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International Journal of Engineering & Technology IJET-IJENS Vol:14 No:05 1 144305-7171-IJET-IJENS © October 2014 IJENS I J E N S AbstractPhytoremediation under greenhouse condition was investigated as an alternative to clean up an industrially- multi-contaminated soil with both petroleum hydrocarbons and heavy metals. The aim of this work was to study the ability of three sunflower (Helianthus annuus) cultivars (M, H2, and P) to absorb/degrade V, Ni, Cu, Pb, benzo(a)pyrene, and TPH (total petroleum hydrocarbons) from multi-contaminated soil. Assays were performed with three sunflower cultivars, suitable for commercial biodiesel production, in pots containing 5 kg of soil for a period of 40 days. All three varieties were able to reduce the concentration of heavy metals, benzo(a)pyrene, and TPH in the soil, however, the contaminants removal varied according to the assay conditions employed. The highest removal percentage of Ni, Pb, and Cu were obtained for H2 cultivar. On the other hand, the highest removal percentage of V was obtained when the soil was treated with M cultivar. TPH removal did not vary according to the use of different sunflower cultivars. Nevertheless, the highest removal percentage of benzo(a)pyrene was obtained with the M cultivar. The results demonstrated the potential of the phytoremediation technique using sunflower for the treatment of soil multi-contaminated by heavy metals and hydrocarbons petroleum. Index Termheavy metals, hydrocarbons, multi- contaminated soil, phytoremediation, sunflower. I. INTRODUCTION The growth of industrial activity in the last century has resulted in a strong increase in anthropogenic substances being introduced into the environment [1]. The accidental or intentional emission of chemical products and wastes can lead to soil pollution, which is a significant problem in modern economic activities, such as in transport and civil construction. It is important to note that appropriate land use appears in the agenda of international discussions due to the increase in population, as well as the need for sustainable development as a strategic safety tool. Furthermore, soil contamination is a concern since it can negatively affect human, animals and plants health [2]. The current world economic model is strongly tied to the use of fossil fuels as energy source. Given that large quantities are necessary, the increase in the number of movements, transformations, and storage operations can also increase the possibility of soil contamination. It is important to remember that petroleum is chemically defined as a mixture of chemical compounds, predominantly hydrocarbons, which also contains heavy metals, semi-metals, and various anions from organic material and rocks from formations and reservoirs [3]. The presence of inorganic material in fuel mixtures and oily wastes is also due to the use of additives, catalysts, and other compounds. Thus, it is clear that the presence of petroleum, petroleum derivatives, and their wastes in the soil can lead to contamination by organic and inorganic compounds. Considering the importance of soil, various techniques, such as excavation, solidification, stabilization, and bioremediation, have been used to decrease the impact of contaminants on the soil environment [4]. Comparatively, operations based on biological processes are considered to be more efficient, less costly, and the least invasive. Biotechnology alternatives, however, are strongly related to the degradation of organic compounds in soil or their solubilization to facilitate extraction processes [5]. Among the biotechnological alternatives, phytoremediation is considered to be low-cost and not harmful to the physical, chemical, and biological characteristics of soil [6]. This technique consists of using plants to remove biota contaminants (phytoextraction), to absorb and convert them into non-toxic forms (phytovolatilisation) or to stabilize an inorganic substance, transforming it into a less soluble form (phytostabilisation) [7]. There are a number of reports that can be found in the literature regarding the use of plants to remove organic or inorganic compounds from soil. Helianthus annuus (sunflower) is one of the target species that has excellent potential as a phytoextractor since it grows quickly, produces large amounts of biomass and is capable of hyper accumulate heavy metals [8; 9]. Many plants have been studied for the phytoremediation of hydrocarbons from soil [10; 11; 12; 13; 14]. However, only Tejeda-Agredano et al. [15] have related the use of sunflower to the phytoremediation of organic contaminants. Many wastes from the petroleum and gas sector have both hydrocarbons and heavy metals, which can contaminate soil Phytoremediation of Soil Multi-Contaminated with Hydrocarbons and Heavy Metals Using Sunflowers Cristiane D. C. Martins 1 ; Vitor S. Liduino 1 ; Fernando J. S. Oliveira 2 ; Eliana Flávia C. Sérvulo 1* 1 Escola de Química, Universidade Federal do Rio de Janeiro UFRJ, Av. Athos da Silveira Ramos, 149, Bloco E Sl E-203 - Ilha do Fundão, RJ - Brasil CEP 21941-909. 2 Petróleo Brasileiro S.A. Gerência de Meio Ambiente. Av. Almirante Barroso 81, Centro, Rio de Janeiro, RJ 20031-004. *Corresponding author: Email: [email protected]

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Page 1: Phytoremediation of Soil Multi Contaminated with ... · phytoremediation of a soil collected from a contaminated area within a petroleum refinery to remove both hydrocarbons and heavy

International Journal of Engineering & Technology IJET-IJENS Vol:14 No:05 1

144305-7171-IJET-IJENS © October 2014 IJENS I J E N S

Abstract— Phytoremediation under greenhouse condition

was investigated as an alternative to clean up an industrially-

multi-contaminated soil with both petroleum hydrocarbons and

heavy metals. The aim of this work was to study the ability of

three sunflower (Helianthus annuus) cultivars (M, H2, and P) to

absorb/degrade V, Ni, Cu, Pb, benzo(a)pyrene, and TPH (total

petroleum hydrocarbons) from multi-contaminated soil. Assays

were performed with three sunflower cultivars, suitable for

commercial biodiesel production, in pots containing 5 kg of soil

for a period of 40 days. All three varieties were able to reduce the

concentration of heavy metals, benzo(a)pyrene, and TPH in the

soil, however, the contaminants removal varied according to the

assay conditions employed. The highest removal percentage of Ni,

Pb, and Cu were obtained for H2 cultivar. On the other hand, the

highest removal percentage of V was obtained when the soil was

treated with M cultivar. TPH removal did not vary according to

the use of different sunflower cultivars. Nevertheless, the highest

removal percentage of benzo(a)pyrene was obtained with the M

cultivar. The results demonstrated the potential of the

phytoremediation technique using sunflower for the treatment of

soil multi-contaminated by heavy metals and hydrocarbon’s

petroleum.

Index Term— heavy metals, hydrocarbons, multi-

contaminated soil, phytoremediation, sunflower.

I. INTRODUCTION

The growth of industrial activity in the last century has

resulted in a strong increase in anthropogenic substances being

introduced into the environment [1]. The accidental or

intentional emission of chemical products and wastes can lead

to soil pollution, which is a significant problem in modern

economic activities, such as in transport and civil construction.

It is important to note that appropriate land use appears in the

agenda of international discussions due to the increase in

population, as well as the need for sustainable development as

a strategic safety tool. Furthermore, soil contamination is a

concern since it can negatively affect human, animals and

plants health [2].

The current world economic model is strongly tied to the

use of fossil fuels as energy source. Given that large quantities

are necessary, the increase in the number of movements,

transformations, and storage operations can also increase the

possibility of soil contamination. It is important to remember

that petroleum is chemically defined as a mixture of chemical

compounds, predominantly hydrocarbons, which also contains

heavy metals, semi-metals, and various anions from organic

material and rocks from formations and reservoirs [3]. The

presence of inorganic material in fuel mixtures and oily wastes

is also due to the use of additives, catalysts, and other

compounds. Thus, it is clear that the presence of petroleum,

petroleum derivatives, and their wastes in the soil can lead to

contamination by organic and inorganic compounds.

Considering the importance of soil, various techniques, such

as excavation, solidification, stabilization, and bioremediation,

have been used to decrease the impact of contaminants on the

soil environment [4]. Comparatively, operations based on

biological processes are considered to be more efficient, less

costly, and the least invasive. Biotechnology alternatives,

however, are strongly related to the degradation of organic

compounds in soil or their solubilization to facilitate

extraction processes [5].

Among the biotechnological alternatives, phytoremediation

is considered to be low-cost and not harmful to the physical,

chemical, and biological characteristics of soil [6]. This

technique consists of using plants to remove biota

contaminants (phytoextraction), to absorb and convert them

into non-toxic forms (phytovolatilisation) or to stabilize an

inorganic substance, transforming it into a less soluble form

(phytostabilisation) [7]. There are a number of reports that

can be found in the literature regarding the use of plants to

remove organic or inorganic compounds from soil.

Helianthus annuus (sunflower) is one of the target species

that has excellent potential as a phytoextractor since it grows

quickly, produces large amounts of biomass and is capable of

hyper accumulate heavy metals [8; 9]. Many plants have been

studied for the phytoremediation of hydrocarbons from soil

[10; 11; 12; 13; 14]. However, only Tejeda-Agredano et al.

[15] have related the use of sunflower to the phytoremediation

of organic contaminants.

Many wastes from the petroleum and gas sector have both

hydrocarbons and heavy metals, which can contaminate soil

Phytoremediation of Soil Multi-Contaminated

with Hydrocarbons and Heavy Metals Using

Sunflowers

Cristiane D. C. Martins1; Vitor S. Liduino

1; Fernando J. S. Oliveira

2; Eliana Flávia C. Sérvulo

1*

1Escola de Química, Universidade Federal do Rio de Janeiro – UFRJ, Av. Athos da Silveira Ramos, 149, Bloco

E Sl E-203 - Ilha do Fundão, RJ - Brasil CEP 21941-909. 2 Petróleo Brasileiro S.A. Gerência de Meio Ambiente. Av. Almirante Barroso 81, Centro, Rio de Janeiro, RJ –

20031-004.

*Corresponding author: Email: [email protected]

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International Journal of Engineering & Technology IJET-IJENS Vol:14 No:05 2

144305-7171-IJET-IJENS © October 2014 IJENS I J E N S

with these two classes of contaminants when the contact or

improper disposal of wastes occurs. It is also important to

emphasize that the literature does not focus on the

phytoremediation of soils co-contaminated with metals and

hydrocarbons. One of the few studies in this field was

published by Sun et al. [16], where Tagetes patula was used to

phytoremediate soil that was artificially contaminated with

benzo(a)pyrene, Cd, Cu, and Pb.

This study analyses the potential of sunflowers for the

phytoremediation of a soil collected from a contaminated area

within a petroleum refinery to remove both hydrocarbons and

heavy metals (V, Ni, Cu, Pb, and Cd). Seeds from three

different, low-cost, commercial cultivars of sunflower were

tested. The 40 days long tests were conducted in greenhouse

scale.

II. MATERIALS AND METHODS

A. Soil

Soil from a contaminated area in a Brazilian petroleum

refinery was used. Approximately 60 kg of multi-

contaminated soil was dried, sifted (<2 mm), and

homogenized before the physical, chemical, and

microbiological analyses were performed to be later used in

the phytoremediation tests.

B. Plants

Three commercial sunflower cultivars (M, H2, and P) were

tested (Dekalb Ltda, Brazil). The growth of the plants was

monitored weekly.

C. Analytical methods

The particle size distribution of the soil were determined

according to the IAC [17], and the pH of the soil was

determined in a 1:1 soil:water suspension (w w-1

) using a

digital pH meter.

The total amounts of nitrogen, phosphorous, and organic

matter were determined according to the methodologies

proposed by USEPA 351.2, USEPA 365.2 [18], and Walkley

and Black [19], respectively. The soil’s water-holding

capacity was determined using the method described by

Veihmeyer and Hendrickson [20].

The concentration of the total petroleum hydrocarbon

(TPH) was determined through the USEPA 8015C method

[18]. Ten priority polycyclic aromatic hydrocarbons (PAH)

described by the Dutch list (naphthalene, phenanthrene,

anthracene, fluoranthene, benzo(a)anthracene, chrysene,

benzo(k)fluoranthene, benzo(a)pyrene, benzo(ghi)pyrene, and

indeno(1,2,3cd)pyrene) were determined by gas

chromatography coupled with mass spectroscopy using the

methodology described by USEPA 8270D [18]. The organic

extracts were obtained using an ultrasound according to the

method described in USEPA 3550C [18].

The concentration of heavy metals in the soil was

determined by spectroscopy using the methodology described

in USEPA 6010C [18]. The digestion was performed

previously in an acidic media using a microwave in

accordance with the USEPA 3051A method [18].

The metal content in the plant biomass was determined by

the methodology described above. Previously, the plant

material was dried in an oven at 60ºC, ground in a porcelain

mill, subjected to acidic digestion, and later analyzed by

atomic absorption spectroscopy.

The total heterotrophic bacteria (THB) and total fungi (TF)

in the soil were determined using the plate counting and the

pour-plate methods [21] in nutrient agar and Sabouraud agar,

respectively. The quantification of hydrocarbon-degrading

microorganisms (HDM) was performed through the Most

Probable Number technique in Bushnell Haas mineral media

with the addition of a drop of light Arabian oil as the only

carbon source [22].

All of the analytical results are reported as the average of

three replicas for all the microbiological, physical, and

chemical assays.

D. Phytoremediation tests

The experiments were conducted in greenhouse scale

(Figure 1). The seeds were germinated directly in the multi-

contaminated soil. The pots used in the tests had a surface area

of 600 cm2. Five kilograms of soil were added to each pot, and

equidistant furrows were made in the soil, with four seeds

being deposited into each hole. When at least one of the

seedlings in each hole reached a height of 10 cm, the plants

were thinned to one plant per hole.

Fig. 1. Greenhouse’s views.

The phytoremediation tests were conducted for 40 days,

during which the soil moisture was maintained at

approximately 80% of its water-holding capacity.

Each soil sampling procedure consisted of taking five

individual samples at equidistant points in each pot to obtain a

representative sample. Individual samples of 50 g of soil each

were mixed to obtain a single sample of 250 g. For each test,

the sampling procedure was performed in triplicate.

At the end of 40 days, the heights of the plants were

measured and the dry weight of the plant biomass was

determined with an analytical balance. The plant drying

process was performed in an oven at 50ºC for three days. All

of the results from the phytoremediation tests were expressed

as the average of three replicas.

The results consider the losses from the biotic and abiotic

control tests. The control tests were performed under the same

conditions, where the abiotic control was made without

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International Journal of Engineering & Technology IJET-IJENS Vol:14 No:05 3

144305-7171-IJET-IJENS © October 2014 IJENS I J E N S

planting sunflowers, and the biotic control, the soil in pots

were treated with an AgNO3 solution (10% w w-1

).

E. Statistical analysis

A two-way analysis of variance (ANOVA) was used in

order to understand the effect of the different sunflower

cultivars in the phytoremediation process. Comparisons of

means were done using the Tukey’s HSD test (p≤0.05). All

statistical analyses were performed using STATISTICA

software v. 7.0 (STATSOFT, USA).

III. RESULTS AND DISCUSSION

A. Soil

Table 1 shows the results of the physical, chemical, and

microbiological characteristics of the multi-contaminated soil

used in this study. The soil was sandy-silty with a neutral pH

in water. The water-holding capacity was low, which is

typical for tropical soils with low clay content. Soil

phosphorous and, especially, N can serve as nutritional

sources for the biota.

Lead, copper, nickel, and TPH were present in the soil at

concentration values above the intervention limit set by local

legislation for industrial used soils with low clay content. The

local legislation uses the same intervention values listed in the

Dutch Legislation [23]. The sum of the concentration of the

10 PAHs listed by the Dutch legislation (naphthalene,

phenanthrene, anthracene, fluoranthene, benzo(a)anthracene,

chrysene, benzo(k)fluoranthene, benzo(a)pyrene,

benzo(ghi)pyrene and ideno(1,2,3cd)pyrene) were found to be

below the intervention values according to the current local

legislation.

Copper was the only metal present at a concentration higher

than the intervention value listed in federal legislation [24].

Benzo(a)pyrene was near the limit of the intervention

concentration (3.5 mg kg-1

), and the other nine monitored

PAHs exhibited concentrations below the limit of detection of

the samples, which was 0.33 mg kg-1

. Therefore, the dataset

justifies the treatment of this soil.

The amount of hydrocarbon-degrading microorganisms

(HDM) was similar to the total number of heterotrophic

bacteria. This fact corroborates the data of increased

concentration of petroleum hydrocarbons and PAHs in the

soil, which can be used as a carbon source by these

microorganisms. The natural selection of the native microbiota

is probably due to the area contamination history by oily

wastes.

The population of fungi found in the soil sample was

elevated prior to the phytoremediation treatment, which also

contributes to the degradation of organic compounds.

According to the lab-scale study published by Atagana et al.

[25], fungi showed the potential to degrade high-molecular-

weight polycyclic aromatic hydrocarbons and other

recalcitrant organic compounds due to their complex

enzymatic systems with the ability to synthesize and excrete

various extracellular enzymes.

Table I

Physical, chemical, and microbiological composition of multi-contaminated

soil

Soil Properties Measurements

Clay (<0.002 mm), % 2

Silt (0.002-0.5 mm), % 33

Sand (0.05-2 mm), % 52

pH (in H2O) 6.81

Water-holding capacity, % 30

Total phosphorus, g kg-1

0.2

Total nitrogen, g kg-1

5.2

Organic matter, g kg-1

129

Cadmium, mg Cd g-1

3.2

Lead, mg Pb g-1

528.1

Copper, mg Cu g-1

684.5

Nickel, mg Ni g-1

109.3

Vanadium, mg V g-1

180.2

10 priority PAH, mg kg-1

5.1

Benzo(a)pyrene, mg kg-1

2.97

TPH, mg kg-1

8.890

THB, CFU g-1

1.8 x 107

TF, CFU g-1

8.7 x 104

HDM, MPN g-1

1.2 x 107

THB: total heterotrophic bacteria; TF: total fungi; HDM:

hydrocarbon-degrading microorganisms.

In the chromatographs of organic extracts from the soils

(Figure 2), compounds were detected mostly as an unresolved

complex mixture (UCM). No linear hydrocarbons, pristane, or

phytane were detected. The absence of pristane and phytane

and the increase in the baseline indicate the weathering of the

oily wastes, increasing their recalcitrance and, consequently,

increasing the difficulty for biotreatment. Weathering refers to

the result of chemical, biological, and physical processes on

the waste that can affect the type of compounds that remain in

the soil. In addition to the described factors, the elevated TPH

concentration makes phytoremediation even more difficult.

Fig. 2. Chromatograph of the total petroleum hydrocarbons present in the

multi-contaminated soil sample from the petroleum refinery.

B. Phytoremediation of multi-contaminated soil

Over the 40 days, all of the cultivated plants showed similar

average heights, measuring approximately 28 3 cm. The

development of the plant height was linear during this time

period. All of the plants had the same number of leaves and

flowers. After the 40 days, the total dry weights of the plants

obtained were 3.08, 4.06, and 2.60 g for the H2, P, and M

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International Journal of Engineering & Technology IJET-IJENS Vol:14 No:05 4

144305-7171-IJET-IJENS © October 2014 IJENS I J E N S

biomasses, respectively. These results were consistent with

those reported in the literature for the treatment of soil

contaminated with different lead concentrations [26].

Results of microbiological analysis of the soil after 40 days

of phytoremediation with sunflowers are represented in Table

2. The results indicate that the presence of the sunflowers was

beneficial to microbial activity, particularly for the fungal

population. Fungi can be related to the release of chemical

substances and enzymes by the plant roots, which favor

microbial growth [27; 28].

The percentage of heavy metals removed from the soil by

the three sunflower cultivars is shown in Figure 3. The largest

percentage of vanadium removal from the soil (39.8%) was

achieved by seeding cultivar P. The removal of nickel and

cadmium was approximately 46 and 100%, respectively, for

the three tested cultivars. The greatest percentage removals of

lead (55.8%) and copper (73.3%) were observed when the H2

cultivar was used.

Usman and Mohamed [29] studied different conditions for

soil phytoremediation by sunflowers for 60 days. They

observed that the Zn removal varied between 2 and 4%, Cu

removal varied between 1.6 and 3%, Cd removal varied

between 2.6 and 4%, and Pb removal varied between 0.4 and

1.3%. The study published by Chen et al. (2004) showed

removals of Pb, Cu, Zn, and Cd of 4, 16, 13, and 23%,

respectively, after 53 days of phytoremediation by sunflowers.

Thus, the promising results for the three tested Brazilian

cultivars were confirmed.

Fig. 3. Removal of V, Ni, Pb, Cd and Cu by the sunflower cultivars (H2, M

and P) in multi-contaminated soil from an industrial area. Values are means ±

SE of three replicates. Different letters indicate significant differences among

the means of different treatments (p<0.05).

The concentrations of heavy metals in the plant biomass

after 40 days of testing are shown in Table 3. There is no

significant difference between the values for vanadium and

cadmium found in the biomasses of the three cultivars. The

greatest concentrations of lead and copper were found in the

biomass of cultivar H2. However, the highest concentration of

nickel was found in the biomass from cultivar M.

c

a

c

a

b b

a a

a

a

a a

b

a

b

0

20

40

60

80

100

120

V Ni Pb Cd Cu

Re

mo

val (

%)

M H2 P

Table II

Soil microbiology after 40 days of sunflower phytoremediation

Cultivar THB (CFU g-1

) HDM (MPN g-1

) TF(CFU g-1

)

H2 5.0 ± 0.2 x 108a 2.0 ± 0.1 x 10

8a 8.6 ± 0.3 x 10

6a

M 2.4 ± 0.1 x 108b 7.8 ± 0.3 x 10

7b 7.1 ± 0.2 x 10

6b

P 1.7 ± 0.1 x 108c 6.9 ± 0.2 x 10

7b 4.3 ± 0.1 x 10

6c

Biotic control 7.2 ± 0.3 x 107 9.5 ± 0.4 x 10

7 7.8 ± 0.3 x 10

4

THB: total heterotrophic bacteria; TF: total fungi; HDM: hydrocarbon-degrading microorganisms; CFU: colony

forming unit; MPN: most probable number. Values are means ± SE of three replicates. Different letters indicate

significant differences among the means of different treatments (p<0.05).

Table III

Heavy metal concentration in plant biomass after 40 days of phytoremediation

Metals (mg kg-1

) Sunflower cultivar

H2 P M

V 0.50 ± 0.01a 0.50 ± 0.01a 0.50 ± 0.01a

Ni 2.61 ± 0.08b 1.72 ± 0.07c 3.43 ± 0.1a

Pb 2.65 ± 0.05a 0.54 ± 0.01c 1.74 ± 0.05b

Cd 0.051 ± 0.010a 0.051 ± 0.010a 0.052± 0.010a

Cu 21 ± 1.5a 14± 1.0b 16± 1.0b

Values are means ± SE of three replicates. Different letters indicate significant differences among the means of

different treatments (p<0.05).

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International Journal of Engineering & Technology IJET-IJENS Vol:14 No:05 5

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Sung et al. [6] studied the accumulation of copper, nickel, and

chromium in various plants, including sunflowers, in

phytoremediation assays with steel tailings. The authors

found concentrations of Cu, Ni, and Cr in the plant tissues

varying between 25-50, 25-80, and 50-80 mg kg-1

,

respectively. Turgut et al. [7] evaluated the accumulation of

Cr, Ni, and Cd in two different cultivars of sunflower, teddy

bear and dwarf sunspot. The two cultivars showed

concentrations of Cr, Ni, and Cd in their plant tissues of

approximately 190; 6.5 and 120 mg kg-1

, respectively. Thus,

it was inferred that the accumulation of metals in plant tissues

is heterogeneous, corroborating the quantitative images of

metals in plant tissues published by Becker et al. [31].

Table 4 shows the removal of benzo(a)pyrene after 40 days

of phytoremediation of multi-contaminated soil by sunflower

cultivars. Benzo(a)pyrene is a high molecular weight PAH,

with a recognized persistence in the soil and half-life values

varying between months to many years [32; 33]. The

metabolites originating from benzo(a)pyrene are considered to

be highly carcinogenic and mutagenic and are classified as

carcinogenic group 1 by IARC [34].

Table IV

Removal of benzo(a)pyrene by sunflower cultivars (H2, M and P).

Initial H2 M P

Concentration

(mg kg-1

soil)

2.97 2.01 ± 0.2 1.55 ± 0.1 2.42 ± 0.2

Removal (%) - 32.1 ± 3.2b 47.7 ± 4.2a 18.3 ± 2.0c

Values are means ± SE of three replicates. Different letters

indicate significant differences among the means of different

treatments (p<0.05).

The largest percentage reductions of benzo(a)pyrene in the

soil were obtained through the cultivation of the M and H2

cultivars, considering the losses in the biotic and abiotic

control reactors. Liste and Alexander [35] performed

phytoremediation tests over 56 days with the Avena sativa and

Brassica napus var. radicola grains, resulting in a reduction in

pyrene of 55.4 and 73.5%, respectively, 18% of which were a

result of the microbial activity and the abiotic losses (control

tests). Through the growth of the herbs Anethumgraveolens,

Capsicum annuum, and Raphanussativus, the same authors

found decreases in the concentration of pyrene varying

between 51.2 and 66.9% after 28 days of phytoremediation, of

which only 6.7% corresponded to losses in the control tests.

Comparing the results now presented in the literature, the

applicability of sunflower cultivars is verified for the removal

of a high-molecular-weight PAH.

Phytoremediation was also investigated for the removal of

TPH after 40 days of sunflower growth on multi-contaminated

soil. In this period, there was a reduction of 10 ± 2% of the

TPH in the soil, regardless of the sunflower cultivar used;

these values were calculated considering the losses in the

control reactors. The petroleum hydrocarbons are a class of

organic compounds in which aromatic compounds, PAH,

alkanes, and other substances that show various

physicochemical properties are included. An increase in the

size of the carbon chain leads to a reduction in the solubility of

the compound and an increase in the partition coefficient in

octanol-water (Kow) and, consequently, a reduction in the

availability of the contaminant to the plants [36].

Siciliano and Germida [37] reported a reduction of

approximately 21% in the concentration of TPH in

phytoremediation tests using forage species. However, these

results were achieved only after 20 months of treatment for

soil contaminated only with hydrocarbons. More recently,

Moreira et al. [38] observed a decrease of 12% in the

concentration of TPH in mangrove sediment also

contaminated with only petroleum hydrocarbons, after 90 days

of phytoremediation with Rizophora mangle L. Thus, the

results in the present study are promising and innovative, as

they demonstrate the concomitant removal of hydrocarbons

and heavy metals in real soil from an industrial area.

Considering Dutch regulations to be among the most

restrictive in the world, the seeding of H2 and M cultivars

provided a reduction in the concentration of heavy metals that

allowed the soil to be classified as rehabilitated for use in an

industrial area (with reference to these parameters). However,

there was insufficient removal of TPH to consider the soil

rehabilitated for use in industrial areas in relation to all of the

analyzed parameters. Nevertheless, considering the low cost

and the short time required, the results prove that the treatment

is promising for the treatment of this type of multi-

contaminated soil.

IV. CONCLUSION

The three tested sunflower cultivars presented the same

growth and biomass under multi-contaminated and non-

contaminated soils. All of them were also capable of removing

benzo(a)pyrene, TPH, and metals from multi-contaminated

soil. The amount of contaminants removed from soil varied

according to the sunflower cultivar used. Sunflower H2, P and

M can hyperaccumulate Cu, Pb, Ni and V from multi-

contaminated soil. Nevertheless, Cd absorption was lower than

other metals. After the treatment with the H2 and M cultivars,

the soil was considered rehabilitated for use in an industrial

area, according to Brazilian federal legislation. Considering

the costs and time required, the phytoremediation with

sunflower proved to be an efficient, convenient, low-cost

process for the treatment of soils contaminated with organic

and inorganic compounds.

ACKNOWLEDGMENTS

The authors thank the Brazilian National Petroleum Agency

(ANP) and Petrobras for providing financial support.

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