염생식물 함초(Salicornia herbacea)로부터 이차 대사 물질들의 분리와 구조결정

Abstract

In general, extreme environments are defined as being extreme in one or several physical parameters such as temperature, salinity or pressure. However, in these systems the external variables remain relatively stable over time. On the other hand, a marine system with a high variability in physical parameters such as temperature, salinity or radiation can be considered to be extreme one. Tidal flats, salt marshes, and mangrove ecosystems belong to this type of extreme environment on a daily, seasonal and annual basis. Salt marshes and tidal flats experience regular and irregular external influences. Also, organisms in these extreme environments have to adapt by developing specific responses which often include the synthesis of unusual secondary metabolites.

Salicornia herbacea (also known as glasswort) is one of the most salt tolerant plants growing on salt marshes and tidal flats along the western coast of Korea. It has been not only eaten as a food but also used in folk medicine such as constipation, obesity, diabetes, etc. Nevertheless, studies on its secondary metabolites have little been done until now.

In our search for novel compounds from the salt marsh plants, we collected the salt wort S. harbacea from Daebudo, Ansan in Korea. The crude extracts of these specimens were evaluated for its potential to scavenge DPPH radical and peroxynitrite and to immunomodulate both a mouse thymus and spleen cells. On the basis of moderate effects obtained in both bioassay systems, the combined crude extracts of the glasswort were fractionated with n-hexane, 85% aq. MeOH, and n-BuOH. Scavenging activities of the crude extracts and their fractions on DPPH radical increased in the order of n-BuOH > 85% aq. MeOH > crude extract > n-hexane > water, showing 85.2, 70.0, 58.2, 27.7, and 25.7% 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 > water and were 71.5, 70.8, 44.2, and 34.9% in their scavenging ratios, respectively, at concentration of 50 ㎍/㎖ while inhibitory activities against the generation of ONOO- differed, 85% aq. MeOH > n-BuOH > water > n-hexane fractions at 96.2, 90.0, 83.9, and 65.9% in their scavenging ratios, respectively.

Also, in a immunomodulatory test using a mouse thymus and spleen cells, treatment of each of the mouse thymus and spleen cells with the four fractions resulted in a significant extension of cell survival in culture in order of n-BuOH > n-hexane > 85% aq. MeOH > water.

Considering all these results, n-BuOH soluble fraction was selected for further separation. C18 flash column chromatography followed by silica gel column chromatography and repeated reversed-phase HPLC resulted in the isolation of the two new triterpene saponins 1-2 and the two known triterpene saponins 3-4 and a known flavonoid glycoside 5. The structure of the isolated compounds were established by extensive 2D NMR experiments, 1H COSY, TOCSY, ROESY, HSQC, and HMBC, and through comparison with published spectral data.

In our measurement for evaluating scavenging effect on DPPH radical and authentic ONOO- and inhibitory activity of ONOO- generated from SIN-1, compound 5 exhibited potent scavenging activity on the DPPH radical and authentic ONOO- as well as inhibitory activity of induced ONOO- from SIN-1, comparable with that of BHT on DPPH radical at 100 ㎍/㎖ and those of penicillamine and L-ascorbic acid on ONOO- at 50 ㎍/㎖. Compounds 1-3 also revealed significant scavenging and inhibitory effects on authentic ONOO- and ONOO- generated from SIN-1, respectively.