|Year : 2021 | Volume
| Issue : 1 | Page : 18-23
Antibacterial potency of extracted essential oils of some plant species against common gram-positive and gram-negative bacteria
Gaffar Sarwar Zaman
Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Khalid University, Abha, Kingdom of Saudi Arabia
|Date of Submission||29-Jan-2021|
|Date of Decision||08-Mar-2021|
|Date of Acceptance||15-Mar-2021|
|Date of Web Publication||31-Jul-2021|
Gaffar Sarwar Zaman
Department of Clinical Laboratory Sciences, College of Applied Medical Sciences, King Khalid University, Abha
Kingdom of Saudi Arabia
Source of Support: None, Conflict of Interest: None
Background: In the last decades, due to the rapid emergence of multidrug resistant pathogens, the antibiotic-resistance phenomenon has become a global health crisis. Therefore, there is a need to find new remedies against pathogenic microbes. Objectives: The main intention of this research was to appraise the antibacterial potency of extracted essential oils (EEOs) from various plant species versus human disease-causing bacterial strains. Materials and Methods: Antibacterial and bactericidal activity of EEOs was tested on human disease-causing strains which included Gram-positive as well as Gram-negative bacteria. Antibacterial analysis for various extracts of the different plants was performed by utilizing the method of disc diffusion and deduction of minimum inhibitory concentration (MIC) by microbroth dilution assays of the EEOs against the bacterial strains. Standard antibiotics (amoxicillin, metronidazole, rifampicin, clarithromycin, oxacillin, and clindamycin) were used to compare with EEO antibacterial activity. Results: Eclipta alba EEO was most effective against Streptococcus Pyogenes (2.06 ± 0.15), Neisseria gonorrhoeae (1.50 ± 0.20), Streptococcus aureus (0.05 ± 0.02), and Pseudomonas aeruginosa (4.56 ± 0.25). Leucas linifolia EEO was most effective against E. coli (3.13 ± 0.25) and Klebsiella Pneumoniae (4.33 ± 0.23). Bactericidal activity EEO from E. alba with minimum bactericidal concentration (MBC) ranged from 0.11 ± 0.03 to 10.60 ± 0.55; Atriplex hortensis (8.73 ± 2.62–12.07 ± 0.65); Hedyotis scandens (9.13 ± 0.50–15.30 ± 0.43); L. linifolia (0.94 ± 0.05–10.73 ± 0.20); Murraya koenigii (9.0 ± 0.55–12.90 ± 0.18); and Phlogacanthus thyrsiflorus (5.96 ± 1.15–13.0 ± 0.52). Bactericidal activity E. alba EEO was highest against S. Pyogenes (4.06 ± 0.15), N. gonorrhoeae (3.06 ± 0.40), and S. aureus (0.11 ± 0.03). L. linifolia EEO was most effective against P. aeruginosa (0.94 ± 0.05) and K. Pneumoniae (8.73 ± 0.41). Against E. coli (5.96 ± 1.15), the bactericidal activity of P. thyrsiflorus EEO was most effective. Conclusions: Comparison to the antibacterial activity of EEOs from six different plant species used in the study was more effective than the tested antibiotics. MIC and MBC values show that E. alba EEO plant species was the most effective against the tested human pathogenic bacterial strains.
Keywords: Antibacterial potency, disc diffusion, human disease, minimal inhibitory concentration
|How to cite this article:|
Zaman GS. Antibacterial potency of extracted essential oils of some plant species against common gram-positive and gram-negative bacteria. King Khalid Univ J Health Sci 2021;6:18-23
|How to cite this URL:|
Zaman GS. Antibacterial potency of extracted essential oils of some plant species against common gram-positive and gram-negative bacteria. King Khalid Univ J Health Sci [serial online] 2021 [cited 2021 Nov 28];6:18-23. Available from: https://www.kkujhs.org/text.asp?2021/6/1/18/322880
| Introduction|| |
There is widespread utilization of antibiotics in agriculture, clinical medicine, and veterinary which elevates the expansion of resistance to antibiotics among contagious strains of microbes and ultimately reviews a very significant issue in the therapy of disease-causing microbes;, this has triggered the pursuance of novel antimicrobial drugs predominantly among extracts of plant to unearth new chemical compositions which overwhelm the given impediments.,
Microbial resistance is one of the most prominent problems to the present antibiotics that have given rise to a sustained exploration for a novel antimicrobial drug with powerful potency and rock-bottom price. A great requirement is there to find possible good compounds for extracting novel antibacterial and antifungal drugs from the plant extracts.
Medicinal plants are an overflowing origin of molecules from which antibiotic compounds may be derived. An extensive range of plant extracts that have medicinal values is utilized to medicate different types of infections as they have inherent antimicrobial potency. Many of these plant extracts are sold in the market as raw material form without screening for the reasons of their herbal properties. Many experts have now turned their concentration back toward researching on these bioactive molecules to obtain advantages from these medicinal plants for humankind, after noticing more side effects of the synthetic drugs contrasting to their advantages.,
Innumerable researches have been performed to check the antimicrobial possibilities of plant extracts and their essential oils, and it is well accepted that numerous essential oils can inhibit or destroy microorganisms. According to Enrico et al., the essential oils, which contrasting to antibiotics, are built up of many different molecules so that mutation of the bacteria cannot give rise to resistance. When utilized in prevention and also in the cure, these are uniquely and remarkably recognized for their potential antioxidant, anti-inflammatory, antiviral, antibacterial, larvicidal, and antifungal effects., Various antimicrobial activities of the extracted essential oil (EEO) in the form of popular and commercially available preparations have been utilized for the treatment of fungal and bacterial infections all over the world.,
Higher plants possess natural products which, in some cases, may be a new origin of antimicrobes with a feasibly novel medium for infectious disease treatments. Active principles are incorporated in medicinal plants which can be utilized as alternative herbal medications against various infections caused by bacteria. Therefore, various plants that might have these medicinal qualities should be investigated thoroughly for their properties and efficiency. The commercial use of essential oils encompasses different industries such as agronomic, pharmaceutical, food, cosmetics sanitary, and perfume. In the medical industry, they are mostly utilized as antitumor, antioxidant, and against bacteria and fungi.
There have been various studies on the antibacterial activities of different plants versus Gram-positive bacteria as well as most Gram-negative bacterial strains, but there are only a few studies on bacterial drug-resistant activity. Hence, the present research was dedicated to gauge and assess the in vitro bacterial demolition activity of EEO extracted from six different plant species against six pathogenic bacterial strains, i.e., Neisseria gonorrhoeae (Gram-negative), Streptococcus Pyogenes (Gram-positive), and also Pseudomonas aeruginosa (Gram-negative), Acinetobacter baumannii (Gram-negative), Klebsiella pneumoniae (Gram-negative), and Escherichia coli (Gram-negative) and explained their inhibitory consequences as compared to the commercially available antibiotics.
| Materials and Methods|| |
Organisms and chemicals
In the current study, the antibacterial activity of EEO from six various plant species was tested against six human pathogenic strains including S. Pyogenes, N. gonorrhoeae, Streptococcus aureus, E. Coli, K. Pneumoniae, and P. aeruginosa.
Six plants from which EEO were extracted; Eclipta alba, Family– Asteraceae, Local name– Kenharaj; Atriplex hortensis, Family– orache, Local name-Pahari palang; Hedyotis scandens, Family– Rubiaceae, Local name-Bhedeli-lota; Leucas linifolia, Family– Lamiaceae, Local name-Doron bon; Murraya koenigii, Family– Rutaceae, Local name–Narasingha; Phlogacanthus thyrsiflorus, Family– Acanthaceae, Local name– Tita-phul. Standard antibiotics (Amoxicillin, Metronidazole, Rifampicin, Clarithromycin, Oxacillin, Clindamycin) purchased from Sigma (Sigma-Aldrich, Switzerland).
Preparation of plant extracts
Plant extracts of six plants used in the study were extracted as per standard method. In brief, the plant substances were powdered in a grinding machine and prepared a 5% watery extract of the plant substance (w/v) in ultrapure sterile water by heating (at 80°C) for 3 min in three successive cycles. Filtration of the solutions was done utilizing a 0.24 μm membrane. For the preparation of extracts in methanol, the plant substances were immersed in methanol concentration of 95% having a ratio of approximately 1:10 (w/v) for a period of 74 h at 24°C with shaking in a robust manner. Afterward, methanol evaporation was done for the extracts, under conditions of specific vacuumed pressure, and resulting particular residual substances were taken as the main source of required methanolic extract. Furthermore, stock solutions preparation was done in DMSO (Merck, Darmstadt, Germany), and ultimate working volumes from them were finally attained by regular dilution of the main stock into the Mueller-Hinton (MH) broth (Oxoid, Hampshire, UK).
Antimicrobial susceptibility assays
The method of disc-diffusion (Paper) was implemented for the ascertainment of antimicrobial activities. All tests were performed five times and repeated twice. EEO from plants was examined to find antimicrobial activity utilizing the diffusion method of agar well. Inoculation of the cultures of bacteria was done in LB broth media for a period of 3 h at a temperature of 37°C, and the turbidity was modified to 0.5 McFarland's index which was in saline of phosphate-buffered. Then, 20 μl of EEO was transferred from various plants (2 mg/mL) into each well contained in the Petri dishes and anaerobic incubation done for 1 day at a temperature of 37°C. Columbia blood agar plates which contained 7% of lake horse blood (from Oxoid, Hampshire, UK) furnished a platform for the Helicobacter and Campylobacter located assays. The plates were then incubated in conditions of microaerophilic utilizing a CampyGen kit (from Oxoid, Hampshire, UK) at 37°C for a period of 2 days. The zone of inhibition diameter of bacterial growth on all sides of each well was calculated in millimeters as described by Rojas et al. and Farooqui et al.,
Minimum inhibitory concentration determination
Determination of minimum inhibitory concentrations (MICs) was done by assays microbroth dilution utilizing MH broth. Assays of microbroth dilution were utilized to ascertain minimum inhibitory concentrations (MICs) of the concerned EEO extracted from various plants versus the different strains of bacteria. The EEO concentration utilized for MICs ranged from 5000 to 50 μg/mL. In brief, two-fold dilutions of 100 μL culture of extracts for each of the strains were loaded in flat-bottom 96-well plate duplicate wells of the polystyrene. The inoculum which was started for each strain was 1.5 × 105 CFU/mL, and those wells which contained bacterial inoculum minus any compound served as a control. Incubation of the plates was done as described above. The least concentration of the compounds that depicted neither visible growth of bacteria nor any turbidity after a period of 24 h of intense incubation in the dilution assay of the microbroth was contemplated as MIC. Triplicate repetitions of the experiments were done for each strain.
Ascertainment of minimum bactericidal concentration
To ascertain the minimum bactericidal concentration (MBC) of the concerned EEO, subculture of 100 μl of MH broth was done from each well of the microbroth assay; they were done on MH agar plates after a period of 1 day of initial incubation. Incubation of the MH plates was done for another 24 h. The least concentration of extract that emanated in no bacterial growth was regarded as MBC. Quadruplicate repetitions of the experiments were done on five different occasions.
| Results|| |
Antibacterial activity by microbroth dilution assay of the EEO from six different plant species demonstrated against six tested pathogenic bacterial strains. Antibacterial activity against six tested pathogenic bacterial strains of EEO from E. alba with MIC values ranged from 0.05 ± 0.02 to 5.0 ± 0.30; A. hortensis with MIC values ranged from 3.53 ± 0.35 to 6.0 ± 0.26; H. scandens with MIC values ranged from 4.60 ± 0.30 to 7.50 ± 0.36; L. linifolia with MIC values ranged from 3.13 ± 0.25 to 5.50 ± 0.36; M. koenigii with MIC values ranged from 4.43 ± 0.32 to 6.30.26; and P. thyrsiflorus with MIC values ranged from 3.13 ± 0.40 to 6.50 ± 0.26.
The bactericidal activity against six tested pathogenic bacterial strains of EEO from E. alba with MBC values ranged from 0.11 ± 0.03 to 10.60 ± 0.55; A. hortensis with MBC values ranged from 8.73 ± 2.62 to 12.07 ± 0.65; H. scandens with MBC values ranged from 9.13 ± 0.50 to 15.30 ± 0.43; L. linifolia with MBC values ranged from 0.94 ± 0.05 to 10.73 ± 0.20; M. koenigii with MBC values ranged from 9.0 ± 0.55 to 12.90 ± 0.18; and P. thyrsiflorus with MBC values ranged from 5.96 ± 1.15 to 13.0 ± 0.52.
Antibacterial activity by paper disc-diffusion method of the EEO from E. alba, H. scandens, L. linifolia, and P. thyrsiflorus plant species shows their antibacterial activity against the six tested pathogenic bacterial strains, i.e., S. pyogenes, N. gonorrhoeae, S. aureus, E. coli, K. pneumoniae, and P. aeruginosa. However, the EEO from A. hortensis shows their antibacterial activity against three tested pathogenic bacterial strains, i.e., S. pyogenes, S. aureus, and E. coli, and the EEO from M. koenigii shows their antibacterial activity against four tested pathogenic bacterial strains, i.e., S. pyogenes, N. gonorrhoeae, S. aureus, and E. coli [Table 1].
|Table 1: Antibacterial activity of different plant extracted oils against pathogenic bacteria strains of humans|
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Of the EEO from six different plant species tested against six pathogenic bacterial strains, the most effective EEO was of E. alba followed by L. linifolia, P. thyrsiflorus, H. scandens, A. hortensis, and M. koenigii.
In comparison to the antibacterial activity by the disc-diffusion method of EEO from six different plant species used in the study was more effective than the tested antibiotics (amoxicillin [Figure 1], metronidazole [Figure 2], rifampicin [Figure 3], clarithromycin [Figure 4], oxacillin [Figure 5], and clindamycin [Figure 6]), as shown in [Table 2]. Antibacterial activity by the disc-diffusion method of the EEO from E. alba was most effective against N. gonorrhoeae (22.60 ± 2.50), K. pneumoniae (19.40 ± 1.50), and P. aeruginosa (13.70 ± 1.49). However, against S. aureus (20.50 ± 3.10) and E. coli (26.20 ± 1.54), EEO from H. scandens was most effective. Against S. Pyogenes (32.90 ± 1.37), EEO from A. hortensis was most effective [Table 1].
|Figure 1: Streptococcus pyogenes and zones of inhibition (mean zones of inhibition in mm) against standard antibiotics|
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|Figure 2: Neisseria gonorrhoeae and zones of inhibition (mean zones of inhibition in mm) against standard antibiotics|
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|Table 2: Zones of inhibition (mean zones of inhibition in mm) against standard antibiotics|
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|Figure 3: Streptococcus aureus and zones of inhibition (mean zones of inhibition in mm) against standard antibiotics|
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|Figure 4: Escherichia coli and zones of inhibition (mean zones of inhibition in mm) against standard antibiotics|
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|Figure 5: Klebsiella pneumoniae and zones of inhibition (mean zones of inhibition in mm) against standard antibiotics|
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|Figure 6: Pseudomonas aeruginosa and zones of inhibition (mean zones of inhibition in mm) against standard antibiotics|
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Antibacterial activity by the microbroth dilution method of the EEO from E. alba was most effective against S. pyogenes (2.06 ± 0.15), N. gonorrhoeae (1.50 ± 0.20), S. aureus (0.05 ± 0.02), and P. aeruginosa (4.56 ± 0.25). However, against E. coli (3.13 ± 0.25) and K. Pneumoniae (4.33 ± 0.23), the EEO from L. linifolia was most effective [Table 1].
The bactericidal activity of the EEO from E. alba was most effective against S. pyogenes (4.06 ± 0.15), N. gonorrhoeae (3.06 ± 0.40), and S. aureus (0.11 ± 0.03). The bactericidal activity of the EEO from L. linifolia was most effective against P. aeruginosa (0.94 ± 0.05) and K. Pneumoniae (8.73 ± 0.41). However, against E. coli (5.96 ± 1.15), the bactericidal activity of the EEO from P. thyrsiflorus was most effective [Table 1].
| Discussion|| |
Medicinal plants possessing antimicrobial properties are being increasingly reported from various parts of the earth. Even the World Health Organization states that extracts from plant or active constituents from them are utilized as folk medicine in various types of traditional remedies of almost 80% of the world's population. In this particular study, the author investigated the antibacterial and also the bactericidal activity of the EEO from six different plant species by paper disc-diffusion and microbroth dilution assays. Comparison of antibacterial activity of EEO from six different plant species with standard antibiotics used against the tested human pathogenic bacterial strains was done and was confirmed to be more effective than the tested antibiotics.
Antibacterial activity by the microbroth dilution method of the EEO from E. alba was most effective against S. pyogenes (2.06 ± 0.15), S. aureus (0.05 ± 0.02), N. gonorrhoeae (1.50 ± 0.20), and P. aeruginosa (4.56 ± 0.25). The minimal MICs were procured in the particular alkaloid compound which was having 57 μg/ml versus S. aureus, 42 μg/ml versus E. coli, 61 μg/ml versus S. boydii, 82 ug/ml versus P. aeruginosa, and 89 μg/ml versus S. faecalis. In this study, the E. coli and S. aureus are found to be more susceptible than the other selected human pathogenic bacteria.
In my study, the EEO from L. linifolia was most effective against E. coli (3.13 ± 0.25) and K. pneumoniae (4.33 ± 0.23). Begum et al. reported the antibacterial activity of the isolated secondary metabolites from L. lavandulaefolia Rees. (Family-Labiatae) Syn. L. linifolia Spreng against the tested B. subtilis, S. aureus, E. coli, and C. herbarium. Tahareen et al. studied antibacterial activity on L. aspera leaves showed that E. coli was inhibited at all concentrations, followed by Klebsiella and Pseudomonas., Gowrish et al. studied antibacterial activity of L. marrubioides, and the results depicted that petroleum ether extract was very much effective against Gram-positive bacteria (B. cereus, S. Pyogenes, and B. subtilis) and Gram-negative bacteria (Proteus mirabilis, K. pneumoniae, and V. cholera).,
One of the major alternatives to overwhelm the elevated levels of resistance to drugs is the discovery of newer antimicrobial metabolites from novel medicinal plants. Newer antimicrobial substance from various medicinal plants is a substitute to overthrow the elevated levels of drug resistance posed by many human pathogens. Novel antibiotics and related chemotherapeutic agents' research is increasing in the chemistry of medicinal plants. Medicinal plants are an important source of antioxidants. Various natural antioxidants discovered from plant sources elevate the antioxidant capacity of the plasma and lead to the reduction of the danger from certain diseases. Antioxidant activity of the plasma is increased by natural antioxidants that help in reducing the danger of diseases.,
Screening of antibacterial properties of medicinal plants is being increasingly reported from all over the world. The above results of antibacterial activity showed that all the EEOs from six different plant species tested were significant against studied pathogenic microorganisms. In all cases, the plant extracts were more potent compared to antibiotics used against the microorganisms. Overall, the antibacterial and bactericidal activity of the EEO of E. alba was noticeably more effective against the growth of bacterial strains compared to the EEO of other plant species studied.
The affirmation of antibacterial activity found versus both Gram-positive and also Gram-negative bacteria by the source E. alba could perhaps indicate that broad-spectrum antibiotic compounds can be manufactured from it., The present research upholds the claimed utilization of E. alba in the traditional system of medicine to cure different infections which are caused by the microbes. The present research supports the cultivation of this valuable medicinal plant to meet the elevated demand from the traditional medicinal system.
| Conclusions|| |
Comparison to the antibacterial activity of EEOs from six different plant species used in the study was more effective than the tested antibiotics. MIC and MBC values show the EEO from E. alba followed by L. linifolia, P. thyrsiflorus, H. scandens, A. hortensis, and M. koenigii. Plant species was the most effective against the tested human pathogenic bacterial strains. Properly utilized, the medicinal characteristics of different plant species will make an extraordinary benefaction to the origin and evolution of many different traditional herbal medicines.
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Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6]
[Table 1], [Table 2]