LC–HRMS and Chemical Derivatization Strategies for the Structure Elucidation of Caribbean Ciguatoxins: Identification of C-CTX-3 and -4

Abstract

Ciguatera poisoning is linked to the ingestion of seafood that is contaminated with ciguatoxins (CTXs). The structural variability of these polyether toxins in nature remains poorly understood due to the low concentrations present even in highly toxic fish, which makes isolation and chemical characterization difficult. We studied the mass spectrometric fragmentation of Caribbean CTXs, i.e., the epimers C-CTX-1 and -2 (1 and 2), using a sensitive UHPLC–HRMS/MS approach in order to identify product ions of diagnostic value. We found that the fragmentation of the ladder-frame backbone follows a characteristic pattern and propose a generalized nomenclature for the ions formed. These data were applied to the structural characterization of a pair of so far poorly characterized isomers, C-CTX-3 and -4 (3 and 4), which we found to be reduced at C-56 relative to 1 and 2. Furthermore, we tested and applied reduction and oxidation reactions, monitored by LC–HRMS, in order to confirm the structures of 3 and 4. Reduction of 1 and 2 with NaBH4 afforded 3 and 4, thereby unambiguously confirming the identities of 3 and 4. In summary, this work provides a foundation for mass spectrometry-based characterization of new C-CTXs, including a suite of simple chemical reactions to assist the examination of structural modifications.

Liquid Chromatography Coupled to High-Resolution Mass Spectrometry for the Confirmation of Caribbean Ciguatoxin-1 as the Main Toxin Responsible for Ciguatera Poisoning Caused by Fish from European Atlantic Coasts

“…In HRMS spectra ( Figure 1A Figure 1B and Figure S1, Table S1). As with CTX3C and CTX1B [17], the opening of the G-and H-rings allowed the formation of the ion at m/z 563.3220 followed by water losses at m/z 545.3115, m/z 527.3009, and m/z 509.2903, these fragments were also detected in [20] ( Figure 1B and Figure S1, Table S1). Table S1).…”

“…Despite this, the applicability of HRMS has been demonstrated in particular for CTX1B and CTX3C as the main toxins responsible of CP in the Pacific Ocean, the fragmentation pathways have been identified, increasing the knowledge on how these complex toxins fragment in the mass spectrometer [17][18][19]. Recent studies published by Kryuchkov et al [20] reported strategies for the structure elucidation of Caribbean ciguatoxin analogues (C-CTX3 and C-CTX4).…”

Effects of pH and Nutrients (Nitrogen) on Growth and Toxin Profile of the Ciguatera-Causing Dinoflagellate Gambierdiscus polynesiensis (Dinophyceae)

“…Indeed, numerous studies have shown the ability of Gambierdiscus to produce a wide variety of P-CTX analogs as well as their bioaccumulation at each trophic level, from herbivorous to carnivorous fish or marine invertebrates [28,92]. However, despite the identification of several C-CTX and I-CTX analogs in carnivorous fish and sharks [93][94][95][96][97][98][99][100], no study has yet been able to demonstrate the toxin production of C-CTXs or I-CTXs in Gambierdiscus.…”

Stereoselective Synthesis of the I–L Fragment of the Pacific Ciguatoxins

“…Pacific ciguatoxins are the most numerous and well-characterised of the ciguatoxins and this group accounts for more than three quarters of the ciguatoxins that are known [4][5][6]. Four Caribbean ciguatoxins have been characterized-C-CTX-1 and C-CTX-2 are equilibrating lactols and C-CTX-3 and C-CTX-4 are diastereomers that arise by reduction of C-CTX-1/C-CTX-2 [7,8]. A further eight Caribbean ciguatoxins have been detected by mass spectrometry but have yet to be characterised and the structures of the six ciguatoxins-I-CTX-1-6-isolated from samples collected in the Indian Ocean have yet to be elucidated [6,9,10].…”