Egyptian Journal of Aquatic Research 43 (2017) 185–193

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Egyptian Journal of Aquatic Research

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Full length article

Crude oil spilled water treatment with Vetiveria zizanioides in floating

wetland
Hefni Effendi a,⇑, Ani Munawaroh b, Inna Puspa Ayu b
a
Center for Environmental Research, Bogor Agricultural University (IPB), Indonesia
b
Department of Aquatic Resources Management, Faculty of Fisheries and Marine Science, Bogor Agricultural University (IPB), Indonesia

a r t i c l e i n f o a b s t r a c t

Article history: Floating wetlands on a container (25 L) using Vetiveria zizanioides (L) Nash was applied to remediate
Received 30 December 2016 crude oil spill in water for four weeks. Three plants (height 10 cm) were inserted in a pot containing rock-
Revised 20 June 2017 wool, floated on crude oil spiked water. Treatment included 3 and 6 pot plants with 1% crude oil concen-
Accepted 8 August 2017
tration. Six pot plants during 4 weeks of experiment could reduce the oil content of 91.39%, 84.60% COD,
Available online 24 August 2017
and 84.25% BOD. Meanwhile, 3 pot plants could decrease the oil content of 90.28%, 81.69% COD, and
81.10% BOD. DO at week 4 decreased. The BOD and COD did not show any significant reduction. Plants
Keywords:
grew better in crude oil contaminated water. Those were indicated by 47.22% parent and 100% shoots
Phytoremediation
Crude oil
could live until the study end in 1% crude oil spiked water. Meanwhile, in crude oil uncontaminated
Floating wetland water, 30.56% parent and 40% shoots were able to live until the study end. Wet plant biomass
Vetiveria zizanioides (5.1070 g) in the crude oil contaminated water was greater than the plant biomass (4.8362 g) in crude
oil uncontaminated water. RGR of V. zizanioides in crude oil contaminated water was 5.10 g/week and
4.83 g/week in uncontaminated water.
Ó 2017 National Institute of Oceanography and Fisheries. Hosting by Elsevier B.V. This is an open access
article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Introduction The remediation of oil contaminated soils and waters has

Increased production and distribution of crude oil has been
going on for several years in Indonesia. Oil spills incidence all over
the world is also occasionally occurring. Oil spills often bring about
devastated effects on aquatic and land ecosystems. Consequently
will ruin the aquatic ecosystem homeostasis (Effendi, 2016). Crude
oil consists of variety proportions of hydrocarbon such as alkanes,
aromatics and polycyclic aromatic hydrocarbons (PAHs) as well as
non-hydrocarbon namely sulphur, nitrogen, oxygen and traces of
metals, particularly vanadium, nickel, iron and copper (Brandt,
2003; Wilberforce, 2016).
Each year enormous quantities of crude oil are disposed into the
environment, either deliberate or accident during crude oil produc-
tion and transportation. Between 1.7 and 8.8 million metric tons of
crude oil are estimated being released in the water environment
each year (NRC, 1985), of which >90% as deliberate waste disposal,
directly related to human activities (Ndimele, 2010).
Peer review under responsibility of National Institute of Oceanography and
Fisheries.
⇑ Corresponding author.
E-mail addresses: hefni_effendi@yahoo.com (H. Effendi), 49anie@gmail.com
(A. Munawaroh), innapuspa@gmail.com (I. Puspa Ayu).
become a crucial problem to be solved in several oil producing
countries. Usage of plants as phytoremediator to clean such con-
taminated soils and waters can be applied (Aurangzeb et al.,
2014; Njoku et al., 2009). Oil spill clean-up techniques convention-
ally apply physical and chemical processes that sometimes cause
another negative effect to the aquatic ecosystem. Water bodies
are contaminated, leading to degradation of aquatic lives, uncul-
tivable because of soil fertility loss. Clean-up of such damage is
extremely difficult and cannot be undertaken without substantial
time and monetary commitment (Lin and Mendelssohn, 1998;
Khorphage et al., 2003).
Phytoremediation involving the plant usage to remediate and
detoxify polluted site, appears to be challenging. It is eco-
friendly, cost-effective, and efficient for the abatement and decon-
tamination of hydrocarbon pollution and many other pollutants,
but might take more duration than the conventional technique
because of its natural process (Chappell, 1997; Khan et al., 2000;
Ndimele, 2010; Yakubu, 2007). Oil polluted soil phytoremediation
involves several process namely crude oil uptake from soil or
water, accumulation or processing of crude oil through lignifica-
tion, volatilization, metabolization, mineralization, and enzymes
usage for breaking down complex organic molecules into simpler
molecules (ultimately carbon dioxide and water), furthermore

http://dx.doi.org/10.1016/j.ejar.2017.08.003
1687-4285/Ó 2017 National Institute of Oceanography and Fisheries. Hosting by Elsevier B.V.
This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
186 H. Effendi et al. / Egyptian Journal of Aquatic Research 43 (2017) 185–193
the increases of carbon and oxygen concentration around the roots
could promote microbial or fungal activity (Hong et al., 2001).
The use of grasses for phytoremediation is considered suitable
since they provide rhizosphere zone increase due to their multiple
ramified root systems. Hence, this supports site for more microbial
activity around the root zone (Aprill and Sims, 1990). In the tropics
since climatic conditions favor microbial growth and activity,
bioremediation is frankly a promising technology for coping with
pollution (Merkl et al., 2004). Several plants are able to render a
contaminant in soil harmless via extraction or stabilization, hence
making it unavailable for other creatures and reducing harm to the
environment, in a process called phytoremediation (Cunningham
et al., 1996).
Some types of plants used for phytoremediation of oil contam-
ination including Branchiaria decumbens (Gaskin et al., 2008), Lan-
doltia punctata (Ertekin et al., 2015), Vetiveria zizanioides on land
(Brandt et al., 2006; Nisa and Rashid, 2015), V. zizanioides in water
(Darajeh et al., 2014), Cynodon dactylon and Eleusine indica (Oyedeji
et al., 2013). Aprill and Sims (1990) researched eight prairie grasses
(Andropogon gerardii, Schizachyrium scoparium, Sorghastrum nutans,
Panicum virgatum, Elymus canadensis, Bouteloua curtipendula, Bou-
teloua gracilis and Pascopyrum smithii) in sandy loam soils to deter-
mine whether the degradation of four polycyclic aromatic
hydrocarbons (PAHs) was stimulated by plant growth. Research
on two years plant based soil clean-up using Cymbopogon citratus
was performed at agricultural site spilled with crude oil in Niger
Delta (Nwaichi et al., 2016).
The selection of Vetiver grass (V. zizanioides) as the experimen-
tal plant because of its successful application in diverse soil and
water conservation practices and was regarded to promise a high
potential suitability for phytoremediation (Brandt, 2003). V. zizan-
ioides is originally native to northern India. They grow wildly or
traditionally being cultivated in low, damp wetlands (NRC, 1993).
Vetiver grass has been used for the benefit of human, its aromatic
essential oil content can be extracted from the vetiver’s roots. This
oil could be as an ingredient of a fragrance, medicine or insect-
repellent. The world-wide interest of vetiver nowadays is mostly
aimed at environmental conservation application (Grimshaw,
1990).
V. zizanioides belongs to the grass family (Graminiae) the same
as corn, sorghum, and sugarcane. This plant is the first time used
for the purpose of conservation of soil and water by the World
Bank. V. zizanioides has a strong ecological adaptability and produc-
tivity of large biomass, easy in maintenance and can grow on a
variety of contaminated environmental conditions (Darajeh et al.,
2014).
The research on usage of Vetiver in phytoremediation of oil con-
taminated water in an artificial floating wetland has not been
much reported. Based on this, the goal of this study was to analyze
the ability of V. zizanioides in reducing crude oil spill in water with
artificial floating wetland system in a mesocosm simulation. The
uniqueness of this research was application of V. zizanioides which
is actually a terrestrial plant in artificial floating wetland for treat-
ing oil spill in water.
Materials and method
The study was carried out in February–March 2016, at the Lab-
oratory of Environment, Center for Environmental Research, and
the Laboratory of Productivity and Aquatic Environment, Bogor
Agricultural University (IPB).
Crude oil originated from Oil and Gas Institute (Lemigas)
Jakarta. V. zizanioides was obtained from the cultivation nearby
campus. Containers (45
35
25 cm) were designed for floating
wetland mesocosm. V. zizanioides (10 cm height) was inserted into
a plastic pot (7.5 cm diameter) with rockwool inside. Culture
media of hydroponic nutrient solution (AB mix) was fed into the
water with a ratio of 5 mL/1 L. The research consisted of two con-
trols namely container of oil spiked water only and containers con-
taining plants only. Meanwhile, the treatment consisted of three
and six pot plants with oil concentration of 1% of the total water
volume. Each treatment was triplicates. Oil concentration of 1%
referred to Ertekin et al. (2015) and Kosesakal et al. (2015) and it
was also based on preliminary experiment indicating that 1% crude
oil content did not bring about any prominent visual effect to the
plant. Crude oil was apparently not mixed with the water. How-
ever some fractions of crude oil were dissolved in the water.
Containers were filled with 25 L water and 125 mL hydroponic
nutrients of A and B. After two days, 2.5 ml of crude oil was spiked
into each container. Control and treatment of V. zizanioides were
then floated into the container. Roots submerged into water would
absorb crude oil dissolved in the water (White and Cousins, 2013;
Darajeh et al., 2014) (Fig. 1).
Every week for 4 weeks, water was sampled from the water col-
umn in the middle of container to avoid undissolved crude oil on
the water surface. Several water qualities measurement including
oil and grease content (partition gravimetric method), COD (closed
reflux, colorimetric method), BOD (incubation), DO (DO meter),
temperature (thermometer) and pH (pH meter) according to
APHA (2012), were performed. Determination of oil and grease
applied gravimetric method, extracted by the solvent (APHA,
2012). In addition, measurements of V. zizanioides leaf length was
performed every 2 days. The root and leaf were sliced crosswise
and observed under a microscope.
Plant height and relative growth rate
Plant height measurement was performed every two days for
4 weeks. Measurement was carried out from the stem to the tip
of the highest leaf. The number and height of shoots were also cal-
culated and measured. The number of stem and bud at the begin-
ning and end of the observation was determined to find out the
percentage change (Darajeh et al., 2014).
ab
% Alteration ¼
100
a
where a = initial number, and b = end number.
Relative growth rate was analyzed to determine growth rate of
V. zizanioides (Ertekin et al., 2015).
ðln W 2  lnW 1 Þ
RGR ¼
ðt2  t 1 Þ
where RGR = Relative growth rate (g/week), W1 and W2 = initial
and end weight of biomass, and ðt 2  t 1 Þ = observation duration
(week).
RAL in time and Pearson correlation with SPSS 16.0 software
were used for data analysis. This study applied ANOVA with Dun-
can test to determine the effect of different number of plants and
sampling time on the response of oil and grease content, DO,
COD and BOD. If the significant value of p < 0.05, the diversity
affects very significant on the response. If significant value of
p < 0.10, the diversity significantly affects the response.
V. zizanioides
Pot
Floating stuff 1% spiked crude oil water
Root
Fig. 1. Schematic of experimental design of floating wetland mesocosm.
 H. Effendi et al. / Egyptian Journal of Aquatic Research 43 (2017) 185–193
Linear relationship between two variables was measured by
Pearson correlation. Correlation value ranges from 1 to 1. Strong
positive linear relationship between the variables is indicated by a
value close to 1. Conversely, a value close to 0 addresses that no
relationship between the two variables (Khambete and Cristian,
2014).
Results and discussion
Oil and grease concentration decrease
The declines in oil and grease content in the water from the first
week to the second week on the control, treatment of three pot
plants and six pot plants were 43.70%, 56.07%, and 62.47% respec-
tively. The decline in oil content from week 1 to week 2 was higher
than that of the other weeks (3 and 4), indicating that V. zizanioides
quickly adapted to water environments contaminated by crude oil.
Overall oil content in the water decreased by the end of the obser-
vation. The more plants in containers, the higher the oil content
reduction. The percentage alteration of oil and grease concentra-
tion in the water of 90.29% in 3 pot plants and 91.39% in 6 pot
plants, at the end of 4 weeks experiment, was greater than the
change in the control (84.77%) (Fig. 2a).
Our finding of oil and grease reduction percentage was far much
higher than that the research of Brandt (2003) using Vetiver for oil
contaminated soil remediation. A decreased of 15% oil and grease
content in the medium and 11% oil and grease content in the high
fertilizer medium was observed. The initial oil and grease content
was 5.1% of the total soil dry weight. Oil and grease concentration
decrease was detected in all crude oil contaminated treatments
over 6 months experiment (Brandt, 2003).
In control without plant, oil and grease content also declined.
This is in accordance with Brandt (2003), in oil contaminated soil
without plants, the oil and grease content decrease illustrated
irregular pattern.
Average oil and grease content absorbed by the roots was
2.19 mg/L (3 pot plants) and 2.31 mg/L (6 pot plants). Literatures
reported that THC (Total Hydrocarbon Concentration) content in
0.2
Oil and Grease (mg/L)
0.15
0.1
0.05
0 M6
T0 T1 T2
Sampling me (week)
(a)
COD (mg/L)
Fig. 2. Water quality parameters of (a) Oil and Grease, (b) BOD, and (c) COD. (K = Control/1% crude oil only, M3 = 3 pot plants and 1% crude oil, M6 = 6 pot plants and 1% crude
oil).
187
plant tissues increased in parallel with crude oil concentration in
soil. The oil that has been decomposed and absorbed by plant roots
can serve as nutrients to grow and thrive. V. zizanioides is able to
live and thrive in polluted environment (Oyedeji et al., 2013; Xia
et al., 2004).
A research by Kosesakal et al. (2015) reported that the growth
rate of Azolla filiculoides was not negatively affected by 0.005%
and 0.01% v/v crude oil concentration in the media. A. filiculoides
fresh weight in the control medium after one week cultivation
increased by 117%. The presence of crude oil up to 0.5% v/v reduced
50% plant growth (Kosesakal et al., 2015). However in our research
of 1% crude oil content in the media, no sign of growth retardation
was noticed. The decontamination of oil spiked water by plant
could be through direct uptake of petroleum hydrocarbons into
their tissues (roots and leaves), and direct biochemical transforma-
tion of petroleum hydrocarbons by bacterial consortia living in
root. The process of oil content decrease in this floating wetland
might be analog to the phytoremediation of oil contaminated soil
through phytodegradation and rhizodegradation. In phytodegrada-
tion, contaminants are converted to less harmful substances by
mean of special enzyme process that take up, store and degrade
the oil. The accumulated metabolites are converted into new bio-
mass or released and broken down further by a consortium of
microorganism (ITRC, 1999). In rhizodegradation, plants supply
oxygen to the roots, through the decaying organic material and
the release of exudates, hence the microbial activity around their
roots could be more active (Schnoor et al., 1995). Similar explana-
tion was suggested by (Kathi and Khan, 2011), the successful plant
application in the remediation of a wide range of contaminated
soils, was mainly due to their two ways of function namely creat-
ing favorable conditions for microbial degradation and the crude
oil uptake by roots. Frick et al. (1999) illustrated that petroleum
hydrocarbons uptake by plants occurs at their roots and accumu-
lates small amount in their roots and shoots.
The use of grass for phytoremediation of oil contaminated soil
was also reported by Reilley et al. (1996), the existence of grasses
(tall fescue, sudangrass, and switchgrass) and legumes (alfalfa)
could enhance PAHs (pyrene and anthracene) removal from con-
taminated soils. Planted soils had significantly lower (30–40%)
25
20
BOD (mg/L)
15
K K
10
M3 M3
5
0 M6
T3 T4 T0 T1 T2 T3 T4
Sampling me (week)
(b)
200
150
100 K
50 M3
M6
0
T0 T1 T2 T3 T4
Sampling me (week)
(c)
188 H. Effendi et al. / Egyptian Journal of Aquatic Research 43 (2017) 185–193
PAHs concentrations than the unplanted soils. Hence plant usage
for removal of crude oil pollutant is considered much more effi-
cient than the unplanted applications (Kosesakal et al., 2016).
Meanwhile the research of Frick et al. (1999) summarized pri-
mary mechanisms of plants and microorganisms in petroleum-
contaminated soil and groundwater remediation, namely degrada-
tion, containment, and transfer of the hydrocarbons from the soil
to the atmosphere. In containment process, plants reduce or elim-
inate contaminant bioavailability to other biota. Plant provides
root exudates (enzymes, simple sugars, amino acids, aliphatics
and aromatics) to stimulate the growth of root-associated micro-
organisms (Khan et al., 2000).
BOD is the amount of oxygen required by aerobic microorgan-
isms to degrade organic matter in the water within a certain time.
BOD decreased during four weeks of study. However, by week four
(T4), BOD remained constant, even tended to increase (Fig. 2b).
This shows that V. zizanioides was able to reduce organic material
represented by the lowering BOD in the water optimally for
3 weeks. During the study period, V. zizanioides was able to reduce
BOD by 54.06% (control), 81.10% (3 pot plants), and 84.25% (6 pot
plants). Derajeh et al. (2014) stated that the more number of plants
in containers the higher BOD reduction.
At high BOD, COD tended to be higher as well (Fig. 2b and c).
Despite BOD was generally lower than COD (Alwan et al., 2013).
In the control, COD reduction was 44.38%, in tank spiked by crude
oil was 81.69% (3 pots) and 84.60% (6 pots). The decrease of BOD,
COD and oil content was likely affected by several factors such as
the presence of microbes in the roots of plants that utilized organic
matter as a source of nutrients for growth (Omokeyeke et al.,
2013).
The solubility of oxygen in water is an important factor in sup-
porting the fertility of waters. Main source of dissolved oxygen in
the water is air diffusion and photosynthesis of plants possessing
chlorophyll that live in the waters. Levels of dissolved oxygen
(DO) from the first week to the second week declined, but in the
third week increased and decreased again in the end week. Accord-
ing to Reddy et al. (2015) DO enhancement affected the decrease of
BOD and COD. During the observation period, the DO increased
from the initial of 1.71 mg/L to 8.86 mg/L in the control, in the
oil contaminated water DO reached 6.7 mg/L (3 pot plants), and
3.8 mg/L (6 pot plants). The number of plants in the container
might affect the solubility of oxygen. The more plants, the lower
the DO (Fig. 3a). This was because in the roots of plants in contain-
ers live various microorganisms decomposing organic matter, thus
the oxygen was utilized by microorganisms to degrade organic
matter.
Boonsong and Monchai (2008) mentioned that the DO in the
control (without plant) was higher because of the major factors
(such as microbes) influencing oxygen translocated through the
plant roots of V. Zizanioides. In addition Omokeyeke et al. (2013)
also mentioned that the diffusion of oxygen by photosynthetic
microalgae and wind could cause high DO in control, because there
was nothing blocking the sunlight and the air going into the water
column.
Another parameter that becomes the determining indicators of
water quality are temperature and pH. Temperature affects the
processes occurring in the water. It is closely related to the degra-
dation of organic materials. The higher the water temperature, the
lower the oxygen dissolved in water (Effendi, 2003; Enujiugha and
Nwanna, 2004). Temperatures fluctuated every week (Fig. 3b). The
water temperature during the observation period ranged from
25.86–29.03 °C in oil contaminated water, and 25.08–27.08 °C in
the water without any oil contamination. Fluctuating temperatures
were caused by environmental factors, such as weather, long sun-
shine and rainfall, so the temperature in the water container
adjusted to outdoor conditions (Effendi, 2003). The temperature
was still within the optimum range (30 °C) for the bioremediation
process (Omokeyeke et al., 2013) and the normal range of water
temperature (Effendi et al., 2016).pH during the observation period
fluctuated either in oil contaminated water (6.1–8.5) or in water
with no oil contamination (5.7–7.1) (Fig. 3c). According to
Omokeyeke et al. (2013) the optimum pH for the bioremediation
process ranged 6–8. A decrease in pH in oil treatment, was associ-
ated with the presence of acidic carbon dioxide resulting from bac-
terial activity in the decomposition of hydrocarbons. In the soil
environment, a similar condition occured, in which soil acidity
increased (lower pH) with increasing crude oil concentration in
soils (Ighovie and Ikechukwu, 2014). In crude oil phytoremediation
with Eichhornia crassipes, reduction of turbidity, temperature, pH,
conductivity, and salinity of the water might be due to pollutants
absorption by the plant (Bernard and Blessing, 2013).
Statistical analysis and correlation
Number of pot plants had significant effect on the concentration
of oil and grease in water (p < 0.1). Treatment of 6 pot plants
reduced the average oil and grease concentration from week 0 to
week 4, ranging from 0.151 mg/L to 0.023 mg/L for control, from
0.151 mg/L to 0.015 mg/L for 3 pot plants, and from 0.151 mg/L
to 0.013 mg/L for 6 pot plants. The more number of pot plants in
containers, the more oil content was absorbed by plants (Zhou
et al., 2011). Research duration affected very significant (p < 0.05)
on the concentration of oil in water. This is consistent with the
research of Nisa and Rashid (2015) which stated that the longer
time of the study, the lower oil content reduction in the water.
Interaction between number of pot plant treatment and time had
no significant effect (p > 0.1) on the presence of crude oil in water.
Number of different pot plants had very significant effect
(p < 0.05) on the DO. Average DO control without plant was higher
than that of treatment. The more plants, the lower the DO. This
might relate to natural microbes living in rhizosfer plants. Microor-
ganisms required DO to degrade the oil, and utilized oil as carbon
source and energy (Zulfiqar and Safia, 2012). Research duration
very significantly influenced on the DO in water (p < 0.05). Highest
DO (9.5 mg/L control, 9.4 mg/L M3, and 4.73 mg/L M6) occurred at
week 3. By week 4, DO began to decrease, it was likely used by
microorganisms living in the roots of plants to decompose crude
oil in water. The interaction between the number of pot plants
and the observation time influenced very significantly on DO
(p < 0.05).
Number of pots plant significantly (p < 0.1) affected COD. 6 pot
plants could reduce COD from 145,78 mg/L to 22,44 mg/L, 3 pot
plants from 146,89 mg/L to 26,89 mg/L, and control from
160,22 mg/L to 89,11 mg/L. Observation time had very significant
effect on COD reduction (p < 0.05). The highest COD reduction
occurred in 6 pot plants at week 4. Similar phenomenon was
reported by Darajeh (2014). The interaction between the number
of pot plants and the observation time influenced very significantly
on COD reduction (p < 0.05).
Six pot plants could reduce the BOD up to 3.52 mg/L from the
initiation of 22.350 mg/L accounting for 84.25% reduction consti-
tuting the highest reduction. The interaction between observation
time and number of pot plants had highly significant effect
(p < 0.05) on BOD decrease. Correlation between water quality
parameters (temperature, pH, DO, BOD, oil content) and leaf length
of the plant is presented in Table 1.
The highest correlation (0.88) was indicated by COD and oil and
grease content. The lower the COD, the lower the oil and grease
content in the water. The lowest correlation (0.03) of pH and
BOD means that the relationship did not have closeness. The
negative correlation existed between COD, BOD, oil and grease
content, and DO, although it was not such a close correlation.
 H. Effendi et al. / Egyptian Journal of Aquatic Research 43 (2017) 185–193 189

12 40

Temperature (oC)
10
30

DO (mg/L)
8
6 K 20 K
4 M3 M3
10
2
M6 M6
0 0
T0 T1 T2 T3 T4 T0 T1 T2 T3 T4
Sampling me (week) Sampling me (week)
(a) (b)

10
8
6
K
pH
4
M3
2
M6
0
T0 T1 T2 T3 T4
Sampling me (week)

(c)
Fig. 3. Water quality parameter of (a) DO, (b) Temperature, and (c) pH. (K = Control/1% crude oil only, M3 = 3 pot plants and 1% crude oil, M6 = 6 pot plants and 1% crude oil).

Table 1
Pearson correlation between water qualities and plant leaf length.

Parameter Temperature pH DO COD BOD Oil and Grease Leaf Length

Temperature 1
pH 0.41 1
DO 0.44 0.56 1
COD 0.30 0.38 0.28 1
BOD 0.15 0.03 0.38 0.66 1
Oil and Grease 0.31 0.13 0.43 0.88 0.50 1
Leaf length 0.12 0.18 0.34 0.68 0.61 0.49 1

The similar phenomena was found by Khambete and Cristian
(2014) and Khatoon et al. (2013). High BOD was accompanied by
decreasing DO, because oxygen was used by aerobic bacteria for
survival. Similarly, the higher COD the lower DO. High DO was fol-
lowed by the decline of oil and grease content.
Variable that has directional relationship to the length of the
leaf was DO. Increased DO brought about an effect on leaf length
of V. zizanoides. This was an indication that most organic materials
derived from oil had decomposed into inorganic materials being
used as nutrients by plants.
Oil and grease correlation with leaf length was 0.49. Negative
correlation pointed out that high oil and grease content would be
followed by short leaf length, although the correlation was not
strong. Brandt (2003) also reported a quite similar result namely
a very small Pearson correlation coefficients of oil and grease and
plant root biomass.
anymore, dry out shoots, and rot shoots. The percentage of shoots
number living in the water without contamination of crude oil was
0% (6 pot plants) and 80% (3 pot plants), and in water contami-
nated with crude oil was 100% for both treatment since no death
shoots until the end of observation (Fig. 4). Vetiver cultivated on
crude oil contaminated water tended to grow better, because it
could take advantage of crude oil undergoing decomposition into
nutrients. Brandt et al. (2006) stated that V. zizanioides was a sort
of plant tolerant to adverse conditions. V. zizanioides was able to
120.00
100.00
80.00
60.00 % parent number
%

40.00
Vetiveria zizanioides parent and shoot 20.00 % new shoots
0.00 number
During observation, total number of shoots in water contami-
M3 M6 TM3 TM6
nated crude oil was 10 shoots (3 pot plants) and 25 shoots (6 pot
plants) lived up to the end of observation. Meanwhile, in the water Treatment
without contamination of crude oil, 5 shoots (3 pots of plants) and Fig. 4. Percentage of parent and shoots of V. zizanioides number alive till
1 shoot (6 pot plants) lived. On the water without contamination of experiment end. (M3 = 3 pot plants and 1% crude oil, M6 = 6 pot plants and 1%
the oil existed 1 dead shoots, characterized by budding growing crude oil, TM3 = 3 pot plants only, TM6 = 6 pot plants only).
190 H. Effendi et al. / Egyptian Journal of Aquatic Research 43 (2017) 185–193
live in crude oil contaminated soil, and new buds could emerge. V.
zizanioides reproduces asexually by budding multiplication. In
addition, Brandt et al. (2006) stated that the shoots thriving in
crude oil contaminated soil had low mortality percentage (2%),
even lower than that of plants in crude oil uncontaminated soil
(4%).
Growth of V. zizanioides showed differences in crude oil contam-
inated water and in the water without any crude oil contamina-
tion. As much as 47.22% parent and 100% new shoots could live
until the end of the study in the 1% crude oil spiked media. Mean-
while, in the water without oil contamination, as much as 30.56%
parent and 40% new shoots were able to live until the study end,
which was less than in the oil contaminated media (Fig. 4). This
likely related to the presence of crude oil that could be utilized
by plants as source of nutrients for growth. A similar result was
reported by Bernard and Blessing (2013), morphological parame-
ters (plant height, number of leaves and leaf area) significantly
increased after 2 weeks in oil contaminated soil.
In soil, it was acknowledged that 1% of oil and grease content as
a practical threshold above which PHC (Petroleum Hydrocarbon)
becomes detrimental to plant life (Reis, 1996). In our experiment
of 1% crude oil in water, average relative growth rate (5.10 g/week)
of plants in crude oil contaminated water was greater than the
average relative growth rate (4.84 g/week) of plant in water with-
out crude oil contamination. The successful growth of V. zizanioides
in our research was also supported by the research of Omokeyeke
et al. (2013) who reported that, the interactions between plants
and microorganisms that live in the roots could likely cause an
increase in plant biomass and V. zizanioides tolerant of oil.
In addition, the effect of different levels (0, 2.50, 5.00, 7.50,
10.00, 12.50 and 15.00%) of crude oil contamination on the growth
of Paspalum scrobiculatum, a common weed in Nigeria was investi-
gated by Ogbo et al. (2009). The weed could grow luxuriantly in
crude oil contaminated sites in Delta State, Nigeria. No significant
reduction in the dry weights of the plant in 3 plants per pot and 6
plants per pot treatments of Paspalum scrobiculatum. Phytotoxicity
level strictly depended on the type and constituents of oil.
Kosesakal et al. (2016) reported another result, although the
growth rate of the plants were not negatively influenced by the
presence of 0.005% and 0.01% v/v crude oil in the media, a gradual
impeding effect emerged at concentrations of 0.05–0.1% crude oil
in the media. More than 0.1% crude oil in the media could retard
the growth. As much as 0.2% oil in the media reduced growth
approximately 50%, and the presence of crude oil at concentrations
0.3% or more were lethal.
Wet plant biomass (5.1070 g) in the crude oil contaminated
media was greater than the plant biomass (4.8362 g) in the med-
ium without crude oil contamination. Grasses were regarded to
be superior for phytoremediation due to high level of resistance,
since grasses had extensive, fibrous root systems, and inherent
genetic diversity, quite competitively established in oil contami-
nated soils (Alves et al., 2014; Frick et al., 1999).
Relative growth rate (RGR)
RGR values of V. zizanioides on 3 pot plants and 6 pot plants in
water contaminated with crude oil were 5.10 g/week and 5.11 g/
week, respectively. Meanwhile, RGRs on 3 and 6 pot plants in
water without contamination of crude oil were 4.84 g/week and
4.82 g/week, respectively (Fig. 5). Plant growth in water contami-
nated with crude oil was higher than that of plants in water only.
This is in accordance with the statement of Xia et al. (2004) that
crude oil presence in the water could be utilized by plants as nutri-
ents for their growth.
Based on the growth analysis, the 1% of crude oil in the water
could increase the growth rate of V. zizanioides. This is consistent
5.5
RGR ( g/week)
5
4.5
4
M3 M6 TM3 TM6
Treatment
Fig. 5. Relative growth rate of V. zizanioides. (M3 = 3 pot plants and 1% crude oil,
M6 = 6 pot plants and 1% crude oil, TM3 = 3 pot plants only, TM6 = 6 pot plants
only).
with research of Nisa and Rashid (2015) which showed that the
biomass V. zizanioides growing in water contaminated with crude
oil differed significantly from biomass V. zizanioides in the water
without contamination of crude oil.
Added biomass at the end of the observation was shown by the
length of roots. In 3 pot plants in water contaminated with crude
oil, root length at the end of observation was 8.13 cm, and
6.21 cm (6 pot plants). Meanwhile in water without contamination
of crude oil, root length at the end of observation on 3 pot plants
was 6.57 cm and 4.63 cm (6 pot plants). Treatment of 3 pot plants,
either in crude oil contaminated water or without contamination
of crude oil, generated roots that are longer than that of 6 pot
plants. This might due to at 6 pot plants, nutrient competition
was more intense.
Morphological structure of root and leaf
V. zizanioides root histology (Fig. 6) showed no histological dif-
ferences between plant roots in crude oil contaminated water and
plant roots in crude oil uncontaminated water. The presence of 1%
of crude oil did not cause roots cell damage, as did on the xylem
and phloem (See Fig. 7).
Viewed microscopically (Fig. 6), there were differences in leaf
structure. M3 and M6 image shows that the upper epidermis in
plants contaminated with oil was thicker when compared with
before treatment and treatment without crude oil (Figure TM3
and TM6). Bright green leaves occurred on the treatment of water
contaminated with crude oil. In the treatment without crude oil,
leaves colored dark green. Crude oil affected their leaf structure.
Although it had no significant damage and cell death.
Leaf morphology in crude oil contaminated water during the
study did not show any symptoms of significant damage. This is in
accordance with the research of Brandt et al. (2006) and Xia et al.
(2004) plants grew freshly and optimally until the end of the study.
The existence of V. zizanioides affected the decline of oil concentra-
tion in water. Plants were able to remove harmful chemicals from
the place where the roots take up the nutrients in the water column.
Chemicals were stored in the roots, stems and leaves of plants, then
changed into less harmful chemicals in gaseous form, and released
into the air in the process of transpiration (EPA, 2001).
A research by Alves et al. (2014) compiled that root growth
could extend into deeper soil, allowing access to air and water,
and therefore altering carbon dioxide concentration, pH, redox
potential, osmotic potential, moisture content, and oxygen concen-
tration of the soil. This might lead to a better environment to sup-
port high microbial biomass. Consequently, microbes might
function for phytotoxicity reduction of the soil contaminants, or
augment the capacity of the plant to detoxify and degrade contam-
inants. In our floating wetland, roots of V. zizanioides grew in water
taking up more nutrients constituting decomposition products of
crude oil. V. zizanioides growth performance was better in crude
oil spiked water than that in crude oil uncontaminated water.
 H. Effendi et al. / Egyptian Journal of Aquatic Research 43 (2017) 185–193 191

TM3 TM6

M3 M6
Fig. 6. Root structure of Vetiveria zizanioides. (Initial = before treatment, M3 = 3 pot plants and 1% crude oil, M6 = 6 pot and 1% crude oil, TM3 = 3 pot plants in water only,
TM6 = 6 pot plants in water only).

TM3 TM6

M3 M6
Fig. 7. Leaf structure of Vetiveria zizanioides. (Initial = before treatment, M3 = 3 pot plants and 1% crude oil, M6 = 6 pot plants and 1% crude oil, TM3 = 3 pot plants only,
TM6 = 6 pot plants only).
192 H. Effendi et al. / Egyptian Journal of Aquatic Research 43 (2017) 185–193
Conclusion
On the floating wetland system, 6 pots V. zizanioides during
4 weeks of study could reduce oil and grease content of 91.39%,
84.60% COD, and 84.25% BOD. Meanwhile, three pots of V. zizan-
ioides could reduce oil and grease content of 90.29%, 81.69% COD,
and 81.10% BOD. DO at week 4 decreased, while the BOD and
COD did not show any significant reduction. Therefore, the dura-
tion of V. zizanioides application as an optimal phytoremediator
of oil spill in water was recommended for 3 weeks.
As much as 47.22% parent and 100% new shoots were able to
live until the end of study in the 1% crude oil spiked water. Mean-
while, in the water without oil contamination, as much as 35.56%
parent and 40% new shoots were able to live until the study end.
Average RGR of V. zizanioides in water contaminated with crude
oil was 5.10 g/week and 4.83 g/week in the water without contam-
ination of crude oil.
It can be concluded that crude oil spiked water could support
growth of V. zizanioides better than that in the water without crude
oil contamination. Vetiver could flourish in crude oil contaminated
water by utilizing crude oil undergoing decomposition, as nutri-
ents for growth.
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