Rapid in vivo screening system for anti-oxidant activity using bacterial redox sensor strains.

摘要

Aim: To develop a faster and easier in vivo method to screen compounds for anti-oxidant activity using a microbial system. Methods and Results: Bacterial redox sensor-based assay systems were applied. The activities of SoxR and OxyR, the bacterial redox sensors, were monitored to probe the intracellular redox status through two reporter strains, Escherichia coli soxS_p-lacZ and oxyS_p-lacZ fusions, which specifically respond to paraquat, a superoxide generator, and H₂O₂, respectively, with practically no cross reactivity. For the test screening, 27 natural compounds including phenolics and flavonoids that are putatively considered anti-oxidant nutritional supplements were collected and assayed for their capability to alleviate oxidative stress in these bacterial systems. Among them, rutin, kaempferol and quercetin had significant anti-H₂O₂ activity, and betaine, glycyrrhizic acid and baicalin had weak anti-superoxide activity. While rutin, kaempferol and quercetin significantly reduced the H₂O₂ stress at low concentrations, betaine, glycyrrhizic acid and baicalin required higher concentration for their anti-superoxide effects. In vitro, only quercetin protected DNA in a metal-catalysed oxidation system, suggesting that the other compounds might indirectly exert their anti-oxidant activities through other biological functions. Finally, quercetin, rutin and kaempferol significantly restored the viability of a superoxide dismutase mutant that has limited viability because of defective defence against oxidative stress. Conclusion: These bacterial systems could provide a more efficient method for measuring the activity of compounds affecting cellular oxidative stress and viability. Significance and Impact of the Study: The demand for anti-oxidant and anti-ageing activities is increasing in one of the fastest growing segments of the functional food market, but the screening for these activities is currently very laborious, expensive and time consuming. This study suggests a basis for a high throughput screening method for these activities.