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Title: RAPID PURIFICATION OF FUMONISINS B3 AND B4 WITH SOLID STATE PHASE EXTRACTION COLUMNS

Author
item Poling, Stephen
item Plattner, Ronald

Submitted to: Journal of Agricultural and Food Chemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/7/1996
Publication Date: N/A
Citation: N/A

Interpretive Summary: Fumonisins are a group of toxins produced by a mold that grows on corn. Fumonisins are responsible for diseases in horses and pigs caused by the consumption of moldy feed. Fumonisins are commonly found in grains used for animal feed and in processed corn products consumed by humans. Fumonisin B1 occurs in the greatest amounts in naturally contaminated corn and is the best studied of the fumonisins. Fumonisin B1 is the only fumonisin tested in purified form and shown to be toxic. There is preliminary evidence that the other fumonisins may be as toxic as fumonisin B1. We found a strain of the mold that makes fumonisins B3 and B4 as the main components. They occur in naturally contaminated corn but in amounts less than that of fumonisin B1. We developed a simple two-step method to isolate and purify fumonisin B3 and B4 from this stain. The first step recovers 95% of the fumonisins from extracts of moldy corn and gives a fumonisin sample that is 70% fumonisins. The second step separates fumonisin B3 from B4 and increases the purities to 95% fumonisins. The method does not require any expensive equipment, takes only a few hours to complete, and can produce enough fumonisin B3 and B4 to begin testing their toxicity and comparing it to that of fumonisin B1. Additional purification can provide analytical standards to be used to measure the amounts that occur in naturally contaminated corn. Corn products for human consumption and animal feeds can then be analyzed for these fumonisins to ensure they do not occur at levels that are dangerous to health of humans or farm animals.

Technical Abstract: A simple method was developed to isolate fumonisin B3 and B4 (FB3 and FB4) from cultures of a novel strain of F. moniliforme which produces FB3 and FB4 but does not make FB1 or FB2. Undiluted extracts from the culture material are loaded onto NH2 solid phase extraction cartridges and the fumonisins eluted with 5% HOAc/MeOH. The NH2 cartridges are very efficient at removing pigments from the samples. The 5% HOAc/MeOH eluate was diluted with 1.5 volumes of water and loaded onto a tC18 cartridge. The fumonisins were eluted with increasing amounts of CH3CN in H2O. Complete separation was obtained between the FB3 fraction and the FB4 fraction. The resulting fractions were analyzed for purity by three methods: fluorescence detection of the OPA derivatives, evaporative light-scattering detection of the underivatized fumonisins and by electrospray-MS. The fractions from the combined NH2/tC18 method contained at least 90% and probably about 95% fumonisins. Recovery of the fumonisins from the extracts exceeded 95%. A 277 mg FB3 fraction and a 62 mg FB4 fraction were obtained from the combination of a 10 g NH2 cartridge and a 10 g tC18 cartridge. The method is amenable to scale-up for purification of larger quantities. It should be useful for providing FB3 and FB4 for toxicity studies or for further purification by semi-preparative HPLC.