A magnetic beads-based ligand fishing method for rapid discovery of monoterpene indoles as monoamine oxidase A inhibitors from Hunteria zeylanica.

In this study, a novel magnetic bead-based ligand fishing method was developed for rapid discovery of monoterpene indoles as monoamine oxidase A inhibitors from natural products. In order to improve the screening efficiency, two different magnetic beads, i.e. amine and carboxyl terminated magnetic beads, were comprehensively compared in terms of their ability to immobilize monoamine oxidase A (MAOA), biocatalytic activity and specific adsorption rates for affinity ligands. Carboxyl terminated magnetic beads performed better for MAOA immobilization and demonstrated superior performance in ligand fishing. The MAOA immobilized magnetic beads were applied to screen novel monoamine oxidase inhibitors in an alkaloid-rich plant, Hunteria zeylanica. Twelve MAOA affinity ligands were screened out, and ten of them were identified as monoterpene indole alkaloids by HPLC-Obitrap-MS/MS. Among them, six ligands, namely geissoschizol, vobasinol, yohimbol, dihydrocorynanthenol, eburnamine and (+)-isoeburnamine which exhibited inhibitory activity against MAOA with low IC50 values. To further explore their inhibitory mechanism, enzyme kinetic analysis and molecular docking studies were conducted.

Effect of astragaloside IV against rat myocardial cell apoptosis induced by oxidative stress via mitochondrial ATP-sensitive potassium channels.

Astragaloside is one of the most common traditional Chinese medicines and is derived from Astragalus membranaceus. Astragaloside IV (AsIV) is a monomer located in an extract of astragaloside. The current study investigated the protective effects of AsIV against hydrogen peroxide (H2O2)-induced injury in cardiocytes and elucidated the mechanisms responsible for this protective effect. Cultured neonatal rat cardiocytes were divided into five experimental groups as follows: i) Dimethyl sulfoxide; ii) H2O2; iii) AsIV+H2O2; iv) AsIV+H2O2+5-hydroxydecanoate (5-HD); and v) nicorandil+H2O2. Cardiocyte survival was analyzed using an MTT assay. Lactate dehydrogenase (LDH) release was also assessed to evaluate the viability of the cells. Intracellular reactive oxygen species (ROS) were measured by 2,7-dichlorodihydrofluorescein diacetate staining. The apoptotic rate was measured by flow cytometry. Mitochondrial membrane potential (ΔΨm) and intracellular calcium were observed using a laser confocal microscopy system. The results indicated that AsIV promoted the survival of cardiocytes (P<0.05), attenuated LDH release (P<0.05), ROS production (P<0.01) and apoptosis (P<0.01), stabilized the ΔΨm and reduced intracellular calcium overload (P<0.01) compared with the H2O2 group. The mitochondrial adenosine triphosphate-sensitive potassium channel (mitoKATP) inhibitor 5-HD was observed to partially reverse the protective effect of AsIV. Following treatment with 5-HD, the survival of cardiocytes was reduced (P<0.05), LDH release (P<0.01) and ROS production (P<0.05) were stimulated, ΔΨm and intracellular calcium change were increased (P<0.01) and apoptosis was increased (P<0.01) compared with the AsIV+H2O2 group. Thus, AsIV has potential for use in the suppression of apoptosis resulting from H2O2 exposure, and mitoKATP activation may underlie this protective mechanism.