Enrichment of Olive Oils with Natural Bioactive Compounds from Aromatic and Medicinal Herbs: Phytochemical Analysis and Antioxidant Potential

Olive oil and herbs, two key components of the Mediterranean diet, are known for their beneficial effects on humans. In our study, we incorporated aromatic and medicinal herbs into local monovarietal olive oils via maceration procedures for enrichment. We identified the herbal-derived ingredients that migrate to olive oils and contribute positively to their total phenolic content and functional properties, such as radical scavenging activity. Thus, we characterized the essential oil composition of the aromatic herbs (GC-MS), and we determined the phenolic content and antioxidant capacity of the additives and the virgin olive oils before and after enrichment. The herbal phenolic compounds were analyzed by LC-LTQ/Orbitrap HRMS. We found that olive oils infused with Origanum vulgare ssp. hirtum, Rosmarinus officinalis and Salvia triloba obtained an increased phenolic content, by approximately 1.3 to 3.4 times, in comparison with the untreated ones. Infusion with S. triloba led to a significantly higher antioxidant capacity. Rosmarinic acid, as well as phenolic glucosides, identified in the aromatic herbs, were not incorporated into olive oils due to their high polarity. In contrast, phenolic aglycones and diterpenes from R. officinalis and S. triloba migrated to the enriched olive oils, leading to a significant increase in their phenolic content and to an improvement in their free radical scavenging capacity.


Introduction
Olive oil and aromatic herbs are important components of the Mediterranean diet, one of the healthiest diets worldwide [1].Both have beneficial effects on human health, attributed mainly to their bioactive plant secondary metabolites.Olive oil comprises a saponified fraction (97-99%), consisting mainly of glyceryl fatty acid esters (mainly oleic acid) and a small non-saponified one (1-3%), containing bioactive compounds that affect the olive oil shelf life and its organoleptic properties and provide health benefits.In fact, extra virgin olive oil has been characterized as a functional food with high nutraceutical value [2].On the other hand, aromatic plants, which are widely distributed in the Mediterranean region, constitute a specific category, producing essential oils.These compounds provide the characteristic odor and taste to the plant.Aromatic plants with medicinal properties are also known as medicinal and aromatic plants.Some species are used as herbs for flavoring foods but also as natural preservatives in the food industry due to their antimicrobial and antioxidant properties.The utilization of extra virgin olive oil and aromatic culinary herbs has been associated, among others, with lower mortality rates from cardiovascular disease, lower cancer indices due to their chemopreventive activity and a reduced risk from neurodegenerative diseases [3].The enrichment of olive oils with aromatic herbs, which is a traditional practice in the Mediterranean region, has had significant impact in the market in recent years.Except for the improvement of organoleptic properties, the addition of aromatic herbs is expected to have a positive effect on the nutritional value of the enriched olive oils, upgrading their biofunctional properties, mainly attributed to phenolic compounds [4].These belong to one of the main classes of secondary metabolites, which provide protection to the plant itself [5] by contributing to their adaptation under abiotic stress conditions and providing protection from biotic agents.These compounds are characterized by the presence of at least one phenolic hydroxyl group in their molecule.Their beneficial value in the human's body is mainly attributed to their ability to protect cells (or tissues) from oxidative stress damages.According to our best knowledge, little is known about the migration of phenolic compounds from aromatic plants to olive oils during the enrichment and enhancement of their biofunctional properties.The aim of this study was to identify the ingredients of selected aromatic herbs that contribute to an upgraded phenolic content and antioxidant capacity of the enriched olive oils.For this purpose, we incorporated selected aromatic herbs into monovarietal olive oils from the region of Epirus, Greece and determined the phenolic content and antioxidant capacity of the additives and the olive oils before and after the enrichment.In addition, we proceeded to the characterization of phenolic compounds migrating from plants to the olive oils.

Total Phenolic Content
The total phenolic compounds of the tested aromatic plants (Figure 1A) ranged from 128.40 ± 0.89 to 139.98 ± 1.87 mg GAE g −1 DW.R. officinalis and S. triloba showed a significantly higher yield in phenolic content in comparison to O. vulgare ssp.hirtum.Concerning the three types of olive oils used in the experiment, the values of the total phenolic compounds varied between 105.84 ± 15.26 and 187.85 ± 18.53 mg GAE g −1 DW.Higher values were noticed for "Lianoelia Prevezas" (Figure 1B) compared to "Konservoelia Artas" (Figure 1C) and "Lianoelia Kerkyras" (Figure 1D).It is noteworthy that the infusion with aromatic plants significantly enhanced their content in polyphenols from 233.65 ± 18.65 to 427.50 ± 10.78 mg GAE Kg −1 for "Lianoelia Prevezas", 172.68 ± 12.58 to 357.89 ± 13.66 mg GAE Kg −1 for "Konservoelia Artas" and 194.95 ± 15.61 to 375.40 ± 18.67 mg GAE Kg −1 for "Lianoelia Kerkyras" compared to the relevant untreated control olive oils.
According to the results of the DPPH assay, the antioxidant activity of the untreated and enriched extra virgin olive oils ranged from 132.6 ± 2.61 to 185.8 ± 4.1 and 167.3 ± 3.2 to 672.6 ± 22.2 mM gallic acid Kg −1 olive oil for the three olive oils, respectively (Table 5).All the aromatic plants used in the experiment were found to enhance the scavenging radical activity of the "Konservoelia Artas" (Figure 6A), "Lianoelia Prevezas" (Figure 6B) and "Lianoelia Kerkyras" (Figure 6C) olive oils.It is noted that the three infused olive oils differed in their antioxidant capacity.In particular, it was found that while O. vulgare ssp.hirtum enhanced the antioxidant capacity of the enriched olive oils, R. officinalis was relatively more effective, but the infused olive oils with S. triloba exhibited the highest antioxidant capacity.
According to the results of the DPPH assay, the antioxidant activity of the untreated and enriched extra virgin olive oils ranged from 132.6 ± 2.61 to 185.8 ± 4.1 and 167.3 ± 3.2 to 672.6 ± 22.2 mM gallic acid Kg −1 olive oil for the three olive oils, respectively (Table 5).All the aromatic plants used in the experiment were found to enhance the scavenging radical activity of the "Konservoelia Artas" (Figure 6A), "Lianoelia Prevezas" (Figure 6B) and "Lianoelia Kerkyras" (Figure 6C) olive oils.It is noted that the three infused olive oils differed in their antioxidant capacity.In particular, it was found that while O. vulgare ssp.hirtum enhanced the antioxidant capacity of the enriched olive oils, R. officinalis was relatively more effective, but the infused olive oils with S. triloba exhibited the highest antioxidant capacity.Table 5. Antioxidant activity of the untreated and infused olive oils as determined by DPPH assay.The mean values ± SD of IC50 values are expressed as mg oil and as mM GAE/Kg olive oil.Different letters within the same column state that the mean values present statistically significant differences (Duncan's test, p < 0.05).

Discussion
Foods, which, besides their basic ingredients and initial nutritional value, contain components of natural origin, providing health-beneficial properties to consumers and well-being, are characterized as biofunctional [10].Olive oil is characterized as functional food due to its biofunctional chemical composition.Spices and various aromatic herbs are widely used as flavoring agents in olive oil.Aromatization enhances the flavor of the enriched olive oil, increases its qualitative characteristics and expands its shelf-life.Besides the traditional aromatization of olive oil with aromatic herbs in the Mediterranean region, infused olive oils have been more frequently available on the market during the last few years.Most studies deal with the effects of the additives on the sensory characteristics of the enriched olive oils and the extension of their self-life during storage [8,[11][12][13][14][15].
The traditional procedure (maceration) for flavoring olive oils is to add the dry, ground aromatic herb into the olive oil and to keep it at room temperature.The mixture is frequently agitated in order to facilitate the diffusion of the compounds, while the flavored olive oil is obtained after filtration [12,16].Another practice is to first isolate the essential oil from the aromatic plant and to subsequently incorporate it into the olive oil [16].In this case, olive oil is enriched only with the plant volatile compounds, and the

Discussion
Foods, which, besides their basic ingredients and initial nutritional value, contain components of natural origin, providing health-beneficial properties to consumers and well-being, are characterized as biofunctional [10].Olive oil is characterized as functional food due to its biofunctional chemical composition.Spices and various aromatic herbs are widely used as flavoring agents in olive oil.Aromatization enhances the flavor of the enriched olive oil, increases its qualitative characteristics and expands its shelf-life.Besides the traditional aromatization of olive oil with aromatic herbs in the Mediterranean region, infused olive oils have been more frequently available on the market during the last few years.Most studies deal with the effects of the additives on the sensory characteristics of the enriched olive oils and the extension of their self-life during storage [8,[11][12][13][14][15].
The traditional procedure (maceration) for flavoring olive oils is to add the dry, ground aromatic herb into the olive oil and to keep it at room temperature.The mixture is frequently agitated in order to facilitate the diffusion of the compounds, while the flavored olive oil is obtained after filtration [12,16].Another practice is to first isolate the essential oil from the aromatic plant and to subsequently incorporate it into the olive oil [16].In this case, olive oil is enriched only with the plant volatile compounds, and the advantage is that there is no need to separate the plant material from the olive oil.Another proposed practice for flavoring is the addition of the plant material at the olive mills during the malaxation extraction process [12,16].The monovarietal olive oil samples used in this study were obtained from cultivated olive varieties of the region of Epirus, Greece from the production of 2022.Two samples belong to recognized geographical indication (PGI) varieties: "Lianoelia Prevezas", which is used to produce extra virgin olive oil, and "Konservoelia Artas", which is an edible olive cultivar.The "Konservoelia Artas" olive oil is produced from olives that do not fit to the market standards as edibles.The third sample of olive oil was obtained from the old Venetian origin variety "Lianoelia Kerkyras" grown in the region of Parga.In our study, infused olive oils with the selected aromatic herbs O. vulgare ssp.hirtum, R. officinalis and S. triloba were prepared through maceration.Studies on the migration of plant tissue ingredients to olive oils, enhancing their biofunctional potential, are limited [14].The aromatic herbs' bioactive constituents belong to the plant's secondary metabolites.Their chemical composition depends not only on the plant species that produces them but also on other factors, including the environmental conditions during plant growing, the stage of harvest and the drying and storage conditions.For this reason, we initially conducted phytochemical analysis of the plant material used in the present study.The selected aromatic plants with medicinal properties (Greek oregano, rosemary, Greek sage) are commonly used for food flavoring in the Mediterranean cuisine and are also used traditionally for their healing properties.As known, the bioactivity and medicinal properties of aromatic herbs are mainly attributed to the production of phenolic compounds [17] and volatile essential oil ingredients [18], which mainly belong to mono-and sesquiterpenes.The therapeutic value of the phenolic compounds is mainly attributed to their ability to protect from oxidative stress damage by different mechanisms due to the presence of at least one phenolic hydroxyl group [3].
First, we analyzed the methanol extracts of aromatic plants and we found higher levels in S. triloba and R. officinalis compared to O. vulgare ssp.hirtum.Although S. triloba και R. officinalis extracts presented an almost equal content of phenolic compounds, S. triloba possessed a higher antioxidant activity.This might be due to the structure of its phenolic compounds [19] or even due to synergistic or antagonistic effects with the ingredients of the methanol extract [20].GC-MS analysis of the essential oils revealed that, in O. vulgare, ssp.hirtum and S. triloba carvacrol and 1,8-cineole were most abundant.The monoterpenes camphor, α-pinene, borneol and verbenone were at almost equal percentages in R. officinalis essential oil.The high content in carvacrol (>80%), which is known to possess a significant free radical scavenging activity [21], contributes to the reactivity of O. vulgare ssp.hirtum essential oil toward the DPPH radical.On the other hand, S. triloba and R. officinalis essential oils showed a much weaker antioxidant activity.These findings indicate the presence of nonvolatile compounds with high antioxidant activities in the methanol extracts of S. triloba and R. officinalis.For the evaluation of our data, we used the antioxidant activity index (AAI) proposed by Scherer and Godoy (2009) [22] to avoid fluctuations in the expression of the results due to different reaction conditions used by researchers.According to this index-based classification, the antioxidant activities of Origanum, Salvia and Rosmarinus methanol extracts, as well as that of Origanum essential oil, were characterized as moderate.
Regarding the hydromethanolic extracts of the infused olive oils, our results showed that all the selected aromatic herbs enhanced the phenolic content and improved the antioxidant capacity of all the infused olive oils.In particular, the total phenols were increased by approximately 2.5 to 3.4 times in Origanum and 2.3 to 3.2 times in sageflavored olive oils, while, with rosemary, where the impact was weaker, the increase was about 1.3-1.6 times.
The greatest increase in the phenolic content of infused oils was observed for those with lower levels of phenolic compounds.Indeed, while the methanol extracts of R. officinalis and S. triloba presented a higher total phenolic content compared to the respective O. vulgare ssp.hirtum, we did not observe an equivalent increase in the hydromethanolic extracts of the relevant enriched olive oils.This could be due to a difference in the polarity of phenolic compounds, depending upon the number of hydroxyl groups attached to the aromatic ring [23], that influenced their solubility both in the olive oil as well as in the hydromethanolic mixtures used for their recovery from the infused olive oils.
However, it was remarkable that mainly the S. triloba-enriched olive oils and, subsequently, R. officinalis, presented significantly higher antioxidant activity compared to the O. vulgare ssp.hirtum-enriched ones.It is mentioned that the determination of the radical scavenging activity toward DPPH was conducted on the olive oil per se, and not on the revealed hydromethanolic extract, meaning that the sample contained all the ingredients of the enriched olive oils.
The above-mentioned results indicate that the aromatic herbs S. triloba and R. officinalis contain less polar phenolic ingredients that do not migrate to the hydromethanolic phase during the extraction of the infused oils (where the determination of total phenolic content takes place), but are present in the crude infused olive oils.
These observations are in consistency with the results of the LC-MS analysis used for the identification of the phenolic compounds.For example, rosmarinic acid, a polar phenolic acid compound with a high DPPH free radical scavenging activity [24], was identified in all the methanol extracts of the selected herbs used in this study, but it was not detected in the enriched olive oils.In contrast, salicylic acid, an O. vulgare ssp.hirtum phenolic acid with moderate polarity, migrated into the infused olive oils, enhancing their phenolic content.None of the phenolic acid glucosides were extracted into the olive oils.On the contrary, all the phenolic aglycones identified in the extracts of the aromatic herbs utilized, as well as the phenolic diterpenes present in R. officinalis και S. triloba, significantly increased the phenolic content of the enriched olive oils and contributed to their radical scavenging activity.
The phenolic compounds identified by LC-MS belong to the classes of phenolic acids, flavonoids and phenolic diterpenes.Bibliographic data suggest that specific phenolic compounds that have been established to migrate from the plant material to the olive oil may exhibit additional biofunctional properties, apart from their antioxidant activity.Reports mention the anti-inflammatory and anti-tumor properties of the flavones apigenin [25], genkwanin [26], luteolin [27], the flavanone eriodictyol [28], the flavanonol taxifolin [29], the flavonol quercetin [30], the phenolic diterpenes carnosol and carnosic acid [31] and rosmanol [32].Among other beneficial properties, luteolin also has anti-diabetic, antihypertensive, anti-asthmatic and anti-viral properties [27].Several studies demonstrate the beneficial effect of genkwanin against many diseases, including cardiometabolic diseases, type 2 diabetes and neurodegenerative disorders.Taxifolin also shows antimicrobial, cardiovascular hepatoprotective, anti-Alzheimer and antiangiogenic properties [29].Naringenin has also been mentioned to possess antidiabetic, antibacterial, gastroprotective, immunomodulator, cardioprotective, nephroprotective and neuroprotective effects [33].The bioactive flavonol quercetin exerts a variety of health-beneficial effects, such as antihypertensive, anti-hypercholesterolemic and anti-atherosclerotic, as well as neuroprotective, antibacterial, antiviral and antiallergic activities, and provides protection against cardiovascular diseases [30].

Plant Material and Olive Oils
The aromatic plants were collected from the region of Epirus, Greece.The aerial part of Origanum vulgare ssp.hirtum was collected at the flowering stage (municipality of Ziros, Preveza, Greece) and Rosmarinus officinalis shoots at the end of the autumn flowering period, (municipality of Arta, Arta, Greece), while shoots of Salvia triloba were collected after the fruiting period (municipality of Hygoumenitsa, Thesprotia, Greece).Voucher specimens are kept at the herbarium of the OPENSCREEN-GR infrastructure, at the University of Ioannina-Greece.The plant material was dried by freeze-drying process (lyophilization) and pulverized using an appropriate mixing and milling equipment (Polymix, Kinematica, Bohemia, NY, USA) to particles less than 2 mm.The Protective Geographical Indication (PGI) packaged olive oil "Lianoelia Prevezas" was provided by the olive oil mills of "Zalongo" and the extract of the Protective Geographical Indication (PGI) olives "Konservoelia Artas" olive oil was provided by a local olive mill (municipality Nikolaos Skoufas, Arta, Greece), while a sample of olive oil derived by the variety Lianoelia Kerkyras, grown in the region of Parga (municipality of Parga, Preveza, Greece), was provided by a producer.

Preparation of Infused Olive Oils
The infused olive oils were prepared by natural maceration for 30 days.In more detail, the dried and ground aromatic plant (0.25 g) was added to the olive oil sample (5 mL) in a screw-capped glass tube and was vigorously agitated for 1 min and kept for one month in the dark at room temperature.Olive oil samples without additives (untreated olive oils), stored in the same conditions, were used as control samples.After the extraction period, the enriched olive oils were obtained by centrifugation (2700× g, 25 min).

Aromatic Plant Extracts
The pulverized dry plant material was extracted with methanol (10 mg mL −1 ) for 10 min × 3 using an ultrasound bath (Badelin Electronic, Berlin, Germany) as previously described [34], aliquoted and stored at −20 • C until use.The essential oil was isolated by hydro-distillation (2 h) using a Clevenger-type apparatus.Subsequently, the obtained essential oil was dried over anhydrous sodium sulfate (NaSO 4 ) and stored at −20 • C until use.

Extraction of Untreated and Infused Olive Oils
Liquid-liquid extraction using MeOH-H 2 O (80:20, v/v) as solvent was applied in order to receive the phenolic fraction from the olive oil's samples according to the International Olive Council method (2009) [35].In brief, 2.0 g of olive oil and 5 mL of MeOH-H 2 O (80:20, v/v) were mixed in a screw-capped glass tube under agitation for 1 min.Subsequently, the tube was sonicated with an ultrasonic bath (3 × 5 min) and the MeOH-H 2 O phase was recovered by centrifugation (2700× g, 25 min).

Phytochemical Analysis 4.5.1. GC-MS Analysis of the Essential Oil
The essential oils, isolated from the aromatic plants, were analyzed by a gas chromatograph (GC) coupled with a mass spectrometer (MS) (GC-2030, GCMS-QPSERIES, Shimadzu, Kyoto, Japan).The essential oil constituents were separated using a Mega 5-MS capillary column (30 m × 0.25 mm × 0.25 µm), helium as carrier gas with a flow rate of 0.9 mL min −1 and a temperature program, previously described [36].For the analysis, 1 µL of the essential oil was diluted in n-hexane (1:200) and then injected by autosampler (AOC-20i/s Shimadzu), while the injector was set at split mode (split ratio: 30).Each sample was analyzed twice.The injector, interface and ion source temperatures were set at 250 • C, 300 • C and 240 • C, respectively, while the mass spectrometer was operated at the electron ionization mode (70 eV), with the mass scan ranging from 50 to 550 amu, with 0.5 spectra s −1 acquisition rate.The compounds were identified based on the similarity of the acquired mass spectra with the Nist Library data and the comparison of their retention indices (RIs) relative to n-alkanes (C8-C20), with data obtained through the literature.The contribution of each identified ingredient to the essential oil was calculated as a percentage (%) of the total compounds.

LC-LTQ/Orbitrap HRMS Analysis
An ultra-high-performance liquid chromatography (UHPLC) system coupled to a linear trap quadrupole (LTQ)/Orbitrap HRMS detector was employed for the identification of the phenolic compounds in all extracts.It was equipped with an autosampler (Accela AS autosampler model 2.1.1),an automatic sample flow pump (Accela quaternary gradient U-HPLC-pump model 1.05.0900) and a hybrid LTQ/Orbitrap XL 2.5.5 SP1 mass spectrometer from Thermo Fisher Scientific (Bremen, Germany).An Ion Max electrospray ionization (ESI) probe was included in the system.Compounds' separation was performed on a reversed-phase Hypersil GOLD analytical column (100 mm × 2.1 mm, 1.9 µm) from Thermo (Bremen, Germany).The mobile phase consisted of a phase A (0.1% formic acid in water) and phase B (methanol), following a gradient elution program.Flow rate was set at 0.4 mL min −1 .The injection volume was 10 µL, while the tray and oven temperatures were set at 15 and 40 • C, respectively.The system operated in negative ionization mode at a mass range of 100-1000 m/z.The ESI source conditions were: 55 and 20 arbitrary units (au) of sheath gas and aux gas flow rates, respectively; 350 • C capillary temperature; spray voltage, 2.7 kV. Full-scan mass spectra of high resolution were acquired in the Orbitrap analyzer with data-dependent MS/MS mode with parallel acquisition of top 6 intense ions scanned in the linear ion trap.A normalized collision energy (NCE) of 35% was used throughout the analysis (CID, collision induced dissociation) to obtain the compounds' fragmentation pattern.The phenolic compounds were identified on the basis of their molecular ion formation and their characteristic fragments were compared to either the existing literature or to the NIST Mass Spectral Library 2020.The instruments' control and the mass spectra processing were carried out with Xcalibur v.2.2 software (Thermo Electron, San Jose, CA, USA).

Free Radical Scavenging Capacity Assay
The determination of the free radical scavenging activity (RSA) of the aromatic plant extracts and the essential oil samples was performed according to Conforti et al. (2008) [37], after adaptation.Initially, the extract or the essential oil was diluted in methanol at concentrations ranging from 0.5 to 6.0 mg mL −1 for the essential oils and 0.05 to 0.40 mg mL −1 for the MeOH extracts.Subsequently, 100 µL of the solutions was added to 900 µL DPPH solution (0.1 mM final concentration in DPPH), the mixture was vigorously agitated and the absorbance was measured at 517 nm after staying for 30 min in the dark.Regarding the untreated and infused olive oils, the DPPH assay was performed as described by Minioti et al. (2010) [38].In brief, olive oil samples (20, 80, 120 or 180 mg) were added to a DPPH solution in ethyl acetate (0.1 mM final concentration in DPPH).The mixture was vigorously agitated and, after 1 h in the dark, the absorbance was measured against blank (ethyl acetate solution) at 515 nm.Standard curve of gallic acid, used as a positive control, was plotted for concentrations ranging from 0.5 to 5.0 mg/mL of gallic acid.The scavenging activity was calculated by the following formula, using the absorbance of the DPPH solution (without the oil sample) as control absorbance.% DPPH scavenging activity = [(control absorbance − sample absorbance)/control absorbance] × 100 The concentration of the aromatic plant extracts, or the amount of the olive oil samples required to achieve 50% inhibition of the DPPH free radical activity (IC 50 ), was calculated from a plot of percent inhibition (%) against the sample concentration.The total antioxidant capacity of the olive oil samples was expressed as mL −1 gallic acid equivalent (GAE) kg −1 olive oil using the gallic acid standard curve.The antioxidant activity index (AAI) for the aromatic plant extracts was calculated through the following equation (Scherer and Godoy, 2009) [22]: AAI = final concentration of DPPH µg mL −1 /IC 50 µg mL −1 .

Figure 2 .
Figure 2. Phenolic aglycons migrating from the aromatic herbs to the olive oils analyzed by LC-LTQ/Orbitrap HRMS.

Figure 3 .
Figure 3. Diterpenic phenols migrating from the aromatic herbs to the olive oils analyzed by LC-LTQ/Orbitrap HRMS.

Figure 3 .
Figure 3. Diterpenic phenols migrating from the aromatic herbs to the olive oils analyzed by LC-LTQ/Orbitrap HRMS.

Figure 3 .
Figure 3. Diterpenic phenols migrating from the aromatic herbs to the olive oils analyzed by LC-LTQ/Orbitrap HRMS.

Figure 4 .
Figure 4. Antioxidant capacity of the methanolic extract obtained by the selected aromatic plants St: S. triloba, Ov: O. vulgare ssp.hirtum, Ro: R. officinalis, as assessed by DPPH assay.Values are expressed as means of four experiments.

Figure 4 .
Figure 4. Antioxidant capacity of the methanolic extract obtained by the selected aromatic plants St: S. triloba, Ov: O. vulgare ssp.hirtum, Ro: R. officinalis, as assessed by DPPH assay.Values are expressed as means of four experiments.

Figure 5 .
Figure 5. IC50 values of the selected aromatic plant species for radical scavenging.Ov: O. vulgare ssp.hirtum, Ro: R. officinalis, St: S. triloba, methanolic extracts.Values are expressed as mean ± SD of four experiments.Means with different letter indications are significantly different (Duncan's test, p < 0.05).

Figure 5 .
Figure 5. IC 50 values of the selected aromatic plant species for radical scavenging.Ov: O. vulgare ssp.hirtum, Ro: R. officinalis, St: S. triloba, methanolic extracts.Values are expressed as mean ± SD of four experiments.Means with different letter indications are significantly different (Duncan's test, p < 0.05).

Table 1 .
Chemical composition of essential oils analyzed by GC-MS a .

Table 2 .
Phenolic compounds in the extracts # of O. vulgare ssp.hirtum, and the relevant enriched olive oils, as identified by LC-LTQ/Orbitrap HRMS.

Table 3 .
Phenolic compounds in the extracts # of R. officinalis and the relevant enriched olive oils, as identified by LC-LTQ/Orbitrap HRMS.

Table 4 .
Phenolic compounds in the extracts # of S. triloba and the respective enriched olive oils, as identified by LC-LTQ/Orbitrap HRMS.

Table 5 .
Antioxidant activity of the untreated and infused olive oils as determined by DPPH assay.The mean values ± SD of IC 50 values are expressed as mg oil and as mM GAE/Kg olive oil.Different letters within the same column state that the mean values present statistically significant differences (Duncan's test, p < 0.05).