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Species Sensitivity Distributions for Nonylphenol to Estimate Soil Hazardous Concentration Jin Il Kwak, Jongmin Moon, Dokyung Kim, Rongxue Cui, and Youn-Joo An Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.7b04433 • Publication Date (Web): 08 Nov 2017 Downloaded from http://pubs.acs.org on November 9, 2017
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Species Sensitivity Distributions for Nonylphenol to Estimate Soil Hazardous Concentration Jin Il Kwak, Jongmin Moon, Dokyung Kim, Rongxue Cui, Youn-Joo An* Department of Environmental Health Science, Konkuk University, Seoul, Korea * Corresponding author. Tel.: +82-2-2049-6090 Fax: +82-2-2201-6295 E-mail:
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Abstract
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Nonylphenol is an endocrine-disrupting chemical that mimics estrogenic activity. Few studies
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have investigated the soil ecotoxicity of nonylphenol in the environment, based on
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probabilistic approaches. The present study generated soil toxicity data for nonylphenol
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through bioassays that determined the acute and chronic species sensitivity distributions and
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estimated the hazardous concentrations of nonylphenol in soil in order to protect soil
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ecosystems. We used eight soil-based organisms from six taxonomic groups for acute assays
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and five soil-based organisms from four taxonomic groups for chronic assays. The hazardous
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concentration values of nonylphenol in soil, based on acute and chronic species sensitivity
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distributions, were estimated using compiled data from the present study, as well as previous
25
studies. This is the first study that generated sufficient data to develop species sensitivity
26
distributions for nonylphenol in soil, and to determine hazardous concentrations of
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nonylphenol for soil environments.
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Keywords; Nonylphenols; 4-nonylphenol; endocrine-disrupting chemical; species sensitivity
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distribution; soil; terrestrial species
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INTRODUCTION
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Nonylphenols are used in pesticides, packaging, surfactant cleaners, and detergents1,
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with high production volumes2. Nonylphenols are endocrine-disrupting chemicals classified
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as xenoestrogens3,4. Due to increasing concerns about the release of nonylphenols in the
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environment and their endocrine-disrupting properties, the use of nonylphenol in items such
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as domestic cleaning products has been restricted in Europe5 and Korea6.
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Nonylphenols (CAS No. 25154–52–3, C6H4(OH)C9H19) include isomers with a straight
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chain, such as p-nonylphenol and 4-n-nonylphenol (CAS No. 104-40-5), and those with
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branched chains, such as 4-nonylphenol (CAS No. 84852-103), among others1,4,7,8. In the
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environment, the most common form is branched 4-nonylphenol1,7, which is detected more
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frequently than nonylphenols with straight chains4.
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Nonylphenols are classified as H400 (very toxic to aquatic life) and H410 (very toxic to
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aquatic life, with long-lasting effects) according to the United Nations’ globally harmonized
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system of classification and labelling of chemicals (GHS)8,9. There are considerable data on
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the toxic effects of nonylphenol on aquatic species, including fish10,11, water fleas12,13, and
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freshwater algae14,15.
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However, there have been few studies using environmental risk assessments of soil
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ecotoxicity of nonylphenols, as shown in Tables 1 and 2, even though nonylphenols can be
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released into the soil environment and persist there8,16. Nonylphenol concentrations have been
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measured at 68–8834 µg/kg soil in the United States17, 2720 µ/kg soil in Canada16, and < 0.5
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µg/kg soil in Korea18.
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The present study aimed to estimate the hazardous concentration of nonylphenol in soil
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ecosystems, according to environmental risk assessments. Because previous soil ecotoxicity
55
data are not sufficient to determine reliable hazardous concentrations based on probabilistic
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approaches, we conducted a battery of bioassays using soil-based organisms of taxonomically
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similar groups, including species that coexist in a community, and then generated datasets to
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derive species sensitivity distributions. Finally, we estimated acute and chronic hazardous
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concentrations for nonylphenol in soil. This is the first study evaluating the hazardous
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concentration of nonylphenol in soil using probabilistic ecological risk assessment
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approaches.
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MATERIALS AND METHODS
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Test Nonylphenol and Soil Preparation
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Nonylphenol (4-n-nonylphenol ≥ 98 % purity) in powder form was obtained from Alfa
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Aesar (Alfa Aesar, Thermo Fisher Scientific Inc., Seoul, Korea). The test soil was organic
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natural soil LUFA 2.2 (Landwirtschaftliche Untersuchungs und Forschungsanstalt).
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Physicochemical soil properties were pH 6.17, organic matter 4.7%, total nitrogen 1188
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mg/kg, total phosphate 265 mg/kg, cation exchange capacity 7 Cmol+/kg, and water-holding
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capacity (WHC) 0.473 mL/g. Dried soil was spiked with nonylphenol solution in acetone
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(99.9% purity, DUCKSAN, Daejeon, Korea) to make a stock soil concentration of 2000
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mg/kg dry soil, after which the acetone was allowed to evaporate at least overnight, with a
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maximum evaporation time of 24 h. Subsequently, each test concentration was prepared by
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diluting the stock soil using solvent control soil. The solvent control soil was prepared by
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spiking dried soil with acetone only, after which the acetone was allowed to evaporate at least
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overnight (maximum evaporation time of 24 h). Each test soil for test species was thoroughly
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mixed with a roller mixer at least overnight (maximum mixing time of 24 h) before bioassays
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were conducted. Control soil was spiked with deionized water.
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Bioassays
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A battery of bioassays was performed to estimate the hazardous concentrations of
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nonylphenol in soil, using probabilistic approaches. To investigate short-term effects of
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nonylphenol, eight soil species, including plants, algae, and invertebrates, were tested: mung
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bean (Vigna radiata), rice (Oryza sativa), two green algae species (Chlorococcum infusionum
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and Chlamydomonas reinhardtii), nematode (Caenorhabditis elegans), earthworm (Eisenia
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andrei), and two springtail species (Lobella sokamensis and Folsomia candida). These
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organisms represent six taxonomic groups: Magnoliopsida, Liliopsida, Chlorophyceae,
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Secernentea, Clitellata, and Collembola. To investigate long-term effects of nonylphenol, five
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soil species were assessed: V. radiata, O. sativa, C. infusionum, C. reinhardtii, and F.
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candida. These species represent four taxonomic groups: Magnoliopsida, Liliopsida,
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Chlorophyceae, and Collembola. Ecotoxicity test methods followed the OECD guidelines19-21
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and SCI articles22-25.
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Plant Assays
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The crop plants mung bean (V. radiate, Magnoliopsida, Namyangju, Korea) and rice (O.
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sativa, Liliopsida, Chungju, Korea) were tested to assess the phytotoxicity of nonylphenol.
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Plant assays were conducted according to the modified OECD test guidelines20. Acute and
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chronic effects of nonylphenol were investigated after 14 d and 21 d, respectively20. The tests
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used nonylphenol at concentrations of 100, 500, 1000, 1500, or 2000 mg/kg dry soil for both
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plant species. A 200-g amount of each test soil was added to a square pot (9 × 9 × 9 cm), and
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deionized water was added up to 67% of the WHC. Five seeds were placed in each pot, and
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each treatment was tested in triplicate. Test pots were maintained in a plant growth chamber
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(Vision Scientific Co. Ltd., Daejeon, Korea) at 25°C, 70% humidity, and a 16:8 h light:dark
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cycle. During exposure, deionized water was replenished every two or three days. Changes in
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shoot growth of mung bean and rice were measured after 14 d and 21 d.
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Soil Algae Assays
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Soil algae are producers and food sources in soil ecosystems and are used in soil
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ecotoxicity studies26,27. Test species for the soil algae assays were Chlorococcum infusionum
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(Chlorophyceae) and Chlamydomonas reinhardtii (Chlorophyceae), which inhabit soil
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environments. C. infusionum and C. reinhardtii were purchased from the University of
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Göttingen (Germany) and the University of Texas (Austin, USA), respectively. Toxicity of
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nonylphenol to soil algae in the soil media was investigated following methods that were
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previously developed in our laboratory. In brief, 0.5 g of each test soil was added to a 12-well
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plate, and then deionized water was added up to 65% of the WHC. Subsequently, algae cell
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suspensions were placed on the soil surface. The initial algal cell density was 3×106 cells/mL,
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and final moisture content was 90% of the WHC. C. infusionum was exposed to nonylphenol
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at 25, 50, 100, 200, and 300 mg/kg dry soil. C. reinhardtii was exposed to nonylphenol at 300,
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400, 500, 600, 800, and 1000 mg/kg dry soil. Test well plates were maintained in the
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incubator at 24°C, 100 rpm, 16:8 h light:dark cycle, and 4400–8900 lux. The change in
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chlorophyll-a, which indicates changes in soil algal growth, was measured after 6 d. After 6 d
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of exposure, ethanol (DUCKSAN, Daejeon, Korea) was spiked to each well to extract the
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chlorophyll-a from soil algae. Chlorophyll-a was extracted for 3 h at 24°C in darkness.
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Chlorophyll-a was measured using a fluorescence microplate reader (Gemini, Molecular
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Devices, USA; excitation 420 nm and emission 671 nm). The levels of 50% effect
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concentration (EC50) and 10% effect concentration (EC10) were considered to constitute acute
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effects and chronic effects, respectively28.
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Soil Nematode Assay
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Caenorhabditis elegans, a soil nematode in the class Secernentea, is a common chemical
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test organism29-31. Soil nematode assays were modified and performed according to Kim et
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al.24. C. elegans were exposed to nonylphenol at concentrations of 10, 100, 500, 1000 and
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2000 mg/kg dry soil. There were four replicates for each concentration. First, 3 g of each test
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soil was added to a 24-well plate, followed by 0.2 mL of K-medium (0.032 M KCl, 0.051 M
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NaCl). Subsequently, 10 young adult C. elegans were added to each well. After 24 h of
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exposure, soils in each well were moved to an NGM agar plate (NaCl, 3 g/L; peptone, 2.5
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g/L; agar, 17 g/L; 1 M potassium phosphate, 25 mL; 1 M CaCl2·2H2O, 1 mL; 1 M
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MgSO4·7H2O, 1 mL; cholesterol, 1 mL). After an additional 6 h, the number of offspring
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was counted. Test units were maintained in an incubator at 20°C under dark conditions. In the
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soil nematode assay, 24 h exposure was considered acute exposure, based on the ASTM
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(American Society for Testing and Materials) international standards31.
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Earthworm Assay
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Earthworms are an indicator organism for soil ecotoxicity32. In the earthworm assay,
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Eisenia andrei (Clitellata) was tested based on modified OECD test guidelines19,23.
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Earthworms were exposed to nonylphenol at concentrations of 1000, 1300, 1500, 1700, and
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2000 mg/kg dry soil. Each concentration was tested with 20 replicates. First, 10 g of each test
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soil was added to a glass vial (ID 25 mm, H 50 mm, volume 20 mL), followed by deionized
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water up to 60% of the WHC. An adult earthworm (300–600 mg) was added to each test vial
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and the vial was capped with a silicon stopper. Test units were maintained in an incubator at
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20°C in dark conditions for 7 d. Normalcy of earthworms (survival, mucous secretion,
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bleeding, swelling, thinning, and fragmentation) was measured at 7 d. In the earthworm assay,
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7 d of exposure was considered acute exposure23.
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Collembola Assay
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In the Collembola assay, the soil-dwelling species Lobella sokamensis and Folsomia
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candida were selected as test species. Collembola chronic and acute exposure assays were
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conducted according to the OECD test guidelines21 and followed An et al.22. Both
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Collembola species were exposed to nonylphenol at concentrations of 25, 50, 100, 150, and
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250 mg/kg dry soil. Each concentration was tested in triplicate. First, 24 g of each test soil
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was added to a flat-bottom glass vial (volume 80 mL), followed by 6 mL of deionized water.
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A total of 10 adult L. sokamensis were exposed for 5 d, after which survival was recorded. In
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this assay, 5 d exposure was considered acute exposure. In the F. candida assay, acute effects
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and chronic effects of nonylphenol were evaluated at 14 d and 21 d, respectively21. A total of
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10 juvenile F. candida (aged 9–10 days) were exposed, and adult survival was recorded after
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14 d. Subsequently, the number of F. candida offspring was observed after 28 d. Test vials
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were maintained in an incubator at 20°C under dark conditions.
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Statistical Analysis
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To estimate the endpoint values, the solvent control normalized datasets of all test
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species were analyzed using Probit analysis33, Trimmed Spearman-Karber methods34,35, and
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Dunnett’s tests36. EC10 values and EC50 values with 95% significance level (p < 0.05) were
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estimated by Probit analysis and Trimmed Spearman-Karber analysis, respectively. Dunnett’s
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tests were used for calculating no-observed-effect concentration (NOEC) values based on
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multiple comparisons33.
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Estimation of Hazardous Concentration (HC) of Nonylphenol in Soil
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HCx refers to the concentration that protects (100 - x)% of species in an ecosystem. The
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acute and chronic hazardous concentrations for x% (HCx) were estimated based on species
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sensitivity distributions (SSD)28. In the present study, HCx for nonylphenol based on SSD
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were derived from two different sources: (1) using only our data and (2) using compiled data
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(ours and previous data), using the USEPA SSD generator (ver. 1.0)37. Previous soil toxicity
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results of nonylphenols are presented in Table 1. Direct comparisons between data from the
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present and previous studies were not possible because soil toxicity depends on soil
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properties such as organic matter, pH, and clay and moisture content38. Therefore, the soil
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organic matter normalized endpoint value was used to generate soil SSD38. In the present
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study, the common organic matter content of Korean soil (3.11%)39 was applied to normalize
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toxicity results.
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RESULTS AND DISCUSSION
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Plant Assays
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No inhibition of germination in mung bean (≥ 93% germination) or rice (≥ 87%
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germination) was observed. In the acute plant assay, after 14 d exposure (Fig. 1), shoot
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growth of mung bean was not affected by nonylphenol at concentrations below 2000 mg/kg,
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while shoot growth of rice was reduced by 31% at 2000 mg/kg dry soil based on solvent
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control normalized results. Shoot growth-EC50 (14 d) values for mung bean V. radiata and
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rice O. sativa were calculated as > 2000 and 1793 mg/kg dry soil, respectively (Table 3).
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Shoot growth of mung bean was also inhibited after chronic exposure for 21 d. Shoot growth
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of rice was reduced by 38% at 2000 mg/kg dry soil after chronic exposure for 21 d. As shown
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in Table 3, the shoot growth-EC10 and NOEC values (21 d) for mung bean V. radiata were
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estimated as 1822 and > 2000 mg/kg dry soil, respectively. For rice, the shoot growth-EC10
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and NOEC values (21 d) were estimated as 1433 and 1500 mg/kg dry soil, respectively. In
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this plant assay, rice showed a more sensitive response to nonylphenol than did mung bean.
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Previously, phytotoxicity of nonylphenol was investigated in soil40-42 as well as in
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solution42-47. Soil phytotoxicity tests were conducted in lettuce Lactuca sativa42, wheat
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Triticum aestivum, rape Brassica napus41, bok choy Brassica rapa, and perennial ryegrass
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Lolium perenne40 (Table 1). These previous studies showed that 14 to 15 d exposure to
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nonylphenol in soil caused an acute phytotoxicity response. The weight of L. sativa was
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reduced, and the EC50 (14 d) value for L. sativa was estimated to be 625 mg/kg soil42. Roberts
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et al.41 observed that nonylphenol caused inhibition to shoot height of wheat (T. aestivum) at
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10000 mg/kg soil, and rape (B. napus) at 1000 mg/kg dry soil. Domene et al.40 observed
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adverse effects of nonylphenol on germination and growth of bok choy (B. rapa) and
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perennial ryegrass (L. perenne). More recent studies found that these phytotoxicities of
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nonylphenol were associated with vacuolar fragmentation, a decrease in chlorophyll and root
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hair, an increase in oxidative stress, and damage to plasma membrane and chloroplasts43-45,47.
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Soil Algae Assay
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Exposure to nonylphenol affected both soil algae species. Both C. infusionum and C.
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reinhardtii showed inhibition of growth compared with controls, as shown in Figure 1C and
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1D. Based on solvent control normalized data, growth of C. infusionum was significantly
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stunted by nonylphenol at 200 and 300 mg/kg dry soil (p < 0.05). Exposure to nonylphenol
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significantly suppressed growth of C. reinhardtii at 800 and 1000 mg/kg dry soil (p < 0.05).
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Therefore, the growth-EC50 (6 d) values for soil algae C. infusionum and C. reinhardtii were
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calculated as 108 and 907 mg/kg dry soil, respectively. The growth-EC10 and NOEC values
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(6 d) for C. infusionum were estimated as 17 and 100 mg/kg dry soil, respectively. For C.
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reinhardtii, the growth-EC10 and NOEC values (6 d) were estimated as 449 and 600 mg/kg
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dry soil, respectively (Table 3). In this soil algal assay, C. infusionum appeared to be more
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susceptible to nonylphenol than did C. reinhardtii. A previous study also showed that C.
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infusionum was more sensitive than C. reinhardtii to silver nanoparticles48. These differences
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in sensitivity between the soil algae C. infusionum and C. reinhardtii are due to differences in
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cell walls49 and exopolymeric substances50. Previous studies evaluated the toxicity of
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nonylphenol to freshwater algae and confirmed that the toxicity was related to antioxidant
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responses15 and photosynthesis inhibition14,15. However, the mode of actions of effects of
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nonylphenol on soil algae must be investigated further, as no previous studies have evaluated
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the toxicity of nonylphenol on soil algae in soil media.
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Soil Nematode Assays
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In the soil nematode acute assays, adverse effects were observed in the control and
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solvent control groups after 24 h. The number of offspring in the controls (deionized water
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amended) and solvent controls (acetone amended) were 162 ± 23 and 158 ± 23, respectively.
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However, in the exposed groups, a nonylphenol concentration–dependent decrease in
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reproduction was observed. The number of offspring was significantly reduced starting at 100
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mg/kg dry soil of nonylphenol (Fig. 2A). The numbers of offspring were 133 ± 25, 95 ± 8, 12
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± 4, 4 ± 2, and 0.3 ± 0.5 at 10, 100, 500, 1000, and 2000 mg/kg dry soil, respectively.
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Reproduction-EC50 (24 h) value for C. elegans was calculated as 140 mg/kg dry soil (Table
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3).
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Our results are consistent with a previous study that reported a reduction in nematode
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reproduction in response to alkylphenol and organotin compounds51. Tominaga et al.51
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performed a multigenerational study and observed a decrease in reproduction and fecundity
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in C. elegans. The reduction in reproduction and fecundity in nematodes exposed to
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nonylphenol appeared to be associated with inhibition of estrogen binding52. In contrast, Höss
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et al.53 reported increases in growth and reproduction of C. elegans due to hormesis at lower
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concentrations of nonylphenol. In other previous studies on nematodes30, including early
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studies that used experiments with solution or sediment, nonylphenol exposure caused
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decreased survival54, changes in growth, reduced rates of reproduction, and reproductive
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system abnormalities51,53, as well as alterations in nematode species composition55.
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Earthworm Assays
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In the earthworm soil acute test (7 d of exposure), earthworms appeared to tolerate
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nonylphenol concentrations up to 1700 mg/kg dry soil, and no toxicological effects were
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observed in the control and solvent control groups. However, earthworm abnormalities,
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including mortality and mucous excretion, were observed at 1700 mg/kg dry soil of
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nonylphenol (Fig. 2B). The normalcy-EC50 (7 d) value for E. andrei was calculated as 1828
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mg/kg dry soil (Table 3). As shown in Tables 1 and 2, there have been some studies on
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toxicity or bioaccumulation of nonylphenol in earthworms (Dendrobaena octaedra, Eisenia
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fetida, and E. andrei) and white worms (Enchytraeus albidus and Enchytraeus crypticus) in
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soil media. In a previous study56, a lower survival-LC50 value (14 d, 308 mg/kg soil) was
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estimated for the earthworm D. octaedra, than we estimated using our data, because Jensen et
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al.56 tested soil with lower organic matter content and a longer exposure period. Hu et al.57
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observed no mortality and no changes in cellulose, ATPase, acetylcholinesterase, glutathione
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reductase, and glutathione content at nonylphenol concentrations of 10 mg/kg soil. However,
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significant damage to the DNA in coelomocytes of E. fetida was observed after 7 d exposure
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at 10 mg/kg soil. Domene et al.40 conducted a long-term exposure test and observed a
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decrease in reproductive activity of E. andrei after 56 d of exposure to nonylphenol. It was
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also reported that nonylphenol may accumulate in earthworms and white worms58-61.
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However, it remains unclear what the mode of action of nonylphenol is (for example, through
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endocrine-mediated responses in earthworm species).
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Collembola Assays
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Figure 3 shows acute and chronic effects of nonylphenol on two collembola species (L.
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sokamensis and F. candida). In the adult collembola L. sokamensis acute test, a 5-d exposure
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of adult L. sokamensis to nonylphenol did not induce mortality up to 250 mg/kg dry soil (Fig.
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3A). However, the number of surviving F. candida declined, starting at nonylphenol
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concentrations of 150 mg/kg dry soil and 14 d exposure (acute). In addition, reproduction of
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F. candida was affected, starting at nonylphenol concentrations of 150 mg/kg dry soil after
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28 d of exposure (chronic) (Fig. 3B). At concentrations of 200 mg/kg dry soil, very severe
287
effects of nonylphenol on F. candida were observed. The survival-EC50 (5 d) value for L.
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sokamensis and survival-EC50 (14 d) value for F. candida were calculated as > 250 and 123
289
mg/kg dry soil, respectively. The reproduction-EC10 and NOEC values (28 d) for F. candida
290
were estimated as 88 and 100 mg/kg dry soil, respectively (Table 3). Because the tested F.
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candida (juvenile) were younger than the L. sokamensis (adult), and the exposure duration of
292
F. candida (28 d) was longer than that of L. sokamensis (5 d), lower toxicity values were
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estimated for F. candida than for L. sokamensis.
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As shown in Tables 1 and 2, the collembola F. candida and F. fimetaria were used to
295
evaluate soil toxicity of nonylphenol. Like our study, previous studies reported that
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nonylphenol inhibited reproduction of collembola40,62-65 with similar ranges of reproduction-
297
EC10 values.
298 299
Estimation of the Soil Hazardous Concentration (HC) for Nonylphenol
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Based on soil organic matter normalized toxicity data, acute and chronic SSDs are
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described in Figure 4. Figure 4A shows acute SSD of nonylphenol using the compiled soil
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data from Tables 1 and 3. Figure 4B shows chronic SSD of nonylphenol using the compiled
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soil data from Tables 2 and 3. Total species data on 14 species from six taxonomic groups
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were used to determine acute SSD (Fig. 4A), and total species data on seven species from
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five taxonomic groups were used to determine chronic SSD (Fig. 4B). The soil alga C.
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infusionum and the collembola F. candida were the two species that were most sensitive to
307
nonylphenol. However, plant species such as wheat (T. aestivum) and mung bean (V. radiate)
308
tolerated nonylphenol in the soil tests, as shown in Fig. 4A and 4B, possibly because the
309
biomass of these plants was larger than that of soil algae and Collembola.
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Regarding HC values, acute and chronic HC for nonylphenol based on SSD were
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generated using two sources: (1) using only our data and (2) using compiled data (ours and
312
previous data) in order to estimate soil HC for nonylphenol using as much available data as
313
possible. Each estimated acute and chronic hazardous concentration (HC5, HC10, HC20, and
314
HC50) for soil ecosystem protection is shown in Table 4. Goodness of fit was also confirmed.
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As expected, chronic HC values were lower than acute HC values. According to our data, we
316
estimated HC values of 16, 29, 61, and 252 mg/kg soil for 95%, 90%, 80%, and 50% soil
317
species protection, respectively. In addition, we estimated HC values of 58, 93, 165, and 496
318
mg/kg soil for 95%, 90%, 80%, and 50% soil species protection, respectively, for acute
319
exposure to nonylphenol in soil.
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In conclusion, we propose that HC values from compiled data are more reliable than HC
321
values from our data alone because of the improved quantity and quality of the larger dataset.
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Previously, predicted no-effect-concentrations (PNEC) for bisphenol A66,67 and triclosan68,
323
which are known to be endocrine-disrupting chemicals, were suggested to protect terrestrial
324
species. Although there have been concerns regarding the release of nonylphenol in soil
325
environments, there are no studies evaluating hazardous concentrations of nonylphenol for
326
soil ecosystem protection.
327
We conducted a battery of bioassays using soil algae, invertebrates, and plants, and
328
then estimated HC values of nonylphenol based on probabilistic approach methods (species
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sensitivity distribution approach). The chronic HC5, HC10, HC20, and HC50 values were
330
estimated to be 16, 29, 61, and 252 mg/kg soil, respectively. The acute HC5, HC10, HC20, and
331
HC50 values were estimated to be 58, 93, 165, and 496 mg/kg soil, respectively.
332
Recently, the Canadian government established soil quality guidelines on nonylphenol
333
and its ethoxylates to protect environmental and human health69 as follows: 5.7 mg/kg dry
334
weight for agricultural, residential, and parkland soil, and 14 mg/kg mg/kg dry weight for
335
commercial and industrial soil. However, some countries (including Korea, Japan, USA,
336
Australia, and New Zealand) have no guideline values on nonylphenol in soil for ecosystem
337
protection. Therefore, we propose that the soil HC values for nonylphenol based on our SSDs
338
be used as fundamental data for conducting environmental risk assessments, and that the
339
results of the present study be applied to establish nonylphenol soil guideline values for soil
340
ecosystem protection.
341
There are some limitations in the present study. Future investigations should move
342
beyond taxonomically similar groups to include coexisting species in a community73 that
343
might be affected by nonylphenol concentrations in the soil environment.
344 345
ACKNOWLEDGMENTS
346
This research was supported by Basic Science Research Program through the National
347
Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT and future
348
planning (2016R1A2B3010445). This study was funded by the Korea Ministry of
349
Environment (MOE) as the Environmental Health Action Program (1485014458).
350 351
REFERENCES
352 353 354
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aquatic ecological communities and assessing impacts of chemical exposures. Integr. Environ. Assess. Manag. 2017. 13(4), 664-674.
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Table 1. List of acute endpoint values from previous studies. Each endpoint value was normalized using the soil organic matter value of 3.11%. Species sensibility distributions were created using the species geomean values. This table was adapted from Kwak et al.70. Test soil species
Test Soil organic duration matter (%) 21 d 3
Endpoints (mg/kg soil)
References
Survival
64
7d 14 s 14 d
1.4
Growth
Soil A: 2.9 Soil B: 8.9 Soil C: 7.96 Soil D: 5.61
Growth Soil A Growth Soil B Growth Soil C Growth Soil D Growth Soil A Growth Soil B Growth Soil C Growth Soil D Survival Reproduction Reproduction Weight
NOEC: 50 LOEC: 100 LC10: 67 LC50: 133 EC50: 559 EC50: 625 EC50: >10000 EC50: >10000 EC50: >10000 EC50: 9500 EC50: 800 EC50: 650 EC50: 1000 EC50: 3500 LC50: 308 EC50: 53 LOEC: 30 EC10: 739 EC50: 4012 EC10: 1386 EC50: 7501 EC10: 575 EC50: 1449 EC10: 696 EC50: 8159 OECD EC10: 664 EC50: 907 EC10: 24 EC50: 226 PRA EC10: 215 EC50: 316 EC10: 456 EC50: 640 UAB EC10: 227 EC50: 615 EC10: 197 EC50: 213 LC50: 90 EC50: 50 NOEC: 75 LOEC: 100 LC50: 124 NOEC: 2000
-
-
O. sativa
14-d
EC50
1793
-
1645 b
C. elegans
24-h
Shoot growth inhibition Shoot growth inhibition Reproduction
EC50
140
-
128 b
E. andrei
7-d
Normalcy
EC50
1828
(1779-1878)
1677 b
L. sokamensis
5-d
Survival
EC50
> 250
-
-
F. candida
14-d
Survival
EC50
123
(115-132)
113 b
C. infusionum
6-d
Growth inhibition
EC10
17
(2.7-37)
16 b
NOEC
100
-
92
EC10
449
(249-548)
412 b
NOEC
600
-
552
EC10
1822
-
1671 b
NOEC
> 2000
-
-
EC10
1433
(1315-1515)
1314 b
NOEC
1500
-
1376
EC10
88
(1.2-125)
81 b
NOEC
100
-
92
C. reinhardtii Chronic V. radiata
568 569 570
95% confidence interval
Normalized a value (mg/kg dry soil)
6-d 21-d
Growth inhibition Shoot growth inhibition
O. sativa
21-d
Shoot growth inhibition
F. candida
28-d
Reproduction
a
Soil organic matter normalized value. Soil organic matter normalized value = endpoint value × 3.11% / 3.39% b Values used for estimating species sensitivity distribution.
571
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Table 4. The acute and chronic hazardous concentrations for x% (HCx) were estimated based on species sensitivity distributions (SSD). HCx for nonylphenol based on SSD were derived from two sources: (1) using only our data and (2) using compiled data (ours and previous data), using the USEPA SSD generator (ver. 1.0)37. We estimated the HCx with lower and upper prediction intervals based on the compiled data from the current and previous studies. HCx represents the concentration that protects (100 - x) % of species in an ecosystem.
578 Data from the present study
Compiled data from this study and previous research
Acute
Chronic
Acute
Chronic
HC5
31 (4–242)
9 (1–66)
58 (28–118)
16 (6–42)
HC10
53 (8–363)
18 (3–119)
93 (46–186)
29 (11–73)
HC20
105 (18–617)
45 (8–254)
165 (84–323)
61 (25–145)
HC50
386 (74–2000)
259 (52–1289)
496 (258–953)
252 (110–576)
(mg/kg soil)
579 580
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Fig. 1. Shoot growth inhibition of (A) mung bean (Vigna radiate) and (B) rice (Oryza sativa). Biomass inhibition of (C) soil algae (Chlorococcum infusionum) and (D) soil algae (Chlamydomonos reinhardtii) due to nonylphenol exposure. (A–B) Plants were exposed to nonylphenol in soil for 14 days (acute) and 21 days (chronic). (C–D) Soil algae were exposed to nonylphenol in soil for 6 days. The asterisks (*) indicate significant differences from the solvent control (p < 0.05).
589 590 591 592 593
Fig. 2. Inhibition of (A) reproduction of soil nematode Caenorhabditis elegans and (B) normalcy of earthworm Eisenia andrei after nonylphenol exposure. (A) Nematodes were exposed to nonylphenol in soil for 21 hours (acute). (B) Earthworms were exposed to nonylphenol in soil for 7 days (acute). The asterisks (*) indicate significant differences from solvent control (p < 0.05).
594 595 596 597 598 599
Fig. 3. Inhibition of (A) survival of Collembola Lobella sokamensis and (B) survival (acute) and reproduction (chronic) of Collembola Folsomia candida exposed to nonylphenol in soil. (A) L. sokamensis was exposed to nonylphenol in soil for 5 days (acute). Adult L. sokamensis tolerated nonylphenol well. (B) F. candida was exposed to nonylphenol in soil for 14 days (acute) and 28 days (chronic). The asterisks (*) indicate significant differences from solvent control (p < 0.05).
600 601 602 603 604 605 606 607 608 609
Fig. 4. Species sensitivity distributions (SSDs) of nonylphenol based on the soil toxicity data. (A) Acute SSD of nonylphenol using the compiled data from Tables 1 and 3. A total of 14 species from six taxonomic groups were used, and acute HCx values (Table 4) were estimated based on acute SSD. Blue dots indicate data from the present study, and green dots indicate data from previous studies. (B) Chronic SSD of nonylphenol using the compiled data from Tables 2 and 3. A total of seven species from five taxonomic groups were used, and chronic HCx values (Table 4) were estimated based on chronic SSD. Red dots indicate data from the present study and yellow dots indicate data from previous studies. The 95% confidence interval is shown by a dashed line around the curve.
Figure Captions
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Figure 1.
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Figure 2.
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Figure 3.
619
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Figure 4.
621
622
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