ARTICLE IN PRESS

Ecotoxicology and Environmental Safety 70 (2008) 294–299

www.elsevier.com/locate/ecoenv

Relationships between physicochemical parameters and the toxicity of

leachates from a municipal solid waste landfill
Jesús Olivero-Verbel, Carlos Padilla-Bottet, Orlando De la Rosa
Environmental and Computational Chemistry Group, Department of Chemistry, University of Cartagena, Cartagena, Colombia
Received 5 January 2007; received in revised form 10 May 2007; accepted 27 May 2007
Available online 21 August 2007

Abstract

Landfills are used to dispose municipal solid wastes, and although on-site recycling in these places is an extensive practice in Latin
America, diverse pollutants are incorporated into the leachates. The objective of this work was to establish relationships between
composition and toxicity of leachates from the landfill of the city of Cartagena, Colombia. Leachates were characterized measuring Cd,
Ni, Hg, Mn, Cu, and Pb concentrations, and physicochemical parameters including pH, conductivity, chemical oxygen demand (COD),
and hardness. Bioassays were conducted diluting with synthetic sea water, recording toxicity against Artemia franciscana as median lethal
concentrations (LC50 values) after 24 and 48 h exposure. Average LC50 values oscillated between 3.20% and 39.33% (v/v). Multivariate
analysis showed that toxicity was dependent on Cd and COD. The slope of the concentration–response curve correlated with Ni
concentration independently from toxicity. Results suggest toxicity of these leachates depends on Cd concentrations associated with
organic matter, this effect being modulated by Ni.
r 2007 Published by Elsevier Inc.

Keywords: Artemia franciscana; Bioassay; Cadmium; Chemical oxygen demand; Colombia; Heavy metals; Landfill; Leachate; Nickel

1. Introduction In general, municipal landfill leachate composition is

The landfill is the most common technology used to dispose
municipal solid residues in undeveloped countries. It consisted
of a series of deposition cells in which solid wastes are confined
in the lowest possible surface area, after being reduced to a
minimum volume by compactation. Cartagena city landfill
(Henequen) was operated from 1965 to October 2001 for the
disposal of municipal waste. The capped landfill comprised
approximately 50 acres and it had a total capacity of
1,671,100 m3 of solid waste (daily production, 600 ton). It
had 10 levels with 225 cm depth separation between them, and
each one was covered with 25 cm of sand. Leachates generated
in the landfill were conducted through PVC tubes to a storage
zone. These leachates were recirculated within the landfill as a
treatment procedure.
Corresponding author. Fax: +57 5 6698323.
E-mail addresses: joliverov@unicartagena.edu.co,
jesusolivero@yahoo.com (J. Olivero-Verbel).
URL: http://www.reactivos.com (J. Olivero-Verbel).
complex. It is known to be rich in metals, ammonia,
organic compounds, and other toxicants (Pivato and
Gaspari, 2006) that can be of great concern for aquatic
environments. Therefore, its toxicity has been studied in
several plant and animal models (Sang et al., 2006; Deguchi
et al., 2006; Osaki et al., 2006; Bloor et al., 2005).
Due to the possibility of finding toxic compounds in the
leachate of Cartagena landfill, a study was conducted to
evaluate its toxicity using the cladocerous Artemia francis-
cana, commonly referred to as ‘‘Artemia salina’’ (Thomp-
son and Abramovitz, 1956). This organism has been used
for the evaluation of toxicity derived from industrial
chemicals such as phenolic compounds, antifouling paints,
surfactants (Barahona and Sanchez-Fortun, 1996; Loschau
and Kratke, 2005; Liwarska-Bizukojc et al., 2005),
pesticides (Lieberman, 1999), dental materials (Pelka
et al., 2000), natural products such as plant coumarins
(Ojala et al., 1999), industrial effluents (Guerra, 2001), and
leachates from landfills (Svensson et al., 2005), among
others. The goal of this work was to evaluate the acute and

0147-6513/$ - see front matter r 2007 Published by Elsevier Inc.

doi:10.1016/j.ecoenv.2007.05.016
 ARTICLE IN PRESS
J. Olivero-Verbel et al. / Ecotoxicology and Environmental Safety 70 (2008) 294–299 295

sublethal aquatic toxicity of the leachate from a municipal analysis was employed. Frequently used in a variety of environmental
and toxicological studies (Ren et al., 2004; Zeng and Rasmussen, 2005),
landfill in a Latin American city, relating toxicity to
factor analysis condenses the variables with a minimum loss of
chemical composition. information in a new reduced group of conceptually meaningful variables
named factors, where each one of them is a linear combination of the
original variables. Variances extracted by the factors are called the
2. Methods
eigenvalues. For all statistical analysis, significance was set at Po0.05.

Between May and September 2001, 10 leachate samples were collected

from the landfill approximately each 15 days apart. Composite samples
were taking directly from the leachate pool to complete a final volume of
1 L. Samples were stored on glass recipients and immediately transported
to the lab where they were filtered twice. First, they were passed through a
Whatman 40 cat. no. 1440 filter, 125 mm diameter to separate particles,
followed by a membrane filtration using a 47-mm diameter filter with
0.45 mm pore. A subsample was acidified to pH 2 with HNO3 and kept to
20 1C for later metal analysis.
Acute toxicity experiments using A. franciscana (San Francisco Bay
Brand) were conducted following EPA recommendations (Weber, 1993).
The experimental design consisted of at least seven treatments (leachate
concentrations) and one control, having three replicates per treatment.
Each assay was conducted using 10 mL glass vials containing leachates
and artificial sea water (Hartl and Humpf, 2000), as a dilution solvent, to
obtain the required testing concentration. Physicochemical properties
measured after the dilutions of the leachates with the synthetic sea water
varied as follows: conductivity, 49.1–57.5 mS/cm; pH, 7.6–8.0, and
salinity, 32–38%.
Mortality was recorded each 24 h and a 16:8 h light:dark photoperiod
was used. After 24-h incubation of eggs, a total of 10 healthy, swimming
animals were transfer to the vials. After 24 and 48 h, dead and living
organisms in each well were counted (Lieberman, 1999). The LC50 values
were calculated using probit transformation of the mortality data (Hlywka
et al., 1997). Bioassays were repeated four times using synthetic sea water
as a control.
Physicochemical analysis such as total hardness and chemical oxygen
demand (COD) were done using the techniques presented in the Standard
Methods for the Examination of Water and Wastewater (APHA, AWWA,
and WEF, http://www.standardmethods.org/). pH was measured with a
pH-meter E250 Metrohm, Herisau, Switzerland. Conductivity was read
with a WTW LF 320 conductimeter. Total Cd, Mn, Ni, Pb, and Cu
concentrations in the leachate were determined by atomic absorption
spectroscopy (AAS) in a Unicam 969 Solaar, after acid digestion,
following the US EPA method 3050. Hg was measured by cold vapor
AAS. Detection limits of 0.015, 0.10, 0.04, 0.025, 0.03, and 0.06 mg/L were
obtained for Hg, Pb, Cd, Cu, Mn, and Ni, respectively.
For all cases, data are presented as mean7S.E. Data normality was
checked using the Kolmogorov–Smirnov test. Bivariate relationships were
conducted using Pearson correlation and the association between LC50
and physicochemical variables was performed using multiple linear
regression. In order to analyze multiple interrelationships among all the
variables, including physicochemical and toxicological ones, factor
3. Results
Physicochemical characterization of leachate samples is
shown in Table 1. Leachate pH was always alkaline, with a
conductivity lower than sea water, highly variable in
hardness, and with a high-organic content. In terms of
metals, Cd, Mn, and Ni were always present, whereas Hg,
Cu, and Pb were almost always below the detection limit.
Toxicity values (LC50) assessed using A. franciscana,
as well as the slope of the concentration-probit line (SCPL),
for leachate samples, at both 24 and 48 h are provided in
Table 2.
The LC50 of the leachate had an average of
18.3074.36% (v/v) at 24 h with a maximum and minimum
of 39.33% (v/v) and 3.41% (v/v), respectively. At 48 h, the
average LC50 was 17.8674.29% (v/v) with a maximum
and minimum of 39.02% (v/v) and 3.20% (v/v), respec-
tively. No mortality occurred for A. franciscana exposed to
synthetic sea water during the experimental exposure time.
The slope of the concentration–response curve represents
the change in the response (mortality percentage trans-
formed to probit) vs. the increase in the stimulus (leachate
concentration). Maximum and minimum values for the
slope of the concentration-probit line (SCPL) at 24 h were
obtained for sample 10 (SCPL ¼ 28.6) and sample 4
(SCPL ¼ 5.47), respectively (Table 2).
Bivariate correlation analyses between toxicity (LC50)
and physicochemical parameters are shown in Table 3. It is
clear the significant associations between LC50 at 24 and
48 h, and also with SCPL. LC50 was inversely correlated
with COD, which suggests that an increase in the organic
content generates a more toxic leachate. Interestingly, SCPL
correlated with Ni concentration.
Regression analyses performed in order to obtain relation-
ships between toxicity and two significant physicochemical

Table 1
Physicochemical properties of the leachate from the landfill of Cartagena city

Sample pH Conductivity Hardness (mg/ COD Cd (mg/L) Hg (mg/L) Mn (mg/L) Cu (mg/L) Ni (mg/L) Pb (mg/L)
(mS/cm) L as CaCO3) (mg O2/L)

1 8.60 26.3 3000 2880 0.113 o0.015 0.069 0.053 0.173 o0.10
2 8.55 23.5 1400 4000 0.135 o0.015 0.094 o0.025 0.237 o0.10
3 8.60 23.6 1150 4000 0.217 o0.015 0.052 o0.025 0.194 o0.10
4 8.40 16.4 900 3360 0.078 o0.015 0.030 o0.025 0.209 o0.10
5 8.70 23.5 1250 4480 0.295 o0.015 0.079 o0.025 0.194 o0.10
6 8.50 20.2 1000 3680 0.119 o0.015 o0.030 o0.025 0.180 0.137
7 8.50 17.5 1000 2720 0.057 o0.015 0.031 0.040 0.180 o0.10
8 8.65 26.0 1100 3360 0.078 o0.015 0.165 o0.025 0.209 o0.10
9 8.55 23.3 1250 3200 0.039 o0.015 0.035 o0.025 0.216 o0.10
10 8.60 25.3 450 2400 0.103 o0.015 o0.030 o0.025 0.359 o0.10
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296 J. Olivero-Verbel et al. / Ecotoxicology and Environmental Safety 70 (2008) 294–299

variables are shown in Table 4. Both predicted models have
the conductivity as a significant parameter that decreases the
toxicity of the leachates, whereas COD and Cd increase it.
Factor analysis from all variables included in the data set
(toxicity at 24 h, metal content, and physicochemical
properties of the leachate), led to the extraction of three
factors with eigenvalues greater than 1 (Table 5). The first
factor explained 37.71% of the total variance, the second
29.53%, and the third 17.94%, yielding a total of 85.18%.
To obtain a more interpretable results from the factor
analysis, the extracted factors were rotated applying the
Varimax method, maximizing the variance of the square
factor loadings. This procedure permitted to generate
values of the correlation coefficients between the variables
and the factors to be either higher or lower than those in
the unrotated matrix, with numbers close to 1 or 0,
allowing better identification of the relationships between
them (Table 6). The first factor was highly correlated with
LC50 at 24 h, COD, and to a lesser extent with Cd, the
second factor was associated with conductivity and pH,
and the last factor linked to Ni concentration and SCPL.
4. Discussion
The toxicity of Cartagena city landfill leachates on A.
franciscana varied from sample to sample. These differ-
ences might be associated with pollution load or solid-
residue composition (Ozkaya et al., 2006), change in

Table 4
Table 2 Regression analysis for leachate toxicity using two variables
LC50, confidence limits and slope of the concentration-probit line (SCPL ),
for bioassays performed at 24 and 48 h using the leachate of the landfill Physicochemical Regression parameters
from Cartagena city variable
Regression coefficient t-value P-value
Sample LC50 Confidence SCPL LC50 Confidence SCPL
24 h limits 48 h limits Model 1
COD 0.1770.003 5.666 0.001
1 37.2 33.8–41.0 7.28 35.7 32.5–39.1 7.64 Conductivity 1.83970.575 3.197 0.015
2 8.81 7.80–9.95 8.20 8.59 7.62–9.68 8.30 Constant 35.912716.65 2.157 0.068
3 9.04 8.14–10.0 9.50 8.73 7.90–9.65 9.90
4 10.1 8.89–11.5 5.47 9.72 8.77–10.8 6.90 R2 ¼ 0.816, F ¼ 20.933, P ¼ 0.001
5 3.41 2.98–3.89 7.50 3.20 2.74–3.73 6.50 Model 2
6 5.15 4.47–5.94 7.00 5.03 4.36–5.80 7.02 Conductivity 2.44970.964 2.541 0.039
7 17.6 16.1–19.3 7.82 16.8 15.5–18.3 8.72 Cd 118.583742.453 2.793 0.027
8 17.5 15.9–19.4 7.08 17.2 15.5–19.0 7.09
Constant 22.308721.311 1.047 0.330
9 34.8 33.4–36.2 24.9 34.7 33.3–36.1 24.5
10 39.3 38.1–40.6 28.6 39.0 37.7–40.4 28.4 R2 ¼ 0.513, F ¼ 5.747, P ¼ 0.033

Table 3
Correlations between toxicity slope of the concentration-probit line (SCPL) and physicochemical parameters for leachates from the landfill of Cartagena
city

LC50 24 h LC50 48 h SCPL 24 h SCPL 48 h pH Conductivity Hardness COD Cd Mn Ni

LC50 24 h 1.00
LC50 48 h 1.000a 1.00
(0.001)b
SCPL 24 h 0.709 0.728 1.00
(0.022) (0.017)
SCPL 48 h 0.724 0.742 0.997 1.00
(0.018) (0.014) (0.000)
pH 0.108 0.109 0.132 0.071 1.00
(0.767) (0.764) (0.715) (0.845)
Conductivity 0.447 0.449 0.327 0.284 0.820 1.000
(0.195) (0.194) (0.357) (0.427) (0.004)
Hardness 0.269 0.247 0.335 0.344 0.205 0.383 1.000
(0.453) (0.492) (0.344) (0.330) (0.569) (0.275)
COD 0.805 0.801 0.491 0.534 0.277 0.013 0.010 1.000
(0.005) (0.005) (0.150) (0.111) (0.439) (0.972) (0.977)
Cd 0.522 0.524 0.279 0.327 0.582 0.242 0.066 0.750 1.000
(0.122) (0.120) (0.435) (0.357) (0.078) (0.500) (0.856) (0.012)
Mn 0.211 0.216 0.427 0.457 0.529 0.485 0.268 0.355 0.143 1.000
(0.558) (0.550) (0.219) (0.184) (0.116) (0.156) (0.453) (0.314) (0.693)
Ni 0.459 0.476 0.762 0.767 0.113 0.295 0.501 0.414 0.129 0.246 1.000
(0.182) (0.165) (0.010) (0.010) (0.756) (0.408) (0.140) (0.235) (0.723) (0.493)
a
Significant correlations are shown in bold type.
b
P-values are given in parenthesis.
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Table 5 to exist as a divalent species (Cd2+), the soft character of

Eigenvalues and percent of variance explained for each factor extracted this atom makes it suitable for binding to sulfur-containing
from the data set
compounds or other soft ligands. In this case, Cd will not
Factor Eigenvalue Percent of Cumulative follow a toxicity mechanism in which it is in free form. Due
number variance percentage to the fact that leachate toxicity increases with organic
matter, it is possible that Cd has to be associated with
1 3.394 37.708 37.708
organic compounds in order to get absorbed. In fact,
2 2.658 29.529 67.237
3 1.615 17.942 85.179 leachate models suggests that Cd is found forming
4 0.786 8.734 93.913 complexes with organic compounds (Halim et al., 2004).
5 0.247 2.749 96.662 Moreover, it is well known that Cd can form complexes
6 0.228 2.538 99.200 with humic or fulvic acid present in municipal solid waste
7 0.057 0.636 99.836
8 0.014 0.153 99.989
(Kaschl et al., 2002). Several reports have shown that Cd
9 0.000 0.011 100.000 bioavailability and toxicity increase with organic matter
(Laegreid et al., 1983; Winner, 1984, 1986; Kozuch and
Pempkowiak, 1996). However, there are some other reports
that argue that organic matter can reduce bioavailability
Table 6 (Stackhouse and Benson, 1989; Hollis et al., 1996) and
Varimax rotated factor matrix for toxicity and physicochemical para- toxicity (Kukkonen et al., 1998). According to Table 3, the
meters of leachates obtained from the landfill of Cartagena city correlation between SCPL and Ni concentration was 0.762
Variable Factor 1 Factor 2 Factor 3 (P ¼ 0.01). Partial correlation between these two para-
meters, but adjusting to LC50 showed the same trend
LC50 24 h 0.908 0.304 0.231 (R ¼ 0.76, Po0.05), suggesting that although Ni does not
SCPL 24 h 0.489 0.186 0.790
have a direct relationship to toxicity (R ¼ 0.459,
pH 0.225 0.928 0.088
Conductivity 0.170 0.960 0.081 P ¼ 0.182), it can increase the sensitivity of Artemia to
Hardness 0.301 0.408 0.751 other contaminants.
COD 0.900 0.163 0.212 Data from factor analysis were in agreement with the
Cd 0.787 0.416 0.034 regression information previously provided, and basically
Mn 0.228 0.577 0.444
reinforce the idea that the toxicity of studied landfill
Ni 0.262 0.172 0.871
leachates is statistically related with the presence of organic
compounds and Cd levels, being other variables such as
conductivity and Ni concentration, modulatory factors
caudal, sediment perturbation, and tropical seasonal that encode valuable information about the nature of the
variations (Trankler et al., 2005), among other factors. leachate.
However, toxicity was similar at 24 and 48 h exposure Despite the significant statistical relationships presented
(18.3074.36% (v/v) vs. 17.8674.29% (v/v)), probably in this study, between toxicity and physicochemical
meaning that those larvae that did not die at 24 h became parameters of the leachates, other variables derived from
tolerant to the environment and no further toxicity was the solid waste deposited on the site, can also be
observed. The overall mean LC50 value observed in this responsible for the observed effects on A. franciscana.
study was slightly greater than those reported for the For instance, toxicity in the leachates from several landfills
juvenile macro-invertebrate Asellus aquaticus (12.3% (v/v)) in Sweden and Lithuania have been attributed to ammo-
using landfill leachates in deionized water with COD of nium/ammonia components and to organic pollutants
1200 mg/L (Bloor et al., 2005), and also with Ceriodaphnia (Svensson et al., 2005), with similar results also found in
dubia (o10%) exposed to leachates from several sites in Brazil (Silva et al., 2004) and Italy (Pivato and Gaspari,
Florida, USA (Ward et al., 2002). 2006). However, toxicity in leachates from Taiwan was not
Negative relationships between LC50 and COD might dependent on ammonia content and a significant degree of
indicate that the substances responsible for the toxic effects variation was detected on several factors that may influence
require the formation of complexes with organic matter to leachate toxicity (Fan et al., 2006). Among the vast number
elicit their biological activity, or alternatively, that some of organic pollutants that can contribute to the toxicity of
organic compounds are inducing toxicity. Two parameter landfill leachates, some of the most frequently found
regression equations provide additional information on include bisphenol A (Coors et al., 2003), phthalates
how physicochemical parameters act together to promote (Marttinen et al., 2003), phenol (Mor et al., 2006),
toxicity (Table 4). According to both linear regression pesticides (Bras et al., 2000), polycyclic aromatic hydro-
models, toxicity decreases with an increase in ionic carbons (PAHs) (Marttinen et al., 2003), biphenyls,
substances whereas organic matter (COD) and Cd in- organic phosphates, and polychlorinated biphenyls (Thor-
creases it. This observation implies that toxicity might neby et al., 2006), among others.
depend on the formation of organic complexes rather than The fact that Cd and Ni were two important metals in
on free ionic compounds or elements. Although Cd tends the toxicity of the leachates of the landfill in Cartagena is a
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298 J. Olivero-Verbel et al. / Ecotoxicology and Environmental Safety 70 (2008) 294–299
first observation. Main sources of these metals in municipal
solid waste could include food cans, metal alloys, electronic
objects, cement and construction debris, and nickel–
cadmium batteries, among several others. Considering that
these metals are strongly complexed and difficult to remove
from landfill leachates (Cecen and Gursoy, 2000), the use
of Cd should be restricted in applications where leach-
ability of the disposed product is an issue. Meanwhile, one
efficient way to avoid their presence in these mixtures is
through recycling processes both in house and on the
landfill site.
Taken together, these results indicate that A. francisca-
na-based bioassay is an useful and non-expensive tool for
the rapid and reliable determination of leachate toxicity. In
addition, municipal leachate toxicity in some Latin
American cities could, in part, be decreased if the
incorporation of heavy metals into the garbage is banned
or prevented. This must be done by educational campaigns
on recycling and waste management issues that should be
undertaken by local authorities with the participation of
the local community.
Acknowledgments
This work was sponsored by the Environmental and
Computational Chemistry Group of the University of
Cartagena, Cartagena, Colombia. Special thanks to LIME
consortium and to Eng. Manuel Garcia.
Funding to support this work was provided by the
University of Cartagena. All the bioassays were conducted
in agreement with national laws and institutional guidelines
for animal used in experimental bioassays.
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