Duocarmycin A, C_1 and C_2 produced by Streptomyces sp. are novel antitumor antibiotics which are effective against murine lymphocytic leukemia p388 and murine Sarcoma 180 in mice. In this report, we describe the structural determination of the Duocarmycins (Fig. 1). The ^1H NMR spectrum of Duocarmycin C_1 (1) revealed the presence of three aromatic protons, four methoxyl groups, one tert. methyl group, two methylene groups, one methine proton and three protons which disappeared in the presence of D_2O (Table 2). The observation of ^1H-^<13>C long range couplings through COLOC spectrum of 1 exhibited three partial structures (I, II and III) shown in Fig. 2. The attachment of the amino group between C-2 and C-9a, and of the phenolic OH group to C-9 were also established by COLOC experiment of a permethylate 4, where the long range couplings from N-methyl and O-methyl protons were observed (Fig. 3). Treatment of 1 with triethylamine in methanol at room temperature in the presence of Ag_2CO_3 produced a quinoline derivative 6, indicating the presence of nitrogen linked to C-6 and C-7a. The attachment of the chlorine to C-5 was confirmed by alkaline treatment of 4 producing a dehydrochloride 5. The partial structure III was defined by isolation of the alkaline degradation product 7. The observation of NOE between 1-NH and 9-OH confirmed the attachment of N-1 to C-9a and C-3 to C-3a in the partial structure I and II. The long range couplings between 6-H_2 and C-2'α determined by LSPD experiment established the connection of C-2'α to N-7, thus confirming the whole structure of 1 as shown in Fig. 1. The structure of Duocarmycin C_2 (2) is different from that of 1 with respect to the chlorine-containing moiety. The ^1H NMR spectrum showed that chloromethyl protons were coupled to a methine, which in tern was coupled to a nitrogen-bearing methylene. This indicated that 2 had a dihydroindole skeleton on the corresponding partial structure II of 1. The structural differences between Duocarmycin A (3) and Duocarmycin C_1 are also related to the partial structure II. The COLOC spectrum revealed the structure of 3 having dienone conjugated with the cyclopropane ring (Fig. 4). Treatment of 3 with hydrochloric acid in acetone gave 1 and 2 in the ratio ca. 1:4 (Fig. 5). This provided the evidence for the structures of Duocarmycins and meant that 1 and 2 seemed to be artifact products of 3.
