Fish and Daphnia Toxicity as Surrogates for Aquatic Vascular Plants and Algae Eugene E. Kenaga" and Robert J. Moolenaar Health and Environmental Sciences, The Dow Chemical Company, Midland, Mich. 48640
The relative degree of acute toxicity of thousands of chemicals of heterogeneous structures was compared for fishes, daphnids, aquatic vascular plants, and algae. Aquatic animals were more sensitive indicators of toxic effects than were plants or algae. Compounds that were toxic a t under 1 ppm for daphnids, fishes, aquatic vascular plants, and algae represented 2.4, 1.3,0.12, and 0.02%, respectively, of the total tested for each of these four groups of organisms. Of the 27 781 chemicals tested on aquatic plants and 49 082 tested on the alga Chlorella, all but 3 compounds were either not toxic to these organisms under 1 mg/L, or were toxic to fish a n d Daphnia a t 1 ppm or less. During the manufacture, distribution, use, and disposal of chemicals, the potential exists for some of them t o reach the water environment. The water environment, in contrast to the air environment, has a relatively small volume, is generally not rapidly mixed, and is more unreactive toward chemicals. The potential impact of chemicals on aquatic organisms is therefore usually greater than their potential impact on terrestrial organisms exposed to contact with air. Consequently, laboratory ecotoxicity testing programs have stressed studies on aquatic organisms rather than terrestrial organisms. T h e choice of organisms that are indicative of toxicity to aquatic species must be very selective. The number of organisms living in the aquatic environment is very large (20 000 species of fish alone), so the selection of appropriate indicator organisms is critical. Kenaga ( I ) correlated acute toxicity data for eight organisms commonly used for evaluation of environmental hazards, and concluded that both Daphnia and fish were appropriate indicators for the toxicity of chemicals t o aquatic animals. This is consistent with proposed guidelines for the registration of pesticides in the United States (2). Plants are also important in the ecology of the aquatic environment. Biologists generally regard aquatic plants as less sensitive than aquatic animals to chemical toxicants, but documentation for a wide spectrum of chemicals is lacking. For a number of years screening tests against a variety of aquatic organisms have been carried out at The Dow Chemical Company. Included was a daphnid, four species of fish, an alga. and five species of vascular plants. Tests performed on these organisms were the basis for the comparison of the relative acute toxicities of chemicals to aquatic plants and animals reported in this paper. It was felt that this large data base o f chemicals was very useful for initial hazard evaluation even though not meeting the strict standards of present day LCso studies.
Tt..st M e t h o d s The chemicals tested were obtained sequentially from many sources inside and outside The Dow Chemical Company over 0013-936X/79/0913-1479$01 .OO/O
a period of more than 20 years, and routinely screened a t Dow for many different types of biological activity. The chemicals tested include those originally prepared for screening or for use as insecticides, fungicides, nematocides, fumigants, terrestrial herbicides, aquatic herbicides, coccidiostats, plasticizers, drugs, algacides, microbiocides, deflocculation, chelation nitrogen stabilizers, catalysts, solvents, ion exchange materials, resins, cleaners, latexes, coatings, pigments, lubricants, preservatives, flotation agents, emulsifiers, organic and inorganic chemical intermediates, inhibitors, stabilizers, antifreezes, hydraulic fluids, plastic monomers, plastic surfactants, adhesives, flame retardants, highway products, metals, and other inorganic and organic chemicals. The Dow Chemical Company screening list of over 100 000 compounds can roughly be divided chemically as 0.86% inorganic, 52.0% with benzene rings, 35.7% with heterocyclic rings, 8.2% with carbocyclic rings (excluding benzene), 5.3% containing P, 63.0% containing N, 6.2% containing Br, and 27.2% containing C1 compounds. Most, but not all, of the compounds included in the smaller lists were tested on the same organisms as those in the longer list of compounds. Chemicals were formulated for testing as ball-milled powders or acetone-water suspensions. Dilutions of the chemical were made in chlorinated Lake Huron water, which has a p H of 7.6 and total alkalinity of 85 mg/L. The water was aerated for several days before being used as static test water for fish and Daphnia. Concentrations were not replaced during the test. Untreated and solvent controls on the organisms rarely showed mortality and if in excess of 10% were discounted. Normally compounds showing 100% activity at the initial concentration were reduced in concentration by one-half until less than 100% activity was shown. Tests were conducted a t 65-70 O F and were not replicated. The following species of fish were tested as alternates for each other in about equal frequency in the screening tests: emerald shiner, Notropis atherinoides; fathead minnow, Pimephales promelas; goldfish, Carrasius auratus; and carp, C'tprinus carpio. Two fish were placed in a plastic box containing 500 mL of water usually exposed to an initial concentration of 2 mg/L. Mortality counts were taken after 24-h exposure. About 50 Daphnia magna were used for each test as mixed adults and nymphs using the same test conditions as those used for fish. The following five species of aquatic submersed and floating vascular plants were tested: Carolina fanwort, Cabomba caroliniana; waterweed, Anacharis (Elodea)spp.; moneywort, Lysimachia nummularia; coontail, Ceratophyllum spp.; and Salvirzia rotundifolia. Plants were usually tested in 500 mL of water in concentrations of 10 or 100 ppm for 24 h, and held in untreated water for approximately 3 weeks before final mortality readings were taken.
@ 1979 American Chemical Society
Volume 13, Number 12, December 1979
1479
Table 1. Comparative Toxicity of Chemicals Screened on Daphnia, Fish, Aquatic Plants, and Algae concn tested, ppm
1-1.99 0.1-0.99 0.01-0.09
total no. of compds tested a
compounds giving 100% control of organism fishes aquatic plants
Daphnia
Chlorella
no.
%a
no.
%a
no.
%a
no.
%a
992 823 203 33 909
2.9 2.4 0.6
1529 473 51 35 305
4.3 1.3 0.14
196 33 0 27 781
0.71 0.12 0
263 12 3 49 082
0.54 0.02 0.006
Percent of total compounds tested giving 100% mortality (control) as defined in the text.
The alga was tested in an aqueous nutrient solution (Hoagland’s)beginning a t 10 mg/L. Vapor losses were reduced by use of a Saran film over the opening of the glass container. Mortality was determined visually by change in color (death) or by use of colorimetry to measure chlorophyll density comparative to the control after 1-week exposure to the test compound. Tabulation of Data. Using the lowest concentration giving 100% mortality as the comparative index of biological activity, data from each of the four groups of organisms (fish, daphnids, aquatic vascular plants, and Chlorella) were divided into three toxicity classes as follows: (1) 0.01-0.09 mg/L, (2) 0.1-0.99 mg/L, (3) 1.0-1.99 mg/L. All compounds in toxicity classes 1 and 2 also appear in class 3.
Results and Discussion The number of chemicals tested against the four groups of organisms listed in Table I varied from 27 781 to 49 032, and thus represented a much larger number and more varied sample of chemical structures than is ordinarily available for comparison. A t concentrations below 2 mg/L, a greater percentage of chemicals showed toxicity toward fish or daphnids than toward aquatic plants or alga, confirming the generally held view that aquatic animals are more sensitive to chemical toxicants than aquatic plants. None of the 22 781 compounds tested were toxic to aquatic plants below 0.1 mg/L. Chlorella was generally less sensitive than aquatic vascular plants despite the fact that exposures were for 7 days compared to l day for the other organisms. There were only 3 chemicals out of 49 082 tested on Chlorella that were toxic a t concentrations below 0.1 mg/L. Recent studies of organic contaminants in water have shown their concentrations rarely exceed 1mg/L. Sheldon and Hites ( 3 )identified 100 chemical contaminants in Delaware River water, but reported none a t concentrations higher than 15 pg/L. In a study by the EPA ( 4 ) of priority pollutants in wastewater effluents, concentrations reported were generally below 10 pg/L, with a few in the 10 to 100 pg/L range. In view of these observations, it seemed appropriate to examine further the relative toxicities of chemicals to aquatic plants and animals for those compounds t h a t are toxic to plants below 1 mg/L. Thirty-three chemicals (or 0.12% of the total tested) were lethal to aquatic plants below 1 mg/L, as shown in Table I. Two of these compounds were not tested for toxicity to fish or daphnids. Of the remaining 31 compounds, 22% were toxic (compounds killing 100% of the organisms) to daphnids, 62% to fish, and 68% to fish or daphnids a t concentrations below 1 mg/L. At
