A new alkaloid containing a nonbrominated pyrrole and a guanidine moiety, clathrodin, has been isolated from the MeOH extract of the Caribbean sea sponge Agelas clathrodes. The structure of clathrodin was determined to be 1 on the basis of its spectral data. Several C1-N5 compounds containing a bromopyrrole and a guanidine moiety have been isolated from marine sponges belonging to the families Agelasidae and Axinellidae (1-11). However, not many related nonbrominated compounds are known from these families (12). During a recent expedition in search of physiologically active substances of marine organisms from Puerto Rico, we have examined the antimicrobial and cytotoxic actions of an MeOH extract of a sea sponge Agelas clathrodes (Schmitt) which was collected near Desecheo Island, Puerto Rico in March 1989. The material from the MeOH extract showed weak activity against Proteus vulgaris, Staphylococcus aureus, and Shigella fixneri (MIC = 1 mg/ml) using the standard disk assay. Similarly, the cytotoxicity assays inhibited the growth of SW-480 cells (ED50 = 5.3 μg/ml). We report here the isolation and the structure elucidation of the cytotoxic constituent clathrodin [1]. The MeOH extract of the sponge was suspended in H2O and extracted with CHCl3 (3×250 ml) and n-BuOH (2×250 ml), successively. After concentration, the n-BuOH-soluble portion (3.30 g) was chromatographed on a reversed-phase (C18, 20 g) column with H2O followed by a Si gel column (48 g) with CHCl3-MeOH (4:1) saturated with NH3. Combination of like fractions on the basis of tlc analyses gave pure clathrodin [1] as a colorless semisolid (840 mg). The hrfabms of 1 showed an intense (M + H)+ peak at m/z 232.1198, indicating a molecular formula of C11H14N5O (Δ 0.3 mmu) for the compound. The spectral data recorded indicated that clathrodin [1] was the 3-debromo derivative of hymenidin [2], a potent antiserotonergic constituent of the Okinawan marine sponge Hymeniacidon sp. (7). The 1H-nmr spectrum (Table 1) and the 1H-1H COSY spectrum revealed the partial structure -CO-NH-CH2-CH=CH- (trans). The signals for two aromatic protons at δ 6.96 (H-2 and H-4, overlapping multiplets) and another at δ 6.12 (H-3, complex multiplet) indicated the existence of a monosubstituted pyrrole ring (9). The connection of the pyrrole ring to the amide carbonyl at C-5 was argued on the presence of an intense cross peak between H-4 and H-7 in the 2D nOe spectrum (13,14). An aminoimidazole unit was suggested by the 13C-nmr signals at δ 127.35 (C-11), 151.79 (C-13), and 117.52 (C-15). Furthermore, the 13C chemical shift values closely resembled those reported for hymenidin [2] (7) and oroidin [3] (1,2) and thus confirmed the structure 1 for clathrodin. Clathrodin [1] is biogenetically related to bromine-containing antimicrobial alkaloids keramadine (5), oroidin [3] (2), sceptrin (3), and dibromoagelaspongin (9), which have been isolated from the same genus Agelas. Other related compounds reported are mono- and dibromophakelin from Phakellia flabellata (10), hymenialdisine from Hymeniacidon aldis (4,11) and hymenin and hymenidin [2] from Hymeniacidon sp (6-8).