Background
Methods
Plant collection, extraction and fractionation
Extraction of crude saponins
Gas chromatography–mass spectrometry (GC/MS) analysis
Chemical and drugs
Collection and identification of bacteria
Standardization of bacterial suspension
Antibacterial investigations
Bacterial susceptibility pattern
Disc diffusion assay
Determination of Minimum Inhibitory Concentrations (MICs)
Antifungal investigations
Results
Antibacterial activity
Bacterial susceptibility pattern
Disc diffusion assay
Samples | Conc. μg/ml | Diameter of the inhibitory zone (mm) Mean ± SEM (n = 3) | ||||||
---|---|---|---|---|---|---|---|---|
Enterococcus faecalis
|
Klebsiella pneumonia
|
Escherichia coli
|
Proteus mirabilis
|
Staphylococcus aureus
|
Salmonella typhi
|
Pseudomonas aeruginosa
| ||
Crude (Ph.Cr) | 25 | 5.33 ± 0.88 | 6.33 ± 0.66 | 0.00 ± 0.00 | 11.00 ± 0.00 | 10.00 ± 0.57 | 11.33 ± 1.20 | 15.66 ± 1.85 |
50 | 14.00 ± 0.57 | 10.33 ± 0.33 | 0.00 ± 0.00 | 17.33 ± 0.88 | 14.33 ± 0.33 | 14.00 ± 0.57 | 20.33 ± 0.88 | |
100 | 17.33 ± 0.88 | 14.00 ± 0.00 | 9.00 ± 0.57 | 34.00 ± 1.15 | 18.66 ± 0.66 | 21.33 ± 0.88 | 25.33 ± 1.45 | |
Ethyl acetate (Ph.EtAc | 25 | 9.33 ± 0.33 | 0.00 ± 0.00 | 0.00 ± 0.00 | 11.66 ± 1.20 | 8.33 ± 0.88 | 8.00 ± 0.57 | 13.00 ± 0.57 |
50 | 18.33 ± 0.88 | 7.00 ± 0.57 | 8.33 ± 0.88 | 18.33 ± 0.66 | 13.66 ± 1.20 | 15.66 ± 0.66 | 18.33 ± 0.66 | |
100 | 27.00 ± 0.57 | 13.33 ± 0.88 | 15.00 ± 0.00 | 27.66 ± 2.18 | 18.33 ± 0.88 | 20.73 ± 1.00 | 24.00 ± 0.57 | |
Chloroform (Ph.Chf) | 25 | 13.66 ± 0.33 | 0.00 ± 0.00 | 8.00 ± 0.57 | 12.66 ± 3.40 | 7.33 ± 0.88 | 17.33 ± 0.66 | 18.66 ± 1.20 |
50 | 19.00 ± 0.57 | 8.00 ± 0.57 | 14.33 ± 0.33 | 15.33 ± 2.02 | 11.66 ± 1.45 | 23.00 ± 0.57 | 23.00 ± 1.00 | |
100 | 31.66 ± 1.20 | 15.60 ± 1.15 | 18.00 ± 1.52 | 21.00 ± 1.73 | 16.00 ± 1.15 | 29.33 ± 0.88 | 31.66 ± 2.18 | |
Butanol (Ph.Bt) | 25 | 7.00 ± 1.15 | 0.00 ± 0.00 | 0.00 ± 0.00 | 14.00 ± 0.57 | 13.66 ± 1.20 | 11.33 ± 0.66 | 7.33 ± 0.88 |
50 | 11.00 ± 1.15 | 0.00 ± 0.00 | 0.00 ± 0.00 | 19.66 ± 1.20 | 18.66 ± 1.45 | 14.00 ± 1.52 | 12.66 ± 0.66 | |
100 | 21.00 ± 0.00 | 8.00 ± 0.00 | 11.33 ± 0.88 | 29.00 ± 0.00 | 23.00 ± 1.15 | 19.00 ± 0.57 | 16.00 ± 1.52 | |
n-Hexane (Ph.Hex) | 25 | 11.66 ± 0.66 | 0.00 ± 0.00 | 0.00 ± 0.00 | 0.00 ± 0.00 | 11.00 ± 1.15 | 8.66 ± 1.66 | 12.66 ± 0.33 |
50 | 17.66 ± 1.66 | 9.66 ± 0.33 | 7.66 ± 1.20 | 9.33 ± 0.66 | 17.33 ± 2.18 | 14.00 ± 1.00 | 17.00 ± 0.57 | |
100 | 30.00 ± 0.57 | 15.00 ± 0.57 | 12.00 ± 0.57 | 14.00 ± 0.57 | 26.33 ± 0.88 | 20.33 ± 0.88 | 21.00 ± 0.00 | |
Saponins (Ph.Sp) | 25 | 6.60 ± 1.15 | 0.00 ± 0.00 | 11.00 ± 1.00 | 0.00 ± 0.00 | 10.66 ± 2.02 | 22.00 ± 0.57 | 15.00 ± 0.00 |
50 | 10.33 ± 1.45 | 00.00 ± 0.00 | 16.00 ± 0.57 | 0.00 ± 0.00 | 15.00 ± 0.57 | 26.33 ± 0.33 | 22. 33 ± 1.20 | |
100 | 26.66 ± 1.20 | 7.33 ± 0.88 | 23.00 ± 0.00 | 8.66 ± 1.20 | 22.00 ± 1.52 | 30.66 ± 1.20 | 27.00 ± 0.57 | |
Aqueous (Ph.Aq) | 25 | 4.00 ± 0.00 | 0.00 ± 0.00 | 6.00 ± 1.15 | 0.00 ± 0.00 | 0.00 ± 0.00 | 0.00 ± 0.00 | 6.33 ± 0.66 |
50 | 13.00 ± 0.57 | 0.00 ± 0.00 | 10.33 ± 0.33 | 0.00 ± 0.00 | 7.00 ± 0.57 | 10.33 ± 0.66 | 10.88 ± 0.33 | |
100 | 20.33 ± 0.33 | 10.33 ± 1.45 | 26.66 ± 1.20 | 11.33 ± 1.45 | 14.33 ± 2.02 | 15.60 ± 1.15 | 13.00 ± 0.66 | |
Positive Control | 25 | 19.33 ± 2.02 | 24.00 ± 0.57 | 23.33 ± 1.45 | 22.57 ± 0.57 | 16.66 v 0.88 | 20.33 ± 0.88 | 22.33 ± 0.66 |
50 | 25.00 ± 0.57 | 29.33 ± 0.66 | 29.66 ± 1.20 | 27.66 ± 0.88 | 23.33 ± 1.52 | 27.00 ± 0.57 | 29.00 ± 0.57 | |
100 | 31.66 ± 1.45 | 35.33 ± 0.88 | 34.66 ± 0.88 | 31.00 ± 1.52 | 27.00 ± 1.15 | 34.66 ± 1.20 | 36.66 ± 1.20 |
Minimum Inhibitory Concentrations (MICs)
Bacterial strains | Minimum inhibitory concentrations (MICs) in μg/ml. | |||||||
---|---|---|---|---|---|---|---|---|
Crude (Ph.Cr) | n-Hexane (Ph.Hex) | Ethyl acetate (Ph.EtAc) | Butanol (Ph.Bt) | Chloroform (Ph.Chf) | Aqueous (Ph.Aq) | Saponins (Ph.Sp) | Ceftriaxone | |
Enterococcus faecalis
| 128.00 ± 0.00 | 64.00 ± 0.00 | 26.66 ± 5.33 | 21.33 ± 5.33 | 32.00 ± 0.00 | >512 | 10.66 ± 2.66 | 8.00 ± 0.00 |
Klebsiella pneumoniae
| 53.33 ± 10.66 | 42.66 ± 10.66 | 13.33 ± 2.66 | 26.66 ± 5.33 | 13.33 ± 2.66 | 256.00 ± 0.00 | 32.00 ± 0.00 | 4.00 ± 0.00 |
Escherichia coli
| 64.00 ± 0.00 | 128.00 ± 0.00 | 64.00 ± 0.00 | 128.00 ± 0.00 | 10.66 ± 2.66 | > 512 | 26.66 ± 5.33 | 4.00 ± 0.00 |
Proteus mirabilis
| 21.33 ± 5.33 | 256.00 ± 0.00 | 32.00 ± 0.00 | 128.00 ± 0.00 | 5.33 ± 1.33 | 128.00 ± 0.00 | 6.66 ± 1.33 | 8.00 ± 0.00 |
Staphylococcus aureus
| 512.00 ± 0.00 | 128.00 ± 0.00 | 512.00 ± 0.00 | >512 | 64.00 ± 0.00 | 256.00 ± 0.00 | 128.00 ± 0.00 | 16.00 ± 0.00 |
Salmonella typhi
| 64.00 ± 0.00 | 128.00 ± 0.00 | 64.00 ± 0.00 | 13.33 ± 2.66 | 8.66 ± 0.66 | 128.00 ± 0.00 | 53.33 ± 10.66 | 16.00 ± 0.00 |
Pseudomonas aeruginosa
| 26.66 ± 5.33 | 32.00 ± 0.00 | 64.00 ± 0.00 | 32.00 ± 0.00 | 10.66 ± 2.66 | 512.00 ± 0.00 | 6.66 ± 1.33 | 4.00 ± 0.00 |
Antifungal activity
Fungal strains | Inhibition of fungal growth | |||||||
---|---|---|---|---|---|---|---|---|
Crude (Ph.Cr) | n-Hexane (Ph.Hex) | Ethyl acetate (Ph.EtAc) | Butanol (Ph.Bt) | Chloroform (Ph.Chf) | Aqueous (Ph.Aq) | Saponins (Ph.Sp) | Positive Control | |
Aspergillus fumigatus
| +++ | ++ | +++ | +++ | +++ | ++ | +++ | +++ |
Aspergillus flavus
| +++ | ++ | ++ | ++ | +++ | + | +++ | +++ |
Aspergillus niger
| ++ | ++ | +++ | ++ | +++ | - | ++ | +++ |
Fusarium oxysporum
| ++ | +++ | +++ | ++ | +++ | - | +++ | +++ |
Samples/Fractions | Fungal strains (DIZ in mm n = 3 SEM) | ||||
---|---|---|---|---|---|
Conc. μg/ml |
Aspergillus fumigatus
|
Aspergillus flavus
|
Aspergillus niger
|
Fusarium oxysporum
| |
Crude (Ph.Cr) | 125 | 6.33 ± 0.33 | 9.00 ± 1.15 | 5.00 ± 0.00 | 9.66 ± 1.76 |
250 | 10.00 ± 0.57 | 16.00 ± 0.57 | 8.50 ± 1.00 | 13.33 ± 0.88 | |
500 | 16.00 ± 0.00 | 20.33 ± 0.88 | 12.00 ± 0.57 | 18.00 ± 0.00 | |
n-Hexane (Ph.Hex) | 125 | 6.66 ± 0.66 | 8.00 ± 0.57 | 8.00 ± 1.15 | 4.00 ± 0.50 |
250 | 9.33 ± 0.88 | 11.00 ± 1.15 | 11.33 ± 0.88 | 7.30 ± 1.50 | |
500 | 11.00 ± 1.15 | 16.33 ± 0.33 | 16.00 ± 1.00 | 10.00 ± 1.00 | |
Ethyl acetate (Ph.EtAc) | 125 | 14.00 ± 0.57 | 7.00 ± 1.15 | 13.66 ± 0.66 | 6.33 ± 1.45 |
250 | 16.00 ± 0.33 | 9.33 ± 0.88 | 17.33 ± 2.02 | 9.00 ± 1.15 | |
500 | 21.33 ± 0.88 | 17.00 ± 0.00 | 19.66 ± 1.76 | 15.00 ± 0.00 | |
Butanol (Ph.Bt) | 125 | 13.00 ± 0.50 | 9.00 ± 0.50 | 15.00 ± 0.00 | 12.33 ± 0.66 |
250 | 18.00 ± 1.70 | 14.00 ± 1.73 | 21.66 ± 1.45 | 18. 00 ± 0.50 | |
500 | 22.33 ± 1.45 | 17.33 ± 1.45 | 26.00 ± 2.30 | 26.00 ± 1.73 | |
Chloroform (Ph.Chf) | 125 | 13.00 ± 1.15 | 13.00 ± 1.15 | 14.33 ± 0.88 | 9.33 ± 2.02 |
250 | 20.33 ± 0.88 | 17.00 ± 2.30 | 19.00 ± 1.45 | 13.00 ± 0.57 | |
500 | 26.00 ± 1.73 | 22.66 ± 1.20 | 23.00 ± 1.15 | 16.00 ± 2.30 | |
Aqueous (Ph.Aq) | 125 | 10.33 ± 0.88 | 15.00 ± 1.15 | 12.00 ± 00 | 12.30 ± 0.33 |
250 | 13.00 ± 1.15 | 18.00 ± 1.73 | 17.00 ± 1.45 | 17.00 ± 1.73 | |
500 | 17.00 ± 2.30 | 24.33 ± 1.45 | 21.00 ± 0.57 | 28.66 ± 0.88 | |
Saponins (Ph.Sp) | 125 | 8.33 ± 0.88 | 11.00 ± 0.00 | 7.00 ± 0.00 | 9.00 ± 2.30 |
250 | 11.66 ± 0.66 | 16.00 ± 0.50 | 12.33 ± 1.20 | 12.66 ± 1.20 | |
500 | 16.00 ± 0.57 | 18.33 ± 1.15 | 15.00 ± 1.15 | 15.00 ± 0.00 | |
Positive Control | 125 | 19.33 ± 0.88 | 23.66 ± 1.76 | 17.00 ± 2.88 | 22.66 ± 1.20 |
250 | 26.00 ± 1.15 | 30.66 ± 2.33 | 22.00 ± 2.00 | 28.66 ± 2.60 | |
500 | 33.00 ± 0.57 | 36.00 ± 1.73 | 29.00 ± 1.15 | 35.33 ± 2.02 | |
N. Control | ---- | ----- | ---- | ---- | ---- |
Fungal strains | Minimum Fungicidal concentrations (MFCs) in μg/ml | |||||||
---|---|---|---|---|---|---|---|---|
Crude (Ph.Cr) | n-Hexane (Ph.Hex) | Ethyl acetate (Ph.EtAc) | Butanol (Ph.Bt) | Chloroform (Ph.Chf) | Aqueous (Ph.Aq) | Saponins (Ph.Sp) | Positive Control | |
Aspergillus fumigatus
| 73.33 ± 6.66 | 116.66 ± 8.33 | 16.66 ± 3.33 | 33.33 ± 6.66 | 16.66 ± 3.33 | 500.00 ± 0.00 | 20.00 ± 0.00 | 8.33 ± 1.66 |
Aspergillus flavus
| 250.00 ± 0.00 | 73.33 ± 6.66 | 60.00 ± 0.00 | 125.00 ± 0.00 | 23.33 ± 8.81 | > 1000 | 53.33 ± 6.66 | 6.66 ± 1.66 |
Aspergillus niger
| 100.00 ± 0.00 | > 1000 | 40.00 ± 0.00 | 93.33 ± 6.66 | 125.00 ± 0.0 | 500.00 ± 0.00 | 33.33 ± 6.66 | 16.66 ± 3.33 |
Fusarium oxysporum
| 86.66 ± 6.66 | 500.00 ± 0.00 | 93.33 ± 6.66 | 250.00 ± 0.00 | 46.66 ± 6.66 | >1000 | 40.00 ± 0.00 | 20.00 ± 0.00 |
Samples/Fractions | Total Insects | Conc. (μg/ml) | Tribolium castraneum killed | Percent mortality | LC50 μg/ml | Rhyzopertha dominica Killed | Percent mortality | LC50 μg/ml |
---|---|---|---|---|---|---|---|---|
Crude (Ph.Cr) | 30 | 125 | 12.00 ± 0.57 | 40.00*** | 255 | 19.00 ± 0.00 | 63.33** | 25 |
30 | 250 | 14.33 ± 0.66 | 47.76*** | 21.00 ± 1.15 | 70.00** | |||
30 | 500 | 18.33 ± 0.33 | 61.10** | 22. 67 ± 0.88 | 75.56** | |||
n-Hexane (Ph.Hex) | 30 | 125 | 12.66 ± 0.33 | 42.20*** | 360 | 13.00 ± 0.57 | 43.33*** | 190 |
30 | 250 | 15.66 ± 0.66 | 62.20** | 16.66 ± 0.33 | 55.53*** | |||
30 | 500 | 18.67 ± 0.33 | 62.23** | 19.00 ± 0.58 | 63.33** | |||
Ethyl acetate (Ph.EtAc) | 30 | 125 | 17.66 ± 0.66 | 58.86*** | 80 | 15.33 ± 0.88 | 51.10*** | 110 |
30 | 250 | 20.00 ± 0.57 | 66.66** | 23.00 ± 1.15 | 76.66** | |||
30 | 500 | 24.00 ± 0.57 | 80.00* | 26.33 ± 0.33 | 87.76* | |||
Butanol (Ph.Bt) | 30 | 125 | 21.33 ± 0.33 | 71.10** | 20 | 19.66 ± 1.88 | 65.53** | 57 |
30 | 250 | 24.00 ± 0.57 | 80.00* | 25.00 ± 1.15 | 83.33* | |||
30 | 500 | 26.67 ± 0.89 | 88.90* | 28.00 ± 0.00 | 93.33 | |||
Chloroform (Ph.Chf) | 30 | 125 | 4.66 ± 0.88 | 15.53*** | >500 | 5.00 ± 0.57 | 16.55*** | >500 |
30 | 250 | 6.33 ± 0.33 | 21.10*** | 9.00 ± 1.15 | 30.00*** | |||
30 | 500 | 8.67 ± 0.89 | 28.90*** | 11. 67 ± 0.88 | 38.90*** | |||
Aqueous (Ph.Aq) | 30 | 125 | 14.00 ± 0.00 | 46.66*** | 130 | 9.66 ± 0.88 | 32.20*** | 300 |
30 | 250 | 21.00 ± 1.15 | 70.00** | 14.00 ± 0.57 | 46.66*** | |||
30 | 500 | 23.33 ± 1.21 | 77.77** | 18.33 ± 0.33 | 61.10** | |||
(Saponins) Ph.Sp | 30 | 125 | 15.33 ± 0.88 | 51.10*** | 110 | 21.33 ± 1.20 | 71.10** | 28 |
30 | 250 | 22.00 ± 0.00 | 73.33** | 24.00 ± 0.00 | 80.00* | |||
30 | 500 | 25.00 ± 0.00 | 83.33* | 27.00 ± 0.00 | 90.00 | |||
Positive Control | 30 | 125 | 24.00 ± 0.00 | 80.00 | 15 | 24.66 ± 0.33 | 82.20 | 9 |
30 | 250 | 27.66 ± 0.88 | 92.20 | 27.33 ± 0.88 | 91.10 | |||
30 | 500 | 30.00 ± 0.00 | 100.00 | 30.00 ± 0.00 | 100.00 | |||
Negative Control | 30 | --- | 0 ± 0.00 | 0 ± 0.00 | --- | 0 ± 0.00 | 0 ± 0.00 |
Samples | Dose (mg/ml) | Total treated | No. Repeated | Average lethality | %lethality mean ± SEM | LC50 (mg/ml) |
---|---|---|---|---|---|---|
Crude (Ph.Cr) | 12.5 | 25 | 3 | 17.66 ± 0.66 | 70.64 | 0.93 |
25 | 25 | 18.66 ± 0.33 | 74.64 | |||
50 | 25 | 20.00 ± 0.57 | 80.00 | |||
n-Hexane (Ph.Hex) | 12.5 | 25 | 3 | 8.66 ± 0.88 | 34.64 | 27.32 |
25 | 25 | 12.33 ± 0.88 | 49.32 | |||
50 | 25 | 14.66 ± 1.20 | 58.64 | |||
Chloroform (Ph.Chf) | 12.5 | 25 | 3 | 19.33 ± 0.88 | 77.32 | 1.16 |
25 | 25 | 22.33 ± 1.20 | 89.32 | |||
50 | 25 | 23.33 ± 0.66 | 93.32 | |||
Ethyl acetate (Ph.EtAc) | 12.5 | 25 | 3 | 15.66 ± 1.33 | 62.64 | 6.35 |
25 | 25 | 17.66 ± 2.33 | 70.64 | |||
50 | 25 | 21.00 ± 1.15 | 84.00 | |||
Aqueous (Ph.Aq) | 12.5 | 25 | 3 | 9.66 ± 1.20 | 38.64 | 53.24 |
25 | 25 | 11.00 ± 1.15 | 44.00 | |||
50 | 25 | 12.33 ± 0.88 | 49.32 | |||
Butanol (Ph.Bt) | 12.5 | 25 | 3 | 16.66 ± 0.33 | 66.64 | 2.28 |
25 | 25 | 17.33 ± 0.88 | 69.32 | |||
50 | 25 | 19.66 ± 2.33 | 78.64 | |||
(Saponins) Ph.Sp | 12.5 | 25 | 3 | 23.66 ± 0.33 | 94.64 | <0.01 |
25 | 25 | 24.00 ± 0.00 | 96.00 | |||
50 | 25 | 25.00 ± 0.00 | 100.00 | |||
Positive Control | 12.5 | 25 | 3 | 25.00 ± 0.00 | 100.0 | <0.01 |
25 | 25 | 25.00 ± 0.00 | 100.0 | |||
50 | 25 | 25.00 ± 0.00 | 100.0 | |||
Negative Control | --- | 25 | 3 | 0.00 | 0.00 | 0.00 |
Samples | Dose (mg/ml) | Total treated | No. Repeated | Average lethality | Percent lethality | LC50 (mg/ml) |
---|---|---|---|---|---|---|
Crude (Ph.Cr) | 12.5 | 20 | 3 | 8.66 ± 0.66 | 43.30 | 33.54 |
25 | 20 | 9.00 ± 1.15 | 45.00 | |||
50 | 20 | 12.66 ± 2.33 | 63.30 | |||
n-Hexane (Ph.Hex) | 12.5 | 20 | 3 | 5.00 ± 0.57 | 25.00 | 54.82 |
25 | 20 | 7.66 ± 0.88 | 38.30 | |||
50 | 20 | 9.33 ± 0.88 | 46.65 | |||
Chloroform (Ph.Chf) | 12.5 | 20 | 3 | 16.66 ± 1.33 | 83.30 | <0.01 |
25 | 20 | 17.33 ± 0.66 | 86.65 | |||
50 | 20 | 19.33 ± 0.88 | 96.65 | |||
Ethyl acetate (Ph.EtAc) | 12.5 | 20 | 3 | 12.00 ± 0.00 | 60.00 | 5.91 |
25 | 20 | 14.00 ± 1.15 | 70.00 | |||
50 | 20 | 15.66 ± 0.33 | 78.30 | |||
Aqueous (Ph.Aq) | 12.5 | 20 | 3 | 9.66 ± 1.20 | 48.30 | 17.17 |
25 | 20 | 10.66 ± 1.33 | 53.30 | |||
50 | 20 | 12.33 ± 0.88 | 61.65 | |||
Butanol (Ph.Bt) | 12.5 | 20 | 3 | 12.00 ± 1.15 | 60.00 | 6.13 |
25 | 20 | 13.66 ± 0.88 | 68.30 | |||
50 | 20 | 15.66 ± 0.66 | 78.30 | |||
(Saponins) Ph.Sp | 12.5 | 20 | 3 | 18.66 ± 0.33 | 93.30 | <0.01 |
25 | 20 | 20.00 ± 0.00 | 100.00 | |||
50 | 20 | 20.00 ± 0.00 | 100.00 | |||
Positive Control | 12.5 | 25 | 3 | 25.00 ± 0.00 | 100.0 | <0.01 |
25 | 25 | 25.00 ± 0.00 | 100.0 | |||
50 | 25 | 25.00 ± 0.00 | 100.0 | |||
Negative Control | --- | 25 | 3 | 0.00 | 0.00 | 0.00 |