Among 40 plant-derived biflavonoids with inhibitory potential against Eg5, morelloflavone from Garcinia dulcis leaves was selected for further testing based on in silico analysis of binding modes, molecular interactions, binding energies and functional groups that interact with Eg5. Computational models predicted that morelloflavone binds the putative allosteric pocket of Eg5, within the cavity surrounded by amino acid residues of Ile-136, Glu-116, Glu-118, Trp-127, Gly-117, Ala-133, Glu-215, Leu-214 and Tyr-211. Binding energy was −8.4 kcal/mol, with a single hydrogen bond formed between morelloflavone and Tyr-211. The binding configuration was comparable to that of a reference inhibitor, S-trityl-L-cysteine. Subsequent biochemical analysis in vitro confirmed that morelloflavone inhibited both the basal and microtubule-activated ATPase activity of Eg5 in a manner that does not compete with ATP binding. Morelloflavone also suppressed Eg5 gliding along microtubules. These results suggest that morelloflavone binds the allosteric binding site in Eg5 and thereby inhibits ATPase activity and motor function of Eg5.

This study investigated the protective effect of quercetin against cyclophosphamide‐induced immunosuppressive indoleamine 2,3‐dioxygenase (IDO) via the mechanism of oxidative‐inflammatory stress and behavioral indices. Cyclophosphamide (CYP) was administered to male Wister rats at a dose of 100 mg/kg with or without quercetin 50 mg/kg every other day for 7 days. Experimental techniques including western blotting, immunohistochemistry analysis, and inflammatory and oxidative stress marker assays were carried out. We also conducted behavioral analyses such as open field, tail suspension, and Y‐maze tests for cognitive assessment. The results indicated that quercetin attenuated oxidative‐inflammatory stress induced by CYP in the hippocampus and cerebral cortex of male Wister rats by augmenting the activities of antioxidant enzymes and suppressing lipid peroxidation as well as inflammatory mediators such as interleukin‐6 and interferon‐γ. Concomitantly, quercetin partially prevented the alteration in brain tissue histological architecture and mitigated the activities of IDO/tryptophan 2,3‐dioxygenase (TDO) and protein expression of IDO1. This was corroborated by the IDO‐quercetin model obtained in silico, revealing a favorable inhibitory interaction between quercetin and the enzyme. Finally, the results of behavioral tests suggested that quercetin significantly prevented the depressive‐like posture of the CYP‐treated rats. Our study for the first time revealed that quercetin ameliorates the effect of CYP‐instigated IDO/TDO activities in the cerebral cortex and hippocampus via restoration of antioxidant enzymes and preventing oxidative‐inflammatory stress.

Clerodenrum volubile is an important locally used medicinal plant. It is one of the essential herbs nature has provided for mankind to be consumed as spices, vegetable and also used in the treatment of diseases. Qualitative phytoconstituent screening of the plant revealed the presence of steroids, flavonoids, tannins, saponins, and phenolic compound while chalcones, alkaloid and anthraquinone were absent. Quantitative phytochemical evaluation showed values of 8.29±1.26mg/g rutin equivalent for flavonoids, 3.53±0.05mg/g gallic acid equivalent for phenol, 3.97±0.03mg/g tannic acid equivalent for tannins and 13.67±1.27% for saponin per dry weight. As it is imperative to extend research work on therapeutic effects of the arsenal of plants nature has given to us in Africa so as to obtain a cure for various diseases attacking human’s health, antioxidant properties of the plant was evaluated. Antioxidant models used include iron chelating, DPPH radical, superoxide ion, hydrogen peroxide, ABTS radical, hydroxyl radical scavenging activities and ferric ion reducing properties. The obtained IC50 values against DPPH radical were 141.342 and 120.349µg/mL for Clerodendrum volubile and trolox respectively. The chelating effect of the plant extract at 50% inhibition was close to that of ascorbic acid (standard) with 134.34 and 131.19µg/mL concentration respectively. Overall, the aqueous extract of the plant showed antioxidant potential which was close to the effects exerted by known standard antioxidants (ascorbic acid, trolox and EDTA). The plant could hence provide natural antioxidants which are needed to combat numerous free radical-mediated diseases and complications such as aging, cancer, atherosclerosis, which are linked with oxidative stress.

(+)-Morelloflavone (MF) is an antitumor biflavonoid that is found in the Garcinia species. Recently, we reported MF as a novel inhibitor of ATPase and microtubules-gliding activities of the kinesin spindle protein (Eg5) in vitro. Herein, we provide dynamical insights into the inhibitory mechanisms of MF against Eg5, which involves binding of the inhibitor to the loop5/α2/α3 allosteric pocket. Molecular dynamics simulations were carried out for 100 ns on eight complexes: Eg5-Adenosine diphosphate (Eg5-ADP), Eg5-ADP-S-trityl-l-cysteine (Eg5-ADP-STLC), Eg5-ADP-ispinesib, Eg5-ADP-MF, Eg5-Adenosine triphosphate (Eg5-ATP), Eg5-ATP-STLC, Eg5-ATP-ispinesib, and Eg5-ATP-MF complexes. Structural and energetic analyses were done using Umbrella sampling, Molecular Mechanics Poisson–Boltzmann Surface Area (MM/PBSA) method, GROMACS analysis toolkit, and virtual molecular dynamics (VMD) utilities. The results were compared with those of the known Eg5 inhibitors; ispinesib, and STLC. Our data strongly support a stable Eg5-MF complex, with significantly low binding energy and reduced flexibility of Eg5 in some regions, including loop5 and switch I. Furthermore, the loop5 Trp127 was trapped in a downward position to keep the allosteric pocket of Eg5 in the so-called “closed conformation”, comparable to observations for STLC. Altered structural conformations were also visible within various regions of Eg5, including switch I, switch II, α2/α3 helices, and the tubulin-binding region, indicating that MF might induce modifications in the Eg5 structure to compromise its ATP/ADP binding and conversion process as well as its interaction with microtubules. The described mechanisms are crucial for understanding Eg5 inhibition by MF.

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