The Yield of the crude leaf and stem bark extracts of C. macrostachyus
The yield of crude leaf and stem bark extracts of Croton macrostachyus is presented in Table 1. According to the data in Table 1, the high yield of methanolic leaf extract of C. macrostachyus was 2.1% (1.89 gm) on dry weight bases followed by ethanolic crude leaf extract and ethyl acetate crude leaf extract with a percentage value of 2.1% (1.74 gm) and 1.2% (0.97gm) respectively. Similarly, the methanolic extract of the stem bark of the experimental plant showed a high yield percentage (4.1%), and the lowest yield was obtained from the ethyl acetate stem bark crude extract (3.1%).
Table 1
Crude extract yield of Croton macrostachyus
S.No. | Name of the Solvent | Crude extract yield of C. macrostachyus |
Yield of leaf crude extract (gm) | Yield of leaf crude extract (%) | Yield of stem bark crude extract (gm) | Yield of stem bark crude extract (%) |
1. | Ethanol | 1.74 | 2.1 | 3.19 | 3.9 |
2. | Methanol | 1.89 | 2.3 | 3.33 | 4.1 |
3. | Ethyl Acetate | 0.97 | 1.2 | 2.54 | 3.1 |
Qualitative phytochemical analysis of crude leaf and stem bark extracts of C. macrostachyus
The data presented in Table 2 shows the qualitative phytochemical analysis of the crude leaf and stem bark extracts of C. macrostachyus prepared by using three different solvent systems. The presented results in Table 2 reveal that the methanolic and ethanolic crude leaf extract was rich in various kinds of phytochemicals such as tannins, alkaloids, saponins, flavonoids, glycosides, and phenolics. The ethyl acetate crude leaf extract contains most of the phytoconstituents except alkaloids. The phytochemical investigation of stem bark crude extracts showed that the phytochemicals such as alkaloids and flavonoids were absent. The phytochemicals such as tannins, saponins, and phenolics were positive in ethanolic and methanolic crude stem bark extracts. Most of the phytochemicals were absent in ethyl acetate crude stem bark extract (Table 2).
Table 2
Qualitative phytochemical analysis of crude leaf and stem bark extracts of C. macrostachyus
S. No. | Phytochemicals | Leaf crude extract | Stem bark extract |
Ethanol | Methanol | Ethyl acetate | Ethanol | Methanol | Ethyl acetate |
1 | Tannins | + | + | + | + | + | - |
2 | Alkaloids | + | + | - | - | - | - |
3 | Saponins | + | + | + | + | + | - |
4 | Flavonoids | + | + | + | - | - | - |
5 | Glycosides | + | + | + | + | + | + |
6 | Phenolic | + | + | + | + | + | - |
The antibacterial activity of crude leaf extracts of C. macrostachyus against some selected pathogenic bacteria
The crude leaf extracts of the experimental plant C. macrostachyus exhibited diverse inhibition activity against the selected ATCC bacterial pathogens (Table 3, 4, 5); and the results were expressed in the form of a millimeter (mm) in diameter of the zone of bacterial growth inhibition. The data presented in Tables 3, 4, and 5 clearly showed that the selected bacterial pathogens such as Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus were susceptible to the ethanolic, methanolic, and ethyl acetate crude leaf extracts. However, there were significant differences (p < 0.05) in the mean diameter zone of inhibition between the extractions as well as between the bacterial pathogens.
In terms of ethanolic crude leaf extract, 100mg/ml concentration produced a higher zone of inhibitions against all the three selected bacterial pathogens such as Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus with 18.17 ± 0.65 mm, 20.07 ± 0.22 mm and 22.00 ± 1.33 mm respectively. The zone of inhibition produced by the ethanolic crude leaf extracts at 100 mg/ml was nearly similar to the zone developed by the positive control (tetracycline) against P. aeruginosa and S. aureus (Table 3).
Table 3
The antibacterial activities of ethanolic crude leaf extracts of C. macrostachyus against some selected pathogenic bacteria
Name of pathogenic Bacteria | Ethanolic crude leaf extracts of C. macrostachyus in different concentrations | Positive Control (Tetracycline 30 µg/disc) | Negative Control (DMSO) |
25 mg/mL | 50 mg/mL | 100 mg/mL |
Mean Zone of Inhibition (ZOI) in mm |
E. coli | 10.23 ± 0.38b | 13.92 ± 0.71b | 18.17 ± 0.65c | 26.85 ± 0.21a | 0 |
P. aeruginosa | 12.05 ± 0.32b | 14.13 ± 0.23b | 20.07 ± 0.22b | 19.50 ± 0.5b | 0 |
S. aureus | 15.25 ± 0.35a | 17.55 ± 0.32a | 22.00 ± 1.33a | 25.86 ± 1.54c | 0 |
Note: All results shown are the mean triplicates ± Standard deviation values. Means bearing a different letter of superscripts in a column differ significantly, but the same letters in superscripts indicate an insignificant difference at P < 0.05. |
In terms of methanolic crude leaf extracts, the lowest concentration 25mg/ml produced less zone of inhibition against selected bacterial pathogens, while 50mg/ml and 100mg/ml concentrations produced higher inhibition zones. The pathogenic bacterium S. aureus showed significantly more susceptibility (25.47 ± 0.90 mm) than the pathogenic bacteria E. coli (21.8 ± 0.83 mm) and P. aeruginosa (19.61 ± 0.33 mm). The methanolic crude leaf extracts at 100 mg/ml concentration produced a significantly similar zone of inhibition developed by the positive control (tetracycline) against S. aureus (Table 4). The ethyl acetate crude leaf extract of C. macrostachyus expressed a moderate level of antibacterial activity even at higher concentrations of the crude extracts against all three selected pathogenic bacteria (Table 5). The methanolic and ethanolic crude leaf extracts showed higher activity than ethyl acetate extraction against the selected bacterial pathogens.
Table 4
The antibacterial activities of methanolic crude leaf extracts of C. macrostachyus against some selected pathogenic bacteria
Name of pathogenic Bacteria | Methanolic crude leaf extracts of C. macrostachyus in different concentrations | Positive Control (Tetracycline 30 µg/disc) | Negative Control (DMSO) |
25 mg/mL | 50 mg/mL | 100 mg/mL |
Mean Zone of Inhibition (ZOI) in mm |
E. coli | 14.15 ± 0.17b | 18.17 ± 0.16a | 19.61 ± 0.33c | 26.85 ± 0.21a | 0 |
P. aeruginosa | 14.27 ± 0.59b | 14.93 ± 0.45b | 21.8 ± 0.83b | 19.50 ± 0.5b | 0 |
S. aureus | 12.59 ± 0.44a | 15.81 ± 0.48b | 25.47 ± 0.90a | 25.86 ± 1.54c | 0 |
Note: All results shown are the mean triplicates ± Standard deviation values. Means bearing a different letter of superscripts in a column differ significantly, but the same letters in superscripts indicate an insignificant difference at P < 0.05. |
Table 5
The antibacterial activities of ethyl acetate crude leaf extracts of C. macrostachyus against some selected pathogenic bacteria
Name of pathogenic Bacteria | Ethyl acetate crude leaf extracts of C. macrostachyus in different concentrations | Positive Control (Tetracycline 30µg/disc) | Negative Control (DMSO) |
25mg/mL | 50mg/mL | 100mg/mL |
Mean Zone of Inhibition (ZOI) in mm |
E. coli | 10.76 ± 0.91c | 12.67 ± 0.56b | 13.59 ± 0.72b | 26.85 ± 0.21a | 0 |
P. aeruginosa | 6.42 ± 1.2b | 11.45 ± 0.84b | 11.48 ± 0.95a | 19.50 ± 0.5b | 0 |
S. aureus | 12.74 ± 0.22a | 13.62 ± 0.76a | 14.14 ± 0.44b | 25.86 ± 1.54c | 0 |
Note: All results shown are the mean triplicates ± Standard deviation values. Means bearing a different letter of superscripts in a column differ significantly, but the same letters in superscripts indicate an insignificant difference at P < 0.05. |
The antibacterial activity of crude stem bark extracts of C. macrostachyus
The data received from the antibacterial activity of ethanolic, methanolic, and ethyl acetate crude stem bark extracts are presented in Tables 6, 7, and 8. The prepared crude stem bark extracts using different solvent systems showed antibacterial activity against the selected bacterial pathogens.
Table 6
The antibacterial activities of ethanolic crude stem bark extracts of C. macrostachyus against some selected pathogenic bacteria
Name of pathogenic Bacteria | Ethanolic crude stem bark extracts of C. macrostachyus in different concentrations | Positive Control (Tetracycline 30µg/disc) | Negative Control (DMSO) |
25mg/mL | 50mg/mL | 100mg/mL |
Mean Zone of Inhibition (ZOI) in mm |
E. coli | 9.73 ± 0.36b | 11.50 ± 0.50b | 13.35 ± 0.61b | 26.85 ± 0.21a | 0 |
P. aeruginosa | 9.01 ± 0.24b | 12.12 ± 0.27b | 13.92 ± 0.28b | 19.50 ± 0.5b | 0 |
S. aureus | 11.69 ± 0.53a | 13.20 ± 0.57a | 15.15 ± 0.47b | 25.86 ± 1.54c | 0 |
Note: All results shown are the mean triplicates ± Standard deviation values. Means bearing a different letter of superscripts in a column differ significantly, but the same letters in superscripts indicate an insignificant difference at P < 0.05. |
There were no significant differences in the mean zone of inhibition between ethanolic and methanolic stem bark extracts, while the ethyl acetate crude stem bark extracts showed a significant decrease in the mean diameter inhibition zone. However, the methanolic crude stem bark extracts of C. macrostachyus at 100mg/ml concentration showed the highest activity against S. aureus (15.45 ± 0.45 mm), which was lower than the positive control (Tetracycline, 25.86 ± 1.54 mm) (Table 7). The least zone of inhibition was found to be with E. coli (6.49 ± 0.48 mm) at 100 mg/ml concentration of Ethyl acetate crude stem bark extracts (Table 8).
Table 7
The antibacterial activities of methanolic crude stem bark extracts of C. macrostachyus against some selected pathogenic bacteria
Name of pathogenic Bacteria | Methanolic crude stem bark extracts of C. macrostachyus in different concentrations | Positive Control (Tetracycline 30µg/disc) | Negative Control (DMSO) |
25mg/mL | 50mg/mL | 100mg/mL |
Mean Zone of Inhibition (ZOI) in mm |
E. coli | 10.66 ± 0.54b | 12.40 ± 0.47b | 14.01 ± 0.81b | 26.85 ± 0.21a | 0 |
P. aeruginosa | 9.52 ± 1.80b | 11.14 ± 0.68b | 13.64 ± 0.43b | 19.50 ± 0.5b | 0 |
S. aureus | 11.61 ± 0.47a | 13.95 ± 0.38a | 15.45 ± 0.45b | 25.86 ± 1.54c | 0 |
Note: All results shown are the mean triplicates ± Standard deviation values. Means bearing a different letter of superscripts in a column differ significantly, but the same letters in superscripts indicate an insignificant difference at P < 0.05. |
Table 8
The antibacterial activities of ethyl acetate crude stem bark extracts of C. macrostachyus against some selected pathogenic bacteria
Name of pathogenic Bacteria | Ethyl acetate crude stem bark extracts of C. macrostachyus in different concentrations | Positive Control (Tetracycline 30µg/disc) | Negative Control (DMSO) |
25mg/mL | 50mg/mL | 100mg/mL |
Mean Zone of Inhibition (ZOI) in mm |
E. coli | 4.72 ± 0.99b | 4.9 ± 0.60b | 6.49 ± 0.48b | 26.85 ± 0.21a | 0 |
P. aeruginosa | 3.69 ± 0.38b | 4.85 ± 0.47b | 5.68 ± 0.73b | 19.50 ± 0.5b | 0 |
S. aureus | 6.26 ± 0.62a | 8.20 ± 0.74a | 10.48 ± 0.77a | 25.86 ± 1.54c | 0 |
Note: All results shown are the mean triplicates ± Standard deviation values. Means bearing a different letter of superscripts in a column differ significantly, but the same letters in superscripts indicate an insignificant difference at P < 0.05. |
Minimum Inhibitory concentration (MIC) and Minimum bactericidal concentration (MBC) of crude leaf extracts of C. macrostachyus
The MIC results of the crude leaf and stem bark extracts of C. macrostachyus presented in Table 9 showed that all the selected bacterial pathogens were very susceptible to the minimum inhibitory concentrations ranging from 0.625–2.5 mg/ml. The minimum inhibitory concentration of ethanolic crude leaf extracts of C. macrostachyus was 0.625mg/ml for S. aureus, and 1.25 mg/ml for E. coli and P. aeruginosa. In terms of methanolic crude leaf extracts, the minimum inhibitory concentration was 0.625 mg/ml for S. aureus, E. coli and P. aeruginosa. The minimum inhibitory concentration of ethyl acetate crude leaf extracts was 1.25 mg/ml for S. aureus, E. coli, and P. aeruginosa. The minimum inhibitory concentration of crude stem bark extracts ranged from 1.25–2.5 mg/ml. In terms of ethanolic and methanolic crude stem bark extracts of C. macrostachyus was 1.25 mg/ml for S. aureus, E. coli and P. aeruginosa. The minimum inhibitory concentration of ethyl acetate crude stem bark extract was 1.25 mg/ml for all three selected bacterial pathogens (Table 9).
Table 9
Minimum Inhibitory concentration of crude extracts of C. macrostachyus
S. No. | Nature of the crude extracts | Name of the bacterial pathogens |
E. coli | P. aeruginosa | S. aureus |
mg/mL |
1. | Ethanolic leaf | 1.25 | 1.25 | 0.625 |
2. | Methanolic leaf | 0.625 | 0.625 | 0.625 |
3. | Ethyl acetate leaf | 1.25 | 1.25 | 1.25 |
4. | Ethanolic stem bark | 1.25 | 1.25 | 1.25 |
5. | Methanolic stem bark | 1.25 | 1.25 | 1.25 |
6. | Ethyl acetate stem bark | 2.5 | 2.5 | 2.5 |
The data obtained through the determination of MBC of crude leaf and stem bark extracts are presented in Table 10. The ethanolic crude leaf extract showed a bactericidal effect at 1.25 mg/ml for S. aureus and 2.5 mg/ml for both E. coli and P. aeruginosa. In terms of methanolic leaf crude extract showed bactericidal effect at 1.25 mg/ml for all the three selected pathogens. The MBC of ethyl acetate crude leaf extracts was 2.5 mg/ml for both E. coli and P. aeruginosa, and 1.25 mg/ml for S. aureus. The MBC of ethanolic, methanolic and ethyl acetate crude stem bark extracts ranged from 2.5 to 5 mg/ml for gram-negative bacterial pathogens. In terms of Gram-positive bacteria S. aureus, the ethanolic and methanolic stem bark crude extracts at 1.25 mg/ml concentration showed a bactericidal effect, and ethyl acetate stem bark crude extract at 5 mg/ml concentration showed a bactericidal effect (Table 10).
Table 10
Minimum Bactericidal concentration of crude extracts of C. macrostachyus
S. No. | Nature of the crude extracts | Name of the bacterial pathogens |
E. coli | P. aeruginosa | S. aureus |
mg/mL |
1. | Ethanolic leaf | 2.5 | 2.5 | 1.25 |
2. | Methanolic leaf | 1.25 | 1.25 | 1.25 |
3. | Ethyl acetate leaf | 2.5 | 2.5 | 1.25 |
4. | Ethanolic stem bark | 2.5 | 2.5 | 1.25 |
5. | Methanolic stem bark | 2.5 | 2.5 | 1.25 |
6. | Ethyl acetate stem bark | 5 | 5 | 5 |