International Journal of Environmental Science and Technology

https://doi.org/10.1007/s13762-018-1719-6

ORIGINAL PAPER

The assessment of the genotoxicity of e‑waste leachates from e‑waste

dumpsites in Metro Manila, Philippines
Z. F. Alam1   · A. J. V. Riego1 · J. H. R. P. Samson1 · S. A. V. Valdez1

Received: 30 March 2017 / Revised: 21 July 2017 / Accepted: 9 May 2018

© Islamic Azad University (IAU) 2018

Abstract
The extraordinary growth in the usage of electronic gadgets and their improper disposal has led to the massive problem of
electronic waste management in the Philippines. The improper disposal of e-waste can lead to the exposure to the hazardous
substances that can be harmful to the humans as well as to the environment. Genotoxicity of e-waste from three different
e-waste landfills sites in and around Metro Manila was investigated using the Allium cepa bioassay. Morphological modifica-
tions of the Allium cepa roots, inhibition of root growth, mitotic index and the induction of high frequency of chromosomal
aberrations were observed in the Allium cepa roots exposed to e-waste leachates prepared from the soil samples collected
from all three sites tested as compared to the control. Since heavy metals are one of the hazardous components released
during e-waste recycling, environmental contamination was further detected by Atomic Absorption Spectroscopy to detect
the presence of heavy metals (Cd, Cu, Ni, Pb and Zn) in the soil samples of the e-waste recycling sites and in the hair of the
informal recyclers. Our results are among the first few in the Philippines that establish the genotoxicity of e-waste as well as
trace the route of the heavy metal component of e-waste into the human subjects through the exposure to the environmental
contamination by e-waste. Hence, a proper e-waste management is an absolute priority to minimize the risk to the Filipino
population and the environment.

Keywords  E-waste · Heavy metals · Philippines · Chromosomal aberrations · Allium cepa · Genotoxicity

Introduction Cambodia, Vietnam, Philippines, Bangladesh and Pakistan

Electronic waste generation and its management is one of
the global issues that need immediate attention and action.
According to the Solving the E-waste Problem Initiative
(StEP) estimation, 49.8 million tons of used electrical and
electronic equipment was discarded as e-waste in 2012 and
the e-waste generation is further estimated to be a stagger-
ing 65.4 million tons by 2017 (UNU 2013). Many develop-
ing economies located in the vast continent of Asia have
become the major e-waste contributor to the world with an
estimated 16 million tons of e-waste generation in the year
2014 itself, thereby making the Asian continent the emerg-
ing “E-waste Hotspot.” The countries like China, India,
Editorial responsibility: M. Abbaspour.
* Z. F. Alam
zeba.alam@dlsu.edu.ph
1
Biology Department, De La Salle University, 2401 Taft
Avenue, Manila, Philippines
have not only witnessed increase in the domestic e-waste
generation but are also the recipients of the e-waste gener-
ated in the developed countries due to cheap labor and lack
of stringent regulations.
Philippines with the distinction of being known as the
“Texting Capital of the World” has also seen tremendous
increase in the generation of e-waste in the past few years
(Alam 2016). The increase in the e-waste is from two
sources—the locally generated e-waste comprising of the
obsolete electronic gadgets discarded by the end users and
the foreign e-wastes that end up in Philippines due to the
transboundary movement of the e-waste from developed
countries for recycling purposes. Though the Basel Conven-
tion treaty has made efforts to provide a legitimate frame-
work to regulate the transboundary movement of hazardous
wastes, many developing countries including Philippines end
up as recipients of the hazardous wastes from the developed
countries for recycling purposes (Gutierrez and Agarrado
2011).

13
Vol.:(0123456789)

Most of these informal e-waste-recycling activities are
located at the e-waste dumpsites at the Smoky Mountain,
Pier 18 and Tondo in Manila, Dreamland in Rosario, Cavite
and Valenzuela cities (Gutierrez and Agarrado 2011). The
e-waste consists of over 1000 toxic substances; the more
prevalent are heavy metals like lead, mercury, arsenic, cad-
mium and organic pollutants such as brominated diphenyl
ethers, polybrominated diphenyls, dibenzofurans dioxins,
hexavalent chromium besides glass, plastics, wood, iron
and steel (Lundgren 2012). Some of these toxic substances
such as cadmium, hexavalent chromium and beryllium are
known to be carcinogenic, and others such as the organic
pollutants like polybrominated diphenyl ethers, polycyclic
aromatic hydrocarbons and aluminum cause genotoxicity.
The improper e-waste management leads to the exposure of
human populations to these hazardous substances as well as
the contamination of the environment.
In the developing countries including Philippines, a sub-
stantial portion of recycling of e-waste is carried out by
small junk shop owners or by informal recyclers (scavengers,
waste pickers and junk shop and scrap dealers) who use very
primitive and often-unsafe practices such as segregating and
sorting by bare hands and thereby getting exposed to the
dangerous substances present in the e-waste. At the e-waste
landfill sites, the e-waste is either dumped or buried in the
trenches that are later covered with a thick layer of soil. This
method often leads to the contamination of the surround-
ing areas including the groundwater due to the leaching out
of these hazardous substances and heavy metals, especially
after the heavy precipitation (Pant et al. 2012). Further, the
practice of open burning of e-waste to gather precious metals
leads to the emission of heavy metals in air that results in
air pollution and contamination of the environment besides
causing health issues to the recyclers.
In order to come up with suitable guidelines and proper
e-waste management practices, it is essential to gather data
and relevant information regarding the full potential of the
toxicity impacts of e-waste which at the moment is very
scant and mainly from the samples and case studies from
China, India and Nigeria (Li et al. 2007; Caravanos et al.
2011). There is an urgent need to gather data and informa-
tion from e-waste dumpsites located at other countries and
also design new and sensitive models (both in vitro and
in vivo) to evaluate the efficacy of the existing standard assay
methods to study the full cytotoxic and genotoxic potential
of e-waste. Since Philippines is one of the biodiversity hot
spots, it is of utmost importance to gather relevant infor-
mation regarding the e-waste toxicity if any, so that timely
action can be taken to protect the environment. This research
is one of the very first few attempts to study the genotoxicity
of the e-waste in the Philippines.
This study uses the Allium cepa assay to determine the
genotoxicity of e-waste in terms of inhibition of root growth
13
International Journal of Environmental Science and Technology
and mitotic index and induction of chromosomal aberrations.
Genotoxicity refers to the capacity of any clastogenic agent
to cause lesions in the genetic material or mutations in DNA
and thereby interruption in the genetic transmission. The gen-
otoxins are any such agents that possess the ability to alter
the DNA. The genotoxic potential is vital, as it is a tool to
determine the magnitude of genetic risk to humans by an envi-
ronmental agent/chemical due to unintentional or accidental
exposure. Due to ethnic, logistic and practical reasons, often
it is unfeasible to carry out direct assessment in humans; con-
sequently, other test systems and assay methods are needed.
Allium cepa assay has proven to be a fast, very efficient and
sensitive method, which involves both macroscopic and micro-
scopic parameters to detect environmental genotoxicity. For
this reason, the Allium cepa assay has been classified as one
of the most well-established plant assay systems for environ-
mental biomonitoring by the USEPA (Grant 1982b). However,
there are very limited studies where the e-waste genotoxicity
has been established using the Allium cepa assay (Bakare et al.
2013). The test is also done in vivo, which means data can be
extrapolated to other plants and even animals (Tedesco and
Laughinghouse 2012). For this reason, in this study, we are
using Allium cepa assay to evaluate the suitability of this assay
method to study the e-waste genotoxicity.
Since the heavy metals such as lead, cadmium, chromium,
mercury, copper, manganese, nickel, arsenic, zinc, iron and
aluminum are also reported to be present in the e-waste (Jin-
hui et al. 2011), the soil samples and leachates will be sub-
jected to Atomic Absorption Spectroscopy to determine their
presence in the soil samples and in the hair of the informal
e-waste recyclers. Leachate is a liquid that contains dissolved
and suspended substances that can percolate through soil due
to rainfall or surface drainage and contaminate the environ-
ment, for example the groundwater. The Global Environmen-
tal Monitoring System (GEMS) of the United Nations Envi-
ronment Program has recommended the use of human hair
as one of the important materials for biological monitoring
of environmental contamination (Dodd 2010). The presence
of heavy metals in the hair of the informal e-waste recyclers
would be indicative of exposure to the e-waste. The data
obtained could be the baseline for any future studies related
to the consequences, toxicity and risk assessment of e-waste
exposure in the Philippines and elsewhere. The study was
conducted during the period of April 2016 to February 2017.
Materials and methods
Collection of samples
Soil samples were collected from three different sites in
Tondo, Manila, where a substantial amount of e-waste is
being dumped before being recycled and processed for
International Journal of Environmental Science and Technology
further use. After the preliminary survey and extensive
discussions with the Barangay chief and the inputs from
the Sustainable Project Management personnel operat-
ing in Tondo, three sites were selected to carry out our
experiments. These were the sites where the e-waste was
found to be dumped (Fig. 1c, Site 3), the e-waste was burnt
(Fig. 1c, Site 2) and a site which was close to the water body,
namely—Estero De Vitas (Fig. 1c, Site 1).
For control, the soil samples were collected from sites
away from e-waste landfills (Fig. 1a, b). To keep the control
samples absolutely neutral, the soil samples for control were
taken from two sites—Tagaytay (74.5 km) and Batangas
(109 km) away from Metro Manila, respectively.
Hair samples were collected from the informal e-waste
recyclers located in Tondo as well as from healthy indi-
viduals belonging to professions where there is no history
of e-waste recycling. All recommended guidelines by the
Research Ethics Committees (RECs) of De La Salle Univer-
sity were followed such as explanation about our study and
use of consent and survey forms to obtain permission from
the participants. The interview was conducted face-to-face,
and the information regarding their personal details, work
and health was carefully collected using the questionnaire
(“Appendix I”).
Laboratory analysis
Preparation of soil leachates
The soil leachates were prepared using the soil samples col-
lected from the different sites following the standard pro-
cedure—the modified American Society for Testing and
Materials (ASTM-D 3987-35) method (Bakare et al. 2007).
Exactly 250 g of each soil sample was measured using an
analytical balance and then placed in a beaker containing
Fig. 1  a, b Location of control site. c Location of e-waste soil samples (three sites)
1000 mL of distilled water. The beakers were covered and
placed in a shaker with a temperature of 30–60 °C for 48 h.
Filtration was done 4–5 times or until soil particles were no
longer observable. All the samples were carefully labeled
and stored in the refrigerator until further use.
Preparation of simulated e‑waste
The simulated e-waste was prepared by collecting several
old and obsolete electronic devices such as cellular phones,
cameras, MP3 players. These devices were crushed into
smaller pieces using a hammer and then boiled with distilled
water for 30 min.
Allium cepa test
The genotoxicity of the soil samples was assessed using the
standard Allium cepa test described by Tedesco and Laugh-
inghouse (2012). Onion bulbs were carefully examined in
terms of size and condition before being purchased from a
local market in Makati. The outermost scales and old roots
of each bulb were eliminated to allow the development of
new roots. All bulbs were placed in approximately 60 mL
plastic containers with tap water ensuring that the basal
stems were completely submerged. After the growth period
of 48 h, the roots of each bulb were measured. The bulbs
were then exposed to different concentrations (30, 50, 70,
100%) of the previously prepared e-waste soil leachates from
different sites. Tap water served as the negative control while
10 ppm of methylmethanesulfonate (MMS) served as the
positive control. Five replicates were used for each set of
treatments. After 24 h of exposure to the individual treat-
ments, the root length of the onion bulbs was recorded. The
tips of the roots that were observed to be the healthiest were
cut out and fixed in a 3:1 mixture of ethanol: acetic acid.
13
 International Journal of Environmental Science and Technology

After 24 h, the fixing solution was replaced with 70% etha-
nol and stored in the refrigerator until used.
For mitotic index and chromosomal aberrations analysis,
the root tips were treated with 1 N HCl for 12–20 min at
60 °C. After hydrolysis, the roots were rinsed with distilled
water for 2–3 times and were immersed in acetocarmine stain
for 30 min at 60 °C. They were then placed on the glass slides
and gently squashed in 45% glacial acetic acid. The slides
were examined under a light microscope with a total magni-
fication of 100×. Approximately 1000 cells were scored per
treatment for mitotic index and chromosomal aberrations.
The formulas used to calculate for the mitotic index and
the proportion of chromosomal aberrations are as follows:
P+M+A+T
Mitotic index = , Statistical analysis
Total number of cells
Pa + Ma + Aa + Ta
Chromosomal aberrations = . tica software version 13. The differences in the mean ± SE
P+M+A+T
Heavy metals analysis
Atomic Absorption Spectroscopy was carried out using the
available AA-6300 Shimadzu spectrophotometer to check
for the presence of heavy metals in the soil and hair samples.
The metals that were tested include cadmium (Cd), copper
(Cu), lead (Pb), nickel (Ni) and zinc (Zn). The multi-element
working solution of 100 ppm and the multi-element stand-
ard solutions (0.5, 1, 5, 7 and 10 ppm) were prepared using
distilled water for dilution.
Preparation of the hair samples
(AAS) following the standard method described by Cortez
and Ching (2014). Samples were put into 30 mL crucibles
and oven-dried for 2–3 min. These were ground using mor-
tar and pestle, then placed inside the furnace and ignited at
450 °C in order to eliminate organic matter. Exactly 0.5 g of
each sample was weighed in the 30 mL crucibles and then
placed on a hot plate inside the fume hood. Samples were
digested twice with 10 mL of 1:1 mixture of concentrated
­HNO3 and ­H2O2. Heating was done until the solution was
completely dissolved; then, 20 mL of 2 M ­HNO3 was added
to the residue. The contents of each crucible were transferred
to 100 mL Erlenmeyer flask, labeled carefully and stored in
the refrigerator until used for analysis.
The results were analyzed using the standard open Statis-
values for various treatments were subjected to multivari-
ate and univariate single-factor ANOVA at the 0.05 sig-
nificance level. The Tukey–Kramer test was carried out
to further arrive at the concentrations of e-waste leachate
causing significant genotoxicity as compared to the negative
and positive controls. Linear regression analysis was done
to determine whether there were any trends between root
growth, mitotic index and chromosomal aberrations versus
increasing concentrations of the e-waste leachate treatments.
T test was done to compare the means of the control and the
experimental group with the maximum permissible limits
set by World Health Organization (WHO) for each metal for
soil as well as hair samples.

Acid digestion of the hair samples was performed using the

procedure described by Peter et al. (2012). The collected hair
Results and discussions
samples of the healthy individuals (control group) and infor-
Root growth
mal e-waste recyclers were cut into approximately 0.3 cm;
then, exactly 0.1065 g was weighed for every hair specimen.
The absolute concentrations (100%) of e-waste simulated as
The samples were washed once in a mixture of detergent and
well as the e-waste soil leachates collected from the three
water then twice in deionized water. These were placed into
sites induced significant inhibition of root growth in the
30 mL crucibles in which 8 mL of concentrated nitric acid
Allium cepa roots exposed to the leachates for 24 h (Fig. 2,
­(HNO3) was added. The containers were placed on a hot
Graph I), and 72 h (Fig. 2, Graph IV) as compared to the
plate inside the fume hood, and hair samples were digested
negative control (tap water) and the negative control (Batan-
for about 15 min. As the solutions became clear, 1 mL of
gas soil leachate). The maximum inhibition of root growth
30% ­H2O2 was added and heating was done until the volume
observed after 24- and 72-h exposure was comparable to
was decreased to about 2.5 mL. Each digested hair sample
the root growth inhibition induced by the positive control
was transferred to a 100-mL Erlenmeyer flask, carefully
(MMS, 10 ppm) at p < 0.05 (Fig. 2, Graph I and Fig. 2,
labeled and stored in the refrigerator until used.
Graph IV). The regression analysis performed for all three
sites shows that there is a concentration-dependent inhibi-
Preparation of the soil samples
tion of root growth upon 24-h exposure with e-waste lea-
chates from site 2 having the highest R2 = 0.83756, followed
Soil samples were digested prior to the analysis of heavy
by site 1 with R2 = 0.76802 and site 3 with R2 = 0.73110,
metal concentration by Atomic Absorption Spectroscopy
respectively (Fig. 3, Graphs I, II, III). For the 72-h exposure,

13
International Journal of Environmental Science and Technology

Fig. 2  Comparison between different treatments (100% concentration) from all three sites in terms of inhibition of root growth, mitotic index
and induction of chromosomal aberrations in the Allium cepa root cells upon 24- and 72-h exposure

Fig. 3  Root growth inhibition with increase in e-waste soil leachates concentration treatment from all three sites observed in the Allium cepa
root meristem with 24- and 72-h exposure

site 3 had the highest R2 = 0.94502 followed by site 2 with
R2 = 0.92260 and site 1 with R2 = 0.78646 (Fig. 3, Graphs
IV, V, VI). Further for 24-h exposure, site 3, and for 72-h
exposure, the site 2 had the lowest slopes for root growth
inhibition. From the graphs, it can be inferred that the E
­ C50,
which is the effective concentrations at which root growth
amounts to 50% of the control treatment, was close to 30%
concentrations, which is similar to the ­EC50 value of 37.5%
concentration of e-waste leachate reported elsewhere (Baba-
tunde and Anabuike 2015). The morphological alterations

13

in the roots upon exposure to absolute concentrations of the
e-waste leachates were observed to be very severe (Fig. 4).
Mitotic index
The decrease in the mitotic index in the Allium cepa root
cells (which is indicative of the blocked G­ 2 phase in the cell
cycle) was observed following 24-h exposure (Fig. 2, Graph
II) and 72-h exposure (Fig. 2, Graph V) to the absolute con-
centration (100%) of the e-waste leachates from all three
sites as well as the e-waste simulated. The decrease in the
mitotic index induced by 100% concentrations of e-waste
leachates from all three sites for 24- and 72-h exposure
was almost comparable to the decrease in the mitotic index
induced by the positive control (MMS, 10 ppm); however,
the e-waste leachate (100% concentration) from site 2 caused
more inhibition (though statistically insignificant at p < 0.05)
than the other two sites tested in the 24-h exposure period.
Similar trend was observed in the 72-h exposure; however,
there was no significant difference between the levels of
inhibition caused by the e-waste leachates from the three
sites. With regard to the regression analysis for inhibition
of mitotic index performed for all three sites, no predictable
trend was observed as evident by the R2 values. At 24-h
exposure, Site 1 had the highest R2 = 0.59811, followed
by Site 2 with R2 = 0.56030 and Site 3 with R2 = 0.55316,
respectively (Fig. 5, Graphs I, II, III). Conversely, Site 2
was observed to have the highest R2 = 0.85715, followed
by Site 3 with R2 = 0.78926 and Site 1 with R2 = 0.71973,
respectively, at 72-h exposure (Fig. 5, Graphs IV, V, VI). The
Site 1 (24-h exposure) and the Site 2 (72-h exposure) had
the lowest slopes, which indicates that their mitotic index is
decreasing at a faster rate compared to the other two sites.
The observed trends and the E ­ C50 value of 30% concen-
tration are calculated for root length point toward the need
to further test the concentrations lower than 30% e-waste
leachates and the extent of induced genotoxicity. It would
be noteworthy to mention that the site 2 is where the e-waste
was regularly burnt to minimize the volume of the e-waste
Fig. 4  Inhibition of root growth in Allium cepa exposed to various treatments: a negative control—tap water, b control—Batangas soil leachate,
c1–3 e-waste leachates—initial, 24- and 72-h exposure and d positive control—(MMS, 10 ppm)
13
International Journal of Environmental Science and Technology
as well as to extract the precious metals such as palladium,
gold, silver, copper from the discarded mother boards, elec-
tric wires, etc.
Chromosomal aberrations
All concentrations of e-waste soil leachates tested as well
as the simulated e-waste induced chromosomal aberrations
such as fragments, vagrant chromosomes, anaphase and
telophase bridges and laggards in the Allium cepa root cells
exposed to the leachates for 24 and 72 h (Fig. 7, Table 1).
The absolute concentration of e-waste leachates from all
three sites induced significantly higher percentage of chro-
mosomal aberrations than the positive control at p < 0.05
for 24-h exposure (Fig. 2, Graph III). For 72-h exposure,
though the extent of chromosomal damage was almost
comparable, only for site 1 it was statistically significant as
compared to the positive control (Fig. 2, Graph VI). With
regard to the regression analysis for concentration-depend-
ent increase in the chromosomal aberrations performed for
all three sites, no predictable trend was observed for 24-h
exposure as evident by the R2 values where Site 2 had the
highest R2 = 0.62844, followed by Site 1 with R2 = 0.58434
and Site 3 with R2 = 0.44316, respectively (Fig. 6, Graphs I,
II, III). However, for 72-h exposure, a strong concentration-
dependent correlation was observed as evident by the R2
values for the e-waste leachates concentrations from all three
sites tested with the site 3 being the highest, R2 = 0.89458,
followed by Site 2 with R 2 = 0.86891 and Site 1 with
R2 = 0.78650, respectively (Fig. 6, IV, V, VI). The positive
slope indicates the increase in the chromosomal aberrations
with the increase in the concentration of e-waste soil lea-
chates (Fig. 6).
Interestingly in our study, the simulated e-waste exposure
resulted in the ghost cells in the root meristem of Allium
cepa (Fig. 7). Ghost cell is a dead cell in which outline is
visible but nucleus and cytoplasmic structures are not stain-
able and ultimately result in cell death or apoptosis. The
high concentrations of genotoxins and cytotoxins have been
International Journal of Environmental Science and Technology

Fig. 5  Inhibition of mitotic index for e-waste soil leachates treatments from the three sites for 24- and 72-h exposure. Trend line is for concentra-
tions from 30 to 100% compared to negative- and positive-control treatments

Table 1  Breakdown of Treatment Abnormalities Total Chromosomal

results for total chromosomal abnormali- aberrations (%)
aberrations for three e-waste P M A T ties
sites at different concentrations
of soil leachates tested for 24- Hours of exposure 24 72 24 72 24 72 24 72 24 72 24 72
and 72-h exposure
Negative control (tap water) 21 15 24 9 25 19 4 12 74 55 20.10 17.90
Site 1 (30%) 18 10 32 13 27 20 15 18 92 61 44.90 33.30
Site 2 (30%) 6 58 13 17 25 48 20 12 64 135 40.50 40.80
Site 3 (30%) 18 20 30 14 33 24 24 9 105 67 54.12 36.80
Site 1 (50%) 13 36 49 19 42 23 6 12 110 90 49.50 45.20
Site 2 (50%) 2 11 23 9 36 42 14 12 75 74 46.60 41.80
Site 3 (50%) 23 56 20 22 37 25 4 17 84 120 48.55 51.70
Site 1 (70%) 1 19 35 23 47 28 8 14 91 84 67.40 46.90
Site 2 (70%) 19 41 21 20 44 68 21 22 105 151 53.80 58.10
Site 3 (70%) 29 50 42 19 32 67 22 27 125 163 60.39 63.20
Site 1 (100%) 10 27 49 17 35 29 19 23 113 96 74.30 49.50
Site 2 (100%) 13 49 25 23 37 59 17 10 92 141 62.60 62.90
Site 3 (100%) 15 86 30 20 54 42 13 22 112 170 58.64 66.10
Batangas 4 20 23 16 58 24 4 14 89 74 21.00 20.10
Positive control (MMS, 10 ppm) 11 79 18 8 52 31 18 17 99 135 50.00 79.90

reported to induce the ghost cells (Celik and Aslanturk
2006). The exposure to toxins also results in the partial dis-
banding of nucleoproteins, condensation of chromosomes
and depolymerization of DNA, which result in sticky chro-
mosomes. Many studies have reported the occurrence of
sticky chromosomes in Allium roots after treatment with
various heavy metals such as Hg, Ni and Cu (Fiskesjö 1988).
In our study, all e-waste soil leachates from the three sites
induced genotoxicity in terms of inhibition of root length
and mitotic index and induction of chromosomal aberra-
tions; hence, we conclude that Allium cepa assay can be

13

Fig. 6  Chromosomal aberration increase observed in Allium root cells exposed for 24 h and 72 h to various e-waste leachates treatments
used for environmental biomonitoring of e-waste toxicity
for preliminary screening.
Heavy metals analysis
AAS of soil samples
The assessment of the Atomic Absorption Spectroscopy
results confirmed the presence of heavy metals tested—
Cd, Cu, Ni, Pb and Zn in the soil samples of the e-waste
dumpsites and e-waste recycling sites located at Tondo,
Metro Manila (Table 2). Since some studies have reported
the heavy metal contamination around Metro Manila and
Manila Bay (Prudente et al. 1994; Sia et al. 2009), the con-
trol soil samples were collected from two sites away from
Manila—namely Batangas and Tagaytay. There is a varia-
tion to a certain extent regarding the maximum permissible
limits of heavy metals in the soil samples and suggested
thresholds for industrial, residential and agricultural soils,
by various sources such as Dutch pollutant standards, US
Environmental Protection Agency (US EPA), European
Environmental Agency (EEA), World Health Organiza-
tion (WHO) and Department of Environment and Natural
Resources (DENR), which is the local screening authority
in Philippines. In this study, the maximum permissible limits
prescribed by World Health Organization (WHO) were taken
into consideration for the comparison of the data, as it is the
most widely accepted.
The multivariate test (i.e., Wilks, Pillai’s, Hotellng
and Roy’s) suggests no significant difference between the
13
International Journal of Environmental Science and Technology
concentration of heavy metals present in the control soil
samples and the maximum permissible limits set by WHO
with the exception of copper which was above the refer-
ence value (Table 2). The concentration of all the five heavy
metals (Cd, Cu, Ni, Pb and Zn) was higher in the e-waste
soil samples than the control site though statistically insig-
nificant. The concentration of metals in the e-waste site was
found to be in the order of Ni > Cu > Pb > Zn > Cd. The con-
centrations of heavy metals in the e-waste soil samples were
found to be many folds higher than the permissible values by
WHO; however, it was statistically insignificant (Table 2).
Therefore, AAS results establish the presence of heavy
metals in the soil samples of the e-waste, which could be
one of the probable causative agents for the genotoxicity in
Allium cepa roots exposed to e-waste leachates. Our findings
show trends similar to the ones reported from other e-waste
dumpsites though the concentration of the heavy metals was
found to be less as compared to other such e-waste dump-
sites (Bakare et al. 2013; Zhang et al. 2012). One of the
reasons for low concentrations of heavy metals could be the
presence of organic matter in soil which when combined
with the toxic metal cations, reduces the availability of the
heavy metals (Beck et al. 1993). The burning of e-waste
to minimize the volume as well as to extract the precious
metals leads to the leakage of heavy metals and toxins into
the environment, which is evident by the presence of these
heavy metals in the soil samples.
The leaching of heavy metals and other hazardous
e-waste-associated chemicals can contaminate the ground-
water and rivers, thereby posing serious threat to the
International Journal of Environmental Science and Technology

Fig. 7  Chromosomal aberrations observed in Allium cepa root cells in anaphase, e, f sticky chromosomes, g anaphase bridge, h C-mito-
grown in various e-waste leachates treatments. a, b Vagrant chromo- sis, i simulated e-waste treatment-induced Ghost cells—no viable
somes and laggards, c chromosome fragment, d spindle abnormalities cells were observed in e-waste treatment, j normal mitotic cell

Table 2  Distribution and concentration of heavy metals (ppm) observed in the soil samples from control and e-waste sites using Atomic Absorp-
tion Spectroscopy (AAS)

Soil sample Cd Cu Ni Pb Zn
Mean ± SE Mean ± SE Mean ± SE Mean ± SE Mean ± SE

Control 0.00845 ± 0.02005 1.7873 ± 0.078 1.67845 ± 0.42375 − 1.0088 ± 0.904 7.348 ± 2.2618

E-waste 0.3395 ± 0.1998 22.0413 ± 13.2196 63.0374 ± 58.0076 19.7175 ± 14.3591 13.58555 ± 1.24515
Maximum permissible 0.005 0.05 0.5 0.05 5
limit (WHO)
*p value 0.891519 0.028563 0.219752 0.449895 0.488097
**p value 0.342781 0.34457 0.476089 0.401477 0.091689
***p value 0.574052

Bold value refers to statistically significant (P < 0.05)

*p value: difference between the means of control group versus maximum permissible limit
**p value: difference between the means of experimental group versus maximum permissible limit
***p value: difference between the means of the control and the experimental groups

13

environment and to human populations. There are conflict-
ing reports regarding the presence of heavy metals in the
water samples and soil samples in Philippines. Sia (2008)
reported no significant difference between the heavy metal
concentration (Cr, Pb, Cd) in the groundwater samples of
control and Payatas site, Manila; however, the presence of
heavy metals such as lead and cadmium in excess of allow-
able limits set by the US Environmental Protection Agency
was reported in community tap water samples of Manila
(Solidum and Solidum 2010). High fluctuation in the con-
centration of the heavy metals in the surface and sediment
samples from the Manila Bay was reported suggesting the
inflowing rivers to be the source of pollution (Prudente et al.
1994). The possible source of these heavy metals could be
the leaching out of the heavy metals from these e-waste
recycling sites and the e-waste dumpsites as suggested by
our findings where one of the e-waste dumpsites is located
close to the Estero De Vitas, which is connected to the other
water bodies in Manila. Since these heavy metals can bio-
accumulate in the aquatic organisms and fish is one of the
main components of the Filipino diet, further investigations
of the soil leachates from e-waste dumpsites located in other
areas such as Cavite and Laguna should be carried out not
only to detect the presence of heavy metals but also to detect
the background contamination at various locations far away
from landfills and industrial zones.
AAS of hair samples
The Atomic Absorption Spectroscopy results confirmed
the presence of heavy metals in the hair samples of the
informal e-waste recyclers (Table 3). Surprisingly, the lev-
els of all heavy metals tested (Cd, Cu, Ni and Zn), except
Pb, in the hair samples of the control group were found to
Table 3  Distribution and concentration of heavy metals (ppm) in the hair samples of control group and informal e-waste recyclers
Hair Sample Cd Cu
Mean ± SE Mean ± SE
Control 0.663442857 ± 0.104718 0.480714286 ± 0.120576
E-waste 0.974928571 ± 0.05572 0.369228571 ± 0.065058
Maximum permissible 0.06 0.1
limit (WHO)
*p value 0.001191 0.019628
**p value 0.000003 0.00609
***p value 0.25751
*p value: difference between the means of control group (per metal) vs maximum permissible limit
**p value: difference between the means of experimental group (per metal) versus maximum permissible limit
***p value: difference between the means of the control and the experimental group
13
International Journal of Environmental Science and Technology
be above the reference values set by the WHO at p < 0.05.
Levels of all heavy metals in rank order according to their
means were: Zn > Ni > Cd > Cu > Pb for both the control and
experimental groups. The concentrations of heavy metals
found in the hair samples of the informal e-waste recyclers
were comparable to the concentrations in the hair samples
of e-waste recyclers reported in other studies (Wang et al.
2009). However, statistically, there was no significant dif-
ference between the concentrations of heavy metals in the
hair samples of informal e-waste recyclers when compared
with the control group not exposed to e-wastes (p < 0.05).
Both the control and the e-waste recycler groups showed
the concentration of the heavy metals many folds higher
than the permissible limits recommended by WHO with the
exception of Pb, which was absent in the samples of both
the control and the e-waste recycler groups. The presence
of heavy metals in the control group far in excess than the
permissible limit suggests the presence of these metals in the
environment and the workplace of these subjects. Our results
indicate the absence of Pb in the scalp hair of the control and
the informal e-waste recyclers though some other studies
conducted in Philippines have reported high levels of blood
lead (Pb) in the adult women (Zhang et al. 1998) and chil-
dren (Sharma and Reutergardh 2000). The anomaly in our
results could be due to small sample size and the influence
of other socio-demographic factors such as gender-specific
differences, consumption of vitamins/mineral tablets and
herbal concoctions, use of indoor lead paints, unsafe hair
and cosmetic products and exposure to pollution.
The presence of heavy metals and other toxins in the hair
samples is the consequence of direct exposure to these tox-
ins. The acute and chronic exposure to the e-waste toxic-
ity has been well documented in many studies from China
(Wang et al. 2009; India (Eguchi et al. 2012) and Ghana
Ni Pb Zn
Mean ± SE Mean ± SE Mean ± SE
1.430785714 ± 0.204886 − 1.887542857 ± 0.106622 4.972085714 ± 0.737191
0.995285714 ± 0.044538 − 1.262414286 ± 0.702921 3.312142857 ± 0.478714
0.1 15
0.000002 0.00001
0.10069 0
International Journal of Environmental Science and Technology
(Asante et al. 2012). In Philippines, as anywhere else, the
management of e-waste is carried out by formal sector and
informal sector. According to the Department of Environ-
ment and Natural Resources, 23 registered facilities formally
known as Treatment, Storage and Disposal (TSD) are sup-
posedly engaged in handling e-waste; however, most of it
is indirect where it is a part of the industrial or solid waste
(Yoshida et al. 2016). The informal sector consists of more
than 2300 registered and unregistered junkshops with the
Metropolitan Manila Development Authority; however,
there are numerous scavengers, waste pickers, women and
children who are engaged in e-waste recycling although no
reliable estimate regarding the actual numbers is available
as of now. Our study is the first in Philippines that focuses
on the evaluation of genotoxicity of e-waste using labora-
tory data (Allium cepa assay) and establishes the correlation
between the possibility of its hazardous impact on humans
by determining the presence of heavy metals in the soil sam-
ples of the e-waste dumpsites as well as in the hair samples
of recyclers. Though our investigations involved careful
selection of the informal recyclers after consultations and
confirmation of the relevant information about the recyclers
from various stakeholders such as the Barangay chief, some
NGO workers active in the e-waste dumpsites, the study
also suffers from the limitation of the inability to exclude
other such contributory factors toward the genotoxic effects
induced by e-waste. However, the presence of heavy metals
in the soil and the hair samples of both the control group
and the recyclers confirms the possibility of the chronic
exposure to the heavy metals and their possible role in pos-
ing health risk to the human subjects. Further investigations
using AAS and comet assay are being carried out to increase
the sample size, including the gender-specific difference and
occupational link criteria for the control group to arrive at a
baseline value for the frontline heavy metals including lead
(Pb) in the hair and blood samples of the general public as
well as in the informal and formal e-waste recyclers.
Conclusion
The inhibition of root growth, mitotic index and the induc-
tion of chromosomal aberrations in the Allium cepa root
meristems exposed and grown in the e-waste leachates col-
lected from the e-waste dumpsites located in and around
Metro Manila are indicative of the genotoxic potential of
the e-waste. Our results confirm the sensitivity of the Allium
cepa to the hazardous chemicals present in the e-waste tox-
ins; hence, this plant assay system can be recommended to
be used for environmental biomonitoring of e-waste toxicity
to carry out preliminary screening. The AAS results con-
firmed the presence of five heavy metals tested—cadmium,
copper, nickel, lead and zinc in the soil samples from the
three e-waste recycling and e-waste dumpsites more than the
control sites as well as the standard permissible limits. The
presence of heavy metals in the hair samples establishes the
occupational hazards and the possibility of long-term effects
of toxicity of these heavy metals on the health of the control
group as well as the e-waste recyclers. In our investigation,
the site 2 was deliberately selected for our study as burn-
ing of e-waste is one of the common practices often used
by informal recyclers to minimize the volume of e-waste
and also to extract precious metals in the Philippines. Our
interaction during the interview with the informal recyclers
also revealed that most of the recyclers were aware of the
presence of these precious metals and their value. Based
on our results, it is recommended that the practice of burn-
ing to manage the e-waste should be strongly discouraged
to minimize the toxic effects of e-waste to the humans and
environment. Just like Philippines, many ASEAN (Asso-
ciation of South East Asian countries) member countries
like Vietnam, Cambodia have shown rapid increase in the
volume of e-waste generation. Malaysia, Hong Kong, China
and Singapore are engaged in the import of e-waste from the
European Union to be used as source of secondary raw mate-
rials (Ibitz 2012). It is important that these ASEAN coun-
tries should engage in drafting a common policy response to
address the issue especially that can control the transbound-
ary movement of e-waste among these countries. To formu-
late the guidelines and proper legislation, it is of critical and
vital importance that more investigations are carried out at
other e-waste locations and reliable and sensitive in vitro
and in vivo assays are designed and tested to detect the full
impact and mechanism of the genotoxicity of e-waste. We
propose collaboration and coordination between different
research groups of these countries where similar experi-
ments and assay systems can be devised to generate data
that can be shared and investigated to come up with common
framework to address this issue in this entire region.
Acknowledgements  We would like to thank Ms. Anita Celdran, Direc-
tor, Sustainable Project Management (SPM), Philippines, and Ms. Joyet
Castor, Project Manager and Community Liaison, (SPM, Philippines)
for their continuous support and help in our field survey and identify-
ing the e-waste recyclers located at Tondo, Manila, Dr. Frumencio Co,
Mathematics Department, De La Salle University, for his help in carry-
ing out the statistical analysis of the results and Dr. Maria Carmen Tan,
Mr. Michael Dominic Ajero and Mr. Irving Chiong from the Chemistry
Department for their assistance in carrying out the AAS. This study
was funded by the University Research Coordination Office (URCO),
De La Salle University, Project No-59 FU3TAY14-3TAY15 with Dr
Zeba F. Alam as Principal Investigator.
13
 International Journal of Environmental Science and Technology

Appendix I

Questionnaire

13
International Journal of Environmental Science and Technology

13
 International Journal of Environmental Science and Technology

13
International Journal of Environmental Science and Technology

13
 International Journal of Environmental Science and Technology

13
International Journal of Environmental Science and Technology

References Ibitz A (2012) Environmental policy coordination in ASEAN: the case

of waste from electrical and electronic equipment. ASEAS – Aus-
trian J South-East Asian Stud 5(1):30–51
Alam Z (2016) The assessment of the of e-waste management gener-
Jinhui L, Huabo D, Pixing S (2011) Heavy metal contamination of
ated from cellular phones, laptops, and personal computers in the
surface soil in electronic waste dismantling area: site investi-
Philippines. Manila J Sci 9:27–42
gation and source-apportionment analysis. Waste Manag Res
Asante K, Agusa T, Biney C, Agyekum W, Bello M, Otsuka M, Itai
29(7):727–738
T, Takahashi S, Tanabe S (2012) Multi-trace element levels and
Li H, Yu L, Sheng G, Fu J, Peng P (2007) Severe PCDD/F and PBDD/F
arsenic speciation in urine of e-waste recycling workers from
pollution in air around an electronic waste dismantling area in
Agbogbloshie, Accra in Ghana. Sci Total Environ 424:63–73
China. Environ Sci Technol 41(16):5641–5646
Babatunde B, Anabuike F (2015) In vivo cytogenotoxicity of electronic
Lundgren K (2012) The global impact of e-waste: addressing the chal-
waste leachate from iloabuchi electronic market, Diobu, Rivers
lenge. International Labour Organization. http://ilo.org/wcmsp​5/
State, Nigeria on Allium cepa. Challenges 6(1):173–187
group​s/publi​c/—ed_dialo​gue/—secto​r/docum​ents/publi​catio​n/
Bakare A, Pandey A, Bajpayee M, Bhargav D, Chowdhuri D, Singh
wcms_19610​5.pdf. Accessed 16 Mar 2017
K, Murthy R, Dhawan A (2007) DNA damage induced in human
Pant D, Joshi D, Upreti M, Kotnala K (2012) Chemical and biologi-
peripheral blood lymphocytes by industrial solid waste and munic-
cal extraction of metals present in e-waste: a hybrid technology.
ipal sludge leachates. Environ Mol Mutagen 48(1):30–37
Waste Manag 32(5):979–990
Bakare A, Alabi O, Gbadebo A, Ogunsuyi O, Alimba C (2013) In vivo
Peter O, Eneji I, Ato R (2012) Analysis of heavy metals in human hair
cytogenotoxicity and oxidative stress induced by electronic waste
using atomic absorption spectrometry (AAS). Am J Anal Chem
leachate and contaminated well water. Challenges 4:169–187
3:770–773
Beck A, Jones K, Hayes M, Mingelgrin U (1993) Organic substances
Prudente M, Ichihashi H, Tatsukawa R (1994) Heavy metal concentra-
in soil and water: natural constituents and their influences on con-
tions in sediments from Manila Bay, Philippines and inflowing
taminant behavior. R Soc Chem, Cambridge, p 7
rivers. Environ Pollut 86(1):83–88
Caravanos J, Clark E, Fuller R, Lambertson C (2011) Assessing worker
Sharma K, Reutergardh L (2000) Exposure of preschoolers to lead in
and environmental chemical exposure risks at an e-waste recycling
the Makati area of Metro Manila, the Philippines. Environ Res
and disposal site in Accra, Ghana. J Health Pollut 1:16–25
83:322–332
Celik T, Aslanturk O (2006) Antimitotic and anti-genotoxic effects
Sia Su (2008) Assessing the effect of a dumpsite to groundwater quality
of Plantago lanceolata aqueous extract on Allium cepa root tip
in Payatas, Philippines. Am J Environ Sci 4(4):276–280
meristem cells. Cell Mol Biol (Noisy-le-grand) 61:693–697
Sia Su, Martillano K, Alcantara T, Ragragio E, De Jesus J, Hallare
Cortez L, Ching J (2014) Heavy metal concentration of dumpsite soil
A, Ramos G (2009) Assessing heavy metals in the waters, fish
and accumulation in Zea mays (corn) growing in a closed dump-
and macro invertebrates in Manila Bay, Philippines. J Appl Sci
site in Manila, Philippines. Int J Environ Sci Dev 5(1):77–80
Environ Sanit 4(3):187–195
Dodd G (2010) Heavy metal poisoning and hair analysis. Holist Vet.
Solidum J, Solidum G (2010) Assessment and remediation of heavy
https​://holis​ticve​tpetc​are.net/pdf/Heavy​_Metal​_Posio​n_Hair_
metals in community tap water from Manila, Philippines. In:
Analy​sis-2.pdf. Accessed 16 Mar 2017
International conference on environment science and engineer-
Eguchi A, Nomiyama K, Devanathan G, Subramanian A, Bulbule K,
ing IPCBEE, vol 32, pp 1–6
Parthasarathy P, Takahashi S, Tanabe S (2012) Different profiles
Tedesco S, Laughinghouse H (2012) Bioindicator of genotoxicity: the
of anthropogenic and naturally produced organohalogen com-
Allium cepa test. In: Srivastava J (ed) Environmental contamina-
pounds in serum from residents living near a coastal area and
tion. InTechOpen. https​://doi.org/10.5772/31371​
e-waste recycling workers in India. Environ Int 47:8–16
United Nations University (2013) StEP launches interactive world
Fiskesjö G (1988) The Allium-test an alternative in environmental stud-
e-WasteMap. U.S. environmental protection. http://unu.edu/media​
ies: the relative toxicity of metal ions. Mutat Res 197:243–260
-relat​ions/relea​ses/step-launc​hes-inter​activ​e-world​-e-waste​-map.
Grant W (1982) Chromosome aberration assays in Allium. A report of
html#info_. U.S. Environmental Protection Agency (USEPA).
U.S. Environmental Protection Agency Gene-Tox Program. Mutat
Accessed 16 Mar 2017
Res 99(3):273–291
Wang T, Fu J, Wang Y, Liao C, Tao Y, Jiang G (2009) Use of scalp hair
Gutierrez R, Agarrado G (2011) The vanishing e-wastes of the Phil-
as indicator of human exposure to heavy metals in an electronic
ippines. Ban Toxics. http://banto​xics.org/v2/image​s/downl​oads/
waste recycling area. Environ Pollut 157(8–9):2445–2451
ewast​e_repor​t.pdf. Accessed 16 Mar 2017
Yoshida A, Terazono A, Ballesteros F, Nguyen D, Sukandar S, Kojima
M, Sakata S (2016) E-waste recycling processes in Indonesia, the

13
 International Journal of Environmental Science and Technology

Philippines, and Vietnam: a case study of cathode ray tube TVs Zhang W, Hua W, Marie-Odile S (2012) Soil contamination due to
and monitors. Resour Conserv Recycl 106:48–58 e-waste disposal and recycling activities: a review with special
Zhang Z, Subida R, Agetano M, Nakatsuka H, Inoguchi N, Watanabe focus on China. Pedosphere 22(21):434–455
T, Shimbo S, Higashikawa K, Ikeda M (1998) Nonoccupational
exposure of adult women in Manila, the Philippines, to lead and
cadmium. Sci Total Environ 215:157–165

13