| Coix seed(Coix lachrymal-jobi L.),a homologous traditional “cereal medicine”,is honored as the “king of health and nourishing cereals” due to abundant coixenolides,active polysaccharides and triterpenoids.All these healthful components contribute to the pharmacological effects including anti-tumor,immune regulation and lower blood sugar.Guizhou,as one of the origin regions yield coix seed,is the largest planting and processing cluster area in China.The molds and their mycotoxins contamination occur frequently in processing due to the heavily rainy days in harvest season as well as inappropriate storage and transportation of coarse coix seed.Therefore,two varieties of coix seeds such as small-grain white shell in Guizhou and large-grain black shell in Laos,accounting for the highest proportion,were taken as the research objects for the first time in this paper,which accounts for the highest proportion in coix seed planting.Firstly,the fungi in coix seed,coarse coix seed and polished coix seed were separated and identified.Moreover,the simultaneous detection methods of four potential mycotoxins(AFB1,DON,ZEN and T-2)was established.The mycotoxins of coix seed from different processing enterprises were analyzed,which revealed the mycotoxins contamination in coix seed products.The results showed that:1.Mold floras in the small-grain white shell coix seed yield in Guizhou and large-grain black shell coix seeds in Laos were evaluated.The results indicated that mold floras in coix seeds varied from several different producing areas and the difference of diversity was not significant.Fifteen molds were isolated and purified by morphological identification,and they were mainly Aspergillus,Fusarium,Rhizopus and Penicillium.Among them,the Aspergillus was predominant.Eight molds were verified by inoculated verification test.Meanwhile,on the basis of the morphological identification and the biological ITS sequence test(similarity of more than 99%),the eight molds were identified as Aspergillus oryzae(C1),Aspergillus niger(C2),Asgillus nidulans(C3),Aspergillus fumigatus(C4),Aspergillus cristatus(C5),Fusariumproliferatum(C6),Fusarium equiseti(C7)and Penicillium aethiopicum(C8).The phylogenetic tree was constructed by the NJ method,revealing that there was a significant difference between the ITS sequences,while the interspecific difference was trivial.2.The analysis method was established to detect the four mycotoxins in coix seeds by ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)of multiple reaction monitoring detection mode(MRM).The results suggested that the detection limits of AFB1,ZEN,T-2 and DON by UPLC-MS/MS were 0.6,3.2,3.2 and 40 ?g/kg,respectively.The curve linearity was good,and all the correlation coefficients(R)were greater than 0.995.In addition,the recovery rate was in the range of 84.8% ~ 101.7%,with the relative standard deviation from 1.3% to 7.3%.3.AFB1,ZEN,T-2 and DON of 19 samples from 11 processing enterprises were tested by UPLC-MS/MS.The results showed that the content of T-2 was not detected in all samples.The positive rate of AFB1 was 42%,with a range of 0.09 to 15.91 ?g/kg.The positive rate of DON was 32%,with a range of 19.17 to 338.32 ?g/kg.The positive rate of ZEN was 100%,with a range of 14.19 to 626.84 ?g/kg.4.The content of ZEN in the samples from Laos with long-term storage reached to 626.84?g/kg.The average contents of ZEN in polished and coarse coix seed were 76.12 ?g/kg and233.65 ?g/kg,respectively.The contents of ZEN were found to reduce by 67% during milling coarse coix seed into polished coix seed.It was noteworthy that AFB1,T-2 and DON were not detected in the pre-cooked coix seed through heat treatment as well as ZEN was merely 14.19?g/kg. |
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