Volume 9, Issue 3 (Journal of Clinical and Basic Research (JCBR) 2025)
jcbr 2025, 9(3): 22-26 |
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Jafari-Sales A, Golestani A, Ghahremani Z, Pashazadeh M. Evaluation of the antibacterial properties of methanolic pomegranate peel extract against standard strains of Staphylococcus aureus, Bacillus cereus, Pseudomonas aeruginosa, and Escherichia coli: An in Vitro Study. jcbr 2025; 9 (3) :22-26
URL: http://jcbr.goums.ac.ir/article-1-532-en.html
URL: http://jcbr.goums.ac.ir/article-1-532-en.html
1- Department of Microbiology, Kaz.C., Islamic Azad University, Kazerun, Iran; Infectious Diseases Research Center, TaMS.C., Islamic Azad University, Tabriz, Iran
2- Infectious Diseases Research Center, TaMS.C., Islamic Azad University, Tabriz, Iran; Department of Cellular and Molecular Biology, Ta.C., Islamic Azad University, Tabriz, Iran
3- Infectious Diseases Research Center, TaMS.C., Islamic Azad University, Tabriz, Iran; Department of Laboratory Sciences and Microbiology, TaMS.C., Islamic Azad University, Tabriz, Iran ,Mehrdadpashazadeh85@gmail.com
2- Infectious Diseases Research Center, TaMS.C., Islamic Azad University, Tabriz, Iran; Department of Cellular and Molecular Biology, Ta.C., Islamic Azad University, Tabriz, Iran
3- Infectious Diseases Research Center, TaMS.C., Islamic Azad University, Tabriz, Iran; Department of Laboratory Sciences and Microbiology, TaMS.C., Islamic Azad University, Tabriz, Iran ,
Keywords: Pomegranate, Anti-infective agents, Plant extracts, In vitro techniques, Microbial sensitivity tests
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Discussion
Currently, infectious diseases remain a leading cause of morbidity and mortality around the globe. The discovery of effective treatments for microbial diseases has grabbed increasing attention for disease control, particularly with regard to multidrug-resistant infections. Alkaloids, anthraquinones, saponins, tannins, and polyphenols, which are abundantly present in plants, are rich sources for developing novel antimicrobial agents (22). These natural sources, especially plant-derived bioactive chemicals, have been studied as complements or substitutes for traditional antimicrobial medications in order to address the rising issue of antimicrobial resistance and the pressing need for efficient therapies for bacterial infections (23,24). Accordingly, the present study confirmed the notable performance of PPME as a potential therapeutic agent to fight against pathogenic bacterial strains. In a study by Dahham et al., the antibacterial effects of the methanolic extracts of different parts of pomegranate (Seed, fruit, peel, and juice) were evaluated on some bacteria, including S. aureus, reporting that the peel extract of pomegranate had the strongest antimicrobial activity (25). Similarly, we found that PPME exhibited the MIC and MBC values of 50 and 100 mg/mL against S. aureus, respectively, with a mean inhibition zone of 23.7 mm. These results are consistent with some previous reports but differ from others. For example, Malviya et al. reported an inhibition zone of 24.5 ± 0.53 mm for pomegranate methanolic extract against S. aureus (26), which was close to the value obtained in our study. In another study by Alam Khan et al. in India, the PPME exhibited a minimum inhibition zone of 22 mm against P. aeruginosa (27), which was consistent with our findings.
In contrast, Yassin et al. (2021) (28) reported considerably lower MIC and MBC values for the PPME against S. aureus (MIC and MBC of 0.125 and 0.250 mg/mL, respectively) compared to our study. Moreover, in the recent report, E. coli revealed an MIC of 50 mg/mL and an inhibition zone diameter of 23.7 mm. Likewise, Hamrita et al. reported the MIC and MBC values of 9.37 and 18.75 mg/mL, respectively, for P. aeruginosa (29), which differed from our results. In the study conducted by Al-Hassnawi, at the concentrations of 50 and 100 mg/mL, the smallest inhibition zones were related to P. aeruginosa (8.33±0.57 mm and 12.16±0.57 mm, respectively), while the largest inhibition zones at these concentrations were related to S. aureus (19±1 mm and 21±1 mm, respectively) (30). These results partially aligned with our findings. In another study by Sweidan et al., the highest antimicrobial activity of pomegranate methanolic extract was observed against P. aeruginosa (MIC50 = 7.5 mg/mL) (31), which contradicted our findings. Alnees et al. tested the antimicrobial activity of three types of pomegranate extract against P. aeruginosa, asserting that the most potent effects belonged to the methanolic extract (32). In another study, the antibacterial activity (Based on growth inhibition zones) of pomegranate peel extract was considerable against S. aureus and P. aeruginosa, but not against E. coli (33). According to Akarca et al., the PPME possessed the highest antimicrobial effects, where B. cereus was susceptible to all extracts except aqueous seed extract. In this study, the inhibition zone diameter for PPME was 25.68 ± 0.45 mm against B. cereus and 25.14 ± 0.75 mm against E. coli, indicating that the greatest antimicrobial effect belonged to B. cereus (Mean zone diameter: 21.00 ± 0.05, P > 0.05). In another experiment, the MIC and MBC values of PPME for B. cereus were reported as 62.5 and 15.63 μg/mL, respectively (34), showing some deviations from the findings of the present study. Meléndez and Capriles (2006) used the disk diffusion technique to assess the antibacterial activity of a number of tropical plants from Puerto Rico against S. aureus and E. coli, reporting that pomegranate extract generated growth inhibitory zones of 20 and 11 mm for these bacteria, respectively (35). The antibacterial activity variations observed between our findings and previous reports may be attributed to several factors, including harvest seasons, plant age, geographical origins, extraction methods, drying techniques, and bacterial growth stage.
The most salient finding of this study was the unexpected and significantly higher sensitivity of Gram-negative bacteria, E. coli and P. aeruginosa, to PPME, as demonstrated by remarkably lower MIC and MBC values compared to Gram-positive strains. These findings suggest that particular bioactive chemicals in the extract, namely punicalagins, ellagitannins, and flavonoids, may target structural weaknesses peculiar to Gram-negative bacteria’s envelope, while they cannot penetrate the thick peptidoglycan layer of Gram-positive bacteria. By interacting with and destabilizing the lipopolysaccharides that anchor to structural components, these polyphenols, which are well-known for their metal-chelating and membrane-disrupting capabilities, can efficiently permeabilize the outer membrane. The strong bactericidal effects observed may be the consequence of this first breach, which subsequently allow additional antimicrobial components to enter and enhance damage to the cytoplasmic membrane and the underlying thin peptidoglycan layer. The superior efficacy against Gram-negative organisms can be partly explained by this phenomenon, highlighting the role of outer membrane disruption rather than targeting the peptidoglycan layer, further elaborating on the novelty of our work.
Conclusion
In summary, this study not only confirmed the potent efficacy of the PPME against bacterial pathogens but also revealed a remarkable finding, namely the unique susceptibility patterns of Gram-negative vs. Gram-positive bacteria, as demonstrated by notably lower MIC and MBC values of the former. This discovery opens new avenues for future research into the mechanisms of action and the development of novel antibacterial agents derived from natural compounds. Future studies should include a detailed phytochemical profiling of PPME using techniques such as GC-MS or HPLC to identify and quantify the specific bioactive compounds responsible for the observed antibacterial effects. Furthermore, our findings establish a foundation for designing innovative antimicrobial formulations to cope with infections caused by Gram-negative pathogens and manage antimicrobial resistance.
Acknowledgement
None.
Funding sources
This study received no institutional or university-provided funding.
Ethical statement
Not applicable.
Conflicts of interest
Authors declare no conflict of interests.
Author contributions
Concept/Design: A.J.S, M.P; Data acquisition: A.J.S; Data analysis and Interpretation: A.G, Z.G; Drafting the manuscript: A.G, Z.G; Critical revision of the manuscript: AJS; Final approval and Accountability: MP; Technical or material support: A.J.S, M.P; Supervision: MP; Securing funding (If available): N/A.
Data availability statement
The authors confirm that the data supporting the findings of this study are included in the article.
Full-Text: (88 Views)
Introduction
The use of medicinal plants was one of humanity's earliest innovations for treating diseases, nurturing a close relationship between humans and plants throughout human evolution (1-4). Natural plant derivatives, despite their biodiversity, represent an essential resource for drug discovery. With their broad applications, these compounds serve both as direct therapeutic agents and as building blocks for designing and synthesizing novel drugs (5-7). Plants and their extracts, which have long been used in many areas of traditional medicine, abundantly contain a wide range of structurally diverse secondary metabolites with multiple biological functions (8).
A major healthcare challenge today is the treatment of antimicrobial-resistant infectious diseases, drawing significant attention toward medicinal plants as safer and natural alternatives for synthetic agents. Consequently, global research is steadily expanding to divulge the antibacterial properties of various plant species (9-12). Medicinal plants have gained widespread acceptance in human societies for numerous reasons, including their natural origin, excellent safety, and cost-effectiveness. These natural remedies typically exhibit fewer side effects and, due to their superior compatibility with the human body's genetic and physiological makeup, represent a suitable option for most people (13-17). Research has established that the bioactive compounds of medicinal plants are not limited to their fruits but are also abundant in non-edible parts such as leaves, peels, and seeds. A notable example is pomegranate (Punica granatum L.), a member of the Lythraceae family native to Central Asia. The peel, seeds, and leaves of pomegranate contain substantial levels of phenolic compounds, contributing to its biological functions as an effective therapeutic agent. Scientific studies have confirmed the potent antioxidant, anti-inflammatory, antifungal, antibacterial, and antimicrobial properties of pomegranate (18,19). Pomegranate extracts prepared via different techniques exhibit antimicrobial activity against various Gram-positive and Gram-negative bacteria, including Staphylococcus aureus (S. aureus), Bacillus subtilis (B. subtilis), and Bacillus cereus (B. cereus), as well as against yeasts and molds such as Candida albicans (C. albicans) and Fusarium oxysporum (F. oxysporum) (20). Pomegranate peel methanolic extract (PPME) has shown significant antibacterial effects, particularly against Gram-positive bacteria such as S. aureus, which has been attributed to its rich punicalagin and polyphenol content. Also, PPME has been effective against Gram-negative bacteria like Escherichia coli (E. coli), indicating its capacity to disrupt the cellular integrity of these microorganisms (21). The objective of this study was to evaluate the in vitro antibacterial activity of PPME against four standard pathogenic bacteria, S. aureus, B. cereus, Pseudomonas aeruginosa (P. aeruginosa), and E. coli.
Methods
This study was conducted in the research laboratory of the Islamic Azad University, Tabriz Medical Sciences Branch. Pomegranate (Punica granatum) peels were collected in autumn from the city of Marand. The formal identification of the plant was undertaken by Ms. Fatemeh Chobineh, a botanist at the Islamic Azad University, Ardabil Branch. The methanolic extract was prepared using a Soxhlet apparatus using 500 mL of methanol as the solvent and 300 grams of powdered pomegranate peel at 40°C for 8 hours. The solvent was then removed using a rotary evaporator. Standard bacterial strains, S. aureus (ATCC: 25923), B. cereus (ATCC: 1247), P. aeruginosa (ATCC: 27853), and E. coli (ATCC 25922), were obtained from the University of Tehran Microorganism Collection Center. The extracts prepared were stored in dark containers in a refrigerator until testing.
The extracts were concentrated to 50, 100, 200, and 400 mg/mL using 5% dimethyl sulfoxide (DMSO) as the solvent. Bacterial strains were exposed to these concentrations to assess antibacterial activity using the well diffusion assay. The microorganisms were cultured on Mueller-Hinton agar (MHA) (Merck, Germany) one the day prior to the experiment. To prepare the bacterial suspension, several colonies from a fresh young bacterial culture were transferred to MHA. The turbidity of the microbial suspension was adjusted to the 0.5 McFarland standard, equivalent to a concentration of 1.5 × 106 CFU/mL, which was achieved by diluting the bacterial suspension to a 1:100 ratio. Next, the antibacterial activity of PPME was evaluated using the agar well diffusion method. The bacterial suspension with a turbidity equivalent to 1.5 × 106 bacteria/mL was spread over three different directions on MHA plates using a sterile cotton swab. Subsequently, wells with a diameter of 6 mm and a depth of 5 mm were created on the agar, maintaining an appropriate distance of 2.5 cm from each other. Each well was exposed to 100 μL of PPME at different concentrations (50, 100, 200, and 400 mg/mL). The negative control contained only the solvent (5% DMSO), and the positive control included streptomycin antibiotic (10 μg) (Padtan Teb Iran).
The plates were incubated at 37°C for 24 hours, after which the diameters of bacterial growth inhibition zones were measured and recorded in millimeters. To ensure the reliability of the findings, all assays for each extract concentration and each bacterium were performed in triplicate. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined using Mueller-Hinton broth (MHB) and MHA (Merck, Germany) and by preparing the serial dilutions of 0.78, 1.56, 3.125, 6.25, 12.5, 25, 50, 100, and 200 mg/mL, where culture media containing bacteria without extract and culture media without bacteria served as positive and negative controls, respectively. The test tubes used for MIC determination were incubated at 37°C for 24 hours, and the lowest extract concentration preventing visible bacterial growth (i.e., no turbidity) was recorded as MIC. To determine MBC, samples were taken from the three tubes preceding the MIC point and cultured on MHA for 24 hours at 37°C, and the concentration showing no growth was considered as the MBC. To minimize experimental errors, each assay was performed in triplicate. SPSS version 26 (SPSS Inc., Chicago, IL, USA) was used to statistically analyze the data using one-way ANOVA.
Results
This study comprehensively investigated the antibacterial activity of PPME against two Gram-positive (S. aureus and B. cereus) and two Gram-negative (E. coli and P. aeruginosa) bacteria. The findings revealed that PPME possessed significant growth inhibitory effects on the bacteria tested at the 50, 100, 200, and 400 mg/mL concentrations (P < 0.0001 for all studied bacteria) (Table 1, Figure 1).
The use of medicinal plants was one of humanity's earliest innovations for treating diseases, nurturing a close relationship between humans and plants throughout human evolution (1-4). Natural plant derivatives, despite their biodiversity, represent an essential resource for drug discovery. With their broad applications, these compounds serve both as direct therapeutic agents and as building blocks for designing and synthesizing novel drugs (5-7). Plants and their extracts, which have long been used in many areas of traditional medicine, abundantly contain a wide range of structurally diverse secondary metabolites with multiple biological functions (8).
A major healthcare challenge today is the treatment of antimicrobial-resistant infectious diseases, drawing significant attention toward medicinal plants as safer and natural alternatives for synthetic agents. Consequently, global research is steadily expanding to divulge the antibacterial properties of various plant species (9-12). Medicinal plants have gained widespread acceptance in human societies for numerous reasons, including their natural origin, excellent safety, and cost-effectiveness. These natural remedies typically exhibit fewer side effects and, due to their superior compatibility with the human body's genetic and physiological makeup, represent a suitable option for most people (13-17). Research has established that the bioactive compounds of medicinal plants are not limited to their fruits but are also abundant in non-edible parts such as leaves, peels, and seeds. A notable example is pomegranate (Punica granatum L.), a member of the Lythraceae family native to Central Asia. The peel, seeds, and leaves of pomegranate contain substantial levels of phenolic compounds, contributing to its biological functions as an effective therapeutic agent. Scientific studies have confirmed the potent antioxidant, anti-inflammatory, antifungal, antibacterial, and antimicrobial properties of pomegranate (18,19). Pomegranate extracts prepared via different techniques exhibit antimicrobial activity against various Gram-positive and Gram-negative bacteria, including Staphylococcus aureus (S. aureus), Bacillus subtilis (B. subtilis), and Bacillus cereus (B. cereus), as well as against yeasts and molds such as Candida albicans (C. albicans) and Fusarium oxysporum (F. oxysporum) (20). Pomegranate peel methanolic extract (PPME) has shown significant antibacterial effects, particularly against Gram-positive bacteria such as S. aureus, which has been attributed to its rich punicalagin and polyphenol content. Also, PPME has been effective against Gram-negative bacteria like Escherichia coli (E. coli), indicating its capacity to disrupt the cellular integrity of these microorganisms (21). The objective of this study was to evaluate the in vitro antibacterial activity of PPME against four standard pathogenic bacteria, S. aureus, B. cereus, Pseudomonas aeruginosa (P. aeruginosa), and E. coli.
Methods
This study was conducted in the research laboratory of the Islamic Azad University, Tabriz Medical Sciences Branch. Pomegranate (Punica granatum) peels were collected in autumn from the city of Marand. The formal identification of the plant was undertaken by Ms. Fatemeh Chobineh, a botanist at the Islamic Azad University, Ardabil Branch. The methanolic extract was prepared using a Soxhlet apparatus using 500 mL of methanol as the solvent and 300 grams of powdered pomegranate peel at 40°C for 8 hours. The solvent was then removed using a rotary evaporator. Standard bacterial strains, S. aureus (ATCC: 25923), B. cereus (ATCC: 1247), P. aeruginosa (ATCC: 27853), and E. coli (ATCC 25922), were obtained from the University of Tehran Microorganism Collection Center. The extracts prepared were stored in dark containers in a refrigerator until testing.
The extracts were concentrated to 50, 100, 200, and 400 mg/mL using 5% dimethyl sulfoxide (DMSO) as the solvent. Bacterial strains were exposed to these concentrations to assess antibacterial activity using the well diffusion assay. The microorganisms were cultured on Mueller-Hinton agar (MHA) (Merck, Germany) one the day prior to the experiment. To prepare the bacterial suspension, several colonies from a fresh young bacterial culture were transferred to MHA. The turbidity of the microbial suspension was adjusted to the 0.5 McFarland standard, equivalent to a concentration of 1.5 × 106 CFU/mL, which was achieved by diluting the bacterial suspension to a 1:100 ratio. Next, the antibacterial activity of PPME was evaluated using the agar well diffusion method. The bacterial suspension with a turbidity equivalent to 1.5 × 106 bacteria/mL was spread over three different directions on MHA plates using a sterile cotton swab. Subsequently, wells with a diameter of 6 mm and a depth of 5 mm were created on the agar, maintaining an appropriate distance of 2.5 cm from each other. Each well was exposed to 100 μL of PPME at different concentrations (50, 100, 200, and 400 mg/mL). The negative control contained only the solvent (5% DMSO), and the positive control included streptomycin antibiotic (10 μg) (Padtan Teb Iran).
The plates were incubated at 37°C for 24 hours, after which the diameters of bacterial growth inhibition zones were measured and recorded in millimeters. To ensure the reliability of the findings, all assays for each extract concentration and each bacterium were performed in triplicate. Minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC) were determined using Mueller-Hinton broth (MHB) and MHA (Merck, Germany) and by preparing the serial dilutions of 0.78, 1.56, 3.125, 6.25, 12.5, 25, 50, 100, and 200 mg/mL, where culture media containing bacteria without extract and culture media without bacteria served as positive and negative controls, respectively. The test tubes used for MIC determination were incubated at 37°C for 24 hours, and the lowest extract concentration preventing visible bacterial growth (i.e., no turbidity) was recorded as MIC. To determine MBC, samples were taken from the three tubes preceding the MIC point and cultured on MHA for 24 hours at 37°C, and the concentration showing no growth was considered as the MBC. To minimize experimental errors, each assay was performed in triplicate. SPSS version 26 (SPSS Inc., Chicago, IL, USA) was used to statistically analyze the data using one-way ANOVA.
Results
This study comprehensively investigated the antibacterial activity of PPME against two Gram-positive (S. aureus and B. cereus) and two Gram-negative (E. coli and P. aeruginosa) bacteria. The findings revealed that PPME possessed significant growth inhibitory effects on the bacteria tested at the 50, 100, 200, and 400 mg/mL concentrations (P < 0.0001 for all studied bacteria) (Table 1, Figure 1).
|
Table 1. Bacterial Growth inhibition zone (mm) of pomegranate peel methanolic extract at various concentrations
.PNG) *The results of one-way ANOVA showed statistically significant differences between various concentrations for all cases (p < 0.0001 for all bacteria). The F-values were as follows: S. aureus: 285.65, B. cereus: 104.87, E. coli: 75.23, and P. aeruginosa: 31.15. .PNG) Figure 1. Zone of growth inhibition related to pomegranate peel methanolic extract against S. aureus, B. cereus, E. coli and P. aeruginosa isolates |
Regarding the MIC and MBC values calculated, there was a distinct sensitivity pattern among these bacteria. Gram-negative strains (E. coli and P. aeruginosa) showed remarkably higher sensitivity to PPME, evidenced by much lower MIC values (E. coli: 12.5 mg/mL and P. aeruginosa: 25 mg/mL) compared to Gram-positive bacteria (S. aureus: 50 mg/mL and B. cereus: 100 mg/mL) (Figure 2). This pattern was also evident in MBC values, where E. coli (With MBC equivalent to 25 mg/mL) was identified as the most sensitive bacteria, and B. cereus (With MBC equivalent to 200 mg/mL) was the most resistant microorganism (Table 2).
.PNG)
.PNG)
Discussion
Currently, infectious diseases remain a leading cause of morbidity and mortality around the globe. The discovery of effective treatments for microbial diseases has grabbed increasing attention for disease control, particularly with regard to multidrug-resistant infections. Alkaloids, anthraquinones, saponins, tannins, and polyphenols, which are abundantly present in plants, are rich sources for developing novel antimicrobial agents (22). These natural sources, especially plant-derived bioactive chemicals, have been studied as complements or substitutes for traditional antimicrobial medications in order to address the rising issue of antimicrobial resistance and the pressing need for efficient therapies for bacterial infections (23,24). Accordingly, the present study confirmed the notable performance of PPME as a potential therapeutic agent to fight against pathogenic bacterial strains. In a study by Dahham et al., the antibacterial effects of the methanolic extracts of different parts of pomegranate (Seed, fruit, peel, and juice) were evaluated on some bacteria, including S. aureus, reporting that the peel extract of pomegranate had the strongest antimicrobial activity (25). Similarly, we found that PPME exhibited the MIC and MBC values of 50 and 100 mg/mL against S. aureus, respectively, with a mean inhibition zone of 23.7 mm. These results are consistent with some previous reports but differ from others. For example, Malviya et al. reported an inhibition zone of 24.5 ± 0.53 mm for pomegranate methanolic extract against S. aureus (26), which was close to the value obtained in our study. In another study by Alam Khan et al. in India, the PPME exhibited a minimum inhibition zone of 22 mm against P. aeruginosa (27), which was consistent with our findings.
In contrast, Yassin et al. (2021) (28) reported considerably lower MIC and MBC values for the PPME against S. aureus (MIC and MBC of 0.125 and 0.250 mg/mL, respectively) compared to our study. Moreover, in the recent report, E. coli revealed an MIC of 50 mg/mL and an inhibition zone diameter of 23.7 mm. Likewise, Hamrita et al. reported the MIC and MBC values of 9.37 and 18.75 mg/mL, respectively, for P. aeruginosa (29), which differed from our results. In the study conducted by Al-Hassnawi, at the concentrations of 50 and 100 mg/mL, the smallest inhibition zones were related to P. aeruginosa (8.33±0.57 mm and 12.16±0.57 mm, respectively), while the largest inhibition zones at these concentrations were related to S. aureus (19±1 mm and 21±1 mm, respectively) (30). These results partially aligned with our findings. In another study by Sweidan et al., the highest antimicrobial activity of pomegranate methanolic extract was observed against P. aeruginosa (MIC50 = 7.5 mg/mL) (31), which contradicted our findings. Alnees et al. tested the antimicrobial activity of three types of pomegranate extract against P. aeruginosa, asserting that the most potent effects belonged to the methanolic extract (32). In another study, the antibacterial activity (Based on growth inhibition zones) of pomegranate peel extract was considerable against S. aureus and P. aeruginosa, but not against E. coli (33). According to Akarca et al., the PPME possessed the highest antimicrobial effects, where B. cereus was susceptible to all extracts except aqueous seed extract. In this study, the inhibition zone diameter for PPME was 25.68 ± 0.45 mm against B. cereus and 25.14 ± 0.75 mm against E. coli, indicating that the greatest antimicrobial effect belonged to B. cereus (Mean zone diameter: 21.00 ± 0.05, P > 0.05). In another experiment, the MIC and MBC values of PPME for B. cereus were reported as 62.5 and 15.63 μg/mL, respectively (34), showing some deviations from the findings of the present study. Meléndez and Capriles (2006) used the disk diffusion technique to assess the antibacterial activity of a number of tropical plants from Puerto Rico against S. aureus and E. coli, reporting that pomegranate extract generated growth inhibitory zones of 20 and 11 mm for these bacteria, respectively (35). The antibacterial activity variations observed between our findings and previous reports may be attributed to several factors, including harvest seasons, plant age, geographical origins, extraction methods, drying techniques, and bacterial growth stage.
The most salient finding of this study was the unexpected and significantly higher sensitivity of Gram-negative bacteria, E. coli and P. aeruginosa, to PPME, as demonstrated by remarkably lower MIC and MBC values compared to Gram-positive strains. These findings suggest that particular bioactive chemicals in the extract, namely punicalagins, ellagitannins, and flavonoids, may target structural weaknesses peculiar to Gram-negative bacteria’s envelope, while they cannot penetrate the thick peptidoglycan layer of Gram-positive bacteria. By interacting with and destabilizing the lipopolysaccharides that anchor to structural components, these polyphenols, which are well-known for their metal-chelating and membrane-disrupting capabilities, can efficiently permeabilize the outer membrane. The strong bactericidal effects observed may be the consequence of this first breach, which subsequently allow additional antimicrobial components to enter and enhance damage to the cytoplasmic membrane and the underlying thin peptidoglycan layer. The superior efficacy against Gram-negative organisms can be partly explained by this phenomenon, highlighting the role of outer membrane disruption rather than targeting the peptidoglycan layer, further elaborating on the novelty of our work.
Conclusion
In summary, this study not only confirmed the potent efficacy of the PPME against bacterial pathogens but also revealed a remarkable finding, namely the unique susceptibility patterns of Gram-negative vs. Gram-positive bacteria, as demonstrated by notably lower MIC and MBC values of the former. This discovery opens new avenues for future research into the mechanisms of action and the development of novel antibacterial agents derived from natural compounds. Future studies should include a detailed phytochemical profiling of PPME using techniques such as GC-MS or HPLC to identify and quantify the specific bioactive compounds responsible for the observed antibacterial effects. Furthermore, our findings establish a foundation for designing innovative antimicrobial formulations to cope with infections caused by Gram-negative pathogens and manage antimicrobial resistance.
Acknowledgement
None.
Funding sources
This study received no institutional or university-provided funding.
Ethical statement
Not applicable.
Conflicts of interest
Authors declare no conflict of interests.
Author contributions
Concept/Design: A.J.S, M.P; Data acquisition: A.J.S; Data analysis and Interpretation: A.G, Z.G; Drafting the manuscript: A.G, Z.G; Critical revision of the manuscript: AJS; Final approval and Accountability: MP; Technical or material support: A.J.S, M.P; Supervision: MP; Securing funding (If available): N/A.
Data availability statement
The authors confirm that the data supporting the findings of this study are included in the article.
Article Type: Research |
Subject:
Microbiology
References
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