DIN EN 14176:2017-03

The amnesic shellfish poisoning (ASP) toxin, domoic acid (DA), belongs to a group of amino acids, called the kainoids, which are classed as neuroexcitants or excitoxins that interfere with the neurotransmission mechanisms in the brain. The toxin can be accumulated in shellfish feeding on a number of toxic Pseudonitzschia species. Ingestion of seafood contaminated with DA can lead to an intoxication which symptoms include (among others) abdominal cramps, vomiting, disorientation and memory loss (amnesia) and can become severe in certain cases. This European Standard specifies methods for the quantitative determination of domoic acid in raw bivalve molluscs and finfish as well as in cooked mussels. The limit of detection is about 10 ng/ml to 80 ng/ml (0,05 mg/kg to 0,4 mg/kg), depending on the UV detector sensitivity. Method A has been validated for the determination of DA in different raw matrices such as mussels, clams, scallops and anchovies, spiked and/or naturally contaminated at levels ranging from 2,7 mg/kg to 85,1 mg/kg. Method B has been validated for the determination of DA at levels ranging from 5 mg/kg to 12,9 mg/kg in cooked blue mussels. This European Standard is based on two comparable procedures: one procedure for the quantitative determination of DA and its isomers, for example epi-domoic acid (epi-DA) in unsalted raw seafood (Method A) and another method for the quantitative determination of DA and its isomers, for example epi-domoic acid (epi-DA) in cooked mussel (Method B). Method A uses a single-step extraction with 50 % aqueous methanol and an optional selective clean-up and concentration step with strong anion exchange solid phase extraction (SPE). Taking into account results of the validation procedure, the optional clean-up step of Method A described in the original publication is not described in this standard. Analytes are determined by high performance liquid chromatography (HPLC) under isocratic conditions with ultraviolet absorbance detection. Method B uses a single-step extraction with 50 % aqueous methanol and an optional heating step which allows a better decanting of the supernatant. However, it has been observed that heating can degrade DA and epi-DA. DA and epi-DA are determined by HPLC with binary gradient and ultraviolet absorbance detection. Both methods can be applied for the quantitative determination of DA. This document (EN 14176:2017) has been prepared by Technical Committee TC 275 "Food analysis - Horizontal methods" (secretariat: DIN, Germany) of the European Committee for Standardization (CEN) after detailed preliminary work by Working Group 14 "Marine biotoxins" (secretariat: DIN, Germany). The responsible German working committee is DIN Working Group NA 057-01-03-01 AK "Algentoxine" ("Algal toxins") of DIN Standards Committee Food and Agricultural Products (NAL).