Reactive oxygen species (ROS) such as hydrogen peroxide, singlet oxygen, superoxide anion radical, hydroxy radical, alkoxy radical, and peroxy radical from the autoxidation process of lipids as well as reactive nitrogen species (RNS) such as peroxynitrite are generated under normal and/or pathological conditions. ROS and RNS are likely to damage several cellular components such as lipids, proteins, nucleic acids, and DNAs through oxidation and nitration process. In addition, these reactive species cause inflammation or legions on various organs and are associated with various degenerative diseases.
As part of our search for novel antioxidant compounds from marine organisms, we have collected the brown alga Sargassum thunbergii along the shore of Busan, which is widely distributed in the southern coastal area of the Korean Pennisula. The collected samples of Sargassum thunbergii were briefly dried under shade and repeatedly extracted for 2 days with a mixture (1:1) of acetone-CH2Cl2 (1.5 L X 2) and MeOH (1.5 L X 2), respectively. The combined crude extracts of Sargassum thunbergii were fractionated into n-hexane, 85% aq. MeOH, n-BuOH, and water fractions.
The antioxidant activities of n-hexane, 85% aq. MeOH, n-BuOH, and water fractions including crude extracts of Sargassum thunbergii were evaluated using 1,1-diphenyl-2-picryl-hydrazyl (DPPH) radical, authentic ONOO-, and ONOO- generated from SIN-1. Scavenging activities of the individual fractions on DPPH increased in the order of 85% aq. MeOH > n-BuOH > H2O > n-hexane, showing 91.9, 50.0, 34.8, and 9.4% in their scavenging ratios at a concentration of 100 ㎍/㎖, respectively. The scavenging activities of these four fractions on authentic ONOO- increased in the order of 85% aq. MeOH > n-BuOH > n-hexane > H2O fractions and were 96.3, 63.5, 45.6, and 41.1% in their scavenging ratios, respectively, at concentration of 50 ㎍/㎖.
On the basis of the results, Further purification of the 85% aq. MeOH fraction with significant scavenging activity on DPPH and authentic ONOO- using C18 reversed-phase column, followed by HPLC, resulted in the isolation of compounds 1-8. In our measurement for evaluating antioxidant effect of compounds 1-5 using DPPH radical, they exhibited significantly high activities with IC50 value of 26, 27, 25, 30, and 31 ㎍/㎖, respectively, compared with BHT (IC50, 32 ㎍/㎖) and L-ascorbic acid (IC50, 11.4 ㎍/㎖). Also, in our measurement for the scavenging/inhibitory activity of compounds 1-5 on authentic/induced ONOO- from SIN-1, the scavenging activities of compounds 1-5 on authentic ONOO- increased in the order of 1 > 3 > 5 > 4 > 2, with scavenging ratios of 90.3, 88.1, 60.0, 57.1, and 50.6% at 12.5 ㎍/㎖ respectively. However, inhibitory activities against the generation of ONOO- from SIN-1 differed, 5 > 2 > 4 > 1 > 3 > at 98.6, 95.8, 90.6, 85.8, and 76.6% in their scavenging ratios, respectively. Scavenging/inhibitory activities of L-ascorbic acid and penicillamine, positive controls, on authentic/induced ONOO- were 98.1 and 90.4%, and 93.5 and 88.2%, respectively. These results suggest that the antioxidative activity in the crude extract of Sargassum thunbergii was partially attributable to these compounds 1-5.