P-13 有毒ラン藻,Anabaena flos-aquae NRC 525-17がanatoxin-a(s)およびmicrocystinとともに生産する新規環状ペプチド(ポスター発表の部)

抄録

It is well known that cyanobacteria produce several toxic compounds. Microcystins, nodularin and cylindrospermopsin are known as the lethal hepatotoxins and the neurotoxins, anatoxin-a, anatoxin-a(s) and aphantoxins (saxitoxins) are also produced by some species of cyanobacteria. One strain of Anabaena flos-aquae called NRC 525-17 is a unique cyanobacterium that can simultaneously produce anatoxin-a(s) and microcystins. In a previous report we found that two unidentified peaks were detected by LC/MS along with those of microcystins in the toxic fraction from the cyanobacterium. The two compounds were purified by repeated silica gel and TOYOPEARL HW-40F chromatographies and designated as anabaenopeptins A (1) and B (2). Usual acid hydrolysis of 1 and 2 gave four amino acids, Val, Phe, Hty and MeAla having the L configuration, as determined by Marfey's method combimned with Frit-FAB or ESI LS/MS and confirmed by direct comparison with authentic samples. Application of the technique composed of Marfey's method and ESI LC/MS provided an additional advantage that it can sensibly detect trace amounts of the remaning constituent amino acids, Lys and Tyr for 1 and Lys and Arg for 2 in the hydrolysates. Further, the technique indicated clearly that 1 possesses L-Tyr and D-Lys and 2 possesses L-Arg and D-Lys. The common cyclic pentapeptide moiety, cyclo-(Lys-Val-Hty-MeAla-Phe), for 1 and 2 was deduced by 2D NMR techniques such as ^1H H^1COSY, HSQC, HMBC and NOESY. The results of the HMBC correlations and NOE correlations suggested that Arg is bound with α-NH of Lys through an ureido linkage in 2 and that 1 also has an ureido linkage between Lys and Tyr (Fig. I). Both 1 and 2 at 10-400μg/ml produced concentration-dependent relaxations (2-57% of norepinephrine-induced contraction) in rat aortic preparations with endothelium precontracted with norepinephrine. Recently, Kobayashi et al. reported the isolation and structural determination of keramamide A and konbamide from Okinawan marine sponges and the proposed structures for 1 and 2 have a resemblance to those for these two compounds. Since they also suggested that these cyclic peptides are produced by symbiotic microorganisms, the present case provides additional evidence that marine natural products are also produced by terrestrial microorganisms.