Methylmercury determined by HPLC-ICP-MS in marine food and feed: in-house method validation and inter-laboratory comparison

Rie Romme Rasmussen (Lecturer)

Activity: Talks and presentationsConference presentations


Abstract to RAFA 2013 submitted for the category: Industrial contaminants

Methylmercury determined by HPLC-ICPMS in marine food and feed;
in-house method validation and interlaboratory comparison

Rie R. Rasmussen1, Maja E. Svendsen1, Heidi Amlund2, Martijn van der Lee3, Inge Rokkjær4 and Jens J. Sloth1*

1) National Food Institute (DTU Food), Technical University of Denmark, Soeborg, Denmark
2) National Institute of Nutrition and Seafood Research (NIFES), Bergen, Norway
3) RIKILT – Institute of Food Safety, Wageningen, The Netherlands
4) Danish Veterinary and Food Administration, Laboratory Aarhus (Chemical), Lystrup, Denmark
*) Corresponding author e-mail:; Phone +45 35887625

Mercury (Hg) is one of the most hazardous compounds in our environment and is released through natural and anthropogenic processes. It occurs as elemental mercury (metallic), inorganic mercury or organic mercury. Inorganic mercury and the organic mercury compound, methylmercury, are the two major mercury species found in the environment. The toxicity is highly dependent on the chemical form and methylmercury is particularly toxic as it affects the functions of enzymes and is toxic to the nervous system (with the developing brain as most the most sensitive target).

Currently only total mercury in foodstuffs is regulated by the European Union but accurate estimation of methylmercury exposure is needed for better evaluation of food safety. European Food Safety Authority established in 2012 a maximum tolerable weekly intake at 1.3 µg/kg body weight for methylmercury. Fully validated and standardized methods for determination of organic mercury levels in foods are currently missing.

Here results from an in-house validation and an interlaboratory comparison study are presented for a simple HPLC-ICPMS method developed for methylmercury in marine based food and feed extracted with 5 M hydrochloric acid by sonication. The extraction step was carried out twice, the pH was increased and the extracts were diluted in mobile phase and filtrated (0.45 m) prior to HPLC-ICP-MS analysis. Quantification of methylmercury (m/z 202) in the sample extracts was achieved by cation exchange separation (Hamilton PRP-X200, 150×2.1mm, 10 μm) and calibration standards prepared in mobile phase. The mobile phase (0.20 ml/min) consisted of 0.5% (w/v) L-cysteine, 50 mM pyridine, 0.8% (v/v) formic acid, 5% (v/v) MeOH and had a pH 3.

The in-house validation included certified reference materials of marine origin (TORT-2, DORM-2 and DORM-3) and 4 other marine samples which were analysed in triplicates on 3 different days. The individual results were within the certified ranges. The limit of detection was 0.004 mg/kg and the in-house reproducibility standard deviations were less than ≤20% for samples containing 0.06 to 4.47 mg/kg.

The small-scale collaborative study included four different laboratories that analysed 6 different marine food and feed samples in duplicate on two different days. For some samples one outlying dataset were excluded from the evaluation as Cochran or Grubbs outlier (following ISO 5725-2). The results were in general satisfactory in the tested range (0.2-5.7 mg/kg). The reproducibility standard deviations were less than ≤13%, HorRat values below 0.9 and the overall means for the reference materials (DORM-3, TORT-2, CE464 Tunafish) were in agreement with the certified values.

Pitfalls for application of the method are; carry over between HPLC injections, calculations (exact extraction volume is critical for re-extraction) and careful interpretation of methylmercury certificates (which can be given as e.g. Hg, as CH3Hg or CH3HgCl).

Keywords: Metylmercury, speciation, HPLC-ICPMS, validation, interlaboratory comparison

Funding from the European Community's Seventh Framework Programme (FP7/2007-2013) under grant agreement n° 211326: CONffIDENCE project (
Period8 Nov 2013
Event title6th International Symposium on Recent Advances in Food Analysis
Event typeConference
Conference number6
LocationPrague, Czech Republic