3.1 Drug likeness screening
The drug-likeness screening revealed that thirteen (13) out of the twenty-four bioactive compounds breached one or more of the five rules (Lipinski's, Ghose's, Veber's, Egan's, and Muegge's) and were excluded from subsequent analysis.
The drug-likeness screening indicated that thirteen (13) out of the twenty-four bioactive compounds violated one or more of the five rules (Lipinski's, Ghose's, Veber's, Egan's, and Muegge's) and were eliminated from further analysis. Consequently, the eleven phytocompounds (Castincin, Rotundifuram, Curcumin, Demethoxycurcumin, Bisdemethoxycurcumin, Cyclocurcumin, Gingerol, Shogaol, Gingerdiol, Paradol, and Ajoene) without any violation of the rule, were considered for molecular docking analysis (Table 2). Moreover, among the eleven (11), three (3) bioactive compounds had lower binding energies compared to the control drug and were also removed and not considered for the pharmacokinetic screening. The remaining eight (8) ligands adhered to all the five drug-likeness screening rules and had higher or equal binding affinity with the control drug (Table 3).
Table 2
Drug-likeness screening of phytocompounds and the control drug using the SwissADME online tool.
S/N
|
Molecule
|
Formula
|
MW
|
XLO GP
|
TPSA
|
Lipinski
#violation
|
Ghose
#violation
|
Veber
#violation
|
Egan
#violation
|
Muegge
#violation
|
Bioavailability
|
1
|
Agnuside
|
C22H26O11
|
466.44
|
-1.15
|
175.37
|
2
|
1
|
1
|
1
|
3
|
0.17
|
2
|
Casticin
|
C19H18O8
|
374.34
|
3.12
|
107.59
|
0
|
0
|
0
|
0
|
0
|
0.55
|
3
|
Vitexin
|
C21H20O10
|
432.38
|
0.21
|
181.05
|
1
|
0
|
1
|
1
|
2
|
0.55
|
4
|
Rotundifuran
|
C22H34O4
|
362.5
|
5
|
59.67
|
0
|
0
|
0
|
0
|
0
|
0.55
|
5
|
Curcumin
|
C21H20O6
|
368.38
|
3.2
|
93.06
|
0
|
0
|
0
|
0
|
0
|
0.55
|
6
|
Demethoxycurcumin
|
C20H18O5
|
338.35
|
3.32
|
83.83
|
0
|
0
|
0
|
0
|
0
|
0.55
|
7
|
Bisdemethoxycurcumin
|
C19H16O4
|
308.33
|
3.26
|
74.6
|
0
|
0
|
0
|
0
|
0
|
0.55
|
8
|
Turmerone
|
C15H22O
|
218.33
|
3.33
|
17.07
|
0
|
0
|
0
|
0
|
1
|
0.55
|
9
|
Cyclocurcumin
|
C21H20O6
|
368.38
|
3
|
85.22
|
0
|
0
|
0
|
0
|
0
|
0.56
|
10
|
Gingerol
|
C17H26O4
|
294.39
|
2.76
|
66.76
|
0
|
0
|
0
|
0
|
0
|
0.55
|
11
|
Shogaol
|
C17H24O3
|
276.37
|
3.99
|
46.53
|
0
|
0
|
0
|
0
|
0
|
0.55
|
12
|
Gingerdiol
|
C17H28O4
|
296.4
|
3.27
|
69.92
|
0
|
0
|
0
|
0
|
0
|
0.55
|
13
|
Paradol
|
C17H26O3
|
278.39
|
4.11
|
46.53
|
0
|
0
|
0
|
0
|
0
|
0.55
|
14
|
Zingiberene
|
C15H24
|
204.35
|
5.17
|
0
|
1
|
0
|
0
|
0
|
2
|
0.55
|
15
|
Allicin
|
C6H10OS2
|
162.27
|
1.31
|
61.58
|
0
|
1
|
0
|
0
|
1
|
0.55
|
16
|
Diallyl sulfide
|
C6H10S
|
114.21
|
2.16
|
25.3
|
0
|
3
|
0
|
0
|
2
|
0.55
|
17
|
Diallyl disulfide
|
C6H10S2
|
146.27
|
2.2
|
50.6
|
0
|
2
|
0
|
0
|
1
|
0.55
|
18
|
Diallyl trisulfide
|
C6H10S3
|
178.34
|
2.64
|
75.9
|
0
|
1
|
0
|
0
|
1
|
0.55
|
19
|
Thiosulfinate
|
C6H10OS2
|
162.27
|
1.47
|
61.58
|
0
|
1
|
0
|
0
|
1
|
0.55
|
20
|
Ajoene
|
C9H14OS3
|
234.4
|
1.71
|
86.88
|
0
|
0
|
0
|
0
|
0
|
0.55
|
21
|
Flavonoids
|
C26H28O6
|
436.5
|
5.87
|
100.13
|
0
|
1
|
0
|
0
|
1
|
0.55
|
22
|
Galactomannan
|
C18H32O16
|
504.44
|
-6.33
|
268.68
|
3
|
2
|
1
|
1
|
4
|
0.17
|
23
|
Paeoniflorin
|
C23H28O11
|
480.46
|
-1.02
|
164.37
|
1
|
2
|
1
|
1
|
2
|
0.55
|
24
|
Paeonol
|
C9H10O3
|
166.17
|
1.98
|
46.53
|
0
|
0
|
0
|
0
|
1
|
0.55
|
25
|
Ulipristal Acetate
|
C30H37NO4
|
475.62
|
3.47
|
63.68
|
0
|
2
|
0
|
0
|
0
|
0.55
|
3.2 Molecular docking and interaction of progesterone receptor (1a28) and ligands
Table 3 shows the binding energy and molecular interaction among the bioactive compounds and the amino acid residues located at the catalytic site of the Progesterone Receptor. Additionally, it illustrates the binding efficiency, electrostatic energy, hydrophobic, and hydrogen bond interactions for each compound during molecular docking with the Progesterone Receptor (1A28).
Casticin, Curcumin, Demethoxycurcumin, Bisdemethoxycurcumin, and Cyclocurcumin had a binding energy of -7.6, -7.0, -7.5, -7.4, and − 8.0 kcal/mol, respectively with the Progesterone Receptor (Table 3). Moreover, Gingerol, Gingerdiol, Shagaol, and the control drug Ulipristal Acetate had a binding energy of -7.0, -6.7, -6.7, and − 6.7, respectively (Table 3).
Table 3
Binding energy and molecular interaction of phytocompounds and control drugs with varying residues
S/N
|
Plant
Source
|
Molecule
|
Binding
energy
(kcal/mol)
|
Number of
hydrogen
bond (s)
formed
|
Residues involved
in hydrogen bond
formation (Å)
|
Residues involved
in hydrophobic
interaction
(Å)
|
Residues
involved in
π-stacking
(Å)
|
Residues
involved in
π-cation interaction
(Å)
|
1.
|
Vitex agnus castus
|
Casticin
|
-7.6
|
4
|
ASP697(2.91), VAL6983.03), TRP765(3.88), ARG766(4.04).
|
VAL698(3.77, 3.86)
|
|
ARG766(4.62)
|
2.
|
Curcumin longa
|
Curcumin
|
-7.0
|
0
|
|
LEU715(3.68), LEU889(3.84), TYR890(3.49, 3.80)ASN893(3.76), LEU901(3.44).
|
|
|
3.
|
|
Demethoxycurcumin
|
-7.5
|
4
|
SER757(3.12), THR829(3.63), LYS(3.59), LYS885(3.14).
|
THR829(3.80), HIS881(3.81), ASP882(3.64), VAL884(3.43), ILE920(3.55), LEU929(3.62)
|
HIS881(4.92)
|
|
4.
|
|
Bisdemethoxycurcumin
|
-7.4
|
5
|
THR829(3.19), LYS875(3.03), ASN879(3.55), ASP882(3.43), LYS885(3.25).
|
THR829(3.48), ASP878(3.98), HIS881(3.69), ASP882(3.59), ILE920(3.61), VAL925(3.81), LEU929(3.58).
|
HIS881(4.98)
|
|
5.
|
|
Cyclocurcumin
|
-8.0
|
3
|
SER757(2.73, 2.73), LYS885(3.69).
|
THR829(3.74), HIS881(3.45), ASP882(3.69), LYS885(3.95), ILE920(3.29), PRO927(3.83), LEU929(3.68).
|
HIS888(5.08)
|
|
6.
|
Zingiber officinale
|
Gingerol
|
-7.0
|
3
|
ILE699(3.10), ARG766(3.04), HIS770(3.00).
|
PRO696(3.69), VAL698(3.65), ILE699(3.75), GLN725(3.66), VAL729(3.58, 3.56), TRP732(3.68), LEU758(3.65), ARG766(3.97), LYS822(3.63).
|
|
|
7.
|
|
Gingerdiol
|
-6.7
|
7
|
ILE699(3.77), SER728(3.25), VAL729(3.55), TRP765(3.27), ARG766(3.29), HIS770(3.68)
|
PRO696(3.67), VAL698(3.75), ILE699(3.47), GLN725(3.65), VAL729(3.51), TRP732(3.71), LEU758(3.46), ARG766(3.85), LYS822(3.28).
|
|
|
8.
|
|
Shogaol
|
-6.7
|
4
|
ILE699(3.27), TRP765(3.04), HIS770(3.80), GLN815(3.83).
|
VAL698(3.62), ILE699(3.30), GLN725(3.80), LEU758(3.96), ARG766(3.77), LYS822(3.90)
|
|
|
9.
|
|
Ulipristal Acetate
|
-6.7
|
0
|
|
LEU797(3.97), LEU889(3.80), ASN893(3.77).
|
TYR890(4.30)
|
|
Casticin, the bioactive compound in Vitex agnus castus has a binding energy of -7.6 kcal/mol. The binding configuration of the compound in the progesterone active site is shown in Fig. 2. It established four hydrogen bonds with ASP697, VAL698, TRP765, ARG766; had single hydrophobic interaction with VAL698 and also formed a π-cation interaction with ARG766 (Fig. 2).
Analysis of binding interactions was conducted using a protein-ligand interaction profiler. Blue dashed line - Hydrogen bond; Grey dotted line – Hydrophobic interaction.
Curcumin demonstrated hydrophobic interaction with LEU715, LEU889, TYR890, ASN893, and LEU901 (Fig. 3). Using hydrogen bonds, Demethoxycurcumin interacted with SER757, THR829, LYS875, LYS885 and showed hydrophobic interaction with THR829, HIS881, ASP882, VAL884, ILE920, and LEU929. Moreover, it established an interaction with π-stacking HIS881. Bisdemethoxycurcumin established five hydrogen bonds with THR829, LYS875, ASN879, ASP882, LYS885 while hydrophobic interaction was established with THR829, ASP878, HIS881, ASP882, ILE920, VAL925, LEU929. It also engaged in a π-stacking interaction with HIS881 (Fig. 3). Cyclocurcumin interacts with SER757 and LYS885 through hydrogen bonds while hydrophobic interaction with THR829, HIS881, ASP882, LYS885, ILE920, PRO927, LEU929 were also established. It also formed a π-stacking interaction with HIS881 (Fig. 3).
Analysis of binding interactions was conducted using a protein-ligand interaction profiler. Blue dashed line - Hydrogen bond; Grey dotted line – Hydrophobic interaction; Green dotted line – Pi stacking.
Shogaol established four hydrogen bonds with ILE699, TRP765, HIS779, GLN815 and interacted hydrophobically with VAL698, ILE699, GLN725, LEU758, ARG766, LYS822 (Fig. 4). Gingerol demonstrated three hydrogen bond interactions with ILE699, ARG766, and HIS770. Gingerol also interacted hydrophobically with PRO696, VAL698, ILE699, GLN725, VAL729, TRP732, LEU758, ARG766 and LYS822. Gingerdiol interacted with ILE699, SER728, VAL729, TRP765, ARG766 and HIS770, using hydrogen bonds and established hydrophobic interaction with PRO696, VAL698, ILE699, GLN725, VAL729, LEU758, ARG766 and LYS822 (Fig. 4).
Ulipristal Acetate penetrates the active sites of metabolism in the target protein. It also established hydrophobic interaction with LEU797, LEU889, and ASN893 present at the catalytic site of the Progesterone Receptor. It also possessed π-stacking interaction TYR890 but showed no hydrogen bond (Fig. 5).
Analysis of binding interactions was conducted using a protein-ligand interaction profiler. Blue dashed line - Hydrogen bond; Grey dotted line – Hydrophobic interaction.
Analysis of binding interactions was conducted using a protein-ligand interaction profiler. Grey dotted line - Hydrophobic interaction; Green dotted line - Pi stacking.
The screened phytocompounds together with the control drug were predicted to be highly active as enzyme inhibitors. They were also predicted to be moderately active as GPCR ligand and ion channel modulators since their bioactivity scores ranged between − 5.0 to 0.0. However, Gingerol. Shogaol, and Gingerdiol were considered highly active as GPCR ligands, ion channel modulators, and nuclear receptor ligands. Notably, among all bioactive compounds, Gingerdiol was predicted as the most active, as it shows high activity (> 0.0) at different levels of the measured parameters (GPCR ligand, ion channel modulator, kinase inhibitor, nuclear receptor ligand, protease inhibitor, and enzymes inhibitor).
Table 4
Predicted bioactivity score for bioactive compounds and the control drug
S/N
|
Compound name
|
GPCR ligand
|
Ion channel modulator
|
Kinase inhibitor
|
Nuclear receptor ligand
|
Protease inhibitor
|
Enzymes inhibitor
|
1.
|
Casticin
|
-0.14
|
-0.27
|
0.13
|
0.01
|
-0.34
|
0.12
|
2.
|
Curcumin
|
-0.06
|
-0.20
|
-0.26
|
0.12
|
-0.14
|
0.08
|
3.
|
Demethoxycurcumin
|
-0.04
|
-0.20
|
-0.26
|
-0.18
|
-0.14
|
0.10
|
4.
|
Bisdemethoxycurcumin
|
0.00
|
-0.14
|
-0.26
|
-0.25
|
-0.08
|
0.15
|
5.
|
Cyclocurcumin
|
-0.04
|
-0.31
|
-0.28
|
0.10
|
-0.11
|
0.09
|
6.
|
Gingerol
|
0.16
|
0.04
|
-0.33
|
0.20
|
0.15
|
0.38
|
7.
|
Shogaol
|
0.06
|
0.01
|
-0.50
|
0.20
|
-0.05
|
0.29
|
8.
|
Gingerol
|
0.31
|
0.28
|
0.01
|
0.48
|
0.24
|
0.49
|
9.
|
Ulipristal Acetate
|
-0.04
|
-0.08
|
0.58
|
1.03
|
-0.05
|
0.26
|
3.3 Pharmacokinetics properties prediction
The In Silico ADMET profiling of both phytocompounds and the control drug is shown in Table 5. For drug absorption, all the hit ligands except the control drug Ulipristal Acetate were predicted to penetrate the blood-brain barrier (BBB). Also, all lead ligands except Ulipristal Acetate had low absorption in the intestine through Caco-2 permeability. In terms of drug metabolism, the majority of the compounds and the control drug exhibited no inhibition of certain key enzymes (CYP450 2C9 and CYP450 2C19). Bisdemethoxycurcumin, Gingerol, Shogaol, and Gingerdiol were substrates of CYP450 3A4, while Casticin, Curcumin, Demethoxycurcumin, Cyclocurcumin and the control drug might likely inhibit CYP450 3A4 (as they were predicted as substrates) (Table 5). Only the control drug Ulipristal Acetate was found to be a potential substrate of CYP450 2C9. For toxicity, Casticin, Gingerol, Shogaol, and Gingerdiol were predicted to cause hepatotoxicity in humans. Curcumin, Gingerol, Shogaol, Gingerdiol, and Ulipristal Acetate were predicted to show toxicity for Salmonella typhimurium reverse mutation assay. Casticin, Gingerol, Shogaol, and Gingerdiol were predicted to be carcinogenic (Table 5).
Table 5
Predicted ADMET properties of bioactive compounds and the control drug
S/N.
|
Class
|
Casticin
|
Curcumin
|
Demethoxycurcumin
|
Bisdemethoxycurcumin
|
Cyclocurminin
|
Gingerol
|
Shogaol
|
Gingerdiol
|
Ulipristal Acetate
|
1.
Absorption
|
BBB
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
NO
|
|
Caco-2
Permeability
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
NO
|
|
Pgp-inhibitor
|
NO
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
NO
|
|
Pgp
Substrate
|
YES
|
YES
|
YES
|
YES
|
YES
|
NO
|
YES
|
NO
|
YES
|
2.
Distribution
|
PPB
|
84.41%
|
99.80%
|
100.02%
|
99.29%
|
99.17%
|
87.15%
|
96.35%
|
91.17%
|
93.60%
|
3.
Metabolism
|
CYP450 1A2
Inhibition
|
NO
|
NO
|
NO
|
NO
|
NO
|
YES
|
NO
|
YES
|
YES
|
|
CYP450 3A4
Inhibition
|
NO
|
NO
|
NO
|
NO
|
NO
|
YES
|
NO
|
YES
|
NO
|
|
CYP450 3A4
Substrate
|
NO
|
NO
|
NO
|
YES
|
NO
|
YES
|
YES
|
YES
|
NO
|
|
CYP450 2C9
Inhibition
|
NO
|
NO
|
NO
|
NO
|
NO
|
YES
|
NO
|
YES
|
NO
|
|
CYP450 2C9
Substrate
|
NO
|
NO
|
NO
|
NO
|
NO
|
NO
|
NO
|
N0
|
YES
|
|
CYP450 2C19
Inhibition
|
NO
|
YES
|
NO
|
NO
|
NO
|
YES
|
NO
|
YES
|
NO
|
|
CYP450 2D6
Inhibition
|
YES
|
NO
|
YES
|
YES
|
YES
|
YES
|
NO
|
YES
|
NO
|
|
CYP450 2D6
Substrate
|
NO
|
NO
|
NO
|
NO
|
NO
|
NO
|
NO
|
NO
|
YES
|
4. Toxicity
|
Acute oral
toxicity
|
NO
|
NO
|
NO
|
NO
|
NO
|
YES
|
YES
|
YES
|
YES
|
|
hERG
inhibitor
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
YES
|
|
Human
hepatotoxicity
|
YES
|
NO
|
NO
|
NO
|
NO
|
YES
|
YES
|
YES
|
NO
|
|
Ames
mutagenicity
|
NO
|
YES
|
NO
|
NO
|
NO
|
YES
|
YES
|
YES
|
YES
|
|
Carcinogenicity
|
YES
|
NO
|
NO
|
NO
|
NO
|
YES
|
YES
|
YES
|
NO
|
BBB - blood-brain barrier, PPB - plasma protein binding, hERG - human ether-a-go-go.