Acute Toxicity of Gambierone and Quantitative Analysis of Gambierones Produced by Cohabitating Benthic Dinoflagellates

Abstract

Understanding the toxicity and production rates of the various secondary metabolites produced by Gambierdiscus and cohabitating benthic dinoflagellates is essential to unravelling the complexities associated with ciguatera poisoning. In the present study, a sulphated cyclic polyether, gambierone, was purified from Gambierdiscus cheloniae CAWD232 and its acute toxicity was determined using intraperitoneal injection into mice. It was shown to be of low toxicity with an LD₅₀ of 2.4 mg/kg, 9600 times less toxic than the commonly implicated Pacific ciguatoxin-1B, indicating it is unlikely to play a role in ciguatera poisoning. In addition, the production of gambierone and 44-methylgambierone was assessed from 20 isolates of ten Gambierdiscus, two Coolia and two Fukuyoa species using quantitative liquid chromatography-tandem mass spectrometry. Gambierone was produced by seven Gambierdiscus species, ranging from 1 to 87 pg/cell, and one species from each of the genera Coolia and Fukuyoa, ranging from 2 to 17 pg/cell. The production of 44-methylgambierone ranged from 5 to 270 pg/cell and was ubiquitous to all Gambierdiscus species tested, as well as both species of Coolia and Fukuyoa. The relative production ratio of these two secondary metabolites revealed that only two species produced more gambierone, G. carpenteri CAWD237 and G. cheloniae CAWD232. This represents the first report of gambierone acute toxicity and production by these cohabitating benthic dinoflagellate species. While these results demonstrate that gambierones are unlikely to pose a risk to human health, further research is required to understand if they bioaccumulate in the marine food web.

Keywords: Coolia; Fukuyoa; Gambierdiscus; LD50; ciguatera poisoning; liquid chromatography–tandem mass spectrometry; nuclear magnetic resonance spectroscopy.

Figure 1

Structures of gambierone and 44-methylgambierone (monoisotopic masses 1024.5 and 1038.5 g/mol, respectively).

Figure 2

Full scan mass spectra of gambierone (m/z 850–1150) in (A) −ESI mode showing the [M–H]⁻ ion (m/z 1023.3) and (B) +ESI mode showing the [M+H]⁺ (m/z 1025.3), [M–H₂O+H]⁺ (m/z 1007.3), [M–2H₂O +H]⁺ (m/z 989.3) and [M–SO₃+H]⁺ (m/z 945.3) ions, as well as a series of sulphite plus sequential water-loss ions (m/z 927.3, 909.3, 891.3, 873.3).

Figure 3

Collision-induced dissociation tandem MS spectra (m/z 50–1050) of purified gambierone generated from (A) the [M–H]⁻ parent ion (m/z 1023.3) in –ESI mode, collision energy 70 eV, showing a single dominant fragment ion representing the bisulphate anion (m/z 96.8) and (B) the [M+H]⁺ parent ion (m/z 1025.3) in +ESI mode, collision energy 25 eV, showing a variety of unassigned fragment ions.