Abstract
Background
Buchholzia coriacea is acclaimed to be efficacious in several microbial infections and traditionally used for the treatment of diarrhoea and respiratory infections, among others, thereby, raising its potential relevance in the search for new antimicrobial agents. This study evaluated the phytochemical constituents, in-vitro antimicrobial activities and Minimum Inhibitory Concentration (MIC) of the methanol leaf extract of B. coriacea at lower concentrations in comparison with standard drugs.
Methods
The leaves of B. coriacea were harvested, air-dried, pulverized and extracted exhaustively by cold maceration using methanol. Using agar-well diffusion method, antimicrobial screening was done at various concentrations (ranging between 11.9 mg/ml and 190 mg/ml), after phytochemical screening of the dried extract. Further, antimicrobial effects of the extract on test organisms - Klebsiella spp, Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Candida albicans, were compared to ciprofloxacin and gentamicin.
Results
Tannins, saponins, cardenolides, terpenoids and reducing sugars were present in the extract. Klebsiella spp (0–24 mm) had the highest inhibition zone diametre for the extract, while E. coli (2–20 mm), and C. albican (2–11 mm) were most sensitive at much lower concentration, and no inhibitory effect was observed against Bacillus subtilis (0–0 mm). The MIC was lowest for Klebsiella spp (12.59 mg/ml) and S. aureus (12.59 mg/ml), while E.coli had higher MIC (15.85 mg/ml). Ciprofloxacin and gentamycin had significantly (P < 0.001) lower MIC than the extract for the test organisms.
Conclusion
The methanol leaf extract of B. coriacea comprised several phytochemicals at various concentrations and exhibited appreciable concentration-dependent antimicrobial activities. However, ciprofloxacin and gentamycin showed activity against the test bacteria at significantly lower concentrations than the extract. Nonetheless, the methanol leaf extract of B. coriacea having shown a relatively modest broad-spectrum antimicrobial activity, may be further evaluated for potential pharmaceutical formulations.
Supplementary Information
The online version contains supplementary material available at 10.1186/s12906-025-05057-9.
Keywords: Buchholzia coriacea, Wonderful cola, Leaf extract, Antimicrobial effects, Phytochemical constituents, Nigeria
Introduction
Microbial infections have been a major source of challenge to humanity and a leading cause of morbidity and mortality in some parts of the globe, particularly in developing countries where it is of major public health concern [1, 2]. This has resulted in huge costs to individuals, the immediate community and the global community, thereby, raising the burden of microbial infections, particularly gram-negative bacteria which is highly prevalent [2]. Escherichia coli, Klebsiella pneumonia, and Staphylococcus aureus are also associated with high prevalence of several infections, including urinary tract and wound infections [3–5]. More so, there is increasing threats of antimicrobial resistance to available microbials, as a result of wide misuse of antimicrobial medicines, and the mode of transmission of microbial agents [6–9]. This has raised the risks of spread and harm of pathogenic microbials, urging the need for deliberate preventive measures and the search for new, effective and relevant therapeutic agents. The traditional uses of B. coriacea for several health conditions highlights its potential clinical relevance and positions it for antimicrobial evaluations.
Buchholzia coriacea, (B. coriacea) otherwise called “wonderful cola” is a medicinal plant named after RW Buchholz, and widely used in parts of Nigeria for several purposes [10]. It is a tropical evergreen shrub of forest origin and belongs to the Capparaceae family. Its leaves are large, obstanceolate and glossy, arranged spirally in a cluster at the apex of the tree branches [11, 12]. The plant is commonly used in tropical Africa and acclaimed to possess several health benefits, which range from antimicrobial [13], antihelminthes [14], anti-inflammatory [15, 16], immune-regulatory [12], antidepressants, anxiolytic, analgesics [17], hypoglycemic [18], among others. It is therefore, of potential clinical value and a major source of pharmaceutical drugs. These acclaimed therapeutic potentials of B. coriacea may be associated with its phytochemical components which could vary according to plant parts and plant location [19]. Methods and solvents of extraction may also influence the phytochemical constituents observed during analysis and screening of the plant extract, causing variations in the in vitro activities. Some phytochemicals previously identified in B. coriacea include saponins, cardiac glycosides, carbohydrates, alkaloids, tannins and steroids [14]. Several active plant secondary metabolites have been identified to be very useful to man in diverse ways, including antimicrobial [20, 21]. Hence, plants are continuously explored in the search for new efficacious antimicrobial medicines to reduce the current global burden of antimicrobial resistance.
As a result of the high level and emerging trend of multi-drug resistance associated with the currently available antimicrobial drugs, it is expedient to explore potential new pharmaceutical drugs with comparable or higher efficacy and effectiveness [3–5]. This therefore, urges the scientific evaluation of potential clinically useful plants, to provide alternative new antimicrobial drugs. Previous studies have shown that plants are essentially useful and promising sources of antimicrobial agents [13], positioning them for exploration for potential clinical relevance. Importantly, previous research has focused on parts of B. coriacae such as the seed and bark [11, 13, 15], while there is limited evidence on the antimicrobial properties of the leaves, particularly at lower concentrations, and in comparison with standard antibiotics. Therefore, this study was carried out using lower concentrations of the leaf extract, showed regional specificity, and involved comparison of B. coriacea antimicrobial activity with two specific antibiotics. The study was aimed at evaluating the phytochemical constituents of the leaf extract of B. coriacea, the in-vitro antimicrobial activities of the extract at low doses, and the Minimum Inhibitory Concentration (MIC) against Klebsiella spp, Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Candida albicans, in comparison with ciprofloxacin and gentamycin.
Materials and methods
Sample collection and extraction
With appropriate permission, the leaves of B. coriacea were harvested in Abia State of Nigeria (geographical coordinates - Latitude: 5.4167 Longitude: 7.5000) and authenticated by a botanist - Mr. Alfred Ozioku of the Botany department, University of Nigeria, Nsukka, Nigeria. A voucher specimen of the leaf was deposited in the Faculty of Pharmacy herbarium, Madonna University, Nigeria, with the number MADU 00013. The leaves were thoroughly washed with distilled water to remove adhering particles. Air-drying of the leaves was done at room temperature (between 20 °C and 22 °C) after which they were pulverized. A 100 g of the leaf powder was extracted through cold maceration method by 130 ml methanol. The suspension was left to stand for 24 h, after which it was filtered. This process was repeated until the suspension was colourless (three cycles). The filtrate was dried using rotary evaporator and stored in sterile universal bottle at 4 °C prior to its use. Total percentage yield was calculated as:
 |
Reagents, culture media and equipment
The following reagents were sourced commercially and used for the study: Methanol (BDH materials: repackaged in Nigeria by NAAFCO Akanbi Nnitri close Iganmu industrial Estate, Lagos, Nigeria.), Sulphuric acid (by Fluka Sigma – Aldrich Switzerland), Chloroform (BDH chemicals limited, Poole, England), Dilute ammonia (BDH), Concentrated sulphuric acid (29335 5B General purpose reagent), Ethyl acetate (BDH), Distilled water, Acid alcohol, Acetic acid, Meyer’s reagent, Draggendoff’s reagent, 0.1% ferric chloride (Lab Tech Chemicals) and Fehling solution A & B (BDH). Culture media used were nutrient agar and sabouraud dextrose agar (SDA), other materials included; agar plates, inoculating loop, burnsen burner, petri dishes, conical flasks, cotton wool, test organisms, reference drugs, laboratory oven (surgienfield instrument, England), autoclave (Stainless – steel pressure), rotary evaporator, electric blender and laboratory incubator (Sanfa).
Phytochemical screening
The leaf extract was subjected to phytochemical tests for plants’ secondary metabolites using standard methods [22]. The different qualitative chemical tests were performed to establish the chemical composition of the extracts. Test for alkaloids was performed by diluting 0.5 g of the extract to 10 ml acid alcohol, which was boiled and filtered. To every 5 ml of the filtrate was added 2 ml of dilute ammonia and 5 ml of chloroform, and the mixture was gently shaken to extract the alkaloidal base. The chloroform layer was extracted with 10 ml of acetic acid, and this was divided into two portions. Mayer’s reagent was added to one portion and Draggendorff’s reagent was added to the other. The formation of cream (with Mayer) or reddish-brown precipitate (with Draggendorff’ reagent) indicated the presence of alkaloids.
Test for Cardiac glycosides was performed by diluting 0.5 g of the extract to 5 ml in water and adding 2 ml of glacial acetic acid containing one drop of ferric chloride solution, which was underplayed by 1 ml of concentrated sulphuric acid. A brown ring at the interphase indicated deoxy sugar which is a characteristic of cardenolides. The formation of a violet ring below the brown ring was also anticipated. A greenish ring was also anticipated in the acetic acid layer just above the brown ring which may gradually spread throughout this layer.
Test for tannins was performed by boiling 0.5 g of the extract in 10 ml of water in a test tube, and obtaining the filtrate. Three drops of 0.1% ferric chloride was added and observed for a brownish green or a blue-black coloration.
Test for saponins was done by adding 5 ml of distilled water to 0.5 g of the extract in a test tube. The solution was vigorously shaken and observed for a persistent froth. The frothing was mixed with three drops of olive oil, shaken vigorously and observed for the formation of an emulsion.
Test for flavonoids was performed using two methods. First, 5 ml dilute ammonia was added to 0.5 g of the extract, to which 1 ml of concentrated sulphuric acid was also added. A yellow colouration that disappeared on standing indicated the presence of flavonoids. The second method was performed by heating 0.5 g of the extract with 10 ml of ethyl acetate over a steam bath for 3 min. The mixture was filtered and 4 ml of the filtrate was shaken with 1 ml of dilute ammonia solution. A yellow colouration indicated the presence of flavonoids.
Test for terpinoids (Salkowski test) involved the careful addition of 2 ml of chloroform and 3 ml of concentrated sulphuric acid to 0.5 g of the extract, to form a layer. The appearance of reddish-brown colouration at the interface indicated the presence of terpinoids.
For the test for anthraquinones, 0.5 g of the leaf extract was boiled in 10 ml of hot sulphuric acid and filtered while hot. The filtrate was shaken with 5 ml of chloroform and the chloroform layer was pipette into another test tube where 1 ml of dilute ammonia was added. The resulting solution was then observed for colour changes.
Test for reducing sugar (Fehling test) was performed by adding 0.5 ml of the extract to 5 ml of water, and adding the mixture to a boiling Fehling solution (A and B) in a test tube. The solution was then observed for a colour reaction.
Antimicrobial studies
Test organisms
The micro-organisms used for the study were Klebsiella spp, Staphylococcus aureus, Escherichia coli, Bacillus subtilis and Candida albicans, and were obtained from the clinical collection of the Department of Pharmaceutical Microbiology, Faculty of Pharmacy, Madonna University, Nigeria. The bacteria isolates were first sub-cultured in nutrient broth and incubated at 37 °C for 18 h. These were maintained on nutrient agar slants at 4 °C. The fungi were maintained on sabouraud agar. The antimicrobial study involved the use of test organisms to screen for the inhibitory properties of the extract. Standard antimicrobial agents, which included ciprofloxacin and gentamicin were used as positive controls.
Media preparation
Nutrient agar was prepared using 28 g of the agar which was put in a conical flask. To do this, 1000 ml of distilled water was boiled in volumetric flask using a Bunsen burner and with the aid of wire gauze. The nutrient agar was dispersed into the hot water and allowed to dissolve, after which it was covered with a cotton wool and sterilized by autoclaving at 121 °C for 15 min.
Sensitivity tests and minimum inhibitory concentrations (MIC)
Antimicrobial activity of the crude extract was determined by the agar-well diffusion method [23]. The bacteria isolates were first grown in a nutrient broth for 18 h before use. Nutrient agar was poured into petri dishes and 200 µL of the standardised cell suspension was introduced to the agar plates. Wells were bored into the agar using a 6 mm diametre hole borer to a depth of 4 mm. Five concentrations of the extract; 190 mg/ml, 95 mg/ml, 47.5 mg/ml, 23.8 mg/ml and 11.9 mg/ml, were introduced into the wells, allowed to stand at room temperature for 2 hours and then incubated at 37 °C for 24 h. The plates were observed for zones of inhibition. A plot of log concentrations against IZD2 (square of inhibition zone diametres) was done, and minimum inhibitory concentration (MIC) was determined as antilog of the intercept at log concentration. This procedure was also repeated for the various concentrations of the positive controls. Five concentrations of ciprofloxacin: 50 μm/ml, 25 μm/ml, 12.5 μm/ml, 6.25 μm/ml and 3.13 μm/ml, and five concentrations of gentamycin: 40 μm/ml, 20 μm/ml, 10 μm/ml, 5 μm/ml and 2.5 μm/ml, were used. The fungal isolates were allowed to grow on a sabouraud dextrose agar (SDA) at 25 °C until they sporulated. To harvest the fungi spores, a mixture of sterile glycerol and distilled water was poured to the surface of the plate, then sterile glass rod was used to scrape spores. The harvested fungi spores were standardised by dilution of spores in aqueous solution before use, and 1µL of the standardised fungi spore suspension was evenly spread on the SDA using a glass spreader. Wells were then bored into the agar media to a 4 mm depth using a sterile 6 mm hole borer, and the well was filled with the solution of the extract; taking care not to allow spillage of the solution to the surface of the agar. The plates were allowed to stand on the laboratory bench for 1 h to allow for proper diffusion of the extract into the media. Plates were incubated at 25 °C for 96 h and later observed for zones of inhibition, and the diametres were measured.
Study endpoints
In vitro antibacterial and antifungal effects of the leaf extracts of B. coriacea.
Statistical analysis
Microsoft excel 2007 was used for statistical analysis, using Analysis of Variance (ANOVA). A total of 15 samples were included in the study which was performed in triplicates. Means and standard deviations were calculated to manage data variability. The antibacterial effect was extrapolated from the mean diametre of the inhibition zone diametres of the extract and the standard drugs. A plot of log concentrations against IZD2 (square of inhibition zone diametres) was done, and minimum inhibitory concentration (MIC) determined as antilog of the intercept at log concentration. Bar charts with error bars were also determined. p < 0.05 were considered significant.
Results
Percentage yield of the extract was 15%. The phytochemical screening of the leaf extract showed the presence of terpenoids, tannins, saponins, reducing sugars and cardenolides at varying proportions. However, trepenoids was observed to be more present in the extract than other secondary metabolites. See Table 1.
Table 1.
Classes of phytochemical components measured in the extract
| Constituents | Observations | Inference |
|---|---|---|
| Tannins | ++ | Moderately present |
| Reducing sugars | ++ | Moderately present |
| Saponins | + | Sparingly present |
| Alkaloids | Absent | |
| Terpenoids | +++ | Densely present |
| Flavonoids | Absent | |
| Cardiac glycosides (cardenolides) | ++ | Moderately present |
| Anthraquinones | Absent |
+++ abundant presence, ++ moderate, + slight,– absent
The extract had its highest inhibition zone diametre (IZD) against Klebsiella spp, while E. coli, and C. albican were most sensitive to the extract at a much lower concentration. However, the extract did not show any inhibitory effect against Bacillus subtilis. See Table 2.
Table 2.
Inhibition zones diameters (IZD), mean of IZD (mm) at different concentrations (in triplicates) ± standard error of mean (SEM) and log concentration of the extract
| Test organisms | Inhibition zones diametre (mm) at various concentrations of the extract | ||||
|---|---|---|---|---|---|
| 190 mg/ml | 95 mg/ml | 47.5 mg/ml | 23.8 mg/ml | 11.9 mg/ml | |
| Staphylococcus aureus | 12 | 6 | 3 | 2 | 0 |
| Escherichia coli | 20 | 10 | 5 | 3 | 2 |
| Klebsiella spp | 24 | 12 | 6 | 3 | 0 |
| Bacillus subtilis | 0 | 0 | 0 | 0 | 0 |
| Fungi | |||||
| Candida albicans | 11 | 9 | 5 | 3 | 2 |
| Test organisms | Mean Inhibition zones diametre (mm) of the extract (n = 3) | ||||
|---|---|---|---|---|---|
| 190 mg/ml | 95 mg/ml | 47.5 mg/ml | 23.8 mg/ml | 11.9 mg/ml | |
| Staphylococcus aureus | 13.5 ± 1.5 | 6.5 ± 0.5 | 3.5 ± 0.5 | 2.5 ± 0.5 | 0.0 ± 0.0 |
| Escherichia coli | 21 ± 1.0 | 8.5 ± 1.5 | 6.0 ± 1.0 | 3.5 ± 0.5 | 2.5 ± 0.5 |
| Klebsiella spp | 21.50 ± 2.5 | 13.0 ± 2.0 | 5.5 ± 0.5 | 2.5 ± 0.5 | 0.0 ± 0.0 |
| Candida albicans | 11.5 ± 0.6 | 8.4 ± 0.5 | 5.3 ± 0.9 | 3.5 ± 0.3 | 2 ± 0.4 |
| Square of inhibition zone diametre (IZD)2 in mm | |||||
| Log concentration | 2.2788 | 1.977 | 1.6767 | 1.3766 | 1.0755 |
| Staphylococcus aureus | 182.25 | 42.25 | 12.25 | 6.25 | 0.00 |
| Escherichia coli | 462.25 | 72.25 | 36.00 | 12.25 | 6.25 |
| Klebsiella spp | 156.25 | 169.25 | 30.25 | 6.25 | 0.00 |
0 ± 0 no observed antimicrobial inhibition. n number of repetitions
Ciprofloxacin showed high inhibitory activities against the test bacteria, with highest inhibition seen against B. subtilis, followed by E. coli, Klebsiella spp and S. aureus, respectively. Of all the test organisms, gentamycin showed greatest inhibition against E. coli, B. subtilis, Klebsiella spp and S. aureus, respectively, with minimal differences. Others are as shown in Table 3.
Table 3.
Inhibition zone diametre (mm) of Ciprofloxacin and Gentamycin at different concentrations
| Test organisms | Concentrations (µg/ml) of ciprofloxacin | ||||
|---|---|---|---|---|---|
| 50 | 25 | 12.5 | 6.25 | 3.13 | |
| Staphylococcus aureus | 19 ± 0.8 | 16 ± 1.5 | 14 ± 1.0 | 8 ± 0.8 | 7 ± 0.5 |
| Escherichia coli | 20 ± 0.9 | 17 ± 1.0 | 15 ± 0.9 | 14 ± 0.5 | 11 ± 0.8 |
| Bacillus subtilis | 25 ± 1.0 | 21 ± 1.2 | 13 ± 1.0 | 8 ± 0.5 | 6 ± 0.3 |
| Klebsiella spp | 20 ± 0.8 | 14 ± 0.9 | 10 ± 0.7 | 6 ± 0.4 | 3 ± 0.2 |
| Log concentrations | Square of inhibition zone diametre (IZD)2 in mm | ||||
|---|---|---|---|---|---|
| 1.6989 | 1.39799 | 1.0969 | 0.7959 | 0.4949 | |
| Staphylococcus aureus | 361 | 256 | 196 | 64 | 49 |
| Escherichia coli | 400 | 289 | 225 | 196 | 121 |
| Bacillus subtilis | 625 | 441 | 169 | 64 | 36 |
| Klebsiella spp | 400 | 196 | 100 | 36 | 9 |
| Concentrations (µg/ml) of gentamycin (µg/ml) | |||||
|---|---|---|---|---|---|
| 40 µg/ml | 20 µg/ml | 10 µg/ml | 5 µg/ml | 2.5 µg/ml | |
| Staphylococcus aureus | 20 ± 0.6 | 14 ± 0.4 | 9 ± 0.5 | 7 ± 0.4 | 6 ± 0.5 |
| Escherichia coli | 23 ± 0.8 | 17.5 ± 0.8 | 14 ± 0.7 | 7 ± 0.5 | 6 ± 0.5 |
| Bacillus subtilis | 22 ± 1.0 | 20 ± 0.9 | 18 ± 0.9 | 15 ± 0.4 | 12 ± 0.9 |
| Klebsiella spp | 20 ± 0.8 | 16 ± 0.5 | 14 ± 0.4 | 10 ± 0.5 | 6 ± 0.3 |
| Square of inhibition zone diametre (IZD)2 of gentamycin in mm | |||||
|---|---|---|---|---|---|
| Log concentrations | 1.6021 | 1.3010 | 1.0000 | 0.6989 | 0.3979 |
| Staphylococcus aureus | 400 | 196 | 81 | 49 | 0 |
| Escherichia coli | 529 | 306.25 | 196 | 49 | 86 |
| Bacillus subtilis | 484 | 400 | 324 | 225 | 144 |
| Klebsiella spp | 121 | 81 | 64 | 36 | 0 |
The minimum inhibitory concentration (MIC) of the extract was lowest for Klebsiella spp and S. aureus, while E. coli had a higher MIC. Ciprofloxacin and gentamycin had much lower MIC than the extract for the various test bacteria organisms. See Table 4.
Table 4.
Minimum inhibitory concentrations (MIC) of the extract and the standard drugs
| Test organisms | Extract | Ciprofloxacin | Gentamycin | |||
|---|---|---|---|---|---|---|
| Log concentration | Antilog (MIC) (mg/ml) | Log concentration | Antilog (MIC) (µg/ml) | Log concentration | Antilog (MIC) (µg/ml) | |
| Staphylococcus aureus | 1.2 | 12.59 | 0.08 | 1.20 | 0.54 | 3.47 |
| Escherichia coli | 1.1 | 15.85 | −0.02 | 0.95 | 0.48 | 3.02 |
| Bacillus subtilis | 0.0 | 1.0 | 0.59 | 3.89 | −0.20 | 0.63 |
| Klebsiella spp | 1.1 | 12.59 | 0.61 | 4.07 | 0.4 | 2.51 |
Statistical analysis showed statistical significance in the minimum inhibitory concentrations of the extract and the standard drugs (p < 0.001). Therefore, antibacterial activities of the standard drugs were significantly higher than that of the extract.
Discussion
The extract had high percentage yield and exerted both antibacterial and antifungal effects. Phytochemical screening showed the presence of secondary metabolites which have been previously noted to exert antimicrobial effects, among which were; terpenoids, saponins, reducing sugars, tannins and cardenolides. In comparison with gentamicin and ciprofloxacin, the extract had much higher minimum inhibitory concentration (MIC) and the difference was statistically significant. The extract however, showed broad spectrum antibacterial activity as was also seen in the standard drugs.
High percentage yield of the extract is suggestive of the potentials of methanol as a good solvent for the extraction. Although methanol is a toxic solvent and unsuitable for direct therapeutic use, it possesses the ability to dissolve polar and semi polar plant-based substances, with several polar substances showing affinity for it, which highlights its effectiveness in extracting bioactive compounds for in vitro screening purposes. This supports findings from a previous study that demonstrated methanol extracts to be more effective against test organisms than ethanol extracts [24]. Also, aqueous extract was noted to exhibit lower antimicrobial activity than the methanol leaf extract of B. coriacea in a previous study [25]. Higher presence of secondary metabolites may be associated with the high percentage yield, resulting in higher antimicrobial activities of methanol extract of B. coriacea.
Phytochemical screening of the extract showed several secondary plant metabolites which are known for numerous activities, including antimicrobial. Among these metabolites was saponin which is previously reported to exhibit antibacterial activity by increasing the permeability of bacterial cells, which may result in an enhanced uptake of antibiotics in resistant strains of organisms [20]. However, it has also been observed that saponins, in the presence of certain antibiotics, may not inhibit the growth of some bacterial cells [21]. Although with slight differences, previous studies on the methanol seed extract also showed similar chemical components [11, 15]. Alkaloids were observed to be absent in the studied B. coriaceae leaf extract, which was similar to the findings by Osadebe et al., [25]. Flavonoids was also found to be absent in this study, meanwhile, study by Anowi et al., reported its presence [26]. Geographical variation of the plants source may be a major source of this difference in findings.
Findings from the antimicrobial studies highlighted the presence of broad spectrum antibacterial and antifungal activities associated with the extract. At varying concentrations, the extract was effective against both gram positive and gram negative bacteria and a fungus of the Saccharomycetaceae family (Candida albicans). This is similar to findings in previous studies where C. albicans and some bacteria were inhibited by the methanol seed extract of B. coriacea [15]. Also Osadebe et al., reported greater inhibition of C. albicans with the methanol leaf extract than aqueous extract [25]. However, previous studies have noted greater microbial inhibition with the fresh B. coriacea seed than the extract [14]. This may be associated with higher presence of active phytochemical constituents in the fresh cola, which may be lost to drying or methods of extraction in the extracts.
The antifungal activity of the extract was observed at very low concentration against C. albicans. Previous studies on the aqueous and methanol seed extract also showed appreciable activity against C. albicans [15]. This antifungal effect was concentration dependent as higher zones of inhibition were observed with increase in concentration of the extract. The concentration dependent effect of the extract was also observed with the test bacteria, with increase in concentration resulting in increased inhibition zones. Further, the observed difference in the level of susceptibility of the test microbes to various concentrations of the extract may be associated with the intrinsic tolerance of the microbes to the active ingredients of the extract [27]. The type, nature and quantity of phytochemical constituents of the plants may also be major contributory factor to this. The observed sensitivity of the microbes to lower extract concentrations is particularly important, underscoring the relevance of determining appropriate concentrations for effective antimicrobial activities. This is essential to prevent suboptimal concentrations that may encourage microbial resistance.
E. coli and C. albicans were the most sensitive microbes to the extract, with inhibition zones observed at the lowest concentration of the extract. These are clinically important microbes that result in various human diseases. E. coli is highly infectious to several skin lesions, surgical wounds and highly implicated in septicemia [28]. C. albicans is known to cause vaginal candidiasis, oral thrush and many other infections. The extract therefore, shows potentials for an alternative drug for the management of these infections, especially with several emerging resistant strains to the available therapeutic drugs. Meanwhile, the inhibition of Klebsialla spp surpassed the previous two mentioned organisms, at a higher concentration of the extract. At the concentrations used in this study, Bacillus subtilis was not found to be susceptible to the extract. Meanwhile, a previous study of higher extract concentrations showed S. aureus to be the most susceptible bacteria to the methanol seed extract of B. coriacea while Bacillus cereus was the least susceptible [24]. Findings imply limited susceptibility of Bacillus cereus to lower and higher concentrations of B. coriacea methanol extracts.
Although the extract showed antifungal activity and broad spectrum antibacterial activity, its effects were statistically significant at concentrations higher than ciprofloxacin and gentamicin. The observed MICs of the extract against the test bacteria was not closely related to the MIC of the standard drugs. This implies that the minimum concentrations required for antimicrobial activity of the extract was significantly higher than the minimum required concentration of ciprofloxacin and gentamycin to exert antimicrobial activity. The MIC of the extract was highest for S. aureus in the present study, but much lower than in a previous study of methanol leaf extract of B. coriacea [29]. However, higher MICs of B. coriacea leaf extract was observed by Osadebe et al., for various bacteria [25]. Different geographical locations of the plants may have contributed to different levels of phytochemical compounds and hence, the difference in antibacterial activities. The observed lower MICs for B. coriacae leaf extract in this present study highlights the presence of antimicrobial activities of the extract at low concentrations. Future studies may explore the concentration-dependent antimicrobial activities of extracts on other range of microbial agents, in addition to exploring the concentration-dependent antimicrobial activities of B. coriacae leaf extracts from other regions where the plants may be grown under different conditions.
Although the study has demonstrated the potential clinical relevance of B. coriacea methanol leaf extracts in microbial inhibition, it is not devoid of some limitations. The antifungal evaluation of the extracts was limited to only C. albicans and positive controls were not included which limits the validity of conclusions about its antifungal activity. Other factors such as season of harvest, region of cultivation and processing of the leaves may have affected the level and types of phytochemical compounds in the leaves. The potential effects of this on the evaluated antimicrobial activities of the methanol leaf extract of B. coriacea should be noted. Further, only methanol extract was tested for antimicrobial inhibition, which predisposes findings to risk of bias. The study also lacked time-kill or cherkerboard synergistic studies to ascertain the consistency of the observed antimicrobial activities or predict potential clinical outcomes. In addition, quantitative phytochemical analysis was not done in this study, rather, qualitative method was done. Although positive controls were included in the antibacterial study, negative controls were absent, posing a potential risk of bias in the observed antimicrobial activities.
Conclusion
The methanol leaf extract of B. coriacea comprised several phytochemicals at various concentrations and exhibited appreciable concentration-dependent antimicrobial activities. However, ciprofloxacin and gentamycin showed activity against the test bacteria at significantly lower concentrations than the extract. Nonetheless, the methanol leaf extract of B. coriacea having shown a relatively modest broad-spectrum antimicrobial activity, may be further evaluated for potential pharmaceutical formulations. Further investigations of the leaves' activity against wider range of bacteria and fungi, identification and purification of its chemical constituents in addition to its toxicological studies, represent areas for further research with a view to developing novel drugs for human consumption.
Supplementary Information
Acknowledgements
The authors wish to appreciate the laboratory attendants for technical supports and Mr. Ukaga, Ikechukwu for editing the manuscript.
Abbreviations
- IZD
Inhibition zone diameter
- MIC
Minimum inhibitory concentration
- SDA
Sabouraud dextrose agar
- ANOVA
Analysis of variance
Authors’ contributions
XF and COI conceived and designed the study, and performed the analysis, COI wrote initial draft of the manuscript, and XF and COI revised the manuscript for intellectual content and approved the manuscript.
Funding
Not applicable.
Data availability
Data use in this study is available from the author on reasonable request.
Declarations
Ethics approval and consent to participate
Permission was obtained from the land owner where the plant was grown, prior to harvesting.
Consent for publication
Not applicable.
Competing interests
The authors declare no competing interests.
Footnotes
Publisher’s Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
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Supplementary Materials
Data Availability Statement
Data use in this study is available from the author on reasonable request.