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Abstract Six interlaboratory studies were organised by species (arsenobetaine, arsenocholine, monomethylarson-
the Standard, Measurement and Testing Programme of the ic acid, dimethylarsinic acid, As(III) and As(V)) in marine
European Commission on the determination of arsenic matrices and soil. A step-by-step approach was used and a
meeting was held at the end of each study to help the par-
ticipants to discover errors and to improve their analytical
F. Lagarde · M. B. Amran · M. J. F. Leroy methods. The successive steps investigated the calibration
Laboratoire de Chimie Analytique et Minérale, procedures on various solutions, the separation and de-
URA 405 (CNRS-ECPM-Université Louis Pasteur), rivatisation techniques on solutions and extracts and the
ECPM, 1 rue Blaise Pascal BP 296,
F-67008 Strasbourg Cédex, France extraction on mussel and fish tissues. All materials used
for the study were monitored for their stability. Verified
C. Demesmay · M. Ollé · A. Lamotte calibration solutions and compounds were distributed to
Service Central d’Analyse, Echangeur de Solaize BP22,
F-69390 Vernaison Cédex, France the participants in each exercise in order to trace calibra-
tion problems. The agreement between the results im-
H. Muntau proved regularly and at the end of the six campaigns al-
Joint Research Center,
Commission of the European Communities, I-21020 Ispra, Italy lowed the certification of a reference material of tuna-fish
tissue (BCR-CRM 627) for its total arsenic, arsenobetaine
P. Michel and dimethylarsinic acid contents.
IFREMER, BP 1049, F-44037 Nantes Cédex, France
P. Thomas
Institut Pasteur de Lille, Départment Eaux Environnement,
BP 245, F-59019 Lille Cédex, France
1 Introduction
S. Caroli It is known for a long time that some marine organisms
Istituto Superiore de la Sanita, Lab. Igiene del Teritorio,
299 viale Regina Elena, I-00161 Roma, Italy may contain high amounts of arsenic [1]. Concentrations
generally lie in the range of 10 to 500 µg · g–1 but may
E. Larsen sometimes exceed 1000 µg · g–1. The highest content re-
Institute of food chemistry & nutrition, National Food Agency,
DK 2860 Soeborg, Denmark ported (2739 µg · g–1) was found by Gibbs et al. in a poly-
chaeta Tharyx Marioni living in a contaminated area [2].
P. Bonner The first arsenic compound contained in marine animals
State Laboratory Abbosttown, IRL-15 Dublin, Ireland
was isolated from the western rock lobster Panulirus
G. Rauret Cygnus in 1977 and found to be arsenobetaine [3]. Since
Universidad de Barcelona, Dept. Quimica Analitica, then, other species such as arsenocholine, the tetramethyl-
Av. Diagonal 647, E-08028 Barcelona, Spain
arsonium ion, trimethylarsenoxide, dimethylarsinic acid
M. Foulkes and arsenosugars have been identified [4–11]. Very per-
University of Plymouth, Dept. of Environmental Sciences,
UK-PL4 8AA Plymouth, Great Britain
forming hyphenated techniques involving separation by
liquid chromatography and detection by Inductively Cou-
A. Howard pled Plasma – Mass Spectrometry (ICP-MS) [8, 9, 11–14],
University of Southampton, Dept. of Chemistry,
UK-SO9 Highfield, Southampton, Great Britain
Inductively Coupled Plasma – Atomic Emission Spec-
trometry (ICP-AES) [15], Hydride Generation – Quartz
B. Griepink · E. A. Maier (쾷) Furnace Atomic Absorption Spectrometry (HG-QFAAS)
European Commission, Standards,
Measurement and Testing Programme, rue de la Loi 200, [15, 16], or UV degradation – Inductively Coupled Plas-
B-1049 Brussels, Belgium ma Atomic Emission Spectrometry (UV-HG-ICP-AES)
6
Table 2 Composition of the solutions distributed for the second and third interlaboratory studies (all concentrations in mg · kg–1)
Compound 2nd interlaboratory study 3rd interlaboratory study
Solution 1 Solution 2 Solution 3 Solution 4 Solution 5 Solution 1 Solution 2 Solution 3 Solution 4 Solution 5
Aschol 5 1 1 3 1.5 0.25 – – 0.5 –
Asbet 5 – 10 – 1.5 1 0.5 – 1.25 –
As2O3 5 – 1 2 – 0.3 0.3 0.4 – 1
DMA 5 10 10 0.75 – 0.15 0.3 0.2 0.5 0.75
MMA 5 – 1 – 1 0.15 0.2 0.1 0.5 0.5
As2O5 5 10 – – 2 0.3 0.75 1 0.75 2
8
be avoided by dilution of the extract or by using a C18 pre- quality control points necessary to trace and eliminate
column. On chloride cartridges, As(V) and DMA were them were known and under control for the fish matrix.
partially or totally lost. As difficulties with calibration still As a consequence, certification of at least major As spe-
persisted, it was decided that a common unknown standard cies in a separate fish tissue was considered as possible.
solution to check the calibration step would further be dis- This study is described in this issue [19].
tributed. The main observation reported for the mussel ex- The methods and the results for dried mussel tissue
tract concerned the presence of two unknown compounds. were not considered as sufficiently reliable to envisage
It was suggested that these peaks might be due to arseno- certification. The total As content in mussels was often
sugars as already reported in the literature [4–11]. Only much higher than the calculated total As resulting from
3 laboratories detected a peak at the retention time of ar- the speciation measurements. The extraction efficiency in
senocholine but at a level close to the detection limits. As the mussel tissue did never exceed 80% and was often
the results between the laboratories showed a relative lower than 50%. This was attributed to incomplete extrac-
BLSD comparable to the fourth study, it was decided to tion of the studied compounds or to the presence of some
pass to the next step on a raw mussel extract and dried unidentified forms of As which remained in the solid
shark and mussel tissue. residue.
Arsenobetaine, DMA and traces of arsenocholine found in All the analytical steps (extraction, purification, separa-
the samples, were stable at +4 °C and +20 °C in the dark tion, detection) needed for arsenic speciation in fish and
for the entire duration of the interlaboratory study. The mussel have been studied, optimised and validated in suc-
different purification methods tested by the participants cessive interlaboratory studies. Each step benefited from
on the raw mussel extract were based on solvent extrac- the preparation of adequate samples systematically checked
tion using chloroform, diethylether or petroleum ether, for homogeneity and stability. The overall progress made
column elution on a C18 cartridge, dilution or filtration. by the group of laboratories for arsenobetaine, DMA and
Some of the lower values obtained in the extract for ar- MMA in the various samples is shown in Table 4. As(III)
senobetaine could be attributed to losses during the clean- could never be detected in natural fish or mussel tissue
up process. Some higher values were attributed to a co- samples, therefore participants decided to optimise their
elution of an unknown compound with arsenobetaine and methods towards the determination of the other forms of
identified on similar separation phases by other partici- As. Arsenocholine was only detected by some methods
pants. and at very low concentrations. The results also confirm
Two major forms of As, DMA and arsenobetaine, were the validity of such interlaboratory studies to achieve a
detected in both fish and mussel tissue powders. Inorganic quality of determinations which allows certification of
forms of As and in particular As(III) could not be identi- reference materials. The methods which demonstrated the
fied; traces of As(V) were reported by only few partici- necessary reliability to certify the fish tissue material are
pants but could not be accurately determined (close to de- described in detail in this issue [19].
tection limit). Extraction efficiencies (sum of As in vari- The findings made over the six exercises have been
ous chemical forms compared to total As determination) used to elaborate a detailed analytical protocol. This pro-
were very different from one laboratory to another (50 to tocol included various quality control points to allow to
90%). This could be explained either by the poor determi- trace potential sources of errors in the final certification
nation of the total arsenic content (partial hydride genera- study. In particular, the group recommended to make
tion) or by the extraction method itself. Therefore, it was available in the certification study a solution of all studied
recommended to check that the quantification of total ar- substances in order to verify the calibration [18]. It was
senic is properly validated and to verify its accuracy by recommended to certify an arsenobetaine solution as this
using a certified reference material of similar composition substance is not yet available on the market. Arsenocho-
(BCR-CRM 278, NRCC-DORM-1). Correction for water line could not be determined by all participants in the real
content was also requested. samples studied from the fifth exercise up to the certifica-
Recovery experiments were performed in order to ver- tion of the fish tissue. Therefore, no arsenocholine solu-
ify for potential losses or transformation of the species tion has been certified. Over the study it became clear that
during clean-up/pre-concentration steps. Figure 4 shows off-line techniques with sampling of fractions eluting from
that good agreement between laboratories could be ob- a liquid chromatography system are inadequate for the de-
tained in particular when using ICP-MS. With strong UV termination of several forms of As. On-line methods al-
irradiation, hydride generation techniques (Fig. 4: ICP-AES low much more flexible procedures and achieve better ac-
of laboratory 1) achieved results close to those of ICP- curacy. Capillary zone electrophoresis was rapidly aban-
MS. Two participants did still not achieve to set-up an UV doned because of a lack of sensitivity of the UV detection.
irradiation system which delivered a sufficient irradiation Hydride generation with strong UV irradiation [19] al-
power (Fig. 4: laboratories 1 and 2). At this stage of the lowed to achieve very satisfactory performances for ar-
project, several sources of errors were detected and the senobetaine and was finally applied in the certification of
11
the fish tissue. In general, it became clear over the dura- 7. Cullen WR, Dodd M (1989) Appl Organomet Chem 3 : 79
tion of these studies that ICP-MS was the most adequate 8. Beauchemin D, Bednas ME, Berman S, McLaren JW, Siu
KWM, Sturgeon RE (1988) Anal Chem 60 : 2209
detection for As speciation work provided the chloride 9. Shibata Y, Morita M (1989) Anal Chem 61 : 2116
ions are fully removed. The overall performance of the 10. Edmonds JS, Francesconi KA (1981) Nature 289 : 602
group is demonstrated by the certification of a reference 11. Larsen EH, Pritzl G, Hansen SH (1993) J Anal At Spectrom 8 :
material for some of the studied substances. This certifi- 1075
12. Branch S, Ebdon L, O’Neill P (1994) J Anal At Spectrom 9 : 33
cation is described in details in this issue [19]. 13. Demesmay C, Ollé M, Porthault M (1994) Fresenius J Anal
Chem 348 : 205
14. Thomas P, Sniatecki K (1995) Fresenius J Anal Chem 351 : 410
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