Submitted to: Book Chapter
Publication Type: Book / Chapter
Publication Acceptance Date: March 2, 2008
Publication Date: September 16, 2008
Citation: Molyneux, R.J., Mahoney, N.E., Kim, J.H., Campbell, B.C. 2008. Bioassay-directed Isolation and Identification of Anti-aflatoxigenic Constituents of Walnuts. In: Colegate, S.M., Molyneux, R.J. Editors, Bioactive Natural Products: Detection, Isolation and Structural Identification. Boca Raton, FL: CRC Press. p. 421-437 Technical Abstract: Agricultural crops, especially corn, cotton, peanuts and treenuts, may be infected by Aspergillus flavus and A. parasiticus, resulting in contamination by aflatoxins which can present a risk of hepatocarcinogenicity to humans. All tree nuts are subject to contamination but walnuts are exceptionally resistant to aflatoxigenesis. This chapter describes the detailed experimental design and procedure for bioassay-directed fractionation of the antiaflatoxigenic constituents. The resistance factors are restricted to the seed coat and are not present in the kernel. Chemical analysis of the seed coat has established that the inhibitory activity resides in a complex of hydrolysable tannins common to all walnut cultivars. In vitro experiments showed that tannin from the cultivar 'Tulare' completely suppressed growth of A. flavus at a concentration of 0.5% in the media, with no aflatoxin formed. The evidence indicates that hydrolysable tannins are capable of inhibiting growth of A. flavus and that aflatoxigenicity is phytochemically inhibited by biosynthesis and maintenance of high levels of tannins throughout the growing season. The efficiency of such tannins as antioxidants suggests that they may act by down-regulating a transcriptional activation factor through a signaling pathway involved in the oxidative stress response. This hypothesis is being tested by using complementation analysis of deletion mutants of the yeast Saccharomyces cerevisiae as a model system. The approach allows for high throughput screening of genes involved in oxidative stress response induction of aflatoxin biosynthesis.