International Biodeterioration & Biodegradation xxx (2016) 1e8

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International Biodeterioration & Biodegradation

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Assessment of the bioaccumulation and biodegradation of butyltin

compounds by Thalamita crenata in Kaohsiung Harbor, Taiwan
Chiu-Wen Chen, Chih-Feng Chen, Yun-Ru Ju, Cheng-Di Dong*
Department of Marine Environmental Engineering, National Kaohsiung Marine University, Kaohsiung 81157, Taiwan, ROC

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

Article history: Butyltin compounds (BTs) are considered as endocrine disruptors or environmental hormones. It has
Received 14 January 2016 been applied as an antifouling paint in marine vessel, and extensively in commercial, industrial and
Received in revised form agricultural applications. In order to investigate the decomposition and accumulation of BTs in benthic
24 February 2016
crustacean, concentrations of BTs including TBT (tributyltin) and its metabolic products, i.e. MBT
Accepted 24 February 2016
(monobutyltin) and DBT (dibutyltin), accumulated in Thalamita crenata in Kaohsiung Harbor, Taiwan
Available online xxx
were analyzed. The analysis results indicate different concentrations of BTs contained in hepatopanceas
and muscle for the crustacean trapped at different locations. Additionally, concentration and composi-
Keywords:
Butyltin
tion of BTs found in the crustacean hepatopanceas and muscle also show differences. Hepatopancreas has
Butyltin degradation index higher concentrations of BTs than muscle; the BTs contained in hepatopancreas are mostly metabolic
Biota-sediment accumulation factors products of BTs, whereas the muscle contain higher concentrations of TBT. By the estimates of BDI
Tributyltin (TBT) (butyltin degradation index) and BSAF (biota-sediment accumulation factors), T. crenata shows slow
Thalamita crenata degradation ability of TBT and relatively strong TBT accumulation. The sources of BTs accumulated in
T. crenata include harbor sediment and the food that contains BTs.
© 2016 Elsevier Ltd. All rights reserved.

1. Introduction physiological system in addition to being toxic, extremely stable,

Butyltin compounds (BTs) have been used extensively as a
thermal stabilizer in the manufacturing of PVC (polyvinyl chloride)
(Hoch, 2001). Since 1960, TBT (tributyltin) has been applied as an
antifouling agent in marine vessel paints, and also extensively in
other commercial, industrial and agricultural applications such as
industrial water supply system, agricultural chemistry, glass pro-
cessing, material production (stone, leather and paper), textile
product and animal husbandry (Fent, 1996; Hoch, 2001). The
extensive and multiple applications of BT chemical make the
organic metallic compounds ubiquitous in the environment,
especially in harbors with frequent vessel traffic (Sun et al., 1996;
Mzoughi et al., 2005; Chen et al., 2010). BT chemicals are hydro-
phobic; once entering into water bodies, they are easily adsorbed
on the surface of suspended particles and settle to the bottom
causing relatively high BT concentration in sediment (Hoch, 2001).
BT compounds are endocrine disruptors or environmental hor-
mones that may imitate or disrupt biological endocrine
* Corresponding author.
E-mail address: cddong@mail.nkmu.edu.tw (C.-D. Dong).
and not readily biodegradable in natural environment with strong
characteristics of bio-concentration (Gibbs et al., 1988; Hoch, 2001).
A TBT contaminated water body is unfavorable to the growth of
aquatic creatures even TBT concentration is at the ng/L level (Zhou
et al., 2003); the symptom include imposes of gastropoda,
deformed growth of ostracean, and premature death of shellfish
(Tanabe et al., 2000; Zhou et al., 2003). Hence, the International
Maritime Organization (IMO) has suggested ban the use of TBT in
antifouling paints (Champ, 2000; Michaud and Pelletier, 2006;
Hoch, 2001). Many nations have passed legislations to ban or
regulate TBT applications. In Taiwan, the Environmental Protection
Administration, based on the speculations suggested by IMO, is-
sued a ban on the use of TBT-containing paints for vessels shorter
than 25 m in 2003, and an overall ban for all vessels in 2008.
Benthic crustaceans, which contact bottom sediment, have
limited mobility; they take smaller benthic organisms, and
deposited biological fragments or filter out water-borne organic
matters as foods. Hence, the bioaccumulation of environmental
pollutants in the food chain can be easily reflected by crustacean,
and hence, the organism is often used as an important water quality
biological index (Chou et al., 2000; Mac-Farlane et al., 2000). Tha-
lamita crenata, which is a Thalamite in Portunidac, is the largest of

http://dx.doi.org/10.1016/j.ibiod.2016.02.018
0964-8305/© 2016 Elsevier Ltd. All rights reserved.

Please cite this article in press as: Chen, C.-W., et al., Assessment of the bioaccumulation and biodegradation of butyltin compounds by
Thalamita crenata in Kaohsiung Harbor, Taiwan, International Biodeterioration & Biodegradation (2016), http://dx.doi.org/10.1016/
j.ibiod.2016.02.018
2 C.-W. Chen et al. / International Biodeterioration & Biodegradation xxx (2016) 1e8
the 17 crustacean species found in Taiwan. Its major habitat is in the
tidal flat between reef terrain and bedload with water depth be-
tween 1 and 10 m. Sometimes T. crenata is also found in river up to
tidal reach; it is one of the edible crabs found in the river mouth in
Taiwan. T. crenata has long been considered a superior biological
index in Taiwan because it is the most dominant crustacean species
in Kaohsiung Harbor, and is also widely distributed in the river
mouth (Chen et al., 2005; Dong et al., 2015).
In an aquatic system, TBT may be degraded chemically and
biologically or through UV irradiation. The degradation mechanism
is dealkylation that gradually removes the stannum ions from
organic group of toxic TBT to form DBT and MBT (Hoch, 2001) (see
supplementary content Fig. S1). Hence, the butyltin degradation
index (BDI), which is the ratio of TBT to the metabolic produces
(DBT and MBT), can be used to reveal the fate of TBT in the various
natural and biological media (including water, sediment, and or-
ganism), and how it is degraded (Díez et al., 2002; Burton et al.,
2005; Lee et al., 2005; Diaz et al., 2007; Díez and Bayona, 2009).
BDI is calculated as BDI ¼ [(DBT þ MBT)/TBT]; in which MBT, DBT
and TBT are the concentration of BT species contained in the
specimen. A BDI value greater than 1 indicates that crustacean
tends to accumulate TBT in its body, whereas a BDI less than 1
shows that TBT is well degraded in the body of crustacean in
question (Jadhav et al., 2011).
The bottom sediment is a major sources of pollutants found in
benthic creatures (Hoch, 2001). Hence, the ratio of BTs in bottom
sediment and in benthic crustaceans, also known as BSAF (biota-
sediment accumulation factor), will help to understand the ca-
pacity of living organism to accumulate BTs and resulting accu-
mulation effect (Viglino et al., 2004, 2005; Michaud and Pelletier,
2006). BSAF is calculated as BSAF ¼ [BTtissue]/[Normalized BTsedi-
ment], in which BTtissue is the BTs concentration in living organisms;
and normalized BTsediment is the normalized BT concentration in
bottom sediment for 1% organic matter (Viglino et al., 2004). A BSAF
greater than 1 indicates that the living organism has greater BT
concentration than the surrounding bottom sediment, and that the
living organism possess accumulative effect (Viglino et al., 2004).
Recently, Leung et al. (2006) collected gastropods from coastline
of Hong Kong to analyze the tissue burdens of TBT and demon-
strated that imposex indices were positively correlated with the
TBT in the gastropods. Organotins concentrations in edible marine
organisms collected from the west coast of India were assessed, and
found some organism had the relatively higher organotins burdens
(Bhosle et al., 2004). Harino et al. (2007) surveyed organotin
compounds in coastal areas of Japan, and results showed that
sediment and organisms of mussels had high TBT level. It is sug-
gested that the monitoring of organotin compounds is needed to
continue and evaluate the ecological risk for aquatic organisms
(Harino et al., 2007). The concentration of BT chemicals contained
in the tissues of T. crenata collected from Kaohsiung Harbor channel
was analyzed in order to understand the degree of pollution in
harbor, and to establish a background database before a complete
ban of TBT containing paints in Kaohsiung Harbor. Additionally, the
results will lead to the estimation of BDI and BSAF for investigating
the decomposition and accumulation of BTs in benthic crustacean.
2. Materials and methods
2.1. Region of study and sampling locations
Kaohsiung Harbor is located at the southwest Taiwan seashore;
it controls the strategic point where Taiwan Strait meets Basu Strait.
It is the largest international harbor in Taiwan, and an important
port of transshipment between Indian Ocean and Northeast Asian
Navigation Center with annual throughput capacity ranking 13th
Please cite this article in press as: Chen, C.-W., et al., Assessment of the bioaccumulation and biodegradation of butyltin compounds by
Thalamita crenata in Kaohsiung Harbor, Taiwan, International Biodeterioration & Biodegradation (2016), http://dx.doi.org/10.1016/
j.ibiod.2016.02.018
internationally. The harbor is a long and narrow bay with 12 Km in
length, 1 to 1.5 Km in width, 11e16 m in channel depth and
16.24 Km2 in water area (KHB, 2011). The source of BT pollution
includes vessel operations in harbor, as well as industrial, agricul-
tural and domestic wastewater discharge carried from Love River
and Canon River from the north, Jan-Gen River discharges into the
middle section, and Salt River from the south. Vessels in harbor,
industrial dock, and shipyard may release BTs into harbor water.
Hence, sampling stations were installed at strategic locations for
collecting representative crab specimens. As shown in Fig. 1, the 9
sampling stations are located at: 1. Navy Shipyard, 2. Chyi-Jihn
Fishing Port, 3. Private Shipyard, 4. Jen-Gen River mouth, 5. Jen-
Gen Fishing Port, 6. Salt River mouth, 7. China Shipbuilding (CSB)
Shipyard, 8. Canon River mouth and 9. Love River mouth.
2.2. Sample collection
The crustacean samples were collected in May and September,
2006 by placing wire mesh cages (60 cm long, 45 cm wide and
20 cm high with 2.0 cm mesh) at the sampling points from fishing
boats. The cages were then retreated the next day to collect crus-
taceans. All specimens were temporarily stored in covered light-
proof ice chest and then shipped back to the laboratory to be pre-
served at 20  C before being processed for analyses.
2.3. Reagents
All chemical reagents are of analytical grades. Monobutyltin
trichloride (MBT, 95%) and tetrapentyltin (TePeT, 97%) were pur-
chased from Aldrich Chemical (USA); dibutyltin dichloride (DBT,
96%) and tributyltin chloride (TBT, 96%) standards were obtained
from TCI (Japan). Butyltin stock solution in 1000 mg/L as Sn was
prepared daily with methanol; the prepared stock solution was
stored in 4  C dark container. Sodium tetraethylborate (NaBEt4,
98%) was purchased from Strem Chemicals (USA); the working
solution was prepared daily by dissolving 0.10 g NaBEt4 in 1 mL de-
ionized water and stored in 4  C dark container. Tropolone (98%)
was purchased from Lancaster (UK); nehexane (95%), methanol
(99.8%) and Hydrochloric acid (HCl, 37.8%) were obtained from
Tedia (USA).
2.4. Sample preparation and analyses
Eighty-eight crab specimens were collected with 39 (40%)
specimen belonging to the most predominant T. crenata species. All
Fig. 1. Region of study and sampling locations.
 C.-W. Chen et al. / International Biodeterioration & Biodegradation xxx (2016) 1e8 3

captured creatures were weighed, measured for carapace width;
male and female crabs were identified and separated. Because BT
analyses must be done with sufficient samples, e.g. more than 1 g
dw for muscle and 0.1 g dw for hepatopancreas, only specimens
with carapace greater than 55 mm were selected for analyses
(Table 1). Each specimen was dissected and separated into two
types of tissue samples, i.e. hepatopancreas and muscle; the tissue
was freeze dried for 72 h, and then grounded into powder form. It
was then stored in brown glass bottle sealed with Teflon lined
screw cap to be preserved in a 20  C freezer until analyzed. A total
of 62 samples (31 muscle and 31 hepatopancreas samples) were
obtained for analyses.
The procedures suggested by Dong et al. (2004) were slightly
modified for extracting BTs from rock crab tissue. First, an adequate
amount of the freeze dried and homogenized tissue (more than 1 g
dw for muscle and 0.1 g dw of hepatopancreas) was weighed out
with an accurate balance (accuracy of ±0.0001 g), and was placed in
a 40 Ml brown glass container. Subsequently, 3 mL of methanol,
0.1 mL of 37.8% hydrochloric acid (HCl), and 0.2 mL of 0.5% NaBEt4
derivative were added followed by 3 mL of 0.02% Tropolone/n-
hexane. The mixture was submerged in a water bath and agitated
ultrasonically for 10 min. It was then removed from the water batch
and allowed to settle under a quiescent condition for 10 min. The
organic supernatant was collected with a glass pipette; it was
further extracted 3 times by adding 3 mL of the same solvent fol-
lowed by agitation. All of the organic phase solutions were
collected in the same glass tubing; 0.1 mL of the internal standard
10 mg/L TePeT was added. The mixture was passed through a florisil
purification tubing to be purified. Its final volume was reduced to
0.5 mL by slowly blowing nitrogen gas into the mixture. Quanti-
tative and qualitative analyses were carried out by injecting 0.3 mL
of the purified extract into a gas chromatography equipped with
flame photo metric detector (GC/FPD).
2.5. Quality assurance and quality control (QAQC)
Laboratory results were analyzed using blank samples, cali-
brated standards, and a five-point calibration curve covering the
concentration range of 20e320 pg as Sn. All linear correlation co-
efficients (r) were greater than 0.995; results of all blank analyses
were smaller than the lower detectable limit; recoveries of stan-
dards were between 85 and 108%. The detection limit of analytical
procedure was estimated from 3x standard deviation from analyses
of 7 multiple duplicates (n ¼ 7) containing butyltins of 20 pg as Sn,
and the amount of sample extracted. The lower detectable limits,
after repeated tests, were 20 pg as Sn with standard deviations of
0.56 ng/g for MBT, 0.76 ng/g for DBT and 0.50 ng/g for TBT. The
analytical method used in this research was further tested by using
the same procedures to analyze the certified reference No. 11, i.e.
“Fish Tissue”, provided by Japan National Institute for Environ-
mental Studies. TBT result was 1.4 ± 0.1 mg/g (n ¼ 3) with 106 ± 11%
for the TBT reference with given a TBT concentration of 1.3 ± 0.1 mg/
g as chloride. The validity and accuracy of our laboratory method
for analyzing the BT species were thus confirmed.
2.6. Statistical analyses
Statistical analyses of laboratory results were carried out using
EXCEL 2013 (Microsoft) and SPSS 12.0 (SPSS Inc., USA). ANOVA was
used to analyze the concentration difference of BTs in specimens
collected at various sampling sites and in different tissues of crab.
Pearson correlation coefficient was calculated for analyzing the
degree of correlation among MBT, DBT, TBT, SBTs, BDI, and BSAF in
different tissues.
3. Results and discussion
3.1. The concentrations of BTs in T. crenata in Kaohsiung Harbor
Fig. 2 shows the distribution of MBT, DBT, TBT and SBTs (sum of
TBT, DBT and MBT concentration) in hepatopancreas and muscle
tissues of T. crenata specimens collected at various monitoring
stations in Kaohsiung Harbor. The average SBTs concentration was
between 249 ± 106 and 4694 ± 2220 ng Sn/g dw in hepatopancreas
and between 43.5 ± 40.8 and 572.5 ± 319.1 ng Sn/g dw in muscle
(Table S1). For specimens collected at various stations, SBTs con-
centrations was different (p < 0.05) confirming the difference in the
spatial distribution of BT chemicals in harbor (Chen et al., 2010).
Additionally, the hepatopancreas accumulates more BT chemicals
than muscle tissue, which was commonly observed in wildlife such
as Lizardfish Trachinocephalus myops in Taiwan (Dong et al., 2004),
Lateolabrax japonicus in Japan (Harino et al., 2000), Thunnus thyn-
nus thynnus in Italy (Kannan et al., 1996) and Canadian snow crab
(Chionoecetes opilio) (Viglino et al., 2005).
As shown in Table S1, the hepatopancreas of T. crenata contained
127 ± 54e1761 ± 919 ng Sn/g dw of TBT, 74 ± 22e1441 ± 583 ng Sn/
g dw of DBT, and 33 ± 29e1492 ± 717 ng Sn/g dw of MBT; the
muscle contained 19.0 ± 21.9e410.0 ± 215.0 ng Sn/g dw of TBT,
18.7 ± 3.0e271.2 ± 227.2 ng Sn/g dw of DBT, and
2.4 ± 0.3e75.2 ± 29.5 ng Sn/g dw of MBT. Hence, the order of
magnitude for the various BTs contained in hepatopancreas was
DBT (35.9 ± 15.4%) > MBT (32.1 ± 18.8%) S TBT (32.0 ± 18.5%)
(p ¼ 0.65), whereas in muscle, it was TBT (46.9 ± 14.4%) > DBT
(40.0 ± 10.4%) > MBT (13.0 ± 8.0%) (p ¼ 0.02). Similar results on the
distribution of BT chemicals had also been reported by Viglino et al.
(2005). Because the CYP450 (cytochrome P-450) contained in crab
hepatopancreas is capable of metabolizing TBT into DBT, MBT and
inorganic tin (Kannan et al., 1996), hepatopancreas contains higher

Table 1
Location and water depth of the sampling points in Kaohsiung Harbor, and size and weight of the crab specimens collected.

Site Latitude Longitude Water depth (m) n Carapace width Weight of crabs (g
(mm) wt.)

Mean SD Mean SD
 0 00  0 00
1 Navy Shipyard 22 36 57 120 16 14 8.8 3 60.0 3.0 40.1 4.0
2 Chyi-Jihn Fishing Port 22 350 5600 120 160 7700 5.6 3 73.3 2.9 94.1 2.0
3 Private Shipyard 22 350 0000 120 170 2000 7.2 3 69.3 3.1 77.4 2.5
4 Jen-Gen River mouth 22 350 0900 120 170 4400 7.6 2 56.0 1.4 36.6 0.6
5 Jen-Gen Fishing Port 22 340 2100 120 180 1600 10.2 5 69.2 1.9 74.0 2.4
6 Salt River mouth 22 320 5500 120 200 0800 9.8 2 74.0 1.4 69.6 0.9
7 CSB Shipyard 22 320 3500 120 200 3300 9.4 2 69.0 1.4 66.1 1.5
8 Canon River mouth 22 350 9200 120 170 4800 4.8 4 68.8 2.6 56.0 2.2
9 Love River mouth 22 370 1600 120 160 9300 4.2 7 64.1 2.0 62.7 1.9

Please cite this article in press as: Chen, C.-W., et al., Assessment of the bioaccumulation and biodegradation of butyltin compounds by
Thalamita crenata in Kaohsiung Harbor, Taiwan, International Biodeterioration & Biodegradation (2016), http://dx.doi.org/10.1016/
j.ibiod.2016.02.018
4 C.-W. Chen et al. / International Biodeterioration & Biodegradation xxx (2016) 1e8

Fig. 2. Butyltin compounds contained in the hepatopancreas (A) and muscle (B) of T. crenata for the various sampling locations in Kaohsiung Harbor.

metabolic products (e.g. DBT and MBT) than TBT. Conversely, the
crab muscle is not capable of metabolizing TBT so that it contains
higher concentrations of TBT then DBT and MBT because.
Until now, no study was found for investigating the BTs bio-
accumulation of T. ceranta in other regions of coast. This study
compared the BTs concentrations of benthic species in other re-
gions with those of T. ceranta in Kaohsiung Harbor. Table 2 indicates
BTs concentration in various species at various locations in the
world. Kaohsiung Harbor T. cerana has lower BTs concentration in
muscle tissue than some species in other regions, but higher BTs
concentration than some other species such as gastropods in Spain
coast (Gomez-Ariza et al., 2006), clam in Indian west coast (Bhosle
et al., 2004), clam in Vietnam (Midorikawa et al., 2004; Nhan et al.,
2005) and gastropods in Hong Kong (Leung et al., 2006). Kaohsiung
Harbor T. ceranta accumulates more BTs in hepatopancreas than
most species in other regions including starfish in Korea (Shim
et al., 2005), mussel in Canada (Chau et al., 1997), gastropod spe-
cies living in the strait between Denmark and Sweden (Strand et al.,
2003), and mysid shrimp in Scheldt estuary in the Netherlands
(Verslyckea et al., 2005). The differences of BTs bioaccumulations of
organisms between other regions and this study probably due to
various exposure pathways, degradation pathways, physiological
processes, and environmental factors. Generally, Kaohsiung Harbor
T. ceranta accumulates relatively high concentrations of BTs than
those in other regions organism. The phenomenon might be caused
by a large number of ships entering and leaving Kaohsiung Harbor.

Please cite this article in press as: Chen, C.-W., et al., Assessment of the bioaccumulation and biodegradation of butyltin compounds by
Thalamita crenata in Kaohsiung Harbor, Taiwan, International Biodeterioration & Biodegradation (2016), http://dx.doi.org/10.1016/
j.ibiod.2016.02.018
 C.-W. Chen et al. / International Biodeterioration & Biodegradation xxx (2016) 1e8 5

Table 2
Distributions of BTs in marine organisms found in various regions of the world (ng Sn/g dw.).

Location Organism MBT DBT TBT References

American
Coast, Canada Mussel NDae708 NDe1062 20e1198 Chau et al. (1997)
Coos Bay estuary, USA Shellfish <3.1e5.3 <3.2e7 <2.4e181.6 Elgethun et al. (2000)
European
Strait between Denmark and Sweden Bivalve <2.5e15c 11e267c 24e1316c Strand et al. (2003)
North-western Sicilian coasts, Italy Gastropod (H. Trunculus) NDe167 NDe316 NDe91 Chiavarini et al. (2003)
West coast, Portugal Mussel <10e605 <10e345 11e789 Barroso et al. (2004)
Coast, Portugal Mussel <7.9e41 <2.5e18 <5.7e489 Díez et al. (2005)
Scheldt estuary, The Netherlands Mysid shrimp (Neomysis integer) 8.00e12.0c 21.0e25.0c 927e1209c Verslyckea et al. (2005)
South west coast, Spain Gastropod (B. brandaris) 54e63 67e83 35e45 Gomez-Ariza et al. (2006)
Aegean Sea, Greece Bivalves NDe151 NDe366 NDe109 Chandrinou et al. (2006)
North west coast, Spain Oyster 0.4e12.9 7.6e441 74e193 Üveges et al. (2007)
Mussel 52.8e96.1 20.2e25.7 52.8e96 Üveges et al. (2007)
Asian
Coast, Vietnam Clam (Meretrix spp.) 0.86e29 0.7e6.1 1.4e47 Midorikawa et al. (2004)
Clam (Meretrix meretrix) 2.8e18 4.4e27 3.8e15 Nhan et al. (2005)
Coastal, Malaysian Fish 2.3e7.4c <1.3e13c 2.4e190c Sudaryanto et al. (2004)
West coast, India Oyster (S. cucculata) NAb NDe88 ± 8 124e732 Bhosle et al. (2004)
Fish (Mugil sp.) NA 134 ± 3 96 ± 3 Bhosle et al. (2004)
Clam (P. malabarica) NA 215 ± 46 64 ± 7 Bhosle et al. (2004)
Mussel (Perna viridis) NA 358 464 Bhosle et al. (2004)
Crab (Scylla serrata) 25.30 ± 1.25 8.26 ± 1.23 79.10 ± 5.68 Jadhav et al. (2011)
Crab (N. suguinolentus) 16.12 ± 0.56 10.03 ± 0.54 85.23 ± 2.18 Jadhav et al. (2011)
Crab (N. pelagicu) 18.20 ± 0.51 8.90 ± 0.52 15.18 ± 1.13 Jadhav et al. (2011)
Coastal, Korea Starfish (A. pectinifera) 51e2860 8e139 NDe323 Shim et al. (2005)
Starfish (A. amurensis) 103e1520 49e750 21e797 Shim et al. (2005)
Bivalves NDe461 23e699 16e1610 Shim et al. (2005)
Coastline of Hong Kong, China Gastropod (T. clavigera) NDe336 NDe197 NDe18 Leung et al. (2006)
Gastropod (T. Luteostoma) NDe51 NDe85 3.8e170 Leung et al. (2006)
Sanricu coast, Japan Mussel 4e32 3e92 3e287 Harino et al. (2007)
Kaohsiung Harbor, Taiwan Crab muscle (T. crenata) 11.8e410.0 9.3e271.2 1.2e75.2 This study
Crab hepatopancreas (T. crenata) 127e1761 74e1441 33e1492 This study
a
ND: below detection limit.
b
NA: no data available.
c
ng organotin instead of Sn.

BTs probably were released into the waterbody of harbor through
the antifouling paints of ships (Mzoughi et al., 2005).
3.2. Comparison of BTs concentration in water, sediment and
T. crenata
The data of BTs concentration of water and sediment were come
from Chen et al. (2010). The SBTs concentrations ranged
12.1e270 ng Sn/L, 1.7e150.2 ng Sn/g dw, 249e4694 ng Sn/g dw, and
43.5e572.5 ng Sn/g dw in water, sediment, T. crenata hepatopan-
creas and muscle, respectively (Table S1). Fig. 3 shows the SBTs
concentration in bottom sediment and the distribution of SBTs in
hepatopancreas (Fig. 3A) and muscle tissues (Fig. 3B) of T. ceranta in
Kaohsiung Harbor at various stations. The SBTs concentration in
bottom sediment and crab hepatopancreas showed similar distri-
bution profiles (r ¼ 0.94, p < 0.001). However, similar relationship
was not observed between the distribution of SBTs in bottom
sediment and crab muscle (r ¼ 0.44, p > 0.05). In addition, the
distribution of SBTs in water, crab muscle (r ¼ 0.01, p > 0.05), and
hepatopancreas (r ¼ 0.02, p > 0.05) were also not observed sig-
nificant correlation. These observations indicated that the BTs
concentration in hepatopancreas of T. ceranta reflected the BTs
pollution in harbor sediment than those in crab muscle. Moreover,
it is found that the BTs accumulation in tissues of T. ceranta was
unable to reflect the pollution in water, which might be caused by
the primary source of BTs from sediments for T. ceranta.
3.3. Degradation of BTs in Kaohsiung Harbor T. crenata
The crustacean species collected at various stations have the BDI
value between 0.6 and 6.3 for hepatopancreas and 0.4e2.9 for
muscle tissue (Table S1). Except hepatopancreas of the specimen
collected from Station #1 (Navy Shipyard), and muscle tissue of the
specimens collected from Station # 2 (Chyi-Jihn Fishing Port) and
Station # 8 (China Shipbuilding Shipyard), the specimens collected
from all other stations have a BDI value greater than 1 (Fig. 4A). This
indicates that harbor rock crab is capable of degrading toxic TBT
into less toxic products. Additionally, the crustacean species at all
stations except Station #1 have 1 to 6 times higher BDI in hepa-
topancreas than muscle. These results conform to the observations
published in the literature that crab species are capable of
decomposing TBT. For example, Rice et al. (1989) fed blue crabs
with TBT contaminated food to demonstrate that blue crabs could
effectively decompose TBT so that their tolerance to TBT was
elevated. Oberdo € rster et al. (1998) conducted similar studies using
blue crabs and observed that when blue crabs were exposed to high
concentration of TBT, their hepatopancreas produced higher
CYP450 to assist in metabolizing and eliminating the ingested TBT.
Effective decomposition of TBT by hepatopancreas CYP450 was also
observed by Rouleau et al. (1999) with snow crab.
Table 3A shows the Pearson correlation coefficient between the
BTs contained in hepatopancreas and muscle of T. crenata. Corre-
lations were observed among MBT, DBT, TBT, and SBTs contained in
crab hepatopancreas (p < 0.01) with correlation coefficients be-
tween 0.62 and 0.90, and also in muscle (p < 0.05) with correlation
coefficients between 0.37 and 0.88. These observations revealed
that MBT and DBT in crab hepatopancreas and muscle may be
originated from TBT metabolism. Results of numerous previous
studies also indicated that DBT and MBT contained in living or-
ganisms came mainly from the biological degradation of TBT inside

Please cite this article in press as: Chen, C.-W., et al., Assessment of the bioaccumulation and biodegradation of butyltin compounds by
Thalamita crenata in Kaohsiung Harbor, Taiwan, International Biodeterioration & Biodegradation (2016), http://dx.doi.org/10.1016/
j.ibiod.2016.02.018
6 C.-W. Chen et al. / International Biodeterioration & Biodegradation xxx (2016) 1e8
Fig. 3. Comparisons of SBTs in bottom sediment and the SBTs contained in the
hepatopancreas (A) and muscle (B) of T. crenata for the various sampling locations in
Kaohsiung Harbor.
the body of living organisms (Quintela et al., 2000; Shim et al.,
2000; ten Hallers-Tjabbes et al., 2003). Both the TBT contained in
muscle, and the MBT, DBT and TBT contained in hepatopancreas
had correlation relationships with SBTs (p < 0.01). However, the
MBT, DBT, and SBTs contained in muscle did not have correlation
with the MBT, DBT, TBT, and SBTs contained in hepatopancreas
(p > 0.05). This shows that after TBT is absorbed by the living or-
ganisms, it is degraded at different rates in hepatopancreas and also
in muscle so that the DBT and MBT contained in muscle and
hepatopancreas are not correlated.
3.4. Biota-sediment accumulation factor (BSAF)
The average TBT and SBTs concentration at the surface sediment
reported by Chen et al. (2010) were used in this research to
calculate the BSAF values because both studies shared the same
sampling times and locations. As shown in Table S1, for T. crenata,
BSAF values of the TBT (BSAFTBT) was between 100 and 834 for
hepatopancreas and between 4 and 326 for muscle tissues. BSAF
value of the SBTs (BSAFSBTs) contained in hepatopancreas and
muscle tissue was between 21 and 219 and between 3 and 91,
respectively. The maximum BSAF value was found in specimens
collected from Station #8 (Canon River mouth) and #9 (Love River
mouth). Specimens collected from all stations had a BSAF value
greater than 1 (Fig. 4B and C) indicating that the rock crab living in
Kaohsiung Harbor was capable of accumulate BTs primarily in
hepatopancreas. Furthermore, hepatopancreas and muscle had
Please cite this article in press as: Chen, C.-W., et al., Assessment of the bioaccumulation and biodegradation of butyltin compounds by
Thalamita crenata in Kaohsiung Harbor, Taiwan, International Biodeterioration & Biodegradation (2016), http://dx.doi.org/10.1016/
j.ibiod.2016.02.018
Fig. 4. BDI and BSAF in T. crenata for the various sampling locations in Kaohsiung
Harbor.
higher BSAFSBTs value than BSAFTBT value revealing that the
metabolic products, i.e. DBT and MBT, remained to be accumulated
in tissues of the crab.
Table 3C shows that both BSAFTBT and BSAFBTs for hepatopan-
creas and muscle tissue of T. crenata species do not have correla-
tions with SBTs and TBT concentrations of the normalized 1%
organic matter for bottom sediment (p > 0.05). This implies that in
addition to the bottom sediment, other sources also contribute to
the BTs accumulated in crab. Viglino et al. (2005) based on their
research suggested that TBT in crustaceans was caused by the foods
ingested and also by direct exposure to the polluted environment.
Benthic crusteceans, which feed on smaller benthic crustaceans,
enchinodermata, polychaeta, and smaller benthic fish (Bre ^thes
et al., 1994; Viglino et al., 2005), accumulate different quantities
of BTs in their bodies. Hence, the difference in BSAF for Kaohsiung
Harbor T. crenata may be caused by the food intake because the BTs
accumulated in crab come primarily from the various sources of
food.
Our BSAF values was higher than the BSAF value of less than 1 as
reported by Viglino et al. (2005) for snow crab. This comparison
reveals that various crab species have different efficiencies of pro-
ducing CYP450 enzyme; the difference may also be caused by
different food intake. Regardless of the cause, higher TBT accumu-
lation in the body of T. crenata indicates that this crab species is
capable of tolerating relatively high TBT and can mirror the BTs
pollution in sediment. Hence, T. crenata is suitable to be used as a
bioindicator for TBT pollution in sediment of Kaohsiung Harbor.
4. Conclusions
The distribution of BTs (including TBT and its metabolic prod-
ucts, i.e. MBT and DBT) accumulated in T. crenata in Kaohsiung
Harbor water body that has high risks of BTs pollution were
 C.-W. Chen et al. / International Biodeterioration & Biodegradation xxx (2016) 1e8 7

Table 3
Correlations among concentration of BTs, butyltin degradation index (BDI), and biota-sediment accumulation factor (BSAF) for Kaohsiung Harbor T. crenata.

(A) n ¼ 31
Itema MBT (H) DBT (H) TBT (H) SBTs (H) MBT (M) DBT (M) TBT (M)
DBT (H) 0.77b
TBT (H) 0.62b 0.56b
SBTs (H) 0.90b 0.88b 0.85b
MBT (M) 0.17 0.27 0.04 0.14
DBT (M) 0.03 0.01 0.03 0.02 0.69b
TBT (M) 0.67b 0.56b 0.62b 0.70b 0.44c 0.37c
SBTs (M) 0.32 0.30 0.26 0.33 0.77b 0.88b 0.75b
(B) n ¼ 9
MBT (H) DBT (H) TBT (H) SBTs (H) MBT (M) DBT (M) TBT (M) SBTs (M)
BDI (H) 0.03 0.04 0.57 0.27 0.30 0.01 0.29 0.14
BSAFSBTs (H) 0.06 0.17 0.24 0.18 0.59c 0.76c 0.04 0.38
BSAFTBT (H) 0.23 0.26 0.03 0.18 0.35 0.53 0.02 0.26
BDI (M) 0.36 0.36 0.57 0.48 0.38 0.42 0.48 0.09
BSAFSBTs (M) 0.20 0.14 0.24 0.21 0.70c 0.87b 0.03 0.48
BSAFTBT (M) 0.28 0.26 0.29 0.30 0.61 0.76c 0.08 0.45
Nor. TBT 0.82b 0.91b 0.89b 0.95b 0.04 0.02 0.75c 0.48
Nor. SBTs 0.79c 0.87b 0.91b 0.94b 0.02 0.06 0.72c 0.44
(C) n ¼ 9
BDI (H) BSAFSBTs (H) BSAFTBT (H) BDI (M) BSAFSBTs (M) BSAFTBT (M) Nor. TBT
BSAFSBTs (H) 0.15
c
BSAFTBT (H) 0.36 0.77
BDI (M) 0.43 0.68c 0.36
BSAFSBTs (M) 0.01 0.91b 0.79c 0.57
BSAFTBT (M) 0.13 0.78c 0.78c 0.41 0.93b
Nor. TBT 0.26 0.45 0.38 0.58 0.44 0.50
Nor. SBTs 0.32 0.48 0.37 0.61 0.47 0.53 0.99b
a
H: Hepatopancreas; M: Muscle; Nor. TBT: Normalized TBTsediment; Nor. SBTs: Normalized SBTssediment.
b
Correlation is significant at the 0.01 level (2-tailed).
c
Correlation is significant at the 0.05 level (2-tailed).

analyzed. The crab specimens collected from various stations in
harbor had different BTs concentration (p < 0.05) reflecting that BTs
had different spatial distributions in harbor. Additionally, T. crenata
accumulated more BT in hepatopancreas than muscle; the BT
metabolic products, i.e. DBT and MBT, were the major BT species
contained in hepatopancreas whereas TBT was the major BT species
in muscle tissue. Based on the BDI and BSAF values, it is clear that
TBT is effectively degraded by T. crenata although a relatively high
concentration of TBT is stilled accumulated in crab body. Addi-
tionally, the results also indicated that harbor bottom sediment and
the food ingested by crab contributed to the accumulation of BT in
T. crenata.
Acknowledgments
This research was supported by the Taiwan International Ports
Corporation (TIPC), Taiwan. The authors would like to thank the
personnel of the TIPC for their support throughout this project
(2006 Monitoring survey for sediments, water, and ecology in
Kaohsiung Harbor).
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ibiod.2016.02.018.
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