1、Designation: E 1192 97 (Reapproved 2003)Standard Guide forConducting Acute Toxicity Tests on Aqueous AmbientSamples and Effluents with Fishes, Macroinvertebrates, andAmphibians1This standard is issued under the fixed designation E 1192; the number immediately following the designation indicates the
2、year oforiginal adoption or, in the case of revision, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon (e) indicates an editorial change since the last revision or reapproval.1. Scope1.1 This guide covers procedures for obtaining laborato
3、rydata concerning the adverse effects of an aqueous effluent oncertain species of freshwater and saltwater fishes, macroinver-tebrates, and amphibians, usually during 2 to 4-day exposures,depending on the species, using the static, renewal, andflow-through techniques. These procedures will probably
4、beuseful for conducting acute toxicity tests on aqueous effluentswith many other aquatic species, although modifications mightbe necessary.1.2 Other modifications of these procedures might be justi-fied by special needs or circumstances. Although using appro-priate procedures is more important than
5、following prescribedprocedures, results of tests conducted using unusual proceduresare not likely to be comparable to results of many other tests.Comparison of results obtained using modified and unmodifiedversions of these procedures might provide useful informationconcerning new concepts and proce
6、dures for conducting acutetoxicity tests on aqueous effluents.1.3 This guide is based in large part on Guide E 729. Themajor differences between the two guides are (1) the maximumtest concentration is 100 % effluent or ambient sample, (2)testing is not chemical specific, and (3) the holding time ofe
7、ffluent and ambient samples is often considerably less thanthat for chemicals and other test materials. Because the sampleis often a complex mixture of chemicals, analytical tests cannotgenerally be used to confirm exposure concentrations.1.4 Selection of the technique to be used in a specificsituat
8、ion will depend upon the needs of the investigator andupon available resources. Static tests provide the most easilyobtained measure of acute toxicity, but should not last longerthan 48 h. Renewal and flow-through tests may last longer than48 h because the pH and concentrations of dissolved oxygenan
9、d effluent are maintained at desired levels and degradationand metabolic products are removed. Static tests might not beapplicable to effluents that have a high oxygen demand, orcontain materials that (1) are highly volatile, (2) are rapidlybiologically or chemically transformed in aqueous solutions
10、, or(3) are removed from test solutions in substantial quantities bythe test chambers or organisms during the test. Flow-throughtests are generally preferable to renewal tests, although in somesituations a renewal test might be more cost-effective than aflow-through test.1.5 In the development of th
11、ese procedures, an attempt wasmade to balance scientific and practical considerations and toensure that the results will be sufficiently accurate and precisefor the applications for which they are commonly used. Amajor consideration was that the common uses of the results ofacute tests on effluents
12、do not require or justify stricterrequirements than those set forth in this guide. Although thetests may be improved by using more organisms, longeracclimation times, and so forth, the requirements presented inthis guide should usually be sufficient.1.6 Results of acute toxicity tests should usually
13、 be reportedin terms of a median lethal concentration (LC50) or medianeffective concentration (EC50). In some situations, it might benecessary only to determine whether a specific concentration isacutely toxic to the test species or whether the LC50 or EC50is above or below a specific concentration.
14、1.7 This guide is arranged as follows:SectionReferenced Documents 2Terminology 3Summary of Guide 4Significance and Use 5Hazards 7Apparatus 6Facilities 6.1Special Requirements 6.2Construction Materials 6.3Metering System 6.4Test Chambers 6.5Cleaning 6.6Acceptability 6.7Dilution Water 8Requirements 8.
15、1Source 8.2Treatment 8.3Characterization 8.4Effluent 91This guide is under the jurisdiction of ASTM Committee E47 on BiologicalEffects and Environmental Fate and is the direct responsibility of SubcommitteeE47.01 on Aquatic Assessment and Toxicology.Current edition approved Oct. 1, 2003. Published O
16、ctober 2003. Originallyapproved in 1988. Last previous edition approved in 1997 as E 1192 97.1Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959, United States.Sampling Point 9.1Collection 9.2Preservation 9.3Treatment 9.4Test Concentration(s) 9.5Test O
17、rganisms 10Species 10.1Age 10.2Source 10.3Care and Handling 10.4Feeding 10.5Disease Treatment 10.6Holding 10.7Acclimation 10.8Quality 10.9Procedure 11Experimental Design 11.1Dissolved Oxygen 11.2Temperature 11.3Loading 11.4Beginning the Test 11.5Feeding 11.6Duration of Test 11.7Biological Data 11.8O
18、ther Measurements 11.9Analytical Methodology 12Acceptability of Test 13Calculation or Results 14Report 151.8 This standard does not purport to address all of thesafety concerns, if any, associated with its use. It is theresponsibility of the user of this standard to establish appro-priate safety and
19、 health practices and determine the applica-bility of regulatory limitations prior to use. Specific hazardstatements are given in Section 7.2. Referenced Documents2.1 ASTM Standards:2E 380 Practice for Use of the International System of Units(SI) (the Modernized Metric System)3E 724 Guide for Conduc
20、ting Static Acute Toxicity TestsStarting with Embryos of Four Species of SaltwaterBivalve MolluscsE 729 Guide for Conducting Acute Toxicity Tests on TestMaterials with Fishes, Macroinvertebrates, and Amphib-iansE 943 Terminology Relating to Biological Effects and En-vironmental FateE 1203 Practice f
21、or Using Brine Shrimp Nauplii as Food forTest Animals in Aquatic ToxicologyE 1604 Guide for Behavioral Testing in Aquatic Toxicology3. Terminology3.1 The words “must,” “should,” “may,” “can,” and “might”have very specific meanings in this guide. “Must” is used toexpress an absolute requirement, that
22、 is, to state that the testought to be designed to satisfy the specified condition, unlessthe purpose of the test requires a different design. “Must” isonly used in connection with factors that directly relate to theacceptability of the test (see 13.1). “Should” is used to statethat the specified co
23、ndition is recommended and ought to bemet if possible. Although violation of one “should” is rarely aserious matter, violation of several will often render the resultsquestionable. Terms such as “is desirable,” “is often desirable,”and “might be desirable” are used in connection with lessimportant f
24、actors. “May” is used to mean “is (are) allowed to,”“can” is used to mean “is (are) able to,” and “might” is used tomean “could possibly.” Thus the classic distinction between“may” and “can” is preserved, and “might” is never used as asynonym for either “may” or “can.”3.2 The term “effluents” refers
25、 to aqueous discharges regu-lated under the National Pollutant Discharge EliminationSystem (NPDES) collected at the sampling point specified inthe NPDES permit.3.3 The term “ambient samples” refers to water samplescollected from the environment. Examples include surfacewaters, storm waters, leachate
26、s, and ground water.3.4 For definitions of other terms used in this guide, refer toGuide E 729 and Terminology E 943. For an explanation ofunits and symbols, refer to Practice E 380.4. Summary of Guide4.1 In each of two or more treatments, test organisms of onespecies are maintained for 2 to 8 days
27、in one or more testchambers. In each of the one or more control treatments, theorganisms are maintained in dilution water to which no effluenthas been added in order to provide (1) a measure of theacceptability of the test by giving an indication of the qualityof the test organisms and the suitabili
28、ty of the dilution water,test conditions, handling procedures, and so forth, and (2) thebasis for interpreting data obtained from the other treatments.In each of the one or more other treatments, the organisms aremaintained in dilution water to which a selected concentrationof effluent has been adde
29、d. Data on effects on the organisms ineach test chamber are usually obtained periodically during thetest and analyzed to determine LC50s or EC50s for variouslengths of exposure.5. Significance and Use5.1 An acute effluent toxicity test is conducted to obtaininformation concerning the immediate effec
30、ts on test organ-isms of a short-term exposure to an effluent under specificexperimental conditions. One can directly examine acuteeffects of complex mixtures of chemicals as occurs in effluentsand some ambient waters. Acute effluent toxicity tests can beused to evaluate the potential for designated
31、-use or aquatic lifeimperiment in the receiving stream, lake, or estuary. An acutetoxicity test does not provide information about whetherdelayed effects will occur, although a post-exposure observa-tion period, with appropriate feeding if necessary, mightprovide such information.5.2 Results of acut
32、e effluent tests might be used to predictacute effects likely to occur on aquatic organisms in fieldsituations as a result of exposure under comparable conditions,except that (1) motile organisms might avoid exposure whenpossible, (2) toxicity to benthic species might be dependent onsorption or sett
33、ling of components of the effluent onto the2For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.3Withdrawn.E 1192
34、97 (2003)2substrate, and (3) the effluent might physically or chemicallyinteract with the receiving water.5.3 Results of acute effluent tests might be used to comparethe acute sensitivities of different species and the acutetoxicities of different effluents, and to study the effects ofvarious enviro
35、nmental factors on results of such tests.5.4 Acute tests are usually the first step in evaluating theeffects of an effluent on aquatic organisms.5.5 Results of acute effluent tests will depend on thetemperature, composition of the dilution water, condition of thetest organisms, exposure technique, a
36、nd other factors.6. Apparatus6.1 FacilitiesAlthough some small organisms can be heldand acclimated in static or renewal systems, most organismsare held, acclimated, and cultured in flow-through systems.Test chambers should be in a constant-temperature room,incubator, or recirculating water bath. A d
37、ilution-water tank,which may be used to store receiving water, or a headbox isoften elevated so dilution water can be gravity-fed into holdingand acclimation tanks and test chambers. Pumps are often usedto deliver dilution water and effluent to headboxes and tanks.Strainers and air traps should be i
38、ncluded in the water supply.Headboxes and holding, acclimation, culture, and dilution-water tanks should be equipped for temperature control andaeration (see 8.3). Air used for aeration should be free offumes, oil, and water; filters to remove oil and water aredesirable. Filtration of air through a
39、0.22 m bacterial filtermight be desirable (5). The facility should be well ventilatedand free of fumes. To further reduce the possibility of contami-nation by components of the effluent and other substances,especially volatile ones, holding, acclimation, and culture tanksshould not be in a room in w
40、hich toxicity tests are conducted,effluent is stored, test solutions are prepared, or equipment iscleaned. During holding, acclimation, culture, and testing,organisms should be shielded from disturbances with curtainsor partitions to prevent unnecessary stress. A timing deviceshould be used to provi
41、de a 16-h light and 8-h dark photope-riod. A 15 to 30-min transition period (6) when the lights go onmight be desirable to reduce the possibility of organisms beingstressed by large, sudden increases in light intensity. A transi-tion period when the lights go off might also be desirable.6.2 Special
42、RequirementsSome organisms require specialconditions during holding, acclimation, and testing. For ex-ample, burrowing mayfly nymphs should be provided a sub-strate suitable for burrowing (7); immature stream insectsshould be in a current (8); and crabs, shrimp, and bottom-dwelling fish should be pr
43、ovided a silica-sand substrate.Because cannibalism might occur among many species ofdecapod crustaceans, the claws of crabs and crayfish should bebanded, or the individuals should be physically isolated bymeans of screened compartments.6.3 Construction MaterialsEquipment and facilities thatcontact e
44、ffluent samples, test solutions, or any water into whichtest organisms will be placed should not contain substancesthat can be leached or dissolved by aqueous solutions inamounts that adversely affect aquatic organisms. In addition,equipment and facilities that contact effluent samples or testsoluti
45、ons should be chosen to minimize sorption of effluentcomponents from water. Glass, Type 316 stainless steel, nylon,and fluorocarbon plastics should be used whenever possible tominimize dissolution, leaching, and sorption, except that stain-less steel should not be used in tests on metals in salt wat
46、er.Concrete and rigid plastics may be used for holding, acclima-tion, and culture tanks and in the water supply, but they shouldbe soaked, preferably in flowing dilution water, for a week ormore before use (9). Cast iron pipe should not be used with saltwater and probably should not be used in a fre
47、shwater-supplysystem because colloidal iron will be added to the dilutionwater and strainers will be needed to remove rust particles. Aspecially designed system is usually necessary to obtain saltwater from a natural water source (see Guide E 729). Brass,copper, lead, galvanized metal, and natural r
48、ubber should notcontact dilution water, effluent, or test solutions before orduring the test. Items made of neoprene rubber or othermaterials not mentioned above should not be used unless it hasbeen shown that either (1) unfed individuals of a sensitiveaquatic species (see 8.2.3) do not show more si
49、gns of stress,such as discoloration, unusual behavior, or death, when heldfor at least 48 h in static dilution water in which the item issoaking than when held in static dilution water that does notcontain the item, or (2) their use will not adversely affectsurvival, growth, or reproduction of a sensitive species.6.4 Metering System:6.4.1 For flow-through tests, the metering system should bedesigned to accommodate the type and concentration(s) of theeffluent and the necessary flow rates of test solutions. Thesystem should mix the effluent with the dilution water imme-diately befo
