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Toxins 2014, 6(2), 402-415; doi:10.3390/toxins6020402
Article

Detection of Anatoxin-a and Three Analogs in Anabaena spp. Cultures: New Fluorescence Polarization Assay and Toxin Profile by LC-MS/MS

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Received: 4 November 2013; in revised form: 23 December 2013 / Accepted: 9 January 2014 / Published: 24 January 2014
(This article belongs to the collection Marine and Freshwater Toxins)
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Abstract: Anatoxin-a (ATX) is a potent neurotoxin produced by several species of Anabaena spp. Cyanobacteria blooms around the world have been increasing in recent years; therefore, it is urgent to develop sensitive techniques that unequivocally confirm the presence of these toxins in fresh water and cyanobacterial samples. In addition, the identification of different ATX analogues is essential to later determine its toxicity. In this paper we designed a fluorescent polarization (FP) method to detect ATXs in water samples. A nicotinic acetylcholine receptor (nAChR) labeled with a fluorescein derivative was used to develop this assay. Data showed a direct relationship between the amount of toxin in a sample and the changes in the polarization degree of the emitted light by the labeled nAChR, indicating an interaction between the two molecules. This method was used to measure the amount of ATX in three Anabaena spp. cultures. Results indicate that it is a good method to show ATXs presence in algal samples. In order to check the toxin profile of Anabaena cultures a LC-MS/MS method was also developed. Within this new method, ATX-a, retention time (RT) 5 min, and three other molecules with a mass m/z 180.1 eluting at 4.14 min, 5.90 min and 7.14 min with MS/MS spectra characteristic of ATX toxin group not previously identified were detected in the Anabaena spp. cultures. These ATX analogues may have an important role in the toxicity of the sample.
Keywords: anatoxin-a; nicotinic acetylcholine receptor; fluorescence polarization; liquid chromatography-mass spectrometry; Anabaena spp. anatoxin-a; nicotinic acetylcholine receptor; fluorescence polarization; liquid chromatography-mass spectrometry; Anabaena spp.
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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MDPI and ACS Style

Sanchez, J.A.; Otero, P.; Alfonso, A.; Ramos, V.; Vasconcelos, V.; Aráoz, R.; Molgó, J.; Vieytes, M.R.; Botana, L.M. Detection of Anatoxin-a and Three Analogs in Anabaena spp. Cultures: New Fluorescence Polarization Assay and Toxin Profile by LC-MS/MS. Toxins 2014, 6, 402-415.

AMA Style

Sanchez JA, Otero P, Alfonso A, Ramos V, Vasconcelos V, Aráoz R, Molgó J, Vieytes MR, Botana LM. Detection of Anatoxin-a and Three Analogs in Anabaena spp. Cultures: New Fluorescence Polarization Assay and Toxin Profile by LC-MS/MS. Toxins. 2014; 6(2):402-415.

Chicago/Turabian Style

Sanchez, Jon A.; Otero, Paz; Alfonso, Amparo; Ramos, Vitor; Vasconcelos, Vitor; Aráoz, Romulo; Molgó, Jordi; Vieytes, Mercedes R.; Botana, Luis M. 2014. "Detection of Anatoxin-a and Three Analogs in Anabaena spp. Cultures: New Fluorescence Polarization Assay and Toxin Profile by LC-MS/MS." Toxins 6, no. 2: 402-415.



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