Environmental Research 135 (2014) 253–261

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Chemical pollution and toxicity of water samples from stream receiving leachate from controlled municipal solid waste (MSW) landfill A. Melnyk a,n, K. Kuklińska a, L. Wolska a,b, J. Namieśnik a a

Department of Analytical Chemistry, Chemical Faculty, Gdansk University of Technology (GUT), 11/12 G. Narutowicza Street, 80-233 Gdansk, Poland Medical University of Gdansk, Faculty of Health Sciences with Subfaculty of Nursing, Department of Environmental Toxicology, Dębowa Street 3, 80-204 Gdańsk, Poland


art ic l e i nf o

a b s t r a c t

Article history: Received 26 June 2014 Received in revised form 18 August 2014 Accepted 9 September 2014

The present study was aimed to determine the impact of municipal waste landfill on the pollution level of surface waters, and to investigate whether the choice and number of physical and chemical parameters monitored are sufficient for determining the actual risk related to bioavailability and mobility of contaminants. In 2007–2012, water samples were collected from the stream flowing through the site at two sampling locations, i.e. before the stream's entry to the landfill, and at the stream outlet from the landfill. The impact of leachate on the quality of stream water was observed in all samples. In 2007–2010, high values of TOC and conductivity in samples collected down the stream from the landfill were observed; the toxicity of these samples was much greater than that of samples collected up the stream from the landfill. In 2010–2012, a significant decrease of conductivity and TOC was observed, which may be related to the modernization of the landfill. Three tests were used to evaluate the toxicity of sampled water. As a novelty the application of Phytotoxkit F™ for determining water toxicity should be considered. Microtoxs showed the lowest sensitivity of evaluating the toxicity of water samples, while Phytotoxkit F™ showed the highest. High mortality rates of Thamnocephalus platyurus in Thamnotoxkit F™ test can be caused by high conductivity, high concentration of TOC or the presence of compounds which are not accounted for in the water quality monitoring program. & 2014 Elsevier Inc. All rights reserved.

Keywords: Landfill leachate Water pollution Toxicity tests

1. Introduction An inherent human function is the generation of waste, therefore the precise waste utilization and management is one of the most important issues in environmental protection. The dominant method of waste management in Poland is disposal. A typical landfill may be a source of three phases of waste products: solid (wastes), liquid (leachate) and gaseous (landfill gas), which may pose a threat to the individual elements of the environment, including fresh water resources (Kjeldsen et al., 2002; Lisk, 1991). The list of landfill sites that are known to be leaching contaminants into underlying aquifers is still growing (Reinhard et al., 1984). The degree of water contamination risk depends on the construction of the site, the type and level of fragmentation of the waste, the method of storage, as well as hydro-geological and


Corresponding author. E-mail address: [email protected] (A. Melnyk).

http://dx.doi.org/10.1016/j.envres.2014.09.010 0013-9351/& 2014 Elsevier Inc. All rights reserved.

hydrographic conditions at the landfill location (Rapti-Caputo et al., 2006). Old landfills, often unlined, were very frequently localized in wetlands and near watercourses (Ford et al., 2011; Lorah et al., 2009), so that ammonia (NH3) (Camargo and Alonso, 2006), xenobiotic organic compounds (XOCs) (Baun et al., 2004), dissolved organic carbon (DOC), nutrients (Diaz, 2001) and heavy metals (Christensen et al., 2001) penetrated into water. Some of these substances are toxic, persistent and likely to bioaccumulate therefore their emissions, discharges and losses have to be eliminated. As a result, decisions about the modernization of old landfills are made and new landfills are designed in accordance with the system of best available technology (BAT). Assessing the actual impact of functioning landfills on the quality of surface waters is not an easy task. A variety of waste deposited in landfills cause the penetration of various substances, that are not subject of the periodic analytical studies or are not covered by continuous monitoring, into surface and ground waters. As a result, the presence of these compounds in the environment may go unnoticed for a long time, and the biological


A. Melnyk et al. / Environmental Research 135 (2014) 253–261

effects resulting from the exposure of organisms to these pollutants may be irreparable. Still too little is known about the health effects due to exposure to complex mixtures such as those often found in landfill leachate. In many countries, including Poland, the monitoring of the surface water quality is still based on physical and chemical analyses, and the legislation does not require toxicity evaluation. As a consequence, the ecological quality of waters in Poland is unknown (Drobniewska et al., 2007). The two primary aims of this study were to determine the actual impact of a functioning landfill on the water quality in the stream flowing through it, and to evaluate usefulness of a toxicity classification system based on a battery of microbiotests in the context of routine monitoring of surface waters. In order to achieve a realistic estimation of the hazard associated with the contaminants derived from the landfill, the values of parameters which are the overall measure of the level of pollution and certain concentration levels of individual substances being monitored have been supplemented by the results of the ecotoxicological studies. The ecotoxicologal studies allow determining the presence of compounds which have toxic properties and have not been identified during monitoring studies. The results obtained during the toxicological bioassays, combined with studies on physicochemical parameters, can be used to help determine the potential impact of pollutants from landfills on both humans and other living organisms (Dumont and Shultz, 1980). The physical, chemical and toxicological investigations of water samples from the stream flowing through the landfill, collected up and down the stream from the landfill, will allow for optimal elimination of other factors affecting the quality of tested water as well as for determining the impact of the landfill on the surface water quality. The use of a battery of biotests allows determining the magnitude of adverse effects due to landfill leachate-induced water pollution in relation to live organisms from different trophic levels, and thus assessing human impact on the natural environment as a result of the currently used methods of waste management.

Fig. 1. Hydrogeographical conditions in the study area, and the location of water sampling sites. In November 2007, the construction of pumping station at the Kozacki reservoir was completed and the station became operational. This allowed the transfer of water surplus from the reservoir to the stream via a pipeline. The implementation of the aforementioned construction project was aimed to prevent the inflow of stream water under the landfill. In July 2008, works aimed at inhibiting the infiltration process of stream water under the landfill, i.e. pumping out the water from the reservoir and lining its bottom with geomembrane, were completed. 2.2. Water sample collection Surface water samples were collected quarterly during the two time periods, namely, from the beginning of the fourth quarter of 2007 until the first quarter of 2009, and from the beginning of the fourth quarter of 2010 until the first quarter of 2012 in accordance with the guidelines contained in PN-EN 5667-6:2003. Samples were collected into 1 L amber bottles with Teflon-lined caps. Bottles were completely filled with water with no headspace left. The samples were processed immediately after arrival to the laboratory or they were stored at 4 1C until all assays were performed. Samples were not filtered before the toxicity assessment.

2. Material and methods

2.3. Physicochemical analysis

2.1. Study area

The collected water samples were subjected to physicochemical analysis. These measurements were performed by the accredited analytical laboratory which conducts periodic surveys of water quality. The scope of the investigation included:

The subjects of research were water samples taken from the stream flowing through the municipal solid waste landfill in Gdansk (Poland). The landfill has regularly been exploited since 1973 (Małaczyński et al., 2002). It consist of three landfilling areas:

 sector 800/1–12.2 ha (period of landfilling: 2010 till now);  sector 800/2–12.2 ha (period of landfilling: 1973–2009);  sector 800/3–8.8 ha (reserve of land). The bottom liner of sector 800/1 is 56–60 cm of loam/bentomat/2 mm of PEHD. The sector 800/2 does not have a bottom liner. Over the last 20 years, the landfill was subjected to a systematic modernization program. At present, it occupies an area of about 70 ha, and its core business is focused on the disposal of municipal solid waste, mainly by storage. In 2010 leachate treatment plant has been launched (Andrzejewska et al., 2006). An average annual tonnage of accepted waste is as follows:


210  109 g of municipal waste; 50  109 g of industrial waste; 20  109 g of rubble; 600  106 g of waste undergoing biodegradation processes, pyrolysis, or stored in burials.

The Kozacki stream flows through the center of the landfill site (Fig. 1). Rain and ground water collect at the eastern boundary of the site (WP2). On the western side, the surface reservoir for septic purposes, so-called Kozacki reservoir, was created (WP1). The stream has been channelized within the landfill area in order to protect the surface waters against the negative impact of the landfill. However, due to corrosion and cyclical loads, the channelization deteriorated and the stream water has been contaminated with landfill leachates for nearly a quarter of a century.

 determination of pH and conductivity;  determination of heavy metal content, i.e. Pb, Cd, Cr(VI), Cu, Zn and Hg contents;

 determination of Total Organic Carbon (TOC);  determination of the total amount of polycyclic aromatic hydrocarbons (PAHs). The aforementioned parameters are considered to be indicators of water quality as set out in the relevant Polish legislation. The measurements of pH and conductivity were carried out during sampling by means of potentiometric and conductometric methods, respectively. The concentrations of heavy metals in samples were determined by using a Flame Atomic Absorption Spectrometry (FAAS) method according to PN-ISO 8288:2002 method A standard, while the content of PAHs was determined by a liquid chromatography with fluorescence detection (HPLC-FLD) technique according to PAF/PB-07 (5th edition of 16.10.2012 r.) standard. Each analysis was performed in triplicate. 2.4. Toxicity assessment The toxicity estimation was performed with the use of a battery of biotests. Three species belonging to different trophic levels in the food chain were used as indicator organisms, as follows:

 Producers: grass Sorghum saccharatum (Phytotoxkit F™ test);  Consumers: crustacean Thamnocephalus platyurus (Thamnotoxkit F™ test);  Decomposers: bacterium Vibrio fischeri (Microtoxs test). Parameters such as turbidity, color, pH and temperature may interfere with the test results thus they were checked and controlled before every test.

9.80 7 10.91 8.717 12.11

7.1–7.9 459–1133 4–43 0.3–6 3–9 0.019–0.029 n.d. n.d. 2.5–31.7 n.d. 7.62 7 0.20 454 7 104 11.5 7 7.6 2.22 7 1.23 5.83 7 3.49 0.0137 0.0067

7.3–7.9 286–584 4–24 0.3–4 2–9 0.007–0.024 n.d.c n.d. 1.2–33.2 – 0.4 5867 – – 2 0.01 – – 114.72 –


Toxicity assessment with Microtoxs test was carried out according to the PN-EN ISO 11348-3:2008 standard using a Microtox model 500 instrument (ISO, 2008). The pH of the samples was measured to ensure it was ranged between 6 and 8. Number of dilutions prepared were 4 and dilution factor was 2. Test time was 30 min. Calculations were performed using the MicrotoxOmni program. The measured parameter was the bacterial luminescence inhibition (%effect). For toxic samples EC50 was determined. The test was performed in triplicate for each water sample. Thamnotoxkit FTM test was carried out in accordance with the procedure (Thamnotoxkit FTM, 1995). Cysts were incubated for 22 h at 25 1C under continuous illumination. 10 larvae of T. platyurus were added to each well containing 1 ml of water sample. The test was performed in triplicate for each water sample. Multiwell plates were incubated at 25 1C in darkness for 24 h. The measured parameter was the mortality of test organisms. The application of Phytotoxkit F™ for determining water toxicity should be considered a novelty. Originally, the test has been designed to assess toxicity in soil samples. In the present study, the test was modified by using a layer of cotton wool (100% pure cotton) as a substratum mimicking the soil. The cotton substratum was soaked in sample water (50 mL), and then covered with a filter paper on which 10 seeds of S. saccharatum were placed. The test was performed in triplicate for each water sample. The measurements of root growth were carried out after a 72-h incubation at 25 1C using the Image Tool 3.0 program for Windows (UTHSCSA, San Antonio, USA). The control sample consisted of a layer of cotton wool wetted with 50 mL of spring water of the same composition as previously described for Thamnotoxkit F™ test.

0.2 221 6 0.83 1.84 0.01 – – 1.09 –

Rangea WP2 Mean 7sdb Rangea WP1 Difference between WP1 and WP2 mean values

IV qrt. 2010 – I qtr. 2012

Mean 7 sdb

7.42 70.31 6757 247 17.5 7 13.4 3.05 7 2.01 7.67 72.34 0.023 7 0.0043

Difference between WP1 and WP2 mean values

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2.5. Statistical analysis Statistical analysis was performed using the R-3.0.1. program for Windows. Pearson rank correlation analysis was performed, in order to investigate significant relationships among results of toxicity tests and physicochemical parameters. Twoway ANOVA statistical analysis was done to compare the physicochemical parameters and toxicity tests at two sampling locations during different periods. The independent variables were sampling locations (WP1 and WP2) and sampling periods (first from 2007 to 2009 and second from 2010 to 2012). The dependent variables were values of selected parameters. The p-Value shows the significance of the F ratio with a confidence level α of 0.05.

122.5 7 38.6

3.1. Physicochemical parameters and metal content The values of chemical parameters determined in this study are listed in Table 1. The two-way ANOVA results of each physicochemical parameter are summarized in Table 2. The F/Fc value indicate the extent to which the F value is significantly greater than the critical F (Fc) values. Where the F/Fc value is below 1, there is no interaction between two studied factors and the higher the F/

7.1–7.8 3325–7806 4 0.3 2–18 0.05–0.11 0.010–0.011 n.d. 73–186 n.d.

Table 2 Two-way ANOVA results for physicochemical parameters and metal content in surface water samples.

7.78 7 2.19



Mean values based on 3 replicates. n¼ 6. n.d., under quantification limit (LOQ ¼ 0.01 mg/L).



7.95 7 0.38 464 7 127 47 0 0.3 70 4.337 5.72 0.05 70 0.0102 70.00041

pH Conductivity Pb Cd Cu Zn Cr(VI) Hg TOC ∑PAHs

Sampling point

– [μS/cm] (20 1C) [μg/L] [μg/L] [μg/L] [mg/L] [mg/L] [μg/L] [mg/L] [μg/L]

Range WP1

7.4–8.4 279–617 4 0.3 2–16 0.05 0.010–0.011 n.d. 4.4–10.3 n.d.

Mean7 sd

b a

IV qrt. 2007 – I qtr. 2009 Date of collection

Table 1 Physicochemical parameters and metal content in surface water samples.

Range WP2


Mean 7sd


7.55 7 0.24 63317 1597 47 0 0.3 7 0 6.337 6.98 0.06 7 0.02 0.01027 0.00041

3. Results

pH Conductivity Pb Cd Cu Zn Cr Hg TOC ∑PAHs a

– [μS/cm] (20 1C) [μg/L] [μg/L] [μg/L] [mg/L] [mg/L] [μg/L] [mg/L] [μg/L]



Location–period interaction

F/Fcb p





1.53 19.4 0.21 0.17 0.20 0.88 0.08e n.c. 10.5 n.c.

0.91 16.8 2.57 5.38 0.11 8.81 n.c.f n.c. 9.79 n.c.

0.061 0.000 0.003 0.000 0.493 0.000 – – 0.000 –

0.26 16.7 0.21 0.17 0.00d 0.00 n.c. n.c. 10.1 n.c.

0.300 0.000 0.350 0.398 0.968 0.915 – – 0.000 –

0.018 0.000c 0.350 0.398 0.357 0.064 0.549 – 0.000 –

First period: IV qrt. 2007 –I qtr. 2009, second period: IV qrt. 2010–I qtr. 2012. F/Fc, F value to critical F value ratio. Less than 0.001. d Less than 0.01. e Value calculated using one-way ANOVA for WP1 and WP1 for only first period; f n.c., not calculated. b c


A. Melnyk et al. / Environmental Research 135 (2014) 253–261

Fc value (and if the p-Value is equal or less than the α level) the bigger the interaction. All surface water samples showed the impact of leachate. The pH values showed slight variation throughout the five years of research; they ranged from 7.4 to 8.4 at sampling site WP1, and from 7.1 to 7.8 at WP2. The pH was influenced by the location (F/ Fc ¼ 1.53, p o0.05); however there was no significant difference between period. The amount of PAHs was at a very low level (below 0.1 mg/L) in all samples. On the basis of the measured values of conductivity and TOC, two periods in the functioning of the landfill can be distinguished: (I). from 2007 to 2009; (II). from 2010 to 2012.

In the first period, the conductivity and TOC values measured in water samples collected down the stream from the landfill (site WP2) were very high, ranging from 3.3 to 7.8 mS/cm and from 72.6 to 186 mg/L, respectively. In the second period, a significant decrease in conductivity was observed, while the TOC values after 2010 did not exceed the level of 10 mg/L. Statistical analysis indicates that there is a significant difference of the conductivity and TOC between locations (F/Fc ¼ 19.4, p o0.05 and F/Fc ¼10.5, p o0.05, respectively) and periods (16.8, p o0.05 and F/Fc ¼9.79, p o0.05, respectively). The influence of sampling location on conductivity and TOC was highly depended on period (F/Fc ¼16.7, p o0.05 for conductivity and F/Fc ¼10.1, p o0.05 for TOC). These data suggest that a huge amount of organic and ionic compounds entered the stream water with the leachate. Starting in 2011, a large increase in the level of lead and cadmium in stream water collected from both sites, i.e. up and down the stream from the landfill was observed. Pb and Cd concentrations were significantly influenced by period (F/Fc ¼2.57, p o0.05 for Pb and F/Fc ¼5.38, p o0.05 for Cd). This finding indicates the presence of external sources of water pollution. Apart from the sample taken in the fourth quarter of 2011, the leachate contributed to the increase in concentration of these metals in stream water. An increased zinc content in the samples collected down the stream from the landfill was also observed. Zn concentration, like in case of Pb and Cd, was significantly influenced by period (F/Fc ¼8.81, po 0.05). Trace amounts of chromium(VI) and mercury were present in the stream water at concentrations below the level of quantification of the analytical procedure.

3.2. Toxicity tests of water samples 3.2.1. Microtoxs Results of Microtoxs test are shown in Fig. 2. The results showed that only four water samples were toxic for a bacterium V. fischeri. Three of those samples were taken at site WP2 in the first, second and third quarters of 2008; the respective EC50 values were 80, 53 and 76 mL/L. Only in one sample collected in the third quarter of 2011 at site WP1, the reduction in bacterial bioluminescence intensity was observed, while the measured EC50 value reached 52 mL/L. Other water samples were not toxic for the test organism. According to the results the toxicity of samples to bacteria was not influenced both by location and period (Table 3).

3.2.2. Thamnotoxkit FTM Based on the results of water toxicity testing by Thamnotoxkit FTM, it could be concluded that the samples collected at site WP1 were in most cases less toxic than those collected at site WP2 (Fig. 3); however the toxicity towards T. platyurus was not influenced by location (Table 3). A 100% mortality in crustaceans of the species T. platyurus was only observed in the case of two samples collected at site WP1 in the second and fourth quarters of 2008. On the other hand, all samples collected at site WP2 in the period 2007–2009 showed high toxicity and caused the death of all test organisms. In the following years (since 2011) the toxicity of water samples collected at site WP2 decreased, and the crustacean mortality dropped down to 23%. Thus, obtained results of toxicity were significantly influenced by period (F/Fc ¼ 8.93, po 0.05). Moreover the influence of location depended on period (F/Fc ¼1.62, p o0.05). Table 3 Two-way ANOVA results for toxicity of surface water samples. Parameter [%]

Location F/Fc


Phytotoxkit F™ 1.96 Thamnotoxkit F™ 0.76 Microtoxs 0.73 a b c



Location–period interaction




0.008 0.28 0.283 1.41 0.085 8.93 0.000c 1.62 0.089 0.55 0.138 0.88

p 0.022 0.015 0.064

First period: IV qrt. 2007 –I qtr. 2009, second period: IV qrt. 2010–I qtr. 2012. F/Fc, F value to critical F value ratio. Less than 0.001.

Fig. 2. Toxicity of water samples towards Vibrio fischeri determined by Microtoxs test.

A. Melnyk et al. / Environmental Research 135 (2014) 253–261


Fig. 3. Toxicity of water samples towards Thamnocephalus platyurus determined by Thamnotoxkit F™ test.

Fig. 4. Relationships between the mortality of Thamnocephalus platyurus crustacea and TOC content and conductivity in sampled stream water.

Statistical analysis showed quite high dependence of mortality T. platyurus on TOC (Pearson r ¼0.70, po 0.001) and conductivity (r ¼ 0.70, p o0.001). For high values of TOC concentration (above 70 mg/L) and conductivity above 3 mS/cm, a 100% mortality in crustaceans was observed (Fig. 4). On the other hand, the TOC values lower than 11 mg/L and conductivity below 1150 μS/cm did not have an adverse effect on the life functions of test organisms. No correlation was found between the determined toxicity values and metal content in water. An increased content of heavy metals such as lead and cadmium (reaching up to 43 and 6 mg/L, respectively) did not adversely affect the survival of T. platyurus.

3.2.3. Phytotoxkit FTM As in the aforementioned bioassays, the samples of surface water collected down the stream from the landfill (WP2) between 2007 and 2009 showed greater toxicity than samples collected after 2010 (Fig. 5); however the toxicity towards S. saccharatum was not influenced by period but by location (F/Fc ¼1.96, p o0.05)

(Table 3). The influence of location depended on period (F/Fc ¼1.41, po 0.05). The root growth inhibition in the samples taken at site WP1 generally did not exceed the level of 26%. Only three samples displayed greater root length compared to control; such situation was observed in the samples collected in the first quarters of 2008 and 2009, and in the fourth quarter of 2011. The water samples collected at site WP2 in 2008–2009 were characterized by the highest toxicity (85–99%). In subsequent years, the percentage inhibition of root growth ranged between 15% and 40%. In comparison to control, only the surface water sample collected down the stream from the landfill in the fourth quarter of 2010 had a stimulating effect on the growth of S. saccharatum roots. Based on the Phytotoxkit F™ test results, it can be concluded that there is a strong influence of conductivity on the growth of S. saccharatum roots (Pearson r ¼0.87, po0.001) (Fig. 6). For the conductivity values above 1 mS/cm, very strong inhibition of root growth was observed. The correlation between the growth inhibition and TOC concentration was 0.80 (p o0.001) and the strong growth inhibition (over 85%) was observed for TOC concentration above 70 mg/L. As in the case of Thamnotoxkit F™ test, the Phytotoxkit F™ test results also did not indicate the existence of a correlation between toxicity and metal content. The growth of S. saccharatum root was greater compared to control even at high levels of lead and cadmium in water samples.

4. Discussion Surface water contamination is a population stressor of high significance because many live organisms depend on water for reproduction (Bruner et al., 1998). Environmental samples of surface water can include a variety of toxic substances. Based on the results obtained by Chen and Zoltek (1995), organic contaminants present in leachates are being degraded or metabolized by microorganisms, or become diluted to such an extent that they cannot be detected. In addition, biological and chemical transformations of xenobiotics present in water can lead to the formation of toxic compounds from relatively inoffensive substrates. In this way, a large number of compounds pass through a monitoring system and, unnoticed, negatively affect the aquatic ecosystem. While chemical analysis gives information about the chemical composition, the ecotoxicological analysis provides a direct functional


A. Melnyk et al. / Environmental Research 135 (2014) 253–261

Fig. 5. Toxicity of water samples towards Sorghum saccharatum determined by Phytotoxkit F™ test.

Fig. 6. Relationship between the growth inhibition in S. saccharatum roots and conductivity in sampled stream water.

response that relates to the overall toxicity of samples, supplies valuable information on bioavailability aspects, and integrates the effect of all contaminants, including additive, synergistic and antagonistic effects (Kalciková et al., 2011; Pandard et al., 2006; Wolska et al., 2007). Each test organism can be sensitive to different chemicals (Wenzel et al., 1997) therefore in order to assess the real hazard to the environment, a battery of tests should be used (Persoone et al., 2003). In the past two decades, bioassays were subjected to numerous modifications, so the need for culturing test organisms has been eliminated, the methods of measurement have been standardized, and cost-effectiveness was achieved (Blaise, 1998). Movement of toxic compounds from a landfill to surface waters is mainly due to the migration of landfill leachate. The landfill leachate contains a variety of compounds, inter-alia, aromatic hydrocarbons, fatty and aliphatic acids, phenols, heavy metals and others (Bortolotto et al., 2009; Isidori et al., 2003). Therefore striving to properly secure the landfill area against the migration of pollutants from the landfill to the surrounding areas becomes extremely important. As demonstrated by Yusof et al. (2009), even

in the controlled landfills the impact of leachate on the surface water system can be observed. However, the leachate impact is smaller than in the uncontrolled landfill, which may be the result of the landfill surface runoff or the improper treatment being practiced at the landfill. The landfill investigated in this study does not have the appropriate security system for protecting the surface waters adjacent to the landfill against contamination. Landfill leachate entering a surface water system may pose a serious threat to the living organisms because of the bioavailability and toxicity of iron(II), accumulation of iron(III), the toxicity of inorganic trace elements and xenobiotic organic compounds (XOCs) (Thomsen et al., 2012). The measured pH values indicated that water samples were slightly alkaline, while, in most cases, the samples collected down the stream from the landfill had lower pH than those collected up the stream from the landfill. Alkalinity of stream water below the landfill may be an effect of leachate which has migrated to the stream from both closed and currently exploited sectors of the landfill. In the case of old and middle-aged landfills, the leachate pH is alkaline and usually 47.5 (Frascari et al., 2004; Li and Zhao, 2001; Wu et al., 2004). However, the leachates from young landfills (o 5 years old) are slightly acidic, with a pH ca. 6.5 (Chian and DeWalle, 1976; Ozturk et al., 2003). In this study, the lower pH values of stream water below the landfill suggest that this may be related to the migration of leachate from the 800/1 sector, which has a bottom liner and has been exploited since 2010. The migration of leachate from this sector can also be deduced from the increased levels of metals (i.e. Cu, Cd, Pb and Zn) and high TOC values in water samples that had been collected up the stream from the landfill since the fourth quarter of 2010. Only the leachates from young landfills are characterized by high content of metals and organic compounds as reported by Tatsi et al. (2003). Microtoxs is the only fast toxicity test used worldwide in both scientific laboratories and routine operations (Ribo and Kaiser, 1987) because the test is characterized by a short time of analysis, simplicity of operation, and there is no need to culture the bacteria as they are provided by manufacturers (Drzewicz et al., 2004). The luminescent bacterium V. fischeri, used as a test organism, is susceptible to many different compounds (Kaiser and Palabrica, 1991); however in the present study, it showed the lowest sensitivity. In this case, the application of Microtoxs test only was not sufficient to estimate the exposure of live organisms to harmful pollutants present in the studied samples. The number of

A. Melnyk et al. / Environmental Research 135 (2014) 253–261


Table 4 Hazard classification system for natural waters according to Persoone et al. (2003). Class

PE indicator value

Threat degree


o20%; none of the tests shows a toxic effect 20% rPE o 50%; a toxic effect is reached in at least one test, but the effect level is below 50% 50% rPE o 100%; the PE is reached or exceeded in at least one test, but the effect level is below 100% 100%; the PE is reached in at least one test 100%; the PE is reached in all tests

No acute hazard Slight acute hazard Acute hazard High acute hazard Very high acute hazard

toxic samples identified with the use of Microtoxs test was only 27.8%; this value was calculated as the percentage ratio of samples exhibiting toxicity towards the specified test organism to the total number of samples tested with the use of three tests, i.e. Microtoxs, Phytotoxkit F™ and Thamnotoxkit F™. Many samples which were toxic to T. platyurus and S. saccharatum did not cause a reduction in bacterial luminescence of V. fischeri. A possible explanation for this finding may be the migration of substances characterized by insecticidal and herbicidal activity from the landfill to the stream. Lower frequency of toxic response to surface water samples in V. fischeri compared to other biotests has also been reported by other researchers (Mankiewicz-Boczek et al., 2008). In this study, the incubation time of bacteria in sample water was 30 min therefore a decrease in luminescence of V. fischeri resulted not only from the harmful effects of organic compounds, but also from the presence of heavy metals in water (Kaiser and Palabrica, 1991). However, increased levels of lead and cadmium in water samples did not result in reduced intensity of bacterial bioluminescence. The number of toxic samples identified by using Thamnotoxkit F™ test was 50%, which suggests that T. platyurus is more sensitive to surface water pollution than V. fischeri. The results of numerous studies indicate that, in general, T. platyurus is more sensitive than other organisms used in bioassays (Bortolotto et al., 2009; Isidori et al., 2003; Tsarpali et al., 2012). High mortality of crustaceans was observed in samples with high conductivity. There is very limited published information on the conductivity requirements of T. platyurus. Generally, when the concentration of dissolved ions in solution is outside the tolerance range of the organism, osmotic stress occurs (Munns and Tester, 2008; Yin et al., 2013). The osmotic stress may result in disrupted osmoregulation (Zehmer et al., 2002), growth inhibition (Rukke, 2002), lower reproduction and increased mortality (Ashforth and Yan, 2008). Among aquatic invertebrates, ions showing a considerable effect are sodium, chloride and formate ions (Corsi et al., 2009; Sulej et al., 2014). The tolerance range of T. platyurus for conductivity is quite wide. Conductivity 752 mS/cm does not adversely affect this organism (Latif and Licek, 2004) while high mortality of other crustacean Echinogammarus ischnus was observed at 108 μS/cm (Kestrup and Ricciardi, 2010). Samples characterized by high values of TOC contain large amounts of various organic compounds. The intensity of the toxic effect of compound on living organism depends on the type of pollutant and its specific concentration. High mortality of T. platyurus in samples with high TOC Is observed because concentration of some organic compounds adversely affecting the test species has been reached. This can be an example of species-specific nature of toxicity (Latif and Licek, 2004). Considering the lack of sensitivity in this crustacean to elevated concentrations of heavy metals in water samples, high mortality rates of T. platyurus exposed to the samples with correspondingly low values of conductivity and TOC content (1150 mS/cm and 11 mg/L, respectively) are probably due to the presence of compounds, including environmental pollutants, which have not been accounted for in the monitoring program. Based on the results obtained in this study, it can be concluded that Phytotoxkit F™, a test originally intended for evaluating the

toxicity in soil and sediment, was most sensitive. The test was characterized by the highest number of identified toxic samples (55.6%). Although Phytotoxkit F™ is a chronic test, no food or organic material was added during incubation. Thus the influence of organic material on the bioavailability of toxic compounds, which has been proven by Nałęcz-Jawecki and Sawicki (2002), was eliminated. The obtained test results indicated a strong dependence of the root growth inhibition on conductivity. The degree of availability of mobile fractions and thus of water-soluble compounds to plants is very high. It is the water pollution that most strongly affects producers, and thus the basic level in a trophic chain. It is worth noting that different exposure times are used in Thamnotoxkit F™ and Phytotoxkit F™ tests (24 h for T. platyurus vs. 72 h for S. saccharatum) which could affect the obtained results. Also, the results of Phytotoxkit F™ test may vary depending on the plant species used as a test organism (Oleszczuk and Hollert, 2011). Based on the data presented in Sections 3.2.1, 3.2.2 and 3.2.3, an attempt was made to classify the surface water samples according to the five-level classification system for acute toxicity proposed by Persoone et al. (2003). Classification is conducted on the basis of a percentage value of an observed effect (PE), estimated during the test conducted on an undiluted sample. Table 4 summarizes the information which is the basis of this classification. Table 5 contains information about allocation of water samples to appropriate toxicity classes (in accordance with the proposed toxicity scale) and water quality classes based on the measured values of physicochemical parameters of these samples. Water of the highest quality is classified as class I, while that of the lowest quality, as class V. The surface water samples collected down the stream from the landfill (WP2) in 2007–2009 were classified as toxicity class IV. Only three samples taken at site WP1 were characterized by a lack or low level of hazard to live organisms (toxicity classes I and II). In the case of samples collected during the second study period (2010–2012), a significant reduction in the toxicity of surface water was observed. Almost all of those samples belonged to toxicity classes I and II, with an exception of samples taken up the stream from the landfill (WP1) in the first and third quarters of 2011, which have been classified as toxicity class III. Acute hazard classification showed no correlation with the classification based on physicochemical parameters, as has also been reported by others (Mankiewicz-Boczek et al., 2008; Matejczyk et al., 2011). In general, the classification based on the results of physicochemical analysis indicated a better water quality in comparison to the outcome of toxicological classification. Thus the physicochemical analysis may give a false impression of good water status, whereas the results of bioassays on the same water sample will indicate its toxicity towards live organisms. A review of risk assessments, specifically regarding landfills, provides a basis for the conclusion that there is no comprehensive risk assessment methodology that would apply to all landfill types taking into account such parameters as, the type of waste deposited at the site, and hydro-geographical conditions in the area surrounding the landfill, and that would include risks and quantification of risks of all possible impacts of the landfill on the


A. Melnyk et al. / Environmental Research 135 (2014) 253–261

Table 5 Determination of toxicity in the analyzed surface water samples based on the values of physicochemical parameters and the toxicity classification system proposed by Persoone et al. (2003). Sampling date

IV qtr. 2007 I qtr. 2008 II qtr. 2008 III qtr. 2008 IV qtr. 2008 I qtr. 2009 IV qtr. 2010 I qtr. 2011 II qtr. 2011 III qtr. 2011 IV qtr. 2011 I qtr. 2012

Sample number


Scorea Microtoxs

Thamnotoxkit F™

Phytotoxkit F™

0 0 0 2 0 2 0 2 0 1 0 1 NP 0 0 0 0 0 2 0 0 0 0 0

0 3 0 2 3 3 2 3 3 3 1 3 NP NP 2 0 0 0 0 0 0 0 0 1

NPc NP 0 3 NP NP 1 2 0 2 0 3 NP 0 1 1 1 1 0 1 0 0 0 0

Acute hazard classification

Classification based on physical and chemical parametersb



a Score: r19,9% – 0; 20–49,9% – 1; 50–99,9% – 2; 100% – 3. For Microtoxs – % effect; for Thamnotoxkit F™ – % of mortality, for Phytotoxkit F™ – % of growth inhibition of roots. b Based on limits of water quality indicators relating to the surface water such as streams and rivers specified in the Polish legislation. c NP, not performed.

ecosystem (Butta and Oduyemi, 2003). However, supplementing the results of chemical analysis with the outcome of ecotoxicological studies allows us to obtain a more complete picture of the quality of individual elements of the environment.

5. Conclusions The presented comprehensive study of water samples, which had involved determining a number of physical parameters, the concentrations of chemical pollutants and water toxicity, points to the fact that good chemical status of surface water is not equivalent to its good ecological status. The obtained results are also the basis for concluding that bioassays can perfectly complement the monitoring system based on chemical research conducted in the area of the municipal waste landfill. However it is necessary to use a battery of biotests because most organisms are sensitive to a selected group of chemicals. It is important to emphasize that S. saccharatum showed high sensitivity to pollutants present in the sampled water. A modification of Phytotoxkit™, a test originally developed for evaluating toxicity in soil, and its adaptation to evaluate toxicity in water samples have proved very beneficial. The low sensitivity of V. fischeri to pollutants present in the analyzed water samples indicates that Microtoxs test is not adequate for toxicological evaluation of surface water. However, the application of an extended battery of bioassays provides important information about the water quality. High mortality of T. platyurus may indicate high values of TOC and conductivity of water. Consequently toxicity tests, including Microtoxs, Thamnotoxkit F™ and Phytotoxkit F™ tests, should be permanently incorporated into the practice of environmental monitoring in

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Chemical pollution and toxicity of water samples from stream receiving leachate from controlled municipal solid waste (MSW) landfill.

The present study was aimed to determine the impact of municipal waste landfill on the pollution level of surface waters, and to investigate whether t...
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