The certification of the mass concentration of As, Cd, Co, Cu, Mn, Mo, Ni and Pb, and density in seawater : ERM®-CA403 : certification report.
Date: 2018-10-16
Institutions: Joint Research Centre
Subjects: Community certification, environmental monitoring, environmental standard, quality control circle, quality of the environment, research method, research report, saltwater, spectrometry, territorial waters, testing

CERTIFICATION REPORT 

The certification of the mass concentration of As, Cd, Co, Cu, 
Mn, Mo, Ni and Pb, and density in seawater:  
ERM®- CA403 

EUR 29055  EN 

 
 
  
 
 
 
 
 
JRC-IRMM promotes a common and  

provides reference measurements. 

European Commission  
Joint Research Centre 
Directorate F – Health, Consumers and Reference Materials 

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All images © European Union 2018 

JRC 110329 

EUR 29055 EN 

ISBN 978-92-79-77706-6 (PDF) 
ISSN 1831-9424 (online) 
doi: 10.2760/405177 

Luxembourg: Publications Office of the European Union, 2018 
© European Union, 2018 

Reproduction is authorised provided the source is acknowledged. 

Abstract 

This report describes the production of ERM®-CA403, a seawater material certified for the density and the mass concentrations of As, Cd, Co, Cu, Mn, Mo, 
Ni, and Pb. The material was produced following ISO Guide 34:2009. The starting material was 1500 L of seawater collected at Southern Bight just inside 
of Belgian territorial waters. The sample was filtered, acidified and spiked before filling into 500 mL high density polyethylene (HDPE) bottles which were 
placed into an aluminised PET/M (metallized polyethylene terephthalate) sachets. The material was sterilised by gamma irradiation. 
Between unit-homogeneity was quantified and stability during dispatch and storage were assessed in accordance with ISO Guide 35:2006. Due to the 
inherent homogeneity of filtered water samples, determination of minimum sample intake (within-unit heterogeneity) was not required. 
The material was characterised by an inter-comparison among laboratories of demonstrated competence and adhering to ISO/IEC 17025:2005. Technically 
invalid results were removed but no outlier was eliminated on statistical grounds only.  
Uncertainties of the certified values were calculated in compliance with the Guide to the Expression of Uncertainty in Measurement (GUM) and include 
uncertainties related to possible inhomogeneity, and instability and to characterisation. 
The material is intended for the quality control and assessment of method performance. As any reference material, it can also be used for control charts 
or validation studies. The certified reference material (CRM) is available in bottles containing 500 mL of seawater. 

The CRM was accepted as European Reference Material (ERM®) after peer evaluation by the partners of the European Reference Materials consortium.   

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
CERTIFICATION REPORT 

The certification of the mass concentration of As, Cd, Co, Cu, 
Mn, Mo, Ni and Pb, and density in seawater: 
ERM®- CA403 

R. Sánchez, E. de Vos, J. Snell, H. Emteborg, A. Held 

European Commission, Joint Research Centre 

Directorate F – Health, Consumers and Reference Materials 
Geel, Belgium 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Disclaimer 

Certain commercial equipment, instruments, and materials are identified in this paper to specify adequately the 
experimental  procedure.  In  no  case  does  such  identification  imply  recommendation  or  endorsement  by  the 
European Commission, nor does it imply that the material or equipment is necessarily the best available for the 
purpose.  

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
Summary 

This  report  describes  the  production  of  ERM®-CA403,  a  seawater  material  certified  for  the 
density  and  the  mass  concentrations  of  As,  Cd,  Co,  Cu,  Mn,  Mo,  Ni,  and  Pb.  The  material 
was produced following ISO Guide 34:2009 [1]. The starting material was 1500 L of seawater 
collected at Southern Bight just inside of Belgian territorial waters. The sample was filtered, 
acidified  and  spiked  before  filling  into  500  mL  high  density  polyethylene  (HDPE)  bottles 
which  were  placed  into  an  aluminised  PET/M  (metallized  polyethylene  terephthalate) 
sachets. The material was sterilised by gamma irradiation. 

Between  unit-homogeneity  was  quantified  and  stability  during  dispatch  and  storage  were 
assessed  in  accordance  with  ISO  Guide  35:2006  [2].  Due  to  the  inherent  homogeneity  of 
filtered  water  samples,  determination  of  minimum  sample  intake  (within-unit  heterogeneity) 
was not required. 

The material was characterised by an inter-comparison among laboratories of demonstrated 
competence  and  adhering  to  ISO/IEC  17025:2005  [3].  Technically  invalid  results  were 
removed but no outlier was eliminated on statistical grounds only.  

Uncertainties  of  the  certified  values  were  calculated  in  compliance  with  the  Guide  to  the 
Expression  of  Uncertainty  in  Measurement  (GUM)  [4]  and  include  uncertainties  related  to 
possible inhomogeneity, and instability and to characterisation. 

The material is intended for the quality control and assessment of method performance. As 
any  reference  material,  it  can  also  be  used  for  control  charts  or  validation  studies.  The 
certified reference material (CRM) is available in bottles containing 500 mL of seawater. 

The  CRM  was  accepted  as  European  Reference  Material  (ERM®)  after peer  evaluation  by 
the partners of the European Reference Materials consortium. 

The following values were assigned: 

Density (at 20 °C) 

As1 
Cd1 
Co1 
Cu1 
Mn1 
Mo1 
Ni1 
Pb1 

Density 

Mass concentration 

Certified values 2) 
[g/mL] 

1.02352 

Certified values 1,2) 
[µg/L] 

1.90 
0.094 
0.074 
0.87 
2.47 
12.0 
1.04 
0.098 

Uncertainty 3) 
[g/mL] 

0.00005 

Uncertainty 3) 
[µg/L] 

0.13 
0.011 
0.011 
0.13 
0.11 
0.6 
0.16 
0.010 

1) as obtained by measurement methods with quantification by Inductively Coupled Plasma Mass Spectrometry 

2)  Certified  values  are  values  that  fulfil  the  highest  standards  of  accuracy.  The  given  values  represent  the 
unweighted mean value of the means of accepted sets of data, each set being obtained in a different laboratory 
and/or  with  a  different  method  of  determination.  The  certified  value  and  its  uncertainty  are  traceable  to  the 
International System of units (SI). 

3) The uncertainty is the expanded uncertainty of the certified value with a coverage factor, k, corresponding to a 
level of confidence of about 95 % estimated in accordance with ISO/IEC Guide 98-3, Guide to the Expression of 
Uncertainty in Measurement (GUM:1995), ISO, 2008. 

 
 
 
 
 
 
 
Table of contents 

Summary .............................................................................................................................. 1 
Table of contents ................................................................................................................. 3 
Glossary ............................................................................................................................... 5 
Introduction .............................................................................................................. 9 
1 
Background ............................................................................................................... 9 
1.1 
Choice of the material ...............................................................................................10 
1.2 
Design of the project .................................................................................................10 
1.3 
Participants .............................................................................................................10 
2 
Project management and evaluation .........................................................................10 
2.1 
Processing ................................................................................................................10 
2.2 
Homogeneity study ...................................................................................................10 
2.3 
Stability study ...........................................................................................................10 
2.4 
Characterisation ........................................................................................................11 
2.5 
Material processing and process control .............................................................12 
3 
Origin of the starting material ....................................................................................12 
3.1 
Initial characterisation and spiking ............................................................................12 
3.2 
Processing ................................................................................................................12 
3.3 
Homogeneity ...........................................................................................................13 
4 
Between-unit homogeneity........................................................................................13 
4.1 
4.2  Within-unit homogeneity and minimum sample intake...............................................19 
Stability....................................................................................................................20 
5 
Short-term stability study ..........................................................................................20 
5.1 
Long-term stability study ...........................................................................................23 
5.2 
Estimation of uncertainties ........................................................................................24 
5.3 
Characterisation .....................................................................................................25 
6 
Selection of participants ............................................................................................25 
6.1 
6.2 
Study setup ...............................................................................................................26 
6.3  Methods used ...........................................................................................................26 
6.4 
Evaluation of results .................................................................................................26 
6.4.1  Technical evaluation .................................................................................................26 
6.4.2  Statistical evaluation .................................................................................................27 
Value Assignment ...................................................................................................29 
7 
Certified values and their uncertainties .....................................................................30 
7.1 
Indicative values and their uncertainties....................................................................31 
7.2 
Additional material information ..................................................................................31 
7.3 
8 
Metrological traceability and commutability .........................................................32 
8.1  Metrological traceability ............................................................................................32 
Instructions for use ................................................................................................32 
9 
Safety information .....................................................................................................32 
9.1 
9.2 
Storage conditions ....................................................................................................32 
Preparation and use of the material ..........................................................................32 
9.3 
9.4  Minimum sample intake ............................................................................................32 
Use of the certified value ..........................................................................................33 
9.5 
Acknowledgments ..................................................................................................34 
10 
References ..............................................................................................................35 
11 

3 

 
4 

 
 
Glossary 

AFS  

Atomic fluorescence spectrometry 

ASTM 
international 

ASTM international (formerly American Society for Testing and 
Materials) 

ANOVA  

Analysis of variance 

b 
BCR® 

BIPM 

CC  

CCT 

CEN 

CI 

CRM 

EC 

EN 

EQS 
ERM® 

EU 

GRP 

GUM 

HDPE 

HEPA 

Slope in the equation of linear regression y = a + bx 

One of the trademarks of CRMs owned by the European Commission; 
formerly Community Bureau of Reference 

Bureau International des Poids et Mesures (International Bureau of 
Weights and Measures) 

Collision cell 

Collision cell technology 

European Committee for Standardization 

confidence interval 

Certified reference material 

European Commission 

European norm (standard) 

Environmental quality standard 

Trademark of European Reference Materials 

European Union 

Glass fibre reinforced plastic 

Guide to the Expression of Uncertainty in Measurements [4] 

High density polyethylene 

High-efficiency particulate arrestance 

HG-AFS 

ICP-OES 

ICP-MS 

ICP-QMS 

Hydride generation-atomic fluorescence spectrometry 

Inductively coupled plasma optical emission spectrometry 

Inductively coupled plasma-mass spectrometry 

ICP-Quadrupole mass spectrometry  

ICP-SFMS 

ICP-Sector field mass spectrometry  

ISO 

IUPAC 

JRC 

k 

LOD 

LOQ 

MS 

International Organization for Standardization  

International Union of Pure and Applied Chemistry  

Joint Research Centre of the European Commission 

Coverage factor 

Limit of detection 

Limit of quantification 

Mass spectrometry 

MSbetween 

Mean of squares between-unit from an ANOVA 

5 

 
 
MSr  

MSwithin  

MUMM 

N 

N 

n.a. 

n.c. 

n.d. 

NIST 

NRC 

OD 

P 

PET/M 

PFA 

PP 

PTFE 

Pyc 

QA 

QC 

rel 

RM 

RSD 

RSE 

r2 

s 

sbb

sbetween 

se 

SF-MS 

SI 

sr 

swithin 

swb 

T 

Mean of squares between-run from an ANOVA 

Mean of squares within-unit from an ANOVA 

Management Unit of the North Sea Mathematical Models and the 
Scheldt estuary 

Number of replicates per unit 

Number of samples (units) analysed 

Not applicable 

Not calculated 

Not detectable 

National Institute of Standards and Technology (USA) 

National Research Council (Canada) 

Oscillating type density meter 

Number of technically valid datasets 

Metallized polyethylene terephthalate 

Perfluoroalkoxy copolymer resin 

Polypropylene 

Polytetrafluoroethylene 

Pycnometer (for density measurement) 

Quality assurance 

Quality control 

Index denoting relative figures (uncertainties etc.) 

Reference material 

Relative standard deviation 
Relative standard error (=RSD/(cid:214) n) 

Coefficient of determination of the linear regression 

Standard deviation 

Between-unit standard deviation; an additional index "rel" is added when 
appropriate 

Standard deviation between groups as obtained from ANOVA; an 
additional index "rel" is added as appropriate 

Standard error 

Sector-field mass spectrometry 

International System of Units 

Between-run standard deviation 

Standard deviation within groups as obtained from ANOVA; an additional 
index "rel" is added as appropriate 

Within-unit standard deviation 

Temperature 

6 

 
t 

ti 

ta

, df 

tsl 

u 

U 
u*

bb  

ubb 

uc 

ucal 

uchar  

uCRM 

UCRM  

u∆ 

ults 

umeas 

Umeas 

urec  

usts 

ut 

VIM 

Time 

Time point for each replicate 
Critical t-value for a t-test, with a level of confidence of 1-a
degrees of freedom 

 and df 

Proposed shelf life 

standard uncertainty  

expanded uncertainty 

Standard uncertainty related to a maximum between-unit inhomogeneity 
that could be hidden by method repeatability; an additional index "rel" is 
added as appropriate 

Standard uncertainty related to a possible between-unit inhomogeneity;  
an additional index "rel" is added as appropriate 

combined standard uncertainty; an additional index "rel" is added as 
appropriate 

Standard uncertainty of calibration 

Standard uncertainty of the material characterisation; an additional index 
"rel" is added as appropriate 

Combined standard uncertainty of the certified value; an additional index 
"rel" is added as appropriate 

Expanded uncertainty of the certified value; an additional index "rel" is 
added as appropriate 

Combined standard uncertainty of measurement result and certified 
value 

Standard uncertainty of the long-term stability; an additional index "rel" is 
added as appropriate 

Standard measurement uncertainty 

Expanded measurement uncertainty 

Standard uncertainty related to possible between-unit inhomogeneity 
modelled as rectangular distribution; an additional index "rel" is added as 
appropriate 

Standard uncertainty of the short-term stability; an additional index "rel" 
is added as appropriate 

Standard uncertainty of trueness 

Vocabulaire International de Métrologie – Concepts Fondamentaux et 
Généraux et Termes Associés (International Vocabulary of Metrology – 
Basic and General Concepts and Associated Terms) [ISO/IEC Guide 
99:2007]  

WFD 

Water Framework Directive 

x  

nsx  

refx

a  

Arithmetic mean 

Arithmetic mean of all results of normal stock samples  

Arithmetic mean of results of reference samples 

significance level 

7 

 
 
D meas 

n s,meas 
n

MSwithin

Absolute difference between mean measured value and the certified 
value 

Degrees of freedom for the determination of the standard deviation smeas 

Degrees of freedom of MSwithin 

8 

 
 
1 

Introduction  

1.1  Background 

In the year 2000, the European Commission adopted the Water Framework Directive (WFD; 
[5]).  Since  then,  a  number  of  directives  that  provide  the  legislative  framework  for  the 
protection  of  surface  waters  and  other  water  resources  have  been  added  to  the  initial 
Directive.  

The  WFD  constitutes  a  strategy  against  the  chemical  pollution  of  European  water  bodies 
(rivers,  lakes,  ground  and  coastal  waters)  and  requires  all  member  states  to  implement 
specific  monitoring  programmes  covering  the  chemical  status  of  surface  waters  and  the 
assessment  of  significant,  long-term  pollution  trends  resulting  from  human  activities. 
Directive  2013/39/EU  [6]  on  environmental  quality  standards  in  the  field  of  water  policy 
contains a list of priority I substances, including cadmium, nickel, lead and their compounds 
for which maximum allowable concentrations and annual average values were set for surface 
waters (Table 1). In addition, the directive lists values for Zn, Cu, Cr, Se, As, Mo, and Co as 
priority  II  substances.  The  quality  and  comparability  of  analytical  results  generated  by  the 
Member States to perform water chemical monitoring should be ensured. This is guaranteed 
by  Directive  2009/90/EC  [7],  which  lays  down  the  technical  specifications  for  chemical 
analysis and monitoring of the water status. Among other criteria, the Directive 2009/90/EC 
establishes  that  EU  laboratories  shall  ensure  their  competences  in  analysing  relevant 
physico-chemical  or  chemical  parameters  by  the  analysis  of  available  certified  reference 
materials  (CRMs)  that  are  representative  of  collected  samples  which  contain  appropriate 
levels of concentrations in relation to relevant Environmental Quality Standards (EQS). 

Table 1. Environmental quality standards for Cd, Pb and Ni.  

Annual average – 
EQS [µg/L] 

Annual average 
– EQS [µg/L] 

Inland surface 
waters 

Other surface 
waters 

Maximum 
allowable 
concentration – 
EQS [µg/L] 

Maximum 
allowable 
concentration – 
EQS [µg/L] 

Inland surface 
waters 

Other surface 
waters 

< 0.08 (Class 1) 

0.20 

< 0.45 (Class 1) 

< 0.45 (Class 1) 

0.08 (Class 2) 

0.09 (Class 3) 

0.15 (Class 4) 

0.25 (Class 5) 
1.2¥ 

4¥ 

0.45 (Class 2) 

0.45 (Class 2) 

0.60 (Class 3) 

0.60 (Class 3) 

0.90 (Class 4) 

0.90 (Class 4) 

1.5 (Class 5) 

1.5 (Class 5) 

1.3 

8.6 

14 

34 

14 

34 

Cd and its 
compounds 
(depending on 
water hardness 
classes)* 

Pb and its 
compounds 

Ni and its 
compounds 

*: Class 1: < 40 mg CaCO3 /L, Class 2: 40 to < 50 mg CaCO3 /L, Class 3: 50 to < 100 mg 
CaCO3 /L, Class 4: 100 to < 200 mg CaCO3 /L and Class 5: ≥ 200 mg CaCO3 /L. 
¥: These EQS refer to bioavailable concentrations of the substances.  

Moreover,  the  European  Union  has  adopted  two  instruments,  the  EU  Recommendation  on 
Integrated  Coastal  Zone  Management  [8]  and  the  Marine  Strategy  Framework  Directive 
(2008/56/EC)  [9]  as  complimentary  tools  to  the  protection  of  marine  waters.  The  Marine 

9 

 
 
 
Strategy Framework Directive aims to achieve good environmental status of the EU's marine 
waters  and  to  protect  the  resource  base  upon  which  marine-related  economic  and  social 
activities  depend.  The  Commission  Decision  on  criteria  and  methodological  standards  on 
good  environmental  status  of  marine  waters  [10]  contains  a  number  of  criteria  and 
associated  indicators  for  assessing  good  environmental  status,  in  relation  to  the  eleven 
descriptors  of  good  environmental  status  laid  down  in  Annex  I  of  the  Marine  Strategy 
Framework  Directive.  Among  the  descriptors,  contaminants,  such  as  metals  toxic  at  trace 
concentrations, are listed. 

1.2  Choice of the material 

ERM-CA403  was  developed  as  replacement  for  the  seawater-based  certified  reference 
material  BCR®-403.  The  concentration  levels  that  were  targeted  in  the  new  material  were 
based on those of the WFD and the water had to be spiked with Cd, Cr, Ni and Zn following 
assessment  of  the  trace  element  composition  of  the  raw  material  that  was  collected.  The 
spiking aimed to increase the concentrations of the elements to around their respective EQS. 
The  target  uncertainties  for  the  mass  concentrations  of  the  elements  was  below  20 %,  to 
ensure  that  the  material  was  suitable  for  the  quality  control  of  quantitative  methods  for 
measuring elements in seawater. 

1.3  Design of the project 

The  certification  of  the  element  concentrations  of  ERM-CA403  was  performed  by 
interlaboratory comparison. 

2  Participants 

2.1  Project management and evaluation 

European Commission, Joint Research Centre, Geel (BE) 
(accredited to ISO Guide 34 for production of certified reference materials, BELAC No. 268-RM) 

2.2  Processing  

European Commission, Joint Research Centre, Geel (BE) 
(accredited to ISO Guide 34 for production of certified reference materials, BELAC No. 268-RM) 

2.3  Homogeneity study 

European Commission, Joint Research Centre, Geel (BE) 
(accredited to ISO Guide 34 for production of certified reference materials, BELAC No. 268-RM; measurements 
under the scope of ISO/IEC 17025 accreditation BELAC No. 268-TEST)  

Agenzia  Regionale  per  la  Prevenzione  e  Protezione  Ambientale  del  Veneto,  Mestre  (IT) 
(measurements under the scope of ISO/IEC 17025 accreditation ACCREDIA No. 0838) 

Laboratorios Tecnológicos del Levante, Paterna (ES)  
(measurements under the scope of ISO/IEC 17025 accreditation ENAC No. 121/LE1782) 

2.4  Stability study 

Agenzia  Regionale  per  la  Prevenzione  e  Protezione  Ambientale  del  Veneto,  Mestre  (IT) 
(measurements under the scope of ISO/IEC 17025 accreditation ACCREDIA No. 0838) 

Helmholtz  Zentrum  München  –  Forschungszentrum  für  Gesundheit  und  Umwelt  GmbH, 
München (DE) 
(measurements under the scope of ISO/IEC 17025 accreditation DACH; accreditation number DAC-PL-0141-01-
10) 

10 

 
 
 
 
Laboratorios Tecnológicos del Levante, Paterna (ES)  
(measurements under the scope of ISO/IEC 17025 accreditation ENAC No. 121/LE1782) 

Reagecon Diagnostics Ltd, Shannon (IE) 
(measurements under the scope of ISO/IEC 17025 accreditation NSAI, accreditation number 19.2769) 

2.5  Characterisation 

AGQ Labs & Technological Services S.L, Burguillos (ES) 
(measurements under the scope of ISO/IEC 17025 accreditation ENAC; accreditation number 305/LE1322) 

ALS Laboratory Group, ALS Scandinavia AB, Luleå (SE) 
(measurements under the scope of ISO/IEC 17025 accreditation SWEDAC; accreditation number 1087) 

Agenzia  Regionale  per  la  Prevenzione  e  Protezione  Ambientale  del  Veneto,  Mestre  (IT) 
(measurements under the scope of ISO/IEC 17025 accreditation ACCREDIA No. 0838) 

BEV - Bundesamt für Eich- und Vermessungswesen, Wien (AT) 

Brooks Rand Labs L.L.C., Seattle (USA) 
(measurements under the scope of ISO/IEC 17025 accreditation NELAP; accreditation number E87982) 

EVANS Analytical group SAS, Tournefeuille (FR) 
(measurements under the scope of ISO/IEC 17025 accreditation COFRAC; accreditation number E879821-1993) 

Helmholtz  Zentrum  München  –  Forschungszentrum  für  Gesundheit  und  Umwelt  GmbH, 
München (DE) 
(measurements under the scope of ISO/IEC 17025 accreditation DACH; accreditation number DAC-PL-0141-01-
10) 

International Atomic Energy Agency - Marine Environmental Studies Laboratory (MC) 

INM - National Institute of Metrology, Bucharest (RO)  

Institut "Jozef Stefan", Ljubljana (SI) 
(Measurements performed under ISO/IEC 17025 accreditation; Slovenska Akreditacija LP-090) 

I.N.RI.M - Istituto Nazionale di Ricerca Metrologica, Pavia (IT) 
(measurements performed under the scope of ISO/IEC 17025 accreditation, BELAC, accreditation number 015-
TEST) 

INTA - Instituto Nacional de Tecnica Aeroespacial "Esteban Terradas" (ES) 
(measurements under the scope of ISO/IEC 17025 accreditation ENAC; accreditation number 16/LC10.007) 

IPQ - Instituto Português da Qualidade, Caparica (PT) 

Marine Institute, Cork (IE)   
(measurements performed under the scope of ISO/IEC 17025 accreditation, INAB, accreditation number 130T) 

Paragon Scientific Limited, Birkenhead (GB) 
(measurements performed under the scope of ISO/IEC 17025 accreditation, UKAS, accreditation number 0640) 

Reagecon Diagnostics Ltd, Shannon (IE) 
(measurements under the scope of ISO/IEC 17025 accreditation NSAI, accreditation number 19.2769) 

VSL - Dutch Metrology Institute, Delft (NL) 
(measurements under the scope of ISO/IEC 17025 accreditation RvA, accreditation number K 999) 

11 

 
 
 
 
 
 
 
 
 
 
 
 
 
3  Material processing and process control 

3.1  Origin of the starting material 

The  starting  material  for  the  seawater  based  reference  material  was  collected  during  a 
sampling campaign from 18th to 21st October 2010 in the Southern Bight just inside of Belgian 
territorial  waters  in  collaboration  with  Management  Unit  of  the  North  Sea  Mathematical 
Models  and  the  Scheldt  estuary  (MUMM).  The  sample  was  pumped  using  an  immersible 
pump,  Kärcher  DFP  14000,  equipped  with  polytetrafluoroethylene  (PTFE)  tubing  into  three 
pre-cleaned vessels of 550 L each. The vessel wall is a sandwich construction and consists 
of  GRP  (glass  fibre  reinforced  plastic)  as  outer  providing  structure,  ruggedness  and 
protection  and  a  perfluoroalkoxy  copolymer  resin  (PFA)  as  an  inner  liner.  In  this  way  all 
surfaces  inside  the  drums  are  made  of  PFA  which  is  a  highly  inert  material  that  can  resist 
harsh cleaning protocols with concentrated acids if necessary. Prior to pumping the seawater 
sample into the tanks, the tubing was rinsed with several litres of seawater. The final volume 
collected was 1500 L. 

Upon  arrival  at  the  JRC  facilities  in  Geel,  the  starting  material  was  filtered  using  Versaflow 
capsule  filters  (0.8/0.45  µm,  Pall  corporation,  Ann  Arbor,  Michigan  USA),  acidified  to 
1 < pH < 2 with ultrapure hydrochloric acid (Merck) and stored at 4 °C.  

3.2  Initial characterisation and spiking 

An  initial  characterisation  of  the  seawater  based  material  was  required  to  determine  the 
concentration  level  of  trace  elements  and  to  assess  the  need  for  spiking  to  reach  target 
concentrations.  The  sample  was  analysed  by  inductively  coupled  plasma  sector  field  mass 
spectrometry (ICP-SFMS) for all target elements (As, Cd, Co, Cr, Cu, Fe, Mn, Mo, Ni, Pb, Se 
and Zn). The seawater based material required spiking with Cd, Cr, Ni and Zn. Appropriate 
volumes of liquid reference materials (1000 mg/L, Merck) were added to the seawater base 
material. Prior to filling of the bottles the concentrations of Mn and Mo were checked by ICP-
SFMS. 

3.3  Processing  

The  CRM  containers  are  500  mL  narrow-mouth  HDPE  bottles  with  polypropylene  (PP) 
closure (Nalgene, Rochester, New York, USA). Before filling, they were washed in a clean-
cell with high-efficiency particulate arrestance (HEPA) filtered air. Each bottle was filled with 
approximately 100 mL of 2 g/100 g nitric acid, and shaken for 60 minutes by a 3-dimensional 
mixer (Dynamix-CM 200, WAB, Basel, Switzerland). This was followed by two rinsing steps 
with  ultrapure  water  (18.2  MΩ·cm,  0.053  µS·cm−1,  maximum  of  50  µg  L-1  of  total  organic 
carbon,  Merck  Millipore,  Billerica  USA).  The  homogenisation  of  the  seawater  material  was 
ensured  by  recirculating  the  water  between  three  550  L  tanks  before  and  during  the  filling 
into bottles using IWAKI FS-30HT2 inert bellow pumps (Tokyo, Japan). In total about 40 full 
mixing  volumes  were  effected  and  the  spike  was  added  after  20  such  cycles  and 
subsequently  mixed  for  another  20  cycles.  A  peristaltic  pump  (Watson-Marlow,  Falmouth, 
UK)  was  used  to  pump  the  homogenised  water  from  the  three  tanks  simultaneously  using 
PTFE tubing (Bohlender, Grünsfeld, DE) to a 20 L polycarbonate demijohn. Aliquots of 500 
mL  were  filled  into  the  cleaned  HDPE  bottles  from  the  demijohn  placed  in  a  clean  bench 
(Nuair 156, Plymouth, MN, USA). By manually opening the tap at the bottom of the demijohn 
water  was  flowing  out  by  gravity  into  the  sample  bottle  at  sufficient  flow  rate.  At  regular 
intervals  the  demijohn  was  topped  up  by  switching  on  the  peristaltic  pump.  A  total  of  1513 
bottles were filled, which were subsequently labelled, placed into aluminised PET/M sachets 
and  dispatched for  sterilisation  by  gamma  irradiation  (Isotron  NV,  Ede,  NL)  at  15 kGy. The 
residual  bacterial  activity  was  checked  (in-house)  after  the  irradiation  of  the  material.  The 
results confirmed that the material showed no residual bacterial activity. 

12 

4  Homogeneity 

A key requirement for any reference material is the equivalence between the various units. In 
this respect, it is relevant whether the variation between units is significant compared to the 
uncertainty of the certified value. In contrast to that, it is not relevant if this variation between 
units is significant compared to the analytical variation. Consequently, ISO Guide 34 requires 
RM producers to quantify the between unit variation. This aspect is covered in between-unit 
homogeneity studies. 

The within-unit inhomogeneity does not influence the uncertainty of the certified value when 
the minimum sample intake is respected, but determines the minimum size of an aliquot that 
is  representative  for  the  whole  unit.  Quantification  of  within-unit  inhomogeneity  is  therefore 
necessary to determine the minimum sample intake. 

4.1  Between-unit homogeneity 

The between-unit homogeneity was evaluated to ensure that the certified values of the CRM 
are valid for all units of the material, within the stated uncertainty. 

The number of selected units corresponds to approximately the cubic root of the total number 
of  the  produced  units.  Fourteen  units  (trace  elements)  and  twelve  units  (density)  were 
selected  using  a  random  stratified  sampling  scheme  covering  the  whole  batch  for  the 
between-unit homogeneity test. For this, the batch was divided into fourteen or twelve groups 
(with a similar number of units in each group) and one unit was selected randomly from each 
group. Three independent samples were taken from each selected unit, and analysed by the 
oscillating U-tube method (density) and by ICP-SFMS (trace elements). The set of fourteen 
units for the trace element measurements were also used for the short-term stability study as 
described in section 5.2. 

The  measurements  were  performed  under  repeatability  conditions  for  density,  and  in  a 
randomised manner to be able to separate a potential analytical drift from a trend in the filling 
sequence. The measurements for the trace elements were performed in a randomised block 
design due to instrumental constraints, as the number of individual measurements required 
could  not  be  performed  on  a  single  measurement  occasion.  In  this  circumstance  improved 
precision (measured as the within-unit standard deviation) was obtained using several short 
runs in a randomised block design, and applying 2-way ANOVA to account for between-run 
variance in addition to variances between-unit and within-unit. For three replicates on each of 
14 units of ERM-CA403, the simplest randomised block design involves three measurement 
runs and each unit is measured once in random order. Runs were randomised individually in 
a  manner  to  be  able  to  separate  a  potential  analytical  drift  from  a  trend  in  the  production 
sequence.  For  selenium  the  results  of  the  study  on  trace  elements  were  unsatisfactory,  as 
the  multi-element  method  used  did  not  have  adequate  sensitivity.  Therefore,  a  dedicated 
study  was  conducted  for  selenium  using  the  same  samples  tested  for  short-term  stability. 
This study also included fourteen units and was performed under repeatability conditions on 
a single occasion (thus avoiding the need for 2-way ANOVA). The results of all studies are 
shown as graphs in Annex A. 

Density 

Regression analyses were performed to evaluate potential trends in the analytical sequence 
as  well  as  trends  in  the  filling  sequence.  No  trends  in  the  filling  sequence  or  the  analytical 
sequence were visible.  

The dataset was tested for consistency using Grubbs outlier tests on a confidence level of 99 
% on the individual results and the unit means. No outlying individual results and outlying unit 
means were detected. 

13 

 
Quantification  of  between-unit  inhomogeneity  was  accomplished  by  analysis  of  variance 
(ANOVA),  which  can  separate  the  between-unit  variation  (sbb)  from  the  within-unit  variation 
(swb).  The  latter  is  equivalent  to  the  method  repeatability  if  the  individual  samples  are 
representative for the whole unit.  

Evaluation by ANOVA requires unit means which follow at least a uni-modal distribution and 
results for each unit that follow uni-modal distributions with approximately the same standard 
deviations.  Distribution  of  the  unit  means  was  visually  tested  using  histograms  and  normal 
probability  plots.  Too few  data  are  available  for each  unit  to make  a  clear  statement  of  the 
distribution of the individual results. Therefore, it was visually checked whether all individual 
data follow a uni-modal distribution using histograms and normal probability plots. 

One  has  to  bear  in  mind  that  sbb,rel  and  swb,rel  are  estimates  of  the  true  standard  deviations 
and  therefore  subject  to  random  fluctuations.  Therefore,  the  mean  square  between  groups 
(MSbetween)  can  be  smaller  than  the  mean  squares  within  groups  (MSwithin),  resulting  in 
negative  arguments  under  the  square  root  used  for  the  estimation  of  the  between-unit 
variation,  whereas  the  true  variation  cannot  be  lower  than  zero.  In  this  case,  u*
bb,  the 
maximum  inhomogeneity  that  could  be  hidden  by  method  repeatability,  was  calculated  as 
described by Linsinger et al. [11]. u*
bb is comparable to the limit of detection of an analytical 
method,  yielding  the  maximum  inhomogeneity  that  might  be  undetected by  the  given  study 
setup.  
Method  repeatability  (swb,rel),  between–unit  standard  deviation  (sbb,rel)  and  u*
calculated as:  

bb,rel  were 

s

,wb

rel

=

MS

within

y 

s

,bb

rel

=

u

*
,bb

rel

=

MS

between

MS

within

n
y

MS
within
n

4

ν

2

MSwithin

y

Equation 1 

Equation 2 

Equation 3 

MSwithin 

mean square within a unit from an ANOVA  

MSbetween  mean squares between-unit from an ANOVA 

y  

n 

ν

MSwithin

mean of all results of the homogeneity study 

mean number of replicates per unit 

degrees of freedom of MSwithin  

The results  of the  evaluation  of the  between-unit  variation  are  summarised  in  Table  2. The 
resulting value from the above equation was converted into a relative uncertainty.  

Table 2: Results of the homogeneity study for density. 

Measurand  

Density 

swb,rel  

sbb,rel  

[%] 

0.0013 

[%] 

n.c. 

u*

bb,rel 

[%] 

urec,rel 

[%] 

ubb,rel 

[%] 

0.0004 

n.a. 

0.0004 

1) n.c.: cannot be calculated as MSbetween < MSwithin 
2) n.a.: not applicable 

14 

 
-
 
 
 
 
 
The homogeneity study showed no outlying unit means or trends in the filling sequence. The 
u*

bb is adopted as uncertainty contribution to account for potential inhomogeneity. 

Selenium 

The  homogeneity  was  evaluated  by  using  the  data  of  the  short  term  stability  study.  The 
measurements were performed under repeatability conditions, and in a randomised manner 
to  be  able  to  separate  a  potential  analytical  drift  from  a  trend  in  the  filling  sequence.  The 
results are shown as graphs in Annex A.  

Regression analyses were performed to evaluate potential trends in the analytical sequence 
as well as trends in the filling sequence. Some significant (95 % confidence level) trends in 
the  analytical  sequence  were  visible  for  selenium,  pointing  at  a  signal  drift  in  the  analytical 
system.  Correction  of  trends  is  therefore  expected  to  improve  the  sensitivity  of  the 
subsequent  statistical  analysis  through  a  reduction  in  analytical  variation  without  masking 
potential  between-unit  heterogeneities.  As  the  analytical  sequence  and  the  unit  numbers 
were not correlated, trends significant on at least a 95 % confidence level were corrected as 
shown below:  

corrected

result

=

measured

result

ib (cid:215)

Equation 4 

b = slope of the linear regression 

i = position of the result in the analytical sequence 

The  trend-corrected  dataset  was  tested  for  consistency  using  Grubbs  outlier  tests  on  a 
confidence level of 99 % on the individual results and the unit means. No outlying individual 
results and outlying unit means were detected.  

Quantification  of  between-unit  inhomogeneity  was  again  accomplished  using  ANOVA  to 
separate  the  between-unit  variation  (sbb)  from  the  within-unit  variation  (swb).  The  statistical 
distribution was checked for normality as described above for the study on density.  
Method  repeatability  (swb,rel),  between–unit  standard  deviation  (sbb,rel)  and  u*
calculated as using the equations 1, 2 and 3. 

bb,rel  were 

The results  of the  evaluation  of the  between-unit  variation  are  summarised  in  Table  3. The 
resulting value from the equation 3 was converted into a relative uncertainty.  

Table 3: Results of the homogeneity study for selenium. 

Measurand  

Selenium 

swb,rel  

sbb,rel  

u*

bb,rel 

urec,rel 

ubb,rel 

[%] 

14.5 

[%] 

n.c. 

[%] 

4.3 

[%] 

n.a. 

[%] 

4.3 

1) n.c.: cannot be calculated as MSbetween < MSwithin 
2) n.a.: not applicable 

The homogeneity study showed no outlying unit means or trends in the filling sequence. The 
u*

bb is adopted as uncertainty contribution to account for potential inhomogeneity. 

Other trace elements 

For all trace elements other than selenium, a two-way analysis of variance without replication 
was  used  because  not  all  samples  could  be  measured  on  the  same  occasion.  Two-way 
analysis  allowed  us 
the  within-  and  between-unit  standard  deviations 
independently of the between-run effect.  

to  estimate 

The data evaluation was performed in the following order: 

15 

 
 
-
   
  
 
 
 
1) Regression analyses to evaluate potential trends in each analytical run. Some significant 
(95 % confidence level) trends in the analytical sequence were visible for copper in run 1, for 
manganese in run 2 and for chromium and manganese in run 3, pointing at a signal drift in 
the analytical system. 

2) Correction of dataset for significant analytical trend (95% confidence level). The correction 
of  biases,  even  if  they  are  statistically  not  significant,  was  found  to  combine  the  smallest 
uncertainty  with  the  highest  probability  to  cover  the  true  value  [12].  Correction  of  trends  is 
therefore expected to improve the sensitivity of the subsequent statistical analysis through a 
reduction  in  analytical  variation  without  masking  potential  between-unit  heterogeneities.  As 
the  analytical  sequence  and  the  unit  numbers  were  not  correlated,  trends  significant  on  at 
least a 95 % confidence level were corrected as shown below:  

),(x T
ir

=

),(x
ir

rb

)(

i

Equation 5 

i  

r  

b(r)  

x(r,i)  

xT(r,i)  

position of the result in the analytical run 

number of the analytical run from 1 to 3 

slope of the linear regression for the analytical run r 

measurement results on the position i in the analytical run r 

corrected results for analytical trend on the position i in the analytical run r 

3) The analytical trend-corrected dataset was evaluated for statistically significant difference 
between  analytical  runs  (95  %  confidence  level)  using  one  way  ANOVA.  A  statistically 
significant  difference  between  analytical  runs  was  observed  for  all  analytes  on  95  % 
confidence level. 

4) Normalisation of data showing statistically significant difference between analytical run (95 
%  confidence  level).  As  it  is  assumed  that  run-effects  and  unit-effects  are  independent, 
differences  between  analytical  runs  on  at  least  a  95  %  confidence  level  were  corrected  as 
shown below:  

irx
),(
R

=

ir
),(x
T
)(
rx
T

i  

r  

position of the result in the analytical run 

number of the analytical run from 1 to 3 

Equation 6 

)(r

xT
sequence (if necessary) 

mean  results  of  the  analytical  run  r  after  correction  for  trend  in  analytical 

xT(r,i)  

xR(r,i)  

corrected results for analytical trend on the position i in the analytical run r 

normalised results on the position i in the analytical run r 

5)  The  normalised  dataset  was  tested  for  consistency  using  Grubbs  outlier  tests  on  a 
confidence  level  of  99  %  on  the  individual  results  and  the  unit  means.  The  trend-corrected 
dataset  was  used  to  evaluate  significant  trends  in  the  production  sequence.  Outliers  were 
found for  Cu  and  Fe, for  the  remaining  elements  no  outlying  individual  results  and  outlying 
unit  means  were  detected  at  the  99  %  confidence  level  (See  Table  4).  Since  no  technical 
reason for the outliers could be found, all the data were retained for statistical analysis. 

16 

(cid:215)
-
 
 
 
 
 
Table  4:  Results  of  the  statistical  evaluation  of  the  homogeneity  studies  at  95  % 
confidence level 

Measurand 

Arsenic 

Trends 
Analytical 
sequence 
no 

Filling 
sequence 
no 

Outliers 
Individual 
results 
none 

Unit 
means 
none 

Cadmium 

no 

Chromium 

no 

Cobalt 

Copper 

Iron 

Lead 

no 

yes 

yes 

yes 

Manganese 

yes 

Molybdenum  yes 

Nickel 

Zinc 

no 

no 

no 

no 

yes 

yes 

no 

yes 

no 

no 

no 

no 

none 

none 

none 

none 

none 

none 

1 

none 

1 

2 

none 

none 

none 

none 

none 

none 

none 

none 

none 

none 

Distribution 
Individual 
results 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 

Unit means 

normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 
normal/uni-
modal 

6) Quantification of between-unit inhomogeneity was accomplished using the analytical trend 
corrected dataset by two way ANOVA, which can separate the between-run variation (sr), the 
between-unit variation (sbb) and the within-unit variation (swb). The latter is equivalent to the 
method repeatability if the individual samples are representative of the whole unit.  

Evaluation by ANOVA requires unit means which follow at least a uni-modal distribution and 
results for each unit that follow uni-modal distributions with approximately the same standard 
deviations.  Distribution  of  the  unit  means  was  visually  tested  using  histograms  and  normal 
probability  plots.  Minor  deviations  from  uni-modality  of  the  individual  values  were found  but 
do not significantly affect the estimate of between-unit standard deviations (See Table 4).  

Recall that sbb,rel and swb,rel are estimates of the true standard deviations and therefore subject 
to  random  fluctuations.  Therefore,  the  mean  square  between  groups  (MSbetween)  can  be 
smaller  than  the  mean  squares  within  groups  (MSwithin),  resulting  in  negative  arguments 
under the square root used for the estimation of the between-unit variation, whereas the true 
variation cannot be lower than zero. In this case, u*
bb, the maximum inhomogeneity that could 
be hidden by method repeatability, was calculated as described by Linsinger et al. [11]. u*
bb is 
comparable  to  the  limit  of  detection  of  an  analytical  method,  yielding  the  maximum 
inhomogeneity that might be undetected by the given study setup.  

Analysis of variance applied to a randomised block design with one observation per unit per 
run  leads  to  a  between-run  mean  square  MSr  together  with  a  between-unit  mean  square 
MSbetween,  and  a  residual  mean  square  MSwithin.  The  analysis  of  variance  table  also  includes 
associated  degrees  of  freedom  for  each  term.  The  residual  mean  square  MSwithin  is  an 
rs . The results of two way ANOVA are given 
unbiased estimate of the repeatability variance 
in Table 5. 

2

17 

 
 
Table 5: Two way ANOVA results. The two way ANOVA was performed on the results 
corrected only for analytical trends. 

Measurand 
Arsenic 

Iron 

Cobalt 

Copper 

Cadmium 

Between run variation 
Between unit variation 
Within unit variation 
Between run variation 
Between unit variation 
Within unit variation 
Chromium  Between run variation 
Between unit variation 
Within unit variation 
Between run variation 
Between unit variation 
Within unit variation 
Between run variation 
Between unit variation 
Within unit variation 
Between run variation 
Between unit variation 
Within unit variation 
Between run variation 
Between unit variation 
Within unit variation 
Manganese  Between run variation 
Between unit variation 
Within unit variation 
Molybdenum  Between run variation 
Between unit variation 
Within unit variation 
Between run variation 
Between unit variation 
Within unit variation 
Between run variation 
Between unit variation 
Within unit variation 

Nickel 

Lead 

Zinc 

Degrees of freedom  Mean square [(µg/L)]  

2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 
2 
13 
26 

MSr = 1.9 10-2 
MSBetween = 4.2 10-3 
MSwithin = 4.5 10-3 
MSr = 3.0 10-5 
MSBetween = 1.7 10-5 
MSwithin = 1.8 10-5 
MSr = 3.0 10-4 
MSBetween = 1.0 10-6 
MSwithin = 9.9 10-5 
MSr = 7.14 10-6 
MSBetween = 2.4 10-5 
MSwithin = 3.0 10-5 
MSr = 3.16 10-2 
MSBetween = 2.6 10-3 
MSwithin = 1.11 10-3 
MSr = 1.02 10-1 
MSBetween = 1.4 10-2 
MSwithin = 6.5 10-3 
MSr = 1.9 10-6 
MSBetween = 3.3 10-5 
MSwithin = 1.6 10-5 
MSr = 7.4 10-2 
MSBetween = 3.9 10-3 
MSwithin = 4.9 10-3 
MSr = 1.8 10-1 
MSBetween = 7.2 10-3 
MSwithin = 1.3 10-2 
MSr = 8.6 10-4 
MSBetween = 6.9 10-4 
MSwithin = 1.5 10-5 
MSr = 7.6 10-3 
MSBetween = 2.1 10-2 
MSwithin = 4.5 10-2 

With the exception of copper and lead, method repeatability (swb,rel), between–unit standard 
deviation (sbb,rel) and u*

bb,rel were calculated as equations 1, 2 and 3 indicate.  

However, a different approach was adopted for copper and iron for which outlying unit means 
were  detected.  In  this  case  between-unit  inhomogeneity  was  modelled  as  a  rectangular 
distribution limited by the largest outlying unit mean, and the rectangular standard uncertainty 
of homogeneity was estimated by: 

=

urec

outlier

y

3

y

Equation 7 

y  

mean of all results of the homogeneity study 

It  should  be  mentioned  that  the  outlying  unit  means  are  a  result  of  presence  of  outlying 
individual values and do not necessarily reflect the real distribution of these elements in the 
material.  

18 

 
 
(cid:215)
-
 
When  a  trend  in  the  filling  sequence  was  significant  at  least  at  99  %  confidence  level,  the 
uncertainty was assessed in a different way. This applies for lead. Here, urec was estimated 
using  a  rectangular  distribution  between  the  highest  and  lowest  unit  mean.  The  corrected 
uncertainty  in  those  cases  where  there  was  a  significant  trend  in  the  filling  sequence  is 
obtained by using equation 8: 

=

urec

highest re

sult - low

est result

2

3

y 

Equation 8 

The results  of the  evaluation  of the  between-unit  variation  are  summarised  in  Table  6. The 
resulting values from the above equations were converted into relative uncertainties.  

Table 6: Results of the homogeneity study 

Measurand  

Arsenic 

Cadmium 

Chromium 

Cobalt 

Copper 

Iron 

Lead 

Manganese 

Molybdenum 

Nickel 

Zinc 

swb,rel  

sbb,rel  

u*

bb,rel 

urec,rel 

ubb,rel 

[%] 

3.50 

3.70 

3.93 

6.21 

3.49 

2.87 

4.39 

2.69 

0.93 

3.09 

4.89 

[%] 

n.c. 

n.c. 

0.21 

n.c. 

2.35 

1.74 

2.60 

n.c. 

n.c. 

n.c. 

n.c. 

[%] 

1.06 

1.12 

1.20 

1.89 

1.06 

0.87 

1.34 

0.82 

0.28 

0.94 

1.49 

[%] 

n.a. 

n.a. 

n.a. 

n.a. 

5.72 

3.14 

2.96 

n.a. 

n.a. 

n.a. 

n.a. 

[%] 

1.06 

1.12 

1.20 

1.89 

5.72 

3.14 

2.96 

0.82 

0.28 

0.94 

1.49 

1) n.c.: cannot be calculated as MSbetween < MSwithin 
2) n.a.: not applicable 

With the exception of copper, iron and lead, the homogeneity study showed no outlying unit 
means  or trends  in  the filling  sequence. Therefore  the  between-unit  standard  deviation  can 
be used as estimate of ubb. As u*
bb sets the limits of the study to detect inhomogeneity, the 
larger  value  of  sbb  and  u*
bb  is  adopted  as  uncertainty  contribution  to  account  for  potential 
inhomogeneity. Outlying unit means were found for copper and iron, and a trend with filling 
was  found  for  lead.  However,  the  inhomogeneities  quantified  as  urec  were  still  sufficiently 
small to make the material suitable for its intended use. Therefore, urec was used as estimate 
of ubb for these elements. 

4.2  Within-unit homogeneity and minimum sample intake 

The within-unit homogeneity is closely correlated to the minimum sample intake. Due to this 
correlation, individual aliquots of a material will not contain the same amount of analyte. The 
minimum  sample  intake  is  the  minimum  amount  of  sample  that  is  representative  for  the 
whole unit and thus can be used in an analysis. Sample sizes equal or above the minimum 
sample intake guarantee the certified value within its stated uncertainty.  

ERM-CA403 is a true solution and is not expected to have any relevant heterogeneity. This 
assumption  was  confirmed  by  the  characterisation  study,  where  sample  intakes  as  low  as 

19 

 
(cid:215)
(cid:215)
 
 
 
 
2 mL were found to give acceptable repeatability for both density and element concentration 
measurement.  This  demonstrates  that  there  is  no  intrinsic  inhomogeneity  or  contamination 
when using sample intakes at this level. 

5  Stability 

Time, temperature and light were regarded as the most relevant influences on stability of the 
material. The influence of ultraviolet or visible radiation was minimised by the choice of the 
container  and  packaging  which  eliminates  most  of  the  incoming  light.  In  addition,  materials 
are  stored  and  dispatched  in  the  dark,  thus  eliminating  practically  the  possibility  of 
degradation  by  light.  Therefore,  only  the  influences  of  time  and  temperature  needed  to  be 
investigated. 

Stability testing is necessary to establish conditions for storage (long-term stability) as well as 
conditions for dispatch to the customers (short-term stability). During transport, especially in 
summer  time,  temperatures  up  to  60  °C  could  be  reached  and  stability  under  these 
conditions must be demonstrated if transport at ambient temperature will be applied. 

The  stability  studies  were  carried  out  using  an  isochronous  design  [13].  In  that  approach, 
samples  are  stored  for  a  certain  time  at  different  temperature  conditions.  Afterwards,  the 
samples are moved to conditions where further degradation can be assumed to be negligible 
(reference  conditions).  At  the  end  of  the  isochronous  storage,  the  samples  are  analysed 
simultaneously  under  repeatability  conditions.  Analysis  of  the  material  (after  various 
exposure  times  and  temperatures)  under  repeatability  conditions  greatly  improves  the 
sensitivity of the stability tests.  

5.1  Short-term stability study 

For  the  short-term  stability  study,  units  were  stored  at  18  °C  and  60  °C  for  0,  1,  2  and  4 
weeks  (at  each  temperature).  The  reference  temperature  was  set  to  4  °C.  Two  units  per 
storage  time  were  selected  using  a  random  stratified  sampling  scheme.  From  each  unit, 
three  samples  were  measured  by  a  high  performance  calibrated  density  meter  (density), 
ICP-MS  (selenium)  and  by  ICP-SFMS  (all  other  trace  elements).  The  measurements  were 
performed under repeatability conditions for density and selenium, whereas for the remaining 
trace  elements  were  performed  in  a  randomised  block  design  because  the  number  of 
replicates/analytes  on  all  units  cannot  be  included  in  a  single  run  due  to  instrumental 
constraints  (drift  towards  the  end  of  a  long  run).  In  all  the  cases,  the  measurements  were 
performed in a randomised sequence to be able to separate a potential analytical drift from a 
trend over storage time.  

Density 

Samples  were  measured  using  high  performance  calibrated  density  meter.  The 
measurements  were  made  in  accordance  with  ASTM  D4052-09  [14]  which  is  the  standard 
method for Density, Relative Density and API Gravity of Liquids by Digital Density Meter. The 
results of the measurements are shown in Annex B. 

The  obtained  data  were  evaluated  individually  for  each  temperature.  The  results  were 
screened  for  outliers  using  the  single  and  double  Grubbs  test.  One  outlier  was  found  at 
60 ºC. As no technical reason for the outlier could be found the data point was retained for 
statistical analysis.  

Furthermore,  the  data  were  evaluated  against  storage  time  and  regression  lines  of  density 
versus  time  were  calculated.  The  slope  of  the  regression  line  was  tested  for  statistical 
significance (loss/increase due to shipping conditions). The slope of the regression line was 
not significantly different from zero (on 99 % confidence level) at both 18 °C and 60 °C.  

20 

None  of  the  trends  was  statistically  significant  on  a  99  %  confidence  level  for  any  of  the 
temperatures. 

Selenium 

The  measurements  were  performed  under  repeatability  conditions,  and  in  a  randomised 
manner.  Regression  analyses  were  performed  to  evaluate  potential  trends  in  the  analytical 
sequence. Some significant (95 % confidence level) trends in the analytical sequence were 
found for selenium, pointing at a signal drift in the analytical system. Correction of trends is 
therefore expected to improve the sensitivity of the subsequent statistical analysis through a 
reduction in analytical variation without masking potential effect of the time and temperature. 
As the analytical sequence and the unit numbers were not correlated, trends significant on at 
least a 95 % confidence level were corrected as shown below:  

corrected

result

=

measured

result

ib (cid:215)

Equation 9 

b = slope of the linear regression 

i = position of the result in the analytical sequence 

The  obtained  trend-corrected  data  were  evaluated  individually  for  each  temperature.  The 
results were screened for outliers using the single and double Grubbs test. One outlier was 
found  at  18  ºC.  As  no  technical  reason  for  the  outlier  could  be  found  the  data  point  was 
retained for statistical analysis.  

Furthermore,  the  data  were  evaluated  against  storage  time  and  regression  lines  of 
concentration versus time were calculated. The slopes of the regression lines were tested for 
statistical significance (loss/increase due to shipping conditions). For selenium, the slopes of 
the  regression  lines  were  not  significantly  different from  zero  (on  99 %  confidence  level)  at 
both 18 °C and 60 °C.  

The results of the measurements are shown in Annex B. A statistical outlier was detected for 
selenium, and this was retained for the estimation of usts. None of the trends was statistically 
significant on a 99 % confidence level for any of the temperatures.  

Other trace elements 

The measurements for the remaining trace elements were performed in a randomised block 
design because the number of replicates/analytes on all units cannot be included in a single 
run due to instrumental constraints (drift towards the end of a long run). In that circumstance 
better  precision  (measured  as  the  within-unit  standard  deviation)  can  be  obtained  using 
several short runs in a randomised block design than the one obtained in a single run. In a 
randomised  block  design  for  three  replicates  on  each  of  14  units  of  ERM-CA403,  the 
simplest  randomised  block  design  involves  three  measurement  runs  and  each  unit  is 
measured once in random order. Runs were randomised individually. The results are shown 
as graphs in Annex B. 

The  obtained  data  were  evaluated  individually  for  each  temperature.  The  data  evaluation 
was performed in the following order: 

1) Regression analyses to evaluate potential trends in each analytical run. Some significant 
(95 % confidence level) trends in the analytical sequence were visible for copper in run 1, for 
manganese in run 2 and for chromium and manganese in run 3, pointing at a signal drift in 
the measurement system. 

2) Correction of dataset for significant analytical trend (95% confidence level). The correction 
of  biases,  even  if  they  are  statistically  not  significant,  was  found  to  combine  the  smallest 
uncertainty  with  the  highest  probability  to  cover  the  true  value  [12].  Correction  of  trends  is 
therefore expected to improve the sensitivity of the subsequent statistical analysis through a 

21 

 
 
-
   
  
 
 
reduction in analytical variation without masking potential change over time. As the analytical 
sequence  and  the  unit  numbers  were  not  correlated,  trends  significant  on  at  least  a  95  % 
confidence  level  were  corrected  as  for  the  homogeneity  study,  using  equation  5  (Section 
4.1). 

3)  The  analytical  trend-corrected  dataset  was  evaluated  for  significant  difference  between 
analytical  runs  (99  %  confidence  level)  using  one  way  ANOVA.  A  significant  difference 
between analytical runs was observed on 99 % confidence level. 

4)  Normalisation  of  data  showing  significant  difference  between  analytical  run  (95  % 
confidence  level).  As  it  is  assumed  that  run  effects  and  unit  effects  are  independent, 
difference between analytical runs on at least a 99 % confidence level was corrected as for 
the homogeneity study, using equation 6 (Section 4.1).  

5)  The  normalised  dataset  was  tested  for  consistency  using  Grubbs  outlier  tests  on  a 
confidence level of 99 % on the individual results and the unit means. Outliers were found for 
some  at  the  99  %  confidence  level  (See  Table).  Since  no  technical  reason  for  the  outliers 
could be found, all the data were retained for statistical analysis. 

6)  The  normalised  obtained  data  were  evaluated  individually  for  each  temperature. 
Furthermore,  the  data  were  evaluated  against  storage  time  and  regression  lines  of 
concentration versus time were calculated. The slopes of the regression lines were tested for 
statistical significance (loss/increase due to shipping conditions). For all elements, the slopes 
of the regression lines were not significantly different from zero (on 95 % confidence level) at 
both 18 °C and 60 °C.  

The  results  of  the  measurements  are  shown  in  Annex  B.  The  results  of  the  statistical 
evaluation of the short-term stability are summarised in Table 7.  

Table 7: Results of the short-term stability tests 

Measurand 

Arsenic 

Cadmium 

Chromium 

Cobalt 

Copper 

Iron 

Lead 

Manganese 

Molybdenum 

Nickel 

Zinc 

Number  of  individual 
outlying results  

Significance  of  the  trend 
on  a  95  %  confidence 
level 

18 ºC 
1 

2 

0 

0 

1 

0 

0 

0 

2 

0 

0 

60 ºC 

18 ºC 

60 ºC 

0 

1 

0 

0 

1 

2 

0 

0 

0 

0 

0 

no 

no 

no 

no 

no 

yes 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

Statistical  outliers  were  detected  for  some  of  the  analytes,  and  these  were  retained  for  the 
estimation of usts. None of the trends were statistically significant on a 99 % confidence level 
for any of the temperatures. However, the study on Fe at 18 ºC showed a decrease with time 
significant  at  the  95 %  confidence  level.  As  this  trend  was  not  seen  at  the  higher  test 
temperature, and measurement variance was similar for both studies, the trend is considered 
to be a statistical artefact and not due to real degradation of Fe content. 

22 

 
Summary of short term stability: 

Taken  into  account  the  results  for  all  the  analytes  and  the  density,  the  material  can  be 
transported at ambient conditions without special conditions. 

5.2  Long-term stability study 

For  the  long-term  stability  study,  samples  were  stored  at  18  °C  for  0,  4,  8,  12,  16  and  24 
months.  The  reference  temperature  was  set  to  4  °C.  Two  samples  per  storage  time  were 
selected using a random stratified sampling scheme. From each sample, six replicates were 
measured  by  a  high  performance  calibrated  density  meter  (density)  and  by  ICP-SFMS  (all 
trace  elements).  The  measurements  were  performed  under  repeatability  conditions,  in  a 
random  sequence  to  be  able  to  separate  any  potential  analytical  drift  from  a  trend  over 
storage time.  

Regression analyses were performed to evaluate potential trends in the analytical sequence. 
There  were  no  statistically  significant  trends  in  the  filling  sequence  or  the  analytical 
sequence. 

The  obtained  data  were  evaluated.  The  results  were  screened  for  outliers  using  the  single 
and double Grubbs test. Some outlying individual results were found for density (Table). As 
no  technical  reason  for  the  outliers  could  be  found  all  data  were  retained  for  statistical 
analysis.  

Furthermore,  the  data  were  plotted  against  storage  time  and  linear  regression  lines  of 
concentration versus time were calculated. The slope of the regression lines was tested for 
statistical significance (loss/increase due to storage conditions). For all elements, the slopes 
of the regression lines were not significantly different from zero (on 99 % confidence level) at 
18 °C.  

The results of the long term stability measurements are shown in Annex C. The results of the 
statistical evaluation of the long-term stability study are summarised in Table 8.  

Table 8: Results of the long-term stability tests 

Measurand  Number  of  individual  outlying 

results  

Significance  of 
confidence level 

the 

trend  on  a  95 % 

Density 

Arsenic 

Cadmium 

Chromium 

Cobalt 

Copper 

Iron 

Lead 

Manganese 

Molybdenum 

Nickel 

Selenium 

Zinc 

18 ºC 
1 

0 

0 

0 

0 

0 

0 

0 

1 

1 

0 

0 

0 

23 

18 ºC 

no 

no 

yes 

no 

no 

no 

no 

no 

no 

no 

no 

no 

no 

 
 
The study on Cd showed an increase with time significant at the 95 % confidence level. Such 
an effect could only be possible if the matrix degraded or evaporated. However, as this trend 
was not seen for any other element, it is considered to be a statistical artefact and not due to 
real degradation of the sample. The material can therefore be stored at 18 °C. 

5.3  Estimation of uncertainties 

Due  to  the  intrinsic  variation  of  measurement  results,  no  study  can  rule  out  degradation  of 
materials  completely,  even  in  the  absence  of  statistically  significant  trends.  It  is  therefore 
necessary  to  quantify  the  potential  degradation  that  could  be  hidden  by  the  method 
repeatability,  i.e.  to  estimate  the  uncertainty  of  stability.  This  means,  even  under  ideal 
conditions, the outcome of a stability study can only be "degradation is 0 ± x % per time".  

Uncertainties of stability during dispatch and storage were estimated as described in [15] for 
the concentration of each element and for density. For this approach, the uncertainty of the 
linear regression line with a slope of zero is calculated. The uncertainty contributions usts and 
ults are calculated as the product of the chosen transport time/shelf life and the uncertainty of 
the regression lines as: 

u

sts

,
rel

=

∑

RSD
(

x

i

u

lts

,
rel

=

∑

RSD
(

x

i

t

tt

t

sl

)

2

x

)

2

x

Equation 12 

Equation 13 

RSD  

relative standard deviation of all results of the stability study 

xi 

x  

ttt 

tsl 

result at time point i 

mean results for all time points  

chosen transport time (1 week at 60 ºC) 

chosen shelf life (36 months at 18 ºC) 

The following uncertainties were estimated: 

-  usts,rel, the uncertainty of degradation during dispatch. This was estimated from the 60 
°C studies. The uncertainty describes the possible change during a dispatch at 60 °C 
lasting for one week. 

-  ults,rel,  the  stability  during  storage.  This  uncertainty  contribution  was  estimated  from 
the  18  °C  study.  The  uncertainty  contribution  describes  the  possible  degradation 
during 36 months storage at 18 °C.  

The results of these evaluations are summarised in Table 9. 

24 

 
(cid:215)
-
 
(cid:215)
-
 
 
 
 
Table 9: Uncertainties of stability during dispatch and storage. usts,rel was calculated for 
a temperature of 60 °C and 1 week; ults,rel was calculated for a storage temperature of 
18 °C and 3 years 

Measurand 

Density 

Arsenic 

Cadmium 

Chromium 

Cobalt 

Copper 

Iron 

Lead 

Manganese 

Molybdenum 

Nickel 

Selenium 

Zinc 

usts ,rel 
[%] 

At 18 °C 

0.00023 

At 60 °C 

0.0030 

ults,rel 
[%] 

At 18 °C 

0.0022 

0.41 

0.51 

0.44 

0.85 

0.65 

0.48 

0.63 

0.32 

0.085 

0.35 

1.81 

0.60 

0.57 

0.42 

0.57 

0.77 

0.69 

0.45 

0.74 

0.40 

0.14 

0.43 

1.85 

0.62 

2.02 

2.07 

1.88 

4.80 

1.76 

6.91 

2.19 

1.51 

1.54 

6.29 

8.46 

4.74 

After the certification campaign, the material will be subjected to a regular stability monitoring 
programme to control its further stability. 

6  Characterisation  

The material characterisation is the process of determining the property values of a reference 
material. 

The  material  characterisation  was  based  on  an  intercomparison  of  expert  laboratories,  i.e. 
the properties of the material were determined in different laboratories that applied different 
measurement  procedures  to  demonstrate  the  absence  of  a  measurement  bias.  This 
approach aims at randomisation of laboratory bias, which reduces the combined uncertainty. 

6.1  Selection of participants 

Laboratories were selected based on criteria that comprised both technical competence and 
quality management aspects. For density measurements 10 laboratories were selected and 
for  trace  element  measurement,  9  laboratories.  Each  participant  was  required  to  operate  a 
quality system and to deliver documented evidence of its laboratory proficiency in the field of 
density determination in relevant matrices by submitting results for intercomparison exercises 
or method validation reports. Having a formal accreditation was not mandatory, but meeting 
the  requirements  of  ISO/IEC  17025  was  obligatory.  Where  measurements  are  covered  by 
the  scope  of  accreditation,  the  accreditation  number  is  stated  in  the  list  of  participants 
(Section 2). 

25 

 
 
6.2  Study setup  

Each  laboratory  received  two  units  of  ERM-CA403  and  was  requested  to  provide  six 
independent  results,  three  replicates  per  unit.  The  units  for  material  characterisation  were 
selected  using  a  random  stratified  sampling  scheme  and  covered  the  whole  batch.  The 
sample  preparations  (if  necessary)  and  measurements  had  to  be  spread  over  at  least  two 
days to ensure intermediate precision conditions. An independent calibration was performed 
for each independent measurement. 

For  the  trace  elements,  each  participant  received  a  sample  of  NASS-6  (NRC,  National 
Research Council Canada, Canada) as a blinded quality control (QC) sample. The results for 
this sample were used to support the evaluation of the characterisation results. 

Laboratories  were  also  requested  to  give  estimations  of  the  expanded  uncertainties  of  the 
mean value of the six results. No approach for the estimation was prescribed, i.e. top-down 
and bottom-up were regarded as equally valid procedures. 

6.3  Methods used  

Density 

Two  different  methods  without  sample  preparation  were  used  to  characterise  the  material. 
The combination of results from methods based on completely different principles mitigates 
undetected method bias. 

All  methods  used  during  the  characterisation  study  are  summarised  in  Annex  D.  The 
laboratory  code  (e.g.  L01)  is  a  random  number  and  does  not  correspond  to  the  order  of 
laboratories  in  Section  2.  The  lab-method  code  consists  of  a  number  assigned  to  each 
laboratory  (e.g.  L01)  and  abbreviation  of  the  measurement  method  used  (e.g.  L01-OD  for 
oscillating type density meter or L09-Pyc for the determination with pycnometer). 

Trace elements 

All laboratories used measurement methods based on ICP-MS, except for Se for which one 
laboratory used AFS. 

As seen in Annex D, laboratories used different approaches to sample preparation, including 
dilution  or  pre-concentration.  Each  participating  laboratory  reported  that  their  method  was 
validated. Results from any validated method based on ICP-MS should be equivalent to the 
results used in characterisation of the material. 

All  methods  used  during  the  characterisation  study  are  summarised  in  Annex  D.  The 
laboratory  code  (e.g.  L01)  is  a  random  number  and  does  not  correspond  to  the  order  of 
laboratories in Section 2.  

6.4  Evaluation of results 

The  characterisation  campaign  resulted  in  10  datasets  for  density  and  3  to  8  datasets  for 
element  concentrations.  All  individual  results  of  the  participants,  grouped  per  analyte  are 
displayed in tabular and graphical form in Annex E.  

6.4.1  Technical evaluation 
Density 

The  obtained  data  were  first  checked  for  compliance  with  the  requested  analysis  protocol 
and  for  their  validity  based  on  technical  reasons.  The  following  criteria  were  considered 
during the evaluation:  

-  appropriate validation of the measurement procedure 

- 

compliance  with  the  analysis  protocol:  measurements  performed  on  two  days,  and 
the analytical sequence determination. 

26 

-  measurement repeatability < 10%. 

One  dataset  was  rejected  as  not  technically  valid.  Results  provided  by  the  laboratory  L07 
showed variance over 10 times higher than the mean variance of the other laboratories. This 
indicated that the results were not of the same quality as those from the other labs, and the 
dataset was not used in the evaluation. 

Trace elements 

The  obtained  data  were  first  checked  for  compliance  with  the  requested  analysis  protocol 
and  for  their  validity  based  on  technical  reasons.  The  following  criteria  were  considered 
during the evaluation:  

-  appropriate validation of the measurement procedure 

- 

compliance  with  the  analysis  protocol:  sample  preparations  and  measurements 
performed on two days, and the analytical sequence and water content determination 

-  no individual values given as below limit of detection or below limit of quantification 

-  agreement of the measurement results with the assigned value of the QC sample 

-  measurement repeatability < 50% 

- 

results  for  Cd,  Cr,  Ni  and  Zn  equal  to  or  higher  than  the  levels  at  which  they  were 
known to be present following spiking of the seawater for CA403  

The QC sample was a CRM, NASS-6, which was re-bottled and supplied to participants as a 
blind  sample.  The  assigned  values  were  those  certified,  and  the  associated  uncertainties 
were  set  taking  into  account  the  lower  analyte  concentrations  of  certain  elements  in  this 
material.  For  As,  Cr  and  Mo,  which  were  present  in  similar  concentration,  the  certificate 
uncertainty values were assigned. For Cd, Cu, Mn and Ni with concentrations of less than a 
third  of  those  in  CA403 and for  Co, for  which  the  value  was  not  certified,  50 %  uncertainty 
was assigned. For Fe, Pb and Zn with concentrations less than a sixth of those in CA403 and 
for Se, for which no value was available, no QC test was made. 

Based on the above criteria, the following results were rejected as not technically valid: 

L03:  The  laboratory's  complete  data  set  was  not  considered  when  calculating  the  certified 
values  for  the  material,  as  the  reported  results  for  Cd,  Ni  and  Zn  were  below  the 
concentration added by spiking and QC test results for 3 of 8 elements did not agree. This 
strongly indicates poor performance in comparison to the other participating laboratories. 

L05: The results for Co were rejected because measurement of the QC sample did not agree 
with  the  assigned  value.  In  addition,  results  for  Fe,  Se,  Cr  and  Zn  were  not  used  as  they 
were reported as below their LOQ. 

L06: The results for As, Cd, Cr and Cu were rejected as some individual values were below 
the method's respective LOQ. 

L12: The results for Mn were rejected because measurement of the QC sample did not agree 
with the assigned value. 

6.4.2  Statistical evaluation 
Density 

The  datasets  accepted  based  on  technical  reasons  were  tested  for  normality  of  dataset 
means  using  kurtosis/skewness  tests  and  normal  probability  plots  and  were  tested  for 
outlying  means  using  the  Grubbs  test  and  using  the  Cochran  test  for  outlying  standard 
deviations (both at a 99 % confidence level). Standard deviations within (swithin) and between 
(sbetween)  laboratories  were  calculated  using  one-way  ANOVA.  The  results  of  these 
evaluations are shown in Table 10. 

27 

 
Table 10: Statistical evaluation of the technically accepted datasets for ERM-CA403. p: 
number of technically valid datasets 

Analyte  p 

Outliers 

Means  Variances 

Normally 
distributed 

Density  9 

no 

yes 

yes 

Statistical parameters  

Mean 
swithin 
[g/mL] 
[g/mL] 
1.02352  0.00003  0.00002  0.00002 

sbetween 
[g/mL] 

s 
[g/mL] 

The laboratory means follow normal distributions. None of the data contains outlying means, 
but some outlying variances were found. The outlying variances are not method dependent 
and therefore merely reflect the fact that both methods have different intrinsic variability. As 
all  measurement  procedures  were  found  technically  valid,  all  results  were  retained. 
Moreover, closer scrutiny of the data shows that the standard deviations on the mean density 
reported  by  the  laboratories  are  slightly  higher  than  0.1  %.  The  datasets  are  therefore 
consistent  and  the  mean  of  laboratory  means  is  a  good  estimate  of  the  true  value.  The 
uncertainty related to the characterisation is estimated as the standard error of the mean of 
laboratory means (Table). 

Trace elements 

The  datasets  accepted  based  on  technical  reasons  were  tested  for  normality  of  dataset 
means  using  kurtosis/skewness  tests  and  normal  probability  plots  and  were  tested  for 
outlying  means  using  the  Grubbs  test  and  using  the  Cochran  test  for  outlying  standard 
deviations, (both at a 99 % confidence level). Standard deviations within (swithin) and between 
(sbetween)  laboratories  were  calculated  using  one-way  ANOVA.  The  results  of  these 
evaluations are shown in Table 11. 

Table 11: Statistical evaluation of the technically accepted datasets for ERM-CA403. p: 
number of technically valid datasets 

Analyte 

p 

Outliers 

Means 

Variances 

Normally 
distributed 

As 

Cd 

Co 

Cr 

Cu 

Fe 

Mn 

Mo 

Ni 

Pb 

Se 

5 

6 

5 

3 

6 

3 

6 

5 

7 

7 

2 

0 

0 

0 

0 

0 

0 

0 

0 

0 

1 

1 

0 

0 

0 

0 

1 

0 

1 

0 

1 

yes 

yes 

yes 

p < 4* 

yes 

p < 4* 

yes 

yes 

yes 

yes 

Statistical parameters 

Mean 
[µg/L] 
1.90 

s 
[µg/L] 
0.10 

sbetween 
[µg/L] 
0.09 

swithin 
[µg/L] 
0.09 

0.0939 

0.0123 

0.0122 

0.0049 

0.0735 

0.0081 

0.0079 

0.0040 

0.258 

0.875 

3.48 

2.47 

12.0 

1.04 

0.016 

0.015 

0.093 

0.092 

0.014 

0.028 

0.49 

0.08 

0.5 

0.11 

0.48 

0.07 

0.5 

0.11 

0.24 

0.09 

0.5 

0.07 

0.0984 

0.0083 

0.0078 

0.0069 

n.a. 

n.a. 

p < 4* 

0.0771 

0.0105 

0.0102 

0.0059 

Zn 

0 
* For p < 4 it is not possible to test Kurtosis 

4 

0 

yes 

4.59 

0.25 

0.25 

0.14 

The statistical evaluation flags L05 as a high outlier for Pb. However, the difference between 
the  mean  values  of  laboratory  L03  and  the  other  results  is  less  than  the  combined 
measurement uncertainty of laboratory L03 and the uncertainty of the dataset. As the mean 
of L03 agreed with the dataset, the value was retained. The outlying variances merely reflect 

28 

 
the  fact  that  the  method  used  has  different  intrinsic  variability.  As  all  measurement 
procedures were found technically valid, all results were retained. 

For Cr, Fe and Se, the statistical tests cannot be considered reliable, as only 3 laboratories 
mean values were accepted for Cr and Fe, and only 2 for Se. 

For  the  remaining  elements,  the  laboratory  means  follow  normal  distributions.  None  of  the 
data contains outlying means and variances. The datasets are therefore consistent and the 
mean  of  laboratory  means  is  a  good  estimate  of  the  true  value.  With  the  exception  of  Mn, 
standard deviations between laboratories are considerably larger than the standard deviation 
within  laboratories,  showing  that  confidence  intervals  of  replicate  measurements  are 
unsuitable as estimate of measurement uncertainty. 

In  addition  to  the  statistical  tests,  the  agreement  of  individual  laboratories'  results  with  the 
dataset  means  was  tested  according  to  ERM  Application  Note  1.[16]  As  approaches  to 
uncertainty  estimation  differ  between  laboratories,  it  is  possible  that  not  all  sources  of 
uncertainty  are  included  in  the  budgets.    In  addition,  labs  L04,  L05,  L06  and  L10  did  not 
provide measurement uncertainties. Therefore, Um of 20 % were assigned for all results for 
this  test,  as this  was  considered  to  represent  an  acceptable  level  of  Um  for  inclusion  in  the 
datasets. On this basis, all results agreed with the respective dataset means. 

The uncertainty related to the characterisation is estimated as the standard error of the mean 
of laboratory means (Table 12). 

Table 12: Uncertainty of characterisation for ERM-CA403 

Analyte 

Density 

Analyte 

As 

Cd 

Co 

Cu 

Mn 

Mo 

Ni 

Pb  

Zn 

p 

9 

p 

5 

6 

5 

6 

6 

5 

7 

7 

4 

Mean 
[g/mL] 
1.02352 

s 
[g/mL] 
25 • 10-6  8.5 • 10-6 

uchar 
[g/mL] 

Mean 
[µg/L] 

s 
[µg/L] 

uchar 
[µg/L] 

1.90 

0.10 

0.04 

0.0939 

0.0123 

0.0050 

0.0735 

0.0072 

0.0029 

0.875 

0.093 

0.038 

2.47 

12.0 

1.04 

0.08 

0.5 

0.11 

0.03 

0.2 

0.04 

0.0984 

0.0083 

0.0031 

4.59 

0.25 

0.13 

7  Value Assignment 

Certified, indicative and informative values were assigned. 

Certified values are values that fulfil the highest standards of accuracy. Procedures at JRC 
Directorate F require generally pooling of not less than 6 datasets to assign certified values. 
Full  uncertainty  budgets  in  accordance  with  the  'Guide  to  the  Expression  of  Uncertainty  in 
Measurement' [4] were established.  

29 

 
 
 
Indicative  values  are  values  where  either  the uncertainty  is  deemed  too large  or  where  too 
few  independent  datasets  were  available  to  allow  certification.  Uncertainties  are  evaluated 
according to the same rules as for certified values. 

Additional material information refers to values that were obtained in the course of the study. 
For example, results reported from only one or two laboratories or in cases where individual 
measurement uncertainty is high, would fall under this category.  

7.1  Certified values and their uncertainties 

The unweighted mean of the means of the accepted datasets as shown in Table and Table 
was assigned as certified value for each parameter.  

The  assigned  uncertainty  consists  of  uncertainties  related to  characterisation,  uchar  (Section 
6), potential between-unit inhomogeneity, ubb (Section 4.1) and potential degradation during 
transport  (usts)  and  long-term  storage,  ults  (Section  5).  These  different  contributions  were 
combined to estimate the expanded, relative uncertainty of the certified value (UCRM, rel) with a 
coverage factor k as:  

U

CRM,

rel

(cid:215)=
k

u

2
char,

rel

+

u

2
 bb,

rel

+

u

2
 sts,

rel

+

u

2
lts,

rel

. 

Equation 14 

-  uchar was estimated as described in Section 6  

-  ubb was estimated as described in Section 4.1 

-  usts was estimated as described in section 5.3 

-  ults was estimated as described in Section 5.3.  

For  the  concentrations  of  elements  in  CA403,  the  uncertainty  contributions  from  potential 
degradation during transport (usts) were found to be insignificant, and were not retained in the 
combined uncertainty calculation. 

Applying the Welch-Satterthwaite equation [4] to calculate the effective number of degrees of 
freedom yields between 11 and 38 for the elements As, Co, Mo and Zn, for which less than 6 
characterisation datasets were obtained. Because of the sufficient numbers of the degrees of 
freedom of the different uncertainty contributions, a coverage factor k of 2 was applied for all 
elements, to obtain the expanded uncertainties.  

The certified values and their uncertainties are summarised in Table 13. 

30 

 
 
 
 
 
 
 
Table 13: Certified values and their uncertainties for ERM-CA403 

Certified  value 
[g/mL] 

(at 

1.02352 

rel 

uchar, 
[%] 
0.0008 

Analyte 

Density 
20 °C) 

ults, rel  
[%] 

ubb, rel  
[%] 
0.0004  0.0002  0.002 

usts, rel  
[%] 

UCRM, 
[%] 

rel 

UCRM 
[g/mL]1) 

0.005 

0.00005 

Analyte 

Certified value 
[µg/L] 

uchar, rel 
[%] 

ubb, rel [%] 

ults, rel [%] 

UCRM, rel 
[%] 

UCRM [µg/L]1) 

As 

Cd 

Co 

Cu 

Mn 

Mo 

Ni 

Pb 

1.90 

0.094 

0.074 

0.87 

2.47 

12.0 

1.04 

0.098 

2.3 

5.4 

4.9 

4.3 

1.3 

1.9 

4.1 

3.2 

1.1 

1.1 

1.9 

5.7 

0.8 

0.3 

0.9 

3.0 

2.0 

2.1 

4.8 

1.8 

1.5 

1.5 

6.3 

2.2 

6 

12 

14 

15 

4 

5 

15 

10 

0.13 

0.011 

0.011 

0.13 

0.11 

0.6 

0.16 

0.010 

1) Expanded with a factor k=2, and rounded uncertainty. 

7.2  Indicative values and their uncertainties 

An  indicative  value  was  assigned  for  the  Zn  concentration,  as  there  were  less  than  5 
accepted  results.  However,  as  the  laboratories  also  provided  results  for  the  certified 
elements  that  agreed  with  the  dataset  means,  the  results  were  regarded  as  sufficiently 
trustworthy  to  assign  an  indicative  value.  Indicative  values  may  not  be  used  as  certified 
values.  The  uncertainty  budgets  were  set  up  as  for  the  certified  values  and  are  listed 
together with the assigned values in Table 14. 

Table 14: Indicative values and their uncertainties for ERM-CA403 

Analyte 

Indicative 
value [µg/L] 

uchar, rel 
[%] 

ubb, rel [%] 

ults, rel [%] 

UCRM, rel 
[%] 

UCRM [µg/L]1) 

Zn 

4.6 
1) Expanded with a factor k=2, and rounded uncertainty. 

2.8% 

1.5% 

4.7% 

11% 

0.6 

7.3  Additional material information 

Three laboratories in the characterisation exercise provided accepted results for the Fe and 
Cr concentrations, and a single laboratory provided two results for the Se concentration, with 
one  by  an  independent  technique  (Atomic  Fluorescence  Spectrometry  in  addition  to 
ICP-MS). These were used to calculate approximate concentration ranges, and are listed in 
Table 15. These values must be regarded as informative only and can not be, in any case, 
used as certified or indicative values. 

31 

 
 
 
 
 
 
Table 15: Additional information values for ERM-CA403 

Analyte 

Additional information 
range [µg/L] 

Cr 
Fe 
Se 

0.21 – 0.31  
2.7 – 4.2 
0.060 – 0.094 

8  Metrological traceability and commutability 

8.1  Metrological traceability  

Identity 

Density  is  a  clearly  defined  property  that  can  be  measured  with  high  specificity.  The 
participants  used  different  methods  for  the  sample  preparation  as  well  as  for  the  final 
determination, demonstrating absence of measurement bias. The measurands are therefore 
structurally defined and independent of the measurement method. 

The  concentrations  of  the  elements  are  method-defined  measurands  and  can  only  be 
obtained  by  ICP-MS  measurement.  Adherence  to  this  procedure  was  confirmed  by 
agreement of the laboratories' results with the assigned values for the CRM that was used as 
quality control sample. The measurand is therefore operationally defined by method. 

Quantity value 

Traceability  of  the  obtained  results  is  based  on  the  traceability  of  all  relevant  input  factors. 
Instruments  in  individual  laboratories  were  verified  and  calibrated  with  tools  ensuring 
traceability  to  the  International  System  of  units  (SI).  Consistency  in  the  interlaboratory 
comparison  demonstrates  that  all  relevant  input  factors  were  covered.  As  the  assigned 
values  are  combinations  of  agreeing  results  individually  traceable  to  the  SI,  the  assigned 
quantity values themselves are traceable to the SI as well. 

9 

Instructions for use 

9.1  Safety information 

The usual laboratory safety measures apply. 

9.2  Storage conditions 

The material shall be stored at 18 °C ± 5 °C in the dark.  

Please  note  that  the  European  Commission  cannot  be  held  responsible  for  changes  that 
happen  during  storage  of  the  material  at  the  customer's  premises,  especially  of  opened 
bottles. 

9.3  Preparation and use of the material 

The  bottles  shall  be  shaken  by  turning  upside  down  for  at  least  2  min  before  opening  to 
ensure material re-homogenisation.  

9.4  Minimum sample intake 

The minimum amount of sample to be used is 2 mL.   

32 

 
9.5  Use of the certified value 

The  main  purpose  of  this  material  is  to  assess  method  performance,  i.e.  for  checking 
accuracy  of  analytical  results/calibration.  As  any reference material,  it  can  also  be  used for 
control charts or validation studies. 

Use as a calibrant 

It  is  not  recommended  to  use  this  matrix  material  as  calibrant.  If  used  nevertheless,  the 
uncertainty  of  the  certified  value  shall  be  taken  into  account  in  the  estimation  of  the 
measurement uncertainty. 

Comparing an analytical result with the certified value 

A result is unbiased if the combined standard uncertainty of measurement and certified value 
covers the difference between the certified value and the measurement result (see also ERM 
Application Note 1, www.erm-crm.org [16].  

For  assessing  the  method  performance,  the  measured  values  of  the  CRMs  are  compared 
with the certified values. The procedure is described here in brief:  

-  Calculate  the  absolute  difference  between  mean  measured  value  and  the  certified 

value (D meas). 

-  Combine  measurement  uncertainty 

(umeas)  with 

the  uncertainty  of 

the  

certified value (uCRM): 

=D

u

u

+
2
meas u

2
CRM

-  Calculate the expanded uncertainty (UD ) from the combined uncertainty (uD

,) using an 
appropriate coverage factor, corresponding to a level of confidence of approximately 
95 % 
If  D meas  £   UD   no  significant  difference  between  the  measurement  result  and  the 
certified value, at a confidence level of about 95 % exists. 

- 

Use in quality control charts 

The  material  can  be  used  for  quality  control  charts.  Different  CRM-units  will  give  the  same 
result as inhomogeneity was included in the uncertainties of the certified values.  

33 

 
 
 
10   Acknowledgments 

The authors would like to acknowledge the support received from Jean Charoud-Got, Patrick 
Conneely,  Irma  Huybrechts,  Hanne  Leys,  Albert  Oostra,  Katharina  Teipel,  Marie-France 
Tumba  and  Diana  Vernelen  from  JRC  Directorate  F  related  to  the  processing  of  this  CRM 
and from Maria Contreras Lopez concerning the set-up of the required isochronous studies. 

Furthermore, the authors would like to thank Ioannis Dosis and Ingrid Zegers (JRC) for the 
reviewing of the certification report, as well as the experts of the Certification Advisory Panel 
(CAP)  "Elements  and  Inorganic  Ions",  Steve  Balsley  (International  Atomic  Energy  Agency 
(IAEA),  AT),  Thomas  C.  Meisel  (Montanuniversität  Leoben,  AT)  and  Peter  Vermaercke 
(Studiecentrum  voor  Kernenergie  (SCK),  BE),  and  the  Reference  Material  Review  Panel, 
Thomas  Meisel  and  Jens  J.  Sloth  (Technical  University  of  Denmark)  for  their  constructive 
comments. 

34 

11  References 

1  

2  

3  

4  

ISO  Guide  34,  General  requirements  for  the  competence  of  reference  materials 
producers, International Organization for Standardization, Geneva, Switzerland, 2009 

ISO Guide 35, Reference materials – General and statistical principles for certification, 
International Organization for Standardization, Geneva, Switzerland, 2006 

ISO/IEC  17025:2005,  General  requirements  for  the  competence  of  testing  and 
calibration 
for  Standardization,  Geneva, 
Switzerland, 2005 

International  Organization 

laboratories, 

ISO/IEC  Guide  98-3,  Guide  to  the  Expression  of  Uncertainty  in  Measurement,  (GUM 
1995), International Organization for Standardization, Geneva, Switzerland, 2008 

5   European  Commission,  Directive  2000/60/EC  of  the  European  Parliament  and  of  the 
Council of 23 October 2000 establishing a framework for Community action in the field of 
water policy, OJ Eur. Comm. L 327, 22.12.2000. 

6   European  Commission,  Directive  2013/39/EU  of  the  European  Parliament  and  of  the 
Council of 12 August 2013 on Amendment Directives 2000/60/EC and 2008/105/EC as 
Regards  Priority  Substances  in  the  Field  of  Water  Policy,  OJ  Eur.  Union  L  226/1, 
13.8.2013. 

7   European  Commission,  Directive  2009/90/EC  of  the  European  Parliament  and  of  the 
Council of 31 July 2009 on technical specifications for chemical analysis and monitoring 
of water status, OJ Eur. Union L 201/36, 1.8.2009. 

8   European  Commission,  Recommendation  of  the  European  Parliament  and  of  the 
Council  of  30  May  2002  concerning  the  implementation  of  Integrated  Coastal  Zone 
Management in Europe, OJ L 148/24, 6.6.2002. 

9   European  Commission,  Directive  2008/56/EC  of  the  European  Parliament  and  of  the 
Council  of  17  June  2008  establishing  a  framework  for  community  action  in  the  field  of 
marine environmental policy, OJ Eur. Union L 164/19, 25.6.2008. 

10   European  Commission,  Decision  of  1  September  2010  on  criteria  and  methodological 
standards  on  good  environmental  status  of  marine  waters,  OJ  Eur.  Union  L  232/14, 
2.9.2010. 

11   T.P.J.  Linsinger,  J.  Pauwels,  A.M.H.  van  der  Veen,  H.  Schimmel,  A.  Lamberty, 

Homogeneity and stability of reference materials, Accred. Qual. Assur. 6 (2001) 20-25 

12   G.  O’Donnell,  D.  Brynn,  Hibbert,  Treatment  of  bias  in  estimating  measurement 

uncertainty, Analyst 130 (2005) 721-729 

13   A. Lamberty, H. Schimmel, J. Pauwels, The study of the stability of reference materials 

by isochronous measurements, Fres. J. Anal. Chem. 360 (1998) 359-361 

14   ASTM D4052-09, Standard Test Method for Density, Relative Density, and API Gravity 

of Liquids by Digital Density Meter, ASTM International, West Conshohocken, 2009. 

15   T.P.J.  Linsinger,  J.  Pauwels,  A.  Lamberty,  H.  Schimmel,  A.M.H.  van  der  Veen,  L. 
Siekmann,  Estimating  the  Uncertainty  of  Stability  for  Matrix  CRMs,  Fres.  J.  Anal. 
Chem. 370 (2001) 183-188 

16  T.P.J.  Linsinger,  ERM  Application  Note  1:  Comparison  of  a  measurement  result  with 

the certified value, www.erm-crm.org (last accessed on 26.01.2015) 

35 

 
 
 
Annexes 

Annex  A:  Results  of  the  homogeneity  measurements  for  ERM-CA403  for  density  and  trace 
elements as reported by the laboratories. 

A1:  Density  results  of  the  mean  values  against  unit  number.  Vertical  bars  represent  standard 
deviation of the three replicates.  

36 

 
 
 
 
 
 
 
 
A2: Relative mass concentrations of elements against unit number. Points are normalised due to 
between-run effects and are relative to the mean of all points. Vertical bars are the 95 % confidence 
interval of measurement. 

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

As

Cd

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

0

200 400 600 800 1000 1200 1400 1600

0

200 400 600 800 1000 1200 1400 1600

Unit number
Co

Unit number
Cr

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

0

200 400 600 800 1000 1200 1400 1600

0

200 400 600 800 1000 1200 1400 1600

Unit number
Cu

Unit number
Fe

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

0

200 400 600 800 1000 1200 1400 1600

0

200 400 600 800 1000 1200 1400 1600

Unit number

Unit number

37 

 
 
 
 
 
 
 
 
 
 
 
n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

150%

125%

100%

75%

50%

Mn

Mo

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

0

200 400 600 800 1000 1200 1400 1600

0

200 400 600 800 1000 1200 1400 1600

Unit number
Ni

Unit number
Pb

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

0

200 400 600 800 1000 1200 1400 1600

0

200 400 600 800 1000 1200 1400 1600

Unit number

Se

Unit number
Zn

n
o
i
t
a
r
t
n
e
c
n
o
c
e
v
i
t
a
e
r

l

120%

110%

100%

90%

80%

0

200 400 600 800 1000 1200 1400 1600

0

200 400 600 800 1000 1200 1400 1600

Unit number

Unit number

38 

 
 
 
 
 
 
 
 
 
 
 
 
Annex  B:  Results  of  the  short  term  stability  measurements  for  ERM-CA403  for  density  and 
trace elements. 

The data for the short-term stability study at 18 °C. the graphs report unit averages per time point 
and the standard deviation of the measurements per time as reported by the laboratories. 

39 

 
 
 
 
 
 
 
 
 
 
 
 
40 

 
 
 
 
 
 
 
 
 
 
 
 
Annex B – Continued: Results of the short term stability measurements for ERM-CA403 for 
density and trace elements. 

The data for the short-term stability study at 60 °C. the graphs report unit averages per time point 
and the standard deviation of the measurements per time as reported by the laboratories. 

41 

 
 
 
 
 
 
 
 
 
 
42 

 
 
 
 
 
 
 
 
 
Annex  C:  Results  of  the  long-term  stability  measurements  for  ERM-CA403  for  density  and 
trace elements. 

The data for the long-term stability study at 18 °C. the graphs report unit averages per time point 
and the standard deviation of the measurements per time. 

43 

 
 
 
 
 
 
 
 
 
 
 
 
 
 
 
44 

 
 
 
 
 
 
 
 
 
 
Annex D: Summary of analytical techniques used in the characterisation of ERM-CA403 as reported by the laboratories (which may 
not be in line with VIM) 

D.1 Density 

Lab-method code 

Sample pretreatment 

None  

Analytical method 
Oscillating type density 
meter in accordance with 
ASTM D4052-09 
Oscillating type density 
meter  

None 
Temperature stabilisation   Oscillating type density 

meter  

L01-OD 

L02-OD 

L04-OD 

L05-Pyc  

Temperature stabilisation  

Gravimetric density 
determination with 
pycnometer  

L06-OD 

Temperature stabilisation 
(24 hours) 

Oscillating type density 
meter  

L08-OD 

None 

L09-Pyc 

L10-OD 

None 

Not used in certification 

L03-OD 

Samples were stirred 
during 30 minutes at 150 
rpm before collecting one 
subsample 

Oscillating type density 
meter  

Gravimetric density 
determination with 
pycnometer in accordance 
with ASTM D1480  
Oscillating type density 
meter 

Oscillating type density 
meter 

45 

Calibrant 

Instrument 

Air, purified water and 
dodecane 

Anton Paar DMA5000M 

Distilled water 

Anton Paar DMA5000 

Air, distilled water and 
dodecane 
The volume of the 
pycnometer was calibrated 
with primary density 
standard (water) by 
hidrostatic weighing 

2,2,4 trimethyl pentane, 
dodecan, lube oil 8,pure 
water and dimethyl 
phalate 

Anton Paar DMA5000 

Pycnometer made with 
glass 

Anton Paar DMA5000 

Anton Paar DMA5000 

Bingham Pycnometers 

Air and ultra-pure water 

Anton Paar DMA5000 

Air and ultra-pure water 

Anton Paar DMA5000 

 
 
 
 
 
 
L07-OD 

The samples were 
homogenized by shaking 
and adjusted at 20ºC 

Oscillating type density 
meter 

None  

Portable Density Meter 
Densito 30PX, Mettler 
Toledo 

D.2 Mass concentration of As 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L04-ICP-MS 

None  

He collision mode. 

L01-ICP-SFMS 

L02-ICP-MS 

L12-ICP-MS 

L13-ICP-MS 

Not used in certification 

L03-ICP-SFMS 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.5 µg/L 

Dilution and acidification 
with ultrapure 0.5 % 
HNO3. 

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

ICP-MS 

Ge as internal standard. 

ICP-MS 
Sc, Ge, Y, Rh, In as 
internal standards 

ICP-MS in collision 
reaction mode 

Dilution 1:2 with Miili-Q 
purified water 

10 g sample is introduced 
in SeaFAST matrix 
separation and pre-
concentration system and 
retained elements eluted 
with  200 µl 2% HNO3 

Linear, two points: 0.12/ 
5.12 µg/L. Matrix matched. 

0.12 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
25.0 µg/L. 

0.05 µg/L 

External 

1 µg/L 

External with 
gravimetrically diluted 
multi-elemental standards, 
recovery correction with  
NASS-6; SLEW-3 or 
CASS-5. 

0.3 ng/L 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 5/ 10 µg/L. 

0.5 µg/L 

46 

 
 
 
L05-ICP-MS 

L06-ICP-MS 

Pre-concentration step 
with resin chelex-100. 
Elution with 5 mL of HNO3 
2 mol/L. 

Reductive precipitation by 
pH adjustment with 0.2% 
(v/v) HNO3 then prepared 
according to EPA Method 
1640.  

ICP-MS 
Y as internal standard. 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
20.0 µg/L. 

not supplied 

ICP-MS 
Sc, Ge, In, Tm as internal 
standards 

External, 8 calibration 
points 

Varies, reported per 
sample 

D.3 Mass concentration of Cd 

Lab-method code 

Sample pretreatment 

L01-ICP-SFMS 

Acidification with 1 mL ultra high-
purity HNO3 per 100 mL of sample. 

L02-ICP-MS 

Dilution and acidification with 
ultrapure 0.5 % HNO3. 

L04-ICP-MS 

None  

Analytical method 
ICP-SFMS 
In as internal 
standard.  
ICP-MS 
Collision cell mode. 
Rh as internal 
standard. 
ICP-MS 
He collision mode. 
In as internal 
standard. 

Calibration 

External: 0/ 2/ 10 µg/L. Matrix 
matched. 

LOQ 

0.03 µg/L 

Linear, two points: 0.005/ 5.005 
µg/L. Matrix matched. 

0.005 µg/L 

External: 0/ 0.05/ 0.10/ 0.5/ 1.00/ 
2.50/ 5.00/ 10.0/ 25.0 µg/L. 

0.05 µg/L 

L05-ICP-MS 

L10-ICP-MS 

Pre-concentration step with resin 
Chelex®-100. Elution with 5 mL of 
HNO3 2 mol/L. 

ICP-MS 
Y as internal standard. 

External: 0/ 0.05/ 0.10/ 0.5/ 1.00/ 
2.50/ 5.00/ 10.0/ 20.0 µg/L. 

column chelation after adjustment to 
1% (v/v) HNO3 

ICP-MS 
In internal standard 

External, 7 calibration points 

Varies, reported 
per sample 

47 

 
 
 
 
 
 
L13-ICP-MS 

10 g sample is introduced in 
SeaFAST matrix separation and pre-
concentration system and retained 
elements eluted with  200 µl 2% 
HNO3 

ICP-SFMS in low 
resolution mode 

External with gravimetrically diluted 
multi-elemental standards, recovery 
correction with  NASS-6; SLEW-3 or 
CASS-5. 

0.06 ng/L 

Not used in certification 

L03-ICP-SFMS 

Dilution 1:5 with 1 % HNO3 
solution. 

ICP-SFMS 
In as internal standard 

Standard addition, two 
points: 0.05/ 0.10 µg/L. 

0.01 µg/L 

L06-ICP-MS 

Reductive precipitation by pH 
adjustment with 0.2% (v/v) 
HNO3 then prepared 
according to EPA Method 
1640.  

ICP-MS 
Sc, Ge, In, Tm as internal 
standards 

External, 8 calibration 
points 

Varies, reported per 
sample 

D.4 Mass concentration of Co 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L01-ICP-SFMS 

Acidification with 1 mL ultra 
high-purity HNO3 per 100 mL 
of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.03 µg/L 

L02-ICP-MS 

Dilution and acidification with 
ultrapure 0.5 % HNO3. 

L04-ICP-MS 

None  

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

ICP-MS 

He collision mode. 

Ge as internal standard. 

Linear, two points: 0.01/ 
5.01 µg/L. Matrix matched. 

0.01 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
25.0 µg/L. 

L10-ICP-MS 

column chelation after 
adjustment to 1% (v/v) HNO3 

ICP-MS 
In internal standard 

External, 7 calibration 
points 

Varies, reported per 
sample 

48 

 
 
 
 
L13-ICP-MS 

10 g sample is introduced in 
SeaFAST matrix separation 
and pre-concentration system 
and retained elements eluted 
with  200 µl 2% HNO3 

ICP-SFMS in low 
resolution mode 

External with 
gravimetrically diluted 
multi-elemental standards, 
recovery correction with  
NASS-6; SLEW-3 or 
CASS-5. 

0.1 ng/L 

Not used in certification 

L03-ICP-SFMS 

L05-ICP-MS 

Dilution 1:5 with 1 % HNO3 
solution. 

Pre-concentration step with 
resin chelex-100. Elution with 
5 mL of HNO3 2M. 

ICP-SFMS 

In as internal standard 

ICP-MS 
Y as internal standard. 

Standard addition, two 
points: 5/ 10 µg/L. 

0.01 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
20.0 µg/L. 

D.5 Mass concentration of Cr 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L01-ICP-SFMS 

Acidification with 1 mL ultra 
high-purity HNO3 per 100 mL 
of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.15 µg/L 

L02-ICP-MS 

Dilution and acidification with 
ultrapure 0.5 % HNO3. 

L04-ICP-MS 

None  

Not used in certification 

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

ICP-MS 

He collision mode. 

Y as internal standard. 

Linear, two points: 0.01/ 
5.01 µg/L. Matrix matched. 

0.10 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
25.0 µg/L. 

0.05 µg/L 

L03-ICP-SFMS 

Dilution 1:5 with 1 % HNO3 
solution. 

ICP-SFMS 

Standard addition, two 
points: 5/ 10 µg/L. 

0.02 µg/L 

49 

 
 
 
 
L06-ICP-MS 

Reductive precipitation by pH 
adjustment with 0.2% (v/v) 
HNO3 then prepared 
according to EPA Method 
1640.  

In as internal standard 

ICP-MS 
Sc, Ge, In, Tm as internal 
standards 

External, 8 calibration 
points 

Varies, reported per 
sample 

D.6 Mass concentration of Cu 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L01-ICP-SFMS 

Acidification with 1 mL ultra 
high-purity HNO3 per 100 mL 
of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.3 µg/L 

L02-ICP-MS 

Dilution and acidification with 
ultrapure 0.5 % HNO3. 

L04-ICP-MS 

None  

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

ICP-MS 

He collision mode. 

Ge as internal standard. 

Linear, two points: 0.05/ 5.05 
µg/L. Matrix matched. 

0.05 µg/L 

External: 0/ 0.05/ 0.10/ 0.5/ 
1.00/ 2.50/ 5.00/ 10.0/ 25.0 
µg/L. 

0.10µg/L 

L05-ICP-MS 

L10-ICP-MS 

L13-ICP-MS 

Pre-concentration step with 
resin chelex-100. Elution with 
5 mL of HNO3 2 mol/L. 

ICP-MS 
Y as internal standard. 

External: 0/ 0.05/ 0.10/ 0.5/ 
1.00/ 2.50/ 5.00/ 10.0/ 20.0 
µg/L. 

column chelation after 
adjustment to 1% (v/v) HNO3 

ICP-MS 
In internal standard 

10 g sample is introduced in 
SeaFAST matrix separation 
and pre-concentration system 
and retained elements eluted 
with  200 µl 2% HNO3 

ICP-SFMS in low 
resolution mode 

50 

External, 7 calibration points 

External with gravimetrically 
diluted multi-elemental 
standards, recovery 
correction with  NASS-6; 
SLEW-3 or CASS-5. 

Varies, reported per 
sample 

0.0167 µg/L 

 
 
 
 
Not used in certification 

L03-ICP-SFMS 

L06-ICP-MS 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 5/ 10 µg/L. 

0.02 µg/L 

Reductive precipitation by 
pH adjustment with 0.2% 
(v/v) HNO3 then prepared 
according to EPA Method 
1640.  

ICP-MS 
Sc, Ge, In, Tm as internal 
standards 

External, 8 calibration 
points 

Varies, reported per 
sample 

D.7 Mass concentration of Fe 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L01-ICP-SFMS 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

1.5 µg/L 

L02-ICP-MS 

L10-ICP-MS 

Not used in certification 

L03-ICP-SFMS 

Dilution and acidification 
with ultrapure 0.5 % 
HNO3. 

column chelation after 
adjustment to 1% (v/v) 
HNO3 

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

Linear, two points: 0.14/ 
5.14 µg/L. Matrix matched. 

0.14 µg/L 

ICP-MS 
In internal standard 

External, 7 calibration 
points 

Varies, reported per 
sample 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 0.25/ 0.50 µg/L. 

0.05 µg/L 

51 

 
 
 
 
 
 
D.8 Mass concentration of Mn 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.3 µg/L 

L01-ICP-SFMS 

L02-ICP-MS 

Dilution and acidification 
with ultrapure 0.5 % 
HNO3. 

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

ICP-MS 

L04-ICP-MS 

None  

He collision mode. 

Linear, two points: 0.26/ 5.26 
µg/L. Matrix matched. 

0.26 µg/L 

External: 0/ 0.05/ 0.10/ 0.5/ 
1.00/ 2.50/ 5.00/ 10.0/ 25.0 
µg/L. 

External: 0/ 0.05/ 0.10/ 0.5/ 
1.00/ 2.50/ 5.00/ 10.0/ 20.0 
µg/L. 

Ge as internal standard. 

ICP-MS 
Y as internal standard. 

L05-ICP-MS 

Dilution 1:10. 

L10-ICP-MS 

L13-ICP-MS 

Not used in certification 

column chelation after 
adjustment to 1% (v/v) 
HNO3 
10 g sample is introduced 
in SeaFAST matrix 
separation and pre-
concentration system and 
retained elements eluted 
with  200 µl 2% HNO3 

ICP-MS 
In internal standard 

External, 7 calibration points 

Varies, reported per 
sample 

ICP-SFMS in low 
resolution mode 

External with gravimetrically 
diluted multi-elemental 
standards, recovery 
correction with  NASS-6; 
SLEW-3 or CASS-5. 

0.8 ng/L 

L03-ICP-SFMS 

Dilution 1:5 with 1 % HNO3 
solution. 

L12-ICP-MS 

Dilution 1:2 with Miili-Q 
purified water 

ICP-SFMS 

In as internal standard 
ICP-MS 
Sc, Ge, Y, Rh, In as internal standards 

Standard addition, two 
points: 5/ 10 µg/L. 

0.05 µg/L 

External 

1 µg/L 

52 

 
 
 
 
D.9 Mass concentration of Mo 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L05-ICP-MS 

Dilution 1:10. 

L01-ICP-SFMS 

L02-ICP-MS 

L12-ICP-MS 

L13-ICP-MS 

Not used in certification 

L03-ICP-SFMS 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.15 µg/L 

Dilution and acidification 
with ultrapure 0.5 % 
HNO3. 

Dilution 1:2 with Milli-Q 
purified water 

10 g sample is introduced 
in SeaFAST matrix 
separation and pre-
concentration system and 
retained elements eluted 
with  200 µl 2% HNO3 

ICP-MS 

Collision cell mode. 

Rh as internal standard. 

ICP-MS 
Y as internal standard. 

ICP-MS 
Sc, Ge, Y, Rh, In as 
internal standards 

ICP-SFMS in low 
resolution mode 

Linear, two points: 0.98/ 
5.98 µg/L. Matrix matched. 

0.98 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
20.0 µg/L. 

External 

1 µg/L 

External with 
gravimetrically diluted 
multi-elemental standards, 
recovery correction with  
NASS-6; SLEW-3 or 
CASS-5. 

0.01298 µg/L 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 5/ 10 µg/L. 

0.05 µg/L 

53 

 
 
 
 
 
 
 
L01-ICP-SFMS 

L02-ICP-MS 

D.10 Mass concentration of Ni 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.3 µg/L 

Dilution and acidification 
with ultrapure 0.5 % 
HNO3. 

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

ICP-MS 

Linear, two points: 0.03/ 
5.03 µg/L. Matrix matched. 

0.03 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
25.0 µg/L. 

0.05 µg/L 

L04-ICP-MS 

None  

He collision mode. 

Ge as internal standard. 

L05-ICP-MS 

L06-ICP-MS 

L10-ICP-MS 

L13-ICP-MS 

Pre-concentration step 
with resin chelex-100. 
Elution with 5 mL of HNO3 
2M. 
Reductive precipitation by 
pH adjustment with 0.2% 
(v/v) HNO3 then prepared 
according to EPA Method 
1640.  
column chelation after 
adjustment to 1% (v/v) 
HNO3 
10 g sample is introduced 
in SeaFAST matrix 
separation and pre-
concentration system and 
retained elements eluted 
with  200 µl 2% HNO3 

ICP-MS 
Y as internal standard. 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
20.0 µg/L. 

ICP-MS 
Sc, Ge, In, Tm as internal 
standards 

External, 8 calibration 
points 

Varies, reported per 
sample 

ICP-MS 
In internal standard 

External, 7 calibration 
points 

Varies, reported per 
sample 

External with 
gravimetrically diluted 
multi-elemental standards, 
recovery correction with  
NASS-6; SLEW-3 or 
CASS-5. 

5.3 ng/L 

ICP-SFMS in low 
resolution mode 

54 

 
 
 
Not used in certification 

L03-ICP-SFMS 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 0.25/ 0.50 µg/L. 

0.05 µg/L 

D.11 Mass concentration of Pb 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

Lu as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.03 µg/L 

L01-ICP-SFMS 

L02-ICP-MS 

Dilution and acidification 
with ultrapure 0.5 % 
HNO3. 

ICP-MS 

Collision cell mode. 

Rh as internal standard. 

ICP-MS 

L04-ICP-MS 

None  

He collision mode. 

Linear, two points: 0.005/ 
5.005 µg/L. Matrix 
matched. 

0.005 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
25.0 µg/L. 

0.1 µg/L 

External: 0/ 0.05/ 0.10/ 
0.5/ 1.00/ 2.50/ 5.00/ 10.0/ 
20.0 µg/L. 

External, 8 calibration 
points 

Varies, reported per 
sample 

Bi as internal standard. 

ICP-MS 
Y as internal standard. 

ICP-MS 
Sc, Ge, In, Tm as internal 
standards 

ICP-MS 
In internal standard 

External, 7 calibration 
points 

Varies, reported per 
sample 

55 

L05-ICP-MS 

Dilution 1:10. 

L06-ICP-MS 

L10-ICP-MS 

Reductive precipitation by 
pH adjustment with 0.2% 
(v/v) HNO3 then prepared 
according to EPA Method 
1640.  
column chelation after 
adjustment to 1% (v/v) 
HNO3 

 
 
 
 
 
L13-ICP-MS 

Not used in certification 

L03-ICP-SFMS 

10 g sample is introduced 
in SeaFAST matrix 
separation and pre-
concentration system and 
retained elements eluted 
with  200 µl 2% HNO3 

ICP-SFMS in low 
resolution mode 

External with 
gravimetrically diluted 
multi-elemental standards, 
recovery correction with  
NASS-6; SLEW-3 or 
CASS-5. 

0.3 ng/L 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 0.05/ 0.10 µg/L. 

0.01 µg/L 

D.12 Mass concentration of Se 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L01-ICP-SFMS 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

0.3 µg/L 

L11-AFS 

Dilution 1:5 with 1 % 
HNO3 solution. 

AFS according to SS-EN 
ISO 17852:2008 

0.03 µg/L 

Not used in certification 

L03-ICP-SFMS 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 0.05/ 0.10 µg/L. 

0.01 µg/L 

56 

 
 
 
 
 
 
 
D.13 Mass concentration of Zn 

Lab-method code 

Sample pretreatment 

Analytical method 

Calibration 

LOQ 

L01-ICP-SFMS 

Acidification with 1 mL 
ultra high-purity HNO3 per 
100 mL of sample. 

ICP-SFMS 

In as internal standard.  

External: 0/ 2/ 10 µg/L. 
Matrix matched. 

1.5 µg/L 

L02-ICP-MS 

L10-ICP-MS 

L13-ICP-MS 

Not used in certification 

Dilution and acidification 
with ultrapure 0.5 % 
HNO3. 

ICP-MS 

Collision cell mode. 

Ge as internal standard. 

Linear, two points: 0.07/ 
5.07 µg/L. Matrix matched. 

0.07 µg/L 

column chelation after 
adjustment to 1% (v/v) 
HNO3 

10 g sample is introduced 
in SeaFAST matrix 
separation and pre-
concentration system and 
retained elements eluted 
with  200 µl 2% HNO3 

ICP-MS 
In internal standard 

External, 7 calibration 
points 

Varies, reported per 
sample 

ICP-SFMS in low 
resolution mode 

External with 
gravimetrically diluted 
multi-elemental standards, 
recovery correction with  
NASS-6; SLEW-3 or 
CASS-5. 

4.8 ng/L 

L03-ICP-SFMS 

Dilution 1:5 with 1 % 
HNO3 solution. 

ICP-SFMS 

In as internal standard 

Standard addition, two 
points: 0.25/ 0.50 µg/L. 

0.25 µg/L 

57 

 
 
 
Annex E: Results of the characterisation study 

The tables in this annex also contain the data sets that were discarded for technical reasons. These 
data  sets  are  highlighted  in  italics  and  are  given  for  information  purposes  only.  They  are  not 
included in the graphs. 

Error bars represent expanded uncertainties as reported by participating laboratories. The solid line 
represents the certified values (the mean of the laboratory means), while the broken lines represent 
the  expanded  uncertainty  of  the  certified  value.  For  the  trace  element  measurements,  Labs  L04, 
L05,  L06  and  L10  did  not  provide  uncertainties.  For  these  results,  an  indicative  measurement 
uncertainty of 20 % was assigned, as described in section 6.4.2. 

Approaches to number rounding differed between participants. Values presented in this annex were 
rounded to assist document formatting. 

E.1 Density [g/mL] 

U [%] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean  
L01-OD 
L02-OD 
L03-OD 
L04-OD 
L05-Pyc  
L06-OD 
L08-OD 
L09-Pyc 
L10-OD 
Results not used in the certification 
1.0236 
L07-OD 

0.16 
1.023528  1.023526  1.023520  1.023528  1.023512  1.023522  1.023523 
0.003 
1.023507  1.023507  1.023506  1.023508  1.023509  1.023510  1.023508 
1.023506  1.023514  1.023507  1.023504  1.023507  1.023512  1.023508 
0.003 
1.023511  1.023509  1.023510  1.023509  1.023507  1.023509  1.023509  0.0010 
0.002 
1.023523  1.023524  1.023522  1.023525  1.023521  1.023523  1.023523 
0.03 
1.023500  1.023620  1.023620  1.023560  1.023520  1.023640  1.023577 
0.003 
1.023490  1.023481  1.023487  1.023494  1.023492  1.023485  1.023488 
0.010 
1.023520  1.023510  1.023520  1.023510  1.023510  1.023460  1.023505 
0.002 
1.023538  1.023553  1.023518  1.023573  1.023522  1.023519  1.023537 

1.0239  1.023817 

1.0239 

1.0238 

1.0237 

1.024 

58 

 
 
 
 
 
 
 
 
 
E.2 As mass concentration [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
1.930 
L02 
1.900 
L04 
1.743 
L12 
1.940 
L13 
1.780 
Results not used in the certification 
L03 
L06 

1.880  1.847  0.390 
2.100  1.923  0.170 
2.042  2.027 
1.930  1.940  0.120 
1.740  1.773  0.158 

1.840 
1.880 
2.161 
1.910 
1.780 

1.780 
1.870 
2.173 
1.990 
1.830 

1.780 
1.940 
1.961 
1.940 
1.740 

1.870 
1.850 
2.080 
1.930 
1.770 

1.899  1.938  0.082 
1.830  1.722 

1.953 
1.730 

1.908 
1.930 

1.999 
1.590 

1.976 
1.770 

1.891 
1.480 

2.5

L
/
g
µ

2.0

1.5

1.0

13

1

2

12

4

Lab code

59 

 
 
 
 
 
 
 
E.3 Cd mass concentration [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
0.0830 
L02 
0.1050 
L04 
0.0900 
L05 
0.1200 
L10 
0.0884 
L13 
0.0907 
Results not used in the certification 
L03 
L06 

0.0870  0.0782  0.0300 
0.1060  0.1027  0.0110 
0.0820  0.0880 
0.1100  0.1133 
0.0906  0.0897 
0.0950  0.0917  0.0046 

0.0710 
0.1050 
0.0980 
0.1200 
0.086 
0.0925 

0.0740 
0.0970 
0.0810 
0.1100 
0.0913 
0.0923 

0.0720 
0.1040 
0.0850 
0.1100 
0.0875 
0.0903 

0.0820 
0.0990 
0.0920 
0.1100 
0.0946 
0.0895 

0.0545  0.0564  0.0071 
0.1090  0.1013 

0.0569 
0.0800 

0.0558 
0.1100 

0.0505 
0.0940 

0.0619 
0.0990 

0.0589 
0.1160 

0.14

0.12

0.10

L
/
g
µ

0.08

0.06

0.04

0.02

1

4

10

13

2

5

Lab code

60 

 
 
 
 
 
 
 
 
 
 
E.4 Co mass concentration, [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
0.0820 
L02 
0.0800 
L04 
0.0670 
L10 
0.0675 
L13 
0.0644 
Results not used in the certification 
L03 
L05 

0.0800  0.0792  0.0220 
0.0890  0.0838  0.0100 
0.0780  0.0732 
0.0622  0.0657 
0.0681  0.0658  0.0061 

0.0740 
0.0830 
0.0800 
0.0685 
0.0657 

0.0800 
0.0790 
0.0700 
0.0630 
0.0690 

0.0810 
0.0790 
0.0720 
0.0681 
0.0643 

0.0780 
0.0930 
0.0720 
0.0649 
0.0632 

0.0692  0.0727  0.0056 
0.1200  0.1183 

0.0765 
0.0900 

0.0741 
0.0900 

0.0753 
0.1600 

0.0717 
0.1200 

0.0695 
0.1300 

L
/
g
µ

0.12

0.10

0.08

0.06

0.04

0.02

10

13

4

1

2

Lab code

61 

 
 
 
 
 
 
 
 
E.5 Cu mass concentration [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
0.923 
L02 
0.994 
L04 
0.757 
L05 
0.870 
L10 
0.839 
L13 
0.758 
Results not used in the certification 
L03 
L06 

0.977  0.948  0.209 
1.015  1.022  0.080 
0.820  0.809 
0.880  0.868 
0.791  0.817 
0.819  0.785  0.067 

0.950 
1.031 
0.833 
0.880 
0.777 
0.805 

0.968 
1.022 
0.810 
0.910 
0.833 
0.780 

0.944 
1.051 
0.790 
0.870 
0.829 
0.749 

0.923 
1.019 
0.842 
0.800 
0.833 
0.798 

1.868  1.864  0.083 
0.920  0.843 

1.852 
0.840 

1.914 
0.940 

1.919 
0.770 

1.810 
0.920 

1.822 
0.670 

1.2

1.0

L
/
g
µ

0.8

0.6

0.4

13

4

10

5

1

2

Lab code

62 

 
 
 
 
 
 
 
 
 
E.6 Mn mass concentration [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
2.558 
L02 
2.500 
L04 
2.387 
L05 
2.350 
L10 
2.370 
L13 
2.445 
Results not used in the certification 
L03 
L12 

2.559  2.523  0.500 
2.510  2.487  0.220 
2.628  2.567 
2.350  2.357 
2.390  2.392 
2.642  2.508  0.204 

2.484 
2.350 
2.436 
2.380 
2.350 
2.570 

2.521 
2.370 
2.657 
2.400 
2.370 
2.420 

2.464 
2.720 
2.575 
2.300 
2.500 
2.399 

2.552 
2.470 
2.718 
2.360 
2.370 
2.568 

2.428  2.438  0.034 
3.320  3.278  0.180 

2.431 
3.300 

2.468 
3.200 

2.435 
3.160 

2.416 
3.520 

2.449 
3.170 

3.5

3.0

L
/
g
µ

2.5

2.0

1.5

5

10

2

13

1

4

Lab code

63 

 
 
 
 
 
 
 
 
 
 
E.7 Mo mass concentration [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
12.20 
L02 
12.65 
L05 
11.25 
L12 
11.60 
L13 
12.31 
Results not used in the certification 
L03 

12.80  12.32  1.70 
12.56  12.62  1.27 
11.60  11.48 
11.60  11.50  0.60 
10.19  11.89  0.86 

12.00 
12.74 
11.80 
11.10 
12.08 

12.70 
12.42 
11.30 
11.50 
12.89 

11.60 
12.73 
11.50 
11.70 
11.71 

12.60 
12.61 
11.40 
11.50 
12.15 

8.82 

8.45  1.36 

9.70 

8.33 

7.79 

7.68 

8.38 

L
/
g
µ

15

14

13

12

11

10

9

8

5

12

13

1

2

Lab code

64 

 
 
 
 
 
 
 
E.8 Ni mass concentration [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
1.060 
L02 
1.240 
L04 
0.972 
L05 
1.050 
L06 
0.910 
L10 
1.110 
L13 
0.954 
Results not used in the certification 
L03 

1.120  1.026  0.340 
1.340  1.245  0.110 
1.078  1.027 
1.030  1.093 
0.900  0.883 
0.993  1.058 
0.996  0.961  0.086 

0.902 
1.170 
0.990 
1.020 
1.030 
0.972 
0.975 

0.986 
1.200 
1.066 
1.190 
0.900 
1.120 
0.902 

1.060 
1.280 
0.988 
1.150 
0.880 
1.040 
0.932 

1.030 
1.240 
1.068 
1.120 
0.680 
1.110 
1.006 

0.422  0.417  0.045 

0.367 

0.429 

0.422 

0.429 

0.431 

L
/
g
µ

1.4

1.2

1.0

0.8

0.6

0.4

6

13

1

4

10

5

2

Lab code

65 

 
 
 
 
 
 
 
 
 
 
E.9 Pb mass concentration [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
0.104 
L02 
0.100 
L04 
0.096 
L05 
0.120 
L06 
0.103 
L10 
0.092 
L13 
0.092 
Results not used in the certification 
L03 

0.100  0.098  0.029 
0.105  0.097  0.022 
0.099  0.093 
0.110  0.115 
0.117  0.102 
0.090  0.091 
0.092  0.092  0.007 

0.101 
0.080 
0.091 
0.110 
0.086 
0.091 
0.092 

0.091 
0.109 
0.090 
0.110 
0.093 
0.091 
0.093 

0.097 
0.090 
0.091 
0.120 
0.114 
0.094 
0.094 

0.093 
0.100 
0.093 
0.120 
0.100 
0.091 
0.086 

0.071  0.084  0.017 

0.091 

0.082 

0.077 

0.097 

0.085 

L
/
g
µ

0.14

0.12

0.10

0.08

0.06

0.04

10

13

4

2

1

6

5

Lab code

66 

 
 
 
 
 
 
 
 
 
 
 
E.10 Zn mass concentration, indicative value [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
4.92 
L02 
4.84 
L10 
4.34 
L13 
4.23 
Results not used in the certification 
L03 
L05 

4.70  0.98 
4.89  0.55 
4.47 
4.31  0.34 

4.5 
5.00 
4.48 
4.41 

4.53 
4.71 
4.37 
4.19 

4.59 
4.94 
4.42 
4.37 

4.88 
4.89 
4.55 
4.50 

4.78 
4.98 
4.68 
4.18 

0.38  0.11 

0.47 
<2 

0.43 
<2 

0.37 
<2 

0.34 
<2 

0.33 
<2 

0.34 
<2 

L
/
g
µ

6

5

4

3

2

13

10

1

2

Lab code

67 

 
 
 
 
 
 
 
 
 
 
E.11 Cr mass concentration, additional information value [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
0.286 
L02 
0.242 
L04 
0.260 
Results not used in the certification 
L03 
L05 
L06 

0.253  0.271  0.061 
0.244  0.240  0.040 
0.269  0.263 

0.267 
0.218 
0.248 

0.280 
0.270 
0.277 

0.278 
0.235 
0.259 

0.262 
0.228 
0.266 

0.329  0.389  0.068 

0.410 
<0.5 
0.155 

0.394 
<0.5 
0.291 

0.424 
<0.5 
0.163 

0.417 
<0.5 
0.374 

0.358 
<0.5 
0.149 

0.272  0.234 

<0.5 

0.35

0.30

L
/
g
µ

0.25

0.20

0.15

2

4

Lab code

1

68 

 
 
 
 
 
 
 
 
 
 
 
E.12 Fe mass concentration, additional information value [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
4.060 
L02 
2.930 
3.580 
L10 
Results not used in the certification 
L03 
L05 

4.090  3.925  0.810 
2.910  2.960  0.310 
3.130  3.548 

0.414  0.490  0.103 

3.860 
3.010 
4.250 

3.810 
3.100 
3.560 

3.990 
2.920 
3.390 

3.740 
2.890 
3.380 

0.503 
<1 

0.483 
<1 

0.547 
<1 

0.439 
<1 

0.554 
<1 

<1 

/

L
g
µ

5

4

3

2

1

0

2

10

Lab code

1

69 

 
 
 
 
 
 
 
 
 
E.13 Se mass concentration, additional information value [µg/L] 

Lab code  Sample 1  Sample 2  Sample 3  Sample 4  Sample 5  Sample 6  Mean   U 
L01 
0.084 
L10 
0.0654 
Results not used in the certification 
L03 
L05 

0.088 
0.0687 

0.087 
0.0804 

0.077 
0.0729 

0.085 
0.0718 

0.0589  0.070 

0.08 
<1 

0.08 
<1 

0.08 
<1 

0.08 
<1 

0.08 
<1 

<1 

0.086  0.085  0.061 

0.07  0.078  0.009 

0.16

0.12

/

L
g
µ

0.08

0.04

0

11

1

Lab code

70 

 
 
 
 
 
 
 
 
European Commission 

EUR 29055 EN – Joint Research Centre – Directorate F – Health, Consumers and Reference Materials 
Title:   CERTIFICATION REPORT: The certification of the mass concentration of As, Cd, Co, Cu, Mn, Mo, Ni and Pb, 

and density in seawater: ERM®- CA403 

Author(s): R. Sánchez, E. de Vos, J. Snell, H. Emteborg, A. Held 
Luxembourg: Publications Office of the European Union 
2018 – 70 pp. – 21.0 x 29.7 cm 
EUR – Scientific and Technical Research series – ISSN 1831-9424 
ISBN 978-92-79-77706-6 
doi: 10.2760/405177 

 
 
 
 
 
 
 
 
As the Commission's in-house science service, the Joint Research Centre's mission is to provide EU policies 
with independent, evidence-based scientific and technical support throughout the whole policy cycle. 

Working in close cooperation with policy Directorates-General, the JRC addresses key societal challenges 
while stimulating innovation through developing new methods, tools and standards, and sharing its know-
how with the Member States, the scientific community and international partners. 

Key policy areas include: environment and climate change; energy and transport; agriculture and food 
security; health and consumer protection; information society and digital agenda; safety and security, 
including nuclear; all supported through a cross-cutting and multi-disciplinary approach. 

K
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doi: 10.2760/405177 

ISBN: 978-92-79-77706-6