racemosa stem extract required a higher concentration (⩾500 μg/ml) to achieve similar inhibition. Gallic acid was more potent, requiring a lower concentration (50 μg/ml) to reach similar inhibition of LHP production. These results demonstrate that in addition to preventing the formation of MDA, B. racemosa extracts are also able to inhibit the formation of LHP. During redox reaction, Hb enhances the reduction of nitrite ( NO2-) to nitric oxide (NO), converting ferrohaem (Fe2+) to Smad2 phosphorylation ferrihaem (Fe3+), thus causing the formation

of MetHb. MetHb-mediated LDL oxidation has been postulated to promote atherosclerosis (Umbreit, 2007). In this study, the effect of B. racemosa leaf extract, stem extract and gallic acid on Hb oxidation was measured via a NO2-induced MetHb formation method ( Table 3). B. racemosa leaf extract showed a concentration-dependent increase in the inhibition of MetHb formation and the highest

inhibition was seen at 500 μg/ml (79.51%). B. racemosa stem extract showed a slightly different pattern of inhibition, with lower inhibition of MetHb formation at low concentrations (25–100 μg/ml), after which there Selleck Alpelisib was a considerable leap in the inhibition of MetHb formation at concentrations above 250 μg/ml. Silibinin, a flavonoid, was reported to show similar dose–response relationship in Hb oxidation ( Marouf et al., 2011). A threefold lower concentration of B. racemosa leaf extract (116 μg/ml) was needed to inhibit approximately 50% of MetHb formation compared to its stem extract (385 μg/ml).

Gallic acid on the other hand showed pro-oxidant activities by increasing MetHb formation, particularly at high concentrations. High concentrations of gallic acid may increase NO production, NO may form nitrite ( NO2-) via auto-oxidation, further reacting with Hb, leading to formation of MetHb and NO (Umbreit, 2007). This implies that lower Chlormezanone concentrations would be more biologically relevant, especially for pure compounds. In another study, gallic acid, at a concentration of 50 μg/ml prevented lipid peroxidation of erythrocytes and did not exhibit pro-oxidant effects (Hseu et al., 2008). This supports our observation that gallic acid is protective at low concentrations. Overall, B. racemosa leaf and stem extracts could delay the time to achieve maximal MetHb formation as well as the time needed to achieve 50% formation of MetHb. B. racemosa leaf extract was better than stem extract in protecting and delaying the oxidation of Hb to MetHb and was especially protective at concentrations above 100 μg/ml whereas B. racemosa stem extract was protective at concentrations above 500 μg/ml. A similar observation was also reported by Sulaiman and Hussain (2011), whereby Hb treated with anthocyanin delayed the formation of MetHb.