Acute Toxicity and Ecological Risk Assessment of Benzophenone and N,N-Diethyl-3 Methylbenzamide in Personal Care Products

Benzophenone (BP) and N,N-diethyl-3-methylbenzamide (DEET) are two chemicals often used in personal care products (PCPs). There is a lack of systematic ecotoxicological evaluations about the two chemicals to aquatic organisms. In the present study, the acute toxic effects on Chlorella vulgaris, Daphnia Magana, and Brachydanio rerio were tested and the ecotoxicological risks were evaluated. For BP, the 96-h half-maximal effective concentration (EC50) on C. vulgaris was 6.86 mg/L; the 24-h median lethal concentration (LC50) on D. magana was 7.63 mg/L; the 96-h LC50 on B. rerio was 14.73 mg/L. For DEET, those were 270.72 mg/L, 40.74 mg/L, and 109.67 mg/L, respectively. The mixture toxicity of BP and DEET, on C. vulgaris, D. magana, and B. rerio all showed an additive effect. The induced predicted no-effect concentrations (PNECs) for BP and DEET by assessment factor (AF) method are 0.003 mg/L and 0.407 mg/L, respectively. Both are lower than the concentrations detected from environment at present, verifying that BP and DEET are low-risk chemicals to the environment.


Introduction
Personal care products (PCPs) are one of the groups in emerging pollutants and are integral to people's activities of daily life. Pedrouzo et al. [1] sub-classified PCPs into six categories: organic ultraviolet (UV) filters, antimicrobials, preservatives, musk fragrances, insect repellents, and siloxanes. These compounds are not usually considered in routine environmental monitoring, but are becoming of environmental concern because of their increasingly common use and the potential risk to human and ecosystem health. With the improvement in detection techniques, an increasing number of PCPs are being detected from different aquatic environments [2]. A study [3] also indicated that 2-Hydroxy-4-methoxybenzophenone (BP-3), one ingredient of PCPs, has been detected from the B. rerio (AB strains) were ordered from Nanjing YSY Biotech Company Ltd. (Nanjing, China). The average body length was 2.60 ± 0.20 cm, average body weight was 0.33 ± 0.06 g, and potassium dichromate was used to test the sensitivity of B. rerio before the toxicity experiments. The experiments were carried out at: 25 ± 2 • C, pH 7.0-8.0, 12 h:12 h light-dark cycle.

Ninety-Six Hours Acute Toxicity Experiments with C. vulgaris
During the acute toxicity experiments on C. vulgaris, BG 11 liquid medium and BP or DEET stock solution were used to prepare a serial concentration of solution (100 mL) in a glass flask (250 mL). The initial density of C. vulgaris was 10 6 cells/mL, and the experiment lasted 96 h. Each concentration test, including cosolvent and blank control groups, was designed with three parallel samples. Before the experiment, microscopic counting and spectrophotometry were employed to obtain the linear correlation between the C. vulgaris cells density and absorbance at 680-nm wavelength. During the period of the experiment, the absorbance of C. vulgaris was measured every 24 h, then it was converted to cell density to calculate growth rates.

Twenty-Four Hours Acute Toxicity Experiments with D. magna
Aerated tap water was used to prepare a serial concentration of BP or DEET solution. Ten D. Magna were put in a 100-mL glass beaker with 80 mL of solution. The experiments lasted 24 h. Each concentration test including cosolvent and blank control groups were designed with three parallel samples. The numbers of the dead individuals were recorded at the end of the test.

Ninety-Six Hours Acute Toxicity Experiments with B. rerio
Aerated tap water was used to prepare a serial concentration of BP or DEET solution. Ten fish were put in a 5000-mL glass beaker with 4000 mL of solution. The experiments lasted 96 h. Each concentration test including cosolvent and blank control groups were designed with three parallel samples. The numbers of the dead individuals were recorded every 24 h.

The Mixed Toxicity Experiment
The mixed toxicity experiments of BP and DEET were performed in the same method as the independent toxicity experiments. The ratios of BP and DEET used for C. vulgaris, D. magna, and B. rerio were different. All ratios of BP and DEET were randomly prepared, referred to the obtained EC 50 or LC 50 values of each independent chemical, in order to imitate different conditions in the water.
Stock solutions were prepared for C. vulgaris (7.50 mg/L BPand 276.65 mg/L DEET), D. magna (50.10 mg/L BP and 347.20 mg/L DEET), and B. rerio (50.10 mg/L BP and 690.30 mg/L DEET). All stock solutions were set up as 100%; then, a series of solutions at different percentages were made for the test. When the EC 50 or LC 50 results are obtained as a percentage, the toxic units (TU) of BP and DEET can be calculated, respectively, by the following: where C i was the concentration of i in the testing solution at the EC 50 of the mixture. TUi of BP + the TUi of DEET = the total TU of the mixture. If the total TU value at the endpoint of the mixture equaled 1, the toxicity was assumed to be additive; if the total TU value at the endpoint of the mixture was >1, the toxicity was assumed to have an antagonistic effect; if the total TU value at the endpoint of the mixture was <1, the toxicity was deemed to have a synergistic effect.

Statistical Analysis
OriginPro 9 software (OriginLab, Northampton, MA, USA) is used to draw graphs depicting the experimental results. IBM SPSS version 20 software (IBM, Armonk, NY, USA) was applied to calculate the 96-h half-maximal effective concentration (EC 50 ) of C. vulgaris, the 24-h median lethal concentration (LC 50 ) of D. magna, and 96-h LC 50 of B. rerio.

Ecological Risk Assessment
AIST-MeRAM (National institute of Advanced Industrial Science and Technology-Multi-Purpose Ecological Risk Assessment and Management Tool, Japan, English ver. URL: http://en-meram.aistriss.jp/) was employed to assess the ecological risk of the two chemicals. Two methods-assessment factor (AF) and species sensitivity distribution (SSD)-were used to deduce the predicted no-effect concentration (PNEC).

Results
There was no significant difference between the dimethyl sulfoxide control group at the highest concentration and the blank control group for C. vulgaris growth rate, D. magna lethal rate, and B. rerio lethal rate by analysis of variance. This indicated that dimethyl sulfoxide had no repression effect on C. vulgaris, D. magna, and B. rerio in this study. All the results were analyzed by comparing the testing to the blank control groups.
In the present study, the chemical concentrations were set up based on the results of the pre-test, so the concentration ranges are narrow and have good linear relation to the inhibition or mortality rates.

C. vulgaris Growth Inhibition Test of BP
The inhibition effect on C. vulgaris growth increased with the increasing in the concentrations. During the exposure time, although the algal growth rate was inhibited, the algal densities still showed an increasing trend. When the BP concentration was 10.00 mg/L, there was almost no growth between 48 and 72 h, but rapid growth from 72 to 96 h.
The growth inhibition by BP concentration is shown in Figure 1A. The linear regression equation between BP concentration and its inhibition ratio was y = −10.216 + 8.979x, (x : 2.00 mg/L ∼ 10.00 mg/L), where x is the BP concentration, and y is the inhibition ratio on C. vulgaris growth (%). Using SPSS version 20 software, the 96-h EC 50 was calculated as 6.86 mg/L (95% confidence interval: 6.40-7.34 mg/L). This value was classified as high-level toxicity on C. vulgaris [15].
where x is the BP concentration, and y is the mortality of D. magna (%).

Acute Toxicity of BP to D. magna
When the concentrations range from 2.00 to 12.00 mg/L, the relationship between the mortality of D. magna and BP concentration was described by a linear equation: y = −10.802 + 7.989x, (x : 2.00 mg/L ∼ 12.00 mg/L), where x is the BP concentration, and y is the mortality of D. magna (%). The 24-h LC 50 of D. magna for BP was 7.63 mg/L (95% confidence interval: 7.13-8.14 mg/L), the value was also classified as high-level toxicity [15]. The acute toxicity of BP to D. magna is shown in Figure 1B where x is the BP concentration, and y is the mortality of B. rerio (%). The acute toxicity of BP to B. rerio is shown in Figure 1C. The 96-h LC 50 of B. rerio for BP was 14.73 mg/L (95% confidence interval: 14.11-15.39 mg/L), and this value was classified as medium-level toxicity [15].

C. vulgaris Growth Inhibition Test of DEET
When the concentrations of DEET were between 79.04 and 395.20 mg/L, there was an obvious inhibition effect on C. vulgaris. With the increasing in the concentrations, the inhibition effect also increased. The C. vulgaris growth inhibition ratio by DEET concentration is shown in Figure 2A. The linear regression equation used was as follows: where x is the DEET concentration, and y is the inhibition ratio on C. vulgaris (%).

Acute Toxicity of BP to D. magna
When the concentrations range from 2.00 to 12.00 mg/L, the relationship between the mortality of D. magna and BP concentration was described by a linear equation: where x is the BP concentration, and y is the mortality of D. magna (%).
The 24-h LC50 of D. magna for BP was 7.63 mg/L (95% confidence interval: 7.13-8.14 mg/L), the value was also classified as high-level toxicity [15]. The acute toxicity of BP to D. magna is shown in Figure 1B where x is the BP concentration, and y is the mortality of B. rerio (%). The acute toxicity of BP to B. rerio is shown in Figure 1C. The 96-h LC50 of B. rerio for BP was 14.73 mg/L (95% confidence interval: 14.11-15.39 mg/L), and this value was classified as medium-level toxicity [15].

C. vulgaris Growth Inhibition Test of DEET
When the concentrations of DEET were between 79.04 and 395.20 mg/L, there was an obvious inhibition effect on C. vulgaris. With the increasing in the concentrations, the inhibition effect also increased. The C. vulgaris growth inhibition ratio by DEET concentration is shown in Figure 2A where x is the DEET concentration, and y is the inhibition ratio on C. vulgaris (%). The 96-h EC 50 was calculated as 270.72 mg/L (95% confidence interval: 250.40-292.50 mg/L) and showed low-level toxicity on C. vulgaris [15].

Acute Toxicity of DEET to D. magna
The mortality of D. magna by DEET concentration was described by a linear equation: y = −16.160 + 1.605x, (x : 19.76 mg/L ∼ 59.28 mg/L), (6) where x is the DEET concentration, and y is the mortality to D. magna (%).
The 24-h LC 50 for DEET was 40.74 mg/L (95% confidence interval: 37.86-44.25 mg/L), classified as medium-level toxicity [15]. The acute toxicity to D. magna of DEET is shown in Figure 2B where x is the DEET concentration, and y is the mortality to B. rerio (%). The acute toxicity to B. rerio of DEET is shown in Figure 2C. The 96-h LC 50 of DEET was 109.67 mg/L (95% confidence interval: 106.14-141.15 mg/L) and was classified as low-level toxicity [15].

Acute Toxicity of the Mixture to C. vulgaris
Based on the EC 50 values of the independent toxicity experiments, the concentration of the mixture was calculated as a percentage of the total concentration (%). The relationship between the concentration of the mixture and the growth inhibition ratio of C. vulgaris is shown in Figure 3A. There was a good linear relationship between the concentration of the mixture and the growth inhibition ratio of C. vulgaris. The 96-h EC 50 of the mixture of BP and DEET on C. vulgaris was 46.31% (95% confidence interval: 42.04%-51.21%). In this case, the concentration of BP in the mixture was 3.43 mg/L, and the concentration of DEET was 128.44 mg/L. By calculating the TU values of BP and DEET using Equation (1), respectively, the total TU value at this point of the mixture equaled 1. Therefore, the toxicity of BP plus DEET combination showed an additive effect [16]. The 96-h EC50 was calculated as 270.72 mg/L (95% confidence interval: 250.40-292.50 mg/L) and showed low-level toxicity on C. vulgaris [15].

Acute Toxicity of DEET to D. magna
The mortality of D. magna by DEET concentration was described by a linear equation: y = −16.160 + 1.605x, (x: 19.76 mg/L~59.28 mg/L), (6) where x is the DEET concentration, and y is the mortality to D. magna (%). The 24-h LC50 for DEET was 40.74 mg/L (95% confidence interval: 37.86-44.25 mg/L), classified as medium-level toxicity [15]. The acute toxicity to D. magna of DEET is shown in Figure 2B where x is the DEET concentration, and y is the mortality to B. rerio (%).
The acute toxicity to B. rerio of DEET is shown in Figure 2C. The 96-h LC50 of DEET was 109.67 mg/L (95% confidence interval: 106.14-141.15 mg/L) and was classified as low-level toxicity [15].

Acute Toxicity of the Mixture to C. vulgaris
Based on the EC50 values of the independent toxicity experiments, the concentration of the mixture was calculated as a percentage of the total concentration (%). The relationship between the concentration of the mixture and the growth inhibition ratio of C. vulgaris is shown in Figure  3A. There was a good linear relationship between the concentration of the mixture and the growth inhibition ratio of C. vulgaris. The 96-h EC50 of the mixture of BP and DEET on C. vulgaris was 46.31% (95% confidence interval: 42.04%-51.21%). In this case, the concentration of BP in the mixture was 3.43 mg/L, and the concentration of DEET was 128.44 mg/L. By calculating the TU values of BP and DEET using Equation (1), respectively, the total TU value at this point of the mixture equaled 1. Therefore, the toxicity of BP plus DEET combination showed an additive effect [16].  The mortality increased with the increasing in the concentration of mixture and showed a dose-response relationship. The acute toxicity of the mixture to D. magna is shown in Figure 3B.
The 24-h LC 50 of the mixture of BP and DEET to D. magna was 6.61% (95% confidence interval: 6.11%-7.15%), the concentration of BP in the mixture was 3.31 mg/L, and the concentration of DEET was 22.95 mg/L. The total TU value at this point of the mixture equaled 1, so the BP plus DEET combination showed an additive effect [16].

Acute Toxicity of the Mixture to B. rerio
The acute toxicity of the mixture to B. rerio is shown in Figure 3C. The mortality increased with the increasing in the concentrations and showed a dose-response relationship.
The 96-h LC 50 of the mixture of BP and DEET to B. rerio was 9.59% (95% confidence interval: 8.87%-10.36%), the concentration of BP in the mixture was 4.80 mg/L, and the concentration of DEET was 66.20 mg/L. The total TU value at this point of the mixture equaled 1, so the BP plus DEET combination showed an additive effect [16].

Assessment of the Acute Toxicity of BP and DEET
Two methods-the AF method and the SSD method-were used to deduce the PNEC values in the present study. The AF method uses the lowest toxicity data of fish, crustaceans including daphnids, and algae to characterize risk as a margin of exposure (MOE). This is a point risk estimate model for screening or initial risk assessment. The AF is divided by an uncertainty factor (UF) to deduce the PNEC value. The UF is different in different countries and organizations. In this study, the UF was selected as 100, which is recommended by the OECD.
The SSD method uses all of the available toxicity data to describe the differences in species sensitivity to chemicals as a statistical distribution. The HC5 (the concentration at which 5% of the species are affected) is derived from the dose-response curve of species sensitivity distribution. This is a quantitative and probabilistic risk estimate model.
In the present study, UF was suggested as 5 by the producer of the RAM-MeAIST, and the log-normal distribution (LND) estimation method was selected in the assessing results.

Assessment of the Ecotoxicological Effect of BP
The values of 96-h EC 50 of C. vulgaris, the 24-h LC 50 of D. magna, and 96-h LC 50 of B. rerio were applied to AIST-MeRAM. The deduced PNEC value by the AF method was 0.003 mg/L (using the uncertainty factor as 100), and was 0.480 mg/L by SSD (using the uncertainty factor as 5). The results including the former registered data are summarized in Table 1.

Assessment of the Ecotoxicological Effect of DEET
The values of the 96-h EC 50 of C. vulgaris, the 24-h LC 50 of D. magna, and 96-h LC 50 of B. rerio were applied to AIST-MeRAM. The deduced PNEC value by the AF method was 0.407 mg/L (using the uncertainty factor as 100) and was 8.494 mg/L by the SSD method (using the uncertainty factor as 5). The results, including the former studies, are given in Table 2.

Assessment of the PNEC Values by AF and SSD
AF and SSD were employed in the present study to induce the PNEC. The values from SSD are 1-2 orders of magnitude higher than those from the AF method. To obtain reasonable results, at least 8 data (from different species or different tests) are needed for the SSD method, as shown in Tables 1 and 2, there are 22 acute data from 8 species for BP, and 13 acute data from 9 species for DEET. Although the data of BP or DEET are more than 8, the data diversity and numbers were not enough for providing reasonable and trustworthy results, so the deduced value by SSD may underestimate the PNEC values. Therefore, the PNEC values deduced from the AF method are recommended and applied in the discussion for assessing the ecological risk of BP and DEET. We do not deny that, when the value from the AF method is applied as a reference to make criteria for protecting ecosystem, it may overestimate. However, from the point of protecting the ecosystem at the greatest extent, the PNEC value deduced by the AF method is more reasonable when there is no perfect evaluation system at present.
In the present study, the PNEC values of both chemicals were deduced by with and without the present data, respectively, the results show different characteristics. For BP, the PNEC values deduced by the AF method are the same values with or without the present data, while the results by the SSD method are a little different: 0.480 mg/L with present data and 0.443 mg/L without present data, respectively. For DEET, the PNEC values deduced by the AF method are 0.407 and 0.712 with and without the present data, respectively. The results by the SSD method are 8.494 mg/L with present data, but 10.00 mg/L without present data. Although it could not be concluded that these results increased or decreased the PNEC values, it is definitely that the present study enriched the database for the ecological assessment of the two chemicals and further improved the rationality of the assessment conclusion, for the principle of the assessment method is: the more data you provide, the more refined your assessment will be.

The Acute Toxicity and Ecological Risk of BP
According to the acute toxicity results of BP, the 96-h EC 50 for C. vulgaris was 6.86 mg/L, and the 24-h LC 50 for D. magna was 7.63 mg/L. Both were judged as high-level toxicity. The 96-h LC 50 for B. rerio was 14.73 mg/L, which was judged as medium-level toxicity. The sensitivity of C. vulgaris was higher than that of D. magna and B. rerio. To our knowledge, the present study is the first time to use C. vulgaris and B. rerio as organisms to test the acute toxicity of BP. The results provide new species and data for evaluation of the toxicity of BP.
Among all the acute toxicity data of BP registered in AIST-MeRAM, the 72-h EC 50 of Pseudokirchneriella subcapitata (algae) was 3.50 mg/L, which was more sensitive than C. vulgaris in the present study. There were 4 data about the LC 50 of D. magna; one was lower than 1.00 mg/L, the last three were all from 1.00 to 10.00 mg/L (7.60 mg/L, 10.00 mg/L, and 7.63 mg/L), which were similar to the present results. The 96-h LC 50 of Oryzias latipes (fish) was 10.00 mg/L, which is a little lower than the results in the present study. There were 10 data about the LC 50 of Pimephales promelas (fish), 9 of which were between 14.20 and 15.30 mg/L, which is very close to the 96-h LC 50 value (14.73 mg/L) of B. rerio in the present study. These results indicated that the 96-h LC 50 of BP tested by B. rerio was reasonable as compared with the results obtained by other species of fish. For the different trophic levels of biota exposed to BP, the sensitivities were different. Algae were more sensitive than fish. Algae are primary producers in the aquatic trophic web, the harmful effect on algae may affect the stability and function of the ecosystem, so the ecological risk of BP should be fully realized.
According to the assessment results by the AF method, the PNEC value of BP was 0.003 mg/L. It has been reported that the concentration detected in the Zhujiang River in China are 0.10-5.30 ng/L [17], and this value is much lower than the PNEC value. Similarly, the concentrations of the most commonly used UV filters, 4-MBC, BP-3, BP-4, and EHMC measured in coastal seawater samples from Spain ranged between 35 and 164 ng/L [18]. These facts indicated that BP could still be considered as a low environmental risk chemical at present. However, BPs have been detected in urine samples from individuals in the United States and China [3]. Concentrations of BP-3 (one of the derivatives of BP) have been measured at 9.97 mg/L in children and 15.7 mg/L in adults in United States; in China, the concentration of 0.62 mg/L in children and 0.98 mg/L in adults have been recorded. In Europe, concentrations of 4-MBC (4-methylbenzylidene camphor) and OC (octocrylene), two widely used UV filters, were determined in the muscle tissue of fish (brown trout, Salmo trutta fario) from seven small Swiss rivers, which all receiving inputs from wastewater treatment plants [19]. These indicated that the chemicals were transported into human and wild organism bodies through ecological processes and may have a potential health risk. BP has been proved as a type of endocrine disrupter, when it reaches an effective concentration, may cause damage to human health. However, the path of acquisition and how it affects the biota in the ecosystem still remain unknown.
The results in the present study provided most recent ecotoxicological research for the BP. The acute toxic effects on tree typical organisms in the aquatic ecosystem were tested; further, the PNEC were deduced to evaluate the risk to environment. The results can provide references for environmental criteria. Because BPs are widely used in many personal care products in our everyday life, its potential hazard to aquatic organisms and to humans should be further studied.

The Acute Toxicity and Ecological Risk of DEET
In the acute toxicity results of DEET, the 96-h EC 50 to C. vulgaris was 270.72 mg/L, the 24-h LC 50 to D. magna was 40.74 mg/L, and the 96-h LC 50 to B. rerio was 109.67 mg/L. The acute toxicity of D. magna was judged as medium-level toxicity, while the acute toxicity of C. vulgaris and B. rerio were judged as low-level toxicity. The sensitivity of D. magna was higher than that of C. vulgaris and B. rerio. To our knowledge, this is the first time to use C. vulgaris and B. rerio to test the acute toxicity of DEET. The results provided new data for the ecotoxicological assessment of DEET.
Among all the data registered in the AIST-MeRAM about the acute toxicity of DEET, the 72-h EC 50 to Pseudokirchneriella subcapitata (algae) was 100.00 mg/L, which was more sensitive than 96-h EC 50 of C. vulgaris. There were three data of the acute toxicity of DEET to D. magna: 75.00 mg/L, 74.00 mg/L, and 40.74 mg/L. The present result (40.74 mg/L) is similar with these former studies, which indicated the rationality of the present result. There were two data of the LC 50 of DEET to Pimephales promelas (fish)-both are 110.00 mg/L-and two data about the LC 50 of DEET to Gambusia affinis (fish)-both are 235.00 mg/L. For Oncorhynchus mykiss (fish), the LC 50 were 71.25 and 75.00 mg/L, respectively. There was only one LC 50 of DEET to Oryzias latipes (fish) at 100.00 mg/L. The 96-h LC 50 of B. rerio in this study is very close to all of the data reported on fish. According to the results for DEET described above, the sensitivity order is Oncorhynchus mykiss > Daphnia magna > Oryzias latipes = Pseudokirchneriella subcapitata > Pimephales promelas > Gambusia affinis > Chlorella vulgaris. Overall, the sensitivity of daphnids and fish was higher than that of algae. This can be explained that DEET is used as an insect repellent and there were corresponding target mechanisms in daphnids and fish.
The value of PNEC deduced by using the AF method is 0.407 mg/L. In the published papers, the concentrations of DEET detected in the aquatic environment ranged from 0.60 to 1.20 ug/L [20], which is lower than the PNEC value. Therefore, at present, DEET could be classified as chemicals having low toxicity and risk to the environment. To our knowledge, there are no reports about DEET in the human body; however, as DEET is used as repellents in some perfume, the human body cannot avoid being exposed to DEET. Furthermore, as a widely used ingredient in PCPs, it can be discharged to the aquatic environment so that they can reach humans directly or indirectly by processes of enrichment, accumulation, and amplification through the food web. The study of the acute toxicity of DEET can provide a scientific basis for criteria and thus for management of the exposure concentration and the discharge standard.

The Assessment of the Mixing Toxicity of BP and DEET
The mixture toxicity of BP and DEET on C. vulgaris, D. magna, and B. rerio in the present study all showed an additive effect. The results explained these two chemicals' behavior in mixture conditions. Since BP and DEET have different structural compositions, their biological degradability pathways and degradation products may be different, so the interaction of the two chemicals afterwards in an actual environment may behave differently from the results in a laboratory. Therefore, the mixture of their degradation products may perform different interactions such as antagonism, additive effects, irrelevant effects, and synergy.
The results of the present study provided the acute toxic responses of three tropical organisms in the aquatic ecosystem to BP, DEET, and their mixture. The acute toxic data enriched the tested species and acute testing results for the database, thereafter increasing the rationality of the induced PNEC. The PNEC induced from this study evaluated the ecological risk of both chemicals. These will provide a theoretical basis for establishing water quality criteria.
In the future study, attention should be paid to the so-called lower risk, but may be potential disrupt chemicals. Further chronic toxic test and ecosystem level studies should also be carried to evaluate the physiological and developing effects on biota.

Conclusions
The acute toxic effects of BP and DEET on C. vulgaris, D. magna, and B. rerio were tested. The toxicity of BP to C. vulgaris and D. magna were judged as high-level toxicity, but that to B. rerio was judged as medium-level toxicity. For DEET, the acute toxicity of C. vulgaris and B. rerio were judged as low-level toxicity, and the acute toxicity of D. magna was judged as medium-level toxicity. The mixing toxicity of BP and DEET on C. vulgaris, D. magna, and B. rerio displayed an additive effect. The results explained the way of these two chemicals' behavior in mixture conditions. The risk assessment to the environment by the AF method concluded that both chemicals are a low risk to the environment at present. The results in the present study indicated the diversity and complexity of the toxic effect of BP and DEET in PCPs.
Since both BP and DEET are main components in personal care products, they are therefore high exposure sources to humans. BP has been identified as endocrine disruptors [8], and their potential high risk to ecosystems and human health over a long time period should be made widely known. The assessment of the acute toxicity of BP and DEET in the present study clarified the different toxic levels and different sensitivities from different tropical organisms in the aquatic ecosystem, and the results indicated the diversity and complexity of the toxic effect of chemicals in PCPs. Although the BP and DEET are evaluated as low-risk chemicals in the aquatic ecosystem, the identified characteristic as an endocrine disruptor [8] implied the high potential risk of the PCPs over a long time period.