9637

• UCREFRP Catalog Number: 9637
• Author: American Society For Testing And Materials.
• Title: Standard Practice For Conducting Acute Toxicity Tests With Fishes, Microinvertebrates, And Amphibians.
• USFW Year: 1980.
• USFW - Doc Type: Philadelphia, PA.
• Copyright Material: YES

systems that prepare the different test solutions
independently of each other (6).
6.4.2 The metering system should be cali-
brated before and after each test. This should
include determining the flow rate through each
test chamber and measuring either the concen-
tration of toxicant in each test chamber or the
volume of solution used in each portion of the
metering system. The general operation of the
metering system should be checked twice daily
throughout the test.
6.4.3 Selection of the flow rate through the
test chambers should take into account the size
of the test chambers (6.5.5), the size of the test
organisms (10.3), and the loading (I 1.4). The
flow rate should be at least five volume addi-
tions per 24 h, but it is usually desirable to
construct the metering system so that it can
provide at least ten volume additions per day.
The flow rates should not vary by more than
20 % from anyone test chamber to any other
or from one time to another within a test.
6.5 Test Chambers:
6.5.1 Test chambers can be constructed by
welding, not soldering, stainless steel or by
gluing double-strength or stronger window
glass with clear silicone adhesive. Silicone ad-
hesive sorbs some organochlorine and organo-
phosphorus pesticides which are then difficult
to remove. Therefore, as little of the adhesive
as possible should be in contact with water;
extra beads of adhesive should be on the out-
side of chambers rather than on the inside. The
size and shape of the test chamber may affect
the results of tests with toxicants that volatilize
or sorb onto the chambers in significant quan-
tities.
6.5.2 The smallest horizontal dimension of
the test chamber must be at least 1.5 times the
average horizontal dimension of the test orga-
nisms. The depth of the test solution should be
at least 1.5 times the average height of the test
organisms. The test solution should be at least
ISO mm deep for organisms over 0.5 g (wet
weight) each, and at least 50 mm deep for
smaller organisms. Chambers filled to within
ISO mm of the top sometimes need to be cov-
ered to prevent organisms from jumping out.
6.5.3 Embryos are often exposed in glass
cups with stainless steel or nylon screen bot-
toms or cups constructed by welding stainless
steel screen. The cups must be suspended in a

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test chamber in such a way as to ensure that
test solution flows regularly into and out of the
cup and that the embryos are always sub-
merged.
6.5.4 For static tests, organisms over 0.5 g
(wet weight) each are often exposed in 18.9-L
(5-gal) wide-mouth soft-glass bottles (9) con-
taining IS L of solution or in 300 by 600 by
300-mm deep all-glass test chambers. Smaller
organisms are often exposec in 3.8-L (I-gal)
wide-mouth soft-glass bottles or battery jars
containing 2 to 3 L of solution. Daphnids and
midge larvae are often exposed in 250-mL
beakers, which are often covered to prevent
evaporation, containing 200 mL of solution.
6.5.5 For flow-through tests, chambers can
be constructed by modifying glass bottles, bat-
tery jars, or beakers to provide screened over-
flow holes, standpipes, or V -shaped notches.
Organisms over 0.5 g (wet weight) each are
often exposed in 30 L of solution in a 300 by
600 by 300-mm deep all-glass test chamber.
Smaller organisms are often exposed in 2 to 4
L of solution. In tests with daphnids or other
small organisms, the test chambers or metering
system or both must be constructed so that the
organisms are not stressed by turbulence (10).
6.6 Cleaning-Metering systems, test cham-
bers, and equipment used to prepare and store
dilution water, stock solutions, and test solu-
tions must be cleaned before use. New systems,
chambers, and equipment should be washed
with detergent and rinsed with water, pesticide-
free acetone, water, acid (such as 5 % concen-
trated nitric acid), and twice with tap or other
clean water. At the end of every test, all items
that are to be used again should be immediately
(1) emptied, (2) rinsed with water, (3) cleaned
by a procedure appropriate for removing the
toxicant tested (for example, acid to remove
metals and bases; detergent, organic solvent, or
activated carbon to remove organic com-
pounds), and (4) rinsed twice with tap or other
clean water. Acid is useful for removing min-
eral deposits, and 200 mg of hypochlorite per
litre is useful for removing organic matter and
for disinfection. (A solution containing 200 mg
of hypochlorite per litre is conveniently pre-
pared by adding 6 mL of liquid household
chlorine bleach to I L of water.) However, acid
and hypochlorite should not be mixed because
hazardous fumes may be produced. Metering

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