Environmental Forensics

ISSN: 1527-5922 (Print) 1527-5930 (Online) Journal homepage: http://www.tandfonline.com/loi/uenf20

Health risk assessment of polycyclic aromatic

hydrocarbons through aquaculture fish
consumption, Malaysia

Essam Nasher, Lee Yook Heng, Zuriati Zakaria & Salmijah Surif

To cite this article: Essam Nasher, Lee Yook Heng, Zuriati Zakaria & Salmijah Surif (2016) Health
risk assessment of polycyclic aromatic hydrocarbons through aquaculture fish consumption,
Malaysia, Environmental Forensics, 17:1, 97-106, DOI: 10.1080/15275922.2015.1133733

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Published online: 22 Mar 2016.

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ENVIRONMENTAL FORENSICS
2016, VOL. 17, NO. 1, 97–106
http://dx.doi.org/10.1080/15275922.2015.1133733

Health risk assessment of polycyclic aromatic hydrocarbons through aquaculture

fish consumption, Malaysia
Essam Nashera,b, Lee Yook Henga,c, Zuriati Zakariad, and Salmijah Surifa
a
Faculty of Science and Technology, Universiti Kebangsaan Malaysia, Bangi, Selangor, Malaysia; bDepartment of Marine Chemistry and Pollution,
Faculty of Marine Science and Environment, Hodeidah University, Hodeidah, Yemen; cSouth-East Asia Disaster Prevention Research Institute
(SEADPRI), Universiti Kebangsaan Malaysia, Selangor, Malaysia; dMalaysia Japan International Institute of Technology, Universiti Teknologi
Malaysia, Kuala Lumpur, Malaysia

ABSTRACT KEYWORDS
Fish are an important source of proteins and healthy lipids. However, they can accumulate nonpolar PAH; giant sea perch;
pollutants such as polycyclic aromatic hydrocarbons (PAHs) from their aquatic environment Langkawi; Malaysia; health
through absorption and/or adsorption. Human exposure to parent PAHs by fish consumption has risk assessment; fish farm
not yet been determined. This study aims to assess the exposure of Malaysians to PAHs through fish
ingestion and to estimate the lifetime cancer risk using the cancer risk assessment guidelines set by
the U.S. Environmental Protection Agency (US EPA) and previous studies. Giant sea perch
(barramundi) were collected from a fish farm on Langkawi Island. The mean concentration of 18
PAHs in the giant sea perch was 573.66 § 47.56 ng g¡1 dry weight. The abundance of low-
molecular-weight PAHs (63%) was higher than that of high-molecular-weight PAHs (37%),
indicating that petrogenic inputs were higher than pyrogenic inputs. The average daily intake of the
18 PAHs through giant sea perch consumption was 294.47 [ng/kg bwt/day]¡1. The lifetime cancer
risks for the 18 PAHs were 1.06 £ 10¡4, 4.55 £ 10¡5, and 3.69 £ 10¡6 when the frequencies of
exposure were assumed as 365 days year¡1 for people who eat fish seven times a week, 156 days
year¡1 for people who eat fish three times a week, and 52 days year¡1 for people who eat fish once
a week, respectively. These results are within the acceptable criterion of the US EPA (10¡6 to 10¡4).
The present study suggests that the consumption of giant sea perch does not pose a significant
source of PAH exposure to Malaysians.

Introduction
Fish are an important source of proteins (e.g., amino
acids), healthy lipids (e.g., long-chain omega-3 fatty
acids), and vitamins and minerals (e.g., Ca, Fe, Cu, and
Zn) in the diet of Malaysians (He et al., 2004; Taweel
et al., 2013). Malaysia recorded a higher annual con-
sumption rate of fish and seafood with 53.2 kg year¡1
(New Zealand Trade and Enterprise, 2012) compared to
several other countries in the region, such as Indonesia
(23.4 kg year¡1), Thailand (32.9 kg year¡1), United States
(24.2 kg year¡1), and United Kingdom (21.3 kg year¡1)
(Food and Agriculture Organization [FAO], 2009). The
total fish production in Malaysia for 2011 was approxi-
mately 1.7 million tons, 82.43% of which was from sea
fishing and 17.23% from fish farms. The Strait of
Malacca contributes 53% of the total seafood in Malaysia
(Department of Fisheries, 2012).
The rapid growth of marine ecotourism has resulted
in the increasing usage of recreational boats and
passenger ferries, consequently causing hydrocarbon
pollution. The Strait of Malacca on the west coast of pen-
insular Malaysia receives a variety of hydrocarbon pollu-
tants, such as crude oil and polycyclic aromatic
hydrocarbons (PAHs), because of its strategic location.
Furthermore, the strait embraces a major international
shipping lane in addition to agricultural, industrial, and
urbanization activities (Abdullah et al., 1999). Langkawi
Island, which is located in the Strait of Malacca, is
increasingly burdened by these activities as well as by the
high intensity of the tourism industry. PAHs are signifi-
cant pollutants in fish.
PAHs are a group of compounds that contain two or
more fused aromatic rings. The U.S. Environmental Pro-
tection Agency (US EPA) has identified 16 PAH com-
pounds as priority pollutants because of their toxic,
mutagenic, and carcinogenic characteristics. Specifically,
the US EPA classified PAHs into carcinogenic and non-
carcinogenic groups. Some probable human carcinogen

CONTACT Essam Nasher essam_abduh@yahoo.com Department of Marine Chemistry and Pollution, Faculty of Marine Science and Environment,
Hodeidah University, Hodeidah, Yemen.
© 2016 Taylor & Francis
98 E. NASHER ET AL.
compounds include benzo(a)anthracene (BaA), chrysene
(Chr), benzo(b and k)fluoranthene, benzo(a)pyrene
(BaP), dibenz(a,h)anthracene (DBA), and indeno(1,2,3-c,
d)pyrene (InP). Acenaphthylene (Acy), anthracene (Ant),
benzo(g,h,i)perylene (BgP), fluoranthene (Flu), fluorene
(Fl), phenanthrene (Phe), and pyrene (Pyr) are known
noncarcinogenics, but they are toxic (Agency for Toxic
Substances and Disease Registry [ATSDR], 1995). PAHs
are easily and rapidly absorbed by organisms and passed
into the marine food chain (Martinez et al., 2004) because
of their low water solubility and high lipophilicity.
Sources of PAHs can be either petrogenic or pyro-
genic (pyrolitic). The former is from petroleum-related
activities, whereas the latter is from the incomplete com-
bustion of wood, coal, or biomass, forest and grass fires,
waste incinerators, fossil fuels that are used in industrial
operations and power plants, and exhaust from vehicles
(Zeng and Vista, 1997; Boonyatumanond et al., 2007).
PAHs are also widely used in commercial products, such
as intermediaries in pharmaceuticals, agricultural prod-
ucts, photographic products, thermosetting plastics, and
lubricating materials.
Exposures of humans to environmental pollutants,
such as PAHs, can occur by different exposure routes,
such as inhalation of ambient air; ingestion of drinking
water, food (e.g., seafood), and pharmaceutical products;
or dermal contact with an agent (Paustenbach, 2002).
Reports on PAH concentrations in fish from fish farms
(Manan et al., 2011) and mussels from the west coast of
Malaysia (Shahbazi et al., 2011) suggest significant PAH
pollution in the marine environment. However, the
human health risk caused by the consumption of this
seafood has yet to be evaluated.
The present study determined the concentrations and
compositions of PAHs in fish from a fish farm in Lang-
kawi Island, estimated the daily intake (DI) by the Lang-
kawi population, and evaluated the lifetime excess cancer
risk (ECR) of PAHs through fish consumption.
Methodology
Chemicals and reagent
A standard mixture of PAHs was purchased from Restek
Corporation (Bellefonte, PA, USA). The mixture consisted
of naphthalene (Nap), 1-methylnaphthalene (1MNap), 2-
methylnaphthalene (2MNap), Acy, acenaphthene (Ace), Fl,
Phe, Ant, Flu, Pyr, BaA, Chr, benzo[b]fluoranthene (BbF),
benzo[k]fluoranthene (BkF), BaP, InP, DBA, and BgP. p-
Terphenyl-d14 (p-Ter) (Supelco, Bellefonte, PA, USA) was
used as the surrogate internal standard. The standards were
further diluted with hexane to prepare five calibration stan-
dard mixtures. Chromatographic-grade dichloromethane
(DCM), n-hexane, acetone, and cyclohexane and HPLC-
grade distilled pentane were obtained from Merck (Darm-
stadt, Germany).
Study area
The Langkawi Archipelago in the Strait of Malacca,
northwest of peninsular Malaysia, consists of 104 islands,
the largest and most exploited of which is Langkawi
Island, with an area of 478.5 km2. Targeted for ecotour-
ism, Langkawi became a duty-free port in 1987 and was
protected from industrial activities. In 2006, it was
declared a National Geopark; a year later, it became a
UNESCO Global Geopark (Langkawi Development
Authority, 2011). In 2010 Langkawi hosted 2.4 million
tourists (Langkawi Development Authority, 2011). To
cater increased tourism-related activities, the local popu-
lation significantly swelled from approximately 40,000 in
1991 to almost 100,000 in 2010 (Samat, 2010). This boost
in the tourism industry, which involves the use of petro-
leum and diesel for boating activities, significantly
increased the pollution in Langkawi.
Sample collection
Five giant sea perch (Lates calcarifer), with average
length and weight of 34 cm and 601 g, respectively, were
collected in December 2010. Giant sea perch was selected
as the study subject because it is preferred in Malaysia
and is readily available. Giant sea perch is also known as
barramundi or Asian seabass. The study fish farm is
located in the south part of the island near Dayang Bun-
ting Island, approximately 3 km from Porto Malai Jetty.
It is situated at 06
130 northern latitude and at 099
460
eastern longitude with a depth of 11 m. The farm was
selected to represent the fish farms and finfish aquacul-
tures in the coastal waters that surround Langkawi
Island. This fish farm is frequently visited by tourists in
addition to providing fresh seafood to the local restau-
rants. The samples were kept cold at 4
C on ice during
transportation and stored at –20
C until further analysis.
Chemical analysis
Sample preparation and extraction
The frozen fish tissue was thawed at room temperature.
Whole soft tissue from the five fish samples was pooled,
weighed, and macerated using a glass blender (Pensonic)
to obtain a uniform consistency. The macerated tissue
was dried in a freeze-drier (Labconco Lyph Lock 6,
Model: 7530–00, Watertown, PA, USA) and then mixed
in a dry blender until it was turned into powder form.
The extraction process described by Martinez et al.

(2004) was followed. A 1 mL aliquot of the surrogate
standard p-Ter (2 mg mL¡1) was added to dried tissue
samples (10 g) and refluxed with potassium hydroxide in
methanol (1 M; 50 mL) at 80
C for 4 hr. The solution
was allowed to cool and then filtered through a 0.45 mm-
pore size Whatman filter paper. The extract was trans-
ferred into a separation funnel. n-Hexane (30 mL) and
deionized water (10 mL) were added to the extract with
shaking and then allowed to settle for 10 min. The top n-
hexane layer containing the hydrocarbons was decanted
into a 250 mL round-bottomed flask, and the procedure
was repeated three times. The combined extract was
treated with 5 g of anhydrous Na2SO4, activated at
400
C for 4 hr before use to remove residual water, and
concentrated to approximately 3 mL by using a rotary
evaporator. Cyclohexane (10 mL) as an exchange solvent
(US EPA, 1996a) was added to the extract and then con-
centrated to 2 mL by using a rotary evaporator. The
extract was passed through a glass column containing
5 g of activated silica gel (previously activated by heating
at 200
C for 16 hr before use) and 1 g of anhydrous
Na2SO4 (US EPA, 1996a). The PAH fraction was eluted
by using 30 mL of a mixture of DCM/pentane (2:3, v/v)
and concentrated to 2 mL by using a rotary evaporator.
Hexane (10 mL), which was added as an exchange sol-
vent (US EPA, 1996a), was evaporated to 2 mL. The
extract was finally reduced to 1 mL under a gentle stream
of nitrogen gas. All sample extracts were kept in amber
glass vials at –4
C until analyzed within a week.
To determine lipid content in the fish, the freeze-dried
samples (approximately 2 g) were extracted with 50 mL
of hexane in an automatic extraction unit (Soxtherm,
Gerhardt, K€ onigswinter, Germany). Lipid content was
gravimetrically measured. Percentage of lipid content
was calculated according to Shahidi (2001) using the
equation:


w3 ¡ w2
Lipid content ð %Þ D £ 100
w1
where W1 D tissue weight (g) of fish, W2 D weight (g) of
empty cup, and W3 D weight (g) of cup with the lipid
extract
Gas chromatography analysis of PAH
PAH was analyzed on a gas chromatograph (GC) with a
flame ionization detector (FID) in accordance with the
US EPA method 8100 (USEPA, 1986). The final extract
(1 mL) of fish was injected by the injector of an Agilent
7683 auto-sampler into an Agilent 6890N plus gas chro-
matograph with an FID (Agilent Technologies, Santa
Clara, CA, USA) and a fused silica TR-5MS capillary col-
umn (30 m £ 0.25 mm i.d.) with a film thickness of
ENVIRONMENTAL FORENSICS 99
0.25 mm (Thermo Fisher Scientific, Waltham, MA,
USA). Helium (purity 99.9%) was used as a carrier gas,
makeup gas, and purge gas at flow rates of 1.0, 45, and
30.0 mL min¡1, respectively. The flow rates for the FID
were 450 and 45 mL min¡1 for air and hydrogen, respec-
tively. The gas chromatograph was operated in splitless
mode, and separation was conducted with the oven tem-
perature programmed as follows: initial setting at 80
C
(1 min hold), ramped to 180
C at 10
C min¡1 (for
2 min), and finally to 320
C at 5
C min¡1 (10 min hold).
The injector was held at 250
C, and the FID was main-
tained at 350
C. Agilent Chemstation software was used
to obtain the chromatogram and to calculate data.
Quality assurance/quality control
The accuracy of the analytical procedure was examined
through the recovery of spiked dry fish tissue after sub-
traction from the unspiked sample. A known amount of
PAH mixture was spiked into a dry fish tissue sample and
left for a few hours before being similarly extracted and
analyzed. The surrogate internal standard p-Ter was used
throughout the analytical procedure to monitor the losses
and contamination during sample extraction and instru-
mental analysis. Samples were analyzed in triplicate to cal-
culate the precision of the measurements. The solvent and
surrogate internal standard, which was contained in the
method blank, were analyzed to evaluate the contamina-
tion of the used solvents, reagents, and glassware.
An external standard calibration composed of 18
PAH standards was used to identify and quantify each
component peak in the sample chromatogram. Five
PAH mixture standards were run in the GC-FID on the
same day of sample analysis to estimate the regression
equations for the calculation of individual PAH concen-
trations in the samples. The PAH peaks in samples were
matched with PAH standards, and only the peaks that
had the same retention times as the standards were con-
sidered (US EPA, 1986) (Table 1).
Statistical analysis
Pearson correlation analysis was carried out to test the
relationship between the PAHs and lipid content in the
fish using SPSS version 15.0 for Windows. Statistical sig-
nificance was defined as p  0.05 (p value at the 95%
confidence level).
Data analysis
Calculation of DI of PAHs
The dietary intake of PAHs through fish consumption
was calculated by multiplying the average daily
100 E. NASHER ET AL.

Table 1. Average recovery of 18 PAHs spiked dry muscle of fish (mean §RSD %), n D 3, molecular weight, and their retention times.
PAH Abbreviation Retention times Molecular formula Molecular weight Recovery (% RSD)

Naphthalene Nap 8.04 C10H8 128.17 88 § 1

1-methylnaphthalene 1MNap 9.50 C11H10 142.20 87 § 2
2-methylnaphthalene 2MNap 9.80 C11H10 142.21 87 § 2
Acenaphthylene Acy 11.75 C12H8 152.19 86 § 3
Acenaphthene Ace 12.17 C12 H10 154.2 96 § 3
Fluorene Fl 13.90 C13H10 166.23 86 § 0
Phenanthrene Phe 17.71 C14H10 178.23 99 § 0
Anthracene Ant 17.90 C14H10 178.23 102 § 6
Fluoranthene FIu 23.02 C16H12 202.26 110 § 6
Pyrene Pyr 24.06 C16H10 202.25 111 § 6
Benzo[a]anthracene BaA 29.69 C18H12 228.29 113 § 9
Chrysene Chr 29.87 C18H12 228.28 92 § 0
Benzo[b]fluoranthene BbF 34.44 C20H12 252.31 115 § 11
Benzo[k]fluoranthene BkF 34.56 C20H12 252.31 114 § 11
Benzo[a]pyrene BaP 35.77 C20H12 252.31 105 § 11
Indeno[123-cd]pyrene InP 39.90 C22H14 278.35 108 § 12
Dibenzo[ah]anthracene DBA 40.05 C22H14 278.35 108 § 5
Benzo[ghi]perylene BgP 41.00 C22H12 276.33 113 § 13

consumption rate of fish by the corresponding PAH con-
centration. The DI of PAHs through fish was calculated
as
DI D Ci £ IR
DI D ðCi £ IRÞ=BW
where Ci is the PAH concentration in the edible portion
of the fish (ng g¡1 wet weight [wwt] basis). One half of
the reported detection limit was used for results that
were reported as non-detects (i.e., less than the detection
limit). IR is the fish consumption rate (g day¡1) for the
general population; it was set to 147 g day¡1 person¡1
from the annual per capita fish intake of 53.6 kg year¡1
for Malaysia (FAO, 2009). BW is the average body
weight (bwt) for the general population; it was set to
64 kg in accordance with previous studies (Lim et al.,
2000; Taweel et al., 2012).
Calculation of toxic equivalency quotient (TEQ)
TEQ is regarded as a better index for potent toxicity than
concentration (Ni and Guo, 2013). Therefore, the TEQ
of PAHs in the fish was calculated using Equation (3),
which was cited from the reports by Nisbet and Lagoy
(1992) and Ding et al. (2012).
Xn
TEQBaP D .i D 1/
Ci £TEFi
where TEQBaP is the total TEQ level converted as benzo
(a)pyrene by using the toxic equivalency factor (TEF) of
PAHs in fish and Ci is the concentration of each individ-
ual PAH in the edible portion of the fish. TEFBaP is TEF
of the individual PAHs relative to BaP and is used to cal-
culate the toxic potencies of individual PAHs relative to
BaP. The values of TEFBaP were suggested by Nisbet and
Lagoy (1992). The US EPA has established TEFs for only
certain carcinogenic PAHs, whereas Nisbet and LaGoy
established TEFs for both carcinogenic and noncarcino-
genic PAHs because of the effect of noncarcinogenic
(1) PAHs in increasing the tumor incidence of carcinogenic
PAHs (Pfeiffer, 1977; Yoon et al., 2007).
(2)
Calculation of ECR
ECR, also referred to as the incremental lifetime cancer
risk, is induced by dietary exposure to PAHs through
fish consumption. ECR was estimated as follows (Yoon
et al., 2007; Xia et al., 2010; Ding et al., 2012):
ECRi D TEQi £ SF £IR £FEj £ED
£CF=ðBW £ ATÞ (4)
where ECR is the lifetime ECR of the dietary exposure
(dimensionless); TEQi is the TEQ (ng g¡1) of the total
PAH for giant sea perch; SF is the oral cancer slope factor
of BaP, estimated as 7.3 [mg/kg/day]¡1 by the integrated
risk information system of the US EPA (2000); IR is the
average fish consumption rate of 147 [g/day/person]¡1
for the general population; BW is the average bwt
(64 kg) for the general population; CF is a conversion
factor, 10¡6 kg mg¡1; AT is the average lifetime for carci-
nogens (70 years £ 365 days D 25,550 days (US EPA,
(3) 2000; the AT for noncarcinogens is equal to the exposure
duration); ED is exposure duration (30 years; this value
is presented as the 90th percentile for time spent at one
residence; US EPA, 1991); and FEj is the frequency of
exposure with three assumed scenarios, namely,
365 days year¡1 for people who eat fish seven times a
week, 156 days year¡1 for people who eat fish three times
a week, and 52 days year¡1 for people who eat fish once
a week.
 ENVIRONMENTAL FORENSICS 101

Results and discussion
Quality assurance and quality control
The average recovery of the 18 PAHs and p-Ter using the
adopted method was between 86% and 115%, with a rel-
ative standard deviation (RSD) of less than 13% (Table 1).
These values meet the acceptance criteria of the US EPA
(1996b), indicating that the method adopted in this study
is acceptable for the analysis of PAHs in fish samples.
The detection limit values of the individual PAHs using
the method adopted and GC-FID ranged from 3.97 to
17.92 ng g¡1, with the instrumental limit of detection of
the individual PAHs ranging from 0.054 to 0.252 ng m¡1.
Concentration of PAHs
The mean concentration of the 18 total PAHs in giant
sea perch was 124.9 § 24.16 based on wet weight (wwt)
and 573.66 § 47.56 ng g¡1 based on dry weight (dwt).
The concentrations of the PAHs in giant sea perch were
normalized on a lipid weight (lwt) basis to obtain the
real concentration in the lipid content of the fish muscle.
The PAH concentration was also normalized on a wwt
basis for comparison purposes and health risk calcula-
tion. The mean concentrations of the individual PAHs
ranged from 6.85 § 1.20 ng g¡1 dwt for Phe to 165.12 §
19.40 ng g¡1 dwt for Nap (Table 2).
All 18 PAHs, except for 1MNap, Pyr, and Fl, were
detected in the samples of giant sea perch. The possible
accumulation and concentration routes of the PAHs in
fish tissues are through the ingestion of contaminated
food, suspended matter, or sediment, and through the
absorption of these materials through gills and skin
(Saeed et al., 1995). The PAH concentrations in the fish
from this farm were higher than those in the water
(17.86 § 0.43 mg L¡1) (Nasher et al., 2013a) but lower
than those in the sediment (974 § 96 ng g¡1) from the
same fish farm (Nasher et al., 2013b).
European Union regulation (EC) no. 1881/2006 set
allowable maximum levels for PAHs in foodstuffs for
four PAH compounds (chrysene, benzo[a]anthracene,
benzo[b]fluoranthene, and benzo[k]fluoranthene;
European Union, 2011). The total concentration of
these PAHs in the samples of giant sea perch was
14.38 § 4.81 ng g¡1 wwt, which is lower than the
European Union- allowable maximum levels (30.00 mg
kg¡1 wwt) for these four PAHs. This result indicates
that no risk of carcinogenicity can be expected from
consumption of this fish. According to the guidelines
of the British Columbia Ministry of Environment,
Lands and Parks (1993), the maximum allowable con-
centrations of benzo[a]pyrene in fish for people who
consume 100 g of fish per week is 2 mg kg¡1 wwt. The
mean concentration of benzo[a]pyrene in giant sea

Table 2. Concentrations (ng g¡1) of individual PAHs, seven carcinogenic PAHs, and total PAHs based on dry weight (dwt), wet weight
(wwt), and lipid weight (lwt) in the muscle tissues of giant sea perch.
PAH concentration

Dry weight Wet weight Lipid weight

PAH mean § SD mean § SD mean § SD

1 Nap 165.12 § 19.40 35.96 § 4.23 2822.50 § 331.71

2 1MNap nd nd nd
3 2MNap 22.96 § 2.42 5.00 § 0.53 392.55 § 41.37
4 Acy 135.20 § 15.60 29.45 § 3.41 2311.14 § 267.48
5 Ace 16.81 § 2.80 3.66 § 0.61 287.31 § 47.80
6 Fl nd nd nd
7 Phe 6.85 § 1.20 1.49 § 0.27 117.13 § 21.36
8 Ant 14.08 § 4.40 3.06 § 0.95 239.87 § 74.62
9 FIu 45.02 § 12.30 9.80 § 2.68 769.49 § 210.33
10 Pyr nd nd nd
11 BaA 15.51 § 3.94 3.38 § 0.86 265.11 § 67.28
12 Chr 20.89 § 3.76 4.55 § 0.82 357.23 § 64.28
13 BbF 13.10 § 4.95 2.85 § 1.08 223.95 § 84.65
14 BkF 16.54 § 9.44 3.60 § 2.06 282.65 § 161.43
15 BaP 6.86 § 2.10 1.50 § 0.45 117.48 § 35.47
16 InP 15.01 § 8.06 3.27 § 1.75 256.60 § 137.72
17 DBA 54.14 § 12.26 11.79 § 2.67 925.45 § 209.50
18
P BgP 25.61 § 8.26 5.58 § 1.80 437.82 § 141.13
P18 PAHs a 573.66 § 47.56 124.94 § 24.16 9806.30 § 1896.14
7C PAHs 142.07 § 44.48 30.94 § 9.69 2428.48 § 760.33
Lipidb (%) 5.85 § 1.31
Water content (%) 78.22

SD: standard deviation; nd: below the detection limit.

a
BaA, Chr, BbF, BkF, BaP, InP, DBA.
b
In dry weight.
102 E. NASHER ET AL.
perch (1.50 § 0.45 ng g¡1 wwt) was lower than this
guideline, indicating that consuming this fish poses no
health risk.
The lipid content of giant sea perch was 5.85 § 1.31%
dwt. The individual and total concentrations of PAHs
after normalization by lipid content are shown in Table 2.
The total residual level of PAHs on a lipid-normalized
weight basis was 9806.30 § 1896.14 ng g¡1 lwt. As lipo-
philic compounds, PAHs usually prefer to accumulate in
lipids. Hence, a significant positive relationship was
observed (r D 0.99, p < 0.05) between the lipid contents
and total residual level of PAHs. This result indicates
that lipid is an important factor in the accumulation of
lipophilic PAHs by fish. This finding is consistent with
the results of Xu et al. (2011) and Ramalhosa et al.
(2012), who stated that a significant positive relationship
exists (p < 0.001) between lipid content and PAH con-
centration in fish muscle tissues.
Of the 18 PAH compounds studied in giant sea perch,
7 are carcinogenic and 11 are noncarcinogenic, although
they are toxic, according to ATSDR (1995). The mean
concentration of the total residual level of the 7 PAHs,
which are known as high-molecular-weight PAHs
(HMW-PAHs), was 142.07 § 15.51 ng g¡1 dwt (30.94 §
9.69 ng g¡1 wwt). These seven HMW-PAHs are BaA,
Chr, BbF, BkF, BaP, DBA, and InP.
The total concentration of the 18 PAHs in giant sea
perch (124.87 § 24.16 ng g¡1 wwt) is lower than those
obtained in Taiyuan, China (160.30 ng g¡1 wwt; Xia
et al., 2010), Egypt (0.78 to 154 ng g¡1 wwt; Loutfy et al.,
2007), and Ghana (48.75 to 166.8 ng g¡1 wwt; Nyarko
and Klubi, 2011). It is similar to the values obtained in
Hong Kong (15.5 to 118 ng g¡1 wwt; Cheung et al.,
2007), but higher than the values obtained from fish and
shellfish in Spain (2.5 to 22.4 ng g¡1 wwt; Llobet et al.,
2006) and Korea (4.73 to 87.8 ng g¡1 wwt or 12.3 to
243 ng g¡1 dwt; Moon et al., 2010). The total concentra-
tion of the 18 PAHs in giant sea perch (573.66 §
47.56 ng g¡1 dwt) is lower than that found in fish from
the Strait of Malacca, Malaysia (130 to 9610 ng g¡1 dwt;
Manan et al., 2011), but higher than that in Prince Wil-
liam Sound, United States (1.3 to 351 ng g¡1 dwt; Neff
et al., 2006), and Kuwait (83.8 to 473 ng g¡1 dwt; Saeed
et al., 1995). In general, the PAH level in the fish of Lang-
kawi Island is low to moderate in comparison with the
findings from previous studies.
PAH composition
PAHs are divided into two groups based on their physical,
chemical, and biological characteristics (Martinez et al.,
2004). Low-molecular-weight (LMW) PAHs, such as Nap
and Fl with 2 to 3 rings of PAHs, exert significant acute
Table 3. Distribution of high-molecular-weight (HMW) PAHs (4–6
rings) and low-molecular-weight (LMW) PAHs (2–3 rings).
PAH group LMW HMW
ng g¡1
(%) ng g¡1 (%)
2 rings 188.08 33 — —
3 rings 172.89 30 — —
4 rings — — 81.42 16
5 rings — — 44.65 14
6 rings — — 61.01 7
Total 360.97 63 187.08 37
toxicity to aquatic organisms. High-molecular-weight
(HMW) PAHs, such as Chr and coronene with four to
seven rings, do not cause acute toxicity but are carcino-
genic (Neff, 1979). Compared with HMW-PAHs, LMW-
PAHs are the more abundant group in crude oil and oil
products and are more easily degraded in the environment.
The concentration and percentage of PAHs were
grouped according to their ring size (Zhang et al., 2004).
The number of benzene rings in each compound in giant
sea perch is listed in Table 3. The 2-ring PAHs, Nap,
1MNap, and 2MNap, accounted for 33% of the total pro-
files and concentration of 188.08 ng g¡1 dwt. The 3-ring
PAHs, Acy, Ace, Fl, Phe, and Ant, accounted for 30% of
the total profiles and concentration of 172.89 ng g¡1 dwt.
The profiles of 4-ring PAHs (Flu, Pyr, BaA, and Chr),
5-ring PAHs (BbF, BkF, BaP, DBA), and 6-ring PAHs
(InP and BgP) were low, accounting for 16% (81.42 ng g¡1
dwt), 14% (44.65 ng g¡1 dwt), and 7% (61.01 ng g¡1 dwt),
respectively, of the total PAHs in the fish (Table 3).
The concentration of LMW-PAHs (2 and 3 rings) in
giant sea perch was higher (360.97 ng g¡1 dwt) than that
of HMW-PAHs (4 to 6 rings; 187.08 ng g¡1 dwt). The
LMW-PAHs and HMW-PAHs accounted for 63% and
37% of the total PAHs, respectively. The differences in
distribution of HMW-PAH and LMW-PAH in giant sea
perch may be related to the differences in the metabolic
capacities of the fish toward the two types of PAHs (Bau-
mard et al., 1998). The high LMW-PAH content suggests
that the giant sea perch studied can accumulate and
magnify LMW-PAHs; alternatively, the studied fish are
probably more exposed to the water-soluble PAHs and
concentrate the PAHs adsorbed on floating materials,
such as fine suspended particulate matter and phyto-
plankton (Soclo et al., 2008).
The LMW-PAHs may be generated from petrogenic
sources. The most likely inputs of petrogenic PAHs in
this area are the inefficient two-stroke outboard engines
of most boats in Langkawi, which usually involve 20% of
unburned fuel being discharged into the water (Mosisch
and Arthington, 2001). Moreover, oil spill and tanker
accidents in the Strait of Malacca contributed to the dis-
charge of crude oil and oil products directly into the sea
(Zakaria et al., 2000; Sakari et al., 2008). The An Tai
 ENVIRONMENTAL FORENSICS 103

(1997) and Sun Vista (1999) incidents are examples
of such spills. The HMW-PAHs content may suggest
the importance of anthropogenic activities, such as the
incomplete fuel combustion of boats, ships, and vehicle
engines (Nasher et al., 2013b).
Risk assessment
Dietary intake of PAHs through fish consumption
Table 4 shows the calculated PAH intake through giant
sea perch consumption for the general population of
Langkawi, Malaysia. The estimated intake of individual
PAH through fish consumption ranged from 1.34 to
82.60 [ng/kg bwt/day]¡1, and the total intake of 18
PAHs was at 294.47 [ng/kg bwt/day]¡1. The estimated
intake of total carcinogenic PAHs, 71.07 [ng/kg bwt/
day]¡1, was lower than the total noncarcinogenic PAHs
of 223.40 [ng/kg bwt/day]¡1. However, the effect of non-
carcinogenic PAHs in increasing the tumor incidence of
carcinogenic PAHs has been demonstrated (Pfeiffer,
1977). The highest individual estimated intake was
recorded for Nap, 82.60 [ng/kg bwt/day]¡1. The high
intake of PAHs through farmed fish (giant sea perch)
was caused by the high consumption rate of Malaysians
(147 g day¡1 person¡1) and/or the high concentration of
PAHs in the fish. Furthermore, these values are higher
than the median dietary exposure of European countries
for PAHs [51.3 ng/kg bwt/day]¡1, for which seafood and
cereal were the highest contributors.
The average DI of 18 PAHs for the general population
in the present study was 294.47 [ng/kg bwt/day]¡1. This
result is higher than the DI of the Korean population
(men D 16.7 [ng/kg bwt/day]¡1; women D 13.8 [ng/kg
bwt/day]¡1; Moon et al., 2010) and the Atlantic Ocean’s
general population (four PAHs; 9.43 § 3.15 [ng/kg bwt/
day]¡1; Ramalhosa et al., 2012). The higher dietary
intake in the present study than those in other reported
results can be associated with the high consumption rate
of fish in Langkawi, Malaysia.
ECR of PAHs exposed by fish ingestion
The lifetime ECR of PAHs through giant sea perch inges-
tion for the general population, based on three scenarios
of frequency of exposure for the residents of Langkawi,
Malaysia, was calculated (Table 4). After converting
PAH concentrations to their BaP equivalents, risks
were estimated using the BaP cancer slope factor
(Equation (4)).
The first scenario considers the frequency of exposure
as 365 days year¡1 for people who eat fish seven times a
week. On the basis of this assumption, the ECR value
ranged from 9.75 £ 10¡9 to 8.47 £ P 10¡5 for individual
¡4
PAHs and was 1.06 £ 10 for the 18 PAHs. More-
over, the ECR values for the total carcinogenic and non-
carcinogenic PAHs were 1.05 £ 10¡4 and 1.26 £ 10¡6,
respectively. The second scenario assumes that the fre-
quency of exposure is 156 days year¡1 for people who
eat fish three times a week. The ECR values ranged from
4.17 £ 10¡9 to 3.62 £P 10¡5 for individual PAHs and was
¡5
4.55 £ 10 for the 18 PAHs. For the total carcino-
genic and noncarcinogenic PAHs, the ECR values were
4.50 £ 10¡5 and 5.37 £ 10¡7, respectively. The third

Table 4. Concentration of PAHs, estimated total PAH DI, and excess cancer risk (ECR) of PAHs for three fish consumption scenarios for
the residents of Langkawi, Malaysia.
PAH TEFa Conc. TEQb DI ECR ECR ECR
ng g¡1 [ng/kg bwt/day]¡1 (Scen. 1) (Scen. 2) (Scen. 3)

Nap 0.001 35.96 0.04 82.60 6.03 £ 10¡7 2.58 £ 10¡7 2.09 £ 10¡8
1MNap 0.001 2.09 0.001 4.80 3.50 £ 10¡8 1.50 £ 10¡8 1.21 £ 10¡9
2MNap 0.001 5.00 0.01 11.49 8.39 £ 10¡8 3.58 £ 10¡8 2.91 £ 10¡9
Acy 0.001 29.45 0.03 67.64 4.94 £ 10¡7 2.11 £ 10¡7 1.71 £ 10¡8
Ace 0.001 3.66 0.001 8.41 6.14 £ 10¡8 2.62 £ 10¡8 2.13 £ 10¡9
Fl 0.001 0.58 0.001 1.34 9.75 £ 10¡9 4.17 £ 10¡9 3.38 £ 10¡10
Phe 0.001 1.49 0.001 3.43 2.50 £ 10¡8 1.07 £ 10¡8 8.67 £ 10¡10
Ant 0.01 3.06 0.03 7.02 5.12 £ 10¡7 2.19 £ 10¡7 1.78 £ 10¡8
FIu 0.001 9.80 0.01 22.52 1.64 £ 10¡7 7.03 £ 10¡8 5.70 £ 10¡9
Pyr 0.001 0.59 0.001 1.35 9.86 £ 10¡9 4.21 £ 10¡9 3.42 £ 10¡10
BaA 0.1 3.38 0.34 7.76 2.43 £ 10¡6 1.04 £ 10¡6 8.41 £ 10¡8
Chr 0.01 4.55 0.05 10.45 3.27 £ 10¡6 1.40 £ 10¡7 1.13 £ 10¡8
BbF 0.1 2.85 0.29 6.55 2.05 £ 10¡7 8.76 £ 10¡7 7.11 £ 10¡8
BkF 0.1 3.60 0.36 8.27 2.59 £ 10¡6 1.11 £ 10¡6 8.97 £ 10¡8
BaP 1 1.50 1.50 3.44 1.08 £ 10¡5 4.60 £ 10¡6 3.73 £ 10¡7
InP 0.1 3.27 0.33 7.51 2.35 £ 10¡6 1.00 £ 10¡6 8.14 £ 10¡8
DBA 1 11.79 11.79 27.08 8.47 £ 10¡5 3.62 £ 10¡5 2.94 £ 10¡6
BgP
P 0.01 5.58 0.06 12.81 9.35 £ 10¡7 4.00 £ 10¡7 3.24 £ 10¡8
P18 PAHs c 128.20 14.82 294.47 1.06 £ 10¡4 4.55 £ 10¡5 3.69 £ 10¡6
P non-CPAHs 97.26 0.17 223.40 1.26 £ 10¡6 4.37 £ 10¡7 4.36 £ 10¡8
7C PAHsd 30.94 14.64 71.07 1.05 £ 10¡4 4.50 £ 10¡5 3.65 £ 10¡6
a
Toxic equivalency factor of the individual PAH.
104 E. NASHER ET AL.
scenario assumes that the frequency of exposure is
52 days year¡1 for people who eat fish once a week. The
ECR value ranged from 3.38 £ 10¡10 to 2.94 £ 10¡6 Pfor
individual PAHs and was 3.69 £ 10¡6 for the 18
PAHs. Moreover, the ECR values for total carcinogenic
and noncarcinogenic PAHs were 3.65 £ 10¡6 and 4.36
£ 10¡8, respectively. The ECR values of the total PAHs
(carcinogenic and noncarcinogenic) reveal a good pic-
ture of health risk, and we should not forget the carcino-
genic potency of noncarcinogenic PAHs because the
toxicity of PAHs has structure specificity, and most
PAHs are ingested at once, typically in food (Yoon et al.,
2007). The calculation of health risk caused by the con-
sumption of giant sea perch containing different PAHs
showed that the ECR values for the carcinogenic, non-
carcinogenic, and total PAHs ranged from 10¡8 to 10¡5.
This range is within the acceptable criteria of the US
EPA (10¡6 to 10¡4; Sidhu, 2003; Xia et al., 2010). The
priority risk level (10¡4) is considered a serious risk to
health that requires attention from concerned people
(Asante-Duah, 2002; Xia et al., 2010). The values found
in sea perch do not exceed the priority risk level (10¡4),
which means that consuming fish from the study Lang-
kawi fish farm does not pose a health risk to the inhabi-
tants. If the values exceed the priority risk level, the
consumption of this fish may cause adverse health
effects.
In general, the differences in the levels of ECR would
depend not only on the concentration of PAHs but also
on the consumption frequency and intake amount.
These three factors are essential to maintaining the bal-
ance between the health benefits and risks of fish con-
sumption (Domingo et al., 2007).
A few reports have represented the lifetime ECR of
PAHs through seafood ingestion. Yoon et al. (2007) car-
ried out a health risk assessment for the Korean popula-
tion and found that B[a]Peq-based ECRs are 1.89 £
10¡7 for adults and 1.70 £ 10¡7 for the elderly popula-
tion. These values are lower than the results of all three
scenarios in the present study. The results illustrate that
the dietary intake of the Korean population is lower than
that of the Langkawi Island population. Dhananjayan
and Muralidharan (2012) measured the PAH concentra-
tions in fish from Mumbai Harbour, India (from 2006 to
2008). The calculated ECR for the dietary intake of
PAHs through fish ranged from 2.37 £ 10¡7 to 1.43 £
10¡6 for the general population with a bwt of 60 kg. This
result is slightly lower than our results because of the
lower intake rate of the Indian population than the Lang-
kawi population.
The PAH concentration in three pelagic fish samples
collected from Lake Small Bai-Yang-Dian, northern
China, was measured in October 2007. The calculated
ECR for the dietary intake of PAHs in Northern China
was lower than 1 £ 10¡5 for the adult population with a
bwt of 70 kg (Xu et al., 2011). The results of their study
were close to our study results. The results of our study
are within the acceptable excess risk level (1 £ 10¡6 to 1
£ 10¡4) established by the US EPA for carcinogenic
compounds (e.g., PAHs). This level is known as the real-
istic, safe, and protective level for public health (Sidhu,
2003). Thus, the fish from the study Langkawi Island fish
farm are generally safe for consumption.
Conclusion
The concentrations and distributions of carcinogenic
and noncarcinogenic PAHs were measured in giant sea
perch collected from a Langkawi Island fish farm. The
PAH concentrations ranged from 394.6 to 656.8 ng g¡1
dwt. LMW-PAHs were more abundant than HMW-
PAHs in the studied giant sea perch, indicating that pet-
rogenic input was higher than pyrogenic input. The
most possible sources of petrogenic PAHs in this area
are the inefficient two-stroke outboard engines of boats
in Langkawi Island and the oil spill in the Strait of
Malacca. A significant positive relationship exists
between lipid contents and PAHs, suggesting the impor-
tant role of lipids in the accumulation of lipophilic PAHs
in fish. The PAH intake through giant sea perch con-
sumption by the general population was estimated. The
average dietary intake values of PAHs by fish consump-
tion are higher than the median dietary exposure of
European countries to PAHs. However, overall, the ECR
levels from fish consumption for the Langkawi Island
population are generally safe.
Funding
This work was financially supported by grants No. UKM-ST-
06-FRGS0245-2010, UKM-OUP-PLW-11-48/2010, and Grant
UKM-Arus Perdana on Langkawi Geopark.
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