| Summary: | Oil extracted from walnuts leaves behind large amounts of defatted press cake that still retains valuable nutrients. Solid state fermentation (SSF) is a promising method to liberate bioactive compounds from food by-products. <i>Aspergillus oryzae</i>, <i>Rhizopus oligosporus</i>, and <i>Streptococcus thermophilus</i> possess the necessary enzymes to utilize these nutrients from the walnut press cake (WPC) and convert walnut ellagitannins into ellagic acid and urolithins. This study conducted SSF with WPC to release ellagic acid from ellagitannins and form urolithins. The growth of the two filamentous fungi could be observed visually and the growth of <i>Streptococcus thermophilus</i> was confirmed by plate count technique. Extracts from fermented products were subjected to analysis using HPLC–DAD to measure the release of ellagic acid from ellagitannins. Additionally, a more sensitive UHPLC–MS method was employed to screen fermented samples for urolithin A. The ellagic acid content exhibited no perceptible change but was already present in the press cake before and after all fermentations. Urolithin A was undetectable, even with the more sensitive MS method. All studies showing urolithin A formation were conducted under anaerobic conditions. This might be a basic prerequisite for the transformation of ellagic acid to urolithins.
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