Molecules · 8 authors, 9 centres
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This study found that extracts from Alstonia boonei stem bark, particularly the dichloromethane fraction, exhibit antispasmodic activity on isolated rat ileum. The isolated compound boonein showed strong antispasmodic effects, while β-amyrin demonstrated stronger binding to target proteins in silico. These findings suggest that boonein and β-amyrin could serve as starting materials for developing antidiarrheal drugs.
**Background:** Alstonia boonei (Apocynaceae) is a medicinal plant traditionally used in Africa and Asia for muscle relaxation and treating various conditions including gastrointestinal disorders. Spasms are involuntary smooth muscle contractions underlying many GI disorders like colic, diarrhea, and irritable bowel syndrome. Current antispasmodic drugs have side effects, prompting interest in plant-derived alternatives. This study investigated the antispasmodic properties of A. boonei stem bark extracts and isolated compounds using ex vivo and in silico approaches.
**Methods:** Freeze-dried aqueous stem bark extract of A. boonei was partitioned into dichloromethane (DCM), ethyl acetate, and aqueous fractions. Antispasmodic activity was evaluated ex vivo on isolated rat ileum using spontaneous and high-potassium-induced (80 mM) contractions. Two compounds were isolated from the DCM fraction via column chromatography and identified using NMR and mass spectrometry. In silico studies involved molecular docking of isolated compounds with human PPARgamma-LBD (PDBID: 4F9M) and human carbonic anhydrase isozyme 1 (PDBID: 5E2M), followed by 100 ns molecular dynamics simulations and MM-GBSA binding free energy calculations.
**Key Results:** The crude extract, DCM, ethyl acetate, and aqueous fractions showed antispasmodic activity on high-potassium-induced contractions with IC50 values of 0.03 ± 0.20, 0.02 ± 0.05, 0.03 ± 0.14, and 0.90 ± 0.06 mg/mL, respectively. The DCM fraction had the highest activity on both spontaneous (IC50 = 0.31 ± 0.02 mg/mL) and high-potassium-induced (IC50 = 0.02 ± 0.05 mg/mL) contractions. Two compounds were isolated: boonein (C9H14O3, MW 170.0 g/mol) and β-amyrin (C30H50O, MW 426.8 g/mol). Boonein showed concentration-dependent antispasmodic activity on spontaneous (IC50 = 0.29 ± 0.05 µg/mL) and high-potassium-induced (IC50 = 0.09 ± 0.01 µg/mL) contractions. β-amyrin showed spasmodic activity at lower concentrations (0.003–5 µg/mL) and antispasmodic effect at 10 µg/mL (IC50 = 2.2 ± 0.7 µg/mL on spontaneous contractions). In molecular docking, β-amyrin had binding energies of −9.4 Kcal/mol (carbonic anhydrase) and −8.4 Kcal/mol (PPARgamma), comparable to reference inhibitors (−9.4 and −8.2 Kcal/mol, respectively). Boonein had lower binding energies (−6.4 and −6.0 Kcal/mol). MD simulations showed β-amyrin complexes had lower average RMSD (1.972 Å for 4F9M, 2.419 Å for 5E2M) compared to boonein (1.819 Å and 3.630 Å). MM-GBSA binding free energies were higher for β-amyrin (−27.02 Kcal/mol for 4F9M, −12.88 Kcal/mol for 5E2M) than boonein (−0.86 and −4.19 Kcal/mol).
**Clinical Implications:** The study demonstrates that A. boonei stem bark extracts, particularly the DCM fraction, possess significant antispasmodic activity mediated through calcium channel blockade. Boonein emerged as a potent antispasmodic compound, ten times more effective than β-amyrin, suggesting its potential as an antidiarrheal agent. β-amyrin showed stronger and more stable interactions with target proteins in silico, indicating possible therapeutic relevance. These findings support the traditional use of A. boonei for muscle relaxation and gastrointestinal disorders. The isolated compounds could serve as lead molecules for developing safer, plant-derived antispasmodic drugs, potentially offering alternatives to current medications with fewer side effects. Further studies are needed to evaluate efficacy and safety in vivo.