Kratom (Mitragyna speciosa)
Evaluation of selected Malaysian medicinal plants on phase I drug metabolizing enzymes, CYP2C9, CYP2D6 and CYP3A4 activities in vitro
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This study investigated the effects of selected Malaysian medicinal plant extracts towards human recombinant cytochrome P450 (CYP450) enzyme activities in vitro. Five Malaysian medicinal plants were tested on the three main CYP450 enzyme activities of CYP2C9, CYP2D6 and CYP3A4. The abilities of these extracts to inhibit human cytochrome P450 enzyme activities were analyzed using a luminescent assay. Orthosiphon stamineus showed the most potent inhibitory activity against CYP2C9 with an apparent IC50 value of 77.5±1.1 μg mL-1, while Andrographis paniculata, Curcuma xanthorrhiza, Eurycoma longifolia and Mitragyna speciosa extracts showed negligible inhibition. On the metabolism mediated by CYP2D6, Mitragyna speciosa showed the most potent inhibitory activities with IC50 values of 3.6±0.1 μg mL-1, followed by Orthosiphon stamineus, Andrographis paniculata and Curcuma xanthorrhiza with IC50 value of 11.7±1.1, 44.2±4.5 and 215.3±71.6 fig mL-1, respectively. Andrographis paniculata ethanolic extract gave the lowest IC50 value towards CYP3A4 with an apparent IC50 value of 27.6±3.7 μg mL-1, followed by Orthosiphon stamineus (78.4±20.3 μg mL-1), Mitragyna speciosa (142.8±13.8 μg mL-1) and Curcuma xanthorrhiza (285.3±61.7 μg mL-1). Sulfaphenazole, quinidine and ketoconazole were used as positive controls for CYP2C9, CYP2D6 and CYP3A4 respectively. The findings suggest that Orthosiphon stamineus, Mitragyna speciosa and Andrographis paniculata may contribute to herb-drug interactions if they are administered concomitantly with drugs metabolized by CYP2C9, CYP2D6 and CYP3A4 respectively.
1 . POS. CONTROL: Inhibition of CYP2C9 by sulfaphenazole (id=NPDI-H6ZqbA)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified.
- CYP2C9 4309227
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 Not available
Positive control
Results
Positive control
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
Positive controls sulfaphenazole, quinidine, and ketoconazole were purchased from Sigma Chemicals (St. Louis, USA). This assay was carried out using the P450-Glo™ Screening Systems from Promega, USA.
The minus-P450 wells (using luciferin-free water) were negative controls.
Commercially available
100 µL
50 min
MgCl2
NADPH regenerating system
Not available
Not available
0.02-200 µM
2 . Inhibition of CYP2D6 by M. speciosa methanolic extract (id=NPDI-gWxQZg)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified.
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 Not available
Results
% enzyme inhibition versus precipitant concentration:
Values estimated from Figure 1d. In some cases, SEM could not be estimated from the provided figure.
0.01 µg/mL: 25 ± Unknown%
0.1 µg/mL: 21 ± Unknown%
1 µg/mL: 35 ± 1%
10 µg/mL: 78 ± 0%
100 µg/mL: 91 ± 3%
1000 µg/mL: See above data
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
M. speciosa methanolic extract was prepared in-house from the leaves of the plant. This assay was carried out using the P450-Glo™ Screening Systems from Promega, USA.
The minus-P450 wells (using luciferin-free water) were negative controls.
Commercially available
100 µL
65 min
MgCl2
NADPH regenerating system
Not available
Not available
0.01-1000 µg/mL
3 . POS. CONTROL: Inhibition of CYP2D6 by quinidine (id=NPDI-FnVAqw)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified.
- CYP2D6 4173631
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 Not available
Positive control
Results
Positive control
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
Positive controls sulfaphenazole, quinidine, and ketoconazole were purchased from Sigma Chemicals (St. Louis, USA). This assay was carried out using the P450-Glo™ Screening Systems from Promega, USA.
The minus-P450 wells (using luciferin-free water) were negative controls.
Commercially available
100 µL
65 min
MgCl2
NADPH regenerating system
Not available
Not available
0.02-200 µM
4 . POS. CONTROL: Inhibition of CYP3A4 by ketoconazole (id=NPDI-79TG7g)
In Vitro Enzyme Inhibition Experiment
Inhibition was detected. Cutoff used — Not specified.
- CYP3A4 4306811
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 Not available
Positive control
Results
Positive control
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
Positive controls sulfaphenazole, quinidine, and ketoconazole were purchased from Sigma Chemicals (St. Louis, USA). This assay was carried out using the P450-Glo™ Screening Systems from Promega, USA.
The minus-P450 wells (using luciferin-free water) were negative controls.
Commercially available
100 µL
50 min
MgCl2
NADPH regenerating system
Not available
Not available
0.02-200 µM
5 . Inhibition of CYP2C9 by M. speciosa methanolic extract (id=NPDI-jkwBEg)
In Vitro Enzyme Inhibition Experiment
Negligible Inhibition was detected. Cutoff used — Not specified.
- CYP2C9 4309227
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 Not available
Results
IC50 not determined due to the less than 50% of inhibition.
% enzyme inhibition versus precipitant concentration:
Values estimated from Figure 1d. In some cases, SEM could not be estimated from the provided figure.
0.01 µg/mL: 20 ± Unknown%
0.1 µg/mL: 18 ± Unknown%
1 µg/mL: 22 ± 6%
10 µg/mL: 23 ± 6%
100 µg/mL: 26 ± 2%
1000 µg/mL: See above data
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
M. speciosa methanolic extract was prepared in-house from the leaves of the plant. This assay was carried out using the P450-Glo™ Screening Systems from Promega, USA.
The minus-P450 wells (using luciferin-free water) were negative controls.
Commercially available
100 µL
50 min
MgCl2
NADPH regenerating system
Not available
Not available
0.01-1000 µg/mL
6 . Negligible inhibition of CYP3A4 by M. speciosa methanolic extract (id=NPDI-Df2Xyw)
In Vitro Enzyme Inhibition Experiment
Negligible Inhibition was detected. Cutoff used —
Not specified.
- CYP3A4 4306811
Recombinant expression system
Baculovirus-insect cells
Cytochrome B5 Not available
Results
% enzyme inhibition versus precipitant concentration:
Values estimated from Figure 1d. In some cases, SEM could not be estimated from the provided figure.
0.01 µg/mL: 18 ± Unknown%
0.1 µg/mL: 11 ± 4%
1 µg/mL: 9± 0%
10 µg/mL: 18 ± 4%
100 µg/mL: 50 ± 3%
1000 µg/mL: See above data
| Sample | Compound measured | Value | Measurement | Study sequence | Additional information | N replicates |
|---|
Experimental Conditions
M. speciosa methanolic extract was prepared in-house from the leaves of the plant. This assay was carried out using the P450-Glo™ Screening Systems from Promega, USA.
The minus-P450 wells (using luciferin-free water) were negative controls.
Commercially available
100 µL
50 min
MgCl2
NADPH regenerating system
Not available
Not available
0.01-1000 µg/mL