| Ethyl carbamate (EC) was a natural by-product in the process of fermented food production and storage, has been classified as a Group 2A carcinogen by the World Health Organization’s International Agency of Research on Cancer (IARC) in 2007, generally exist in alcoholic beverages, fermented condiment, fermented milk and other foods. Six kinds of typical fermented foods are studied as research object in this paper, two determination methods of EC with optimized sample pretreatment were established by stable isotope dilution LC-MS/MS and GC-MS/MS, and the methods were used in research on EC content of fermented food in market, observing EC content in liquor storage, fermentation and storage of red sufu, changing the environmental factors to determine the impact of various factors on the EC content, so as to put forward control suggestions of reduce EC.1. Two new methods was established for directly determination of EC in fermented food by stable isotope dilution LC-MS/MS and GC-MS/MS. All samples were spiked with EC-d5 as the inner standard in this method. The liquor and vinegar samples were directly analyzed after a micro-pore filtrating. The soy sauce samples were mixed with acetonitrile and stood for minutes until the upper organic layers separating clearly; and the sufu, yoghurt, yellow soybean sauce and bread samples were spiked with ultrapure water, acetonitrile and sodium chloride, mixed and extracted by vortex mixing, and then centrifuged for the layering; next, the upper organic layers of all these samples were injected into the LC-MS/MS or GC-MS/MS after filtration. The evaluation of methodology indicated that the recoveries of the EC was in the range of 92.4%-108.5%for LC-MS/MS and 95.9%-107.9% for GC-MS/MS, the relative standard deviations (RSDs) were 2.18%-7.96%(n=6) for LC-MS/MS and 2.69%-6.31%(n=6) for GC-MS/MS which shows good accuracy and precision. The limit of detection (LOD) was less than 2 ng/mL and the limit of quantification (LOQ) was less than 5 ng/mL which shows good sensitivity. The methods have the advantage of simple operation, environment friendly and suitable for the quantitative detection of large quantities of samples.2.The content of EC in 119 samples of fermented food were determined by LC-MS/MS. The sample survey indicated that the positive rates of EC were from 25.0% to 100% and that the average contents were 2.82 to 221.2 ng/g. Among them the positive rates of bread, bean sauce, yoghurt and cheese were from 25.0% to 45.5% and the average contents were 2.82 to 5.09 ng/g; the positive rates of soy sauce and vinegar were 90.9% and 91.9% and the average contents were 23.45 ng/g and 15.50 ng/g; The positive rates of alcoholic beverages was 100% except 90.0% for blending liquor, the average contents were 8.88 to 74.23 ng/g. The positive rates of Sufu were 100% and the average contents were 7.83 to 221.2 ng/g.3. To investigate the changes of EC content in red sufu and red wine during production and storage. Results indicated that egg flakes product lower content of EC than traditional soy flakes during post fermentation, the content of EC increases with the temperature increasing during post fermentation, the content of EC gradually increases in red sufu in the validity period and start to reduce when beyond the validity period, cold storage and storage avoiding light have a positive effect on inhibiting the formation of EC, low temperature have a positive effect on inhibiting the formation of EC during storage of red wine, heating make the EC content increased for distilled liquor and blending liquor, heating with cover increased the content of EC more than heating without cover. |
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