1 Introduction
Medicinal plants play a significant role in traditional herbal medicine practices worldwide, particularly among impoverished individuals [1,2]. The therapeutic properties of these plants can be attributed to the presence of secondary bioactive compounds. One such plant is the pomegranate (Punica granatum L.), which is consumed widely in various forms such as fresh fruit, juice, jams, and wines [3]. The different parts of the pomegranate fruit, including flowers, leaves, arils, and peel, as well as its products like fresh or sweet juices, possess notable biological activities due to their high content of polyphenols [3], alkaloids, tannins, flavonoids, anthocyanins, terpenoids sterols, ellagic acid, and unsaturated fats [4].
Pomegranate peels (PPs) are characterized by an interior network of membranes comprising almost 26–30% of the total fruit weight and are characterized by substantial amounts of phenolic compounds, including flavonoids (anthocyanins, catechins, and other complex flavonoids) and hydrolyzable tannins (punicalin, pedunculagin, punicalagin, gallic, and ellagic acid) [5]. The extract obtained from PP has demonstrated potent antioxidant properties. The methanol–water extraction process effectively preserves the antibacterial properties of PP against a range of pathogens and organisms, such as Bacillus subtilis, Listeria monocytogenes, Yersinia enterocolitica, Staphylococcus aureus, Candida albicans, Saccharomyces cerevisiae, and Aspergillus [5,6]. Moreover, pomegranate peel extract (PPE) has shown strong antimicrobial activity, as evidenced by its inhibitory effect on the growth of two common human bacteria, S. aureus and Escherichia coli, often associated with foodborne illnesses [7]. The antimicrobial effects of PPEs on cariogenic bacteria have also been highlighted in previous studies [8].
Furthermore, both ethanol and water extracts of pomegranate contain phytochemicals such as flavonoids, phenols, and tannins, which are important constituents responsible for these activities. The advantages of using ethanol as a solvent for extracting substances compared to water include the broad solubility of compounds, including both polar and non-polar substances. This makes it suitable for extracting a diverse array of molecules from various sources and also for the efficient extraction of bioactive compounds. It can extract a higher quantity of target compounds compared to water, resulting in higher yields. Furthermore, ethanol can preserve the stability and integrity of phytochemicals during extraction and the compatibility with analysis techniques However, research on the volatile components of pomegranate and its antibacterial effects is scarce [9]. Therefore, our study aims to investigate the phenolic profile of pomegranate PPE in ethanol and evaluate their potential as an anti-rancidity agent in different oils, including corn, olive, sunflower, and soybean. Additionally, the agar-well diffusion method will be employed to assess the antimicrobial activity of PPE against four microorganisms relevant to food safety, including two Gram-positive bacteria (B. subtilis and S. aureus), two Gram-negative bacteria (E. coli and Pseudomonas aeruginosa), yeast strain (C. albicans), and two fungal strains (Aspergillus flavus and Aspergillus niger). The phenolic content, antioxidant scavenging activity, and total phenolic content of PPE will be quantified to determine its efficacy as an anti-rancidity agent in different oils.
2 Materials and methods
2.1 Preparation of PP
Mature pomegranate fruits were procured from local markets in Saudi Arabia. The peels were manually separated and allowed to cool before being cleaned and dried outdoors for 5 days. Subsequently, the dried peels were fragmented into smaller pieces and processed using an electric mixer until they transformed into a fine powder. The powder was then sieved through a 0.2 mm sieve and stored in a paper bag in a refrigerator at 8°C until further use [10].
2.2 Preparation of pomegranate peel ethanolic extract
The ethanolic extraction procedure of PPE was performed according to the approach outlined by Cai et al. [11]. A known weight (20 g) of the dry powdered PP was mixed with 200 mL of 85% ethanol in a 1,000 mL conical flask. The mixture was extracted for 24 h using a Soxhlet extractor. This extraction procedures were repeated three times, and the resulting extracts were pooled together to ensure maximum yield. To ensure the purity and clarity of the extract, the obtained mixture was subjected to triple filtration using Whatman No. 1 filter paper. Subsequently, the filtrate underwent concentration under reduced pressure at a temperature of 40°C utilizing a rotary evaporator. The obtained powder was then redissolved in distilled water to prepare a solution with a final concentration of 200 ppm of PPE. The prepared extract was the stored in a dark environment at −20°C for future use.
2.3 Phytochemical analyses
2.3.1 Radical scavenging activity (2,2-diphenyl-1-picrylhydrazyl [DPPH]) of PPE
The DPPH˙ assay was employed to evaluate the free-radical scavenging activity as an indicator of the antioxidant potential of PPE (200 ppm), following the methodology proposed by El-Beltagi et al. [12]. The DPPH free radical scavenging activity was calculated and expressed as a percentage (%).
2.3.2 Determination of total phenolic content
The Folin–Ciocalteu method, as described by Danial and Basudan [13], was utilized to determine the total phenolic content of PPE. The total phenolic content of the extracts was quantified using gallic acid as the standard phenol, and the results were expressed in milligrams of gallic acid equivalent per 100 g of dry weight of peel.
2.4 Antimicrobial activity of PPE
2.4.1 Microorganisms and media
The Department of Microbiology at King Abdul-Aziz University in Saudi Arabia provided seven strains of microorganisms relevant to food safety. The strains included C. albicans (yeast), A. niger and A. flavus (mycelial fungi), B. subtilis and S. aureus (Gram-positive bacteria), and E. coli and Pseudomonas aeruginosa (Gram-negative bacteria). The bacterial and fungal strains were kept in nutrient agar and potato dextrose agar (PDA) slants, respectively, at 4°C. Before experimentation, the strains were sub-cultured in a nutrient broth of the corresponding medium and incubated at 37°C for 24 h to activate the microorganisms, which were then used as inoculum for evaluating the antimicrobial activity of PPE.
2.4.2 Antimicrobial activity
The agar well diffusion method, as suggested by Bassiri-Jahromi et al. [14], was employed to assess the antimicrobial activity of PPE (200 ppm) against the provided microbial strains. Microbial suspensions with a concentration of 1.0 × 107 CFU/mL were prepared from the stock cultures. A volume of 0.1 mL of the prepared microbial cell suspensions was aseptically spread onto nutrient agar plates for bacterial strains and PDA plates for fungal strains. After allowing the plates to dry at room temperature for 30 min, wells with a diameter of 8 mm were created in the agar using a sterile stainless-steel borer. Subsequently, 30 µL of PPE was added to each well. The plates were then incubated at 37°C for 24 h for bacterial strains and 30°C for 48 h for fungal strains. The diameter of the resulting inhibition zones was measured to determine the antimicrobial activity of PPE.
2.5 Identification of phenolic compounds by liquid chromatography–electrospray ionization–tandem mass spectrometry (LC–ESI–MS/MS)
Phenolic compounds in PPE were identified using LC–ESI–MS/MS analysis (Shimadzu, Japan). The chromatographic separation was performed on an Xtimate C-18 column (4.6 × 250 mm i.d., 5 μm) with a mobile phase consisting of 2% glacial acetic acid in water (solvent A) and acetonitrile (solvent B). The gradient program started with 5% solvent B and increased linearly to 50% over 30 min, followed by a post-run equilibration period. The flow rate was set at 0.8 mL/min, and the column temperature was maintained at 30°C.
The LC eluent was introduced into the ESI source of the mass spectrometer, operating in both positive and negative ionization modes. The mass spectrometer was operated with the following parameters: capillary voltage, 3.5 kV; nebulizing gas flow, 1.5 L/min; drying gas flow, 15 L/min; and drying gas temperature, 350°C. Mass spectra were acquired in the range of m/z 100–1,000. The phenolic compounds in PPE were identified by comparing their retention times and mass spectra with those of authentic standards and available databases. The identification was further confirmed by analyzing the fragmentation patterns of the compounds.
2.6 Rancidity assessment of utilized edible oils
2.6.1 Selection of oils for testing
The present study included four types of plant-based oils, namely olive, sunflower, soybean, and corn oil. The composition of polyunsaturated fatty acids varies among these oils, with olive oil being rich in oleic acid, sunflower oil containing a high proportion of linoleic acid, soybean oil abundant in α-linolenic acid, and corn oil primarily composed of polyunsaturated fatty acids, particularly linoleic acid, and low levels of saturated fatty acids [15].
2.6.2 Evaluation of oxidative stability using the Rancimat test
The oxidative stability of the oils was determined using a Rancimat apparatus (model 679, Switzerland) according to the guidelines outlined in ISO 6886:1997. A 5 g sample of each oil was subjected to a continuous airflow of 20 L/h at a temperature of 110°C. The induction times, measured with an accuracy of 0.005 s, were automatically recorded by the software of the apparatus [12].
2.6.3 Oven test of PPE compared to butylated hydroxytoluene (BHT) in oils
To evaluate the efficacy of PPE as an antioxidant, the used corn, olive, sunflower, and soybean oils were individually treated with PPE at a fixed concentration of 200 ppm. Negative controls (without any antioxidant) and positive controls (with the synthetic antioxidant BHT at a concentration of 200 ppm) were also included, following the recommendations of Cruz-Valenzuela et al. [16]. Each test was performed three times, and the oils were exposed to ambient air, light, surface, and the testing oil itself in open 500 mL containers placed in an oven at 70°C for 10 days [15].
Before analysis, both PPE and BHT were dissolved in propylene glycol. At intervals of 0, 2, 4, 6, 8, and 10 days, a random 30 mL aliquot was extracted from each treatment to measure the peroxide value (PV) and acid value (AV) of the oil samples. The testing treatments applied to the used oils are summarized in Table 1.
Table 1
List of oil samples treated with PPE or BHT at 200 ppm concentration in the present study
| No | Sample | Code |
|---|---|---|
| 1 | Corn oil | C |
| 2 | Corn oil with PPE | C PPE |
| 3 | Corn oil with BHT | C BHT |
| 4 | Olive oil | O |
| 5 | Olive oil with PPE | O PPE |
| 6 | Olive oil with BHT | O BHT |
| 7 | Sunflower oil | F |
| 8 | Sunflower oil with PPE | F PPE |
| 9 | Sunflower oil with BHT | F BHT |
| 10 | Soybean | B |
| 11 | Soybean oil with PPE | B PPE |
| 12 | Soybean oil with BHT | B BHT |
2.6.4 Determination of PV
The PVs of the tested oils, as well as the oils supplemented with PPE and BHT, were calculated using the method proposed by El-Beltagi et al. [12]. The PV was determined using the following formula:
where M represents the molarity of sodium thiosulfate, B is the volume of titrant (mL) for the blank, S is the volume of titrant (mL) for the test portion, and the sample weight refers to the weight of the oil sample.
2.6.5 Calculation of the AV
The AVs of the tested oils and the oil samples fortified with PPE and BHT were determined based on the method described by El-Beltagi et al. [12]. A known weight of the test oils (56.4 ± 0.20 g) was placed in an Erlenmeyer flask, followed by the addition of 2 mL of indicator and 50 mL of hot neutralized alcohol. The AV was calculated using the following formula:
2.7 Statistical analysis
All experiments were performed in triplicate, and the results were expressed as means ± standard deviations (SD). Statistical analysis was conducted using one-way analysis of variance (ANOVA) by means of CoStat software (CoHort, V. 6.311) to compare the data means, followed by Duncan’s multiple range test. Differences were considered significant at P < 0.05.
3 Results and discussion
3.1 Radical scavenging activity of PPE
In the current study, the PPE showed a significant radical scavenging activity (85.26%) as contrasted to the standard antioxidant ascorbic acid (93.88%) (Figure 1). PPE has gained significant attention as a potential natural source of antioxidants due to its high content of phenolic compounds, flavonoids, and tannins [17]. These bioactive constituents have been reported to exhibit strong radical scavenging activity, which plays a crucial role in preventing oxidative damage caused by reactive oxygen species in various biological systems [18]. In this study, the radical scavenging activity of PPE was compared to the standard antioxidant, ascorbic acid. The results demonstrated that PPE exhibited remarkable radical scavenging activity comparable to that of ascorbic acid. This suggests that PPE possesses potent antioxidant properties and can effectively neutralize free radicals.
Figure 1
DPPH radical-scavenging activity of PPE against the standard ascorbic acid. Different letters denote significant differences at the significance level of 0.05.
Several studies have investigated the antioxidant potential of PPE. For instance, Sihag et al. [19] evaluated the radical scavenging activity of PPE using the DPPH assay and reported a strong antioxidant capacity. Similarly, Cervantes-Anaya et al. [20] found that PPE exhibited significant radical scavenging activity against superoxide and hydroxyl radicals. The high radical scavenging activity of PPE can be attributed to its rich composition of phenolic compounds, such as ellagitannins and flavonoids [21]. These compounds possess strong antioxidant properties and can donate hydrogen atoms or electrons to stabilize free radicals, thereby preventing oxidative damage. Thus, the similar radical scavenging activity observed between PPE and ascorbic acid suggests that PPE can serve as a natural alternative to synthetic antioxidants. Overall, the findings of this study highlight the significant radical scavenging activity of PPE, which makes it a promising natural source of antioxidants. Further research is warranted to elucidate the specific bioactive compounds responsible for PPE antioxidant activity and to explore its potential applications in various industries, including food and pharmaceuticals.
3.2 Phenolic content of PPE
The results of the current study revealed that the PPE possessed a relatively high content of phenolic compounds (32.30 mg/g dw) (Figure 2). This extract is known for its high phenolic content, which contributes to its potent antioxidant and therapeutic properties. Phenolic compounds are bioactive compounds found abundantly in plant-based foods and have been extensively studied for their beneficial effects on human health. The high phenolic content in PPE can be attributed to the presence of various phenolic compounds, including ellagitannins, flavonoids, and anthocyanins [17]. These compounds are known for their antioxidant, anti-inflammatory, and antimicrobial activities. Among the phenolic compounds, ellagitannins, such as punicalagins and punicalins, are the major contributors to the phenolic content in PPE [22].
Figure 2
Total phenolic content of PPE as compared to the standard gallic acid. Different letters denote significant differences at the significance level of 0.05.
The high phenolic content in PPE is of great interest due to its potential health benefits. Phenolic compounds are known to scavenge and neutralize free radicals, thereby protecting cells from oxidative damage [23]. Additionally, they have been associated with various health-promoting effects, including anti-inflammatory, anticancer, and cardioprotective properties [24]. Moreover, the phenolic compounds in PPE have been shown to exhibit antimicrobial activity against a wide range of pathogens, including bacteria, fungi, and viruses [25]. This antimicrobial property is of significant interest in the development of natural alternatives to conventional antimicrobial agents.
3.3 Antimicrobial activity of PPE against the tested microorganisms
The data presented in Table 2 provide evidence regarding the antimicrobial properties of PPE against the microorganisms that were tested in comparison to the standard antimicrobial agents (neomycin as an antibiotic and itraconazole as an antifungal). The results indicate that PPE displayed varying degrees of inhibition zone diameter, ranging from 18.2 to 24.5 mm. More specifically, when tested against Gram-positive bacteria, PPE exhibited inhibition zones measuring 22.4 and 20.6 mm against S. aureus and B. subtilis, respectively. However, when tested against Gram-negative bacteria, PPE demonstrated a stronger inhibitory effect against P. aeruginosa (24.5 mm) compared to E. coli (18.2 mm). Moreover, PPE exhibited an inhibition zone diameter of 20.6 mm against the yeast strain C. albicans, while both Aspergillus strains (A. niger and A. flavus) showed an equal inhibition zone diameter of 18.3 mm. PPE, renowned for its high content of phenolic acids, ellagitannins (such as punicalin and punicalagin), and flavonoids, has attracted considerable attention as a potential source for the development of antimicrobial agents due to its diverse biological functions and potential health benefits [26,27]. Several studies have consistently demonstrated the antibacterial and antifungal activities of PPE [9,27,28,29]. Notably, the antimicrobial efficacy of PPE has been found to surpass that of other plant parts, with its activity being closely associated with the presence of total flavonoids and tannins [27].
Table 2
Inhibition zones diameter (mm) of PPE, antibacterial (neomycin), and antifungal (itraconazole) against the investigated microorganisms
| Microorganism | PPE | Neomycin | Itraconazole |
|---|---|---|---|
| B. subtilis | 20.6 ± 0.4c | 19.6 ± 0.8 | — |
| Staphylococcus aureus | 22.4 ± 0.6b | 17.5 ± 0.7 | — |
| Escherichia coli | 18.2 ± 0.8d | 24.3 ± 1.1 | — |
| Pseudomonas aeruginosa | 24.5 ± 0.5a | 25.4 ± 1.0 | — |
| C. albicans | 20.6 ± 0.4c | — | 27.5 ± 1.1 |
| A. niger | 18.3 ± 0.7d | — | 26.4 ± 0.6 |
| A. flavus | 18.3 ± 0.7d | — | 19.7 ± 0.3 |
| Source of variation | |||
| F | 225.76 | ||
| P | 0.0000 | ||
| LSD at 0.05 | 0.48 | ||
Different letters denate significant differences at the significance level of 0.05.
Furthermore, PPE has been extensively investigated for its antimicrobial effects against various pathogens, including foodborne bacteria such as E. coli and B. subtilis, as well as fungi like F. sambucinum and Penicillium [27,28]. Noteworthy findings include the potent inhibition of spore germination and mycelial growth of F. sambucinum by PPE at a concentration of 20 mg/mL [28]. Furthermore, PPE has exhibited inhibitory effects against Gram-positive bacteria, such as S. aureus, and Gram-negative bacteria, such as Salmonella spp. [30]. In the context of nosocomial infections, Candida species have been of particular concern, and PPE has been investigated for its potential to treat these pathogens. Researchers have explored ethanolic extracts derived from various parts of the pomegranate fruit, including arils, seeds, pericarp, and peels, and have reported their effectiveness against Candida species [31]. Overall, the wealth of research conducted by multiple scholars consistently affirms the antimicrobial activity of PPE. Its ability to inhibit the growth and development of both bacteria and fungi has been demonstrated, with its efficacy attributed to its bioactive constituents, including phenolic acids, ellagitannins, and flavonoids. These findings highlight the potential of PPE as a valuable source for the development of novel antimicrobial agents with broad-spectrum activity.
3.4 LC–ESI–MS/MS identification of PPE phenolic compounds
The LCMS spectra of PPE (as presented in Table 3 and Figure 3a and b) exhibited a diverse array of phenolic compounds within the extract. Both positive and negative modes were utilized for analysis. The results revealed the presence of 16 distinct phenolic compounds in PPE, each with varying retention times and peak areas. Among these compounds, gallic acid was identified as the most abundant, constituting approximately 86.47% of the total phenolic content. Chlorogenic acid and ellagic acid were also present in notable quantities, accounting for 7.35% and 3.64% of PPE composition, respectively. The remaining phenolic compounds were detected in minor proportions, with catechin (0.17%), caffeic acid (0.83%), syringic acid (0.94%), coumaric acid (0.11%), vanillin (0.09%), ferulic acid (0.12%), naringenin (0.17%), rosmarinic acid (0.08%), daidzein (0.01%), cinnamic acid (0.01%), kaempferol (0.02%), and hesperetin (0.01%) being identified. However, three compounds, namely pyrocatechol, rutin, and quercetin, were not detected in the PPE sample under investigation.
Table 3
Identification of phenolic compounds in PPE using LC–ESI–MS/MS technique
| Peak no. | RT (min) | Compound name | Area % | Chemical formula |
|---|---|---|---|---|
| 1 | 3.597 | Gallic acid | 86.47 | C7H6O5 |
| 2 | 4.251 | Chlorogenic acid | 7.35 | C16H18O9 |
| 3 | 4.487 | Catechin | 0.17 | C15H14O6 |
| 4 | 5.501 | Methyl gallate | 0.00 | C8H8O5 |
| 5 | 5.927 | Caffeic acid | 0.83 | C9H8O4 |
| 6 | 6.425 | Syringic acid | 0.94 | C9H10O5 |
| 7 | 6.635 | Pyrocatechol | 0.00 | C6H6O2 |
| 8 | 6.902 | Rutin | 0.00 | C27H30O16 |
| 9 | 7.237 | Ellagic acid | 3.64 | C14H6O8 |
| 10 | 8.681 | Coumaric acid | 0.11 | C9H8O3 |
| 11 | 9.097 | Vanillin | 0.09 | C8H8O3 |
| 12 | 9.735 | Ferulic acid | 0.12 | C10H10O4 |
| 13 | 10.401 | Naringenin | 0.17 | C15H12O5 |
| 14 | 11.842 | Rosmarinic acid | 0.08 | C18H16O8 |
| 15 | 16.001 | Daidzein | 0.01 | C15H10O4 |
| 16 | 17.324 | Quercetin | 0.00 | C15H10O7 |
| 17 | 19.263 | Cinnamic acid | 0.01 | C9H8O2 |
| 18 | 20.620 | Kaempferol | 0.02 | C15H10O6 |
| 19 | 21.209 | Hesperetin | 0.01 | C16H14O6 |
RT = Retention time (min.).
Figure 3
(a) LC–ESI–MS/MS of standard phenolic compounds. (b) LC–ESI–MS/MS of PPE phenolic compounds.
PPE is known to contain a rich assortment of phenolic compounds, which are responsible for its diverse biological activities and potential health benefits. The identification and characterization of these phenolic compounds in PPE have been the focus of numerous studies. Among these compounds, gallic acid was found to be the most abundant, constituting approximately 86.47% of the total phenolic content. Gallic acid is known for its antioxidant properties and potential health benefits, including anti-inflammatory and anticancer activities [32]. Chlorogenic acid, another prominent phenolic compound in PPE, accounted for 7.35% of the composition. Chlorogenic acid has been associated with various health benefits, such as cardioprotective, neuroprotective, and anti-diabetic effects. Its antioxidant and anti-inflammatory properties contribute to its therapeutic potential [33]. Ellagic acid, as a commonly detectable phenolic compound in PPE, present at a concentration of 3.64%, is known for its strong antioxidant activity and potential anticancer properties. It has been studied for its ability to inhibit the growth of cancer cells and induce apoptosis [34].
Although gallic acid, chlorogenic acid, and ellagic acid were the major phenolic compounds identified in PPE, minor quantities of other phenolic compounds were also detected. These include catechin, caffeic acid, syringic acid, coumaric acid, vanillin, ferulic acid, naringenin, rosmarinic acid, daidzein, cinnamic acid, kaempferol, and hesperetin. Each of these compounds possesses unique bioactive properties and contributes to the overall antioxidant and antimicrobial potential of PPE. It is worth noting that three phenolic compounds, namely pyrocatechol, rutin, and quercetin, were not detected in the PPE sample analyzed. These compounds, commonly found in other plant sources, may be present in PPE at concentrations below the detection limit of the analytical method employed. All in all, understanding the composition of phenolic compounds in PPE is crucial for exploring its therapeutic potential and developing novel applications in the fields of medicine, nutrition, and functional foods.
3.5 Oxidative stability (Rancimat) test
The Rancimat test serves as a means to quantify the susceptibility of fats, oils, and fat-containing food products to oxidation, thereby assessing their oxidation stability. This test involves subjecting a sample of fat or oil to an elevated temperature while passing an airstream through it. Consequently, the fat molecules within the sample undergo oxidation, resulting in the formation of volatile organic compounds and other byproducts. The duration of oxidation indicates the level of resistance displayed by the fatty molecules against this process.
In Table 4, the impact of PPE on the oxidative stability of four plant-based oil samples (corn, olive, sunflower, and soybean oils) is summarized. The data reveal that all the oil samples examined exhibited relatively short stability times. Specifically, the order of stability for the oils was as follows: corn > soybean > sunflower > olive oil, with stability times of 6.8, 4.1, 3.5, and 3.2 h, respectively. Introducing PPE to the oil samples at a concentration of 200 ppm significantly enhanced their stability times, surpassing those achieved with the standard antioxidant BHT. For instance, when PPE was added to the corn oil sample, the stability time increased to 12.6 h, compared to 11.0 h with BHT. In the case of olive oil, stability time reached 8.2 h with PPE, whereas it was 7.1 h with BHT. Similarly, supplementation of sunflower oil with PPE resulted in a stability time of 9.2 h, whereas BHT yielded a stability time of 8.3 h. Soybean oil demonstrated a stability time of 10.1 h in the presence of PPE, while BHT achieved 8.5 h. Consequently, the presence of PPE significantly prolonged the stability of the tested oil samples more effectively than the conventionally employed BHT.
Table 4
The oxidative stability of the investigated plant-based oil samples treated with 200 ppm PPE and BHT
| Sample | Stability time (h) |
|---|---|
| C | 6.8 ± 0.2f |
| C PPE | 12.6 ± 0.3a |
| C BHT | 11.0 ± 0.4b |
| O | 3.2 ± 0.1h |
| O PPE | 8.2 ± 0.3e |
| O BHT | 7.1 ± 0.4f |
| F | 3.5 ± 0.1gh |
| F PPE | 9.2 ± 0.7d |
| F BHT | 8.3 ± 0.6e |
| B | 4.1 ± 0.4g |
| B PPE | 10.1 ± 0.8c |
| B BHT | 8.5 ± 0.1de |
| Statistics | |
| F | 129.29 |
| P | 0.0000 |
| LSD at 0.05 | 0.76 |
Different letters denate significant differences at the significance level of 0.05.
When PPE was added to the oil samples at a concentration of 200 ppm, a significant improvement in stability time was observed compared to the standard antioxidant BHT. These findings indicate that PPE has a greater capacity to extend the oxidative stability of plant-based oils compared to BHT, a commonly used synthetic antioxidant. Similar results were reported in many investigations [35,36,37,38]. The polyphenols present in PPE are known for their strong antioxidant properties, which can effectively scavenge and neutralize free radicals, thereby inhibiting the oxidation process. In their study, Bashir et al. [39] determined that PPE demonstrated antioxidant properties that might be utilized to extend the storage duration of fatty products. They showed that adding the extracts at the maximum concentration (1,000 ppm) led to a reduction in free fatty acids (FFAs), iodine value, saponification value, PV, and thiobarbituric acid reactive substances compared to the control sample, which was supplemented with 200 ppm BHT. Furthermore, the study conducted by Hemachandra et al. [40] examined the impact of a PPE (2% w/w) on the oxidative stability of several edible oils (sunflower oil, coconut oil, palm oil, and sesame oil) during the deep frying process of potato strips at a temperature of 170 ± 5°C. This ability to delay oxidation is crucial in preserving the quality, flavor, and nutritional value of oils, as oxidative degradation can lead to rancidity, off-flavors, and nutrient loss. The results of this study suggest that incorporating PPE as a natural antioxidant in the food industry could be a promising alternative to synthetic antioxidants. Not only does PPE offer improved stability times, but it also aligns with the growing consumer demand for natural and clean-label ingredients. Further research and exploration of the specific polyphenols present in PPE and their mechanisms of action would provide valuable insights into harnessing their antioxidative potential for the preservation of plant-based oils and other food products.
3.6 Changes in PV
The determination of PV serves to measure the concentration of hydroperoxide, which represents the primary oxidation products found in plant-based oils. PV is commonly expressed as milliequivalents (meq) of peroxide oxygen per 1 kg of the oil under analysis [41]. This parameter holds significant importance as it is widely employed to assess the oxidation state of plant-based oils. In the present study, Figure 4 displays the PV of four plant-based oils, namely olive, corn, sunflower, and soybean, over a period of ten days. The investigation focuses on the impact of two treatments: PPE and the synthetic antioxidant BHT. The results indicate that the PV exhibited a time-dependent trend, reaching its maximum level on the 10th day for all oils examined under the respective treatments. Furthermore, the peroxidation values showed the following ranking: sunflower oil > olive oil > soybean oil > corn oil, with mean values of 49.0, 40.0, 20.0, and 16.0 meq O2/kg, respectively.
Figure 4
Changes in PV (meq O2/kg) of corn oil (a), olive oil (b), sunflower oil (c), and soybean oil (d) under accelerated oxidation conditions (at 70°C for 10 days) using 200 ppm PPE as a natural antioxidant in comparison with control (without any antioxidant) and positive control using 200 ppm BHT. C = corn oil, O = olive oil, F = sunflower oil, and B = soybean oil. Different letters denote significant differences at the significance level of 0.05.
Interestingly, the treatment of the investigated oils with PPE (200 ppm) led to a significant reduction in the PV compared to the non-treated oil samples throughout the experiment. Specifically, on the 10th day of treatment, the PV of corn, sunflower, olive, and soybean oils exhibited decreases of 166.7, 145.0, 100.0, and 33.3%, respectively, in relation to their untreated counterparts. Notably, the decline in PV resulting from the addition of PPE was more pronounced than that observed with the addition of BHT in most cases during the entire experimental duration for the various oil samples. The fact that the decline in PV due to PPE addition was more significant than that resulting from BHT addition suggests that PPE exhibits superior antioxidant performance in most cases during the experimental period. The same result was obtained by many authors using various oil samples [35,37,38,42]. This finding implies that PPE might be more efficient at inhibiting the formation of hydroperoxides and preventing oil oxidation compared to BHT. This finding has implications for enhancing product stability, improving shelf life, and minimizing potential health risks associated with the consumption or use of oxidized oils.
3.7 Changes in AV
The AV is a widely used parameter for assessing the quality of fats and oils. It quantifies the amount of FFAs present in a given fat or oil sample by measuring the weight of potassium hydroxide (KOH) required to neutralize the organic acids in 1 g of the sample. An increase in FFA content indicates the hydrolysis of triglycerides, and it serves as an indicator of rancidity, as FFAs are typically formed during triglyceride decomposition [43]. In this study, the AVs of four plant-based oils (olive, corn, sunflower, and soybean) were monitored over a 10-day period, focusing on the effects of two treatments: PPE and the synthetic antioxidant BHT. Figure 5 presents the time-dependent trends of the AVs under each treatment. The findings revealed that the AVs reached their highest levels on the 10th day for all oils tested. Moreover, the AVs exhibited the following ranking: corn oil > soybean oil > olive oil > sunflower oil, with mean values of 16.2, 7.0, 6.0, and 4.0 mg KOH/kg, respectively.
Figure 5
Changes in AV (mg KOH/kg) of corn oil (a), olive oil (b), sunflower oil (c), and soybean oil (d) under accelerated oxidation conditions (at 70°C for 10 days) using 200 ppm PPE as a natural antioxidant in comparison with control (without any antioxidant) and positive control using 200 ppm BHT. C = corn oil, O = olive oil, F = sunflower oil, and B = soybean oil. Different letters denote significant differences at the significance level of 0.05.
Interestingly, the application of PPE at a concentration of 200 ppm resulted in a significant reduction in the AVs compared to the untreated oil samples throughout the experiment. Specifically, on the 10th day of treatment, the AVs of corn, soybean, olive, and sunflower oils decreased by 2214.3, 218.2, 66.7, and 65.0%, respectively, in relation to their untreated counterparts. Notably, the decline in AVs due to PPE addition was generally more pronounced than that observed with the addition of BHT across most of the experimental duration for the different oil samples. This suggests that PPE demonstrates superior stability performance compared to BHT in resisting the hydrolytic degradation of the investigated oils. Similar results were affirmed by researchers [35,38,44].
Kaderides et al. [37] demonstrated that the incorporation of PPE into various food products, including meat, fish, poultry, dairy products, edible oils, and fats, resulted in enhanced physicochemical and microbiological stability during processing and storage, leading to extended shelf-life. Importantly, the addition of PPE did not adversely affect the sensory properties or overall acceptability of the final products, indicating promising potential. These findings suggest that PPE may be more effective than the commonly used commercial antioxidant BHT in preventing oil hydrolysis. Because of lipid radicals, including alkoxyl (RO), peroxyl (ROO), and alkyl (R) radicals, are generated during autooxidation of polyunsaturated fatty acids, by factoring in the quantity of free radical scavenging chemicals at the outset and the amount of time needed for oxidation to consume these compounds [12]. These findings hold implications for improving the stability of food products, prolonging their shelf life, and reducing potential health hazards associated with the consumption or use of partially hydrolyzed oils.
4 Conclusion
In conclusion, the significance of PPE in inhibiting the growth of pathogenic microorganisms threatening human life, preventing oil rancidity, and reducing the oils’ hydrolysis cannot be overstated. PPE, rich in antioxidant phenolic compounds, has demonstrated remarkable potential in enhancing food quality and safety. One of the key advantages of PPE is that its inclusion in various food products could enhance their microbiological stability, thereby extending their shelf life. Additionally, PPE, with its high content of antioxidant phenolic compounds, effectively inhibits the hydrolysis of triglycerides, thus reducing the formation of FFAs. This property is crucial for preserving the freshness and quality of oils, as rancidity not only impairs sensory attributes but also poses potential health risks. Consequently, the utilization of PPE represents a promising approach in the food industry for combating microorganisms, reducing plant-based oils’ rancidity, and preventing oil hydrolysis. Further research and exploration of the potential applications of PPE in different food systems are warranted to fully harness their benefits and unlock their diverse functionalities.
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Funding information: Authors state no funding involved.
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Author contributions: All authors have accepted responsibility for the entire content of this manuscript and consented to its submission to the journal, reviewed all the results and approved the final version of the manuscript. GAA conceptualization, visualization, methodology, validation, data curation, software, investigation, writing – original draft preparation, writing – reviewing and editing. ASA conceptualization, visualization, methodology, validation, data curation, investigation, writing – reviewing and editing.
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Conflict of interest: Authors state no conflict of interest.
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Data availability statement: The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.
[1] Mendes PM, Gomes Fontoura GM, Rodrigues LDS, Souza AS, Viana JPM, Fernandes Pereira AL, et al. Therapeutic potential of Punica granatum and isolated compounds: Evidence-based advances to treat bacterial infections. Int J Microbiol. 2023;2023:e4026440. 10.1155/2023/4026440.Search in Google Scholar PubMed PubMed Central [2] Alsaedi S, Aljeddani G. Phytochemical analysis and bioactivity screening of primary and secondary metabolic products of medicinal plants in the Valleys of Medina Region Saudi Arabia. Adv Biol Chem. 2022;12:92–115.10.4236/abc.2022.124009Search in Google Scholar [3] Kalaycıoğlu Z, Erim FB. Total phenolic contents, antioxidant activities, and bioactive ingredients of juices from pomegranate cultivars worldwide. Food Chem. 2017;221:496–507. 10.1016/j.foodchem.2016.10.084.Search in Google Scholar PubMed [4] Amri Z, Zaouay F, Lazreg-Aref H, Soltana H, Mneri A, Mars M, et al. Phytochemical content, fatty acids composition and antioxidant potential of different pomegranate parts: Comparison between edible and non edible varieties grown in Tunisia. Int J Biol Macromol. 2017;104:274–80. 10.1016/j.ijbiomac.2017.06.022.Search in Google Scholar PubMed [5] Rahnemoon P, Jamab MS, Dakheli MJ, Bostan A. Phenolic compounds and antimicrobial properties of pomegranate (Punica granatum) peel extracts. Int J Agric Biosyst Eng. 2016;10:646–51.Search in Google Scholar [6] Rosas-Burgos EC, Burgos-Hernández A, Noguera-Artiaga L, Kačániová M, Hernández-García F, Cárdenas-López JL, et al. Antimicrobial activity of pomegranate peel extracts as affected by cultivar. J Sci Food Agric. 2017;97:802–10. 10.1002/jsfa.7799.Search in Google Scholar PubMed [7] Pagliarulo C, De Vito V, Picariello G, Colicchio R, Pastore G, Salvatore P, et al. Inhibitory effect of pomegranate (Punica granatum L.) polyphenol extracts on the bacterial growth and survival of clinical isolates of pathogenic Staphylococcus aureus and Escherichia coli. Food Chem. 2016;190:824–31. 10.1016/j.foodchem.2015.06.028.Search in Google Scholar PubMed [8] Ferrazzano GF, Scioscia E, Sateriale D, Pastore G, Colicchio R, Pagliuca C, et al. In vitro antibacterial activity of pomegranate juice and peel extracts on cariogenic bacteria. Biomed Res Int. 2017;2017:e2152749. 10.1155/2017/2152749.Search in Google Scholar PubMed PubMed Central [9] Al-Zoreky NS. Antimicrobial activity of pomegranate (Punica granatum L.) fruit peels. Int J Food Microbiol. 2009;134:244–8. 10.1016/j.ijfoodmicro.2009.07.002.Search in Google Scholar PubMed [10] Kumar YR, Narayanaswamy HD, Rao S, Satyanarayana ML, Nadoor P, Rathnamma D. Effects of pomegranate (Punica granatum) juice and peel extract on biochemical parameters in streptozotocin induced diabetic rats. Pharma Innov J. 2022;11:970–7.Search in Google Scholar [11] Cai H, You S, Xu Z, Li Z, Guo J, Ren Z, et al. Novel extraction methods and potential applications of polyphenols in fruit waste: A review. J Food Meas Charact. 2021;15:3250–61. 10.1007/s11694-021-00901-1.Search in Google Scholar [12] El-Beltagi HS, Hussein Y, Hendawy EAA, El-Masry R, Al-Gaby A, Osman A. Improving oxidative stability of corn oil by curcumin and beta-carotene under accelerated oxidation conditions. Pol J Env Stud. 2024;33:117–24. 10.15244/pjoes/171572.Search in Google Scholar [13] Danial EN, Basudan N. Comparative study as antioxidant, antimicrobial activities and total phenolic content between various parts of pomegranate. Res J Life Sci Bioinf Pharm Chem Sci. 2019;5:674–84. 10.26479/2019.0502.49.Search in Google Scholar [14] Bassiri-Jahromi SP, Pourshafie MRP, Mirabzade Ardakani EDVM, Ehsani AHMD, Doostkam AMD, Katirae FP, et al. In vivo comparative evaluation of the pomegranate (Punica granatum) peel extract as an alternative agent to nystatin against oral candidiasis. Iran J Med Sci. 2018;43:296–304.Search in Google Scholar [15] Konsoula Z. Comparative efficacy of pomegranate juice, peel and seed extract in the stabilization of corn oil under accelerated conditions. Int J Nutr Food Eng. 2016;10:556–63.Search in Google Scholar [16] Cruz-Valenzuela MR, Ayala-Soto RE, Ayala-Zavala JF, Espinoza-Silva BA, González-Aguilar GA, Martín-Belloso O, et al. Pomegranate (Punica granatum L.) peel extracts as antimicrobial and antioxidant additives used in alfalfa sprouts. Foods. 2022;11:e2588. 10.3390/foods11172588.Search in Google Scholar PubMed PubMed Central [17] Singh B, Singh JP, Kaur A, Singh N. Phenolic compounds as beneficial phytochemicals in pomegranate (Punica granatum L.) peel: A review. Food Chem. 2018;261:75–86. 10.1016/j.foodchem.2018.04.039.Search in Google Scholar PubMed [18] Fawole OA, Makunga NP, Opara UL. Antibacterial, antioxidant and tyrosinase-inhibition activities of pomegranate fruit peel methanolic extract. BMC Complement Altern Med. 2012;12:e200. 10.1186/1472-6882-12-200.Search in Google Scholar PubMed PubMed Central [19] Sihag S, Pal A, Ravikant, Saharan V. Antioxidant properties and free radicals scavenging activities of pomegranate (Punica granatum L.) peels: An in-vitro study. Biocatal Agric Biotechnol. 2022;42:e102368. 10.1016/j.bcab.2022.102368.Search in Google Scholar [20] Cervantes-Anaya N, Azpilcueta-Morales G, Estrada-Camarena E, Ramírez Ortega D, Pérez de la Cruz V, González-Trujano ME, et al. Pomegranate and its components, punicalagin and ellagic acid, promote antidepressant, antioxidant, and free radical-scavenging activity in ovariectomized rats. Front Behav Neurosci. 2022;16:e836681. 10.3389/fnbeh.2022.836681.Search in Google Scholar PubMed PubMed Central [21] Saparbekova AA, Kantureyeva GO, Kudasova DE, Konarbayeva ZK, Latif AS. Potential of phenolic compounds from pomegranate (Punica granatum L.) by-product with significant antioxidant and therapeutic effects: A narrative review. Saudi J Biol Sci. 2023;30:e103553. 10.1016/j.sjbs.2022.103553.Search in Google Scholar PubMed PubMed Central [22] Berkoz M, Yalin S, Yildirim M, Yalın AE, Çömelekoğlu Ü. Punicalagin and punicalin suppress the adipocyte differentiation through the transcription factors. Acta Endocrinol. 2021;17:157–67. 10.4183/aeb.2021.157.Search in Google Scholar PubMed PubMed Central [23] Diab TA, Donia T, Saad-Allah KM. Characterization, antioxidant, and cytotoxic effects of some Egyptian wild plant extracts. Beni-Suef Univ J Basic Appl Sci. 2021;10:e13. 10.1186/s43088-021-00103-0.Search in Google Scholar [24] Saleem A, Akhtar MF, Sharif A, Akhtar B, Siddique R, Ashraf GM, et al. Anticancer, cardio-protective and anti-inflammatory potential of natural-sources-derived phenolic acids. Molecules. 2022;27:e7286.10.3390/molecules27217286Search in Google Scholar PubMed PubMed Central [25] Tanveer A, Farooq U, Akram K, Hayat Z, Shafi A, Nazar H, et al. Pomegranate extracts: A natural preventive measure against spoilage and pathogenic microorganisms. Food Rev Int. 2015;31:29–51. 10.1080/87559129.2014.961074.Search in Google Scholar [26] Türkyılmaz M, Tağı Ş, Dereli U, Özkan M. Effects of various pressing programs and yields on the antioxidant activity, antimicrobial activity, phenolic content and colour of pomegranate juices. Food Chem. 2013;138:1810–8. 10.1016/j.foodchem.2012.11.100.Search in Google Scholar PubMed [27] Kharchoufi S, Licciardello F, Siracusa L, Muratore G, Hamdi M, Restuccia C. Antimicrobial and antioxidant features of ‘Gabsiʼ pomegranate peel extracts. Ind Crop Prod. 2018;111:345–52. 10.1016/j.indcrop.2017.10.037.Search in Google Scholar [28] Abdollahzadeh S, Mashouf R, Mortazavi H, Moghaddam M, Roozbahani N, Vahedi M. Antibacterial and antifungal activities of Punica granatum peel extracts against oral pathogens. J Dent. 2011;8:1–6.Search in Google Scholar [29] Malviya S, Arvind X, Jha A, Hettiarachchy N. Antioxidant and antibacterial potential of pomegranate peel extracts. J Food Sci Technol. 2014;51:4132–7. 10.1007/s13197-013-0956-4.Search in Google Scholar PubMed PubMed Central [30] Ali A, Chen Y, Liu H, Yu L, Baloch Z, Khalid S, et al. Starch-based antimicrobial films functionalized by pomegranate peel. Int J Biol Macromol. 2019;129:1120–6. 10.1016/j.ijbiomac.2018.09.068.Search in Google Scholar PubMed [31] Alexandre EMC, Silva S, Santos SAO, Silvestre AJD, Duarte MF, Saraiva JA, et al. Antimicrobial activity of pomegranate peel extracts performed by high pressure and enzymatic assisted extraction. Food Res Int. 2019;115:167–76. 10.1016/j.foodres.2018.08.044.Search in Google Scholar PubMed [32] Jiang Y, Pei J, Zheng Y, Miao Y, Duan B, Huang L. Gallic acid: A potential anti-cancer agent. Chin J Integr Med. 2022;28:661–71. 10.1007/s11655-021-3345-2.Search in Google Scholar PubMed [33] Singh AK, Singla RK, Pandey AK. Chlorogenic acid: A dietary phenolic acid with promising pharmacotherapeutic potential. Curr Med Chem. 2023;30:3905–26.10.2174/0929867329666220816154634Search in Google Scholar PubMed [34] Eroglu Ozkan E, Seyhan MF, Kurt Sirin O, Yilmaz- Ozden T, Ersoy E, Hatipoglu Cakmar SD, et al. Antiproliferative effects of Turkish pomegranate (Punica granatum L.) extracts on MCF-7 human breast cancer cell lines with focus on antioxidant potential and bioactive compounds analyzed by LC-MS/MS. J Food Biochem. 2021;45:e13904. 10.1111/jfbc.13904.Search in Google Scholar PubMed [35] Drinić Z, Mudrić J, Zdunić G, Bigović D, Menković N, Šavikin K. Effect of pomegranate peel extract on the oxidative stability of pomegranate seed oil. Food Chem. 2020;333:e127501. 10.1016/j.foodchem.2020.127501.Search in Google Scholar PubMed [36] Abd-Allah IMA, Rabie MA, Sulieman AM, Mostfa DM, El-Badawi AA. Oxidative stability of edible oils via addition of pomegranate and orange peel extracts. Foods Raw Mater. 2018;6:413–20. 10.21603/2308-4057-2018-2-413-420.Search in Google Scholar [37] Kaderides K, Kyriakoudi A, Mourtzinos I, Goula AM. Potential of pomegranate peel extract as a natural additive in foods. Trends Food Sci Technol. 2021;115:380–90. 10.1016/j.tifs.2021.06.050.Search in Google Scholar [38] Iqbal S, Haleem S, Akhtar M, Zia-ul-Haq M, Akbar J. Efficiency of pomegranate peel extracts in stabilization of sunflower oil under accelerated conditions. Food Res Int. 2008;41:194–200. 10.1016/j.foodres.2007.11.005.Search in Google Scholar [39] Bashir S, Gilani SA, Shah F, Khan AA, Ullah S. Preparation of chicken nuggets using pomegranate blended sunflower oil. Asian J Hallied Heal Sci. 2016;1:3–10.Search in Google Scholar [40] Hemachandra TP, Jayathilake RR, Madhujith WM. The effect of antioxidative extracts on mitigating autoxidation of selected edible oils during deep frying. Trop Agric Res. 2017;28:247–55. 10.4038/tar.v28i3.8229.Search in Google Scholar [41] Roselló-Soto E, Barba FJ, Lorenzo JM, Munekata PES, Gómez B, Moltó JC. Phenolic profile of oils obtained from “horchata” by-products assisted by supercritical-CO2 and its relationship with antioxidant and lipid oxidation parameters: Triple TOF-LC-MS-MS characterization. Food Chem. 2019;274:865–71. 10.1016/j.foodchem.2018.09.055.Search in Google Scholar PubMed [42] El-Hadary AE, Taha M. Pomegranate peel methanolic-extract improves the shelf-life of edible-oils under accelerated oxidation conditions. Food Sci Nutr. 2020;8:1798–811. 10.1002/fsn3.1391.Search in Google Scholar PubMed PubMed Central [43] Dijkstra AJ. Vegetable oils: Composition and analysis. In: Caballero B, Finglas PM, Toldrá F, editors. Encycl. Food Heal. Oxford: Academic Press; 2016. p. 357–64. 10.1016/B978-0-12-384947-2.00708-X.Search in Google Scholar [44] Javani-Seraji S, Bazargani-Gilani B, Aghajani N. Influence of extraction techniques on the efficiency of pomegranate (Punica granatum L.) peel extracts in oxidative stability of edible oils. Food Sci Nutr. 2023;11:2344–55. 10.1002/fsn3.3244.Search in Google Scholar PubMed PubMed Central References
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