Aflatoxin Screening by MALDI-TOF Mass Spectrometry Rodrigo Ramos Catharino, † Lygia de Azevedo Marques, † Leonardo Silva Santos, † Antonio S. Baptista, ‡ Eduardo M. Glo ´ ria, ‡ Maria A. Calori-Domingues, ‡ Elizete M. P. Facco, § and Marcos N. Eberlin* ,† Thomson Mass Spectrometry Laboratory, Institute of Chemistry, State University of Campinas, UNICAMP, Campinas, SP Brazil, Division of AgroIndustry, Food and Nutrition, ESALQ/USP, Piracicaba, SP, Brazil, and Division of Food Science, Food Engineering, UNICAMP, Campinas, SP, Brazil Efficient detection of aflatoxins B 1 ,B 2 ,G 1 , and G 2 has been performed by matrix-assisted laser desorption/ ionization time-of-flight mass spectrometry using a UV- absorbing ionic liquid matrix to obtain “matrix-free” mass spectra and addition of NaCl to enhance sensitivity via Na + cationization. Using ionic r-cyano-4-hydroxycinnamic acid (Et 3 N-r-CHCA) as the matrix, matrix-free mass spectra in the m/z range of interest are acquired, and the B 1 ,B 2 , G 1 , and G 2 aflatoxins are readily detected with an LOD as low as 50 fmol. The technique is fast, requires little sample preparation and no derivatization or chromato- graphic separation, and seems therefore to be suitable for high-throughput aflatoxin screening. It should be easily extended to other micotoxins and provide an attractive technique to control the quality of major crops subjected to huge world commercial trades such as peanuts, corn, and rice as well as to monitor bioterrorism threats by micotoxin poisoning. The broad success of matrix-assisted laser desorption/ioniza- tion (MALDI) 1 is related mainly to its ability to ionize relatively heavy or polar molecules such as peptides and proteins. 2 In MALDI, the analyte is co-crystallized with a solid ultraviolet- absorbing organic acid matrix that vaporizes upon laser radiation, carrying with it the analyte. 3 MALDI is not so widely applicable, however, to relatively light molecules owing to severe chemical noise from matrix ions in the low m/z range. Matrix-free laser desorption/ionization has been applied, but its use is restricted owing to rapid analyte degradation due to direct exposure to laser radiation. 4 An interesting approach used in MALDI to minimize matrix interference is to use a UV-absorbing ionic liquid as the matrix. 5,6 Ionic liquids are easily prepared and have many beneficial properties for MALDI, including the broad ability to dissolve organic, inorganic, and polymeric substances, good thermal stability, and low vapor pressures. The pioneering study of Gross and co-workers 6 demonstrated the suitability of ionic liquids as MALDI matrixes. We have also applied ionic liquids as the MALDI matrix for rapid screening of low molecular weight chemicals by thin-layer chromatography and “on-spot” MALDI- TOF MS analysis. 5 Aflatoxins (Chart 1) are carcinogenic mycotoxins that can be ingested via contaminated food or inhaled via dust from mold- infected products. 7 Mycotoxin poisoning is more serious in developing countries where the climatic conditions and the agricultural and storage practices facilitate fungal growth and toxin production. Peanuts (and their products) is the crop most susceptible to aflatoxin contamination. 8 Herein we report that high- throughput MALDI-TOF MS screening of low molecular weight aflatoxins is efficiently performed by using a UV-absorbing ionic liquid as the matrix to nearly eliminate matrix noise and by adding NaCl for enhanced ionization via Na + cationization. EXPERIMENTAL SECTION The Et 3 N-R-cyano-4-hydroxycinnamic acid (R-CHCA) ionic liquid matrix was prepared by adding triethylamine to a solution of R-CHCA in acetonitrile, with subsequent reflux for 1 h, and by * Corresponding author. Phone/Fax: (55-019)3788-3023. E-mail: eberlin@ iqm.unicamp.br. † State University of Campinas. ‡ ESALQ/USP. § UNICANP. (1) Karas, M.; Hillenkamp, F. Anal. Chem. 1988, 60, 2299. (2) (a) Macfarlane, R. D.; Torgerson, D. F. Science 1976, 191, 920. (b) Barber, M.; Bordoli, R. S.; Sedwick, R. D.; Tyler, A. N. Nature 1981, 293, 270. (3) Zenobi, R.; Knochenmuss, R. Mass Spectrom. Rev. 1998, 17, 337. (4) Zenobi, R. Chimia 1997, 51, 801. (5) Santos, L. S.; Haddad, R.; Ho ¨ehr, N. F.; Pilli, R. A.; Eberlin, M. N. Anal. Chem. 2004, 76, 2144. (6) Armstrong, D. W.; Zhang, L.-K.; He, L. F.; Gross, M. L. Anal. Chem. 2001, 73, 3679. (7) Maurer, H. H. J. Chromatogr., B 1998, 713, 3. (8) Aycicek, H.; Aksoy, A.; Saygi, S. Food Control. 2005, 16, 263. Chart 1 Anal. Chem. 2005, 77, 8155-8157 10.1021/ac051111p CCC: $30.25 © 2005 American Chemical Society Analytical Chemistry, Vol. 77, No. 24, December 15, 2005 8155 Published on Web 11/08/2005