Ligustrum japonicum is distributed along the feet of the mountains adjacent to the coasts of Korea, China and Japan. L. japonicum is commonly used as an herbal medicine to strengthen the function of the heart and the liver as well as to treat a ringing in the ears, dizziness and eye pain.
Samples of L. japonicum were purchased from ‘Chungmyungyakcho,’ the pharmaceutical company specializing in herbal medicine. Dried samples were extracted twice: with methylene chloride and with methanol (MeOH), respectively. The combined crude extracts were evaporated in vacuo, and then the residue was partitioned between water and methylene chloride. The aqueous layer was re-partitioned between H2Oandn-butanol (n-BUOH), and then the organic layer (i.e. between 85% aqueous methanol (85% aq. MeOH) and n-hexane).
The crude extract and its solvent-partitioned fractions were evaluated for their antioxidant and antiproliferative effects. In the DPPH radical assay system, only the n-BuOH fraction showed a significant radical scavenging effect. In the peroxynitrite assay system, the 85% aq.MeOH and the n-BuOH fractions showed a strong scavenging effect on both authentic peroxynitrite and peroxynitrite induced from SIN-1. In a cellular system using 2'',7''-dichlorofluorescin diacetate (DCF-DA) as the fluorescence probe in HT-1080 cells, all tested samples remarkably decreased the level of intracellular reactive oxygen species (ROS) at the concentration of 100 μg/ml. Among them, the 85% aq.MeOH fraction revealed the strongest scavenging effect.
In the cytotoxicity bioassay system using the MTT reduction method, on the other hand, the crude extract showed a weak cytotoxic effect on all human cancer cells. For the solvent-partitioned fractions, however, the 85% aq.MeOH fraction showed a potent inhibitory effect on the growth of all human cancer cells ( HT-1080, AGS, HT-29, and MCF-7). The n-BuOH fraction exhibited a good inhibition effect on the growth of AGS cells at the concentration of 100 μg/ml.
Six known compounds were isolated from L. japonicum: Oleanolic acid (1), Maslinic acid (2), Ursolic acid (3), Tyrosol (4), Ligustruoside (5), and 8-(E)-Nuezhenide (6). These six compounds were also evaluated for their antioxidant and antiproliferative effects. In the antioxidant bioassay, compound 6 not only showed a potent scavenging effect on peroxinitrites but also remarkably decreased the level of intracellular reactive oxygen species (ROS) in HT 1080 cells. In the cytotoxicity test, compound 3 of all tested compounds exhibited the strongest inhibitory effect on growth of HT-1080, AGS, HT-29, and MCF-7 cells.
Therefore, these results suggest that L. japonicum may be useful as a potential biomaterial, as a natural antioxidant to alleviate oxygen-induced damage as well as a chemopreventive agent for cancer.