Mass spectrometric studies shed light on unusual oxidative transformations of 1,2-dehydro-N-acetyldopa Adal Abebe 1† , Qun F. Kuang 2† , Jason J. Evans 2 and Manickam Sugumaran 1 * 1 Department of Biology, University of Massachusetts Boston, Boston, MA 02125, USA 2 Department of Chemistry, University of Massachusetts Boston, Boston, MA 02125, USA RATIONALE: Lamellarins are a group of over 70 plus bioactive marine natural compounds possessing a 6,7- dihydroxycoumarin moiety. Although they appear to derive from 3,4-dihydroxyphenylalanine (dopa), practically nothing is known about the metabolic fate of these compounds. Biochemical considerations indicate that they could arise from a N-acetyl-1,2-dehydrodopa precursor through oxidative cyclization reaction. METHODS: To assess the above hypothesis, we synthesized N-acetyl-1,2-dehydrodopa and conducted oxidation studies with commercially available mushroom tyrosinase and evaluated the course of the reaction with reversed-phase liquid chromatography/mass spectrometry (LC/MS). RESULTS: Mushroom tyrosinase readily oxidized N-acetyl-1,2-dehydrodopa – not to the normally expected quinone – but to an unstable quinone methide isomer, which rapidly cyclized to produce the dihydroxycoumarin product, 3-aminoacetyl esculetin. Interestingly, 3-aminoacetyl esculetin was further oxidized to a second quinone methide derivative that exhibited an addition reaction with the parent dihydroxycoumarin generating dimeric and other oligomeric products in the reaction mixture. CONCLUSIONS: LC/MS analysis of the N-acetyl-1,2-dehydrodopa oxidation reaction reveals not only a possible novel oxidative cyclization route for the biosynthesis of coumarin-type dehydrodopa compounds in marine organisms, but also unusual oxidative transformations of dehydro dopa derivatives. Copyright © 2013 John Wiley & Sons, Ltd. Lamellarins are a group of over 70 different polycyclic condensed aromatic compounds. [1] Faulkner and his group first reported the isolation and characterization of four lamellarins from the prosobranch mollusc, Lamellaria sp., and named them lamellarins A–D. [2] Subsequently, four additional members of this group (E–H) were isolated from the didemnid ascidian Didemnum chartaceum. [3] Since then, as many as 70 different, yet structurally closely related, polycyclic aromatic condensed compounds have been isolated from a variety of marine organisms including, ascidians, sponges and other organisms. [1] These novel compounds possess a wide range of biological activities that include but are not limited to cytotoxicity, antibiotic activity, antitumor activity, antioxidant activity, multi drug resistance reversal activity, HIV integrase inhibition, human aldose reductase inhibition, cell division inhibition, immunomodulatory activity, and feeding deterrent activity. [1,4] The majority of the lamellarins possess either a type 1a or 1b structure as shown in Fig. 1. In addition to lamellarins, ningalin A and ningalin B, whose structures are also shown in Fig. 1, possess such a coumarin ring structure. [5] Ningalins, as well as their derivatives, exhibit marked cytotoxicity against several cancer cell lines. They also exhibit significant multi-drug resistance reversal activity at non-cytotoxic concentrations. [1] Hence, there is a considerable interest in the biochemistry of these compounds. In spite of the vast literature available on the isolation, characterization and biological activities of these interesting metabolites, practically nothing is known about their biosynthetic pathways as well as metabolic fate. Examination of their structure (Fig. 1) indicates that the bulk of these compounds possess a 6,7-dihydroxycoumarin skeleton. [1] Coumarins are of widespread occurrence in the plant kingdom but are rarely found in the animal kingdom. [6] In plants, they are usually biosynthesized from p-coumaryl coenzyme A and/or feruloyl coenzyme A. For example, 7-hydroxy-6- methoxycoumarin (or scopoletin) is biosynthesized from feruloyl coenzyme A by the formation of 6’-hydroxyferuloyl coenzyme A and subsequent isomerization and lactonization to produce scopoletin with the release of coenzyme A (Fig. 2). The coenzyme A seems to be necessary for the activation of the carboxyl group and to prepare it for the lactonization reaction. [6,7] However, occurrence of such a reaction has not been demonstrated in marine organisms. In marine organisms, most coumarins are present as 6,7-dihydroxy derivatives (Fig. 1). An abundance of precursor compounds such as dehydrodopyl and dehydrotyrosyl compounds in marine animals [8] calls for a new look at an alternate mode of biosynthesis of coumarins in these organisms. * Correspondence to: M. Sugumaran, Department of Biology, University of Massachusetts Boston, 100 Morrissey Blvd, Boston, MA 02125, USA. E-mail: manickam.sugumaran@umb.edu † These authors contributed equally to this work. Copyright © 2013 John Wiley & Sons, Ltd. Rapid Commun. Mass Spectrom. 2013, 27, 1785–1793 Research Article Received: 14 February 2013 Revised: 15 May 2013 Accepted: 19 May 2013 Published online in Wiley Online Library Rapid Commun. Mass Spectrom. 2013, 27, 1785–1793 (wileyonlinelibrary.com) DOI: 10.1002/rcm.6630 1785