During our studies on bioactive substances from Okinawan marine organisms, we recently examined extracts of the compound tunicate Aplidium multiplicatum, and succeeded in isolating seven novel nucleoside derivatives, shimofuridins A (1)〜G (7), possessing an acylfucopyranoside moiety by careful studies on the HPLC separation of closely related analogs. Shimofuridin A (1), C_<34>H_<44>O_<12>N_4, was a nucleoside consisting of a purine base, two sugar units, and two unsaturated fatty acids, deduced from extensive analysis of the spectral data including several types of 2D NMR spectra (^1H-^1H COSY, HSQC, HMBC, NOESY, and ROESY). The structure of 1 including the stereochemistry of all chiral centers was established by degradation experiments and chiral HPLC and GC analyses. Shimofuridins B (2)〜E (5) were stereoisomers of double bonds of 1, while shimofuridins F (6) and G (7) were homologs of 1 with 2 more carbon units in the second acyl chain. The NMR chemical shift data of unsaturated acyl chain moiety studied in detail here may be helpful to investigate the stereochemistry of these types of unsaturated chain molecules. Shimofuridin A (1) exhibited cytotoxicity against murine lymphoma L1210 cells in vitro (IC_<50>, 9.5μg/mL), and also showed inhibitory activity against protein kinase C (IC_<50>, 20μg/mL).