Cardiovascular diseases (CVDs) are a major health problem and, to date, the principal cause of morbidity and mortality worldwide [1
]. The main condition that exposes people to CVD is represented by atherosclerosis, defined as a progressive inflammatory process caused by an excessive cholesterol deposition in the arterial walls. It is well-known that oxidative stress is strongly implicated in the pathogenesis of atherosclerosis, and oxidized low density lipoproteins (ox-LDL) play a critical role [2
]. Indeed, reactive oxygen species (ROS) can rapidly inactivate nitric oxide (NO) and form reactive nitrogen species (RNS) that damage vascular endothelial cells, creating a prothrombotic environment and an associated inflammatory condition. Therefore, anti-oxidant and anti-inflammatory agents can represent effective tools against atherosclerosis and, consequently, CVD [5
Indeed, in heart failure, inflammatory processes associated with fibrosis and alteration of angiogenesis lead to cardiac hypertrophy [6
]. Moreover, several studies have shown that cardiac dysfunctions such as myocardial infarction are associated with an increase of myocardial oxidative stress [7
]. Finally, coronary heart diseases can be deeply influenced by diet habits, particularly the intake of saturated fatty acids [8
In this context, lifestyle and dietary modifications are strongly recommended as an efficient, early interventional approach to changing these modifiable risk factors, acting especially on ROS and inflammatory markers.
In recent years a large number of scientific reports have pointed out the nutraceutical and nutritional value of the Mediterranean diet, suggesting that its consumption contributes to the reduction in the incidence of oxidative- and inflammatory-related pathologies, such as cardiovascular diseases and cancer. Virgin and extra virgin olive oils (EVOO) are a stronghold of the Mediterranean diet and have been described as functional foods endowed with a high nutritional quality [9
]. Indeed, the bio-functional components of EVOO show positive effects on genes involved in the pathogenesis of most prevalent age- and lifestyle-related human conditions, pointing to a role for these molecules as natural homeostatic and even hormetic factors in applications such as prevention agents used to treat conditions of premature and pathologic aging [14
Olive oil (OO) is a food composed of a major saponifiable fraction (about 98–99%) represented by oleic acid (55–83%) and other saturated and unsaturated acids (linoleic, palmitic and stearic acids, 3–21%), and of a minor unsaponifiable fraction (about 1–2%), including a high number of vitamins (α-, β-, γ- and δ- tocopherols), polyphenols (mainly tyrosol, hydroxytyrosol, and oleuropein) and squalene [15
] (Figure 1
Several reports suggest that the beneficial effects of EVOO are linked to the minor components and in particular to polyphenols; however, further studies have recently shed light on the health effects of the fatty fraction and the other constituents of the unsaponifiable fraction.
Indeed, the concept that saturated fatty acids (SFA) increase serum cholesterol and induce inflammation and insulin resistance, thus contributing to the risk of atherosclerosis and CVD, is generally accepted; on the other hand, various translational studies identify a protective role for unsaturated oils, monounsaturated fatty acids (MUFA), and more widely for polyunsaturated fatty acids (PUFA).
Considered as a whole, this evidence shows that EVOO is a functional food endowed with a healthy profile and the widely-studied phenolic component, as well as tocopherols and the MUFA (represented by oleic acid) fraction, can contribute in different ways and act on different types of molecular targets to ensure interesting pleiotropic effects.
In this regard, in 2004, based on numerous clinical trials carried out in the past few decades [17
], the US Food and Drug Administration (FDA), and more recently the European Food Safety Authority (EFSA), authorized the health claims for olive oil, suggesting a dose of 20–23 g/day as a replacement for the same amount of saturated fats to reduce the risk of coronary diseases [22
However, the quality of EVOO depends on a process that begins with the olive ripening and finishes with the packaging. Thus, agronomical practices, raw materials, harvesting, fruit storage, and extraction technology, and also oxygen, light, and temperature during storage, have to be considered in order to correctly estimate the nutraceutical, nutritional, and sensorial value.
In this context, the aim of this review has been twofold: firstly, an extensive analysis of clinical and preclinical evidence of cardiovascular beneficial effects of both unsaponifiable and saponifiable fractions of EVOO has been carried out; in the second part of the paper, the main operating conditions adopted during EVOO production and/or storage have been pointed out and critically discussed in order to highlight their influence on the concentration of health compounds in extracted oil as well as on their preservation during oil storage.
5. Focus on The Main Operating Conditions Adopted for EVOO Production and/or Storage: Influence on The Initial Concentration of Health Compounds and on The Kinetics of Their Degradation during Storage
According to Nicoli et al. 2012 [170
], “shelf life” can be defined as a finite length of time after production (in some cases, after maturation or aging) and packaging during which the food product retains a required level of quality under well-defined storage conditions.
With regards to EVOO, its shelf-life is directly linked to the occurrence of oxidation processes with a subsequent progressive degradation of the majority of both the saponifiable and the unsaponifiable fraction responsible for the healthy and nutraceutical properties attributed to EVOO. As reported in literature, EVOO shelf life has been assessed at 12–18 months [171
], even if it has been shown that when it is properly stored in well-sealed packages, this product can reach the second year of storage, preserving the concentration of active health compounds and thus maintaining its nutraceutical and sensorial properties unaltered to the greatest possible extent [172
However, the quality of EVOO in terms of both chemical compositions and sensorial expression depends on a process that begins with the olive ripening and finishes with the packaging. Thus, agronomical practices, raw materials, harvesting, fruit storage, and extraction technology, as well as oxygen, light, and temperature during storage, have to be considered in order to correctly estimate the nutraceutical, nutritional, and sensorial value [173
Based on a critical analysis of recent scientific literature, Figure 2
illustrates the main factors that can directly influence the olive oil composition (i.e., saponifiable and unsaponifiable fractions) during production as well as the degradation rate of main health compounds during storage.
5.1. Chemical Composition of Olive oil at Starting of Storage Time
The chemical and organoleptic quality of olive oil depends on several factors, such as the geographical location of the olive grove, the chemical and microbiological composition of the soil, the evolution of the climatic conditions during fruit ripening, and the extraction process [176
Among the several variables that could potentially determine the quality of this product, the oil composition can be greatly affected not only by the cultivar (genetic variability) as well as the ripening degree but also by the cultivation techniques (i.e., irrigation system) and the climatic conditions occurring in a specific crop season.
5.1.1. Characteristic of Raw Materials: Olive Cultivar, Ripening Degree, and Agronomic Practices
The oxidative stability of olive oil with respect to other vegetable oils is mainly due to its fatty acid composition, to the high MUFA/PUFA ratio in particular, and to the presence of minor compounds (i.e., polyphenols, carotenoids) that play a main role in preventing oxidation [173
The expression of phenolic compounds in olive fruit is predominately driven by genetic factors, and large differences exist between olive cultivars [179
]. In all cultivars, Ole and HT are the major phenolic compounds, but their concentrations vary considerably between cultivars at the same degree of ripeness [180
During fruit ripening and processing, many chemical and enzymatic transformations that affect the accumulation of phenols inside the olives may take place [181
]. In particular, due to the transformation of more structured compounds, phenols with a low molecular weight are produced [176
]. As a consequence, the quality, sensory properties, oxidative stability, and the nutritional value of the olive oil can change considerably [177
While the green or turning-color of olives creates a product characterized by bitter notes due to a higher presence of phenolic components (i.e., oleocanthal), the more acute and pungent notes are due to Tyrosol and its derivatives such as deacetoxy-ligstroside. Furthermore, some authors observed that the phenolic concentration of the olive fruit increases with ripening, reaching a maximum at the “half pigmentation” stage, after which it rapidly decreases [176
]. This evolution could explain why some researchers report that the phenolic concentration increased with the ripening degree of the olives [184
], while others observed an opposite evolution [176
Finally, the environmental conditions (especially light) as well as the type of fertilization also deeply influences phenolic biosynthesis in plants [186
]: While the yield of oil extracted from olive fruits belonging to the same cultivar and coming from the same orchard increased with the ripening degree of the milled fruits [180
], according to Caruso and co-workers [187
], the olives harvested on the same date from irrigated plants produced more oil than those coming from non-irrigated trees. Furthermore, agronomical practices seems to also influence the nutraceutical profile of extracted oil: Olives harvested from irrigated plants show a higher total phenol concentration value in the oil extracted than that obtained by milling fruits from non-irrigated trees [188
], and the organic fruits have a higher phenolic content than conventional ones [186
5.1.2. Extraction Technology
One of the most important industrial criticisms in the olive oil production is the low efficiency of current extraction techniques [189
]. Nowadays, several studies have pointed out the importance of the different virgin olive oil processing stages on the extraction yield as well as the minor composition found in the final product, and the most used solution for improve extraction is increased malaxation time and/or temperature [191
Scientific data report that milling and malaxation are the technological unit operations that most affect the quality of EVOO and the concentration of phenolic compounds and carotenoids, which are the main antioxidants of virgin olive oils [193
]. During malaxation, the crushed olive paste is mixed slowly to promote coalescence, thus improving the separation efficiency of the subsequent centrifugation. The most critical point of this step is the possible oxidation of the polyphenolic compounds, leading to an oil with lower sensory and nutritional properties as well as a reduction in shelf-life [197
Recently, Zinnai and co-workers set up an innovative system based on the direct addition of a cryogen (CO2,s
) to olives during pre-milling phase, observing positive effects on the concentration of polyphenols and vitamin E [190
Furthermore, in recent years, the development of new extraction methods based on the production of functional foods enriched with natural antioxidants has been demonstrated to be a promising potential application for the stabilization of olive oil and the increase of its shelf life [89
It is worth mentioning that, due to their healthful and nutritional effects, considerable attention has been recently focused on identifying natural sources of antioxidants and improving their extraction processes—in particular olive oil by-products [200
]—and fruit skin was also considered to produce enriched olive oils with an higher content of antioxidants compounds and, consequently, an improved nutraceutical profile [202
5.2. Main Parameters Affecting the Degradation Rate of Health Compounds During EVOO Storage
Generally speaking, during storage the olive oil chemical composition (i.e., MUFA/PUFA ratio and concentration of minor compounds such as polyphenols and carotenoids) is influenced mainly by the final balance between oxidative degradation and antioxidant activity due to the presence of both tocopherols and phenolic compounds. In this context the lipid fraction shows the highest sensitivity to oxidative degradation with the subsequent development of off-flavors caused by the production of carbonyl and aldehyde compounds and the final occurrence of the typical “oxidative rancidity”. In addition, auto-oxidation based on a free radical mechanism starting from the formation of hydroperoxides induced by the initial oxygen availability further improve the degradation rate of the stored olive oil.
While auto-oxidation can also be ruled out in the absence of light, this process appears to be accelerated by the action of natural photosensitizers such as chlorophyll, which reacts with triplet oxygen to form excited state singlet oxygen. In this context, the storage and packing conditions of olive oil become of primary importance [203
5.2.1. Influence of Storage Atmosphere
Until now, many experimental studies have been carried out to verify the real effectiveness of the use of inert gases (i.e., nitrogen) in the head-space of the containers to improve the stability and the shelf life of the stored olive oil, thus slowing down its oxidative changes [193
In a recent paper, Sanmartin and co-workers verified the possibility of using Ar and CO2
as head-space gases for the long-term storage of olive oil in order to slow down its oxidative degradation [174
]. After 250 days of storage in the dark at a controlled temperature (12 ± 1 °C), the authors concluded that replacing air with Ar or CO2
in the headspace of the container during storage can significantly reduce the oil oxidation rate, thus preserving, as much as possible, the compositional, nutritional, and organoleptic qualities of the oil. In regard to chemical composition, while at the end of the observation period, the oil stored under CO2
appeared to be very similar to that stored in Ar atmosphere, it was significantly different with regard to sensorial characteristics. In particular, CO2
determined a negative organoleptic interference that would not support its use for the long-term storage of EVOO. Therefore, Ar treatment appears to be the best solution alternative to nitrogen to preserve the quality of the EVOO over time.
5.2.2. Characteristics of Packaging and Storage Temperature
As discussed previously, among all the operating conditions that can influence the degradation rate of an olive oil, oxygen availability appears to be of primary importance, followed by the light exposure level. The presence of metal compounds must also be taken in account as they can play the role of activators of oxidative degradative reactions [173
], thus reducing the concentration of active health compounds.
It appears of primary importance, therefore, to carefully select the packaging materials with regard to the specific protection provided, together with the storage conditions to be adopted in order to preserve the nutraceutical features showed by the oil at the start of the storage time to the maximum extent.
The main characteristics of the most widely used packaging materials for the storage of olive oil, together with a description of their specific functionality in terms of olive oil preservation, are given in Table 5
In particular, metal containers can provide total protection against light, oxygen, and water vapor. In order to avoid the activation of oxidation by metallic catalysis, it is possible to opt for tin plate or tin-free steel based on chromium instead of aluminum or aluminum alloys. In addition, while the inside of the tin can be coated with resins to protect the metal surface against corrosion, particular attention should be paid, in this case, to the main concern related to the leaching of unsafe chemical compounds from food contact materials (FCM) into the stored oil. Glass represents a good barrier against moisture and gases without leaching [201
], but transparent bottles cannot protect the olive oil from photo-oxidation [203
]. For this reason, glass containing specific additives to significantly reduce the transmittance of light in the UV range have been created [205
To determine the effects of packaging on the commercial life of olive oil, several studies have been carried out, and different containers such as clear and dark bottles, polyethylene, and tin containers have been taken into consideration [203
], and the storage stability of oils in tin or stainless containers and in dark glass was the highest [203
Besides the type of packaging, storage temperature can also influence the degradation rate of stored olive oil [173
], obtaining a longer shelf life when a lower temperature was adopted during storage.
In a recent paper, Sanmartin and co-workers [173
] evaluated the effects of packaging and storage conditions on an EVOO as it occurs in most points of sale: the storage of oil in tanks under nitrogen for a more or less long time (also for several months), after which the oil is packaged and sold. Interestingly, st the end of the observation period, the authors observed that the storage conditions can not only prevent oxidation processes from occurring but they can even be usefully implemented to slow down or almost block these processes in the case of oil in which the oxidative processes had already started.