Significance and Use
5.1 The term duckweed commonly refers to members of the family Lemnaceae. This family has many species world-wide in 4 genera. This guide is designed for toxicity testing with one particular clone of one species of duckweed that has been extensively studied, Lemna gibba G3, although other species such as Lemna minor or Spirodela spp. can probably also be tested using the procedures described herein.
5.2 Duckweeds are widespread, free-floating aquatic plants, ranging in the world from tropical to temperate zones. Duckweeds are a source of food for waterfowl and small animals and provide food, shelter, and shade for fish. The plants also serve as physical support for a variety of small invertebrates. Duckweed is fast growing and reproduces rapidly compared with other vascular plants (1).3 Under conditions favorable for its growth, it can multiply quickly and form a dense mat in lakes, ponds, and canals, primarily in fresh water, but also in estuaries. It also grows well in effluents of wastewater treatment plants and has been suggested as a means of treating wastewaters (2). A dense mat of duckweed can block sunlight and aeration and cause fish kills (3).
5.3 Duckweed is small enough that large laboratory facilities are not necessary, but large enough that effects can be observed visually.
5.4 Because duckweed is a floating macrophyte, it might be particularly susceptible to surface active and hydrophobic chemicals that concentrate at the air-water interface. Results of duckweed tests on such chemicals, therefore, might be substantially different from those obtained with other aquatic species.
5.5 Results of toxicity tests with duckweed might be used to predict effects likely to occur on duckweed in field situations as a result of exposure under comparable conditions.
5.6 Results of tests with duckweed might be used to compare the toxicities of different materials and to study the effects of various environmental factors on results of such tests.
5.7 Results of tests with duckweed might be an important consideration when assessing the hazards of materials to aquatic organism (see Guide E1023) or when deriving water quality criteria for aquatic organisms (4).
5.8 Results of tests with duckweed might be useful for studying biological availability of, and structure-activity relationships between test materials.
5.9 Results of tests with duckweed will depend on temperature, composition of the growth medium, condition of the test organisms, and other factors. The growth media that are usually used for tests with duckweed contain concentrations of salts, minerals, and nutrients that greatly exceed those in most surface waters.
Scope
1.1 This guide describes procedures for obtaining laboratory data concerning the adverse effects of a text material added to growth medium on a certain species of duckweed (Lemna gibba G3) during a 7-day exposure using the static technique. These procedures will probably be useful for conducting toxicity tests with other species of duckweed and other floating vascular plants, although modifications might be necessary.
1.2 Special needs or circumstances might also justify modification of this standard. Although using appropriate procedures is more important than following prescribed procedures, results of tests conducted using unusual procedures are not likely to be comparable to results of many other tests. Comparison of results obtained using modified and unmodified versions of these procedures might provide useful information concerning new concepts and procedures for conducting tests with duckweed.
1.3 The procedures in this guide are applicable to most chemicals, either individually or in formulations, commercial products, or known mixtures. With appropriate modifications these procedures can be used to conduct tests on temperature and pH and on such other materials as aqueous effluents (see also Guide E1192), leachates, oils, particulate matter, sediments and surface waters. These procedures do not specifically address effluents because to date there is little experience using duckweeds in effluent testing and such tests may pose problems with acclimation of the test organisms to the receiving water. Static tests might not be applicable to materials that have a high oxygen demand, are highly volatile, are rapidly biologically or chemically transformed in aqueous solution, or are removed from test solutions in substantial quantities by the test chambers or organisms during the test.
1.4 Results of toxicity tests performed using the procedures in this guide should usually be reported in terms of the 7-day IC50 based on inhibition of growth. In some situations it might only be necessary to determine whether a specific concentration unacceptably affects the growth of the test species or whether the IC50 is above or below a specific concentration. Another end point that may be calculated is the no observed effect concentration (NOEC).
1.5 The sections of this guide appear as follows:
|   Title | Section |
|
| |
| Referenced Documents | 2 |
| Terminology | 3 |
| Summary of Guide | 4 |
| Significance and Use | 5 |
| Hazards | 6 |
| Apparatus | 7 |
|  Facilities | 7.1 |
|  Test Chambers | 7.2 |
|  Cleaning | 7.3 |
|  Acceptability | 7.4 |
| Growth Medium | 8 |
|  Test Material | 9 |
|  General | 9.1 |
|  Stock Solution | 9.2 |
|  Test Concentration(s) | 9.3 |
| Test Organisms | 10 |
|  Species | 10.1 |
|  Source | 10.2 |
|  Stock Culture | 10.3 |
| Procedure | 11 |
|  Experimental Design | 11.1 |
|  Temperature | 11.2 |
|  Illumination | 11.3 |
|  Beginning the Test | 11.4 |
|  Duration of Test | 11.5 |
|  Biological Data | 11.6 |
|  Other Measurements | 11.7 |
| Analytical Methodology | 12 |
| Acceptability of Test | 13 |
| Calculation of Results | 14 |
| Report | 15 |
1.6 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety, health, and environmental practices and determine the applicability of regulatory limitations prior to use. Specific hazard statements are given in Section 6.
1.7 This international standard was developed in accordance with internationally recognized principles on standardization established in the Decision on Principles for the Development of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.
