Ethylcarbamate (EC, urethane, C2H5OCONH2) is a known genotoxic carcinogen that commonly exists in fermented food and beverages due to the natural biochemical processes in the fermentation process
[1, 2]. EC was re-classified as a carcinogen (Group 2A) by the International Agency for Research on Cancer (IARC) in 2007
 and has already been regulated in several countries such as Germany, USA, Canada, France and the Czech Republic
. A report from a commission by the European Food Safety Authority (EFSA) issued in 2010
 recommended that special attention should be paid to spirits distilled from stone fruits. Furthermore, EC has been detected in various fermented products such as bread, yoghurt, cheese, soy sauce, vinegar and alcoholic beverages
Methylcarbamate (MC, methylurethane, CH3OCONH2) is simplest ester of carbamic acid. MC has a relatively low toxicity, otherwise, there is experimental evidence that MC is mutagenic in Droso phila
 and carcinogenic in rats
EC and MC can co-exist through natural formation during the fermentation processes
. In order to estimate the risk for EC and MC exposure, a sensitive simultaneous analytical method in fermented foods and beverages is required.
Many methods for detecting EC in beverages have been reported, such as high-performance liquid chromatography (HPLC)
[10–12], liquid chromatography tandem mass spectrometry (LC-MS/MS)
, gas chromatography (GC)
[14–17], and gas chromatography mass spectrometry (GC–MS)
[6, 9, 18–27].
Several assay methods have been based on headspace solid-phase micro extraction (HS-SPME)
[14, 15, 22, 28], where the headspace is discriminatory in nature because only the volatile compounds in the injection vials can be transferred to the GC system. Many volatile alcohols and interferences exist in fermented food and beverages, give much interference, and have a short fiber life time. Liquid-liquid extraction (LLE)
[16, 19, 21] and solid phase extraction (SPE)
[20, 21, 26] are often used to determine the EC content in alcoholic beverages. Although it is a traditional extraction technique, LLE represents a convenient method when it is connected with derivatization. Also, 9-xanthydrol has been used to improve the fluorescence of EC in the HPLC method
[10–12] and to improve the sensitivity of EC using the GC-MS
. However, until now, analytical target compounds and matrices were limited to EC and liquid phases such as spirits or beverages. Another drawback with the methods is that EC is derivatized using 9-xanthydrol after extraction and concentration, and in this case volatile MC and EC can be lost during the evaporation process.
GC coupled with mass spectrometry (GC-MS) is the most widely used due to its good resolution, sensitivity and selectivity. Although the GC-MS methods are very selective and sensitive, it is difficult to detect to ng/kg levels without concentration and derivatization.
In this study, the derivatization parameters that enable the direct reaction of MC and EC in food or beverages are established. The xanthyl methylcarbamate or xanthyl ethylcarbamate derivatives that were formed were extracted by LLE and detected by GC–MS. Therefore, the experiment reported in this paper aimed to optimize the parameters of the derivatization, extraction and GC-MS detection in order to simultaneously determine the MC and EC in fermented foods and beverages, and in order to apply the modified method in the analysis of seventeen real samples.