RESUMO
Anatoxin-a, a neurotoxin produced by blue-green algae (BGA) species, can cause death to exposed organisms. In North America, BGA are harvested and sold as food supplements, some of which contain elevated levels of other algal toxins, such as microcystins. Concern that elevated levels of anatoxin-a also may be present in BGA food supplements has led to the development of a simple method to determine the presence of anatoxin-a in BGA. Some researchers have successfully analyzed this compound using liquid chromatography with fluorescence detection by forming a fluorescent derivative with 4-fluoro-7-nitrobenzofurazan (NBD-F) in water and phytoplankton extracts. With this method, the background noise is high in BGA extracts due to the presence of co-extractives. Addition of o-phthaldialdehyde (OPA) and mercaptoethanol to the extract before addition of the NBD-F resulted in the successful removal of primary amines from the background noise when the NBD-F derivatives were detected with fluorescence. Improved chromatograms were obtained when extracts were cleaned up in this manner, leading to a lower detection limit (approximately 50 microg/kg) for anatoxin-a. The detection limits obtained for the 2 degradation products dihydroanatoxin-a and epoxyanatoxin-a in BGA extracts were similarly low (55 and 65 microg/kg, respectively).
Assuntos
Toxinas Bacterianas/análise , Técnicas de Química Analítica/métodos , Cromatografia Líquida/métodos , Cianobactérias/metabolismo , Toxinas Marinhas/análise , 4-Cloro-7-nitrobenzofurazano/análogos & derivados , 4-Cloro-7-nitrobenzofurazano/análise , Aminas/química , Proteínas de Bactérias/análise , Toxinas Bacterianas/química , Cromatografia , Toxinas de Cianobactérias , Fluorescência , Toxinas Marinhas/química , Espectrometria de Massas , Mercaptoetanol/análise , Mercaptoetanol/química , Microcistinas , Fosfatos/química , Fitoplâncton/metabolismo , Spirulina , Fatores de Tempo , Tropanos , o-Ftalaldeído/análiseRESUMO
A colorimetric method and a capillary electrophoresis procedure were developed for quantifying histidyl-leucine and hippurate, respectively. The colorimetric method is sensitive (extinction coefficient = 7.5 mM(-1) cm(-1)) and reproducible (CV = 1.7%, n = 5), which is based on a selective chromogenic reaction for histidyl-leucine (lambda(max) = 390 nm) using o-phthalaldehyde. For samples containing unusually high levels of histidine and/or histidyl peptides, the separation-based approach is preferable. The capillary electrophoresis method makes use of an in-capillary microextraction technique; complicated samples can be measured in less than 4 min without pretreatment. Protocols using both methods to measure angiotensin converting enzyme inhibitory activity were proposed.