Purification of fusaproliferin from cultures of Fusarium subglutinans by preparative high-performance liquid chromatography.

Thirty-six Fusarium strains were grown on cracked yellow corn and evaluated for optimum fusaproliferin production, with Fusarium subglutinans E-1583 producing the highest levels (1600 microg/g). Three solvent systems were tested for extracting fusaproliferin from the cultures of F. subglutinans E-1583. Methanol gave the highest fusaproliferin recovery, followed by methanol/1% aqueous NaCl (55:45, v/v) and acetonitrile/methanol/H(2)O (16:3:1, v/v/v). Hexane partitioning was effective in removing many impurities from the crude fusaproliferin extracts prior to the liquid chromatography step. Fusaproliferin samples were further purified by high-performance liquid chromatography (HPLC) with a C18 preparatory column using a mobile phase of acetonitrile/H(2)O (80:20, v/v). The purity of the fusaproliferin was verified by analytical HPLC, GC/MS, (1)H NMR spectroscopy, and electrospray ionization (ESI) MS. The isolated fusaproliferin was shown to be free of impurities and can be used as a standard for routine analysis. Fusaproliferin was shown to be temperature-sensitive when samples were stored at room temperature (20-24 degrees C) for more than several days. After 30 days at 4 degrees C, approximately 8% of the fusaproliferin had been transformed to deacetyl-fusaproliferin; however, samples stored at -20 degrees C for 1 year contained only trace amounts of the deacetylated form.