8080

8080 SEDIMENT POREWATER TESTING*
8080 A. Introduction
1. Applications
The standard approach for assessing the quality or potential
toxicity of marine or estuarine sediments has been to expose
macrobenthic organisms directly to whole sediments for a
specified time, after which the survival of the test species was
determined. Whole-sediment methods
1,2
have limitations, in-
cluding use of adult macrobenthic organisms and the use of
mortality as the primary endpoint. In addition, the standard
amphipod test protocol
1,2
underestimates the potential toxic-
ity of contaminated sediments because the pore water is
flushed out and replaced with fresh overlying water before
exposure begins.
The porewater toxicity test approach offers several advantages
over the standard whole-sediment method. Sensitive life stages
of sensitive species can be used in tests using sublethal end-
points. There are no artifacts produced by sediment texture. A
dilution series test design can be used easily for better differen-
tiation among highly toxic samples. Whereas whole sediment
preferably should be tested within 2 weeks of collection, studies
indicate that pore water can be stored in the frozen state for
extended periods without any change in toxicity.
3
Most of the
studies on porewater testing have focused on marine and estua-
rine species.
2. References
1. U.S. ENVIRONMENTAL PROTECTION AGENCY & U.S. ARMY CORPS OF
ENGINEERS. 1991. Evaluation of Dredged Material Proposed for
Ocean Disposal, EPA-503/8-91/001. U.S. Environmental Protection
Agency Off. Research & Development & U.S. Army Corps of
Engineers, Washington, D.C.
2. AMERICAN SOCIETY FOR TESTING AND MATERIALS. 2004. Standard test
method for measuring the toxicity of sediment-associated contami-
nants with estuarine and marine invertebrates. E1367-03, Annual
Book of ASTM Standards, Vol. 11.05. American Soc. Testing &
Materials, West Conshohocken, Pa.
3. CARR, R.S. & D.C. CHAPMAN. 1995. Comparison of methods for
conducting marine and estuarine sediment pore water toxicity tests. I.
Extraction, storage and handling techniques. Arch. Environ. Contam.
Toxicol. 28:69.
3. Bibliography
ADAMS, D.D. 1991. Sediment pore water sampling. Chapter 7. In
A. Mudroch & S.D. MacKnight, eds., Handbook of Techniques for
Aquatic Sediments Sampling. CRC Press, Inc., Boca Raton, Fla.
BURTON, G.A., JR., ed. 1992. Sediment Toxicity Assessment. Lewis
Publishers, Inc., Boca Raton, Fla.
CARR,R.S.&M.NIPPER, eds. 2003. Porewater Toxicity Testing: Bio-
logical, Chemical, and Ecological Considerations. SETAC Press,
Pensacola, Fla.
8080 B. Sediment Collection and Storage
Sediment collection methods vary considerably with specific
study objectives. Remove as much overlying water as possible
from the sample before placing it in the sample container or
adding it to a composite sample. Nearly fill sample container to
minimize headspace, allowing some room for sample to be
rehomogenized in its original container. Extract pore water as
soon as possible after sample collection. If the sediment sample
cannot be processed immediately, store on ice or refrigerate at 4°C.
The toxicity of pore water extracted from refrigerated sediments can
change considerably after weeks or even days.
8080 C. Extraction of Sediment Pore Water
Methods that have been used to obtain sediment pore (inter-
stitial) water include centrifugation,
1–3
pressurized (pneumatic or
mechanical “squeezing”) extraction,
4–8
vacuum (suction) meth-
ods,
9,10
and equilibration methods using dialysis membranes or
fritted glass samplers.
4,11,12
Studies comparing recovery efficien-
cies of trace metals and organics for different extraction methods
indicate that substantial losses of nonpolar contaminants (e.g.,
fluoranthene and p,p⬘-DDE) can occur with all methods.
13
Tox-
icity tests with echinoderm gametes and embryos have been
conducted to compare the toxicity of pore water obtained by
various extraction techniques.
14
These studies suggest that
centrifugation minimizes loss of nonpolar contaminants. Loss
of metals is comparable among the various extraction meth-
ods. Centrifugation is preferable to filtration for removing
* Approved by Standard Methods Committee, 1997. Editorial revisions, 2009.
Joint Task Group: 20th Edition—Robert S. Carr (chair), Marion Nipper, Donald
J. Reish.
1

particulates because it minimizes adsorptive loss of contam-
inants.
Sandy sediments do not compact appreciably during centrif-
ugation, making pore water recovery difficult; however, the
pneumatic extraction method (Figures 8080:1 and 2) is particu-
larly effective.
11
The vacuum method is least expensive for
small-scale projects.
Regardless of the method used for initial extraction, centrifuge
the extracted pore water to remove suspended particulates for
fertilization and embryo development assays with echinoderms
and mollusks.
1. Centrifugation
Use a centrifuge equipped with a swinging bucket-type rotor
capable of spinning 100- to 1000-mL bottles at 10 000 ⫻g. Use
glass or polycarbonate tubes or bottles to minimize adsorption of
soluble contaminants on container wall. For some sediments, it
may be possible to decant supernatant without disturbing the
pellet; however, for most sediments, use a pipet to transfer the
supernatant to a separate container.
2. Pressurized Squeeze Extraction
The most common squeeze extraction devices use com-
pressed air (or nitrogen) to pressurize a cylinder containing
the sediment. Normally, use a filter in the bottom of the
cylinder so minimal sediment is introduced into the porewater
sample. Some filters (e.g., glass fiber filters) can adsorb a high
percentage of nonpolar contaminants from solution.
13,14
Other
filter materials (e.g., polyester and nylon) are preferable.
Before use, test any part of the extraction device that contacts
pore water during extraction for toxicity. Fill extraction de-
vice with a small volume of test dilution water and, after a
minimum of 8 h, test dilution water for toxicity. Soak new
filters in deionized water or test dilution water, with several
exchanges for at least 24 h to remove any residual contami-
Figure 8080:1. Pneumatic system for porewater extraction. Source: CARR, R.S. & D.C. CHAPMAN. 1995. Comparison of methods for conducting marine and
estuarine sediment porewater toxicity tests. I. Extraction, storage and handling techniques. Arch. Environ. Contam. Toxicol. 28:29.
SEDIMENT POREWATER TESTING (8080)/Extraction of Sediment Pore Water
2
SEDIMENT POREWATER TESTING (8080)/Extraction of Sediment Pore Water

nants before use. Between samples, acid-wash the parts of the
extraction devices that come in contact with the sample.
3. Vacuum Extraction
The simplest vacuum extraction system is a fused-glass air
stone attached with aquarium air-line tubing to a polypropylene
syringe. Apply vacuum by bracing the syringe plunger or using
a vacuum pump. Modify the system with TFE* tubing and a
glass syringe when loss of contaminants due to adsorption is a
concern. This method is inexpensive and may retain more vol-
atile compounds than centrifugation or pressurized extraction.
Vacuum methods may be more time-consuming than other ex-
traction methods when large (⬎1 L) volumes are needed, par-
ticularly for fine-grained sediments. Thoroughly rinse all system
components before use to remove residual toxicants.
15
Deter-
mine effectiveness of the rinsing procedure by testing the
toxicity of test dilution water held in the vacuum extraction
system for a minimum of 8 h. Pore water extracted via vacuum
methods from sandy sediments has a higher particulate con-
tent than pore water obtained by the other methods; if the
suspended particulates are not removed before testing, they
may produce a response in fertilization and embryo develop-
ment toxicity tests.
4. Equilibration Methods
The most commonly used equilibration technique for collect-
ing pore water involves a small-volume vessel with a membrane
placed in the sediment and allowed to equilibrate with the
surrounding interstitial water.
11,16,17
The limitations to this tech-
nique are that only milliliter volumes can be obtained within a
reasonable time (days). Test toxicity of components used to
construct the equilibration device by soaking device in clean test
dilution water or clean sediment for at least the same length of
time as the longest equilibration period to be used.
5. References
1. EDMUNDS, W.M. & A.H. BATH. 1976. Centrifuge extraction and
chemical analysis of interstitial waters. Environ. Sci. Technol.
10:467.
2. GIESY, J.P., R.L. GRANEY, J.L. NEWSTED, C.J. ROSIU,A.BENDA, R.G.
KREIS & F.J. HORVATH. 1988. Comparison of three sediment bioas-
say methods using Detroit River sediments. Environ. Toxicol.
Chem. 7:483.
3. LANDRUM, P.F., S.R. NIHART, B.J. EADIE & L.R. HERCHE. 1987.
Reduction in bioavailability of organic contaminants to the amphi-
pod Pontoporeia hoyi by dissolved organic matter of sediment
interstitial waters. Environ. Toxicol. Chem. 6:11.
4. BENDER, M., W. MARTIN,J.HESS,F.SAYLES,L.BALL &C.LAMBERT.
1987. A whole-core squeezer for interfacial pore-water sampling.
Limnol. Oceanogr. 32:1214.
5. CARR, R.S., D.C. CHAPMAN, C.L. HOWARD &J.BIEDENBACH. 1996.
Sediment quality triad assessment survey in the Galveston Bay,
Texas system. Ecotoxicol. 5:341.
6. CARR, R.S. & D.C. CHAPMAN. 1992. Comparison of whole sediment
and pore-water toxicity tests for assessing the quality of estuarine
sediments. Chem. Ecol. 7:19.
7. JAHNKE, R.A. 1988. A simple, reliable, and inexpensive pore-water
sampler. Limnol. Oceanogr. 33:483.
8. REEBURGH, W.S. 1967. An improved interstitial water sampler.
Limnol. Oceanogr. 12:163.
9. KNEZOVICH, J.P. & F.L. HARRISON. 1987. A new method for
determining the concentrations of volatile organic compounds in
sediment interstitial water. Bull. Environ. Contam. Toxicol.
38:937.
10. WINGER, P.V. & P.J. LASIER. 1991. A vacuum-operated pore-water
extractor for estuarine and freshwater sediments. Arch. Environ.
Contam. Toxicol. 21:321.
11. DITORO, D.M., J.D. MAHONY, D.J. HANSEN, K.J. SCOTT, M.B. HICKS,
S.M. MAYR & M.S. REDMOND. 1990. Toxicity of cadmium in sedi-
ments: the role of acid volatile sulfide. Environ. Toxicol. Chem.
9:1487.
12. HESSLIN, R.H. 1976. An in situ sampler for close interval pore water
studies. Limnol. Oceanogr. 21:912.
13. SCHULTS, D.W., S.P. FERRARO, L.M. SMITH, F.A. ROBERTS & C.K.
POINDEXTER. 1992. A comparison of methods for collecting intersti-
tial water for trace organic compounds and metals analyses. Water
Res. 26:989.
14. CARR, R.S. & D.C. CHAPMAN. 1995. Comparison of methods for
conducting marine and estuarine sediment pore water toxicity tests.
I. Extraction, storage and handling techniques. Arch. Environ. Con-
tam. Toxicol. 28:69.
* Teflon威, or equivalent.
Figure 8080:2. Detail of porewater extraction cylinder. (For dimensions
in centimeters, multiply dimensions in inches by 2.54.)
Source: CARR, R.S. & D.C. CHAPMAN. 1995. Comparison of
methods for conducting marine and estuarine sediment pore-
water toxicity tests. I. Extraction, storage and handling tech-
niques. Arch. Environ. Contam. Toxicol. 28:29.
SEDIMENT POREWATER TESTING (8080)/Extraction of Sediment Pore Water
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SEDIMENT POREWATER TESTING (8080)/Extraction of Sediment Pore Water

15. PRICE, N.M, P.J. HARRISON, M.R. LANDRY,F.AZAM & K.J.F. HALL.
1986. Toxic effects of latex and Tygon tubing on marine phyto-
lankton, zooplankton and bacteria. Mar. Ecol. Prog. Ser. 34:41.
16. BOTTOMLEY, E.Z. & I.L. BAYLY. 1984. A sediment pore water
sampler used in root zone studies of the submerged macrophyte,
Myriophyllum spicatum.Limnol. Oceanogr. 29:671.
17. CARIGNAN, R. 1984. Interstitial water sampling by dialysis: method-
ological notes. Limnol. Oceanogr. 29:667.
8080 D. Toxicity Testing Procedures
1. General Procedures
Because of the difficulty in obtaining large volumes of pore
water, organisms and life stages that only require small volumes
are most amenable to testing with pore water. For tests requiring
more than7dtocomplete, preferably use a static renewal test
design to ensure acceptable water quality. Short-term toxicity
tests have been used most frequently with pore water. Much of
the general guidance provided in Section 8010 is applicable to
testing with pore water. More specific guidance can be found in
sections for particular species or groups of organisms (e.g.,
Sections 8510, 8610, and 8710).
2. Exposure Chambers
The type of exposure chamber used depends on the test. Most
porewater tests are conducted in relatively small volumes (i.e.,
ⱕ10 mL). Preferably cover test chambers to minimize evapora-
tion and resulting salinity increases during the exposure period.
Scintillation vials (20 mL) with polyethylene or polypropylene
cap liners are ideal inexpensive disposable test chambers for
many species. Avoid caps with urea-formaldehyde liners be-
cause these can be toxic. Stender dishes with ground-glass lids
(20-mL capacity with 10 mL of exposure media) make excellent
exposure chambers for tests that require microscopic examina-
tion of the test organisms without transferring them to another
container (e.g., the Dinophilus gyrociliatus life-cycle test).
1
3. Organisms
Many types of organisms have been used in porewater tests.
Minute species or larval forms are preferable not only for their
small volume requirements, but also because they tend to be the
most sensitive. Most of the studies on porewater testing have
focused on marine and estuarine species.
a. Marine and estuarine species: A commercially available
test system,* which detects changes in the photoluminescence of
the marine bacterium Photobacterium phosphoreum as an end-
point, has been used more frequently in freshwater porewater
studies
2– 4
than in marine or estuarine pore waters. Although the
small sample size required is well suited for limited sample sizes,
the sensitivity of the standard assay of this type for pore water
from freshwater, estuarine, or marine sediments is low compared
to those of other toxicity tests.
Algal studies with Ulva fasciata and Ulva lactuca suggest that
a zoospore germination endpoint is as sensitive as some of the
most sensitive embryological development assays used in pore-
water testing. This test appears to be particularly resistant to
ammonia toxicity. Many algal species used in microplate proce-
dures could easily be adapted for use with porewater samples.
Porewater toxicity testing has been conducted with the
polychaete Dinophilus gyrociliatus.
1,5,6
Other minute polychaetes,
such as Ctenodrilus serratus or Ophryotocha spp.,
7,8
can be tested
in small volumes.
The mollusk tests used most successfully with pore water are
fertilization and embryological development tests with the aba-
lone Haliotes refugens. Other more common embryological de-
velopment tests with oysters
9
and clams
10
could be adapted for
use with porewater samples.
Most of the toxicity testing with marine and estuarine pore
water has been conducted with sea urchin gametes and em-
bryos.
6,11,12
The species most commonly used is the sea urchin
Arbacia punctulata but other species of sea urchin (e.g., Strongy-
locentrotus spp. and Lytechinus spp.), as well as the sand dollar
(e.g., Dendraster spp.), also have been used successfully. Types
of tests include fertilization tests, embryological development
tests, and cytogenetic assay.
13
Fish embryos and larvae of red drum Sciaenops ocellatus also
have been used successfully in porewater testing.
14
b. Freshwater species: Only a limited number of species have
been used in porewater studies with fresh water. A number of
studies with a commercially available system* have been re-
ported.
2,3,15
The freshwater amphipod Hyalella azteca has been
used to test the toxicity of pore water from freshwater sedi-
ments.
3,15
Ceriodaphnia dubia also has been used in life-cycle
tests with pore water.
4. References
1. CARR, R.S., J.W. WILLIAMS & C.T.B. FRAGATA. 1989. Development
and evaluation of a novel marine sediment pore water toxicity test
with the polychaete Dinophilus gyrociliatus.Environ. Toxicol.
Chem. 8:533.
2. GIESY, J.P., R.L. GRANEY, J.L. NEWSTED, C.J. ROSIU,A.BENDA, R.G.
KREIS & F.J. HORVATH. 1988. Comparison of three sediment bioassay
methods using Detroit River sediments. Environ. Toxicol. Chem. 7:483.
3. GIESY, J.P., C.J. ROSIU, R.L. GRANEY & M.G. HENRY. 1990. Benthic
invertebrate bioassays with toxic sediment and pore water. Environ.
Toxicol. Chem. 9:233.
4. ANKLEY, G.T., K. LODGE, D.J. CALL, M.D. BALCER, L.T. BROOKE,
P.M. COOK, R.J. KREIS,JR., A.R. CARLSON, R.D. JOHNSON, G.J.
NIEMI, R.A. HOKE, C.W. WEST, J.P. GIESY, P.D. JONES & Z.C.
FUYING. 1992. Integrated assessment of contaminated sediments in
the lower Fox River and Green Bay, Wisconsin. Ecotoxicol. Envi-
ron. Safety 23:46.
5. CARR, R.S., M.D. CURRAN &M.MAZURKIEWICZ. 1986. Evaluation of
the archiannelid Dinophilus gyrociliatus for use in short-term life-
cycle toxicity tests. Environ. Toxicol. Chem. 5:703.
* Microtox威, or equivalent.
SEDIMENT POREWATER TESTING (8080)/Toxicity Testing Procedures
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SEDIMENT POREWATER TESTING (8080)/Toxicity Testing Procedures

6. CARR, R.S. & D.C. CHAPMAN. 1992. Comparison of whole sediment
and pore-water toxicity tests for assessing the quality of estuarine
sediments. Chem. Ecol. 7:19.
7. REISH, D.J. & R.S. CARR. 1978. The effect of heavy metals on the
survival, reproduction, development and life cycles for two species
of polychaetous annelids. Mar. Pollut. Bull. 9:24.
8. CARR, R.S. & D.J. REISH. 1977. The effect of petroleum hydrocar-
bons on the survival and life history of polychaetous annelids. In
D.A. Wolfe, ed., Fate and Effects of Petroleum Hydrocarbons in
Marine Ecosystems and Organisms. Pergamon Press, New York, N.Y.
9. LONG, E.R., M.R. BUCHMAN, S.M. BAY, R.J. BRETELER, R.S. CARR,
P.M. CHAPMAN, J.E. HOSE, A.L. LISSNER,J.SCOTT & D.A. WOLFE.
1990. Comparative evaluation of five toxicity tests with sediments
from San Francisco Bay and Tomales Bay, California. Environ.
Toxicol. Chem. 9:1193.
10. LAUGHLIN, R.B., JR., R.G. GUSTAFSON &P.PENDOLEY. 1989. Acute
toxicity of tributyltin (TBT) to early life history stages of the hard
shell clam, Mercenaria mercenaria.Bull. Environ. Contam. Toxi-
col. 42:352.
11. CARR, R.S. & D.C. CHAPMAN. 1995. Comparison of methods for
conducting marine and estuarine sediment pore water toxicity tests.
I. Extraction, storage and handling techniques. Arch. Environ. Con-
tam. Toxicol. 28:69.
12. LONG, E.R., R.S. CARR, G.A. THURSBY & D.A. WOLFE. 1995. Sedi-
ment toxicity in Tampa Bay: Incidence, severity, and spatial extent.
Fla. Sci. 58:163.
13. HOSE, J.E., H.W. PUFFER, P.S. OSHIDA & S.M. BAY. 1983. Develop-
mental and cytogenetic abnormalities induced in the purple sea
urchin by environmental levels of benzo(a)pyrene, Arch. Environ.
Contam. Toxicol. 12:319.
14. ROACH, R.W., R.S. CARR, C.L. HOWARD & B.W. CAIN. 1993. An
assessment of produced water impacts in Galveston Bay system.
U.S. Fish Wild. Serv. Rep.
15. WINGER, P.V., R.J. LASIER &H.GEITNER. 1993. Toxicity of sedi-
ments and pore water from Brunswick Estuary, Georgia. Arch.
Environ. Contam. Toxicol. 25:371.
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