Development of an Extraction Method of Aflatoxins and Ochratoxin A from Oral, Gastric and Intestinal Phases of Digested Bread by In Vitro Model.

Validated extraction methods from in vitro digestion phases are necessary to obtain a suitable bioaccessibility study of mycotoxins in bakery products. The bakery industry produces bread with different ingredients to enrich the nutritional properties of this product and protect it from fungal growth. This bread can be contaminated by AFB, AFB, AFG, AFG and OTA, so an extraction method was developed to analyse these five legislated mycotoxins in digested phases of two types of bread, one with wheat and the other with wheat and also enriched with at 20%. The studied "in vitro" digestion model consists of oral, gastric and duodenal phases, each one with different salt solutions and enzymes, that can affect the extraction and most probably the stability of the mycotoxins. The proposed method is a liquid-liquid extraction using ethyl acetate by extract concentration. These analytes and components have an important effect on the matrix effect (MEs) in the analytical equipment, therefore, validating the method and obtaining high sensitivity will be suitable. In the proposed method, the highest MEs were observed in the oral phase of digested pumpkin bread (29 to 15.9 %). Regarding the accuracy, the recoveries were above 83% in the digested duodenal wheat bread and above 76 % in the digested duodenal pumpkin wheat bread. The developed method is a rapid, easy and optimal option to apply to oral, gastric and duodenal phases of digested bread contaminated at a level of established maximum levels by European legislation (RC. 1881/2006) for food.