Xenocoumacin-1 (XCN-1) and Xenocoumacin-2 (XCN-2) are reported antibiotics produced by the mutualistic bacteria Xenorhabdus nematophila. XCN-1 was isolated from the OmpR deletion strain of X. nematophila (ABR2), which preferentially produces XCN-1. Studies of XCN-1 via HPLC/MS/MS indicated the possible formation of an adduct with benzylideneacetone (BZA) at the dihydroxyamine moiety found in XCN-1. We propose that this adduct forms through Michael addition of XCN-1 to BZA. To support this hypothesis, BZA was incubated at room temperature with three amine diols: 3-amino-1,2-propanediol, 2-amino-1,3-propanediol, and N-methyl-3-amino-1,2-propanediol. BZA spontaneously reacted with 3-amino-1,2-propanediol, forming N-(3-oxo-1-phenylbutyl)- 3-amino-1,2-propanediol as verified by UHPLC separation with MS/MS detection. This reaction likely proceeds through the stabilization of the enolate tautomer by the diol (see figure). When BZA was incubated with 2-amino-1,3-propanediol, lacking the adjacent diol, and the more sterically hindered nucleophile, N-methyl-3-amino-1,2-propanediol, the Michael addition did not spontaneously occur. This result provides support for our proposed mechanism of adduct formation, with the adjacent hydroxyl groups on XCN-1 serving to stabilize the tetrahedral intermediate formed during the Michael attack of the amine on the beta carbon of the conjugated ketone in BZA.

Studies of xenocoumacin-1-benzylideneacetone Michael adduct formation

253rd National Meeting (2017)

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