Heike Helmholz 2 , Marijana
1
2
Ru đ er Boškovi ć
Helmholtz-Zentrum
Erk 1 , Daniel Pröfrock 2 , Željka Strižak 1 , Dušica Ivankovi
Geesthacht, Institute for Coastal Research; Marine Bioanalytical ć 1 and Andreas Prange
Institute, Division for Marine and Environmental Research, Zagreb, Croatia
2
Chemistry, Geesthacht, Germany
đ
ć
Division for Marine and Environmental Research
Next to the natural sources, a large number of metals are present in the environment as a result of human activities. Since the ban of organo-tin compounds, Cu is the main component of anti-fouling paints, with the result that boat traffic is a major anthropogenic source of Cu in the coastal aquatic environment. Besides, zinc oxide is generally used in combination with Cu(I) as a booster. As a result of the leaching of chemicals from the painted surface, concentrations of metals are significantly elevated in semi-enclosed marine systems, such as harbours, marinas and estuaries, where the transport, berthing or docking of vessels is extensive.
Aim of the study was the assessment of metal exposure in coastal areas under indirect or direct influence of maritime activities and nautical tourism (port and marinas) using integrated biologicalchemical analytical approach and mussels ( Mytilus galloprovincialis ) as bioindicator species.
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 investigation of metal distribution in the cytosolic fractions of proteins separated by size exclusion chromatography (SE-HPLC) and detected by inductively coupled plasma mass spectrometry (ICP-MS) cellular energy allocation (CEA) methodology applied as a general biomarker of physiological status of mussels non-target bioanalytical approach utilizing 2D gel-electrophoresis will give hints for further potential protein biomarker indigenous mussel populations were sampled in the middle part of the eastern coast of the Adriatic Sea (Croatia) in March and September 2011
– marina Kaštela (KM), further ports and marinas and a reference site
Zabla ć e (ZA)
Kaštela marina (KM) Zabla ć e (ZA)
Copper
Cadmium pH 5
200 kDa
116 kDa
67 kDa
45 kDa
29 kDa
21 kDa
14.3 kDa
V
0
–
OVA dextran blue
–
(M r
= 2000 kDa ) ovalbumine (M r
= 43 kDa )
Distribution of Cu in the cytosolic fractions of proteins separated by SE-HPLC
(column: Superdex 75, flow: 0.3 mL/min, 0.02M Tris, pH 7.4) and detected by ICP-MS
Chy – aMb – chymotripsinogen (Mr= 25 kDa ) apomyoglobine (Mr= 17 kDa )
Distribution of Cd in the cytosolic fractions of proteins separated by SE-HPLC
(column: Superdex 75, flow: 0.3 mL/min, 0.02M Tris, pH 7.4) and detected by ICP-MS
pH
KM gill
8
ZA gill
ZA gland
IEF pH range 5-8
2nd dimension Criterion AnykDa
Comassie staining
Protein load 1mg per IPG strip (11 cm)
KM gland
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Matching and fusion of 2D gels obtained from different stations
(orange + blue)
Detection and selection of differently expressed spots
In-gel digestion and mass spectrometry for protein identification
(Methods see poster WE 317 1020)
This study represents a contemporary and new approach in quantification and assessment of the environmental risk caused by metal exposure, as well as the potential toxic effect of metals in mussels
 An altered profile and concentration of metal-associated proteins can be detected in relation to metal charges in environmental samples
 Whole protein pattern were tissue specific and vary in differently contaminated sampling sites
 Physical environmental conditions and the physiological performance of the organisms have to be taken into account for the assessment of chemical stress factors
 Further progress is needed in the chemical elucidation and identification of the potential biomarker for metal contamination
Helmholtz-Zentrum Geesthacht • Max-Planck-Straße 1 • 21502 Geesthacht • Phone +49 (0)4152 87-1844 • Fax +49 (0)4152 87-1875 • www.hzg.de
Contact: Dr. Heike Helmholz
Ru đ er Boškovi ć heike.helmholz@hzg.de
and Dr. Marijana Erk erk@irb.hr
PK: 50752021
Institute, Division for Marine and Environmental Research, Zagreb, Croatia; http://www.irb.hr/eng/Research/Divisions-and-Centers/Division-for-Marine-and-Environmental-Research/Laboratoryfor-Biological-Effects-of-Metals