Development of a reference material for mercury in fish: certified for total mercury and characterized for methylmercury

Reference materials (RMs) and certified reference materials (CRMs) are essential for ensuring compliance of food products with international standards. In fish, mercury content is closely monitored due to associated human health risks. This study developed and certified a CRM for total mercury (Hg) and methylmercury (MeHg) in striped catfish (Pseudoplatystoma fasciatum) from the Colombian Amazon. The process involved raw material selection, sample preparation, homogeneity, and stability, resulting in the CRM INM-039-1. Total Hg was measured using inductively coupled plasma-mass spectrometry (ICP-MS) and cold vapor atomic absorption spectrometry (CV-AAS), while MeHg was analyzed by gas chromatography-mass spectrometry (GC-MS). Calibration methods varied: ICP-MS used gravimetric standard addition, CV-AAS employed bracketing calibration, and GC-MS applied matrix-matched calibration. Method validation was performed using CRMs DORM-4 and ERM-CE464. The total Hg results were combined using the Levenson approach, yielding a relative standard uncertainty of 1.8%, while MeHg was 2.9%. Other uncertainty components included long-term stability (2.5%), homogeneity (1.8%), and short-term stability under transport conditions (0.02%). The CRM was certified with a total Hg mass fraction of 3.94 mg/kg (relative expanded uncertainty 7.1%, k = 2) and an informative MeHg mass fraction of 3.79 mg/kg (relative expanded uncertainty 8.3%, k = 2). This CRM is a valuable tool for assessing mercury in similar matrices, supporting compliance with safety standards for small and mid-size fish producers in the region.