Apteranthes tuberculata's Antidiabetic Potential: Exploring Phytochemicals, Screening Antioxidant Activity, and Validating DPP-4 Inhibition Using In Vitro and In Silico Approaches

Abstract

Diabetes is a chronic metabolic disorder that affects an increasing number of people worldwide, frequently managed with synthetic drugs that have side effects and can be costly. Apteranthes tuberculata (N.E.Br.) Meve & Liede, a plant with traditional medicinal use in Pakistan to treat diabetes, but its antidiabetic potential has not been scientifically validated. This research assessed the phytochemicals, antioxidant properties, and dipeptidyl peptidase-4 (DPP-4) inhibitory activity of A. tuberculata's methanolic extract. The extract was assessed through in vitro antioxidant assays, DPP-4 inhibition test, and metabolomic analysis via Fourier-transform infrared (FTIR) spectroscopy and liquid chromatography-mass spectrometry (LC-MS). The study used computational tools to visualize compound structures, protein-ligand interactions, and to measure pharmacokinetic parameters. Phytochemical analysis revealed significant levels of total phenols (71.991 ± 0.78 mg/g gallic acid equivalents) and flavonoids (66.216 ± 0.09 mg/g quercetin equivalents). Results showed a robust total antioxidant capacity (70.900 ± 2 mg/g ascorbic acid), total reducing power (72.000 ± 2.00 mg/g gallic acid equivalents), and DPPH IC₅₀ value of 96.54 μg/mL. FTIR spectra showed the presence of carbohydrates and glycosides. The extract exhibited 70% DPP-4 inhibitory activity (IC₅₀ value = 46.761 ± 0.043 μg/mL), comparable to Sitagliptin at 78% (IC₅₀ value = 20.474 ± 0.407 μg/mL). LC-MS identified 24 bioactive compounds, including flavonoids and glycosides, with compounds like Kaempferol-3-O-rutinoside-7-O-glucoside and Kaempferol-7-O-rutinoside showing strong binding interactions with DPP-4. These results underscore the therapeutic potential of A. tuberculata as a natural source of DPP-4 inhibitors for managing diabetes.

Keywords: ADMET; DPP‐4 protein; diabetes mellitus; glycosides; molecular docking.

FIGURE 1

(a) Shows the percentage inhibition of DPPH radical scavenging activity by ATME at different concentrations. (b) Demonstrate the total reducing power of the ATME at various concentrations. (c) Represents the antioxidant capacity of ATME. Antioxidant activities increase with higher concentrations, indicating a dose‐dependent relationship. Concentrations for all activities range from 62.5 to 4000 μg/mL. The results are presented as the mean ± S.D. of three independent biological replicates. Mean values at each point that differ from one another are denoted by different letters and are considered significantly different according to one‐way ANOVA (p < 0.05). ATME, Apteranthes tuberculata methanolic extract.

FIGURE 2

FT‐IR spectra display the functional group of inorganic and organic compounds in A. tuberculata extract. The spectra demonstrate characteristic absorption peaks analogous to different functional groups identified in the plant extract. These functional groups are linked to the plant's therapeutic properties and bioactive potential, including antidiabetic and antioxidant activities.

FIGURE 3

Representative LC–MS chromatogram of analyzed sample. LC–MS untargeted screening of bioactive secondary metabolites was carried out to evaluate the therapeutic potential of A. tuberculata plant.

FIGURE 4

(a) In vitro DPP‐4 inhibition assay of ATME. Percentage inhibition was determined at different concentrations (7.8125–4000 μg/mL). Result shows that percentage inhibition is dose‐dependent, and the activity increased with an increase in concentration. (b) Calculated IC₅₀ value of plant extract and sitagliptin (drug) from the percentage inhibition of DPP‐IV. The results are presented as the mean ± S.D. of three independent biological replicates. Mean values at each point that differ from one another are denoted by different letters and are considered significantly different according to one‐way ANOVA (p < 0.05). ATME; Apteranthes tuberculata methanolic extract.

FIGURE 5

Three‐dimensional structure of dipeptidyl peptidase‐4 (DPP‐IV) protein [PDB accession ID: 5I7U].

FIGURE 6

It shows molecular structures. Two‐dimensional representation of the best‐docked complex of the selected dataset of chemical compounds within the binding pocket of the dipeptidyl peptidase‐4 protein. Hydrophobic residues are highlighted in red while green indicates residues forming hydrogen bonds within 4A of the active binding pocket of DPP‐IV.

FIGURE 7

It shows a three‐dimensional representation of the best‐docked complex of the selected dataset of chemical compounds within the binding pocket of the dipeptidyl peptidase‐4 protein.