Study On Simultaneous Detection Of Alternaria Toxins In Cereal Grains And Dietary Exposure Assessment

Alternaria toxins pose a great threat to the health of animals and human due to their high toxicity and carcinogenicity.Alternaria toxins have been caused widely residual in cereals,causing huge economic loss and food safety problems.Moreover,“masked”mycotoxins can be formed in cereal growth and escape monitoring by conventional analytical methods,causing the underestimation of risk.Based on the above situation,the research contents of this paper are as follows:1.The binding abilities of Alternaria toxins(AOH and AME)to glutenin,as the representative of macromolecules in grains,have been assessed by different spectroscopic techniques including steady-state fluorescence spectrum,ultraviolet absorption spectrum,synchronous fluorescence spectrum and 3D fluorescence spectrum.The experimental results showed that the fluorescence quenching of glutenin by AOH was due to the formation of Alternaria toxins-glutenin complex,while the interaction between AME and glutenin was not strong.Then,the interactions between AOH and glutenin have been revealed by molecular docking and molecular dynamics.The results showed that AOH was stably bound to glutenin,and the conformational changes of glutenin were not significant.The results of theoretical simulation were consistent with experimental results,which further confirmed the existence of glutenin-bound“mask”Alternaria toxins in cereals.2.The simultaneous detection of Alternaria toxins(Te A,AOH,AME and TEN)method in cereals(rice,corn and wheat)has been proposed based on ultra-performance liquid chromatography-tandem mass spectrometry.Alternaria toxins(Te A,AOH,AME and TEN)in cereals,including the matrix-bound“masked”Alternaria toxins,were effectively extracted by pretreatment method.The acidified acetonitrile aqueous solution was utilized to separate the Alternaria toxins from the matrix complex and transfer the Alternaria toxins to organic phase,which were purified by HLB solid phase extraction column.The chromatographic column was ACQUITY UPLC BEH C18(1.7μm,2.1×100 mm),and ammonia aqueous solution(0.55mmol/L)-acetonitrile were used as mobile phase for gradient elution.Qualitative and quantitative were performed in negative ion electrospray ionization mode and multi-reactive ion monitoring mode,respectively.The Alternaria toxins(Te A,AOH,AME and TEN)showed good linear relationships in cereals(R~2>0.990 0).The detection limits were in the range of 0.4-4.0μg/kg,and the limits of quantification were 1.0-10.0μg/kg.The recoveries of three different spiked levels ranged from 62.50%to 113.40%with RSDs less than 7.89%.The established method has been verified by three provinvial-level inspection institutions,which further indicates the accuracy and repeatability of the method.3.The degree of contamination of Alternaria toxins in cereals has been surveyed and dietary exposure assessment for Alternaria toxins has been proceeded.The 52 samples including rice and wheat flour were obtained by major supermarkets in locals.Then the samples were analyzed by the proposed method.The contamination degrees of rice were lower than that of wheat flour.The detection rates of Te A,AOH,AME and TEN in wheat flour were all over80%and the contents were 0.20-261.31μg/kg,0.20-60.32μg/kg,0.20-55.36μg/kg,and 0.20-69.24μg/kg,respectively.Based on the residual data of Alternaria toxins in wheat flour samples,dietary exposure to Alternaria toxins intakes was evaluated by point estimate model in different age groups in southern and northern China.The exposure doses in the northern were higher than that in the southern,and the exposure doses of low age groups were higher than that in the high age groups.The Te A exposure dose of the high consumption population in the less than 6-years old group[99th percentile(P99)intake level]was close to the threshold of toxicological concern(TTC).The exposure doses of AOH and AME of all age groups in the northern and southern populations exceeded the corresponding TTC values.These results indicate that more attention should be paid to the contamination of Alternaria toxins in cereals.