1、Designation: E1193 97 (Reapproved 2012)Standard Guide forConducting Daphnia magna Life-Cycle Toxicity Tests1This standard is issued under the fixed designation E1193; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revision, the year of last
2、 revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.1. Scope1.1 This guide covers procedures for obtaining laboratorydata concerning the adverse effects of a test material (added todilut
3、ion water, but not to food) on Daphnia magna Straus,1820, during continuous exposure throughout a life-cycle usingthe renewal or flow-through techniques. These procedures alsoshould be useful for conducting life-cycle toxicity tests withother invertebrate species, although modifications might benece
4、ssary.1.2 These procedures are applicable to most chemicals,either individually or in formulations, commercial products, orknown mixtures. With appropriate modifications, these proce-dures can be used to conduct tests on temperature, dissolvedoxygen, pH, and on such materials as aqueous effluents (a
5、lsosee Guide E1192), leachates, oils, particulate matter,sediments, and surface waters. The technique, (renewal orflow-through), will be selected based on the chemical charac-teristics of the test material such as high oxygen demand,volatility, susceptibility to transformation (biologically orchemic
6、ally), or sorption to glass.1.3 Modification of these procedures might be justified byspecial needs or circumstances. Although using appropriateprocedures is more important than following prescribedprocedures, results of tests conducted using unusual proceduresare not likely to be comparable to resu
7、lts of standard testprocedures. Comparison of results obtained using modified andunmodified versions of these procedures might provide usefulinformation on new concepts and procedures for conductinglife-cycle toxicity tests with D. magna.1.4 This guide is arranged as follows:SectionReferenced Docume
8、nts 2Terminology 3Summary of Guide 4Significance and Use 5Apparatus 6Facilities 6.1Construction Materials 6.2Test Chambers 6.3Cleaning 6.4Acceptability 6.5Reagents 7Purity of Reagents 7.1Hazards 8Dilution Water 9Requirements 9.1Source 9.2Treatment 9.3Characterization 9.4Test Material 10General 10.1S
9、tock Solutions 10.2Test Concentrations(s) 10.3Test Organisms 11Species 11.1Age 11.2Source 11.3Brood Stock 11.4Food 11.5Handling 11.6Harvesting Young 11.7Quality 11.8Procedure 12Experimental Design 12.1Dissolved Oxygen 12.2Temperature 12.3Loading 12.4Selection of Test System 12.5Beginning the Test 12
10、.6Care and Maintenance 12.7Feeding 12.8Duration 12.9Biological Data 12.10Other Measurements 12.11Analytical Methodology 13Acceptability of Test 14Calculation of Results 15Report 16Keywords 17AppendixesAppendix X1 Statistical Guidance1This guide is under the jurisdiction of ASTM Committee E47 on Biol
11、ogicalEffects and Environmental Fateand is the direct responsibility of SubcommitteeE47.01 on Aquatic Assessment and Toxicology.Current edition approved Dec. 1, 2012. Published December 2012. Originallyapproved in 1987. Last previous edition approved in 2004 as E1193 97 (2004).DOI: 10.1520/E1193-97R
12、12.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States1SectionAppendix X2 Food1.5 The values stated in SI units are to be regarded asstandard. No other units of measurement are included in thisstandard.1.6 This standard does not purport t
13、o 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 health practices and determine the applica-bility of regulatory limitations prior to use. Specific hazardstatements are given in Section 8.2
14、. Referenced Documents2.1 ASTM Standards:2E729 Guide for Conducting Acute Toxicity Tests on TestMaterials with Fishes, Macroinvertebrates, and Amphib-iansE943 Terminology Relating to Biological Effects and Envi-ronmental FateE1023 Guide for Assessing the Hazard of a Material toAquatic Organisms and
15、Their UsesE1192 Guide for Conducting Acute Toxicity Tests on Aque-ous Ambient Samples and Effluents with Fishes,Macroinvertebrates, and AmphibiansIEEE/ASTM SI 10 American National Standard for Use ofthe International System of Units (SI):The Modern MetricSystem3. Terminology3.1 The words “must,” “sh
16、ould,”“ may,” “can,” and “might”have very specific meanings in this guide.3.2 mustused to express an absolute requirement, that is,to state that the test ought to be designed to satisfy the specifiedcondition, unless the purpose of the test requires a differentdesign. “Must” is used only in connecti
17、on with factors thatdirectly relate to the acceptability of the test (see 14.1).3.3 shouldused to state that the specified condition isrecommended and ought to be met if possible. Althoughviolation of one “should” is rarely a serious matter, violation ofseveral will often render the results question
18、able. Terms suchas “is desirable,” “is often desirable,” and “might be desirable”are used in connection with less important factors.3.4 mayused to mean “is (are) allowed to,” “can” is usedto mean “is (are) able to,” and “might” is used to mean “couldpossibly.” Therefore the classic distinction betwe
19、en “may” and“can” is preserved, and “might” is never used as a synonym foreither“ may” or “can.”3.5 For definitions of other terms used in this guide, refer toGuide E729 and Terminology E943. For an explanation ofunits and symbols, refer to IEEE/ASTM SI 10.4. Summary of Guide4.1 A 21-day life-cycle
20、toxicity test for Daphnia magna isdescribed. The test design allows for the test organisms to beexposed to a toxicant using either the renewal technique (withexchange of the total volume of test water and toxicant at leastthree times a week) or the flow-through technique (withcontinual water and tox
21、icant addition, usually at least fourvolume additions per day).At least five concentrations of a testmaterial, a control, and a solvent control (if applicable)replicated at least four times are recommended. Each testconcentration has at least ten Daphnia per treatment. Thetechnique (renewal or flow-
22、through) which uses a minimum often daphnids per treatment has only one daphnid per replicate,whereas the typical technique (renewal or flow-through) uti-lizes four replicates with at least five daphnids per replicate(20 daphnids per treatment).Acontrol consists of maintainingdaphnids in dilution wa
23、ter to which no test material has beenadded to provide (1) a measure of the acceptability of the testby giving an indication of the quality of the test organisms andthe suitability of the dilution water, food, test conditions,handling procedures, and so forth, and (2) the basis forinterpreting data
24、obtained from the other treatments. In each ofthe other treatments, the daphnids are maintained in dilutionwater, to which a selected concentration of test material hasbeen intentionally added. Measurement end points obtainedduring the test include the concentration of the test material andfinal num
25、ber alive, final weight, and number of progeny perdaphnid. Then data are analyzed to determine the effect of thetest material on survival, growth, and reproduction of D.magna.5. Significance and Use5.1 Protection of an aquatic species requires prevention ofunacceptable effects on populations in natu
26、ral habitats. Toxic-ity tests are conducted to provide data that may be used topredict what changes in numbers and weights of individualsmight result from similar exposure to the test material in thenatural aquatic environment. Information might also be ob-tained on the effects of the material on th
27、e health of the species.5.2 Results of life-cycle tests with D. magna are used topredict chronic effects likely to occur on daphnids in fieldsituations as a result of exposure under comparable conditions.5.2.1 Life-cycle tests with D. magna are used to comparethe chronic sensitivities of different s
28、pecies, the chronic tox-icities of different materials, and study the effects of variousenvironmental factors on the results of such tests.5.2.2 Life-cycle tests with D. magna are used to assess therisk of materials to aquatic organisms (see Guide E1023)orderive water quality criteria for aquatic or
29、ganisms (1).35.2.3 Life-cycle tests with D. magna are used to predict theresults of chronic toxicity tests on the same test material withthe same species in another water or with another species in thesame or a different water. Most such predictions take intoaccount the results of acute toxicity tes
30、ts, and so the usefulness2For 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.3The boldface numbers in parentheses
31、 refer to the list of references at the end ofthis guide.E1193 97 (2012)2of the results of a life-cycle test with D. magna is greatlyincreased by also reporting the results of an acute toxicity test(see Guide E729) conducted under the same conditions. Inaddition to conducting an acute toxicity test
32、with unfed D.magna, it may be desirable to conduct an acute test in whichthe daphnids are fed the same as in the life-cycle test to see ifthe presence of that concentration of that food affects theresults of the acute test and the acute-chronic ratio (ACR) (see10.3.1).5.2.4 Life-cycle tests are used
33、 to evaluate the biologicalavailability of, and structure-activity relationships between,test materials and test organisms.5.3 Results of life-cycle tests with D. magna might beinfluenced by temperature (2), quality of food, composition ofdilution water, condition of test organisms, and other factor
34、s.6. Apparatus6.1 FacilitiesCulture and test chambers are often kept in aroom maintained at about 20C but at separate locations.Alternatively, culture and test chambers may be placed in atemperature-controlled water bath or environmental chamberor incubator. The water-supply system should provide an
35、adequate supply of dilution water to the culture tanks and testchambers. The water-supply system should be equipped fortemperature control and aeration, and strainers and air trapsshould be included in the water-supply system. Air used foraeration should be free of fumes, oil, and water; filters tor
36、emove oil and water are desirable. Filtration of air through a0.22-m bacterial filter might be desirable (3). During culturingand testing, daphnids should be shielded from disturbances toprevent unnecessary stress. The test facility should be well-ventilated and free of fumes.Atiming device should b
37、e used toprovide a 16-h light and 8-h dark photoperiod (4).A15to30-min transition period when lights go on might be desirableto reduce the possibility of daphnids being stressed by instan-taneous illumination; a transition period when lights go off mayalso be desirable.6.1.1 When D. magna are fed al
38、gae, a high-light intensitymight cause sufficient photosynthesis to result in an increase ofpH high enough to kill daphnids (5). Therefore, the maximumacceptable intensity is dependent on the buffer capacity of thedilution water, species, and density of algae, and the kind oftest chamber and cover.
39、Light intensities up to 600 lx or afluence rate of 1 w/m2will usually be acceptable, but higherintensities might result in an unacceptably high pH in theculture water.6.2 Construction MaterialsEquipment and facilities thatcontact stock solutions, test solutions, or any water into whichdaphnids will
40、be placed should not contain substances that canbe leached or dissolved by aqueous solutions in amounts thatcan adversely affect daphnids. In addition, equipment andfacilities that contact stock solutions or test solutions should bechosen to minimize sorption of test materials from water.Glass, Type
41、 316 stainless steel, nylon, fiberglass, silicon, andfluorocarbon plastics should be used whenever possible tominimize leaching, dissolution, and sorption. Concrete andrigid (unplasticized) plastics may be used for culture tanks andin the water-supply system, but they should be soaked, prefer-ably i
42、n flowing dilution water, for several days before use (6).Cast-iron pipe may be used in supply systems, but colloidaliron probably will be added to the dilution water and strainerswill be needed to remove rust particles. Copper, brass, lead,galvanized metal, and natural rubber should not contact dil
43、u-tion water, stock solutions, or test solutions before or duringthe test. Items made of neoprene rubber and other materials notpreviously mentioned should not be used unless it has beenshown that their use will not adversely affect survival, growth,and reproduction of D. magna (see Section 14).6.3
44、Test Chambers:6.3.1 Flow-through tests, 500-mL to 2-L glass beakers (orequivalent) with a notch (approximately 4 by 13 cm) cut in thelip may be used to expose the Daphnia to the test material. Thenotch should be covered with 0.33-mm opening (U.S. standardsieve size No. 50) stainless steel or polyeth
45、ylene screeningsmall enough to retain first instar Daphnia. The screen can beattached to the beaker with silicone adhesive. The chambersshould provide at least 30 mL of solution for each of the initialtest daphnid(s).6.3.2 Renewal tests, beaker ranging in size from 100 to 1000mL.Anotched chamber is
46、not required for a renewal test. Eachchamber should provide at least 40 mL of solution for each ofthe initial test daphnid(s).6.3.3 Any container made of glass, Type 316 stainless steel,or a fluorocarbon plastic may be used if (1) each chamber isseparate with no interconnections, (2) each chamber co
47、ntains atleast 30 mL of test solution (see 12.4) per first-generationdaphnid for flow-through tests and at least 40 mL for renewaltests, (3) there is at least 1000 mm2of air to water interface perdaphnid, and (4) the test solution is at least 30 mm deep. Statictest chambers should be covered with gl
48、ass, stainless steel,nylon, or fluorocarbon plastic covers to keep out extraneouscontaminants and to reduce evaporation of test solution. Allchambers and covers in a test must be identical. Covers are notrequired for flow-through studies.6.4 CleaningTest chambers and equipment used to pre-pare and s
49、tore dilution water, stock solutions, and test solutionsshould be cleaned before use. New equipment should bewashed with detergent and rinsed with water, a water-miscibleorganic solvent, water, acid (such as 5 % concentrated nitricacid), and washed at least twice with distilled, deionized, ordilution water. Some lots of some organic solvents might leavea film that is insoluble in water. Also, stronger nitric acid, forexample, 10 %, might cause deterioration of silicone adhesive;an initial rinse with 10 % concentrated hydrochloric acid mightprevent such deterioration.Adichro
