Evaluation and Characterization of Reaction Products from Ozonated Aflatoxin Contaminated Corn

2005 Annual Report for GS02-015

Project Type: Graduate Student
Funds awarded in 2002: $10,000.00
Projected End Date: 12/31/2006
Region: Southern
State: Louisiana
Graduate Student:
Major Professor:
Joan King
LSU Agricultural Center

Evaluation and Characterization of Reaction Products from Ozonated Aflatoxin Contaminated Corn

Summary

The effect of ozone on the degradation of AFB1 in an aqueous model system was determined. Standard solutions of AFB1 suspended in distilled water were treated with ozone. Dichloromethane extracts and the remaining water solution of the samples were evaluated with 2-D TLC. MALDI-MS analysis was conducted to partially identify reaction products. AFB1 was totally degraded after 60 sec in dichloromethane extracts. Water portion extracts showed the presence of seven compounds unrelated to aflatoxin. The discovery of these compounds produced from aflatoxin is important because they may be formed in ozonated corn and they may have toxic properties.

Objectives/Performance Targets

  1. 1.Fractionation of corn samples.

    2.Evaluation of extracts by TLC and HPLC.

    3.Identification of one or more ozone
    reaction products.

Accomplishments/Milestones

Ozonation of Aqueous Solution of Aflatoxin/s (A Model System)

Five-hundred µL of a standard solution containing 100µg of AFB1 were transferred into replicate scintillation vials and evaporated to dryness. Dry materials were suspended in 10ml distilled water in a vial and treated with 12-13 wt.% ozone from 0 to 60 sec at 10 minute intervals. Dichloromethane extracts of the solutions were dried under nitrogen. Water portions were freeze dried, re-dissolved in methanol, and dried under nitrogen. The extracts were re-diluted with 1 ml methylene chloride and 20 µl of each was spotted into a TLC plate. Extracts were evaluated with single and 2-dimentional TLC using ether:methanol:water and chloroform:acetone as developing solvents. Unkown spots were scrapped off the plate and extracted for MALDI-MS analysis which was conducted to partially identify reaction products with 2,5-Dihydroxybenzoic acid (DHB) as a matrix.

Analysis of dichloromethane extracts showed the presence of AFB1 after ozonation for 50 sec and it was totally degraded after 60 sec. Conversely, analysis of the water portion extracts showed the presence of seven compounds having Rf values of 0, 0.07, 0.14, 0.07, 0.25, 0.5, and 0.39, respectively, after ozonation for 60 sec. In comparison, Rf values for AFB1, B2, G1, and G2 were 0.71, 0.62, 0.56, and 0.46, respectively. Results of the study suggested the formation of polar compounds. Result of MALDI-MS showed presence of compounds that have higher molecular weights than AFB1. Mass spectra of dichloromethane extracts from samples ozonated for 50 and 60 sec showed base peaks with molecular weights of ca 459 and 439, respectively. Extracts from water portions showed based peaks with molecular weights of ca 475 and 494. The discovery of these compounds produced from aflatoxin is important because they may be formed in ozonated corn and they may have toxic properties.

Future Work

The isolation of new compounds from actual treated corn solvent extraction fractions is being done by TLC separation and their identification will follow.

Impacts and Contributions/Outcomes

Impact

The discovery of these compounds produced from aflatoxin is important because they may be formed in ozonated corn and they may have toxic properties.

Collaborators:

Alfredo Prudente

aprude1@lsu.edu
student
LSU Agricultural Center
Department of Food Science
111 Food Science Bldg., Louisiana State University
Baton Rouge, LA 70803
Office Phone: 2255785186