Environmental Research 158 (2017) 118–125

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Environmental Research
journal homepage: www.elsevier.com/locate/envres

Electrokinetic – Enhanced ryegrass cultures in soils polluted with organic MARK

and inorganic compounds
Gustavo Acosta-Santoyoa, Claudio Cameselleb, , Erika Bustosc
⁎

a
Center of Research and Technological Development in Electrochemistry (CIDETEQ), Pedro Escobedo 76703, Mexico
b
Department of Chemical Engineering, University of Vigo, Rua Maxwell s/n, Building Fundicion, 36310 Vigo, Spain
c
Center of Research and Technological Development in Electrochemistry (CIDETEQ), Pedro Escobedo 76703, Mexico

A R T I C L E I N F O A B S T R A C T

Keywords: The effect of electric fields on seed germination and development of ryegrass (Lolium perenne L.) was studied in
Perennial ryegrass clean and contaminated soil with heavy metals and/or PAHs. The application of 0.2 DCV/cm in clean soil near
Phytoremediation ryegrass seeds enhanced the germination by 75%. The presence of contaminants in soil hindered the germination
Electric field and growing of ryegrass. However, the application of DC electric field favored the germination and growing of
Direct current
plants compensating the negative effects of the contaminants. The electrode material in anodes has a decisive
Alternate current
influence in the germination and growing of ryegrass. Stable anode materials have to be used to avoid the release
of toxic ions in the soil that affect the development of the plant. Graphite anodes are very appropriate because
they are inexpensive and does not generate toxic effects on plants. The electro-phytoremediation of mixed
contaminated soil with ryegrass showed very promising results, especially AC electric fields. The tests with AC
current showed the highest biomass production in a treatment of 1 month. The more biomass production the
more removal of heavy metals and PAHs from soil.

1. Introduction concentration of contaminants (due to the phytotoxicity of con-

Soil is the habitat for plants, microorganisms and animals. The
biological activity of the organisms in soil has an enormous influence in
soil fertility, and hence in the production of crops that are the base of
the human feeding. The rapid industrialization and the bad manage-
ment of wastes had led to a serious contamination of soil. In general,
the main contaminants in soil are toxic heavy metals (chromium, cad-
mium, mercury, lead,…) and a series of hydrophobic organics (hydro-
carbons, PAHs, PCBs, solvents, pesticides,…). Several remediation
technologies have been proposed to palliate the contamination in soil
(Sharma and Reddy, 2004). Among them, phytoremediation have been
proposed as a benign technology that is able to deal with organic and
inorganic contaminants in soil and groundwater (Batty and Dolan,
2013). Phytoremediation shows several advantages: it can be applied in
situ over large areas, the cost of operation and control is low, and the
soil do not undergo significant damages during remediation. In fact
phytoremediation tend to stimulate the biological activity of soil unlike
other physicochemical technologies. The main drawbacks of phytor-
emediation are related with the long treatment time required to achieve
the remediation goals (the remediation depends on the growth and
biological cycles of the plant), it is limited to low or moderate
taminants at increasing concentrations), and the remediation depth is
limited to the rhizosphere (Sarwar et al., 2017).
The coupled technology electrokinetics-phytoremediation have
been proposed as an attempt to palliate in part the limitation of phy-
toremediation (Cameselle et al., 2013). The application of an electric
field near a growing plant may enhance the remediation capacity of the
plant by mobilization of nutrients and contaminants that will be more
available for plant uptake. The first author in describing the effects of
the electric current on green plants was Lemstrom (1904) who reported
that plants exposed to an electric field (DC) grew greener and produced
more biomass that plants cultured in the same conditions but the
electric field. O'Connor et al. (2003) published the first study about the
influence of a constant DC electric field (30 V; 18 cm electrode gap) in
the phytoremediation of a heavy metal contaminated soil with ryegrass.
These authors concluded that the electric field transported the metals
towards the cathode but there was not accumulation of heavy metals on
the cathode side because ryegrass accumulated the mobilized metals in
its tissues. The pH changes induced in the soil by the application of the
electric field (acid pH in the anode side and alkaline pH in the cathode
side) affected the development and survival of the ryegrass in the soil
near the electrodes. Cang et al. (2011) reported that low potential drops

⁎
Corresponding author.
E-mail addresses: gacosta@cideteq.mx (G. Acosta-Santoyo), claudio@uvigo.es (C. Cameselle), ebustos@cideteq.mx (E. Bustos).

http://dx.doi.org/10.1016/j.envres.2017.06.004

0013-9351/ © 2017 Elsevier Inc. All rights reserved.

Received 15 February 2017; Received in revised form 5 May 2017; Accepted 7 June 2017
G. Acosta-Santoyo et al.
(2 V/cm) favor the growing of the plant and, as a consequence, the
phytoremediation capacity. However, high potential drop (4 V/cm)
affected negatively to the plant because of the phytotoxicity caused by
the pH changes in soil and the increasing bioavailability of heavy me-
tals. High electric potentials also decreased the activity of soil micro-
flora (Cang et al., 2012). Aboughalma et al. (2008) and Bi et al. (2011)
proposed the use of alternate current instead of direct electric current.
These authors hypothesized the AC electric fields would enhance the
plant growth and phytoremediation capacity, avoiding the problems
associated with pH changes in soil because the alternate current (AC)
does not provoke these changes. However, the experimental results in
the phytoremediation tests were not as good as they expected
(Aboughalma et al., 2008; Bi et al., 2011).
The literature reports on the coupled technology electrokinetics-
phytoremediation stand out the benefits of the electric field on the plant
growth and phytoremediation capacity (Cameselle et al., 2013). How-
ever, the literature also proved the negative effects of an intense electric
field in the growing of plants and soil microflora activity (Cang et al.,
2012), as well as the minor effect in plant and phytoremediation if a
weak electric field is applied (Chirakkara et al., 2015). The variety of
test conditions in literature reports makes difficult the selection of the
most appropriate operating conditions in electro-phytoremediation
applications. The main operating conditions are: the strength of the
electric field (voltage drop: 0.5–4 V/cm); the type of current (DC, AC);
the mode of application (continuous, periodic, polarity inversion, etc.)
Furthermore, there is very limited information available in the litera-
ture about the influence of the electric field in the germination and
early stages of plant development (Acosta-Santoyo et al., 2016).
In general, the literature results proved the beneficial effects of the
electric fields in the growing of green plants and their phytoremedia-
tion capacity. However, different authors report very different condi-
tions in terms of type of electric current (AC, DC), mode of application
(continuous, periodic, polarity inversion,…), strength of the electric
field, electrode materials and others. Moreover, most of the studies uses
plants that were grown for 1 month or more before the application of
the electricity, and there is not information available about the influ-
ence of the electric current in the early stages in the developing of the
plants.
In this study, the authors try to elucidate how the application of an
electric field near a growing plant may affect its germination, devel-
opment and remediation capacity. Perennial ryegrass (Lolium perenne
L.) was selected because its fast growth and remedial capacity
(O'Connor et al., 2003; Zhou et al., 2007). The study evaluate the
germination of ryegrass in clean and contaminated soil using different
electrode materials. The anode material may release ions in the inter-
stitial fluid that may affect the plant. Graphite, stainless steel and alu-
minum were selected as anode materials based on its availability and
relatively low cost. The remediation capacity of the ryegrass was stu-
died in mixed contaminated soil with AC or DC electric fields and with
polarity inversion to compare the benefits of each type of current and
mode of operation with the objective to define the most appropriate
operating conditions for large scale applications.
2. Materials and methods
2.1. Plant species
Perennial ryegrass (Lolium perenne L.) seeds were collected directly
from wild populations in an agricultural area in the surroundings of the
city of Vigo (NW Spain). This species is cultivated as pasture and forage
plant. The use of a local variety of perennial ryegrass assures the ac-
climation and surveillance of the plant to local weather and soil con-
ditions used in the tests. Seeds were harvested in June 2015 and were
stored in a fresh and dry place until use.
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Environmental Research 158 (2017) 118–125
2.2. Soil
Soil was sampled in an agricultural field in the surroundings of Vigo
(Spain). The soil characteristics corresponded with a Humic cambisol.
Soil was sampled from the upper layer (between 0 and 0.2 m) in three
different adjacent sites. Then, the soil was thoroughly mixed to obtain a
single sample that was stored in the lab in plastic containers. Soil was
extended in a thin layer of 2–5 cm to let it dry for 72 h. Then, the soil
was sieved through a 2 mm mesh to remove small rocks, roots, seeds
and other possible non-soil components. Soil was stored at 4 °C in the
dark until use.
2.3. Contamination of soil with PAHs and heavy metals
Soil was contaminated in the lab with two PAHs: Anthracene and
phenanthrene; or three heavy metals: Pb, Cr and Cd. For PAHs con-
taminated soil, the necessary amounts of the PAHs for a final con-
centration in soil of 50 mg/kg of anthracene and 100 mg/kg of phe-
nanthrene were weighted and dissolved in 1 L of acetone. The solution
was mixed with 15 kg of dried soil and it was thoroughly mixed. The
soil was periodically mixed until acetone completely evaporated, to
assure a uniform distribution of the PAHs in the soil. For heavy metal
contaminated soil, the necessary amounts of heavy metal salts (Pb
(NO3)2, K2Cr2O7 and CdCl2) for a final concentration in soil of 500 mg/
kg of Pb, 200 mg/kg of Cr and 50 mg/kg of Cd were weighted and
dissolved in 1 L of DI water. This solution was added to 15 kg of dry soil
and it was thoroughly mixed. The soil was let to dry at room tem-
perature while it was periodically mixed to assure a uniform distribu-
tion of the heavy metals. 5 kg of PAH contaminated soil and 5 kg of
heavy metal contaminated soil were mixed to obtain a soil sample with
mixed contamination with the following concentrations: Anthracene,
25 mg/kg; phenanthrene, 50 mg/kg; Pb, 250 mg/kg; Cd, 25 mg/kg and
Cr, 100 mg/kg. Soil was stored in the dark at 4 °C until use.
2.4. Germination tests with DC electric current
The effect of the direct application of DC electric field in the ger-
mination of perennial grass was studied in clean soil saturated in
moisture with tap water (41% of moisture). The initial soil pH was 5.5
and the electric conductivity was 114.8 μS/cm. The germination tests
were carried out in pots with 1.5 kg of moisten soil. 52 seeds of per-
ennial ryegrass were uniformly distributed in the surface of the soil,
separated 2 mm from each other. Seeds were introduced 1–2 mm below
the soil surface. The electrodes used to apply the electric field were
inserted 10 cm deep in the soil on both sides of the pots. The distance
between anode and cathode was 15 cm. The anode was a sheet of
graphite 2 cm wide, and the cathode was a titanium rod of 6 mm of
diameter. Graphite sheet was used because is inexpensive and readily
available, moreover the sheet can be cut to the size and shape required
in each application. The graphite sheet was cut 2 cm wide to have a
similar surface area (20 cm2) than the electrode rods (19 cm2). Right
after the perennial ryegrass was sown, a constant electric field of 0.2
DCV/cm was applied for 3 h. Then, the pots were placed in a growth
chamber with artificial light in a long-day photoperiod (16 h light: 8 h
darkness) at 25 ± 2 °C and the seeds were led to germinate for 7 days
(Fig. 1).
2.5. Plant growth tests with electric current
The same experimental setup used in the germination tests was used
to determine the influence of the electric current in the development of
perennial ryegrass (Fig. 1). Pots with germinated perennial ryegrass
were kept in a growth chamber with artificial light with a long-day
photoperiod (16 h light and 8 h darkness) at 25 ± 2 °C for 3 weeks. The
electric current was applied to the pots using a graphite sheet anode
(2 cm wide, 10 cm length) and two different cathodes: graphite sheet or
G. Acosta-Santoyo et al. Environmental Research 158 (2017) 118–125

Fig. 2. Experimental setup for the growth of Lolium perenne L. in contaminated and un-
contaminated soil under the effect of electric fields.

2.7. Soil analysis

Fig. 1. Experimental set up for the germination and growth of Lolium perenne L. with
1.5 kg of soil (uncontaminated and contaminated) under the effect of electric fields.
Heavy metal concentration in soil was determined by acid digestion
following the EPA method 3050, using 5 g of dry soil and nitric and
titanium rod (6 mm diameter, 10 cm length). Direct current (DC), al- hydrochloric acids. After the digestion, the supernatant was filtered and
ternate current (AC) and direct current with polarity inversion (DC-PI) the metal concentration was determined by ICP/OES. PAHs in soil were
were applied to the different pots and the results were compared with a extracted mixing 1 g of dry soil with 3 mL of cyclohexane for 30 min in
control test with no current. In all the tests with electric current, 1 V/ an ultrasonic bath. The extraction procedure was done twice. Then,
cm was applied for 4 h twice a day (4 h on to 8 h off). In the DC-PI tests, cyclohexane was evaporated with a flow of nitrogen and the extracted
the polarity of the electric current was inverted every time electric PAHs were dissolved in 1 mL of ethanol. PAH concentration was then
current was applied, but despite the inversion of the polarity, the anode measured by HPLC with a UV–vis diode array detector (Kayali-Sayadi
material was always graphite, even in the tests with graphite-titanium et al., 2000).
electrodes. In the DC-PI tests, two pairs of electrodes (graphite-tita-
nium) were used to avoid the electrode of titanium to become the anode 3. Results and discussion
when the polarity was inverted. The plant height was followed during
the treatment. At the end of the tests, the roots and stem of each plant 3.1. Germination and growing of perennial ryegrass with electric current
were collected, and the fresh and dry biomass were determined.
The germination of perennial ryegrass in non-contaminated soil can
be enhanced by the application of a DC electric field right after the
2.6. Ryegrass growth amended with electric fields in contaminated soils sowing of the seeds. In these tests, a constant DC electric field of 0.2 V/
cm was applied to the culture pots for 3 h using electrodes made of
The influence of the electric field in the germination and growth of graphite (anode) and titanium (cathode). As it is shown in Fig. 3, the
perennial ryegrass was studied in clean and contaminated soil with pots with electric field showed a germination percentage of 77%;
PAHs and/or heavy metals. In these tests, each pot was filled with 200 g compared with only 44% of germination in the pots with no electric
of dry soil. The soil was saturated with tap water (moisture: 41%) be-
fore sowing ryegrass seeds. The initial soil pH was 5.5 and the electric
conductivity was 115 μS/cm. The electrodes were inserted in the soil on
both sides of the pot. The distance between anode and cathode was
10 cm. The seeds were planted 1–2 mm below soil surface, in 5 rows
and 5 columns (25 seeds per pot) in the area between the electrodes
(Fig. 2). The cathode was made of titanium. Three different materials
were tested as anode: aluminum, graphite and stainless steel. A constant
DC electric field was applied for 3 h to the main electrodes right after
sowing ryegrass. The voltage gradient ranged from 0 to 0.8 V/cm in the
different tests. Then, the pots were placed in a growth chamber with
artificial light in a long-day photoperiod (16 h light: 8 h darkness) at
25 ± 2 °C and the germination and development of the plant was re-
gistered for 3 weeks.
Fig. 3. Effect of DC electric field in the germination of ryegrass seeds (0.2 V/cm for 3 h,
anode: graphite, cathode: titanium).

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G. Acosta-Santoyo et al.
Fig. 4. Germination of perennial ryegrass amended with DC electric field (0–0.8 V/cm).
current.
The germination and growing of ryegrass with electric current was
assessed in clean and contaminated soils with heavy metals and/or
PAHs. Titanium was used as cathode and three different materials:
aluminum, graphite and stainless steel; were tested as anode. Fig. 4
shows the germination percentage of all the tests after one week of
sowing. In general, the application of a low DC electric field to the
culture pots had a clear benefit in the germination of ryegrass. In clean
soil, graphite anode improved the germination up to 80% at 0.2 V/cm.
Similar behavior was found for stainless steel, whereas aluminum an-
odes hindered the germination probably because of the release toxic
Al3+ to the soil. No significant changes in Al concentration or pH were
observed in the soil at the end of the tests. The pots were watered
periodically to maintain the moisture of the soil, and this may mask pH
or Al changes during the germination and growing of the plant. The
presence of contaminants in soil clearly affected the germination of
ryegrass. The electric field was able to stimulate the germination
reaching percentages as high as 80% in mixed contaminated soil with
0.2 V/cm with graphite anode. In general, the best germination results
were found with graphite anodes with a voltage gradient of 0.2 DCV/
cm. Stainless steel showed the better germination values at 0.2 DCV/cm
but germination ratio was not as good as with graphite. Aluminum
showed an interesting behavior: the germination percentage increased
with the voltage gradient in contaminated soil showing the best results
at 0.8 DCV/cm. Fig. 5 shows the plant height after 3 weeks of growing.
In clean soil, the maximum height of plants was registered at 0.2 VDC/
cm voltage gradient with very similar values for the three anode ma-
terials. The contaminants in soil reduced the growing and developing of
the ryegrass but the electric treatment was able to compensate the
toxicity of the heavy metals and PAHs. Thus, the plant height in some
experiments was even higher than in the control tests. The best results
in contaminated soil were found for voltage gradient ranged from 0.2 to
0.8 VDC/cm. In general, the best results corresponded to graphite and
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Environmental Research 158 (2017) 118–125
stainless steel anodes. These are stable electrodes in the operating
conditions in the tests. No corrosion was observed in the stainless steel
anode, and graphite showed no significant changes in its structure.
These results suggest that the negative effect of the contaminants in
seed germination and plant development could be overcome with the
electric treatment, although the electrode materials and voltage gra-
dient need to be tested for each specific application.
The results of this study proved that the DC electric current shows a
significant influence on the germination of plants. The application of
0.2 VDC/cm for 3 h was enough to boost the germination ratio from
44% to 77% in uncontaminated soil (Fig. 3). This result is in agreement
with a previous study by Acosta-Santoyo et al. (2016) and Cuevas et al.
(2016) who found that the germination of Arabidopsis thaliana was
enhanced when the seeds were exposed to 0.2 VDC/cm. The electric
field also improved the growth of the plantlets in the early stages of
development. These authors claim that the electromigration towards
seeds and roots of essential ionic species is the very reason for the en-
hanced germination and development of the plant. On the other hand,
Acosta-Santoyo et al. (2016) used stable materials in the electrodes (Ti
cathode, IrO2-Ta2O5 anode). Thus, the electric current did not increase
the concentration of harmful ions in the interstitial fluid. In order to
evaluate the possible influence of the electrode materials in the ger-
mination and development of the ryegrass, three anode electrodes were
tested in this work. It has been proved that Al anodes releases harmful
Al+3 ions to the soil that affect the germination and development of
ryegrass (Figs. 4 and 5). Furthermore, the acid environment in the
anode increased the phytotoxicity of aluminum (Kunhikrishnan et al.,
2016). Graphite and stainless steel did not show any significant change
under the operating conditions in this study, although it is expected
some corrosion of these electrodes at longer treatment times. Anyway,
graphite is not expected to show any negative effect in the soil or plant,
and the release of ferric ions is not going to affect the plants as long as
the soil pH was higher than 4 (Zahid et al., 2009). Ferric ions
G. Acosta-Santoyo et al. Environmental Research 158 (2017) 118–125

Fig. 5. Plant height of perennial ryegrass amended with DC electric field (0–0.8 V/cm).

precipitate as hydroxide at that pH (Morel and Hering, 1993), thus the

possible risk of phytotoxicity of the ferric iron is reduced to a minimum.
The germination of ryegrass and its development have been very
affected by the presence of contaminants in soil (Figs. 4 and 5) due to
phytotoxic effects of heavy metals and PAHs. On the other hand, the
application of a constant DC electric field have increased the germi-
nation of ryegrass (Fig. 3). It has been hypothesized that the benefits of
the electric field may overcome the negative effects of the con-
taminants. This hypothesis have been proved in the tests shown in
Figs. 4 and 5. The germination rate and the height of plant was im-
proved with the presence of a DC electric field, but the optimum DC
potential drop was rather different for contaminated and clean soil. The
best growing results of ryegrass in contaminated soil were achieved
with 0.2–0.8 DCV/cm whereas the clean soil only requires 0.2 DCV/cm.
There is not additional information in literature about this topic. As Fig. 6. Effect of DC, AC and DC with polarity inversion (PI) on ryegrass growing with
Acosta-Santoyo et al. (2016) claims, the benefits of the electric field periodic application of electric current (4 h on: 8 h off). Anode: graphite, cathode: tita-
may be related to the electromigration of beneficial ionic species (nu- nium.
trients,…) towards the roots of the growing plants, and that beneficial
effect may compensate the phytotoxic effect of the contaminants.
However, the intrinsic mechanism is not completely understood. Fur-
ther research is necessary to evaluate the benefits of the electric field in
contaminated soil with different plants and various levels of con-
taminants.

3.2. Implications of electric current on phytoremediation with ryegrass

Fig. 6 shows the effects of the electricity in the growing and de-
veloping of ryegrass in uncontaminated soil. In this set of tests, 1 V/cm
of direct current (DC), alternate current (AC) and DC with polarity in-
version (PI) were applied to the pots periodically: 4 h on-8 h off. Gra-
phite was used as anode and titanium was used as cathode. Only the DC
tests showed a clear enhancement of the height of ryegrass by 60%. Fig. 7. Effect of DC, AC and DC with polarity inversion (PI) on ryegrass growing with
Minor influence showed the AC and PI tests with only an increase of periodic application of electric current (4 h on-8 h off). Anode and cathode: graphite.
height of 4% and 10% respectively.

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G. Acosta-Santoyo et al.
Fig. 8. Phytoremediation of mixed contaminated soil with ryegrass. Effect of DC, AC and
DC with polarity inversion (PI) on plant height (periodic application of electric current:
4 h on-8 h off; Anode and cathode: graphite).
The influence of electric field (DC, AC and PI) in the development of
ryegrass was also tested with graphite electrodes in anode and cathode
in uncontaminated soil (Fig. 7). The use of graphite electrodes in anode
and cathode is very interesting for large-scale applications considering
the high cost of titanium. Graphite is cheap, ready available and does
not release ions to the interstitial fluid when used as anode. Other
anode materials can release ions that could be toxic for growing plants
and soil microflora. In these tests (Fig. 7), the plants with electricity
showed better growing compared to the test with no current. The plant
height increased by 43% (DC) 23% (AC) and 17% (PI) for the three
types of electric current/mode of application. The better results in the
tests with graphite electrodes (compared with the tests with graphite-
titanium) can be explained by the uniform distribution of the electric
field in the pot when using two sheets of graphite instead of a rod of Ti
(López-Vizcaíno et al., 2017).
The ryegrass growing amended with electricity was also tested in
mixed contaminated soil (Fig. 8). In the 1st and the 2nd week, the DC
test showed the best growing of ryegrass, similar to the tests with clean
soil (Fig. 7), but at longer treatment time, the AC test showed the
highest development of plants. These results are in agreement with
Chirakkara et al. (2015) that reported an increase in the total biomass
by 28% for the oat plant and 13% for the sunflower in the tests with AC
electric current compared to the control test with no electricity. Bi et al.
(2010) studied the growth of lettuce in hydroponic medium. The ap-
plication of AC electric field resulted in higher biomass production
compared to the test with no electricity in Cd contaminated and non-
contaminated medium. Bi et al. (2011) reported an enhanced biomass
production of rapessed (Brassica napus) in the presence of AC electric
field whereas tobacco plant (Nicotiana tabacum) was not affected. They
claim that the better biomass production of rapeseed (Brassica napus)
when growing in the presence of the AC electric current is due to three
possible factors as it is reported by Bi et al. (2011). Firstly, an increment
in the water of soil due to a bidirectional electro-osmosis flow induced
by the AC electric field. Secondly, an enhanced ionic inflow in the roots
by periodic hyperpolarization and depolarization. Thirdly, the meta-
bolic activity of the plants could be influenced at both the extracellular
and intracellular level as a response of interaction between the AC
electric fields with the ions in the interstitial fluid. Considering the type
of pots used in this study and their periodic watering, the first me-
chanism about the electroosmotic flow in the soil is not likely to exert
any significant influence; so, the explanation of the higher biomass
production in the AC tests compared to the control has to be associated
to the biological/biochemical alterations induced by the electric field.
Fig. 9 shows the biomass production at the end of the tests (4
weeks), expressed as fresh weight (the weight of the stem as collected
from the pot) and dry weight (the weight of the stem after drying at
room temperature for 7 days). All the tests with electric current en-
hanced the growing of plants and the synthesis of biomass, but in this
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Environmental Research 158 (2017) 118–125
Fig. 9. Phytoremediation of mixed contaminated soil with ryegrass. Effect of DC, AC and
DC with polarity inversion (PI) on biomass production (periodic application of electric
current: 4 h on-8 h off; Anode and cathode: graphite).
case, the AC tests resulted in much better biomass production than any
other test. This is a very interesting result considering the possible
application of the electricity amended plant culture for the production
of biomass or phytoremediation applications. Thus, in phytoremedia-
tion, it is accepted that the contaminant removal can be improved with
the biomass production. The difference between the fresh and dry
weight (Fig. 6) is the water content in the plant biomass, and that was
almost constant in all the tests. The water content ranged from 89.5%
(in the PI tests) to 91.7% (in the AC tests and in the control tests
without electricity). These results confirmed that electricity favored the
growth of the plant and the production of biomass, and this biomass
production was not due to water storage in the plant tissue.
Figs. 10 and 11 shows the PAH and heavy metal in soil after the
electro-phytoremediation treatment with ryegrass for 28 days. The re-
sidual heavy metal concentrations in soil showed in Fig. 10 corre-
sponded to the average value of the soil samples from each pot. The
error bars corresponds to the standard deviation. The blank columns in
Fig. 10 corresponded to a control test with no plant and no current. In
general, minor concentrations of the three heavy metals were removed
from soil, probably due to the short treatment time. There is not sig-
nificant differences among the three metals. Pb, Cd and Cr are all toxic
metals with no known biological activity. This is one of the reason for
their acute toxicity and the limited uptake by the plant. The experi-
ments with electricity showed better removal of heavy metals form soil
than the blank test. Moreover, the removal of metals can be correlated
with the biomass production (Chirakkara and Reddy, 2015). Thus, the
AC test showed an average removal of heavy metals around 15%
whereas the concentrations of heavy metals only decreased by 2% in
the test with no electricity. The DC and IP test showed an intermediate
value of 5% and 4% respectively. These results suggest the capability of
L. perenne to remove toxic heavy metals from soil. The removed heavy
metals were accumulated in the roots of the plant and no metals were
Fig. 10. Residual heavy metals in soil in phytoremediation of mixed contaminated soil
with ryegrass.
G. Acosta-Santoyo et al.
Fig. 11. Residual PAHs in soil in phytoremediation of mixed contaminated soil with
ryegrass.
detected in the shoots. Other species from the genus Brassica are well-
known for their capacity to accumulate heavy metals and translocate
them from roots to the shoots. The limited removal results in this study
is explained by the short treatment time. Overall, the results suggest
that the more biomass production the more heavy metal removal, so the
phytoremediation with L. perenne amended with AC electric fields
seems to be a promising process for the remediation of the con-
taminated soil.
Fig. 11 shows the residual concentration of PAH in soil at the end of
the tests. There is a clear removal of the PAHs in the soil planted with L.
perenne. The best removal results corresponds with the AC test, al-
though the differences among all the planted test are relatively small.
The blank test with no current and no electricity also showed a sig-
nificant removal of PAH compared to the initial concentration in soil
(Anthracene, 25 mg/kg; Phenanthrene, 50 mg/kg). This result can be
associated to the organic matter content in the soil and the microbial
activity that may degrade the PAHs (Johnsen et al., 2005). In phytor-
emediation, the removal or degradation of organic contaminants can be
achieved by phytodegradation, phytovolatization, accumulation in the
plant tissues or rhizodegradation (Reddy et al., 2017). The first three
mechanisms imply the uptake of the organic contaminants by the plant.
The absorption of organics is mainly by diffusion through the root cell
membranes, and the diffusion is controlled by the hydrophobicity of the
organic contaminant. The two PAHs tested in this study are very hy-
drophobic with an octanol partition coefficient above 3 (Log Kow of
naphthalene = 3.37; Log Kow of phenanthrene = 4.46). The ideal oc-
tanol partition coefficient for the diffusion of organics into the cells
roots is from 0.5 to 3 (De Maagd et al., 1998). This indicates that is not
likely a phytodegradation of the PAHs in the soil. So, the most probable
mechanism for the reduction in PAH concentration in the soil is the
dissipation/volatilization and the microbial degradation in the rhizo-
sphere but not plant uptake. Phenanthrene seemed to be easier to re-
move than anthracene, and the planted test with AC electric current
showed one of the lower residual concentration. Thus, the AC electric
field appears to be a promising technology for the remediation of mixed
contaminated soils based on the remediation results and the enhanced
biomass production.
4. Conclusions
The use of DC electric field in the vicinity of seeds of ryegrass has a
positive effect in the germination ratio and in the development of the
plant. The same positive effect was found in clean and contaminated
soil with heavy metals and/or PAHs. Contaminated soil tend to hinder
the germination and development of the plant but the application of
electric fields favor the germination and growing of the plant com-
pensating the negative effect of the contaminants. It is supposed that
the mobilization and electromigration of essential ions (nutrients…) are
the responsible mechanisms for the enhanced germination and
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Environmental Research 158 (2017) 118–125
growing. Anyway, further research is necessary to elucidate the in-
trinsic mechanism associated to the interaction of the electric fields and
the growing plants. The electrode materials for the application of the
electric field in soil have a decisive influence in germination and de-
velopment of the plant. Anodes has to be made of stable materials that
do not let toxic ions in the soil.
The phytoremediation of mixed contaminated soil with ryegrass
showed very interesting perspectives with AC electric fields. The al-
ternate current favored the production of biomass and as a result, sig-
nificant amounts of heavy metals and PAHs were removed from soil.
Further experiments at longer treatment time are necessary to stablish
the capacity of ryegrass amended with AC electric field as a practical
remediation technology.
Acknowledgements
Authors would like to thank to the Consejo Nacional de Ciencia y
Tecnología of Mexico (CONACyT, 264785) by the financial support to
develop this research. Gustavo Acosta Santoyo would also like to thank
CONACyT for the grant received to accomplish his doctoral degree at
CIDETEQ and the grant for international mobility and the University of
Vigo for the facilities provided during this project.
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