium/rat hepatic S9).
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2000 -i
Light Value
100
1000-
Genotoxic
Response
1 o
5 2.5 1.2
Concentration [^ig/cuvette]
0.6
Figure 7.2 Genotoxicity of benzo(a)pyrene determined with the activated
Mutatox Assay (Photobacterium/rat hepatic S9) .
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10 n
8-
6-
Dose-Response
Number
4 -
2-
Genotoxic
1 o
Figure 7.3 Genotoxicity of sediment extracts from Indiana Harbor determined
with the activated Mutatox Assay (Photobacterium/rat hepatic S9).
Dose-response number - responses/dilution series; mean (dark bar)
of three replicates with standard deviation (white bar) .
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1000 -i
800-
600-
Light Value
400-
200 -
100
0 -4—
100 50 25 12 6 3 1.5 0.7 0.3
Sediment Extract [mg eq/cuvette]
Genotoxic
Response
Figure 7.4 Genotoxicity of sediment extracts from Indiana Harbor determined
with the activated Mutatox Assay (Photobacterium/rat hepatic S9):
single data set from Station 6.
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Dose-Response
Number
Genotoxic
10
STATION
Figure 7.5 Genotoxicity of sediment extracts from Buffalo River determined
with the activated Mutatox Assay (Photobacterium/rat hepatic S9).
Dose-response number - responses/dilution series; mean (dark bar)
of three replicates with standard deviation (white bar).
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Dose-Response
Number
4-
Genotoxic
2-
10
STATION
Figure 7.6 Genotoxlcity of sediment extracts from the first survey of Saginaw
River determined with the activated Mutatox Assay (Photobacterium-
/rat hepatic S9). Dose-response number = responses/dilution
series; mean (dark bar) of three replicates with standard
deviation (white bar).
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Dose-Response
Number
Genotoxic
STATION
Figure 7.7 Genotoxicity of sediment extracts from the third survey of Saginaw
River (core samples) determined with the activated Mutatox Assay
(Photobacterium/rat hepatic S9). Dose-response number -
responses/dilution series; mean (dark bar) of three replicates
with standard deviation (white bar).
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Dose-Response
Number
Genotoxic
6d
STATION
Figure 7.8 Genotoxicity of sediment extracts from the third survey of Saginaw
River (grab sample) determined with the activated Mutatox Assay
(Photobacterium/rat hepatic S9). Station number and approximate
core depth: 2a, 20 to 50 cm; 2b, 50 to 88 cm; 5a, 0 to 60 cm; 5b,
60 to 120 cm; 6a, 0 to 60 cm; 6b, 12 to 33 cm; 6c, 30 to 81 cm;
6d, 60 to 120 cm. For additional detail on core samples, see
Table 2.1. Dose—response number — responses/dilution series; mean
(dark bar) of three replicates with standard deviation (white
bar).
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Table 7.1 Genotoxicity^- of arylamines and polycyclic aromatic hydrocar-
bons in sediment extracts determined with the activated
Mutatox Assay (Photobacterium/rat hepatic S9).
PROGENOTOXIN
2-ACETAMIDOFLUORENE
2-AMINOANTHRACENE
2-AMINOFLUORENE
2-AMINONAPHTHALENE
BENZO(a)PYRENE
3-METHYLCHOLANTHRENE
PYRENE
MFC
(/ig/cuvette)
5.0
5.0
5.0
5.0
5.0
5.0
5.0
LDC
(/ig/cuvette)
<0.6
<0.6
<0.6
<0.6
<0.6
<0.6
<0.6
DOSE-RESPONSE
NUMBER
5
5
5
5
5
5
5
aMPC = maximum peak concentration; LDC = lowest detected concentration;
Dose-response number = number of genotoxic responses/dilution series;
sediment extract - procedural control.
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Table 7.2 Genotoxicity of progenotoxic chemicals^-2 in complex mixtures deter-
mined with the activated Mutatox Assay (Photobacterturn/rat hepatic S9) .
TREATMENT
CHEMICAL
BOILING
S9
WITHOUT
S9
WITH
S9
PROGENOTOXINS ND'E ND GENOTOXIC
NON-GENOTOXINS^ ND ND ND
PROGENOTOXINS +
NON-GENOTOXINS ND ND GENOTOXIC
aProgenotoxins : 2-acetamLdof 1 uorene , 2—amiiioanthraceiie , 2-aminof luorene ,
2—aminonaphthalene, benzo(a)pyrene, 3—methylcholanthrene, and pyrene.
bND = not detected
cNon—genotoxins: complex mixture of carbofuran (carbamate insecticide), di-
2—ethylphthalate (plasticizer), malathion (organophosphate insecticide).
simazine (triazine herbicide) , and permethrin (synthetic pyrethroid insecti
cide).
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Table 7.3 Comparison of the Mutatox Assay and Ames
HUTATOX AMES
TEST ORGANISM
BACTERIAL
REQUIREMENT
ENDPOINT
EXOGENOUS
ACTIVATION
TEST DURATION
TEST TEMPERATURE
RELATIVE
SENSITIVITY^
STERILITY
PROCEDURE
INSTRUMENTATION
SCIENTIFIC
DEVELOPMENT
Photobacterium
Single isolate
Light emission
Optional
16-24 h
23±1°C
<1.0 /zg/cuvette
Optional
Simple
Luminoineter or
scintillation
counter
Low
Validation phase
Salmonella
Usually one to
four isolates
Colony formation
Optional
48-72 h
37°C
<1.0 /ig/plate
Essential
Complex
Particle counter*'-
1 High
In common use
\8
aPhotobacterium/activation and Salmonella/activation genotoxic assays.
bRat S9 activation with 2-acetamidofluorene, aflatoxin B^
2-aminoanthracene, 2-aminofluorene, 2-aminonapthalene, benzo(a)pyrene, 3-
methylcholanthrene, and pyrene.
cParticle counter is essential for enumeration of large samples.
dLabor ;ind materials factor.
eExtensive literature and validation.
US. GOVEHNMENT PRINTING OffXX: 1993-546-737
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