# Increasing the Bioactive Compound Content of Olive Oil by Acidification of Olive Paste
## 1 Introduction  
Olive oil is highly valued for its exceptional nutritional properties and associated health benefits. It is widely consumed globally, playing a particularly crucial role in Mediterranean countries, which account for a substantial portion of global olive oil production and consumption [1]. Its composition, characterized by unsaturated fatty acids, bioactive compounds, and a distinctive flavor and aroma profile, makes it a unique dietary fat [2]. Several factors influence the phenolic composition and quality of olive oil, including geographical origin, soil composition, climatic conditions, agricultural practices, olive cultivar, fruit ripeness, and processing parameters [3]. Among the minor components in olive oil, phenolic compounds are crucial due to their antioxidant properties and their contribution to the stability, bitterness, and pungency of virgin olive oils. These bioactive molecules have demonstrated potential in mitigating inflammation, cardiovascular diseases,certain cancers, microbial infections, and cellular oxidative stress [4; 5; 6; 7; 8; 9]. The European Commission has authorized a health claim for olive oil, stating that 'olive oil polyphenols contribute to the protection of blood lipids against oxidative damage'. This claim is valid provided a minimum daily intake of 20 g of olive oil containing at least 250 mg/kg of polyphenols, particularly hydroxytyrosol and its derivatives, is consumed [10]. Secoriidoids constitute the most abundant group of phenolic compounds in olive oil, comprising approximately 80% of total phenolics. These compounds are formed during oil extraction through the hydrolysis of oleuropein and ligstroside. Oleacein (3,4-DHPEA-EDA) and oleocanthal (\(p\)-HPEA-EDA) are the predominant secoriidoids, while their precursor aglycones (3,4-DHPEA-EA and \(p\)-HPEA-EA) complete this group [11]. Hydroxytyrosol and tyrosol are the final decomposition products of oleacein and oleocanthal, respectively (Figure 1). Among all antioxidants present in olive oil, hydroxytyrosol (3,4-DHPEA) and its secoriidoid derivatives (3,4-DHPEA-EDA and 3,4-DHPEA-EA) are considered the most potent, exhibiting greater antioxidant activity than even \(\alpha\)-tocopherol [3]. Other phenolic constituents of virgin olive oil include phenolic acids (e.g., vanillic, ferulic, and \(p\)-coumaric acid), lignans (such as pinoresinol), and flavonoids (such as luteolin and apigenin) [12; 13; 14]. The phenolic and volatile profiles of olive oil are largely determined by the enzymatic activity of the olive fruit, which is influenced by factors such as ripening stage, geographical origin, and water stress. Furthermore, the technological conditions employed during oil extraction significantly affect these profiles [15; 16; 17; 18; 19]. In particular, the milling and malaxation steps play a crucial role in regulating the secoiridoid content of olive oil. Lower milling temperatures and controlled malaxation conditions have been associated with higher retention of these valuable phenolic compounds. Optimal conditions include malaxation at temperatures between 25 and 27 \({}^{\circ}\)C, acidic pH, and minimal oxygen exposure, as these conditions favor the enzymatic activity of endogenous lipoxygenase and \(\beta\)-_glucosidase_, which contribute to higher concentrations of volatile compounds and secoriidoids [20; 21; 22]. Conversely, increased olive ripeness leads to the activation of phenol oxidase enzymes, reducing the levels of oleacein, oleocanthal, and their derivatives. Oleocanthal-rich oils have been linked to improved cognitive function in patients with Alzheimer's disease [23]. Additionally, oleocanthal offers cardiovascular benefits and has demonstrated therapeutic potential for treating inflammation, cancer, and neurodegenerative conditions [24; 25]. Similarly, oleacein exhibits antioxidant and anti-inflammatory prop  
Figure 1: Secoriidoid formation from oleuropein/ligstroside hydrolysis and the subsequent conversion to hydroxytyrosol/tyrosol.