| Deoxynivalenol (DON) was also named Vomitoxin because it was found as inducing emesis in swine.It is a secondary metabolite produced by Fusariums which belongs to trichothecene mycotoxins. DON is mainly produced by Fusarium graminearum. It was first found in barley infectedby Fusariumin Japan in 1972 and its structure was identified and named 4-deoxynivalenol in the same year.Vesonder et al also got this compound from corn infected by Fusarium graminearum in 1973. WHO and FAO took trichothecene mycotoxins as the most dangerous contaminants occurring naturally in food in the third food additives and contaminants Conference in 1973,especially DON had a large prevalence harmful to the human beings and animals.DON was also isolated from wheat in Shanghai China which caused the pigeons vomiting in 1976.In recent years,DON had a high relevance rate in the feed which caused greatly negative effects to animal husbandry.Additionally,some studies indicated that esophagus cancer and Kaschin-Beck in human beings were related to DON. So the studies on the DON is becoming more and more popular in many countries.It mainly studied on the DON contamination in feedstuff in China from 2012 to 2013 and the preparation craft of DON.The result of this survey provide a theoretical basis to develope targeted prevention and control measures in the future,while DON purification process research can provide the raw materials for toxical research in the future.Test I Contamination surveys of DON in feed in China from 2012 to 2013199 copies of feed and raw materials were detected by HPLC in this test.The results showed that:the whole relevance rate was up to 87.44%, the rate of exceeding the allowed standards of China was up to 27.14%;From the perspective of region,DON contamination in the middle part of China was the most serious; From the perspective of time,DON contamination in 2013 was lighter than 2012;From the perspective of the sample kind,relevance rate of DON in the complete feed was higher than that in the raw materials,but the rate of exceeding the allowed standards of China was lower,and the contamination of DON in the pig feed was more serious than that in poultry feed among the complete feed.Test II Optimization of culture conditions for DON production with fusarium graminearumIn this study, we firstly made toxin-producing medium optimization, and found that the DON production in corn was significantly higher than that in wheat and rice. Then using the corn as the toxin-producing medium, Plackett-Burman (PB) design was used to evaluate the effects of key factors (moisture content, temperature, time, macroconidial suspension concentration and the medium weight) for DON production by Fusarium graminearum. It was shown by PB test results that there was significance difference (R2=0.9869) within this model in terms of DON yield. The final results showed that moisture content(P<0.0001), temperature(P=0.025),time(P=0.0038)were the key factors affecting DON yield. The final results of optimization were the maximum DON yield reached 61.82 mg after cultured at 37℃ for 25 days with moisture content of 32.5% and increased by 26.45% than that of single-factor experiments.Test Ⅲ Optimization of preparation craft of DON with high purityIn this experiment, gel silica column chromatography was used for the separation and purification of DON. Firstly, we separated the sample roughly by petroleum ether, dichloromethane, ethyl acetate and methanol and founded that DON was just in the ethyl acetate fraction. Then the ethyl acetate fraction was applied to a second silica column chromatography under the methylene chloride-methanol(17:1) system. DON was monitored by TLC and the fractions containing DON was concentrated and crystallized at 4℃. The crystallines were analyzed and identified by HPLC-UV and MS comparison to the standard materials. The crystallines had a great purity of 99% which were obtained by The method. Therefore, it is confirmed that we can purified DON by this simple method successfully. |
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