The Novel Methods For The Determination Of Aflatoxins In Food And Dietary Exposure Assessment In China

Aflatoxins（AFTs）are poisonous secondary metabolites produced by Aspergillus flavus,Aspergillus parasiticus,and Aspergillus nomius under appropriate conditions.The aflatoxin B₁,B₂,G₁,G₂,M₁,M₂ are commonly found in food and feed.Their toxicological effects on human health caused widespread concern.To satisfy the need of the detection of aflatoxins in food and the exposure risk assessment,the study was carried out on the method of the detection of aflatoxins in food,the occurrence of aflatoxins in several typical kinds of foods and the research of dietary exposure assessment.Firstly,a ultra-high performance liquid chromatography coupled with tandem triple quadrupole mass spectrometry（UPLC-MS/MS）method for the determination of aflatoxins in 12 kinds of diet samples was established.Isotope-labeled internal standards were added to the samples first.The diet samples were ultrasonically extracted with acetonitrile-water solution（86:14,v:v）and then purified using Romer Multi Sep 226 multi-functional columns.The purified extracts were dried under nitrogen and reconstructed in proper sample solvent.The samples were detected by UPLC-MS/MS and quantified by internal standard method.The limits of detection（LOD）of 6 aflatoixns were in the range of 0.002 0.005 μg/kg.The limits of quantity（LOQ）of 6 aflatoixns were in the range of 0.006 0.020 μg/kg.The recoveries in the blank spiked diet samples were in the range of 96.1% 106.8% in different matrices,with the relative standard deviations（RSD）less than 13%.The method is sensitive,accurate and suitable for the determination of aflatoixns in diet samples.Secondly,a novel method for the determination of aflatoxins in edible oil by supercritical fluid chromatography tandem triple quadrupole mass spectrometry（SFC-MS/MS）with liquid-liquid extraction（LLE）was established.The samples were added with isotope-diluted internal standards,diluted with n-hexane,and acetonitrile was added to extract AFTs from the oil layer.The extraction was centriduged and the upper layer was directly injected into the SFC-MS/MS for quantification of the AFTs.The method showed good linearity of the calibration curve（r≥0.9996）AFB₁,AFB₂ and AFG₂ in the concentration range of 0.1⁵0 μg/L and AFG₁ in the range of 0.2 50 μg/L.The limits of detection（LOD）of aflatoxin B₁,B₂,G₁ and G₂ were 0.01 μg/L,0.02 μg/L,0.02 μg/L,0.02 μg/L.The limits of quantification（LOQs）were 0.05 μg/L,0.08 μg/L,0.12 μg/L,0.10 μg/L,respectively.The absolute recoveries in the blank spiked diet samples were in the range of 60.8% 82.9%.Isotope-diluted standards of the four AFTs were used and the anlytes were quantified with internal standard method.the relative recoveriese were in the range of 92.8% 104.6% and relative standard deviation RSD were less than 10%.This rapid,accurate,precise and sensitive method is suitable for analysis of aflatoxins in edible oil samples.30 commercial samples were analyzed and 14 samples were contaminated with AFTs,the overall detection rate of aflatoxins is 46.7%.Of the 30 samples detected,AFB₁ was found in 14 edible oil samples,the level is in the range of 0.050 4.03 μg/L,the average is 0.72 μg/.AFB₂ was found in 7 edible oil samples,the level is in the range of 0.087 0.73 μg/L,the average is 0.28 μg/L,AFG1 was found in 3 edible oil samples,the level is in the range of 0.16 0.62 μg/L,the average is 0.35 μg/L.AFG2 was found in one edible oil sample at the concentration of 0.12 μg/L.This implicated the method suitable for real sample detection and with the pollution levels of 0.050 4.03 μg/L.Thirdly,the residue levels of aflatoxins were determined in 12 kinds of 240 samples from 20 provinces collected in the 5th Chinese TDS.Results show that AFs were detected in 37 samples and the total detection rate of aflatoxins was 15.4%.Significant differences were found among the food categories,where legumes had the highest detection rate at 60%,followed by dairy products at 50% and no aflatoxin was detected in the samples of fruits,beverages,liquor and sugars.In all samples detected,positive samples were found for AFB₁,B₂,G₁,G₂,M₁ but for AFM₂.AFB₁ was the most polluted in total dietary samples,mainly in legumes samples,with the highest level at 50.861 μg/kg,followed by AFB₂,also mainly in legumes samples,with the highest level at 9.519 μg/kg and the average content of 1.018 μg/kg.AFG₁ and AFG₂ were only detected in legumes in Zhejiang,the contaminated level were 1.146 μg/kg and 0.280 μg/kg,respectively.AFM₁ was only detected in dairy and legumes samples.The national average daily dietary exposure to aflatoxin BG in China was 9.526 ng/kg bw,the national average MOE value was 18.This revealed that the dietary intake of aflatoxins BG was a potential health hazard in China.Of the various food categories,legumes and cereals are the primary source of exposure to aflatoxins BG,the contribution rate were 8.8% and 35.2%,with the contributions of other food categories being all less than 10%.In summary,this study established the UPLC-MS/MS method for aflatoxins in TDS samples and a total number of 240 samples from 20 provinces collected in the 5th Chinese TDS study were detected.A novel SFC-MS/MS method for determination of AFTs in edible oil was also extablished and 30 commercially available edible oil in Beijing was analyzed.Based on the determination results of the TDS samles,the main source of AFBG were analyzed and food intake risk of AFBG for Chinese residues were assessed with MOE method.