Thai pigmented rice bran extracts inhibit production of superoxide, nitric oxide radicals and inducible nitric oxide synthase in cellular models

Objective: To study the inhibitory effect of rice bran extracts of Thai black Kam Muang and red Hawm Dawk Mali Deang on oxidative stress factors including superoxide (O 2 •- ), nitric oxide (NO • ), and inducible nitric oxide synthase (iNOS). Methods: Bran extracts (40% ethanol) of Kam Muang and Hawm Dawk Mali Deang were obtained and evaluated for in vitro 2-2’-azino-di-(3-ethylbenzthiazoline sulfonate) (ABTS) and NO • scavenging activity. Their inhibitory effects on cellular O 2 •- and NO • were measured in phorbol 12-myristate 13-acetate-stimulated neutrophil-like HL-60 cells and lipopolysaccharide- stimulated RAW264.7 macrophages, respectively, and their viability was monitored using the MTT assay. The effect on iNOS expression was also assessed by the Western blotting assay. Total contents of phenolics, flavonoids, and subtypes were also determined. Results: Hawm Dawk Mali Deang exhibited about 3.5-fold greater cellular O 2 •- inhibitory activity than Kam Muang [EC 50 values of (23.57±4.54) and (81.98±1.45) μg/mL, respectively] in phorbol 12-myristate 13-acetate-stimulated HL-60 cells. Hawm Dawk Mali Deang exhibited about 2-fold higher in vitro ABTS •+ and NO • scavenging activity than Kam Muang, but it exerted cellular NO • inhibitory activity of only about 26% (undetermined EC 50 value) in lipopolysaccharide-stimulated RAW264.7 cells. Conversely, Kam Muang exerted potent cellular NO • inhibitory activity [EC 50 value: (281.13±59.18) μg/mL] and dose-dependently decreased iNOS levels. No cytotoxicity of both extracts was detected in both cell types. As for corresponding contents, Hawm Dawk Mali Deang contained higher contents of phenolics and flavonoids than Kam Muang. Moreover, Kam Muang and Hawm Dawk Mali Deang had a high content of total anthocyanins [(14.73±0.52) mg C3GE/g of extract] and total proanthocyanidins [(115.13±1.47) mg CE/g of extract], respectively. Conclusions: Based on these data, bran extracts of Thai black Kam Muang and red rice Hawm Dawk Mali Deang can help lower oxidative stress and inflammation attributed partly to O 2 •- and NO • .