Dorneles et al 2013 DRCs dolphins Brazil

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High accumulation of PCDD, PCDF, and PCB congeners in marine
mammals from Brazil: A serious PCB problem
Paulo R. Dorneles
a,
⁎, Paloma Sanz
b
, Gauthier Eppe
c
, Alexandre F. Azevedo
h
, Carolina P. Bertozzi
d
,
María A. Martínez
b
, Eduardo R. Secchi
e
, Lupércio A. Barbosa
f
, Marta Cremer
g
, Mariana B. Alonso
a,h
,
João P.M. Torres
a
, José Lailson-Brito
h
, Olaf Malm
a
, Ethel Eljarrat
i
, Damià Barceló
i,j
, Krishna Das
k
a
Biophysics Institute, Federal University of Rio de Janeiro (UFRJ), Brazil
b
Persistent Organic Pollutant Group, Environment Department, CIEMAT, Madrid, Spain
c
Centre for Analytical Research and Technology, Mass Spectrometry Laboratory, Liege University, Belgium
d
Projeto Biopesca, Praia Grande, SP, Brazil
e
Laboratório de Tartarugas e Mamíferos Marinhos, Instituto de Oceanografia e Museu Oceanográfico “Prof. E.C. Rios”, Universidade Federal do Rio Grande (FURG), Rio Grande, RS, Brazil
f
Instituto ORCA, Vila Velha, ES, Brazil
g
Universidade da Região de Joinville, Departamento de Ciências Biológicas, Laboratório de Nectologia, São Francisco do Sul, SC, Brazil
h
Aquatic Mammal and Bioindicator Laboratory (MAQUA), School of Oceanography, Rio de Janeiro State University (UERJ), Brazil
i
Department of Environmental Chemistry, IDAEA, CSIC, Barcelona, Spain
j
Catalan Institute for Water Research (ICRA), Parc Científic i Tecnològic de la Universitat de Girona, Girona, Spain
k
Laboratory for Oceanology, MARE Center, Liege University, Belgium
HIGHLIGHTS
•Dioxin-like PCBs accounted for over 83% of the total TEQ for all cetaceans
•Negative correlations were found between length and TEQ values in franciscanas.
•PCB concentrations found are among the highest ever reported for cetaceans.
abstractarticle info
Article history:
Received 9 March 2012
Received in revised form 3 June 2013
Accepted 3 June 2013
Available online xxxx
Editor: Adrian Covaci
Keywords:
Dioxins
Furans
PCBs
Dolphins
Brazil
Blubber samples from three delphinid species (false killer whale, Guiana and rough-toothed dolphin), as well
as liver samples from franciscana dolphins were analyzed for dioxins and related compounds (DRCs). Sam-
ples were collected from 35 cetaceans stranded or incidentally captured in a highly industrialized and urban-
ized area (Southeast and Southern Brazilian regions). Dioxin-like PCBs accounted for over 83% of the total
TEQ for all cetaceans. Non-ortho coplanar PCBs, for franciscanas (82%), and mono-ortho PCBs (up to 80%),
for delphinids, constituted the groups of highest contribution to total TEQ. Regarding franciscana dolphins,
significant negative correlations were found between total length (TL) and three variables, ΣTEQ-DRCs,
ΣTEQ-PCDF and ΣTEQ non-ortho PCB. An increasing efficiency of the detoxifying activity with the growth
of the animal may be a plausible explanation for these findings. This hypothesis is reinforced by the signifi-
cant negative correlation found between TL and PCB126/PCB169 concentration ratio. DRC concentrations
(ng/g lipids) varied from 36 to 3006, for franciscana dolphins, as well as from 356 to 30,776, for delphinids.
The sum of dioxin-like and indicator PCBs varied from 34,662 to 279,407 ng/g lipids, for Guiana dolphins
from Rio de Janeiro state, which are among the highest PCB concentrations ever reported for cetaceans.
The high concentrations found in our study raise concern not only on the conservation of Brazilian coastal ce-
taceans, but also on the possibility of human health problem due to consumption of fish from Brazilian
estuaries.
© 2013 Elsevier B.V. All rights reserved.
Science of the Total Environment 463–464 (2013) 309–318
⁎Corresponding author at: Universidade Federal do Rio de Janeiro (UFRJ), Centro de Ciências da Saúde (CCS), Instituto de Biofísica Carlos Chagas Filho (IBCCF), Laboratório de
Radioisótopos Eduardo Penna Franca (LREPF), Avenida Carlos Chagas Filho, 373 (Edifício do CCS), sala G0-62, Cidade Universitária, 21941-900 Rio de Janeiro, RJ, Brazil. Tel.: +55 21
25615339; fax: +55 21 22808193.
E-mail addresses: dorneles@biof.ufrj.br,dornelespr@gmail.com (P.R. Dorneles), g.eppe@ulg.ac.be (G. Eppe), azevedo.alex@uerj.br (A.F. Azevedo), carolinabertozzi@hotmail.com
(C.P. Bertozzi), edu.secchi@furg.br (E.R. Secchi), lupercio@orca.org.br (L.A. Barbosa), marta.cremer@univille.br (M. Cremer), alonso.mb@gmail.com (M.B. Alonso),
jptorres@biof.ufrj.br (J.P.M. Torres), lailson@uerj.br (J. Lailson-Brito), olaf@biof.ufrj.br (O. Malm), eeeqam@iiqab.csic.es (E. Eljarrat), krishna.das@ulg.ac.be (K. Das).
0048-9697/$ –see front matter © 2013 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.scitotenv.2013.06.015
Contents lists available at SciVerse ScienceDirect
Science of the Total Environment
journal homepage: www.elsevier.com/locate/scitotenv

1. Introduction
Special concern has been raised since the 1960s about the environ-
mental persistence, bioaccumulative capacity and toxicity of dioxins
and related compounds (DRCs), such as polychlorinated dibenzo-p-
dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), as well as
non-ortho and mono-ortho polychlorinated biphenyls (dioxin-like
PCBs) (Bopp et al., 1991; Cleemann et al., 2000). DRCs constitute a
group of polyhalogenated aromatic hydrocarbons that have similarities
from the structural, chemical andtoxicological point of view (McDonald
et al., 1994). These persistent and bioaccumulative contaminants are
carcinogenic (Chaloupka et al., 1993; Cleemann et al., 2000), immuno-
suppressive (Harper et al., 1993), and induce a number of pathological
effects related to their endocrine disrupting action in mammals, in-
cluding reproductive impairment (Schecter et al., 2006). Despite their
toxicity, studies on environmental levels of DRCs in Brazil are scarce
(de Assunção et al., 2005).
Dioxins are unintentionally-produced contaminants, generated by
different processes, including paper bleaching, synthesis of pesticides,
herbicides, fungicides and PCBs, as well as burning of wastes and veg-
etation (Alcock et al., 1999; Cleemann et al., 2000; McDonald et al.,
1994; Schecter et al., 2006). Taking the latter activity into consider-
ation, it is interesting to mention that intentional burning of sugar
cane crops and forest fires are common in Brazil (Lara et al., 2005;
Santilli et al., 2005).
Polychlorinated biphenyls (PCBs) were used in a number of indus-
trial activities, but mainly as dielectric fluids in capacitors and trans-
formers (Alcock et al., 1999). Manufacture, use and commercialization
of PCBs have been prohibited in Brazil since the beginning of the
1980s (Azevedo e Silva et al., 2007; Penteado and Vaz, 2001). However,
the same restrictive law allows the functioning of already installed ap-
paratuses up to their complete replacement or substitution of their di-
electric fluid (Azevedo e Silva et al., 2007; Penteado and Vaz, 2001).
Industrial and urban developments, at present, spread along the entire
Brazilian coast; however, they are particularly concentrated along the
Southeastern and Southern Brazilian regions (Saboia, 2000). Therefore,
evaluating environmental contamination by DRCs in these two regions
constitutes a matter of interest.
Using samples from cetaceans, the present study constitutes the
first scientific investigation on DRC levels in marine biota from Brazil.
Due to their high level in the food chain, their long life-span, and, for
some species, their year-round presence in polluted and relatively
small areas, cetaceans are important indicators of environmental con-
tamination (Bjørge, 2001; O'Shea and Tanabe, 2003). In addition to
providing information on the pattern and magnitude of DRCs that
are reaching the top positions of the marine food webs in Brazil, anal-
yses of cetacean tissues provide valuable data for conservation pur-
poses. The high marine mammal exposure to DRCs in different
regions of the globe raises concern about the conservation of the spe-
cies, as adverse effects on reproductive and immune systems have
been demonstrated for these animals (Dietz et al., 1989; Reijnders,
1986). The Brazilian economic growth and the subsequent use and
degradation of the coastal zone are a concern for the conservation
of small coastal cetaceans. In the present study, samples from four ce-
tacean species (franciscana dolphin, Pontoporia blainvillei; Guiana
dolphin, Sotalia guianensis; rough-toothed dolphin, Steno bredanensis;
and false killer whale, Pseudorca crassidens) were analyzed for DRCs.
Franciscana and Guiana dolphins are small cetaceans that can
reach up to 1.5 and 2.0 m, respectively. They occur exclusively in
western Atlantic coastal waters of South (franciscana and Guiana dol-
phins) and Central (Guiana dolphins) America (Cunha et al., 2005;
Secchi et al., 2003). Due to their nearshore distribution, these dolphin
species are especially vulnerable to the effects of human activities.
Rough-toothed dolphins and false killer whales occur in tropical to
warm temperate waters all over the world (Baird, 2009; Miyazaki
and Perrin, 1994). In Brazil, these species occur predominantly on
the continental shelf (Bisi, 2011; Dorneles et al., 2010, 2008b,
2007a) and rough-toothed dolphins can even be found in shallow
coastal waters (Flores and Ximinez, 1997). Rough-toothed dolphins
can occasionally reach more than 2.8 m of total length (Jefferson,
2009). Taking into consideration that this continental shelf (CS)
delphinid has many prey species in common with estuarine dolphin
species, such as the Guiana dolphin (Melo et al., 2010, submitted for
publication), the greater size of rough-toothed dolphins means that
these CS delphinids have anatomical characteristics that enable them
to feed on larger fish, and hence, on prey specimens that had a longer
exposure time to pollutants than those consumed by Guiana dolphins.
Although it is still controversial whether or not false killer whales feed
on other cetacean species, they are known to prey on large and
long-lived fish such as tunas and swordfish (Baird et al., 2008). This fea-
ture, combined with the hypothesis that organohalogen compounds
from bays and estuaries of the Southeastern Brazilian region are
transported to the adjacent Atlantic Ocean waters (Dorneles et al.,
2010), makes DRC determination in rough-toothed dolphins and false
killer whales a matter of great scientific interest.
Because of these reasons, cetaceans were used in the present
study as indicators of micropollutant trophic flow, as well as sentinel
species for human health. The latter approach is based on the fact that
many fish species are consumed by humans in Rio de Janeiro at the
size preyed on by the delphinid species analyzed in our study (Melo
et al., 2010, submitted for publication). In addition, it is also of great
interest to investigate if accumulation of persistent bioaccumulative
toxicants can pose an additional threat to these cetacean species.
2. Materials and methods
2.1. Sampling and sample preparation
Liver and blubber samples were obtained by different marine
mammal research groups from five Brazilian states (Fig. 1), including
the Southeastern [Espírito Santo (ES), Rio de Janeiro (RJ) and São
Paulo (SP) states] and the Southern [Santa Catarina (SC) and Rio
Grande do Sul (RS) states] regions of the country. They were collected
through the necropsy of cetaceans either incidentally captured in
fishing operations or stranded on the beaches. Liver samples were
collected from franciscana dolphins, comprising fourteen males
from Rio Grande do Sul (RS, n = 3), Santa Catarina (SC, n = 2), São
Paulo (SP, males, n = 6) and Espírito Santo (ES, n = 3) states, as well
as six female franciscana dolphins from São Paulo (SP, females, n = 6)
state. Blubber samples were collected in RJ from 15 delphinids, com-
prising 11 Guiana dolphins (males, n = 7; females, n = 4), three
rough-toothed dolphins (males, n = 1; females, n = 2) and one false
killer whale (female, n = 1). After dissection, samples were wrapped
in aluminum foil and kept frozen (−20 °C) until analyses.
2.2. Analytical procedure—targeted compounds
The following dioxins and furans were targeted for analysis:
2,3,7,8-Tetra CDD; 1,2,3,7,8-Penta CDD; 1,2,3,4,7,8-Hexa CDD; 1,2,3,
6,7,8-Hexa CDD; 1,2,3,7,8,9-Hexa CDD; 1,2,3,4,6,7,8-Hepta CDD;
Octa CDD (OCDD); 2,3,7,8-Tetra CDF; 1,2,3,7,8-Penta CDF; 2,3,4,7,8-
Penta CDF; 1,2,3,4,7,8-Hexa CDF; 1,2,3,6,7,8-Hexa CDF; 1,2,3,7,8,
9-Hexa CDF; 2,3,4,6,7,8-Hexa CDF; 1,2,3,4,6,7,8-Hepta CDF; 1,2,3,
4,7,8,9-Hepta CDF; Octa CDF (OCDF). Concerning dioxin-like PCBs,
the following congeners (IUPAC numbers) were targeted for analysis:
77; 81; 126; 169; 105; 114; 118; 123; 156; 157; 167 and 189. Toxic
equivalent (TEQ) concentrations of PCDD/Fs and dioxin-like PCBs
were calculated using the World Health Organization (WHO)-2005
toxic equivalency factors (TEF) (van den Berg et al., 2006). Concentra-
tions below detection limits were considered as zero (lower bound
TEQ). For blubber samples, in addition to all the mentioned above
310 P.R. Dorneles et al. / Science of the Total Environment 463–464 (2013) 309–318

DRCs, levels of indicator-PCBs (IUPAC numbers: 28, 52, 101, 138, 153,
180) were also measured.
2.3. Analytical procedure—liver sample analysis
The analytical procedure has been detailed elsewhere (Eljarrat et
al., 2008). Briefly, samples were fortified with
13
C
12
-labeled PCDD/F
and
13
C
12
-labeled PCB quantification standard solutions (Wellington
Laboratories Inc., Canada), EPA 1613 LCS and WP-LCS, and extracted
using a Dionex ASE100 at the following conditions: hexane, 100 °C,
1500 psi, 90% flush volume and two static cycles (Focant et al.,
2001). After extraction, the solvent was removed and, subsequently
fat content was determined gravimetrically. Resulting extracts were
transferred into a separation funnel and liquid-extracted with con-
centrated sulphuric acid to remove organic matter. Clean-up stage
was performed in an automated purification Power Prep™System
(FMS, Inc., USA) including acidic silica gel and basic alumina columns
for mono-ortho PCB purification and an additional carbon column for
PCDD/F and coplanar PCB cleanup. Different mixtures of hexane:DCM
were used to recover mono-ortho PCBs while retaining interfering
compounds. PCDD/Fs and coplanar PCBs were recovered with toluene.
The final extracts were concentrated avoiding dryness, spiked with
EPA1613-ISS and WP-ISS internal standard solutions (Wellington Labo-
ratories Inc., Canada) and further analyzed by GC–HRMS.
2.4. Analytical procedure—blubber sample analysis
The extraction, purification and measurement methods are described
elsewhere (Focant et al., 2001). Briefly, blubber samples were extracted
by pressurized liquid extraction (PLE) using a Dionex (Sunnyvale, CA,
USA) ASE 200 extractor. Conditions were: 33 ml extraction cells filled
approximately 5 g of blubber sample and sodium sulphate, 20 ml of hex-
ane per cycle, 5 min cycle time, two cycles per extraction, pressure of
1500 p.s.i. The fat extracts were dried on sodium sulphate prior to com-
plete solvent evaporation. A solution of hexane/dichloromethane was
added to aliquots of about 200–300 mg of extracted lipids. The lipid ex-
tracts were then spiked with a mixture containing seventeen
13
C-labeled
2,3,7,8-substituted dioxins isomers, 4 coplanar PCBs isomers (EDF-4144,
LGC Promochem) and 8 mono-ortho PCB isomers (Campro Scientific
WP-LCS). An automated multi-column clean-up was performed on the
Power-Prep system (FMS, Waltham, MA, USA). The lipid content was de-
termined gravimetrically. All analyses were performed by GC–HRMS
using a MAT95XL high-resolution mass spectrometer (Finnigan, Bremen,
Germany) and a Hewlett-Packard (Palo Alto, CA, USA) 6890 series gas
chromatograph.
2.5. Statistical treatment
Due to the small sample sizes, non-parametric tests were used
for investigating possible correlations between compounds and
Fig. 1. Brazilian map amplifying Rio Grande do Sul (RS), Santa Catarina (SC), São Paulo (SP), Rio de Janeiro (RJ) and Espirito Santo (ES) states. Guanabara Bay is complementarily
amplified.
311P.R. Dorneles et al. / Science of the Total Environment 463–464 (2013) 309–318

biological parameters (Spearman's correlation test), as well as for in-
vestigating possible differences between species, sexes, and regions,
regarding pollutant levels and profiles (Mann–Whitney U test). Due
to the low number of individuals of some species/groups, only data
from franciscana dolphins from SP and Guiana dolphins from RJ
were used for statistical evaluation. The Statistica 7.0 Statistical Soft-
ware System was used for statistical analyses and the level of signifi-
cance was set at p ≤0.05.
3. Results and discussion
PCDD, PCDF and dioxin-like (non-ortho and mono-ortho) PCB
concentrations and TEQ values (pg.g
−1
, l.w.), as well as percentages
to ΣTEQ, in liver or blubber of cetaceans from Southeastern and
Southern Brazilian regions, are presented in Table 1.
The use of TEFs to achieve TEQ constitutes an accurate procedure
for the assessment of the toxic potential of a complex mixture of pol-
lutants that are capable of triggering Aryl hydrocarbon (Ah) receptor-
mediated effects, such as DRCs (Sanderson and Van Den Berg, 1999).
Considering that metabolization of DRCs is also mediated by the Ah
receptor, as well as that metabolizing rates for xenobiotics may vary
according to age (Reijnders, 2003) and gender (Hall, 2002), possible
correlations with the total length and possible sex-related differences
were investigated not only for DRC concentrations but also for TEQ
values.
3.1. Franciscana dolphins
Dioxin-like PCBs accounted for 83–96% of the total TEQs in the he-
patic tissues of franciscana dolphins. In this context, non-ortho copla-
nar PCBs deserve to be highlighted, since the group contributed to
82% of the total TEQs, on average. Among the latter group, PCB-126
was by far the congener of greatest importance. In fact, of the mea-
sured DRCs, PCB-126 contributed to the majority of the total TEQ in
all franciscana dolphins (mean 78%, SD 11%). However, PCB-118 oc-
curred in the highest concentrations, which varied from 19.7 to
563.9 ng/g (l.w.). PCDDs and PCDFs accounted for, on average, 10%
of the total TEQs. The predominant PCDD/F congener found in the
livers of franciscana dolphins was OCDD, with concentrations ranging
between 32 and 1093 pg/g (l.w.). Concentrations of PCDDs were
greater than those of PCDFs in all individuals. However, regarding
the TEQ values (TEQ pg/g l.w.), levels were in general higher for
PCDFs. Among furans, 2,3,4,7,8-PeCDF provided the highest contribu-
tion to total TEQ.
It is important to keep in mind that while analyzing franciscana
dolphins from RS, SC, SP and ES, individuals from three different
management areas are being considered. It has been proposed that
franciscana should be split into four stocks for management purposes,
three of which occur in Brazilian waters (Secchi et al., 2003). Each
stock inhabits discrete areas named Franciscana Management Areas
(FMA): FMA I, including coastal waters of ES and RJ states; FMA II,
covering SP to SC states; and FMA III, comprising the coastal waters
of RS state and Uruguay (Secchi et al., 2003). A recent study from
our research team (Lailson-Brito et al., 2011) has shown that different
ecological franciscana dolphin populations could exhibit remarkably
distinct organochlorine compound bioaccumulation profiles, charac-
terized by different ΣDDT/ΣPCB ratios, even within the same manage-
ment area (FMA II). However, in our study there was no distinction
among areas. Instead, there was great variation among individuals
(Fig. 2).
According to Ballschmiter and Wittlinger (1991), the environmen-
tal levels of many organohalogen compounds in Northern and South-
ern Hemispheres are a consequence of emissions within each half of
the planet independently. As DRCs are pollutants of urban/industrial
origin, it is worth mentioning that SP houses a human population of
approximately 40 million people (Rudge et al., 2011). In addition,
the metropolitan area of São Paulo city constitutes the most industri-
alized center of Latin America (Saldiva et al., 1994). In this context, it
becomes interesting to compare the organochlorine compound pro-
files between franciscana dolphins from SP state and air samples
from São Paulo City (de Assunção et al., 2005). Considering the
PCDF profiles specifically (Fig. 2), for all individuals from SP state,
with the exception of BP136, a higher contribution of PeCDF to
ΣPCDF in dolphins than in air could be observed. The contribution
of PeCDF in dolphin liver samples (excluding BP136 from the analysis)
varied from 25.3 to 65.5% (50.8 ± 11.7, Mean ± SD), while in air sam-
ples from São Paulo City (de Assunção et al., 2005) a percentage of 12.5
could be observed. A possible explanation for this lies in the selective
biomagnification of 2,3,4,7,8-PeCDF reported in literature (Rolff et al.,
1993).
For investigating possible gender-related differences on DRC
bioaccumulation by franciscana dolphins from SP state, comparisons be-
tween sexes were performed regarding ΣPCDD, ΣPCDF, ΣTEQ-PCDD,
ΣTEQ-PCDF, Σnon-ortho PCB, Σortho PCB, ΣTEQ non-ortho PCB, ΣTEQ
ortho PCB, ΣDL-PCB, ΣTEQ-DRCs, as well as the percent contribution
to total TEQ (% to ΣTEQ-DRCs) of ΣTEQ-PCDD, ΣTEQ-PCDF, ΣTEQ
non-ortho PCB and ΣTEQ ortho PCB. No significant differences were
observed.
Table 1
Hepatic (franciscana dolphins) and blubber (delphinids) PCDD (pg.g
−1
l.w.), PCDF (pg.g
−1
l.w.) and PCB (ng.g
−1
l.w) concentrations, total TEQ (ΣTEQ-DRCs or T-TEQ, pg.g
−1
l.w.)
and percentages to T-TEQ (Mean (Median) ± SD; [n] Min–Max) of cetaceans from Brazil.
Franciscana dolphin Guiana dolphin RT dolphin FK whale
Males Females Males Females Male Females Female
ΣPCDD/DFs
pg/g lipids
349 (141) ± 433
[13] 77.4–1554
448 (492) ± 173
[5] 162–629
52.2 (31.6) ± 35.8
[7] 26.7–123
27.3 (17.5) ± 29.3
[4] 5.13–69.2
109
[1] [2] 17.4–2060
136
[1]
ΣPCDD/DFs % to T-TEQ
percentage
8.89 (8.79) ± 2.92
[11] 4.40–13.5
12.3 (11.4) ± 2.76
[5] 9.76–16.6
0.65 (0.48) ± 0.41
[7] 0.17–1.16
0.20 (0.17) ± 0.13
[4] 0.09–0.38
1.61
[1] [2] 4.82–13.9
0.98
[1]
Σnon-ortho PCBs
ng/g lipids
2.79 (1.81) ± 2.43
[13] 0.21–8.17
4.22 (2.61) ± 4.45
[6] 1.40–13.1
5.49 (3.87) ± 3.81
[7] 2.35–13.1
8.30 (1.78) ± 13.2
[4] 1.49–28.1
13.0
[1] [2] 0.76–5.19
10.3
[1]
Σnon-ortho PCBs to T-TEQ
percentage
81.6 (85.0) ± 13.7
[11] 43.3–91.3
82.9 (83.4) ± 2.24
[5] 79.8–85.0
35.6 (34.0) ± 9.36
[7] 24.9–49.4
28.9 (24.8) ± 12.0
[4] 19.6–46.4
41.6
[1] [2] 13.6–69.4
44.1
[1]
Σortho PCBs
ng/g lipids
429 (170) ± 828
[12] 35.0–3000
139 (87.0) ± 124
[6] 61.5–385
13,080 (13,060) ± 2810
[7] 7551–15,804
14,396 (11,116) ± 11,643
[4] 4606–30,748
11,054
[1] [2] 355–19,844
16,179
[1]
Σortho PCBs to T-TEQ
percentage
9.54 (4.2) ± 12.1
[11] 1.80–43.2
4.81 (4.58) ± 1.51
[5] 3.32–7.06
63.7 (65.1) ± 9.57
[7] 49.5–75.0
70.9 (75.0) ± 12.0
[4] 53.5–80.4
56.8
[1] [2] 25.8–72.5
54.9
[1]
ΣTEQ-DRCs (T-TEQ)
pg/g lipids
103 (67.2) ± 76.4
[11] 38.9–276
86.5 (85.0) ± 49.1
[5] 33.7–164
622 (568) ± 150
[7] 458–886
690 (423) ± 706
[4] 187–1725
583
[1] [2] 41.3–822
885
[1]
ΣPCBs (ind. + DL PCBs)
ng/g lipids
Not determined Not determined 100,290 (78,365) ± 44,751
[7] 56,096–160,355
107,865 (58,695) ± 115,796
[4] 34,662–279,407
74,705
[1] [2] 2510–167,079
122,004
[1]
312 P.R. Dorneles et al. / Science of the Total Environment 463–464 (2013) 309–318

HpCB
HxCB
PeCB
TeCB
RS-CA142
RS-CA143
SC-Pb22
SC-Pb221
SP-BP104
SP-BP116
SP-BP125
SP-BP136
SP-BP156
ES-PON09
ES-PON11
0
20
40
60
80
100
OCDF
HpCDF
HxCDF
PeCDF
TCDF
RS-CA142
RS-CA143
SC-Pb22
SC-Pb221
SP-BP104
SP-BP116
SP-BP125
SP-BP136
SP-BP156
ES-PON09
ES-PON11
0
20
40
60
80
100
OCDD
HpCDD
HxCDD
PeCDD
TCDD
RS-CA142
RS-CA143
SC-Pb22
SC-Pb221
SP-BP104
SP-BP116
SP-BP125
SP-BP136
SP-BP156
ES-PON09
ES-PON11
0
20
40
60
80
100
A
B
C
Fig. 2. Relative contribution of PCBs (Fig. 2-A), PCDFs (Fig. 2-B) and PCDDs (Fig. 2-C) grouped by the number of chlorine atoms in the molecule (e.g. tetra-chlorinated biphenyls,
TeCB; penta-chlorinated dibenzofurans, PeCDF; and hexa-chlorinated dibenzodioxins, HxCDD) to ΣPCB (Fig. 2-A), ΣPCDF (Fig. 2-B) and ΣPCDD (Fig. 2-C), in liver samples of
male franciscana dolphins from the Brazilian states of Rio Grande do Sul (RS), Santa Catarina (SC), São Paulo (SP) and Espírito Santo (ES). The figure exposes the individual
codes of each dolphin (e.g. CA142, CA143, Pb22 and etc.), which are preceded by the identification of the Brazilian state (RS, SC, SP, ES).
313P.R. Dorneles et al. / Science of the Total Environment 463–464 (2013) 309–318

DRC levels in male mammals generally increase with age (O'Shea
and Tanabe, 2003); however, a different accumulation pattern is
commonly observed for females. This difference is a consequence of
the elimination of pollutants through placental and lactational trans-
fer to offspring. In general, concentrations of organohalogen com-
pounds also increase through life in females, but this occurs up to
the start of the reproduction, after which the levels reach a plateau
or even decrease with age (Dorneles et al., 2010; O'Shea and
Tanabe, 2003; Thron et al., 2004). Considering that no significant
sex-related differences were observed, as well as that reproduction
had not started for immature females, groups constituted of all indi-
viduals and composed of males and immature females were also
used in statistical treatment scenarios.
For investigating possible correlations between DRC concentrations
or TEQ values and total length of franciscana dolphins from SP state,
tests were performed in three different scenarios, i.e., regarding only
males, males and immature females and all individuals. The pollutant-
related variables investigated were ΣPCDD, ΣPCDF, ΣTEQ-PCDD,
ΣTEQ-PCDF, Σnon-ortho PCB, Σortho PCB, ΣTEQ non-ortho PCB, ΣTEQ
ortho PCB, ΣDL-PCB, as well as ΣTEQ-DRCs. No significant correlation
was found for DRC concentrations; however, correlations were ob-
served for TEQ values. Significant negative correlations were found
between total length and three TEQ-related variables, i.e., ΣTEQ-DRCs,
ΣTEQ-PCDF and ΣTEQ non-ortho PCB, in three different scenarios: con-
sidering all individuals (males and females) (p = 0.025, Rs = −0.69;
p = 0.005, Rs = −0.81; and p = 0.019, Rs = −0.69; for ΣTEQ-
DRCs, ΣTEQ-PCDF and ΣTEQ non-ortho PCB, respectively); considering
the group formed by males and immature females (p = 0.01,
Rs = −0.83; for ΣTEQ-DRCs and ΣTEQ-PCDF; as well as p = 0.005,
Rs = −0.83, for ΣTEQ non-ortho PCB); and considering only males
(p = 0.014, Rs = −0.95; p = 0.015, Rs = −0.95; and p = 0.011,
Rs = −0.96; for ΣTEQ-DRCs, ΣTEQ-PCDF and ΣTEQ non-ortho PCB,
respectively).
As cited above, TEQ values result from the concentrations of the
different DRCs as well as from the ability of each compound to induce
the Ah-receptor mediated response. Therefore, the fact that correla-
tions with total length were found for TEQ values rather than for con-
centrations, associated with the fact that they constituted negative
correlations, suggests these findings result from chemically-induced
developmental disruption. As mentioned, inhibition of growth and
development is among the effects attributed to exposure to DRCs
(Sanderson and Van Den Berg, 1999). However, an increasing effi-
ciency of the detoxifying activity with the growth of the animal
may be a plausible explanation as well (Reijnders, 2003). The concen-
tration ratios between PCB-77 and PCB-169, as well as between
PCB-126 and PCB-169, have been used for evaluating enzyme induc-
tion and detoxifying activity (Corsolini et al., 2000; Storelli and
Marcotrigiano, 2003). Such use is based on the fact that PCB-169 is
the most stable coplanar congener (Corsolini et al., 2000). The
PCB77/PCB169 and the PCB126/PCB169 concentration ratios of the
franciscana dolphins from the present study averaged 29.1 and 7.4,
respectively. Therefore, possible correlations between these ratios
and total length values of franciscana dolphins from SP were investi-
gated in the same three scenarios mentioned above. A significant neg-
ative correlation was found between total length and PCB126/PCB169
concentration ratio for the group formed by males and immature fe-
males (p = 0.036, Rs = 0.7). The same correlation was not found
for the other groups (only males and all individuals) possibly due to
the small sample size of the male group, as well as due to the in-
creased number of females in the group composed by all franciscanas.
As cited, there are gender-related differences in the metabolizing
rates for xenobiotics (Hall, 2002). Nevertheless, this finding rein-
forces the hypothesis of increasing efficiency of the detoxifying activ-
ity with the growth of the animal.
The DRC levels verified in the present study constitute a matter of
concern for the conservation of the species. The hepatic PCDD and
PCDF concentrations of franciscana dolphins are high when compared
to cetaceans from different regions of the globe (Table 2). In addition,
three franciscana dolphins (two males from SP and SC, and 1 female
from SP) exhibited ΣTEQ concentrations within the range of thresh-
old levels for TEQs in livers of aquatic mammals that are capable of
eliciting physiological effects (160 to 1400 pg.g
−1
, l.w.) (Kannan et
al., 2000). Contamination with pollutants that have been shown to
be risk factors for cancer, immune deficiency and reproductive abnor-
malities (Schecter et al., 2006) constitutes a matter of concern for all
species. However, this is especially important for species facing pop-
ulation decreases due to other man-made causes, which enhances
the apprehension over this dolphin, as franciscana is the cetacean
species most impacted by fisheries in the southwestern Atlantic
(Secchi et al., 2003).
3.2. Guiana dolphins
Dioxin-like PCBs accounted for over 98.8% of the total TEQ for all
Guiana dolphins from RJ. For this cetacean population, mono-ortho
PCBs constituted the group of highest contribution, which varied
from 49 to 80% to the total TEQ. PCB-118 was the compound of
greatest importance among mono-ortho PCB congeners, with an av-
erage contribution of 34% to the total TEQ. This penta-PCB was also
the compound with the highest concentrations in the adipose tissue,
which varied from 2.81 to 11.5 μg/g (l.w.). PCDDs and PCDFs
accounted for less than 1.2% of the total TEQ in all Guiana dolphins.
The predominant PCDD/F congeners found in blubber of Guiana
dolphins were OctaCDD (mean ± SD; 19.4 ± 29.0 pg/g lipids) and
1,2,3,4,7,8-HexaCDF (6.7 ± 5.4 pg/g lipids). Regarding the TEQ
values (TEQ pg/g l.w.), 1,2,3,7,8-PentaCDD provided the highest con-
tribution among PCDD/Fs when the average of all Guiana dolphins
was considered. However, this penta-chlorinated dibenzo-p-dioxin
was not the PCDD/F of greatest contribution to total TEQ for six indi-
viduals (two males and the four females).
For investigating possible correlations between DRC concentra-
tions or TEQ values and total length of Guiana dolphins from RJ,
tests were performed in two different scenarios, i.e., regarding only
males and all individuals. The pollutant-related variables investigated
were ΣPCDD, ΣPCDF, ΣTEQ-PCDD, ΣTEQ-PCDF, Σnon-ortho PCB,
Σortho PCB, ΣTEQ non-ortho PCB, ΣTEQ ortho PCB, ΣDL-PCB, as well
as ΣTEQ DL-PCB. No significant correlations were found for DRC con-
centrations or TEQ values.
For investigating possible gender-related differences on DRC
bioaccumulation by Guiana dolphins from RJ, comparisons between
sexes were performed regarding ΣPCDD, ΣPCDF, ΣTEQ-PCDD,
ΣTEQ-PCDF, Σnon-ortho PCB, Σortho PCB, ΣTEQ non-ortho PCB,
ΣTEQ ortho PCB, ΣDL-PCB and ΣTEQ DL-PCB. Significantly higher
ΣTEQ-PCDD levels were verified in males than in females (p =
0.02). Higher persistent bioaccumulative toxicant concentrations in
male than in female mammals have been legitimately attributed to
the mother-to-calf transfer (O'Shea and Tanabe, 2003). However,
the fact that significant differences between sexes were found for
TEQ values rather than for concentrations suggests gender-related
peculiarities in DRC metabolizing capabilities play an important role
in this process. As mentioned above, males and females metabolize
DRCs at different rates (Hall, 2002).
3.3. Rough-toothed dolphins
On average, dioxin-like PCBs accounted for 93% of the total TEQ for
rough-toothed dolphins from RJ. For this species, the contribution of
mono-ortho PCBs to the total TEQ varied from 26 to 72%, and the
contribution of non-ortho PCBs was between 14 and 69%. PCB-118
and PCB-126 were the compounds of highest contribution among
mono-ortho and non-ortho coplanar PCB congeners, with contribu-
tion percentages to the total TEQ that varied from 13 to 43% as well
314 P.R. Dorneles et al. / Science of the Total Environment 463–464 (2013) 309–318

as from 2.3 to 59%. PCB-118 was the compound of highest concentra-
tion, which varied from 174 to 11,822 ng/g (l.w.). Concentrations of
PCB-126 varied from 189 to 1940 pg/g lipids. The summed contribu-
tion of PCDDs and PCDFs to the total TEQ in rough-toothed dolphins
varied from 1.6 to 13.9%. A different pattern was found in Guiana dol-
phins, for which the contribution of these compound classes was very
low. An extremely wide concentration range was observed for the
predominant PCDD/F congeners found in blubber of rough-toothed
dolphins, since OctaCDD and 1,2,3,4,7,8-HexaCDF levels varied from
6.5 to 713, as well as from 0.4 to 1026 pg/g lipids. This organochlorine
compound (OC) accumulation pattern, in which extremely high con-
centrations are found in only a few individuals, has been reported for
marine mammals (Hall, 2002). Actually, extremely wide concentra-
tion ranges could be found for organohalogen compounds in marine
mammals (Dietz et al., 2004; Hall, 2002; Sonne et al., 2008, 2004,
2006b, 2005, 2006c), even reaching a 70-fold variation (Sonne, 2010).
In fact, the ΣPCDD (747 pg/g lipids) and ΣPCDF (1313 pg/g lipids)
concentrations found in blubber of a female rough-toothed dolphin
are among the highest ever reported for marine mammals (Table 2).
Finding high concentrations of persistent accumulative toxicants in
this dolphin population is not surprising. Rough-toothed dolphins
from RJ feed on top-predator fish (Bisi et al., 2012; Melo et al.,
2010), thus they are more likely to take in high amounts of contami-
nants that have accumulated in the bodies of the prey.
3.4. False killer whales
Concentrations and percent contribution of PCB-118, PCB-126,
OctaCDD and 1,2,3,4,7,8-HexaCDF lie within the range mentioned
for rough-toothed dolphins, with the exception of the PCB-126 con-
centration, which was higher for the false killer whale (2179 pg/g
lipids). It is important to note that, as there was only one individual,
the pollutant levels and profiles may not be representative of the pop-
ulation. Despite this, the ΣPCDD and ΣPCDF concentrations found in
our study compare favorably with data for false killer whales from
North America (Jarman et al., 1996)(Table 2).
3.5. A serious PCB problem
Comparing DRC concentrations in franciscana dolphins from SP
with levels of the same compounds in Guiana dolphins from RJ is a
difficult task, since contaminant determination was performed in
liver and blubber, respectively. Evaluating the body distribution of li-
pophilic organohalogen compounds in cetaceans, Watanabe et al.
(1999) and Ramu et al. (2005) observed that, in general, the highest
levels are found in blubber, even considering lipid normalized con-
centrations. However, different body distribution patterns are ob-
served for compounds of the same chemical class (Kunisue et al.,
2008). An investigation that dealt with DRC concentrations in raccoon
dogs from Japan showed that the HxCB-157 ratio in liver/adipose tis-
sue presented values close to 1.0 (Kunisue et al., 2006). To the au-
thors' knowledge, this approach has not been applied to cetaceans
yet. Although species-specific differences in toxicokinetics of DRCs
may exist, the latter cited investigation turned HxCB-157 into the
most suitable compound for comparison between liver and adipose
tissue concentrations. Therefore, statistical comparison between the
HxCB-157 concentrations of franciscana dolphins and Guiana dol-
phins was performed, limiting it to male dolphins from SP and RJ, re-
spectively. The HxCB-157 concentrations found in Guiana dolphins
were significantly higher than those found in franciscana dolphins
(p = 0.003). Since both data sets presented normal distribution, the
mean values were used as a reference and it could be observed that
HxCB-157 levels of Guiana dolphins were supposedly 75 times
those of franciscana dolphins. Even considering two dolphin species,
this should be done with caution due to possible species-specific dif-
ferences in metabolism. However, it was the most appropriate among
the available tools for comparing the exposure to PCBs between
both odontocete species/populations. Nonetheless, this finding sug-
gests that Guiana dolphins from RJ are more exposed to PCBs than
franciscana dolphins from SP.
PCB levels found in Guiana dolphins in the present study are
among the highest ever reported for cetaceans. For instance, these
concentrations were in the range reported by Kuehl et al. (1991)
and Berggren et al. (1999) in blubber samples from bottlenose,
Table 2
Mean ΣPCDD and ΣPCDF concentrations (pg/g, l.w.) in blubber (B) and liver (L), with standard deviation (±SD), number of individuals of each species/area of sampling of cetaceans
from all over the world.
Species-common name Area ΣPCDD mean ± S.D. ΣPCDF mean ± S.D. Sex Tissue n Ref.
Finless porpoise Japan 60.9 6.0 N.S. B 1 Tanabe et al. (1989)
Killer whale Japan N.D. 345 MB 1Ono et al. (1987)
Killer whale Japan N.D. 436.8–551.7 F B 2 Ono et al. (1987)
Hector's dolphin N. Zealand 38.7 ± 8.1 41.6 ± 11.1 MB 4Buckland et al. (1990)
Hector's dolphin N. Zealand 12.5–37.8 13.6–36.8 F B 2 Buckland et al. (1990)
Killer whale N. America N.D.–26.4 25.3–42.9 N.S. B 2 Jarman et al. (1996)
Killer whale N. America N.D.–16.5 6.6–14.3 MB 2Jarman et al. (1996)
False killer whale N. America 4.5–41.9 2.3–117 MB 2Jarman et al. (1996)
Dall's porpoise N. America 17.4–60.2 50.0–125 MB 2Jarman et al. (1996)
Dall's porpoise N. America 43.9 35.7 F B 1 Jarman et al. (1996)
Harbor porpoise N. America 84.8 ± 112 13.1 ± 14.4 MB 4Jarman et al. (1996)
Harbor porpoise N. America 103–182 41.3–77.2 F B 2 Jarman et al. (1996)
Risso's dolphin Italy 48.9–424 59.2–331 ML 2Jimenez et al. (2000)
Risso's dolphin Italy 208 706 F L 1 Jimenez et al. (2000)
Bottlenose dolphin Italy 72.3 ± 46.0 75.3 ± 44.6 ML 4Jimenez et al. (2000)
Bottlenose dolphin Italy 19.2 22.9 F L 1 Jimenez et al. (2000)
Striped dolphin Italy 271 ± 148 92.3 ± 70.6 ML 3Jimenez et al. (2000)
Striped dolphin Italy 186–403 103–162 F L 2 Jimenez et al. (2000)
Pilot whale Italy 183 196 ML 1Jimenez et al. (2000)
Harbor porpoise North Sea 122 12.9 F B 1 Beck et al. (1990)
Franciscana dolphin S-SE Brazil 236 ± 323 113 ± 113 ML 13 PS
Franciscana dolphin SP, Brazil 348 ± 163 100 ± 53.8 F L 5 PS
Guiana dolphin RJ, Brazil 33.1 ± 39.0 19.0 ± 11.3 MB 7 PS
Guiana dolphin RJ, Brazil 14.2 ± 16.0 13.1 ± 13.5 F B 4 PS
Rough-toothed dolphin RJ, Brazil 27.2 82.0 MB 1 PS
Rough-toothed dolphin RJ, Brazil 9.99–747 7.42–1313 F B 2 PS
False killer whale RJ, Brazil 24.2 112 F B 1 PS
PS, present study.
315P.R. Dorneles et al. / Science of the Total Environment 463–464 (2013) 309–318

common and white-sided dolphins from the U.S. Atlantic Coast
(Kuehl et al., 1991), as well as from male harbor porpoises from the
Baltic Sea, the Kattegat–Skagerrak Seas and the West Coast of Norway
(Berggren et al., 1999). In the study performed by Kuehl et al. (1991),
the total PCB concentrations varied from 17.4 to 195 μg/g lipid, and in
the investigation conducted by Berggren et al. (1999), the levels var-
ied between 2.2 and 78 μg/g lipid (sum of CB52, CB101, CB118,
CB138, CB153 and CB180). The concentration range reported by
Kuehl et al. (1991) comprises the sum of one hundred PCB congeners,
much more than the 18 PCB congeners determined in the present
study. In Kuehl et al. (1991) and Berggren et al. (1999), cetaceans
were collected from 1978 to 1990. During the 1970s and 1980s, the
manufacturing of PCBs was terminated in most industrialized nations.
This implies further that the animals analyzed by Kuehl et al. (1991)
and Berggren et al. (1999) lived not only in a highly polluted area
but also in a period of elevated environmental contamination by
PCBs. The sample set analyzed by Kuehl et al. (1991) included
bottlenose dolphins obtained after an unusual mortality event. Al-
though the impact of these immunosuppressive compounds (Sonne
et al., 2006a) is not fully known, their role as causative agents in
that mass mortality was considered by the authors (Kuehl et al.,
1991), which emphasizes the toxicological importance of the concen-
trations found in Guiana dolphins from RJ. Among cetaceans species,
the highest concentrations of biomagnifying pollutants are usually
found in killer whales, due to their predation on organisms that occu-
py high trophic positions, such as sharks, seals and other cetaceans
(O'Shea and Tanabe, 2003). Therefore, the magnitude of the delphinid
exposure to PCBs in RJ waters can be exemplified with the fact that
concentrations found in these dolphins are comparable to the levels
verified in killer whales from Alaska (Ylitalo et al., 2001).
Interestingly, 10 out of the 11 Guiana dolphins analyzed in the
present study were found in Guanabara Bay, which may explain the
extremely high PCB concentrations found. Guanabara Bay is the
most anthropogenically disturbed area along the Brazilian coast
(Dorneles et al., 2008b). The estuary is bordered by more than
14,000 industries and four cities (Wasserman et al., 2006) with a pop-
ulation of more than 10 million people (Jablonski et al., 2006; Kjerfve
et al., 1997). Despite the anthropogenic pressure, the small Guiana
dolphin population exhibits habitat fidelity to Guanabara Bay, since
the same individuals are found year-round in this site (Azevedo et
al., 2004), where feeding-related activities predominate (Azevedo et
al., 2007). In addition to the expected contamination by PCBs from a
highly urbanized and industrialized area, some activities in Rio de
Janeiro amplify the apprehension. Eleven thousand liters of Aroclor
were stolen from an abandoned building in Rio de Janeiro City in Sep-
tember 2005 (ALERJ, 2005). This raises suspicion on the existence of
an illegal market and usage of PCBs in Brazil, which is a serious reason
for concern in terms of environmental pollution.
The Brazilian economic growth and the consequent use and degra-
dation of the coastal zone are a threat to the conservation of coastal
cetaceans. A large set of construction works is planned for the estab-
lishment of harbors, shipyards and industries on Brazilian coastal
bays and their drainage basins. Many of these estuaries have been
suffering from a large series of different types of pressure, as well as
they have been going through broad dredging and even submarine
demolishing. Some Brazilian bays seem to be on the same degrada-
tion path once followed by Guanabara Bay, which harbors a residual
Guiana dolphin population assessed to be around 40 individuals
only (Azevedo et al., submitted for publication). Environmental con-
tamination by DRCs constitutes one additional obstacle for these
marine mammals to reach their natural longevity and reproduce at
a physiologically normal rate. The concerns generated by the DRC
contamination are amplified when it is taken into account that
these cetaceans are exposed to other persistent bioaccumulative tox-
icants (PBTs) that have been shown to be endocrine disrupters, tumor
promoters and immunosuppressors as well (Schecter et al., 2006;
Simonyte et al., 2006; Sonne et al., 2006a; Whalen et al., 1999). The
high exposure of cetaceans from Brazil to cadmium (Dorneles et al.,
2007a, 2007b), mercury (Lailson-Brito et al., 2012a), organochlorine
pesticides (Lailson-Brito et al., 2011, 2010, 2012b), as well as to
organotin (Dorneles et al., 2008b), perfluoroalkyl (Dorneles et al.,
2008a) and organobrominated (Alonso et al., 2012; Dorneles et al.,
2010) compounds has already been reported. Therefore, the possibil-
ity of synergistic effects among these PBTs should not be ruled out.
The data generated by the present study raises apprehension on
the possibility of human health problem due to consumption of fish
from Brazilian estuaries. This apprehension is augmented when it is
considered that many fish species are consumed by humans in Rio
de Janeiro at the size preyed on by Guiana dolphins from Guanabara
Bay (Melo et al., submitted for publication). Therefore, risk assess-
ment studies on human exposure to PCBs through consumption of
fish from Guanabara Bay are of fundamental importance.
Conflict of interest
The authors state that there is no conflict of interest.
Acknowledgments
This work was supported by the Ministry of Education of Brazil—
CAPES (“Ciências do Mar”—Proc. 23038.051661/2009-18, as well
as fellowship to M.B. Alonso, “Sandwich Programme”—PDEE), by the
Brazilian Research Council—CNPq and by Rio de Janeiro State Govern-
ment Research Agency—FAPERJ (“Pensa Rio”Program). This study was
also supported by a scientific cooperation established between CNPq
and F.R.S.—FNRS (Proc. 490471/2010-2 CNPq). The Yaqu Pacha Founda-
tion is acknowledged for its financial support to research projects that
allowed cetacean carcass recovery. Dr. Lailson-Brito is a researcher of
“Prociência”Program—FAPERJ/UERJ. Dr. Azevedo has a research grant
from CNPq (grant No. 304826/2008-1). Since Dr. Torres is partially
funded by Grant 1D43TW00640 from the Fogarty International Center
of the National Institute of Health, this institution is also acknowledged.
Dr. Torres is also a Research Fellow of CNPq and FAPERJ. Krishna Das is a
F.R.S.—FNRS research associate.
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