INVESTIGADORES
PATRIARCA Andrea Rosana
congresos y reuniones científicas
Título:
UV-MALDI mass spectrometry analysis of AAL toxin TA
Autor/es:
PATRIARCA, A.; ERRA-BALSELLS, R.; GIUDICESSI, S.; TERMINIELLO, L.; VAQUERA, S.; FERNÁNDEZ PINTO, V.
Lugar:
Mendoza
Reunión:
Conferencia; MycoRed ISM conference; 2011
Institución organizadora:
International Society for Mycotoxicology
Resumen:
AAL
toxins belong to the sphinganine-analog mycotoxins (SAMT) due to their
structural similarity to sphinganine. SAMT execute their toxicity
through the competitive inhibiton of sphinganine N-acetyltransferase. The inhibition of this enzyme leads to various diseases in animals and humans. AAL toxins are produced by Alternaria arborescens and are the causal agent of stem-canker disease in tomatos.
Assessment of toxicological properties and health risks posed by AAL
toxins as well as detailed investigations into their biosynthesis
require appropriate methods for screening and structural confirmation. The
ultraviolet matrix-assisted laser/desorption ionization time of flight
mass spectrometry (UV-MALDI-TOF MS) instrumentation is a technique
that enables a highly sensitive and rapid analysis of complex native mixtures obtained from cells and tissues. The objective of this work is to determine if UV-MALDI-TOF MS is an appropriate method for screening and confirmation of AAL toxins in fungal cultures. A representative culture of A. arborescens EGS 39-128 and two native A. arborescens strains isolated from tomato ITEM 8162 and ITEM 8134, were used. For mycotoxin production, the strains were inoculated in autoclaved rice and incubated at 25ºC for 21 days. The extraction and quantification of AAL toxins were made according to Solfrizzo et al 2005. Samples of AAL TA standard and products extracted from the fungal cultures were analyzed by UV-MALDI-MS performed on the Ultraflex II TOF/TOF Bruker Daltonics mass spectrometer.
Matrix selection was performed and the best results were obtained with
9H-[3,4-b]piridoindole (nHo; norharmane) in negative ion mode and 2,5-dihydroxibenzoic acid (DHB) in a positive ion mode. ESI-MS analyses were performed using a Bruker micrOTOF-Q II mass spectrometer.The UV-MALDI-TOF spectrum of the AAL TA standard revealed a molecular ion [M + H]+ at m/z 522, whereas the adducts [M + Na]+ at m/z 544 and [M + K]+ at m/z 560 were also observed with high signal intensity. The same peaks were observed in ESI spectrum. The main fragments of AAL TA showed high intensity and a reproducible pattern: [M + H ? H2O]+ at m/z 504, [M + H ? 2 H2O]+ at m/z 486, [M + H ? TCA]+ at m/z 346, [M + H ? TCA ? H2O]+ at m/z 328, [M + H ? TCA ? 2 H2O]+ at m/z 310, [M + H ? TCA ? 3 H2O]+ at m/z 292, [M + H ? TCA ? 4 H2O]+ at m/z 274. EGS 39-128 and ITEM 8162 cultures of A. arborescens
yielded a MALDI-TOF and ESI spectra similar to that observed for the
AAL TA standard. For the native culture ITEM 8134 no one of the
characteristic ions was observed on both the UV-MALDI-TOF and ESI spectra. It could be concluded from the present results that UV-MALDI-TOF
is an adequate method for screening and confirmation of AAL toxins
from fungal culture extracts. It represents a better option to
chromatographic techniques that require complex and laborious
derivatization of the toxins and it would be a good complement to
HPLC-ESI-MS/MS methods.