*Article* **A Tara Gum/Olive Mill Wastewaters Phytochemicals Conjugate as a New Ingredient for the Formulation of an Antioxidant-Enriched Pudding**

**Umile Gianfranco Spizzirri 1, Paolino Caputo 2, Cesare Oliviero Rossi 2, Pasquale Crupi 3, Marilena Muraglia 4, Vittoria Rago 1, Rocco Malivindi 1, Maria Lisa Clodoveo 3, Donatella Restuccia 1,\* and Francesca Aiello 1**


**Abstract:** Olive mill wastewater, a high polyphenols agro-food by-product, was successfully exploited in an eco-friendly radical process to synthesize an antioxidant macromolecule, usefully engaged as a functional ingredient to prepare functional puddings. The chemical composition of lyophilized olive mill wastewaters (LOMW) was investigated by HPLC-MS/MS and 1H-NMR analyses, while antioxidant profile was in vitro evaluated by colorimetric assays. Oleuropein aglycone (5.8 μg mL−1) appeared as the main compound, although relevant amounts of an isomer of the 3-hydroxytyrosol glucoside (4.3 μg mL−1) and quinic acid (4.1 μg mL−1) were also detected. LOMW was able to greatly inhibit ABTS radical (IC50 equal to 0.019 mg mL−1), displaying, in the aqueous medium, an increase in its scavenger properties by almost one order of magnitude compared to the organic one. LOMW reactive species and tara gum chains were involved in an eco-friendly grafting reaction to synthesize a polymeric conjugate that was characterized by spectroscopic, calorimetric and toxicity studies. *In vitro* acute oral toxicity was tested against 3T3 fibroblasts and Caco-2 cells, confirming that the polymers do not have any effect on cell viability at the dietary use concentrations. Antioxidant properties of the polymeric conjugate were also evaluated, suggesting its employment as a thickening agent, in the preparation of pear puree-based pudding. High performance of consistency and relevant antioxidants features over time (28 days) were detected in the milk-based foodstuff, in comparison with its non-functional counterparts, confirming LOWM as an attractive source to achieve high performing functional foods.

**Keywords:** olive mill wastewater; polyphenols; antioxidant features; tara gum; pudding; rheological properties
