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Challenges of accurate gluten quantitation in selected foods.
K. KONITZER (1), H. Wieser (1), P. Koehler (1). (1) Deutsche Forschungsanstalt fuer Lebensmittelchemie, Freising, Germany

The accurate quantitation of the gluten content in foods and beverages remains a considerable challenge. Sensitive immunological techniques like ELISA are widely used to assess compliance to the Codex Alimentarius threshold of 20 mg/kg for gluten-free foods. However, ELISAs also have limitations due to different reference standards, extraction protocols, and antibodies that are only specific for prolamins and not for all gluten proteins. In addition to PCR, mass spectrometry is the most promising nonimmunological approach for the detection of gluten proteins and peptides. Key considerations to be made are the selection of relevant gluten marker peptides, the conversion of peptide to gluten content and the establishment of reliable databases covering as many protein sequence variations as possible. An important point for all methods is appropriate sample preparation to solubilize all gluten proteins or peptides from the matrix independent of prior heat-treatment, hydrolysis or fermentation. Apart from the PWG-gliadin provided by the Working Group on Prolamin Analysis and Toxicity, a commonly accepted reference material for gluten is not available so far and there are no independent reference methods to verify ELISA results. Wheat starch, rendered gluten-free by extensive washing, is commonly used as ingredient for gluten-free foods. A gel permeation-HPLC method with fluorescence detection allowed the quantitation of both prolamins and glutelins in wheat starch. The analysis of wheat starch prepared from dough followed by three washing steps with water showed that prolamins are removed more extensively than glutelins. The ratio of prolamins to glutelins was <1 already after two washing steps, so that the gluten content in wheat starches may be underestimated after ELISA quantitation and duplication of the prolamin content to obtain the gluten content.

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