Occurrence, exposure assessment and reduction strategies of Deoxynivalenol and T-2 mycotoxins, in wheat and corn

Abstract

Cereals particularly corn and wheat are used as staple food in Pakistan due to rich sources of protein, carbohydrates, minerals and vitamins. From field to spoon, the crops are vulnerable for the attack of fungus. As a result of fungal attack, mycotoxins specially Deoxynivalenol (DON) and T-2 toxin are produced which are considered potent carcinogenic, teratogenic, mutagenic and hepatocellular in nature. Samples of corn, wheat and flour were collected from various localities (semi-urban, rural, and urban) of Faisalabad, Pakistan. Many analytical techniques like TLC, HP-TLC, gas chromatography, liquid chromatography with UV-Vis and fluorescence detectors, tendom mass spectrometry are used for analysis ofthese mycotoxins in agriculture commodities but in undertaken study, HPLCcoupled with UV-Vis detector was preferred due to its high resolution, economical and less time-consuming. The procedure for mycotoxins extraction, clean up and analysis was validated for the purpose of extraction efficiency, limit of quantification (LOQ), limit of detection (LOD), precision and accuracy and linearity. Experiments for percentage recovery were performed by spiking cereals samples with three various concentrations 50, 100 & 1000 µg kg-1 of DON and 5, 10 and 50 µg kg-1 of T-2 toxin standard solutions. The recovery% was 94.13 and 93.66 for DON and T2 respectively. In another trial flour, corn and wheat samples were spiked and found recovery% 91.44 and 91.88 for T-2 and DON respectively. LOD and LOQ of DON using HPLC-UV-Vis were evaluated and noted as 0.8 ng/mL and 0.9ng/mL in the range of 1-50 µg/mL and 100-1000 µg/mL respectively with good repeatability (6 and 8) and reproducibility (9 and 14) whereas for T-2 toxin, 1.2 ng/mL and 1.3 ng/mLin the range of 1-10 µg/mL and 5-50 µg/mL respectively with best repeatability (7 and 8) and reproducibility (10 and 12). Sensitivity of detector plays an important role in the resolution of an analyzed analyte. Good linearity was observed for mycotoxins having the values of the regression coefficient (R2) 0.9987 and 0.9993 for DON in the range of 1-50 µg/mL and 100-1000 µg/mL respectively. Similarly, values of regression coefficient (R2) for T-2 toxin were 0.9787 and 0.9998 in the range of 1-10 µg/mL and 5-50 µg/mL respectively with value of slope 1431.8 and 1225.7. xiii Different organic solvents were tried for the complete extraction of these mycotoxins and acetonitrile with double distilled water was found the excellent choice for the DON and T-2 from collected samples. Corn samples of urban (40), semi-urban (40) and rural (20) were examined for the presence of DON residue by HPLC equipped with UV-Vis detector (220 nm). From the results, it is revealed that 63 samples were affected with DON residue with average values ranging from 1354 to 1512 µg kg-1. Distribution study of DON showed that 21% of samples contained low level as compared to established limits (1250 µg kg-1) whereas 42% had the concentration above the tolerable limits. Wheat samples of the same areas were studied for the presence of DON level and 56% were detected contaminated in the range of 1352 to 1585 µg/kg. Distribution pattern was revealed that 22% samples contained residues of DON less than established limits whereas 34% contained intensity of DON greater than threshold limits i.e. 1250 µg kg1. Trichothecene group A is very important mycotoxin and referred as T-2 toxin.The existence of this toxin was studied in collected samples of corn. The collected data indicated that 28% of samples were contaminated with T-2 toxin having a concentration in the range of 101-106 µg/kg. Out of the contaminated samples, 6% were found below the tolerable limits (100 µg kg-1) whereas 22% had the level exceeding the permissible level. Wheat samples of the Faisalabad territory (urban, semi-urban and rural) depicted the presence of T-2 toxin and 34% were observed affected by toxin in the range of 106-112 µg kg-1. From the contaminated samples, 12% had the level less than the recommended permissible values whereas 22% contained toxin concentration above the maximum limits (100 µg kg-1). Wheat flour of different grinding units (mill processed; MP, China chaki; CC and stone processed; SP) were also evaluated for the incidence of DON and T-2 toxin. Wheat flour from urban areas showed the incidence of DON ranging from 1285 to 1475 µg/kg however the samples of semi-urban region contained 1290-1480 µg/kg. Rural area samples were highly contaminated with DON (1255-1485 µg/kg). The incidence of T-2 toxin in flour was studied and found the level 102-140 µg/kg in urban, 101- µg/kg in semi-urban and 101-145 µg/kg in rural samples. xiv Different approaches were carried out to observe the reduction of DON and T-2 toxin in corn and wheat. Gamma irradiation and ozonization were applied to see the fate of mycotoxins. From irradiation treatment, it was concluded that 20 kGy significantly decrease the level of DON in corn (41%) and wheat (41.14%) in samples of rural areas as compared to others whereas reduction of T-2 toxin was high at 20 kGy in corn and wheat samples of urban areas i.e 38.61% and 40.34% respectively. Ozone treatment was considered viable, cheap and easily available to reduce the toxin of cereal commodities at a massive scale. Ozonization for 30 minutes produced best results for the reduction of DON (46.42%) in corn and 38.92% in wheat of rural samples; however, 54.45% reduction of T-2 toxin was achieved in corn and 42.45% reduction was found in wheat samples belonging to urban area. Human beings are always in danger of mycotoxin exposure through the ingestion of contaminated agriculture produce from field to fork stage. Exposure assessment of mycotoxins was evaluated as estimated daily intake (EDI) of the contaminated food with DON and T-2 toxins. From the results, it is evident that femalesare highly exposed with contaminated corn of DON and the EDI fell in the range of 3.05-3.40 µg/kg bw d-1 and were far below than the threshold level (8 µg/kg bw d-1) so the ingestion of corn belongs to Faisalabad areas has no health hazards to consumers. Wheat samples of the same areas contained high EDI as compared to the recommended level (8 µg/kg bw d-1). Females and adults were highly exposed as compared to males with DON in wheat. The levels of EDI for T-2 toxin in cereals (corn and wheat) samples collected from Faisalabad areas showed values less than the tolerable limits i.e. 60 µg/kg bw d-1) so consumption of corn and wheat contaminated with T-2 toxin pose no health effects.