1、Designation: E1295 01 (Reapproved 2013)Standard Guide forConducting Three-Brood, Renewal Toxicity Tests withCeriodaphnia dubia1This standard is issued under the fixed designation E1295; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revisio
2、n, the year of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () 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 a t
3、est 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 E1193), although modifications wil
4、l 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 tests on
5、temperature, dissolved oxygen, pH, and on such materials asaqueous effluents (see also Guide E1192), leachates, oils,particulate matter, sediments (see also Guide E1383), andsurface waters. Renewal tests might not be applicable tomaterials that have high oxygen demand, are highly volatile,are rapidl
6、y 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 of thes
7、e 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 useful info
8、rmation 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 health prac
9、tices 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 Materials 6.2T
10、est 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 Organ
11、isms 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 Solutions 1
12、2.8Duration of Test 12.9Biological Data 12.10Other Measurements 12.11Analytical Methodology 13Acceptability of Test 14Calculation 15Report 161This guide is under the jurisdiction of ASTM Committee E47 on BiologicalEffects and Environmental Fateand is the direct responsibility of SubcommitteeE47.01 o
13、n Aquatic Assessment and Toxicology.Current edition approved March 1, 2013. Published March 2013. Originallyapproved in 1989. Last previous edition approved in 2006 as E1295 01(2006).DOI: 10.1520/E1295-01R13.*A Summary of Changes section appears at the end of this standardCopyright ASTM Internationa
14、l, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United States1AppendixesFood Appendix X1Culture Techniques Appendix X2Test Chambers Appendix X3Statistical Guidance Appendix X42. Referenced Documents2.1 ASTM Standards:2D1193 Specification for Reagent WaterD3978 Practice for A
15、lgal Growth Potential Testing withPseudokirchneriella subcapitataE729 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 t
16、oAquatic Organisms and Their UsesE1192 Guide for Conducting Acute Toxicity Tests on Aque-ous Ambient Samples and Effluents with Fishes,Macroinvertebrates, and AmphibiansE1193 Guide for Conducting Daphnia magna Life-CycleToxicity TestsE1383 Guide for Conducting Sediment Toxicity Tests withFreshwater
17、Invertebrates (Withdrawn 1995)3E1706 Test Method for Measuring the Toxicity of Sediment-Associated Contaminants with Freshwater InvertebratesIEEE/ASTM SI 10 American National Standard for Use ofthe International System of Units (SI):The Modern MetricSystem3. Terminology3.1 The words “must,” “should,
18、” “may,” “can,” and “might”have 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
19、 with factors that directly relate 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
20、questionable. Terms such as “is 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
21、between “may” and “can” is preserved, 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 br
22、ood should increase over the periodof 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
23、other terms used in this standard, referto Guide E729, Terminology E943, and Guide E1023. 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
24、 (or in separate compartments 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) anduncontami
25、nated site water, to provide a measure of organismsurvival and reproduction based on specific test waterconditions, 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
26、 acceptability of the test 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 ten
27、organisms are maintained in 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
28、 the testedconcentration (% 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 Ce
29、riodaphnia was first used as 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
30、 the number, weight, health, 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 Resu
31、lts of three-brood toxicity 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 service
32、astm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.3The last approved version of this historical standard is referenced onwww.astm.org.4Boldface numbers in parentheses refer to the list of references at the end of thisguide.
33、E1295 01 (2013)25.4 Results of three-brood toxicity tests with C. dubia mightbe compared with the chronic 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 t
34、oxicity tests with C. dubia mightbe useful for predicting the results of chronic tests on the sametest 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
35、of the results of a three-brood toxicity testwith C. dubia might be greatly increased by also reporting theresults of an acute toxicity test (see Guides E729 and E1192)conducted under the same conditions. In addition to conductingan acute test with unfed C. dubia, it might also be desirable toconduc
36、t an acute test in which the organisms are fed the sameas in the three-brood test, to see if the 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 toxici
37、ty test with a known testmaterial, but often all the concentrations in the test will bebelow the 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 sho
38、uld be noted that results from anacute test may not necessarily correspond to those of a chronictest, 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
39、 materials.5.7 Results of three-brood toxicity tests with C. dubia canvary with temperature, quality 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 a
40、ssessing the hazards ofmaterials to aquatic organisms (see Guide E1023), or whenderiving water quality 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 i
41、s not prepared batchwise, itis usually piped directly from the source of an elevatedheadbox so it 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 c
42、ontrol and aeration. Airused for aeration should be free of fumes, oil, and water; filtersto remove 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 con
43、tamination by test materials and othersubstances, especially volatile ones, the culture tanks shouldnot 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
44、 shielded from disturbances with curtains or partitionsTABLE 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 solution.4) Test Durat
45、ion 68 days, when 60% of controlanimals produce 3 broods3) Temperature 25C ( 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 12-h old neonatesto 360 m
46、L 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 2nd brood hasbeen produce
47、d (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 and 1stand 2nd brood you
48、ng 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 experimental design (bloc
49、king 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 eachweek, and the oldest cul
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