# Microbial Application to Improve Olive Mill Wastewater Phenolic Extracts
## 1 Introduction  
The Mediterranean diet is becoming more and more popular due to its nutritional and health benefits, which are associated with lower incidences of atherosclerosis, certain cancers, and cardiovascular and neurodegenerative diseases [1, 2, 3, 4]. These health benefits can be strongly related to the high content of antioxidant molecules present in food products widely consumed within the Mediterranean diet, such as extra virgin olive oil and table olives. Olives and their industrial derived products are rich in unsaturated fatty acids, tocopherols, and phenols. However, the olive oil industry generates large amounts of high-polluting by-products, such as a solid residue and an effluent commonly named olive mill wastewater (OMW) [5], which is a mix of olive vegetation water and water added during oil extraction, above all with the three-phase centrifugation system [6]. Olive waste still contains most of the valuable and interesting compounds. During the olive oil processing, most of the phenolic compounds are found in the aqueous phase, with a hydroxytrosol concentration up to 100-fold higher than that found in olive oil [7]. OMW is rich in phenols, such as hydroxytrosol, tyrosol, oleuropein, flavonoids, and other compounds. These molecules have great potential in the pharmaceutical and nutraceutical fields [8]. In particular, hydroxytrosol is characterized by remarkable antioxidant activity, which is similar to that of the main synthetic antioxidants [7, 9]. In 2011 [10], the scientific opinion on the substantiation of health claims related to olive polyphenols, standardizedby their content of hydroxytyrosol and its derivatives (e.g., oleuropein complex), was published by European Food Safety Authority (EFSA). Substantial interest has been given to the production of hydroxytyrosol both from natural sources and by synthetic procedures [11]. Several techniques have been reported in literature aiming at recovering phenolic compounds from OMW, such as liquid-liquid extraction [12], membrane filtration [13], adsorption on selective polymeric resins [14; 15], and enzymatic reactions [11; 16; 17]. Compared to chemical treatments, enzymatic treatment is advantageous because it requires mild operating conditions, such as pH, temperature, and the absence of toxic organic solvents [18]. To overcome the high costs of enzymes and their poor operational stabilities, new research has focused on enzyme immobilization strategies [19]. Among microorganisms, _Aspergillus_ spp. has been extensively studied for its ability to produce extracellular enzymes that can be easily applied to food processing, because they are recognized as Generally Regarded as Safe (GRAS) by the Food and Drug Administration (FDA) [20]. Within the _Aspergillus_ genus, _A. niger_ is the most efficient producer of \(\beta\)-glucosidase [21]. However, few reports focused on the effect of fungal enzyme addition on OMW phenols have been published, but even less on the direct application of live microorganisms into an OMW extract to improve antioxidant activity. In particular, yeasts and lactic acid bacteria (LAB) appear particularly interesting as they are commonly used as starter cultures in fermented food and widely isolated by OMW or spontaneous fermentation of table olives. During the last two decades, several _Lactiplantiobacillus plantarum_ strains have been extensively described for their ability to degrade eleuropein and for their probiotic potential [22; 23]. [PERSON] et al. [24] for the first time tested the in vivo bioconversion of oleuropein into hydroxytyrosol by oral granules containing probiotic _L. plantarum_ and an olive leaves standardized extract. Their results showed that co-administration of live _L. plantarum_ bacteria with the extract provides for higher amounts of bioavailable hydroxytyrosol, compared to the extract alone. Several yeasts isolated from table olives have been found able to produce noticeable enzymes, such as \(\beta\)-glucosidase, and to exhibit probiotic traits [25; 26].