Enzyme-treated red rice (Oryza sativa L.) bran extracts mitigate inflammatory markers in RAW 264.7 macrophage cells and exhibit anti-inflammatory efficacy greater/comparable to ferulic acid, catechin, γ-tocopherol, and γ-oryzanol

Ethnopharmacological relevance: Rice (Oryza sativa L.), a staple food for a significant portion of the global population, has been recognized for its traditional medicinal properties for centuries. Rice bran, a by-product of rice milling, contains many bioactive compounds with potential pharmaceutical and therapeutic benefits. In recent years, research has highlighted the anti-inflammatory potential of rice bran, contributed by the bioactive components concentrated in their bran but, unfortunately, entrapped in the bran matrix, with limited bioavailability. Previous studies have reported that the enzymatic treatment of rice bran improves the bran's bioactive compound profile but did not investigate its impact on chronic conditions such as inflammation. Aim of the study: This study investigates the anti-inflammatory effects of endo-1,4-beta-xylanase (ERB) and Viscozyme (VRB) treated red rice bran extracts against lipopolysaccharide-induced inflammation in RAW264.7 macrophages in comparison with non-enzyme-treated bran (CRB). Further established their efficacy with known anti-inflammatory compounds-ferulic acid (FA), catechin (CAT), gamma-tocopherol (GTP), and gamma-oryzanol (ORZ). Materials and methods: The RAW 264.7 macrophage cells were pre-treated with non-toxic concentrations (10-200 mu g/mL) of FA, CAT, GTP, ORZ, CRB, ERB, and VRB, followed by inflammatory stimulation with LPS for 24 h. Further, the cell supernatant and pellets were harvested to study the anti-inflammatory effects by evaluating and measuring their efficacy in inhibiting pro-inflammatory cytokines (TNF-alpha, IL-6, IL-10, IL-1 beta) and mediators (ROS, NO, PGE2, COX2, iNOS) through biochemical, ELISA, and mRNA expression studies. Results: The findings showed that both ERB and VRB effectively inhibited the production of pro-inflammatory markers (TNF-alpha, IL-6) and mediators (ROS, NO, PGE2) by downregulating mRNA expressions of inflammatory genes (TNF-alpha, IL-1 beta, IL-6, IL-10, COX2, iNOS) and demonstrated anti-inflammatory efficacy higher than CRB. On comparison, ERB demonstrated exceptional efficacy by causing a reduction of 48% in ROS, 20% in TNF-alpha, and 23% in PGE2 at 10 mu g/mL, surpassing the anti-inflammatory capabilities of all the bioactive compounds, FA and ORZ, respectively. At the same time, VRB exhibited remarkable efficacy by reducing NO production by 52% at 200 mu g/mL and IL-6 by 66% at 10 mu g/mL, surpassing FA, CAT, ORZ, and GTP. Further, ERB downregulated the mRNA expression of IL-10 and iNOS, while VRB downregulated TNF-alpha, IL-1 beta, and COX2 expression. Both extracts equally downregulated IL-6 expression at 10 mu g/mL, demonstrating the efficacy more remarkable/on par with established anti-inflammatory compounds. Conclusions: Overall, enzyme-treated rice bran/extract, particularly ERB, possesses excellent anti-inflammatory properties, making them promising agents for alternatives to contemporary nutraceuticals/functional food against inflammatory diseases.