Davies, I.M.

Gubbins, M.

Roose, P.

Vethaak, D.

Open Marine Archive

ICES Cooperative Research Report No. 315 | 197 
   
28 Technical annex: recommended packages of chemical and 
biological methods for monitoring on a determinant basis 
Matt Gubbins, Patrick Roose, Ian M. Davies, and Dick Vethaak 
28.1 Review of CEMP requirements 
This technical annex was prepared by reviewing the chemical determinants listed in 
the OSPAR Coordinated Environmental Monitoring Programme (CEMP) and pre-
CEMP (ASMO, 2007a) and considering the most appropriate chemical analyses and 
biological effects techniques that could be applied in an integrated fashion to 
monitor for these compounds in the marine environment. 
Some general points concerning integrated monitoring were noted during this 
process: 
• In some cases, the list of contaminants that should be reported under the 
CEMP (and pre-CEMP) may be insufficient for an integrated approach. In 
order to aid interpretation of biological effects measurements, an 
integrated assessment may require data on related contaminants, which 
would elicit a response on the biological effects components of the 
methods packages. Determinants additional to those required under the 
CEMP have, therefore, been added to the packages below. 
• It was felt that a fully ”integrated” approach to monitoring should include 
passive sampling of contaminants as part of the package of methods. This 
will provide information on availability of contaminants in sediments and 
allow for temporally integrated sampling of contaminants in water. 
(Guidelines for the application of passive samplers are available from 
ICES Working Group on Marine Sediments in Relation to Pollution 
(WGMS).) 
• The biological effects techniques applied to these packages of methods are 
listed either in the ICES Working Group on Biological Effects of 
Contaminants (WGBEC) recommended techniques list (ICES, 2007c) or 
form part of the fish and shellfish methods packages proposed in the draft 
Joint Assessment and Monitoring Programme (JAMP) Guidelines for 
integrated monitoring and assessment of contaminants and their effects 
(ASMO, 2007b). The biological effects methods included here are 
separated into those appropriate to monitoring selected fish species, 
shellfish (mussels), and bioassays (sediment, water, and in vitro tests). 
• It should be noted that the biological effects methods listed here are those 
which may form part of an overall integrated monitoring package and are 
likely to be affected by the OSPAR priority contaminants in question. 
Many of the effects measurements listed are ”general” biological effects, 
which are indicative of stress or health status of marine organisms or 
general toxicity in the sediments and water column. These may be affected 
by a wide range of contaminants and are not specific to the contaminants 
in question. Therefore, for each group of substances, the most specific and 
relevant biological effects techniques have also been highlighted. 
• These packages of methods should be considered supplemental to the 
existing JAMP Guidelines for Contaminant-specific (OSPAR 2003–10) and 
General (1997–7) Biological Effects Monitoring and the JAMP Guidelines 
on Contaminants in Biota (OSPAR 1999–2) and Sediment (OSPAR 2002–
198 | ICES Cooperative Research Report No. 315 
 
16). The JAMP Guidelines provide more detailed background on the 
biological effects and chemical analysis methods referred to here and the 
necessary cofactors that should be recorded for these techniques. The 
packages of methods presented here combine contaminant-specific effects 
with the general biological effects methods that are likely to respond to the 
contaminants. They also deal with groups of contaminants not addressed 
by the contaminant specific guidelines and propose further integration of 
techniques, such as passive sampling and invertebrate methods for metals. 
The priority chemical determinants from the OSPAR CEMP and pre-CEMP are as 
follows (taken from ASMO, 2007a). (The appendices referred to are CEMP 
appendices.) 
The following components of the CEMP are to be measured on a mandatory basis: 
• the heavy metals cadmium, mercury, and lead in biota and sediment 
(appendix 2); 
• the PCB congeners CB 28, CB 52, CB 101, CB 118, CB 138, CB 153, and CB 
180 in biota and sediment (appendix 3); 
• the PAHs anthracene, benz[a]anthracene, benzo[ghi]perylene, 
benzo[a]pyrene, chrysene, fluoranthene, ideno[1,2,3-cd]pyrene, pyrene, 
and phenanthrene in biota and sediment (appendix 4); 
• TBT in sediment (biota voluntary/pre-CEMP; appendix 5). 
The following components are currently part of the pre-CEMP and are to be 
measured on a voluntary basis: 
• the brominated flame retardants HBCD and PBDEs 28, 47, 66, 85, 99, 100, 
153, 154, and 183 in biota and sediment, and BDE 209 in sediment 
(appendix 8); 
• the planar PCB congeners CB 77, 126, and 169 in biota. Monitoring of 
those congeners in sediment should be undertaken only if levels of marker 
PCBs are e.g. 100-fold higher than the background assessment 
concentration (appendix 9); 
• the alkylated PAHs C1-, C2-, and C3-naphthalenes, C1-, C2-, and C3-
phenanthrenes, and C1-, C2-, and C3-dibenzothiophenes and the parent 
compound dibenzothiophene in biota and sediment (appendix 10); 
• perfluorooctane sulfonate (PFOS) in sediment, biota, and water (appendix 
12); 
• polychlorinated dibenzodioxins and furans in biota and sediment 
(appendix 13). 
28.2 Methods package for metals 
Although cadmium, mercury, and lead are the only mandatory metal determinants 
under the CEMP, other metal species are needed to interpret the biological effects 
data as part of an integrated package. Additional metal species needed include 
copper and zinc. Metals analysis should be performed on sediments and biota 
collected from the same times and locations, where possible. Cofactors for sediment 
analysis are also required, including aluminium and lithium. Diffusive gradients in 
thin films (DGTs) present the opportunity to undertake passive sampling for metal 
species to allow temporally integrated sampling of water and measure availability of 
metals in sediments. 
ICES Cooperative Research Report No. 315 | 199 
   
Metal-“specific” biological effects measurements include metallothionein, ALA-D, 
and oxidative stress, although both metallothionein and oxidative-stress responses 
are known to be affected by other contaminants. ALA-D is lead-specific and can be 
measured in fish blood, although it has limited use/expertise across the ICES/OSPAR 
community. It is recommended that it be applied only in areas where lead 
contamination is perceived to be a problem or where chemical monitoring indicates 
that concentrations are significantly above background. 
ALA-D is relevant only for fish. Metallothionein can be applied to fish liver and 
mussel digestive glands, although best results are obtained from mussels. There are a 
number of oxidative-stress measurements that can be made in both fish and mussels 
which could add value to an integrated package of metals methods, but owing to the 
lack of standardized methods, quality assurance, and assessment criteria, it is 
suggested that this method is not an essential part of the metals package. 
A number of ”general” biological effects measurements in fish and shellfish will be 
affected by environmental metal contamination and these are shown in Figure 28.1. 
In vivo bioassays are also relevant measurements for the effects of metals. 
Metallothionein in mussels and ALA-D in fish are considered the most 
specific/relevant biological effects methods for metals. 
 
Metals 
Chemistry Biological effects techniques 
Fish Shellfish 
 
Bioassays 
Oxidative stress  
 
Metallothionein  
(liver) 
ALA-D (blood)  
if lead considered 
an issue 
General biological effects  
  Lysosomal stability 
  Externally visible fish diseases 
  Liver histopathology 
  Reproductive success 
MT  
(digestive glands)  
outside of the 
spawning season 
General biological effects 
  Lysosomal stability 
  Mussel histopathology  
  MXR  
  Scope for Growth 
Toxicity tests 
all in vivo bioassays 
potentially relevant 
Toxicity Identification 
Evaluation (T.I.E.) 
using EDTA 
Sediments collected 
from same site as 
biota (except for 
shoreline 
invertebrates?) 
More metal spp. 
than required by 
CEMP should be 
analysed to interpret 
biological effects 
data as a minimum: 
Al, Li (cofactors for 
chemistry), Copper, 
Zinc, Lead, Hg, Cd. 
 Biota chemistry: 
Same metals as 
above. 
Passive sampling 
devices – DGTs in 
water (temporally 
integrated) and 
sediment 
(availability)  
 
Figure 28.1. Package of chemical and biological effects methods relevant to monitoring for 
metals. The most specific/relevant biological effects methods are highlighted (bold, shade). 
28.3 Methods package for PCBs, polychlorinated dibenzodioxins, and furans 
Because of the similarity of their toxicological effects, a single methods package was 
proposed for polychlorinated biphenyls (PCBs), polychlorinated dibenzodioxins, and 
furans. In addition to the OSPAR CEMP required determinants, additional 
chlorinated biphenyls (CBs) may cause biological effects, and their analysis should 
be included in an integrated monitoring approach. These include coplanar CBs 
CB105, and CB156. A variety of passive sampling devices (e.g. silicone rubber) offer 
200 | ICES Cooperative Research Report No. 315 
 
the potential for temporally integrated sampling of these compounds from water and 
investigation of their availability in sediments, and these should be employed where 
possible. 
There are no truly specific biological effects measurements available for PCBs, 
polychlorinated dibenzodioxins, and furans. The most relevant are considered to be 
induction of CYP1A/EROD activity in fish liver and application of the dioxin 
receptor-based in vitro test, DR-CALUX. 
Several other general biological effects measurements in fish and shellfish may 
respond to exposure to these compounds and are shown in Figure 28.2. DR-CALUX 
is considered the most useful in vitro bioassay technique, although chronic in vivo 
bioassays may also be relevant. 
 
PCBs, Polychlorinated dibenzodioxins and furans 
 
Chemistry Biological effects techniques 
Fish Shellfish 
(mussel) 
Bioassays 
Toxicity Identification 
Evaluation methods 
(T.I.E.) using C16 
columns 
 
Some chronic in vivo 
and in vitro tests 
relevant such as DR-
CALUX. 
Specific-No specific 
biological effects 
measurements 
available. 
 
General-These 
measurements can 
be affected by PCBs-
Lysosomal stability, 
Comet assay, MXR, 
GST, histopathology, 
Scope for Growth. 
‘Specific’  
Hepatic CYP1A/EROD, 
comet assay, 
macroscopic liver 
neoplasms 
 
General 
Liver histopathology, 
external fish diseases, 
reproductive success, 
lysosomal stability 
Sediments and biota 
sampled at the same sites 
and time. 
PCBs-CB 28, CB 52, CB 
101, CB 118, CB 138, CB 
153, and CB 180  
Planar CBs (in biota)-CB 
77, 126 and 169  
PC dibenzodioxins and 
furans 
Other CBs may cause 
effects therefore 
recommend co-planar CBs 
105, 156. 
Passive sampling- 
temporally integrated water 
concentrations and 
sediment availability-see 
WGMS Guidelines. 
 
Figure 28.2. Package of chemical and biological effects methods relevant to monitoring for PCBs, 
polychlorinated dibenzodioxins, and furans. The most specific/relevant biological effects 
methods are highlighted (bold, shade). 
28.4 Methods package for PAHs and alkylated PAHs 
Because of their similar toxicological effects, a single package of methods is proposed 
for polycyclic aromatic hydrocarbons (PAHs) and alkylated PAHs (Figure 28.3). The 
package of methods is similar to Figure 28.2 above, although chemical determinants 
should be analysed in sediment and shellfish only for biota. Because of their rapid 
metabolism in finfish, PAHs should be analysed as metabolites in bile rather than as 
parent compounds in liver or flesh. As above, passive sampling should also be 
applied where possible. 
Additional specific biological effects are applicable for PAHs/alkylated PAHs. These 
include PAH metabolites in fish bile and DNA adducts in fish liver. The most 
relevant/specific biological effects techniques are highlighted as induction of hepatic 
CYP1A/EROD, DNA adducts, and the DR-CALUX in vitro bioassay. 
ICES Cooperative Research Report No. 315 | 201 
   
General biological effects measurements will also respond to exposure to these 
compounds and are given in Figure 28.3. 
 
PAH & alkylated PAH 
 
Chemistry Biological effects techniques 
Fish Shellfish 
(mussel) 
Bioassays 
‘Specific’-hepatic 
CYP1A / EROD 
PAH bile metabolites 
DNA adducts, 
macroscopic liver 
neoplasms, comet 
assay 
 
General-Liver 
histopathology, 
externally visible fish 
diseases, reproductive 
success, lysosomal 
stability 
Specific-No specific 
biological effects 
measurements 
available. 
 
General-These 
measurements can 
be affected by PAHs-
Lysosomal stability, 
Comet assay, MXR, 
GST, histopathology, 
Scope for Growth. 
Toxicity 
Identification 
Evaluation 
methods (T.I.E.) 
using C16 columns 
Some chronic in 
vivo and in vitro 
tests relevant such 
as DR-CALUX. 
Sediments and biota 
sampled at the same sites 
and time. 
PAH: anthracene, 
benz[a]anthracene, 
benzo[ghi]perylene, 
benzo[a]pyrene, chrysene, 
fluoranthene, ideno[1,2,3-
cd]pyrene, pyrene and 
phenanthrene. Alkylated 
PAH: C1-, C2-, and C3-
naphthalenes, C1-, C2- and 
C3-phenanthrenes, and C1-, 
C2- and C3-
dibenzothiophenes, 
dibenzothiophene. 
Other PAH than required 
under the CEMP may have 
effects on biological effects 
measurements and would be 
useful to aid interpretation of 
integrated monitoring. 
Chemistry in sediments and 
for biota, shellfish only. 
Passive sampling- 
temporally integrated water 
concentrations and sediment 
availability-see WGMS 
Guidelines.  
 
Figure 28.3. Package of chemical and biological effects methods relevant to monitoring for PAHs 
and alkylated PAHs. The most specific/relevant biological effects methods are highlighted (bold, 
shade). 
28.5 Organotins 
It was felt that the package of methods appropriate to organotin monitoring was 
already very well described by the JAMP Guidelines on organotin-specific 
monitoring and included a suite of parameters relevant to imposex/intersex in 
gastropods, TBT, DBT, MBT, TPhT, DPhT, MPhT in sediments (for offshore 
monitoring), and in biota, where appropriate (voluntary). It was noted that passive 
sampling for organotins may become an option for integrated monitoring of 
organotins in future. It was also noted that bivalve embryo bioassays are sensitive to 
dissolved TBT at the ng l–1 level. 
28.6 BFRs 
It was noted that there are currently very few biological effects methods available 
and tested in a monitoring context for measuring the effects of brominated flame 
retardants (BFRs). The determinants required for CEMP are HBCD and PBDEs 28, 
47, 66, 85, 99, 100, 153, 154, and 183 in biota and sediment, and BDE 209 in sediment. 
Passive sampling is also relevant. 
There are no specific biological effects techniques available. Thyroid hormone 
receptor assays in fish blood are relevant, but have not been well field-tested, nor is 
this an ICES recommended technique. Recent studies on the toxicological properties 
202 | ICES Cooperative Research Report No. 315 
 
of these compounds in fish suggest that there are limited overt effects that can be 
detected by existing techniques. 
28.7 PFOS 
Perfluorooctane sulfonate (PFOS) analysis in sediment, biota, and water is included 
in the list of pre-CEMP determinants; however, no specific biological effects 
techniques are recommended here. It was noted that the compound may have 
endocrine-disrupting effects and that some endocrine disruptor-relevant endpoints 
may be appropriate along with general biological effect measurements, such as 
reproductive success. A battery of short-term, low-volume bioassays (in vitro and in 
vivo) using extracts can be used to perform a first screening/assessment of 
unintended impacts and novel contaminants (see background document on water 
bioassays). These extracts can be derived from water, sediment, biota, and/or passive 
samplers. Information obtained from bioanalysis can also be used as input for the 
design of future monitoring programmes and the development of appropriate higher 
level biological effects techniques biomarkers. However, a package of methods 
relevant to PFOS would require further consideration. 


Technical annex: recommended packages of chemical and biological methods for monitoring on a determinant basis

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