Determination of anatoxin-a in environmental water samples by solid-phase microextraction and gas chromatography-mass spectrometry

Abstract

Anatoxin-a is a neurotoxin produced by various species of cyanobacteria such Anabaena, Planktothrix (formerly Oscillatoria), Aphanizomenon, Cylindrospermum, and Microcystis spp. This toxin acts as a mimic of the neurotransmitter acetylcholine and irreversibly binds the nicotinic acetylcholine receptor (NAChR) therefore affecting the functioning of the nervous system. Experimental studies have shown that in few minutes after drinking contaminated water, animals presented typical symptoms of anatoxin-a intoxication such as muscle fasciculation, gasping, violent convulsions, and death due to respiratory arrest. The estimated dosage needed to produce death in 50% of exposed mice (LD50) after interperitoneal administration was approximately 200 µg/kg. Anatoxin-a was first detected in Canada (1960s) and since then has been responsible for a number of animal and human fatalities, in different parts of the world. The search for efficient and reliable analytical methods for the control of such cyanotoxins is becoming more and more urgent due to their recent appearance and worldwide distribution. The present work, summarizes the development of an automated method combining Solid phase microextraction (SPME) with Gas Chromatography coupled with Mass Spectrometry (GC-MS) for the fast and sensitive determination of Anatoxin-a in water samples. This method is based on the direct derivatization of the analyte by adding isobutylchoroformate to the sample extract in alkaline conditions. The derivatized anatoxin-a was extracted by SPME procedure, submersing a PDMS fiber in an amber vial for 20 min under magnetic stirring. GC-MS is used to identify and quantify the analyte in the SIM mode (quantification ions underlined): 191,164 and 265. Parameters affected to the extraction such as: salt concentration, time of extraction, time of reaction and stirring speed, were initially evaluated. The results obtained in this study let us to conclude that this method is an efficient and fast alternative for the reliable control of anatoxin-a present in contaminated waters.