Unveiling the antioxidant and anticancer potential of local mediterranean olive and fig extracts against breast cancer

Sara Al Srouji; Walid El Kayal; Diana Ismail; Lara Nasreddine; Hala Gali-Muhtasib; Nahla Hwalla

doi:10.1186/s12906-026-05296-4

. 2026 Feb 20;26:116. doi: 10.1186/s12906-026-05296-4

Unveiling the antioxidant and anticancer potential of local mediterranean olive and fig extracts against breast cancer

Sara Al Srouji ^1,^#, Walid El Kayal ², Diana Ismail ³, Lara Nasreddine ¹, Hala Gali-Muhtasib ^3,^✉,^#, Nahla Hwalla ^1,^✉,^#

PMCID: PMC13032439 PMID: 41715047

Abstract

Given the interest in dietary interventions against cancer, this study investigated the antioxidant and anticancer potential of nutraceuticals from key components of the mediterranean diet (MD) against breast cancer considering their rich polyphenolic content and antioxidant profile. Extracts from three Lebanese Ficus carica (FC) varieties Shatawi (Sh), Baqarati (Bq), Asali (As), and two local Olea europea (OE) varieties Green olive (GO) and Black olive (BO) were selected for evaluation based on their traditional dietary relevance and potential bioactivity. Methanolic extracts of these components were analyzed for their total polyphenolic content (TPC), anthocyanin content (TAC), and antioxidant activity. The anticancer effects of the single and combined agents were evaluated on 2 breast cancer cell lines (MCF-7 and MDA-MB-231) and non-tumorigenic intestinal epithelial cell line (FHs-74) using MTT assay and flow cytometry. FC and OE extracts were tested at concentration ranges (2–30 mg/ml and 0.3–2.4 mg/ml), respectively. We showed that OE extracts, particularly from BO, exhibited significantly higher TPC and three-times greater TAC than FC extracts. Compared to all FC extracts, BO extract demonstrated strong anticancer effects at 100-fold lower concentrations, especially against MCF-7 cells. Importantly, BO did not affect the viability of normal FHs-74 cells at doses up to 1.2 mg/mL. Interestingly, combination of FC and OE extracts particularly darker- colored varieties with higher TPC (Sh and BO) resulted in an enhanced 79% and 64% reduction of respectively MCF-7 and MDA-MB-231 BC cell line viability compared to single treatment. This dual treatment induced a greater increase in sub-G1 populations in MCF-7 cancer cells compared to BO alone. These findings highlight the therapeutic potential of Mediterranean fruit extracts, particularly BO and Sh, positioning them as promising candidates to support breast cancer treatment strategies.

Supplementary Information

The online version contains supplementary material available at 10.1186/s12906-026-05296-4.

Keywords: Mediterranean diet, Breast Cancer, Polyphenols, Ficus carica, Olea europea, Cell viability, Synergism

Introduction

According to the World Health Statistics 2023 report, breast cancer (BC) has emerged as the fastest-growing cancer worldwide, posing a significant threat to public health [1]. The WHO reported that BC was responsible for more than 600,000 deaths globally in 2022 [2]. It also showed that population growth and aging alone are projected to lead to nearly 3 million new instances of BC and 1 million deaths annually by 2040 [3]. In the Middle east and north African region, Lebanon, an eastern mediterranean country stands out as having the highest age standardized rate of BC incidence (68.9%) among 21 countries; based on a meta-analysis of reports spanning from 2011 to 2019 [4].

Given the serious adverse effects associated with the synthetic anticancer chemotherapeutic agents, along with the poor responsiveness and sensitivity of triple negative breast cancer cells [5], extensive research efforts have been devoted to identifying adjuvant therapeutic strategies, particularly those that harness the therapeutic potential of medicinal plants. In this context, the Mediterranean culture's long-standing reliance on medicinal plants for treating various ailments provides a well-established foundation for exploring their therapeutic applications against cancer [6, 7]. In line with this perspective, the Mediterranean diet (MD) has been recognized as the best overall Diet according to U.S. News due to its high polyphenol and bioactive compounds content which interfere with the multistage carcinogenesis process [8, 9]. In particular, consumption of flavonoids such as resveratrol, curcumin, lycopene, quercetin, and oleic acid, that are commonly found in fruits and vegetables legumes, and extra virgin olive oil, has been associated with a significant reduction in the incidence of aggressive mammary neoplasms [10]. Moreover, the MD has revealed a superior preventive effect from BC among post-menopausal women upon adhering to a supplemented MD with extra-virgin olive oil compared to a low fat control group [11].

Among the key traditional ingredients of the MD, extra virgin olive oil and olives, have attracted significant attention in cancer research due to their potential therapeutic properties. They are rich in anthocyanins, flavanols, flavones, phenolic acids, phenolic alcohols, secoiridoids, and lignans content [12–14]. In cancer research, major compounds in olives and olive oil such as oleocanthal and oleuropein have been proven to induce apoptosis and decrease leukemia cancer cell viability [15]. Moreover, hydroxytyrosol, a derivative of oleuropein was shown to interact with copper, effectively inhibiting the spread of triple-negative breast cancer [16]. However, compared to olive oil, the olive fruit holds higher total phenolic contents, due to the loss of water-soluble antioxidants in the wash-water step in the extraction process [17] highlighting the importance of the use of the whole fruit.

Another key traditional ingredient of the MD is Ficus, with Ficus carica (FC) being the most commonly consumed in the region and widely recognized for its numerous health benefits. Notably, the highest total polyphenolic content is found in the skin and peel of the FC fruit [18, 19]. Given that this part is also highly edible, it is essential to consider the whole fruit, rather than focusing on a single component, as emphasized by previous studies [20]. Major polyphenols found in FC fruit are apigenin, luteolin, quercetin, hesperetin, prenylated, isoflavones, morin, anthocyanins, catechins, and γ-tocopherols which exhibit anti-inflammatory properties [20, 21].

Although numerous studies have examined the anticancer properties of fig leaf and latex [22–25], olive oil and olive leaf extracts [26, 27] in various tumors, the effects of FC and OE whole fruit extracts on breast cancer remain very limited. To the best of our knowledge, this is the first study to examine the antioxidant and anticancer effects of Lebanese FC and OE variety extracts on BC in single and dual treatment.

In view of the above, we aimed to evaluate the anticancer potential of three Lebanese varieties of FC fruits, Shatawi (Sh), Baqarati (Bq), and Asali (As), as well as two Lebanese varieties of Olea europaea (OE) fruits, green (GO) and black olives (BO), using the whole fruit in both single and combination treatments against BC. We evaluated the total phenolic and anthocyanin contents of the methanolic extract of each fruit and determined their antioxidant activity. We further evaluated their inhibitory effects on cell viability and cell cycle progression in human breast cancer cell lines, MCF-7 and MDA-MB-231 (triple-negative), using both single and combination treatments.

Taking that these fruits are highly consumed in our region, and an integral part of our diet, understanding the mechanism of action of these extracts will provide a potential strategy to BC adjunctive treatment strategy.

Materials and methods

Fruit material and extraction preparation

Fruit material

Three different local varieties of FC fruits were handpicked on from Baalbek (Lebanon) in October 2023, namely: Sh, Bq, and As. Whereas the two varieties of OE fruit, GO and BO were collected from Kfeir, Hasbaya (Lebanon) in November 2023. FC and OE fruits were washed with distilled water and gently dried with a napkin. As for assessing the ripeness, Dr. Walid El Kayal checked for the fruit’s firmness, external color, and maturity stage. Permission to collect both samples of both fruits was granted by the landowners. For the pre-extraction treatment, the fresh fruits were first assessed for ripeness and inspected for any visible defects. The samples were then divided into batches, wrapped in aluminum foil, labeled with the variety name and date, and immersed in liquid nitrogen for 2 min. Immediately after removal, the samples were stored at −80 °C for a couple of days. Prior to processing, the samples were taken out of the freezer, ensuring they did not thaw, and were once again immersed in liquid nitrogen. Finally, they were transferred to a dry freezer (LABCONCO, FreeZone2.5, MO, US), where they were kept for 2–4 days until completely dried. Samples of freeze-dried figs and olives are available at Dr. Walid El Kayal’s lab at the Department of Agriculture at the American University of Beirut.

Extraction process

The extraction process was carried out as described previously by Darwiche et al. (2023) [28] with slight modifications. Briefly, 10 g of each variety of olive and fig were ground separately using a mortar and pestle, and an electric grinder/homogenizer machine (Millmix 20, Domel, Slovenia), respectively to obtain a fine powder. This powder was then mixed with 80% aqueous methanol (1:10 weight/volume) and subjected to periodic shaking in the dark for 24 h via an orbital shaker-incubator ES-20 (Biosan SIA., Latvia). The resulting mixture was filtered using Whatman filter paper no. 1. The purified filtrates were concentrated using a rotary evaporator at 40 °C and reduced pressure, followed by a second freeze-drying step, resulting in a 20% extraction yield. After that, the samples were diluted in PBS to a stock concentration of 200 and 400 mg/mL for OE and FC extracts, respectively. The finished methanolic extracts were stored at 4 °C in sealed, dark containers until the start of the experiment.

Quantitative phytochemical analysis in extracts

Total Polyphenol Content (TPC) Quantification

To determine the TPC of OE and FC extracts, the Folin–Ciocalteu (F–C) colorimetric method was performed using the BQC KB03006 polyphenol quantification assay kit (BQC Redox Technologies, Asturias, Spain) following the manufacturer’s protocol. Briefly, 20 μL of each extract is combined with 100 μL Working solution (F–C reagent) and 80 μL Solution B (Sodium Bicarbonate). The phenolic compound in the extract is oxidized to release an electron producing a reduced form of F–C reagent whose absorbance is measured at 700 nm [29]. Gallic acid was used as the reference standard in this assay and results were expressed as µg gallic acid equivalent (GAE) per ml.

Total anthocyanins content (TAC) quantification

Briefly, 20 μL of each extract were combined with either specific concentration of Reagent A (Hydrochloric Acid) or Reagent B (Sodium phosphate monobasic dihydrate) in 96-well clear bottom plates referring to the manufacturer’s instructions. Subsequently, the plates were kept at room temperature for 10 min, after which absorbance reading was taken at 510 and 700 nm using a multimode microplate reader (ThermoFisher Scientific, USA, Varioskan LUX), known as the PH differential method. The recorded TAC values represent the mean of three replicate measurements and results were expressed as cyanidin 3-glucoside equivalents (mg/L).

Evaluation of antioxidant activity in extracts

Ferric Reducing Antioxidant Power (FRAP) and DPPH Radical Scavenging assays were used (BioQuoChem, Asturias, Spain) using standardized kits. Both assays are indicated to examine the activity of polyphenols as primary antioxidants [30]. Antioxidant potential was measured at increasing concentrations: 0.3, 0.6, 0.9, 1.2, 1.6, 2.4, 2.6 mg/mL for OE extracts, and 2, 5, 7, 10, 20, 30 mg/mL for FC extracts.

Ferric Ion Reducing antioxidant potential (FRAP)

Briefly, add 10 μL of each of the extracts to 220 μL of Reagent A ((NaAc/ddH2O/TPZ/HCl/FeCl3). Absorbance was measured at 593 nm after 4 min at RT, and antioxidant potential of samples was determined based on iron standard curve (R2 = 0.9929) and expressed as µM of iron (II) equivalent. Mechanistically, FRAP incorporates a SET [31] reaction, explained in Eq. (1) below:

2-diphenyl-1-picrylhydrazyl (DPPH)

This antioxidant activity is determined by measuring the absorbance at 517 nm, and it is expressed as Trolox equivalent antioxidant capacity (TEAC, µM). Antioxidant capacity is reported as % Inhibition (Radical scavenging activity). The antioxidants in the extracts can either donate an electron or H-atom to the DPPH radical through SET or HAT mechanism [31] as shown in Eq. (2) and (3), respectively:

Readings of the absorbance for TPC, TAC, FRAP, and DPPH was done using ELISA reader (Multiskan Ex, Thermo Fisher Scientific, Waltham, MA, USA).

Cell culture

Chemicals and reagents

The MCF-7, and MDA-MB-231 breast cancer cells as well as the FHs-74, non-tumorigenic intestinal epithelial cells were purchased from American Type Culture Collection ((ATCC), USA). Dulbecco’s modified Eagle medium (DMEM-High glucose), Gibco Dulbecco's Modified Eagle Medium: Nutrient Mixture F12 (DMEM/F12) media, epidermal growth factor (EGF), sodium pyruvate, Minimum Essential Media (MEM), fetal bovine serum (FBS), trypsin–EDTA, Dulbecco’s Phosphate Buffer Saline (PBS), DMSO, MTT (3-(4,5-Dimethylthiazol-2-yl)−2,5-diphenyltetrazolium bromide), and 1 mg/mL propidium iodide (PI) were purchased from Sigma Aldrich (Sigma-Aldrich, St. Louis, MO, USA). Penicillin/Streptomycin (P/S) was purchased from Biowest (Biowest, France), and 10 mg/mL RNase was purchased from Thermofisher Scientific (Carlsbad, CA, United States).

Cell culture conditions

The cancer cell model includes breast cancer cells (MCF-7 and MDA-MB-231) which were cultured in DMEM-High media. The FHs 74, non-tumorigenic intestinal epithelial cells were cultured in DMEM/F12 media containing 1 mL EGF, 1% sodium pyruvate, and 1% MEM. Culture media supplemented with 10% FBS and 1% P/S. Cells were maintained in an incubator at 37ºC and humidified atmosphere (5% CO₂, 95% O₂). Cells were monitored daily and sub-cultured weekly upon reaching 80–85% confluency.

Cell viability assay (MTT)

The antiproliferative capacity of fig and olive extracts on breast cancer cell lines was assessed by MTT [3-(4,5-dimethylthiazol-2-yl)−2,5-diphenyltetrazolium bromide]) assay. MCF-7, MDA-MB-231, and FHs-74 cells were seeded at 8 × 10³, 5 × 10³, and 2 × 10⁴ seeding density in 96 well plates, respectively. They were then treated with the extracts with a range of corresponding concentrations for 24 h: 0.3, 0.6, 0.9, 1.2, 1.6, 2.4, 2.6 mg/mL for OE extracts, and 2, 5, 7, 10, 20, 30 mg/mL for FC extracts. This process was done in sextuplicate (technical replicates) for each treatment condition and repeated three times (biological replicates). After 24 h treatment period, media was discarded and replaced with 50 µL of MTT solution and 50 µL of serum-free media DMEM (for MCF-7) or DMEM/F12 (for FHs 74). MTT containing media was removed after 3 h incubation at 37 °C in the dark. Subsequently, a volume of 150 µL of MTT solvent (DMSO) was added to each well after which the optical density (O.D) was read at 595 nm using a spectrophotometer (ELISA reader (Multiskan Ex, Thermo Fisher Scientific, Waltham, MA, USA). The results were expressed as percentage of viable cells with respect to the untreated control using this formula:

To quantify the selectivity of a compound for its anticancer activity (ecreasing cell viability in this case) without causing undue toxicity, we have calculated an estimated selectivity index (SI) [32]. SI is often expressed as the ratio of the toxic dose to the effective dose (SI = (IC50 in FHs-74/IC50 in BC cell line).

Combination Index analysis

Combination Index (CI) is a tool generated by CompuSyn software (CompuSyn, Inc. Paramus, NJ, United States) that integrates equations based on calculations [33] (Version1.0). This tool identifies whether dual treatment or drug combinations works in a synergism (if CI < 1) or antagonism (CI > 1) or simply have an additive effect (CI = 1). The study investigated the effect of Sh at concentrations of 10, 20, and 30 mg/ml, in combination with BO at 0.6, 1.2, and 2.4 mg/ml, on BC cells.

Cell cycle analysis

MCF-7 cells were seeded at 10⁶ cells/mL in T25 flasks and kept to reach 50% confluency by the following day. Cells were treated with appropriate concentrations of each extract. All live and dead cells were collected, fixed with 70% cold ethanol for a duration of 6–7 days at −20 °C. Cells were then washed with PBS to remove any traces of ethanol and incubated with 100 µL RNase (0.2 mg/ml) for 40–45 min at 37 °C. Subsequently, a mixture of 20 µL propidium iodide (PI) and 400 µL cold PBS solution was added in the dark and cells were incubated for 1 h at 4 °C. Cell cycle analysis was then performed using Guava EasyCyte8 Flow Cytometer-Millipore. Data of the cell cycle was analyzed by GuavaSoft™ 2.7 Software to quantify the distribution of cells in pre-G1, G1, S, and G2/M phases. It is important to note that extracts were filtered using a filter syringe before treatment on cells.

Statistical analysis

Data are presented as mean ± standard error of mean (SEM) of three independent replicates, unless specified otherwise. Statistical analysis was performed by One-way ANOVA for comparing TPC and TAC, Two-way ANOVA (or mixed model, Bonferroni) when comparing different varieties of the same group using Graph Pad Prism V.7. Software (La Jolla, CA, United States). Pearson’s correlation coefficient (r) was used to measure correlations between variables (TPC, FRAP, DPPH, % Viability). Statistical significance was determined at 95% Confidence Interval (CI) with p < 0.05.

Results

Total polyphenolic content of OE and FC varieties

To determine the TPC of OE and FC varieties, an extract concentration of 10 mg/ml was analyzed using the Folin–Ciocalteu assay. Our results showed that the highest TPC was observed in BO and GO extracts, with concentrations of 45.634 ± 4.765 and 41.481 ± 1.001 µg GAE/mg, respectively. These TPC values were significantly higher than those observed in the FC extracts (p < 0.001), as illustrated in Fig. 1. Within the FC varieties, Sh extract had a relatively higher TPC (9.028 ± 2.584 µg GAE/mg) compared to Bq extract (7.834 ± 1.458 µg GAE/mg) and As (5.052 ± 1.671 µg GAE/mg).

Fig. 1 — Higher Total Phenolic Content (TPC) recorded for OE varieties compared to FC varieties at 10 mg/mg. TPC was determined spectrophotometrically by using Folin Ciocalteu assay and expressed as micrograms of gallic acid equivalents per milliliter of extracts (µg GAE/mL). The experiment was performed in triplicates and data are represented as mean ± SEM (*** p < 0.001, ns denotes non-significant differences (p > 0.05))

Total anthocyanin content of OE and FC varieties

Given the role of anthocyanins in inhibiting cancer proliferation, we also investigated the TAC in the different extracts. Our results showed that the BO extract had the highest TAC among all the five tested extracts (p < 0.001) (Fig. 2). They also reported that among the FC extracts, Sh and Bq extracts exhibited similar TAC, with values of 1.71 ± 0.09 and 1.038 ± 0.135 mg C6GE/L, respectively. These values were significantly higher than that in As (0.193 ± 0.079 mg C6GE/L) and GO (0.082 ± 0.002 mg C6GE/L). It is important to note that GO showed little to no AC.

Fig. 2 — Highest Total Anthocyanin Content (TAC) recorded for BO among the other FC and OE extracts at 10 g/mL dry weight. TAC was determined using anthocyanin quantification assay and expressed as milligrams of cyanidin-6-O-glucoside equivalent per liter of extracts (mg C6GE/L). The experiment was performed in triplicates and data are represented as mean ± SEM (**p < 0.01, *** p < 0.001, ns denotes non-significant difference (p > 0.05))

Antioxidant activity

Ferric reducing antioxidant power

To evaluate the electron-donating ability and overall antioxidant potential of the extracts, the FRAP assay was performed. A dose-dependent increase in FRAP capacity was observed across all extracts, indicating their capacity to act as effective reducing agents (Fig. 3).

Fig. 3 — Dose-dependent increase in FRAP capacity/antioxidant activity with increasing concentrations of (A) Shatawi, (B) Baqarati, (C) Asali, (D) Blak olives, and (E) Green olives. Antioxidant activity was determined using fast FRAP assay kit. The experiment was performed in triplicates and data are represented as mean ± SEM

Among the FC group, Sh and Bq extracts had significantly higher ferric reducing antioxidant capacity than As at concentrations ≥ 5 mg/mL. For instance, the antioxidant activity for Sh and Bq extracts at 30 mg/mL were reported as 1244 ± 37.5 and 1180 ± 23.17 (µM Fe²⁺), respectively, compared to As (500 ± 23.37 µM Fe²⁺) at the same dose (p < 0.001) (Fig. 4 A). As for the OE group, the antioxidant capacity of BO was significantly higher than that of GO at extract concentrations of 0.9 mg/mL and above (p < 0.001). For example, at an extract concentration of 2.4 mg/mL, the antioxidant capacity was 3508 ± 36.25 µM Fe²⁺ for BO and 2908 ± 143 µM Fe²⁺ for GO (Fig. 4 B).

Fig. 4 — BO, Sh, and Bq extracts exhibited higher FRAP capacity among FC and OE variety extracts. FRAP capacity of extracts was measured using fast FRAP assay at increasing concentrations (mg/mL) of (A) FC variety extracts and (B) OE variety extracts. The experiment was performed in triplicates and data are represented as mean ± SEM (*p < 0.05, *** p < 0.001)

Overall, OE variety extracts (BO, GO) had significantly higher antioxidant activity than all Sh, Bq, and As extracts (p < 0.005) (Fig. 5).

DPPH Radical scavenging radical activity

The antioxidant potential of the extracts was assessed using the DPPH assay, which measures the free radical scavenging activity, expressed as percentage inhibition of DPPH radical. The results show that Sh, Bq, and As extracts exhibited similar antioxidant potential. However, BO exhibited significantly higher percentage inhibition compared to GO at all the tested concentrations (0–2.4 mg/mL). These values are higher than the percentage inhibition of all FC varieties, which showed only 28.3 ± 2.60% inhibition at a much higher concentration of 30 mg/mL (p < 0.05) (Fig. 6).

Fig. 6 — OE extracts exhibited higher DPPH radical scavenging activity compared to FC extracts. Assessment of the radical scavenging activity of various concentrations of FC and OE extracts was performed using DPPH radical scavenging activity assay. The experiment was performed in triplicates and data are represented as mean ± SEM (***p < 0.001, while ns denotes non-significant difference (p > 0.05))

Association between total phenolic content and antioxidant capacity among FC and OE extracts

A strong positive linear correlation was shown between TPC for both FC and OE extracts and their antioxidant activities highlighting that antioxidant activity is strongly dependent on the extracts’ TPC as shown in Table 1.

Table 1.

The correlation between TPC and antioxidant activity

Pair	95%CI	Person’s r	p-value
TPC, FRAP	0.743 to 0.955	0.89	< 0.0001¹
TPC, DPPH	0.893 to 0.982	0.956	< 0.0001¹

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¹Associations were analyzed using Pearson’s correlation coefficient (r), and significance was determined at 95% CI (p < 0.05)

Examining the cytotoxic effect of FC and OE extracts on non-tumorigenic intestinal FHs-74 cells

To determine whether the extracts exert any toxic effect on noncancerous cells, we treated FHs-74 cells with FC extracts (0–30 mg/mL) or OE extracts (0–2.4 mg/mL) for 24 h after which cells viability was assessed by MTT assay. Figure 7B showed that treatment with Bq extract resulted in a significant 10% increase in cell viability at concentrations of 2 mg/mL and 5 mg/mL (p < 0.05) compared to the untreated control group. However, Sh and As extracts did not affect the viability of these non-cancerous cells (Fig. 7 A,C).

Similarly, the BO and GO olive extracts demonstrated a protective effect at 0.6 mg/mL and lower concentrations (Fig. 7 D, E). However, higher concentrations of 2.4 mg/mL for BO and 1.6 mg/mL for GO led to over 50% cell death, indicating these doses exceed the toxic threshold.

Notably, at intermediate concentrations of 1.2 mg/mL and 1.6 mg/mL, BO treatment resulted in a significantly greater cell viability by 23% in FHs-74 cells compared to GO (Fig. 7 D, E).

Examining the cytotoxic effect of FC and OE extracts on MCF-7 and MDA-MB-231 breast cancer cells

Results demonstrated a dose-dependent decrease in cell viability upon the single treatment of 0–30 mg/mL FC extracts for 24 h (Fig. 8). No significant difference in MCF-7 cell viability was observed between Sh and Bq extracts. However, the Sh extract caused a significantly greater reduction in cell viability 30 ± 0.23% (p < 0.0001) compared to As extract, which showed a 22 ± 1.76% reduction at the same concentration of 30 mg/mL (Fig. 8). In contrast, when comparing single-dose treatments of OE variety extracts, no significant difference in MCF-7 cell viability was observed between the BO and GO extracts (Fig. 9). Both extracts reduced cell viability to approximately 41.4% at 2.4 mg/mL, suggesting similar cytotoxic effects.

Fig. 9 — OE variety extracts decreased the viability of MCF-7 and MDA-MB-231 cell lines. Cells were treated with increasing concentrations (0- 2.4 mg/mL) of extracts form BO and GO for 24 h after which cell viability was assessed by MTT assay. Data are represented as mean ± SEM of three independent experiments (n = 3; * p < 0.05, *** p < 0.001 compared to untreated control)

On the contrary, the effect of FC variety extracts on MDA-MB-231 cells was almost negligible, where the lowest cell viability achieved was 83% for Sh extract at 30 mg/mL (Fig. 8). The BO extract demonstrated a more pronounced cytotoxic effect on MCF-7 cells compared to MDA-MB-231 cells at concentrations up to 1.2 mg/mL. In contrast, the GO extract showed a comparable effect on both cell lines at 1.2 mg/mL and higher concentrations (Fig. 9).

Figure 10A shows that a 28% reduction in MCF-7 cell viability was achieved with BO at 0.3 mg/mL, a 100-fold lower concentration than that required for Sh (30 mg/mL). Similarly, as shown in Fig. 10B, a 15% reduction in MDA-MB-231 cell viability was obtained with BO at 1 mg/mL, a 30-fold lower concentration than that required for Sh (30 mg/mL).

Fig. 10 — The anticancer potential of increasing concentrations of FC and OE extracts on (A) MCF-7 and (B) MDA-MB-231 breast cancer cell lines. Cancer cells were treated with FC extracts (0–30 mg/mL) or OE extracts (0–2.4 mg/mL) for 24 h after which cells viability was assessed by MTT assay. Data are represented as mean ± SEM of three independent experiments (n = 3)

Synergistic anticancer potential of extracts (BO-Sh) in dual treatment on MCF-7 and MDA-MB-231 breast cancer cells in Vitro

We investigated whether dual treatment would have a more pronounced anticancer effect on MCF-7 cells than single treatments. Different concentrations for combination assay were selected based on MTT results following: (1) the variety of each family that was least cytotoxic on FHs-74 cells, and (2) the concentration that resulted in a range of 30–40% inhibition or 60–70% cell viability against MCF-7 cells.

As a result, Sh and BO were selected at concentrations of 10, 20, and 30 mg/mL (10 folds increment) for Sh and 0.6, 1.2, 2.4 mg/mL (2 folds increment) for BO. A difference in MCF-7 cell viability was observed at 0.6 mg/mL of BO extract in combination treatments, a more significant decrease was evident at 1.2 mg/mL (37 ± 0.408% with 30 mg/mL Sh) and 2.4 mg/mL (21 ± 0.732% with 30 mg/mL Sh) (p < 0.0001) compared to single treatment with BO (Fig. 11 A). Similar results were observed in MDA-MB-231 cells (Fig. 11 B).

Fig. 11 — The combination of Sh and BO extracts enhanced the decrease in cell viability of (A) MCF-7 and (B) MDA-MB-231 cells. Cells were treated with different concentrations of Sh (10, 20, 30 mg/ml) in combination with BO (0.6, 1.2, 2.4 mg/ml) for 24 h after which MTT assay was performed. Data are represented as mean ± SEM of three independent experiments (n = 3)

Combination index of different concentrations of Sh and BO dual extract treatment on breast cancer cells

Combination Index (CI) is a tool generated by CompuSyn that integrates equations based on calculations Chou & Talalay, (1984). This tool identifies whether dual treatment or drug combinations works in a synergism (if CI < 1) or antagonism (CI > 1) or simply have an additive effect (CI = 1). For this purpose, data from MTT assays were analyzed confirming the synergistic interaction at fraction affected (Fa) > 0.538 and > 0.366 in MCF-7 and MDA-MB-231 cells, respectively (Table 2). The dual treatment with BO extract at 2.4 mg/mL and Sh extracts at 30 mg/mL showed the highest anti-tumor activity in MCF-7 cells (CI = 0.315), 79% inhibition of cancer cell viability. MDA-MB-231 cells exhibited a comparable and notable 64% reduction in viability when treated with a combination of 2.4 mg/mL of BO and 10 mg/mL of Sh, as shown in Table 2. This indicates that Sh extract potentiated the cytotoxic effect of BO extract against breast cancer by promoting dose-dependent cell death.

Table 2.

The synergistic effect of Sh and BO extracts on decreasing Breast cancer cell viability

		MCF-7		MDA-MB-231
Dose Sh	Dose BO	Effect	CI¹	Effect	CI
30	2.4	0.791	0.31598	0.60439	0.29576
20	2.4	0.748	0.37934	0.60439	0.19727
10	2.4	0.723	0.39207	0.64234	0.07915
30	1.2	0.631	0.52582	0.55125	0.39839
20	1.2	0.588	0.54769	0.38499	0.66790
10	1.2	0.538	0.55593	0.36657	0.37205
30	0.6	0.275	2.68256	0.20359	3.40826
20	0.6	0.264	2.16902	0.07726	10.4583
10	0.6	0.208	2.13129	0.05268	9.15338

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¹Combination index of different assays at different concentrations of Black olives (BO) (0.6, 1.2, 2.4 mg/ml) and Shatawi (Sh) (10, 20, 30 mg/ml) extracts. CI > 1; CI = 1; CI < 1 indicates antagonism, additive effect, and synergism; respectively. “Effect” refers to the effected fraction (Inhibitory fraction) of cells at each combination assay. Boldface values denote statistically significant synergistic effects (CI < 1) observed at extract concentration > 0.6 mg/ml

Comparing IC50 values of FC and OE extracts in single and dual treatment on breast cancer cells

In line with these results, the calculated half-maximal inhibitory concentration (IC₅₀) values were approximately 2.25 mg/mL for BO extract and 2.3 mg/mL for GO extract (Table 3A), indicating comparable potency. IC₅₀ values of FC variety extracts were estimated based on simple linear regression equations.

Table 3.

The half- maximal inhibitory concentration (IC50) of different FC and OE extracts on Breast cancer cells at 24 h

A	Extract	MCF-7		MDA-MB-231
Single extract treatment		IC50 ¹	95% CI	IC50	95% CI
	Sh	54.99	43.87- 76.25	94.23	81.43–112.6
	Bq	54.05	43.68- 73.02	96.78	74.83–140.9
	As	64.48	52.08—86.84	75.36	64.14–92.34
	BO	2.257	1.917—2.814	2.321	2.159—2.517
	GO	2.302	2.107—2.552	2.210	2.004—2.479

B	Pairs	MCF-7		MDA-MB-231
Combination		IC50 ¹	95% CI	IC50	95% CI
	With 10 Sh	1.486	1.308- 1.698	1.874	1.659–2.155
	With 20 Sh	1.373	1.170- 1.611	1.927	1.699–2.231
	With 30 Sh	1.280	1.074–1.513	1.561	1.267–1.986

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¹The concentration at which FC and OE variety extracts exerts 50% cell viability (IC50) on MCF-7 and MDA-MB-231 cells in single treatment (A) and IC50 of BO extract in dual treatment (B). Sh Shatawi, Bq Baqarati, As Asali, BO black olive, GO green olive

Table 3 B demonstrated that the concentration of BO extract required to induce 50% cell death in combination with Sh extract (30 mg/ml) against MCF-7 cells was reduced to 1.28 mg/mL, compared to 2.25 mg/mL when used alone. A similar trend was observed in MDA-MB-231 cells, where the effective concentration decreased from 2.321 mg/mL in single treatment to 1.561 mg/mL in the combined treatment. The combination of both BO and Sh extracts shows a higher SI (1.875, 1.537) compared to the treatment with BO alone (1.063, 1.034) in MCF-7 and MDA-MB-231 BC cell lines, respectively (Table 1, supplementary files).

Examining the effect of FC and OE extracts on the cell cycle progression of MCF-7 cells

We treated MCF-7 cells with 1.2 mg/mL of BO extract alone or in combination with 30 mg/mL of Sh extracts for 24 h. We subsequently performed cell cycle analysis using flow cytometry with PI staining on DNA to denote whether cell death occurred due to apoptosis or cell cycle arrest (Fig. 12). Accumulation of cells was significant at the sub-G1 phase for both single and dual treatment. However, the dual treatment induced a higher accumulation of cells in sub-G1 phase compared to single treatment. In addition, the increase in the sub-G1 population was associated with a more pronounced reduction in the G0/G1 and G2/M populations with the dual treatment of BO and Sh extracts compared to single treatment with BO extracts.

Fig. 12 — Single and dual treatment of Sh and BO extracts enhanced cell death through an increase in sub-G1 population in MCF-7 cells after 24 h of treatment. Cell cycle analysis performed using flow cytometry with PI staining of DNA on: (1) Control (untreated MCF-7 cells), (2) MCF-7 cells treated with BO extract at 1.2 mg/mL, and (3) MCF-7 cells treated with the combination of (BO-Sh) extracts at 1.2 and 30 mg/mL, respectively. Results are demonstrated as % of cells in each of the cell cycle phases (Sub-G1, G0/G1, S-phase, G2/M). Data are represented as mean ± SD of two replicates and analyzed by a two-way analysis of variance test followed by multiple comparisons test

Discussion

The Mediterranean diet had been previously shown to exert anticancer effects due to the action of several Mediterranean polyphenolic compounds namely gallic acid, tannins, oleocanthal, olive oil phenols, anthocyanins, and quercetin; all of which are as well olive- and fig-derived polyphenols [34]. This study provides the first evidence that the combined Lebanese FC and OE fruit methanolic extracts exert synergistic cytotoxic effects of on BC cells and elucidate their mechanisms of action suggesting potential for dietary -based adjunctive strategies in BC. The central findings of this study are: (1) darker-colored varieties exhibited higher TPC and stronger antioxidant activity compared to their lighter-colored counterparts; (2) both FC and OE extracts exhibited notable anticancer activity; (3) however, their effects were enhanced when applied in combination, while showing no antiproliferative impact on non-cancerous FHs-74 cells.

In this study, we aimed to quantify a broad phenolic profile (polyphenols, and anthocyanins) using a methanolic extraction method. Previous studies have opted for a methanolic extraction for both figs and olives [35, 36] and were successful in retrieving higher TPC, total flavonoid content, DPPH, and FRAP values in figs compared to other extraction solutions. Methanol was also widely used in the extraction of phenolic compounds, flavonoids, anthocyanins, phenolic acids, terpenoids, lignans, polysaccharides, and carotene; which are abundant in fruits and plants [37].

Our findings are consistent with previous studies indicating that figs with darker peels contain greater amounts of polyphenols, particularly flavonoids [38–41]. However, the TPC of FC extracts analyzed in this study were lower than the values reported in the literature [42]. This deviation may be attributed to several factors including geographic origin, harvest timing, processing methods, and extraction techniques which can markedly influence the polyphenolic profile [43]. In our case, the figs were collected from mountainous areas in Lebanon, where altitude, cooler temperatures, and distinct soil composition may influence phenolic biosynthesis compared with figs grown in lowland or coastal regions.

On the other hand, higher TPC was shown among OE varieties compared to FC varieties. This is consistent with the known phenolic profile of olive fruits, whose flesh contains a relatively high proportion of hydrophilic phenols mainly hydroxytyrosol, tyrosol, oleuropein, verbascosides, and flavonoids including luteolin-7-rutinoside, luteolin-7-glucoside, apigenin-7-glucoside and rutin [44–46].This could be related to its non-anthocyanin compounds such as flavonoid content that is likely to contribute significantly to the high TPC in OE. This is in line with previous studies which identified luteolin-7-rutinoside—alongside verbascoside—as major polyphenols in olive fruit, and that their concentrations tend to increase as the fruit matures, while the secoiridoid oleuropein is predominant at early stages of maturation, and declines with ripening [12]. Thus, we hypothesize that these non-anthocyanin phenolic compounds are largely responsible for the elevated TPC in GO as it was harvested at earlier ripening stage than BO and expected to retain higher levels of such phenolics.

Since anthocyanins represent a major fraction of the polyphenolic profile, TAC was quantified to further characterize the qualitative aspect of TPC across the varieties. This study demonstrated a higher TAC only in the BO variety but not in GO extract. This can be explained by the fact that anthocyanins are a major subgroup of flavonoids in plants, and are responsible for the pigmented coloration of fruits and vegetables [47]. Similarly, the dark colored FC varieties (Sh and Bq) expressed significantly higher TAC compared to As and GO varieties. Comparable results were reported by Vallejo et al., (2012) where neither Cyanidin-3-glucoside nor Cyanidin-3-rutinoside anthocyanins were present in green fig cultivars [20].

To examine the relationship between antioxidants and phytochemical properties, this study further demonstrated a dose-dependent relationship between antioxidant activity and the color intensity across different varieties of FC and OE. This association aligns with previous findings by Bayrak et al., (2023) [39], which showed highest antioxidant activity among the dark purple and purple FC variety extracts. Moreover, a previous study examining nine Algerian FC varieties, reported the highest Vitamin C -a strong antioxidant- in the dark peeled variety “Olk Elhama” as compared to lighter variety. The authors also reported higher TPC in the peel of the figs, encouraging the consumption of the whole fruit [48]. Even though all FC extracts had higher anthocyanin levels than GO, they exhibited weaker antioxidant and anticancer activities, while GO maintained strong antioxidant capacity despite lacking detectable anthocyanins supports the notion that that phenolics other than anthocyanins are likely the primary drivers of the observed effects.

We then evaluated the effect of the FC and OE extracts on the viability of normal FHs-74 cells. Although we have used FHs 74 intestinal cells which don’t share do not share the same tissue origin or microenvironment as breast epithelial cells, they were used as non-tumorigenic models in our project to evaluate off-target cytotoxicity and selectivity of OE and FC extracts. Both cell lines are non-tumorigenic, adherent monolayer cells that are derived from solid organs composed of epithelial layers. They were used as proof-of-concept on this project providing a first-line indication of general epithelial safety when exposed to the extracts. On the other hand, primary human mammary epithelial cells are much more complicated to handle due to their shorter lifespan and high variability. As a stable and well-characterized epithelial line, and considering that these fruits are orally ingested, Fh74Ins was chosen to assess for any toxicity since intestinal epithelium would be of the first healthy tissues to get exposed to these compounds. Interestingly, the FC extracts displayed a safe profile. This finding is consistent with the work of Abdel-Rahman et al. (2021) [25], who reported enhanced viability of peripheral blood mononuclear cells at low concentrations (156 µg/mL) of fig leaf and fruit extracts, which are rich in antioxidant polyphenols. Similarly, Zubair et al. (2015) [49] found no cytotoxic effects of fig extracts on mouse epithelial (3T3) cells. As for the OE extracts, they did not affect the viability of normal cells at concentrations up to 0.9 mg/mL, but reduced cell viability at higher concentrations. In the same line, previous research showed that Oleocanthal, one of the phenolic compounds of extra virgin olive oil, did not significantly affect the viability of the human dermal fibroblasts and human mammary epithelial cells [50, 51]. Notably, the favorable safety profile of BO may be linked to its high content in anthocyanins. The latter have been shown to protect human somatic cells and possess anti-mutagenic effects [52].

We next examined the effect of the FC and OE extracts on the viability of breast cancer cells. Extracts with higher TPC and antioxidant activity demonstrated a stronger antiproliferative effect on breast cancer cells. Besides, our results are consistent with Jarwan et al. (2023)[53], who reported an effect of olive extracts, and Soltana et al. (2019), who observed a similar effect of whole-fruit FC extracts in reducing the viability of colorectal cancer cells [54]. Moreover, when comparing the cytotoxic effect of both OE and FC extracts between the two breast cancer cell lines, the anticancer activity was significantly higher in MCF-7 cells than in the triple negative breast cancer cells (MDA-MB-231) indicating higher sensitivity of the MCF-7 cells. Notably, research on triple negative breast cancer is of a major concern, considering that it is an aggressive subtype of breast cancer, that lacks estrogen, progesterone, and HER2 receptors, limiting the options of treatment (such as hormonal therapy) [55]. Although FC extracts showed only a minimal inhibitory effect on the viability of MDA-MB-231 cells, OE extracts exerted a more pronounced effect. It is important to note that several phytochemicals have been shown to target specific signaling pathways of triple negative breast cancer [56]. For instance, Hydroxytyrosol- a derivative of oleuropein- was shown to counteract the propagation of this cancer through the chelation of the copper metal in the cancerous cells [57].

Moreover, this research indicated that the decrease in MDA-MB-231 cell viability was evident only after 0.9 mg/mL of treatment with OE extracts. This might be explained due to the concentration dependent activity of flavonoids to exert pro-oxidative activity as evident by Yen et al., (2003) [58]. León-González et al. (2015) [59] further discusses several mechanisms by which antioxidants exert anticancer effects, including acquiring pro-oxidant potential, activating NADPH oxidases, inducing mitochondrial dysfunction and apoptosis, and participating in transition metal-mediated cytotoxicity.

Even though our results showed that the cytotoxicity effect with BO extract was significant, dual treatment with Sh extract had a greater synergistic cytotoxic activity showing a dose dependent decrease in the proliferation of both BC cells. A previous study [60] conducted in Indonesia investigated the anticancer effect of fig and olive oil extracts on HeLa cervical cancer cells also reported the highest cytotoxic activity for the combination assay at ratio 3:1 (FC: OE oil) with an IC₅₀ of 563.62 μg/mL. Similar results were shown by researchers on T-47D and MCF-7 breast cancer cells at 1:1 ratio [61]. Moreover, the combined administration of olive oil, figs, and date palm fruit exerted a synergistic protective effect in rats during diethylnitrosamine-induced carcinogenesis [62].

Previous studies have shown that fig fruit extract inhibits cancer cell viability, and induce apoptosis through ROS generation, and autophagy [63]. In line with this finding, our results show an enhanced increase in G0-G1 cell population (a sign of apoptosis and/or necrosis) when combining BO and Sh extracts. This is not surprising, considering that the combination of key phytochemicals has been previously shown to exert a superior inhibition of cancer cell viability compared to single component treatment, through inhibiting Notch-1 gene expression and inactivates the NF-κB DNA-binding activity against pancreatic cell lines [64]. Combinatorial effects of key components in fig and olive fruit extracts (anthocyanins, quercetin, oleuropein, hydroxytyrosol,…) have not been tested yet on cancer cell death. However, one study showed an anti-migratory effect of fig and olive extracts on pancreatic cancer cell lines [65].

The findings of this study highlight the potential of integrating adjunctive nutritional strategies into breast cancer management. Importantly, the concentrations of FC and OE fruit extracts tested in vitro can be reasonably translated into fresh weight equivalents, suggesting that the observed biological effects may be achievable through dietary intake. A particularly notable result was the synergistic interaction between BO and Sh extracts. The combined treatment of 1.2 mg of BO extract with 30 mg of Sh extract produced a significantly greater cytotoxic effect on MCF-7 breast cancer cells compared to either extract administered alone, indicating a potentiation of their anticancer activity when used together. When converted to dietary equivalents, these amounts correspond to approximately 0.006 g of BO pulp and 0.07 g of Sh fruit in fresh weight, quantities that are both minimal and realistically attainable in the context of regular consumption (Appendix A; Supplementary material). While some polyphenols from olive extracts have been shown to be absorbed in human studies such as Oleuropein, hydorxytyrosol, their systemic concentrations remain relatively low. In humans, the approximate blood concentration of olive polyphenols (e.g. oleuropein) is 0.7–1.2 ug/ml which is equivalent to 0–4 Um from the oral intake 50 mg aglycone [66]. Another clinical study reported that 250 mg oleuropein-rich olive leaf extract were also detected at a low micromolar range with peak plasma concentrations observed with 1–3 h after oral administration [67].

Although no clinical trials have been done to assess their anticancer efficacy and very few studies have been done to evaluate the fig and olive pharmacokinetic properties, future studies should evaluate in vivo pharmacokinetics of these extracts and their bioavailability to determine levels that may be achievable clinically. The doses used in our in vitro studies are solely as a proof of concept for cytotoxicity screening. These findings also underscore the promise of developing combinatory nutritional approaches that may complement conventional therapeutic strategies against breast cancer.

This present work advocates for the antioxidant and anticancer potential of bioactive compounds in the MD. While this study compared the overall antioxidant and anticancer potential of fig and olive fruit extracts only and related these biological effects to their global phenolic content within the context of the Mediterranean diet, many other nutraceutical components of the MD remain underexplored. This extract-based, and synergistic effects of nutraceuticals on human health closely reflects the way humans actually consume these fruits in their diet rather than isolating or supplementing single nutraceutical compounds.

Several challenges must be addressed before these findings can progress toward clinical application. One critical factor is the bioavailability and kinetics of absorption of active compounds in the human body [68]. This area is currently receiving growing attention, particularly through innovations in enhancing the delivery of nutraceuticals using advanced systems such as metal–organic frameworks (MOFs) [69, 70]. Moreover, the experimental protocol was done on breast cancer cells, however, given that cell lines lack the complexity of a living organism. Additional mechanistic studies are therefore required to determine whether the extract-induced reduction in breast cancer cell viability occurs through apoptotic pathways, necrotic processes, or a combination of both. Further in vivo validation of the extracts and their combination in animal models followed by formulation studies and early-phase clinical trials to assess their pharmacokinetics, safety profile and synergistic therapeutic potential.

Conclusion

In summary, while FC exerted no cytotoxicity on FHs-74 cells and only a statistically irrelevant effect on triple negative breast cancer MDA-MB-231, they had a notable significant effect on MCF-7 cells. Likewise, OE extracts which showed a similar protective effect on FHs-74 up to a certain concentration, exhibited a more prominent anticancer effect on MCF-7 than MDA-MB-231 at lower concentrations. This pattern is in line with their higher TPC and stronger antioxidant capacity, as BO and GO had a significantly more pronounced anticancer effect than FC varieties. In contrast, although all FC extracts contained higher anthocyanin levels than GO, they showed lower antioxidant and anticancer activities and GO retained high antioxidant capacity despite having undetectable anthocyanins, suggesting that non-anthocyanin phenolics are the main contributors to the observed effects. Moreover, the dual treatment of Sh and BO extracts resulted in a more prominent anticancer effect and higher sub-G1 cell death compared to single treatment. Further research on these extracts is required to elucidate the mechanisms of cell death as adjunctive anticancer modulators and to evaluate the potential synergistic effects of fig and olive extract with conventional chemotherapy drugs on tumor behavior, inflammatory and oxidative stress marker, while also assessing DNA damage, and expanding our studies to Xenograft models using SCID-NOD mice.

Supplementary Information

Supplementary Material 1.^{(192.7KB, pdf)}

Supplementary Material 2.^{(2.3MB, docx)}

Acknowledgements

We gratefully acknowledge the funding provided by the American University of Beirut Research Board (URB), the K. Shair Central Research Science Laboratory (CRSL) at AUB for providing the facilities to carry out this work.

Abbreviations

FC: Ficus carica
OE: Olea europea
Sh: Shatawi
Bq: Baqarati
As: Asali
BO: Black olives
GO: Green olives
BC: Breast cancer
ASIR: Age Standardized Incidence Rate
TPC: Total polyphenolic content
F-C: Folin–Ciocalteu
TAC: Total anthocyanin content
RT: Room temperature
C6GE: Cyanidin-6-O-glucoside equivalent
FRAP: Ferric Ion Reducing antioxidant potential
DPPH: 2,2-Diphenyl-1-picrylhydrazyl
MTT: 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide
MCF-7: Michigan Cancer Foundation-7
MDA-MB-231: M D Anderson—Metastatic Breast – 231
MDR: Multidrug resistance
DMEM: Dulbecco’s modified Eagle medium
DMSO: Dimethyl sulfoxide
MEM: Minimal essential medium
FBS: Fetal bovine serum
EGF: Epidermal growth factor
EDTA: Ethylenediaminetetraacetic acid
CI: Combination index
PI: Propidium iodide
GAE: Gallic acid equivalent
IC₅₀: 50% Inhibitory concentration
WHO: World Health Organization
SEM: Standard error of mean

Authors’ contributions

SAa.S, H.G.M, N.H, and W.E.K handled Conceptualization and Methodology; S.A.S carried out lab work as part of her MSc thesis, performed analysis and interpretation of data (e.g., biostatistics, statistical analysis and editing), and wrote the first draft of the manuscript; S.A.S, WEK, DI did Data curation, LN, NH, HGM, WEK, and DI supervised the work; SAS, DI, HGM, and NH handled Interpretation, Manuscript preparation; All authors have read and approved the final manuscript.

Funding

This research was funded by the University Research Board (URB), (Award: 104514, Project 27589).

Data availability

The datasets used and analyzed during the current study are available from the corresponding author upon request.

Declarations

Ethics approval consent to participate

Permission was obtained from the landowners (farmers), who handpicked the fruit samples and delivered them to our facility.

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare no competing interests.

Footnotes

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Hala Gali-Muhtasib and Nahla Hwalla contributed equally to this work.

Contributor Information

Hala Gali-Muhtasib, Email: amro@aub.edu.lb.

Nahla Hwalla, Email: nahla@aub.edu.lb.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

Supplementary Material 1.^{(192.7KB, pdf)}

Supplementary Material 2.^{(2.3MB, docx)}

Data Availability Statement

The datasets used and analyzed during the current study are available from the corresponding author upon request.

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[CR63] 63.Ou A, Zhao X, Lu Z. Autophagy is involved in Ficus carica fruit extract-induced anti-tumor effects on pancreatic cancer. Biomed Pharmacother. 2022;150:112966. [DOI] [PubMed] [Google Scholar]

[CR64] 64.Wang Z, Desmoulin S, Banerjee S, Kong D, Li Y, Deraniyagala RL, et al. Synergistic effects of multiple natural products in pancreatic cancer cells. Life Sci. 2008;83(7–8):293–300. [DOI] [PMC free article] [PubMed] [Google Scholar]

[CR65] 65.Dekdouk N, Ameddah, Souad, Bensouici, Chawki, Ibtissem, Meghboun , Ahmed, Menad, Martel, Fatima. Antioxidant properties and anticancer activity of Olea europaea L. olive and Ficus carica fruit extracts against pancreatic cancer cell lines. Trends in Phytochemical Research. 2024;8(2):80–92.

[CR66] 66.Bucciantini M, Leri M, Nardiello P, Casamenti F, Stefani M. Olive Polyphenols: Antioxidant and Anti-Inflammatory Properties. Antioxidants (Basel). 2021;10(7):1044. [DOI] [PMC free article] [PubMed]

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PERMALINK

Unveiling the antioxidant and anticancer potential of local mediterranean olive and fig extracts against breast cancer

Sara Al Srouji

Walid El Kayal

Diana Ismail

Lara Nasreddine

Hala Gali-Muhtasib

Nahla Hwalla

Abstract

Supplementary Information

Introduction

Materials and methods

Fruit material and extraction preparation

Fruit material

Extraction process

Quantitative phytochemical analysis in extracts

Total Polyphenol Content (TPC) Quantification

Total anthocyanins content (TAC) quantification

Evaluation of antioxidant activity in extracts

Ferric Ion Reducing antioxidant potential (FRAP)

2-diphenyl-1-picrylhydrazyl (DPPH)

Cell culture

Chemicals and reagents

Cell culture conditions

Cell viability assay (MTT)

Combination Index analysis

Cell cycle analysis

Statistical analysis

Results

Total polyphenolic content of OE and FC varieties

Fig. 1.

Total anthocyanin content of OE and FC varieties

Fig. 2.

Antioxidant activity

Ferric reducing antioxidant power

Fig. 3.

Fig. 4.

Fig. 5.

DPPH Radical scavenging radical activity

Fig. 6.

Association between total phenolic content and antioxidant capacity among FC and OE extracts

Table 1.

Examining the cytotoxic effect of FC and OE extracts on non-tumorigenic intestinal FHs-74 cells

Fig. 7.

Examining the cytotoxic effect of FC and OE extracts on MCF-7 and MDA-MB-231 breast cancer cells

Fig. 8.

Fig. 9.

Fig. 10.

Synergistic anticancer potential of extracts (BO-Sh) in dual treatment on MCF-7 and MDA-MB-231 breast cancer cells in Vitro

Fig. 11.

Combination index of different concentrations of Sh and BO dual extract treatment on breast cancer cells

Table 2.

Comparing IC50 values of FC and OE extracts in single and dual treatment on breast cancer cells

Table 3.

Examining the effect of FC and OE extracts on the cell cycle progression of MCF-7 cells

Fig. 12.

Discussion

Conclusion

Supplementary Information

Acknowledgements

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