Marine Pollution Bulletin 69 (2013) 228232

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Marine Pollution Bulletin

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Baseline

Concentration proles of PCB congeners in the blubber and liver of Steller sea lions
(Eumetopias jubatus) from the coast of Hokkaido, Japan
Keiko Kubo a,b,1, Katsuyuki Yamaguchi b,, Masaki Mitsuhashi c, Kaoru Hattori d, Shunitz Tanaka a,1
a
Graduate School of Environmental Science, Hokkaido University, N10-W5, Kita-ku, Sapporo, Hokkaido 060-0810, Japan
b
Institute of Environmental Sciences, Environmental and Geological Research Department, Hokkaido Research Organization, N19-W12, Kita-ku, Sapporo, Hokkaido 060-0819, Japan
c
Kushiro Fisheries Research Institute, Fisheries Research Department, Hokkaido Research Organization, Hama-machi 2-6, Kushiro, Hokkaido 085-0024, Japan
d
Ecosystem Studies Fisheries Resources Division, Hokkaido National Fisheries Research Institute, National Fisheries Agency, Katsurakoi 116, Kushiro, Hokkaido 085-0820, Japan

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

Keywords: The concentrations of PCB congeners in the blubber and liver of male and female Steller sea lions (Eumet-
PCBs opias jubatus; SSLs) collected from the Shakotan Peninsula and the Nemuro Strait, Hokkaido, Japan in
Congeners 2008 and 2010 were measured by HRGCHRMS, in order to express the concentration proles in SSLs
Steller sea lion at these regions. #153 predominated in both organs, followed by #138, #99 and #118. In males, #28
Blubber
and #31 were concentrated particularly in the blubber, while #177 and #199 were accumulated specif-
Liver
Hokkaido
ically in the liver. The differences in these concentration proles might be expressed by the differences in
the organs and their functions. The concentrations of #99, #118, #138, #153, and #180 in the blubber
appeared to correspond signicantly with the differences between genders. The details of the surveys
on PCB congeners in SSLs could clarify the differences in the residue levels of individual congeners for
organs and genders.
2012 Elsevier Ltd. All rights reserved.

Polychlorinated biphenyls (PCBs) continue to be the major con-
taminants in marine mammals at the top of the marine food chain
(e.g., Kajiwara et al., 2006; Tanabe et al., 1994a). The blubber and
livers of marine mammals are thought to accumulate the most
PCBs. There is concerned that a high risk of immunity obstruction
and cacochymia in marine mammals could be caused by the accu-
mulation of organochlorines such as PCBs (e.g., Kajiwara et al.,
2002; Tanabe, 1988). To study PCB congener proles in marine
mammals, the area in which a target lives and feeds is an impor-
tant factor because of their individual manner (Boon et al., 1997).
Therefore, more detailed surveys of PCB congeners are necessary
for marine mammals in various areas that have different habitats
and feeds.
The coastal Hokkaido area in Japan is an important foraging
area for Steller sea lions (Eumetopias jubatus; SSLs). SSLs breed on
the Kuril Islands and on certain Islands in the Okhotsk Sea
(Ishinazaka and Endo, 1999), and migrate to the Hokkaido coast
during the non-breeding winter season to forage.
The residue level of total PCBs in various organs and tissues
including the blubber and liver was reported by Kim et al. (1996)
in SSLs from the Hokkaido coast (Kim et al., 1996). However, there
was no mention of the detail proles of PCB congeners in their
report.
Corresponding author. Tel.: +81 11 747 3588; fax: +81 11 747 3254.
E-mail address: k-yamaguchi@hro.or.jp (K. Yamaguchi).
1
Tel.: +81 11 706 2219.
The purpose of this study was to explain the details of the con-
centration proles of PCB congeners in the blubber and liver of SSLs
from the Hokkaido coast of Japan during the winter season of 2008
and 2010. The concentrations of organochlorines such as total PCB
concentrations detected in the liver were clearly correlated with
those in the blubber (Lee et al., 1996). These two organs are suit-
able samples for a comparison study into the organ-wise differ-
ences of PCB congener proles. Therefore, the detailed survey on
PCB concentrations has the potential to obtain new knowledge
about the accumulation and the distribution of individual PCB
congeners in SSLs.
The samples of SSLs were collected by bycatch on set nets or by
gun-hunting for alleviation of shery damages from harmful ani-
mals, at the Shakotan Peninsula and the Nemuro Strait, Hokkaido,
Japan, during the winter seasons of 2008 and 2010. The liver sam-
ples from 8 males and 3 females, and blubber samples from 9
males and 3 females were used for analysis. These samples were
stored at 30 C until analysis. The biometrical data of the ana-
lyzed SSLs are shown in Table 1, and the sampling location is
shown in Fig. 1.
PCBs of the blubber and liver samples were analyzed according
to a manual on the determination of PCBs in organic samples by
Ministry of the Environment in Japan (Environmental Monitoring
Report on Persistent Organic Pollutants (POPs) in Japan, 2002
2004) and to cleanup methods of previous reports (Suzuki et al.,
2006, 2007). Two grams of the blubber samples were ground with
an adequate quantity of anhydrous sodium sulfate to promote

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http://dx.doi.org/10.1016/j.marpolbul.2012.12.012
 K. Kubo et al. / Marine Pollution Bulletin 69 (2013) 228232 229

Table 1
Biometrical data of analyzed SSLs.

Sampling year and area ID number Sex Whole length (cm) Weight (kg) Analyzed tissue Lipid content (%)
Blubber Liver
Shakotan area in 2008 S-1 Male 310 420 LiverBlubber 75.0 3.4
S-2 Male 310 420 LiverBlubber 79.0 3.6
S-3 Male 302 425 LiverBlubber 83.0 3.7
S-4a Female 285 310 LiverBlubber 80.7 2.1
S-5 Male 290 410 Liver-Blubber 81.6 4.2
S-6 Male 260 310 LiverBlubber 78.0 3.0
S-7 Male 295 210 LiverBlubber 81.7 2.8
S-8 Male 265 310 LiverBlubber 89.9 2.7
S-9 Male 320 600 LiverBlubber 64.2 2.9
S-10 Male 325 520 Blubber 86.3
Nemuro Strait in 2010 N-1a Female 240 LiverBlubber 86.0 2.6
N-2a Female 230 LiverBlubber 93.2 4.6
a
S-4, N-1, N-2 were pregnant.

Fig. 1. Map of the study area and locations.

dehydration, then were extracted three times with diethyl ether-
hexane (1:2 (v/v)) for 20 min at 300 rpm using a reciprocating
shaker (SR-2w:TAITEC, Japan). After concentrating the extracted
solvents and removing all solvents, the lipid content was deter-
mined gravimetrically as taking account of an aliquot of the
extract. This lipid residue was dissolved completely with hexane
in a warm-water bath. In 1% or 2% of the extracted solvent,
13
C12-labeled PCBs were spiked to examine the recovery rate
through sample treatment. 50 ml of 1 mol/L KOH/Methanol solu-
tion was added to this aliquot, stirred calmly for 16 h at room tem-
perature, and then the solution was cleaned using an arranged
multi-layered silica-gel column. For high-resolution gas chromato-
graph/high-resolution mass spectrometry (HRGCHRMS) analysis,
this cleaned solution was converted to about 50 ll nonane solu-
tion, and 13C12-labeled recovery standards were spiked.
Identication and quantication of PCB congeners were per-
formed by HRGC (HP6890, Agilent Technologies, Inc., USA)HRMS
(JMS-700D, JEOL, Japan) according to the procedures described in
the manual (Environmental Monitoring Report on Persistent
Organic Pollutants (POPs) in Japan, 20022004). PCB congeners
were analyzed with a HT8-PCB capillary column (60 m,
0.25 mm i.d., Kanto chemical Co., Inc., Japan). There were 193
peaks in the HRGCHRMS chromatogram, that is, peaks for 178
congeners and 15 peaks for co-elutions including 31 congeners.
Some peaks could not be isolated, so these peaks on the chromato-
gram are shown as a summation of the concentrations. All congen-
ers were referred to by the IUPAC number.
PCBs were quantied using an isotope dilution method to the
corresponding 13C12-labels. The chromatograph peaks with signal
to noise ratios (S/N) higher than three, and with an ion intensity
ratio between the main channel and the subchannel that was with-
in 15%, were identied and determined. We analyzed procedural
blanks, and checked any PCBs that were not detected in these
blanks. The limits of sample detection (LOD) for mono-chlorinated
biphenyls (MoCBs), di-chlorinated biphenyls (DiCBs), tri-chlori-
nated biphenyls (TrCBs), tetra-chlorinated biphenyls (TeCBs), pen-
ta-chlorinated biphenyls (PeCBs), hexa-chlorinated biphenyls
(HxCBs), hepta-chlorinated biphenyls (HpCBs), octa-chlorinated
biphenyls (OcCBs), nona-chlorinated biphenyls (NoCBs) and deca-
chlorinated biphenyl (DeCB) in the liver were 0.060.1 ng/g,
0.070.2 ng/g, 0.050.1 ng/g, 0.030.1 ng/g, 0.030.2 ng/g, 0.06
0.1 ng/g, 0.060.2 ng/g, 0.060.2 ng/g, 0.060.2 ng/g, and 0.08
0.2 ng/g, respectively. And the LOD for MoCBs, DiCBs, TrCBs, TeCBs,
PeCBs, HxCBs, HpCBs, OcCBs, NoCBs and DeCB in blubber were 0.1
0.2 ng/g, 0.20.4 ng/g, 0.10.3 ng/g, 0.10.4 ng/g, 0.20.5 ng/g, 0.2
0.4 ng/g, 0.10.4 ng/g, 0.20.4 ng/g, 0.30.4 ng/g, and 0.30.6 ng/g,
respectively. The recovery rates for most of the 13C12-labeled PCB
congeners ranged from 40% to 120%, except for a few congeners
230 K. Kubo et al. / Marine Pollution Bulletin 69 (2013) 228232

Table 2
RIPCBs (ng/g-fat) in the blubber and liver of SSLs.

Region/sample Male Female

Blubber Liver Blubber Liver
This study (in 20082010) 2700 750 (n = 9) 3000 830 (n = 8) 910 400 (n = 3) 2000 1500 (n = 3)
Western Hokkaido 1994a 2400 2500
Alaskab 9000 6100 (n = 13) 17,000 12,000 (n = 12) 4000 4000 (n = 15) 5300 5400 (n = 17)
Bering seab 4500 1500 (n = 14)
a
Kim et al. (1996).
b
Lee et al. (1996).

#153 of HxCBs was predominated congener in both of the blub-

Table 3
Concentration (the mean SD) of congeners (ng/g-fat); organ-wise and gender ber and liver, and #138 of HxCBs, #99 and #118 of PeCBs, and
differences were explained in blubber and liver of males and females. #180 of OcCBs also existed in both types of samples. Generally,
these congeners are known to be major congeners in marine mam-
Congeners Male Female
mals (Boon et al., 1994; Nakata et al., 1997). A distinct differentia-
Blubber (n = 9) Liver (n = 8) Blubber (n = 3) Liver (n = 3)
tion was not observed between the concentrations of these ve
TrCBs major congeners in the samples of male blubber and liver. In con-
#28 110 96 19 18 72 41 15 2.7 trast, #28 and #31 of TrCBs, #177 of HpCBs, and #199 of OcCBs
#31 95 75 8.4 14 50 30 0.46 0.18
showed signicant differences in the concentration between these
PeCBs two organs (Table 3). In addition, the prole from the number of
#99 180 44 190 72 30 19 94 64
#101 52 38 51 21 23 16 51 69
chlorines in PCBs in the blubber and liver depended on the conge-
#105 62 17 81 29 12 6.9 47 34 ner prole in each organ. The differences in the distribution of the
#118 180 47 160 52 39 24 110 79 type of the number of chlorines between the blubber and the liver
HxCBs were shown in TrCBs, HpCBs and OcCBs. These organ-wise differ-
#138 250 72 290 130 43 19 140 92 ences seem to be appeared from the differences in the function
#153 460 140 380 130 78 36 210 140 of each organ of SSLs.
HpCBs In the female blubber, #153 was the most common congener,
#177 6.1 5.4 120 52 2.1 0.84 62 42 followed by #28, #31, #138, and #118. The concentrations of their
#180 89 24 130 76 19 12 75 52
major congeners were almost at the same levels. These only ve
OcCBs congeners dominated 31% of RPCBs in the female blubber. In the
#199 8.3 3.2 76 57 2.7 1.5 40 39
female liver, #153 was also predominated, followed by #138,
#118, #99 and #180. The concentrations of these ve congeners
dominated 32% of RPCBs in the female liver. In the female, the
in some samples 40118% in liver and 33118% in blubber, organ dependency appeared in the concentration of #28, #31,
respectively. #177 and #199 as the same as those in males (Table 3). It indicated
The total concentrations of PCBs (RPCBs) in the blubber and that the differences in these concentration proles might be due to
liver are shown in Table 2. RPCBs in the blubber and liver of males the differences in organs and their functions.
were at the same level; the average standard deviation (SD) in Fig. 2 showed that there were large differences in the concen-
the blubber was 2700 750 ng/g-fat (n = 9), and the average in trations of some congeners in the blubber between males and
the liver was 3000 830 ng/g-fat (n = 8). Lee et al. (1996) reported females. RPCBs in females were about 66% of that in the blubber
that RPCBs in the liver of SSLs were clearly correlated with those in of male and about 33% in the liver of male. The contamination level
the blubber. RPCBs in SSLs in 2008 from the western Hokkaido of males was reported to be higher than that of females (Wolkers
were in the same range as those found in 1994, as reported by et al., 2004), because females could excrete the lipophilic contam-
Kim et al. (1996) from the same region (Table 2). These results sug- inants by maternal-to-fetal transfer via the placenta and lactation
gest that RPCBs of SSLs in this region continued to be at the same throughout their lifespan (Filho et al., 2009; Wolkers et al.,
levels from 14 years ago. RPCBs of SSLs in this region were lower 2004). The females in this study were pregnant and had fetuses
than those of SSLs in other regions such as the Bering Sea (Kim in their bodies (Table 1), and it could be estimated that these
et al., 1996) and Alaska (Lee et al., 1996). The dependency RPCBs females had experienced the maternal-to-fetal transfer at least
in SSLs in migratory areas is shown clearly. The residue levels of once before.
some organochlorine insecticides have declined slowly, while PCBs The differences in the proles of PCB congeners between males
remain at a steady state in the open oceans for a long time (Tanabe and females were not clear in the liver, but distinct in the blubber
et al., 2003, 1994b). The residue level of PCBs in SSLs from Hokka- (Fig. 2). In the female blubber, the concentrations of #28 and #31
ido will probably remain constant, even in the future. were almost at the same levels as those of #153, #138, #118, and
The concentrations of individual PCB congeners of the blubber #99, all of which were major congeners in males. In particular,
and liver are shown in Table 3. In the blubber #153 is a predomi- major congeners of blubber in males such as #153 and #138
nant congener, the concentration was 460 140 ng/g-fat, which showed signicant gender differences (Fig. 2). The concentration
corresponded to 17% of the average of RPCBs. #153 was followed of #138 in the males was 5.8 times as large as those in the females;
by #138, #99, #118, #28, and #31. The total amount of the six the concentration of #153 in the males was 6.2 times as large as
congeners reached to about 47% of RPCBs in the blubber. In the those in females. On the other hand, the concentration of #28
liver, #153 was the predominate congener where the concentra- and #31, which were major congeners in the females, and the
tions were 380 130 ng/g-fat on average, followed by #138, #99, congeners from OcCBs to DeCB showed no gender differences as
#118, #180, and #177. #153 was 13% of RPCBs. The amount of Imaeda et al. (2009) reported in Baikal Seals. The concentration
these six congeners, including #153, was more than 100 ng/g-fat prole between genders differed with HxCBs such as #138 and
in the liver and reached to 44% of RPCBs. #153 suggests that the behaviors of these congeners could be
 K. Kubo et al. / Marine Pollution Bulletin 69 (2013) 228232 231

700

600
Blubber (male)

Concentration (ng/g-fat)
Blubber (Female)
500

400

300

200

100

0
#28 #31 #99 #101 #105 #118 #138 #153 #177 #180 #199

TrCBs PeCBs HxCBs HpCBs OcCBs

600

500 liver (male)

Concentration (ng/g-fat)

liver(Female)
400

300

200

100

0
#28 #31 #99 #101 #105 #118 #138 #153 #177 #180 #199

TrCBs PeCBs HxCBs HpCBs OcCBs

Fig. 2. The differentiations between major congener concentrations (ng/g-fat) in the blubber and liver of male and female SSLs from the Hokkaido coast.The upper graph
shows those in the blubber and the lower graph shows those in the liver. The concentrations of males and females are indicated as the mean SD.

dependent on lactation from the blubber into milk (Wolkers et al.,
2002, 2004; Imaeda et al., 2009) in SSLs. The study in more detail is
needed on the transfer of the PCB congeners from the mother to
the fetus and to the pups of marine mammals.
It was reported that the half-lives of PCB congeners in marine
mammals were signicantly different depending on each congener
(Imaeda et al., 2009). Although the difference in RPCBs between
the blubber and the liver was not observed, the differences in the
distribution of major congeners and in the type of congeners
between these two organs were observed obviously. This detailed
information could be a useful knowledge for the study on the bio-
logical effect and toxicity of PCBs and PCB hydroxides (OH-PCBs) as
metabolites of PCBs, which have been remain unclear. In order to
assess PCBs in marine mammals, it is necessary to investigate
not only RPCBs but also the detailed concentrations of every con-
gener in the organs. The data of the concentration of every PCB
congeners in marine mammals will be useful and precious infor-
mation to understand the accumulation and movement of PCBs
in marine mammals, and the biological effect on marine lives.
Acknowledgments
The authors wish to express their gratitude to Dr. Orio Yamamura
(Hokkaido National Fisheries Research Institute, Fisheries Research
Agency), Dr. Tsuyoshi Ishinazaka (Shiretoko Nature Foundation),
M.Sc. Katsunori Anezaki and M.Fi. Ryuji Fukuyama (Hokkaido Re-
search Organization) for their support and advice during our inves-
tigation. Samples of SSLs were collected from the Actualization
project for the alleviation of damages from harmful animals com-
missioned by the National Federation of Fisheries Cooperative Asso-
ciations. This study was supported partially by a grant for ordinary
research from the Hokkaido Research Organization.
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