1、Designation: E 1295 01 (Reapproved 2006)Standard Guide forConducting Three-Brood, Renewal Toxicity Tests withCeriodaphnia dubia1This standard is issued under the fixed designation E 1295; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revis
2、ion, 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. Scope*1.1 This guide describes procedures for obtaining dataconcerning the adverse effects of an effluent or
3、a test material(added to dilution water, but not to food) on Ceriodaphniadubia Richard 1894, during continuous exposure throughout aportion of the organisms life. These procedures should also beuseful for conducting life cycle toxicity tests with otherCladocera (Guide E 1193), although modifications
4、 will benecessary.1.2 These procedures are applicable to most chemicals,either individually or in formulations, commercial products, orknown mixtures, that can be measured accurately at thenecessary concentrations in water. With appropriate modifica-tions these procedures can be used to conduct test
5、s ontemperature, dissolved oxygen, pH, and on such materials asaqueous effluents (see also Guide E 1192), leachates, oils,particulate matter, sediments (see also Guide E 1383), andsurface waters. Renewal tests might not be applicable tomaterials that have high oxygen demand, are highly volatile,are
6、rapidly biologically or chemically transformed, or sorb totest chambers. If the concentration of dissolved oxygen fallsbelow 4 mg/Lor the concentration of test material decreases bymore than 20 % in test solution(s) between renewals, morefrequent renewals might be necessary.1.3 Other modifications o
7、f these procedures might be justi-fied by special needs or circumstances. Results of tests con-ducted using unusual procedures are not likely to be compa-rable to results of many other tests. Comparisons of resultsobtained using modified and unmodified versions of theseprocedures might provide usefu
8、l information on new conceptsand procedures for conducting three-brood toxicity tests withC. dubia.1.4 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 healt
9、h practices and determine the applica-bility of regulatory limitations prior to use. Specific hazardstatements are given in Section 8.1.5 This guide is arranged as follows:SectionReferenced Documents 2Terminology 3Summary of Guide 4Significance and Use 5Apparatus 6Facilities 6.1Construction Material
10、s 6.2Test Chambers 6.3Cleaning 6.4Reagents and Materials 7Hazards 8Dilution Water 9Requirements 9.1Source 9.2Treatment 9.3Characterization 9.4Test Material 10General 10.1Stock Solution 10.2Effluent 10.3Test Concentration(s) 10.4Collection 10.5Sample Containers 10.6Preservation 10.7Treatment 10.8Test
11、 Organisms 11Species 11.1Age 11.2Source 11.3Brood Stock 11.4Food 11.5Handling 11.6Quality 11.7Procedure 12Demonstration of Feasibility 12.1Experimental Design 12.2Dissolved Oxygen 12.3Temperature 12.4Preparing Test Solutions 12.5Conditioning Test Chambers 12.6Beginning a Test 12.7Renewing Test Solut
12、ions 12.8Duration of Test 12.9Biological Data 12.10Other Measurements 12.11Analytical Methodology 13Acceptability of Test 14Calculation 151This guide is under the jurisdiction of ASTM Committee E47 on BiologicalEffects and Environmental Fate and is the direct responsibility of SubcommitteeE47.01 on
13、Aquatic Assessment and Toxicology.Current edition approved April 1, 2006. Published May 2006. Originallyapproved in 1989. Last previous edition approved in 2001 as E 1295 01.1*A Summary of Changes section appears at the end of this standard.Copyright ASTM International, 100 Barr Harbor Drive, PO Box
14、 C700, West Conshohocken, PA 19428-2959, United States.Report 16AppendixesFood Appendix X1Culture Techniques Appendix X2Test Chambers Appendix X3Statistical Guidance Appendix X42. Referenced Documents2.1 ASTM Standards:2D 1193 Specification for Reagent WaterD 3978 Practice for Algal Growth Potential
15、 Testing withPseudokirchneriella subcapitataE 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 1023 Guide for Assessing the Hazard of a Material toAquatic Organisms
16、 and Their UsesE 1192 Guide for Conducting Acute Toxicity Tests onAqueous Ambient Samples and Effluents with Fishes,Macroinvertebrates, and AmphibiansE 1193 Guide for Conducting Daphnia magna Life-CycleToxicity TestsE 1383 Guide for Conducting Sediment Toxicity Tests withFreshwater Invertebrates3E 1
17、706 Test Method for Measuring the Toxicity ofSediment-Associated Contaminants with Freshwater Inver-tebratesIEEE/ASTM SI 10 American National Standard for Use ofthe International System of Units (SI): The Modern MetricSystem3. Terminology3.1 The words “must,” “should,” “may,” “can,” and “might”have
18、very specific meanings in this standard. “Must” is used toexpress an absolute requirement, that is, to state that the testhas to be designed to satisfy the specified condition, unless thepurpose of the test requires a different design. “Must” is onlyused in connection with factors that directly rela
19、te to theacceptability of the test (see Section 14). “Should” is used tostate that the specified condition is recommended and has to bemet in most tests. Although a violation of one “should” israrely a serious matter, violation of several will often render theresults questionable. Terms such as “is
20、desirable,” “is oftendesirable,” and “might be desirable” are used in connectionwith less important factors. “May” is used to mean “is (are)allowed to,” “can” is used to mean “is (are) able to,” and“might” is used to mean “could possibly.” Thus the classicdistinction between “may” and “can” is prese
21、rved, and “might”is never used as a synonym for either “may” or “can.”3.2 A brood refers, collectively, to the young neonatesreleased at the time of adult molt by the young/adult animaloriginally exposed to the control and test solutions. Thenumber of young in each brood should increase over the per
22、iodof the test. Animals may be transferred to fresh control or testsolution before completing the release of a brood, resulting insplit broods. Care is needed when interpreting the results todetermine the number of broods released during a test.3.3 For definitions of other terms used in this standar
23、d, referto Guide E 729, Terminology E 943, and Guide E 1023. For anexplanation of units and symbols, refer to IEEE/ASTM SI 10.4. Summary of Guide4.1 At the beginning of the test, at least ten C. dubia lessthan 24-h old are maintained individually in separate testchambers (or in separate compartments
24、 in two or more testchambers), exposed to control water and one (preferably 2 ormore) toxicant concentrations. One or more control treatmentsmay be used. Control treatments may include standard labora-tory water only, or some combination of standard water(s) anduncontaminated site water, to provide
25、a measure of organismsurvival and reproduction based on specific test water condi-tions, such as hardness, alkalinity, and so forth. A controltreatment consists of maintaining organisms in water to whichno test material has been added in order to provide (a)ameasure of the acceptability of the test
26、by giving an indicationof the quality of the test organisms and the suitability of thedilution water, food, test conditions, handling procedures, andso forth, and (b) the basis for interpreting data obtained fromthe other treatments. In each of the other treatments the tenorganisms are maintained in
27、 water to which a selected concen-tration of test material (percentage of effluent or river, or lakewater) has been added. Specified data on the concentration oftest material and the survival and reproduction of C. dubia arecollected and analyzed to determine the effect of the testedconcentration (%
28、 effluent or ambient water) on C. dubia.4.2 Table 1(1)4contains a summary of the conditions usedwhen conducting a three-brood test with C. dubia. Table 2 andSection 14 list the requirements that need to be met for a testto be deemed acceptable.5. Significance and Use5.1 Ceriodaphnia was first used a
29、s a toxicity test organismby Mount and Norberg (4). Introduced for use in effluent andambient water evaluations, Ceriodaphnia have also been avaluable addition to single chemical test procedures.5.2 Protection of a population requires prevention of unac-ceptable effects on the number, weight, health
30、, and uses of theindividuals of that species, or species for which the test speciesserves as a surrogate. A three-brood toxicity test is conductedto help determine changes in survival and the number ofneonates produced that result from exposure to the testmaterial.5.3 Results of three-brood toxicity
31、 tests with C. dubia mightbe used to predict chronic or partial chronic effects on speciesin field situations as a result of exposure under comparableconditions.2For referenced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.org. For Annual Book o
32、f ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.3Withdrawn.4Boldface numbers in parentheses refer to the list of references at the end of thisguide.E 1295 01 (2006)25.4 Results of three-brood toxicity tests with C. dubia mightbe compared with the c
33、hronic sensitivities of different speciesand the chronic toxicities of different materials, and to studythe effects of various environmental factors on results of suchtests.5.5 Results of three-brood toxicity tests with C. dubia mightbe useful for predicting the results of chronic tests on the samet
34、est material with the same species in another water or withanother species in the same or a different water. Most suchpredictions are based on the results of acute toxicity tests, andso the usefulness of the results of a three-brood toxicity testwith C. dubia might be greatly increased by also repor
35、ting theresults of an acute toxicity test (see Guides E 729 and E 1192)conducted under the same conditions. In addition to conductingan acute test with unfed C. dubia, it might also be desirable toconduct an acute test in which the organisms are fed the sameas in the three-brood test, to see if the
36、presence of thatconcentration of that food affects the results of the acute testand the acute chronic ratio (see 10.4.1).5.5.1 A 48 or 96-h EC50 or LC50 can sometimes beobtained from a three-brood toxicity test with a known testmaterial, but often all the concentrations in the test will bebelow the
37、EC50 or LC50. In addition, it is usually desirable toknow the EC50 or LC50 before beginning the three-brood test,as a means to determine the concentrations for use in thechronic test (see 10.4.1). It should be noted that results from anacute test may not necessarily correspond to those of a chronict
38、est, due to the addition of food to the chronic test.5.6 Three-brood toxicity tests with C. dubia might be usefulfor studying biological availability of, and structure activityrelationships between, test materials.5.7 Results of three-brood toxicity tests with C. dubia canvary with temperature, qual
39、ity and quantity of food, quality ofthe dilution water, condition of the test organisms, and otherfactors.5.8 Results of three-brood toxicity tests with C. dubia mightbe an important consideration when assessing the hazards ofmaterials to aquatic organisms (see Guide E 1023), or whenderiving water q
40、uality criteria for aquatic organisms.6. Apparatus6.1 FacilitiesCulture and test chambers should be main-tained in a constant temperature room, incubator, or recircu-lating water bath. If dilution water is not prepared batchwise, itis usually piped directly from the source of an elevatedheadbox so i
41、t can be gravity-fed into culture tanks andcontainers used to prepare test solutions. Strainers and air trapsshould be included in the water supply system. The head-boxshould be equipped for temperature control and aeration. Airused for aeration should be free of fumes, oil, and water; filtersto rem
42、ove oil and water are desirable. Filtration of air througha 0.22 m bacterial filter might be desirable (5). The facilityshould be well ventilated and free of fumes. To further reducethe possibility of contamination by test materials and othersubstances, especially volatile ones, the culture tanks sh
43、ouldnot be in a room in which toxicity tests are conducted, stock ortest solutions are prepared, effluent or test material is stored, orequipment is cleaned. During culture and testing, organismsshould be shielded from disturbances with curtains or partitionsto prevent unnecessary stress. A timing d
44、evice should be usedto provide a 16-h light and 8-h dark photoperiod. A 15- toTABLE 1 Test Conditions for Conducting Three-Brood ToxicityTests with Ceriodaphnia dubiaTest Criteria Specification1) Test Type Whole effluent, receiving water orreference toxicity test, or both, withstatic-renewal of test
45、 solution.4) Test Duration 68 days, when 60% of controlanimals produce 3 broods3) Temperature 25C (6 1C)4) Photoperiod 16 h light: 8 h dark, ambientlaboratory light levels5) Test Chamber Size 30 mL6) Test Solution Volume 15 mL7) Renewal of Test Solution every 24 or 48 hours8) Age of Test Organisms 1
46、2-h old neonatesto 360 mL of reconstituted hard or soft water contained in500-mL jars. Feed these cultures at a rate of 12 mL of 0.4 O.D.algae plus rye grass powder (see X1.2.3) daily. Transferorganisms to fresh reconstituted water on Day 3 prior to theaddition of algae rye grass powder. After the 2
47、nd brood hasbeen produced (generally Day 5 or 6 at 25C), isolate theoriginal females into 100-mL beakers containing 60 mL of theappropriate reconstituted water (50-mL beakers containing 30mL of medium will suffice). To these beakers add 1 mL of 0.4O.D. algae/30 mL of media. Discard the mass culture
48、and 1stand 2nd brood young contained therein.X2.1.1.1 Use the 3rd brood young to start new cultures andexperiments. The isolated females generally produce between10 and 16 third brood neonates for use in starting cultures andexperiments.X2.1.1.2 This technique is particularly useful when thecohort e
49、xperimental design (blocking on females) is used. Onemass culture of this type is generally sufficient to produce atleast 10 females whose young are within 12 h of each other.Cultures may be started on consecutive days to ensure thatsufficient organisms are present on any given day.X2.1.2 Use 1-L glass beakers as culture vessels. Maintaincultures in several separate vessels to provide back-up in caseone is lost due to accident or other problems such as lowdissolved oxygen (D.O.), or lack of food. Fill the 1-L culturevessels with 900 mL of media. A new culture is started eachwe
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