Environmental Toxicology and Chemistry, Vol. 25, No. 1, pp. 265–271, 2006 2006 SETAC Printed in the USA .

00 0730-7268/06 $12.00

Department of Environmental Science and Engineering, International Environmental Research Center, Gwangju Institute of Science and Technology, 1 Oryong-dong, Buk-gu, Gwangju 500-712, Korea ( Received 17 March 2005; Accepted 16 June 2005) Abstract—The overall ecotoxicological effect of pharmaceutically active compounds (PhACs) detected in the effluents of Korean wastewater treatment plants (WWTPs) to Daphnia magna was investigated using biological and chemical analyses. The bioassay results showed median lethal concentrations and no-observed-effect concentrations ranging from a few to tens of ppm levels for nine PhACs in 48-h acute and 21-d chronic tests. The mixture effects of pharmaceuticals also were examined by other acute and chronic tests, which showed no significant toxicity despite a slightly increased combined effect of approximately twofold. The residual concentrations of nine PhACs were analyzed at concentrations ranging from 10 ng/L to 89 g/L in the influents and from 10 ng/L to 11 g/L in the effluents from four metropolitan cities in South Korea between January and November of 2004. Through repeated investigations of the influents and the effluents from different WWTPs, relatively higher removal efficiencies (23.9–91.3%) compared with those of previous surveys performed in other countries were observed for most pharmaceuticals, with the exception of acetaminophen (8.7%). The present study showed no significant risk effects of the effluents from WWTPs containing pharmaceuticals (i.e., hazard quotient 1), even at the 95th percentile contamination range, although a risk assessment factor of 1,000 was applied. Therefore, it can be concluded that the potential risk of pharmaceuticals should be monitored carefully with more bioassay data, because many uncertainties still exist in the determination and toxicity of metabolites in water environments. No significant risk was observed, however, from the selected PhACs in the effluents from WWTPs discharged into surface waters. Keywords—Pharmaceuticals Wastewater treatment plants Toxicity Risk assessment


Although large quantities of pharmaceuticals are produced and used as animal and human medicines, little attention has been paid to the potential risks of pharmaceuticals as toxic contaminants in aquatic environments. Pharmaceuticals have similar physicochemical behaviors and cause biological effects toward nontarget organisms in the environment (i.e., human and animals) even at low concentrations [1,2]. Recent studies have demonstrated that some metabolites are more lipophilic and more persistent than the original drugs from which they were derived [3]. These substances are excreted or discarded from both humans and domestic animals as a mixture of metabolites and unchanged forms via the municipal sewage system to rivers, bank filtrates, and groundwaters. This implies that their parent compounds and metabolites may exist in drinking water through several drug-fate pathways. The balances of the influents and effluents of drug residues detected in wastewater treatment plants (WWTPs) reveal that many pharmaceuticals are not eliminated completely by traditional treatment processes (i.e., coagulation, flocculation, and sedimentation) [4]. Recently, major efforts to clarify the ecological influence and occurrence of pharmaceuticals in the aquatic environment have been attempted in North American and European countries [1,3,5,6]. Diclofenac, ibuprofen, and clofibric acid are medical agents used widely in both human and domestic animal practices. These pharmaceuticals have been detected frequently at concentrations up to several ng/L or g/L levels in
* To whom correspondence may be addressed (sdkim@gist.ac.kr). 265

aquatic environments [7–13]. Many metabolites, such as hydroxy- and carboxy-ibuprofen and carboxy-hydrotrophic acid for ibuprofen [11,12] and 2-(4-chlorophenoxy)-2-methyl propionic acid for clofibric acid [3], have been found in raw sewage water and rivers. Ibuprofen and its metabolites are degraded efficiently by biological treatment in WWTPs, whereas diclofenac and clofibric acid are degraded only minimally by biological reaction [3,5,7]. More than 90% of the diclofenac in Lake Greifensee, Switzerland, was removed by photolytic degradation [7]. Some of the other substances detected in rivers include antipyretics (i.e., acetaminophen [5,6]), -sympathomimetics [14], antibiotics (e.g., erythromycin [15,16]), as well as antiepileptics (e.g., carbamazepine [12,17,18]) and hypolipidemics (i.e., gemfibrozil [5,6,19]). Despite many studies, it remains unclear whether the exposure of aquatic biota to these pharmaceuticals have direct or indirect effects on the environments. Furthermore, no data are available from Korea, which uses large quantities of drugs because of the high population densities and expanding sizes of its cities. The objectives of the present study were to monitor the extent of drug residues in Korean WWTPs and to investigate the ecological risks of target compounds. First, the acute and chronic toxicity of pharmaceuticals, both singularly and as mixtures, were evaluated for Daphnia magna. Using ecotoxicological results, environmental risk assessments were performed for drug residues in Korean rivers receiving waters from WWTP effluents to predict their potential adverse effects.

Sample collection Samples were collected between January and November of 2004 using precleaned, amber-glass containers during periods


Environ. Toxicol. Chem. 25, 2006
Table 1. Overview of pharmaceutically active compounds

G.H. Han et al.


CASa no.

Molecular weight

Molecular structure

Therapeutic use




Nonsteroidal anti-inflammatory




Nonsteroidal anti-inflammatory

Clofibric acid



Lipid regulator (active metabolite)





Salicylic acid



Analgesic, antipyretic




Lipid regulator




Analgesic antipyretic





Tolfenamic acid



Nonsteroidal anti-inflammatory



Chemical Abstracts Service.

of normal WWTP operation. Big cities, including Seoul, Busan, Gwangju, and Daegu, were selected as study areas because of their high population densities, which were assumed to have exposure to large amounts of pharmaceuticals to their aquatic environments because of the discharge of effluents. Treatments at all WWTPs consisted of three main steps: Preliminary clarification, final clarification, and an aeration tank. Gwangju2 had additional treatment steps for phosphate removal and denitrification treatment. The WWTP in Busan also was equipped with sand filtration, followed by an ultraviolet disinfection step, after the final clarification. Water samples were stored in an icebox for their return to the laboratory, adjusted to pH less than 2 by the addition of concentrated HCl, and preserved at a temperature less than 4 C before analysis. Samples were filtered through glass-fiber filters (0.45 m) and sucked through solid-phase extraction cartridges containing 500 mg of Sep-Pak Vac 6cc C18 (Waters, Milford, MA, USA) at a rate of approximately 8 ml/min. Cartridges were dried for 2 h under a moderate stream of nitrogen and then extracted by elution with three successive 1-ml aliquots of high-performance liquid chromatography (HPLC)–grade methanol [20]. After elution, the samples were evaporated to a final 1-ml volume.

Analytical procedure
All solutions of pharmaceutically active compounds (PhACs) were prepared in HPLC-grade methanol, purchased from Sigma-Aldrich Chemical (St. Louis, MO, USA), for analysis. Chemicals were analyzed by gas chromatography–mass

spectrometry (GC-MS; Shimadzu, Kyoto, Japan) in full-scan or selected ion-monitoring mode. Separations were carried out using a DB5-type capillary column (length, 30 m; inner diameter, 0.25 mm; film thickness, 0.25 m) with helium (99.999%) as the carrier gas (flow rate, 1.1 ml/min) and the flow velocity set to 38.1 cm/s in the constant-flow mode. Injections were performed in the split mode (purge time, 2 min) with an injection volume of 1 l. The GC oven was programmed as recommended by Ternes et al. [21]. The methyl esters of the analytes were quantified by monitoring the ions at m/z (mass to charge ratio) 214 and 242 for diclofenac; m/z 161 for ibuprofen; m/z 128 for clofibric acid; m/z 80, 109, and 151 for acetaminophen; m/z 95, 165, and 193 for carbamazepine; m/z 92, 120, and 138 for salicylic acid; and m/z 122 and 107 for gemfibrozil. All samples were analyzed by five-point calibration over the whole procedure. Using this method, limits of detection of 10 and 250 ng/L were obtained for the sewage influents and effluents, respectively. The target pharmaceuticals used in the bioassay experiments were selected with respect to their occurrence, potential ecological effect, and broad application (Table 1). Individual stock solutions of each target pharmaceutical were prepared by dissolving in ethanol for 1 d before use for each experiment, with the concentration of the solvent in the medium, including the control groups, being less than 1%.

Test organisms and culture
The test organism used in the present study, D. magna, was obtained from the Korea Institute of Toxicology (Daejeon,

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South Korea). Daphnia magna was cultured with Selenastrum capricornutum and a mixture of yeast, CEROPHYLL (Ward’s, Rochester, NY, USA), and trout chow (YCT) supplied as food. The organisms were cultured and handled according to the procedures outlined in the U.S. Environmental Protection Agency (U.S. EPA) manual [22]. Neonates of D. magna less than 24 h old were used for acute and chronic tests, which were illuminated with a 16:8-h light:dark photoperiod. Organisms were not fed during the test periods. Reconstituted 10 4 M; CaSO4, 6.98 10 4 M; hard water (MgSO4, 9.98 3 4 10 M; and KCl, 1.07 10 M), with a NaHCO3, 2.28 hardness of 170 5 mg/L (mean SD), an alkalinity of 110 5 mg/L as CaCO3 mg/L, and pH 7.8 0.2 at 25 C throughout the study.

ber, were assigned for each treatment under the same test conditions as used for a single compound.

Statistical analysis
The endpoints of the toxicity tests using D. magna are based on adverse effects on survival and reproduction. The LC50s for the acute tests were calculated using regression analysis following probit analysis. The Dunnett method of pair-wise separation was used to determine the NOEC in the 21-d chronic test. The experimental response used in the statistical analysis was the total number of neonates produced until the end of the experiment or at the time of death. An animal that died before reproducing neonates was included in the analysis, with zero entered as the number of young produced [23].

Single-compound bioassay tests
For 48-h static nonrenewal acute toxicity tests, D. magna was exposed to a control and eight different concentrations of PhACs. Four replicates, containing five organisms per 30-ml test chamber, were assigned to the treatments and control. Test organisms were fed YCT and S. capricornutum for 2 h before the test [22]. Mortality was recorded 48 h later. The 21-d chronic effect of each pharmaceutical was assessed on the reproduction of D. magna in a static renewal test according to the U.S. EPA guidelines [23]. The exposure concentrations to D. magna in 21-d chronic tests ranged from 0.1 to 80 mg/L based on the results of the acute toxicity tests for nine different concentrations and one control. Five replicates, containing one organism per test chamber, were assigned to each concentration. Organisms were fed every 2 d with YCT and S. capricornutum, with all test solutions renewed with fresh culture medium every 2 d. The number of neonates reproduced and the survival of D. magna were counted and recorded on each of the 21 d.

Risk assessment
Ecological risk assessments were performed to characterize the degree of contamination and to evaluate the adverse effects of these chemicals in real aquatic environments [24]. In the present study, results from the D. magna toxicity tests were compared to those from the GC-MS analyses of selected compounds in the influents and effluents of the WWTPs. The hazard quotient (HQ) was calculated to evaluate the degree of risks for each target PhAC, which was mathematically formulated as follows: HQ EC (Exposure concentration) TRV

where TRV represents the toxicity reference value (i.e., LC50 or NOEC). A HQ value of less than 1 when the assessment factor was applied indicates an insignificant risk. In the present study, we employed a quite moderate assessment factor of 1,000, as recommended by the European Union [25] and the Organization for Economic Co-operation and Development [26].

Mixture bioassay tests
To elucidate the combined effects of the pharmaceutical mixtures to which the aquatic ecosystems were exposed, the toxic unit (TU) concept was used as follows: TU TU concentration (mg/L) LC50 concentration (mg/L) NOEC (acute test) (chronic test)

Bioassay tests Single-compound toxicity. The mortality (LC50) and reproduction inhibition (NOEC) were measured in D. magna exposed to nine pharmaceuticals. The results indicated that most pharmaceuticals tested were toxic and harmful to aquatic organisms (Table 2). Generally, toxicity levels of chemicals are classified as follows: very toxic to aquatic organisms for less than 1 mg/L, toxic to aquatic organisms for 1 to 10 mg/L, and harmful to aquatic organisms for 10 to 100 mg/L [25]. Comparing the measured and predicted LC50s using the Ecological Structure–Activity Relationships (ECOSAR) model from the website of the U.S. EPA [1], the toxicity values showed relatively significant differences for high-lipophilic compounds, including diclofenac, ibuprofen, and tolfenamic acid, than for the hydrophilic compounds. For diclofenac, Sanderson et al. [1] used an extremely low octanol–water partition coefficient (e.g., log Kow of 0.7; in the present study, log Kow of 4.51 was used), which caused a large difference in the toxicity value. To investigate the relationship of toxicity with respect to hydrophobicity, the LC50 versus log Kow values were plotted, as shown in Figure 1. This figure indicated a linear relationship between the toxicity values and hydrophobicities (r2 0.79). That is, the LC50s decreased (i.e., toxicity increases) with increasing log Kow of the pharmaceuticals. However, the result for acetaminophen was exceptional, showing high toxicity for an extremely low log Kow of 0.27.

where LC50 is the single-substance concentration causing 50% mortality when applied alone and NOEC is the no-observedeffect concentration. The total combined effect can be calculated by summation of each TU. For mixture acute toxicity tests, the methodology of equitoxic addition of chemicals tested was employed. Daphnia magna was exposed to the control and five TUs, ranging from 0.18 to 1.05, in 48-h static nonrenewal tests. For example, to prepare 0.6 TU of a three-chemical mixture, 0.2 TU for each chemical (diclofenac, ibuprofen, and clofibric acid) was combined. Four replicates, containing five organisms per test chamber, were assigned to the treatments and the control. Other experimental procedures, such as illumination, feeding regime, and endpoint measurement, were the same as those for a single compound. For the mixture chronic toxicity test, D. magna was exposed to less than 1 TU for the five treatments and one control for 21 d. Ten replicates, containing one organism per test cham-


Environ. Toxicol. Chem. 25, 2006
Table 2. Toxicity values of pharmaceuticals to Daphnia magna in 48-h acute and 21-d chronic tests Literature review Chemicals Diclofenac Ibuprofen Clofibric acid Carbamazepine Salicylic acid Gemfibrozil Acetaminophen Bezafibrate Tolfenamic acid
a b c

G.H. Han et al.

Log Kowa 4.51 3.97 2.57 2.45 2.26 4.77 0.27 4.25 5.17

LC50b (mg/L) 80.1 132.6 141.2 111 111.7 10.4 20.1 30.3 7.4

Hazard classc Harmful Nontoxic Nontoxic Nontoxic Nontoxic Harmful Harmful Harmful Toxic

NOECd (mg/L) 10 20 40 — — — — — —

LC50e 5057.0 38.0 293.0 111.0 59.0 6.0 42.0 25.0 1.7

NOEC — 3f — — — — — — —

Values from Howard and Meylan [33] and Sanderson et al. [1]. LC50 median effective concentration. Classified in European Union Directive 93/67/EEC (European Economic Communities) [25] in the basis of LC50 values. d NOEC no-observable-effect concentration. e Values from Sanderson et al. [1] calculated by the structure–activity relationship model using Ecological Structure–activity relationships (ECOSAR). f Value from Halling-Sorensen et al. [28].

In the 21-d chronic toxicity tests, the concentration–response curves for the single substances were sigmoid but had different sharpness and shapes. When D. magna was exposed to 80 mg/L of each PhAC, the mortality rates were 100%. All compounds showed good reproduction rates at 10 mg/L. The observed mortality rate with diclofenac was 100% at the relatively low concentration of 40 mg/L. The single chronic toxicity results indicated that diclofenac was the most toxic pharmaceutical in the present study. The mortality by the end of the experiment never exceeded 10% for the controls. The NOECs were 10, 20, and 40 mg/L for diclofenac, ibuprofen, and clofibric acid, respectively. Compared with the acute toxicities, the NOECs were not low, as shown by the acute to chronic ratio (LC50:NOEC), which ranged from 3.5 to 8. This implies that the tested chemicals (i.e., diclofenac, ibuprofen, and clofibric acid) show no serious long-term effects toward aquatic ecosystems [27,28]. The LC50s, even the chronic values (NOECs), were quite high for single compounds, in the range 10 to 100 mg/L (not

toxic but harmful to aquatic organisms). These values were 5,000- to 100,000-fold higher than the median concentrations detected in aquatic environments [5]. Accordingly, considering the presence of low concentrations of these pharmaceutical residues in real environments, such as rivers and wastewater effluents containing various PhACs within the range 0.1 to 10 g/L, we can conclude that the acute effects, or even chronic effects (NOECs), of single compounds are negligible. Mixture toxicity. To evaluate the combined effect of mixtures, toxicity tests were carried out on D. magna with a combination of the three drugs (diclofenac, ibuprofen, and clofibric acid). The observed and calculated toxicities, based on the TU concept, were compared (Fig. 2). At greater than 0.5 TU, the TU observed in bioassay experiment showed approximately twofold higher toxicity values than the TU predicted by calculation. For example, addition of the mixture concentration corresponding to 1.0 TU (i.e., 50% mortality in calculation) for three pharmaceuticals resulted in 2.0 TU (i.e., an observed 100% mortality) to D. magna. The measured toxicity values

Fig. 1. The relationship between the toxicity (median lethal concentration [LC50]) and hydrophobicity (log Kow) for nine pharmaceuticals in the Daphnia magna 48-h acute toxicity tests. The solid line represents the linear regression of the data (r2 0.79). The closed circle indicates the data for acetaminophen.

Fig. 2. Comparison of the observed and calculated percentage mortality based on the toxic unit (TU) in 48-h acute mixture toxicity tests to Daphnia magna. The dashed line shows a 1:1 slope. The observed TU was determined from the percentage mortality results, with the calculated TU indicating the sum of the TUs of individual pharmaceuticals based on their median lethal concentrations.

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Occurrence of pharmaceuticals from WWTP effluents
Grab samples of the influents and effluents at five WWTPs in metropolitan cities in South Korea were collected between January and November of 2004 (Table 3). Recovery of the pharmaceuticals ranged from 72 to 89%, with standard devi3) between 5 and 20%, following spiking of ations (1 , n the water with 1 g/L of each compound. The present results showed that residual concentrations of seven PhACs were detected at concentrations ranging from 0.01 to 88.99 g/L in the influents and from 0.01 to 10.96 g/L in the effluents. Salicylic acid, ranging between 0.12 and 88.99 g/L, had the highest detected concentrations in the present study. Gemfibrozil appeared at nonquantifiable levels in the multiple samples from all sampling sites because of the low limit of detection. The results in Table 3 also show highly fluctuating removal rates, between 8.7 to 91.3%, indicating that the pharmaceuticals in WWTPs may not be eliminated effectively by the use of traditional treatment processes (i.e., activated sludge, coagulation, flocculation, and sedimentation) at WWTPs [4]. Low removal rates were found for several PhACs in repeated observations. Very low removal rates of 8.7 and 23.9% were observed for acetaminophen and diclofenac, respectively, in all sampling areas, whereas that for carbamazepine in the conventional treatment system, which discharged into surface waters at a concentration as low as 0.16 g/L, was 91.3%. This was a notable result compared to that in another study [5], which showed a very low removal rate (7%) for carbamazepine in WWTPs. However, through analytical investigations on the influents and effluents from different WWTPs in South Korea, relatively high removal efficiencies were observed for most pharmaceuticals, with the exception of acetaminophen, although no notable differences were found between WWTPs compared to results of previous surveys in other countries [3,8]. This may have been caused by differences in the elimination rates of the pharmaceutical residues because of sorption and biodegradation. Meanwhile, treatment technologies, such as oxidation with chlorine and ozone, activated carbon, and membrane filtration, have been demonstrated as effective methods for the elimination of antibiotics [29] and some pharmaceuticals [21,30]. Figure 4 shows the effluent concentration profiles for seven pharmaceuticals following treatment at the WWTPs in Korean metropolitan cities (Gwangju1, Gwangju2, Busan, and Daegu). Most residual PhAC concentrations were in the range 0.01 to 0.4 g/L, including approximately 40% not-detectable points.

Fig. 3. Comparison of the observed and calculated percentage inhibitions, based on the toxic unit (TU), in 21-d chronic mixture toxicity tests to Daphnia magna. The dashed line shows a 1:1 slope. The observed TU was determined from the relative inhibition value corresponding to the no-observed-effect concentration (NOEC) from the mixture tests, with the calculated TU indicating the sum of the TUs of individual pharmaceuticals based on their NOECs.

for the mixture were somewhat underestimated compared with those predicted [27]. In the 21-d chronic mixture tests, the inhibited reproduction rate was used as the endpoint, which can be determined by the ratio of the number of neonates in various mixture series of the three pharmaceuticals to those in the control. Figure 3 shows the expected and observed toxicities toward D. magna of the diclofenac, ibuprofen, and clofibric acid mixtures. The present results illustrate that the measured chronic toxicities for the mixtures were greater than we predicted. In statistical analysis, the observed TU had high standard deviations over all the ranges tested. Overall, the measured toxicities of the mixtures of the three pharmaceuticals in the present study were not significantly different from the predicted toxicities for either the acute or chronic toxicities, although a twofold increase in the combined effect of PhACs on D. magna was observed. Note, however, that the result of the mixture toxicity could be serious if the combined toxic effects of unknown metabolized chemicals and large number of PhACs in real water environments are considered.

Table 3. Concentrations ( g/L) of selected pharmaceuticals in wastewater treatment plant influents and effluents from South Korea during the 2004 survey Influent ( g/L) Pharmaceuticals Diclofenac Ibuprofen Clofibric acid Carbamazepine Salicylic acid Gemfibrozil Acetaminophen
a b

Effluent ( g/L) LOD 14 14 17 9 15 0 14

LOD 0.01 0.01 0.01 0.05 0.1 0.5 0.01


REC 1 (%) 85 79 89 75 73 83 72 7 10 5 15 20 9 18


Samples (n) 17 17 17 16 16 16 16

Mean (max, min) 2.59 (9.87, ND)d 0.30 (0.58, ND) 1.51 (4.38, 0.03) 1.84 (9.42, ND) 23.92 (88.99, 0.12) ND (ND, ND) 0.07 (0.26, ND)


Mean (max, min) 1.97 (10.96, ND) 0.07 (0.31, ND) 0.31 (0.74, ND) 0.16 (0.97, ND) 2.43 (6.73, ND) ND (ND, ND) 0.06 (0.16, ND)


LODc 12 12 15 8 13 0 12

Removal rate (%) 23.9 77.8 79.5 91.3 89.8 ND 8.7

LOD limit of detection. Recoveries (REC) and standard deviation (1 , n 3) of pharmaceuticals, following spiking of the water with 1 c Number of samples having values greater than the LOD. d ND not detectable.

g/L of each compound.


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G.H. Han et al.

Fig. 4. Comparative mean concentration ( g/L) profiles for the target pharmaceuticals in the effluents from metropolitan cities (Gwangju, Busan, and Daegu) in South Korea during 2004. An asterisk indicates no detectable concentration. The data were less than the detection limits in Seoul (South Korea).

Fig. 5. Graphical illustration of the calculated hazard quotients (HQs) for the selected pharmaceuticals in the wastewater treatment plants effluents. A HQ of 1 indicates the critical value to determine a significant risk of pharmaceuticals. A acetaminophen; Carb carbamazepine; Clof clofibric acid; D diclofenac; G gemfibrozil; I ibuprofen; S salicylic acid.

However, several hundredfold higher concentrations were observed for salicylic acid, up to 2.2 and 6.7 g/L at Gwangju1 and Busan, respectively. These high concentrations in the effluents might affect ecosystems because of the large volumes of water discharged daily into the nearby river. Interestingly, none of the PhACs mentioned above were detected in the effluents at the Seoul site (not shown in graph). Although only one sample was taken from Seoul, the results were unexpected, considering Seoul is the largest and most industrialized city in Korea. Many pharmaceuticals containing polar functional groups are thermally stable and do not readily lend themselves to GC analysis. Therefore, most GC analysis of polar pharmaceuticals must incorporate a derivatization step to overcome this limitation [30], which makes the sample preparation laborious and time-consuming, increases the probability of contamination and experimental errors, and leads to degradation of labile compounds. In addition, pharmaceuticals are easily changed to their metabolites by glucuronidation [31] or photodegradation [32], although considerable amounts of their unchanged forms are excreted via urine or feces. Therefore, further measurements and identification of pharmaceuticals and their metabolites in WWTP effluents and rivers will be required to evaluate the contamination and occurrence of drug residues in surface waters.

Risk assessment in the aquatic environment
The toxicity index (i.e., HQ) was calculated by comparing the maximum measured concentrations of field data collected from the WWTPs with the acute toxicity values to D. magna. Figure 5 shows the effect of WWTP effluents that contained pharmaceuticals was not a significant risk, even at the 95th percentile contamination range, although a risk assessment factor of 1,000, as recommended by the European Union and the Organization for Economic Co-operation and Development, was applied. For salicylic acid and diclofenac, a difference of less than one order of magnitude between maximum environmental and effect concentrations was represented by the HQ values. None of the HQ values of the drugs tested in the present study exceeded 1. On average, the HQ values of the pharmaceuticals measured in the WWTP effluents were within the

ranges of 10 1 to 10 4 when an application factor of 1,000 was applied. From our results, it may be concluded that no risks to D. magna were caused by the pharmaceuticals in Korean WWTP effluents. Risk assessments also were conducted by applying a factor of 100 for the chronic effect on the basis of NOECs for the three pharmaceuticals (diclofenac, ibuprofen, and clofibric acid). The present results indicated that the three pharmaceuticals had no significant ecotoxicological impact on long-term chronic exposure (not shown in graph). Moreover, pharmaceuticals may be of less risk if the dilution, because of the mixing of effluents and the receiving stream, is considered. Many uncertainties, however, exist in risk assessments, especially those attributed to the difficulties in identifying the presence of pharmaceuticals and their quantification in water discharged from WWTPs containing a mixture of chemicals. Currently, a method (i.e., extraction and derivatization) used for the detection of pharmaceuticals in WWTP effluents with lower detection limits is being developed. Metabolites can be one of the concerns in the ecological impact of pharmaceuticals; therefore, an analytical method for the detection of their forms should be developed for more accurate risk assessments. In addition, more toxicological data for various pharmaceuticals must be obtained from bioassay experiments using aquatic species, such as algae, daphnids, and fish. Sanderson et al. [1] demonstrated that the susceptibility of aquatic species to pharmaceuticals were in the following order: Algae, daphnids, and fish. According to their results, approximately 10% of pharmaceuticals in the environment had a HQ greater than 1 when multiplied by an application factor of 1,000. Consequently, the potential risk of pharmaceuticals should be monitored carefully, with more bioassay data, although no risk was found in the WWTP effluents from large Korean cities.

The overall ecotoxicological effects of PhACs detected in WWTP effluents were investigated by biological and chemical analyses. The bioassay results showed LC50s and NOECs ranging from levels of several ppm to several tens of ppm in the acute and chronic tests, respectively, for the selected PhACs. The combined toxicity of pharmaceuticals showed

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slight synergistic effects in both the acute and chronic tests. These bioassay results would be valuable in risk assessments with respect to pharmaceutical residues in surface waters, taking into account the insufficient toxicological data regarding pharmaceuticals. The concentrations of PhACs in discharged effluents indicated highly variable distributions depending on the site, although relatively high removal efficiencies for most pharmaceuticals (except for acetaminophen) were observed in Korean WWTPs compared to those from other countries. The PhAC residues in the effluents might affect aquatic ecosystems because of the large volumes of water discharged daily into the nearby river. Finally, the potential risk from the pharmaceuticals detected in the effluents from Korean WWTPs indicated a low biological risk (HQ 1) to aquatic ecosystems when a risk assessment factor of 1,000 was applied to the tests on D. magna. The monitored effluent concentrations did not reach critically harmful levels. It was concluded that no significant risk from the selected pharmaceuticals existed in Korean WWTPs, despite the presence of uncertainty that should be characterized in the determination and toxicity of metabolites.

13. 14. 15. 16. 17.




Acknowledgement—We acknowledge the financial support of the Ministry of Environment as The Eco-technopia 21 project through the Korea Institute of Environmental Science and Technology (KIEST) and a grant (code 4-1-2) from the Sustainable Water Resources Research Center of 21st Century Frontier Research Program. We also thank Dong-il Shimadzu Corporation for technical and financial support.
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