Inhibition effect of thymoquinone and lycopene compounds on glutathione reductase enzyme activity purified from human erythrocytes

Glutathione reductase (GR, EC 1.8.1.7) is a specific antioxidant enzyme that catalyzes oxidized glutathione (GSSG) to reduced glutathione (GSH). GR enzyme maintains the cellular reduced GSH level and plays a central role in cell defense against reactive oxygen species. Herein, GR was purified with affinity chromatography method in one step using 2′,5′-ADP Sepharose 4B from human erythrocytes. The purification rate of glutathione reductase enzyme purified from human erythrocytes was 6224 fold and specific activity was calculated as 9.586 EU/mg protein. The molecular weight of GR was determined to be 53 kDa by SDS-PAGE. The effect of thymoquinone and lycopene compounds on the GR activity purified from human erythrocytes was researched. Both compounds showed an inhibitory effect on GR activity. IC50 values for thymoquinone and lycopene were calculated as 62.12 µM and 35.79 µM, respectively. Inhibition type and Ki values were determined from the Lineveawer-Burk graph. The type of inhibition for thymoquinone and lycopene was found to be non-competitive inhibition. Ki value was calculated as 57.71 µM for thymoquinone and 46.65 µM for lycopene. In this study, it was concluded that antioxidant compounds thymoquinone and lycopene, which have an inhibitory effect on GR activity, may have a therapeutic effect on cancer disease.Communicated by Ramaswamy H. Sarma.

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Preparation of glutathione loaded nanoemulsions and testing of hepatoprotective activity on THLE-2 cells

To improve bioavailability and stability of hydrophobic and hydrophilic compounds, nanoemulsions are good alternatives as delivery systems because of their nontoxic and nonirritant nature. Glutathione (GSH) suffers from low stability in water, where its encapsulation in nanoemulsions is a powerful strategy to its stability in aqueous systems. The aim of this study was to obtain nanoemulsions from the hydrophobic/hydrophilic contents of N. sativa seed oil so as to improve GSH stability along with bioavailability of N. sativa seed oil. Then, the prepared nanoemulsions were tested for in vitro hepatoprotective activity against ethanol toxicity. To the best of our knowledge, there is no study on the test of nanoemulsions by the combination of Nigella sativa seed oils and GSH in hepatoprotective activity. Here, nanoemulsions with different contents were prepared using Nigella sativa seed oils. Content analyses and characterisation studies of prepared nanoemulsions were carried out. In order to investigate the protective effects against to ethanol exposure, THLE-2 cells were pretreated with nanoemulsions for 2 h with the maximum benign dose (0.5 mg/mL of nanoemulsions). Ethanol (400 mM) was introduced to pretreated cells and nontreated cells for 48- or 72-h periods, followed by cell viability assay was carried out. Fluorescence microscopy tests revealed the introduction of the nanoemulsions into THLE-2 cells. The findings show that nanoformulations have promising in vitro hepatoprotective effects on the THLE-2 cell line against ethanol exposure.

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