Structure Elucidation of Fucan Sulfate from Sea Cucumber Holothuria fuscopunctata through a Bottom-Up Strategy and the Antioxidant Activity Analysis

Fucan sulfate I (FSI) from the sea cucumber Holothuria fuscopunctata was purified and its structure was clarified based on a bottom-up strategy. The unambiguous structures of a series of oligosaccharides including disaccharides, trisaccharides, and tetrasaccharides, which were released from mild acid hydrolysis of FSI, were identified by one-dimensional (1D)/two-dimensional (2D) nuclear magnetic resonance (NMR) and mass spectrometry (MS) analysis. All the glycosidic bonds in these oligosaccharides were presented as α1,3 linkages confirmed by correlated signals from their (1)H-(1)H ROESY and (1)H-(13)C HMBC spectra. The structural sequence of these oligosaccharides formed by Fuc(2S4S), Fuc(2S), and non-sulfated ones (Fuc(0S)), along with the general structural information of FSI, indicated that the structure of FSI could be elucidated as: [-L-Fuc(2S4S)-α1,3-L-Fuc((2S))-α1,3-L-Fuc(2S)-α1,3-L-Fuc(0S)-α1,3-1-](n). Moreover, the L-Fuc(0S-)α1,3-L-Fuc(2S4S) linkage in FSI was susceptible to be cleaved by mild acid hydrolysis. The antioxidant activity assays in vitro showed that FSI and the depolymerized product (dFSI′) had potent activities for superoxide radical scavenging activity with IC(50) of 65.71 and 83.72 μg/mL, respectively, while there was no scavenging effect on DPPH, hydroxyl and ABTS radicals.