Chem. Pharm. Bull. 53(7) 788—791 (2005)

sphingolipids (GSLs) from echinoderms, a series of studies on the isolation and structural elucidation of the GSLs from sea cucumber species have been performed in our laboratory. In the study of the GSLs of the sea cucumber Holothuria pervicax, we reported the isolation and structure of ten glucocerebrosides revealing a very close resemblance in structure. However, all these compounds still exist as a mixture of regio-isomers for terminal methyl groups in the side chain of the long-chain base (LCB) moiety, namely a mixture of iso and ante-iso isomers as shown in Fig. 1. Continuing the previous studies, the separation of the mixture of the regio-isomers was conducted. In this paper, we report on the isolation of an ante-iso type regio-isomer of glucocerebroside from its parent glucocerebroside molecular species HLC-2, an abundant compound obtained from the whole bodies of the sea cucumber Holothuria leucospilota (Nisekuronamako in Japanese). The less polar lipid fraction, which was obtained from the chloroform/methanol extract of the whole bodies of H. leucospilota, was subjected to repeated silica gel column chromatography to give three cerebroside molecular species, HLC-1, HLC-2, and HLC-3, each showing a single spot on silica gel thin-layer chromatography (TLC). In the IR and positive-ion FAB mass spectra of HLC-1, HLC-2, and HLC-3, strong hydroxy and amide absorptions and a series of molecular ion peaks are observed. Their CNMR spectra (Fig. 1, Table 1) exhibit the characteristic signals of a sphingosine-type b-glucocerebroside possessing an unsubstituted fatty acid (HLC-1), a sphingosine-type b-glucocerebroside possessing a 2-hydroxy fatty acid (HLC-2), and a phytosphingosine-type b-glucocerebroside possessing a 2-hydroxy fatty acid (HLC-3), respectively. Therefore, they are suggested to be the molecular species of three typical types of glucocerebrosides. Their structures shown in Fig. 1 were characterized by comparison of their C-NMR spectral data with those of known glucocerebrosides hitherto obtained, and by means of the results of their chemical degradations, namely methanolysis followed by the GC-MS analysis of the methanolysis products, fatty acid methyl ester (FAM), and LCB, as shown in Fig. 2 and Experimental. The geometry (Z) of the double bond in the unsaturated fatty acyl moiety of HLC-1—HLC-3 was determined from the d value (27.4, 27.5) of the allylic carbon atoms obtained from their H-detected heteronuclear multiple-bond connectivity (HMBC) spectra (Fig. 3), since allylic carbon signals of Z788 Chem. Pharm. Bull. 53(7) 788—791 (2005) Vol. 53, No. 7