Application Of Multiple Headspace SPME Followed By GC To Quantitative Analysis Of Ethyl Carbamate In Various Alcoholic Beverages And Bread

Ethyl carbamate (EC) is known to cause cancer in animals and was classified as a carcinogen (Group 2A) by the International Agency for Research on Cancer. EC is naturally formed during fermentation processes and was detected in various fermented products. This has forced the scientific community to develop analytical procedures that can be used to determine not only the presence of EC in different food samples but also their concentrations with a good accuracy.In general, the determination of EC in fermented foods poses problems due to its low concentration and matrix interferences in the samples The problems can be solved by application of suitable sample clean-up procedures and specific detectors. Solid-phase microextraction (SPME) is a fast, convenient, versatile and solvent-free technology that can perform sampling, clean-up, and concentration into a single step. Unlike traditional sample preparation methods, SPME is a non-exhaustive extraction technique in which only a small portion of the target analyte is removed from the sample matrix. The composition of the sample matrix usually affects the extraction of the analyte and the sensitivity of the SPME-based method by the so-called "matrix effect". Therefore, calibration of SPME for quantitative analysis is very important. Multiple headspace solid-phase microextraction (MHS-SPME) combined with gas chromatography is proposed to determine EC in various alcoholic beverages by nitrogen-phosphorus detector and in breads by flame ionization detector. The use of MHS-SPME enables a complete recovery of the target compound and therefore the matrix effect is avoided. These methods offer several advantages including simplicity, reliability, rapidity and reduction of cost, especially in the analysis of a large number of samples in different matrices. The main contents in this thesis are described as follows:1. A novel polyethylene glycol/hydroxy-terminated silicone oil (PEG/OH-TSO) fiber was prepared for the first time and subsequently used instead of commercial ones because of its high extraction ability, good operational stability and long lifetime (above 200 uses).2. MHS-SPME technique coupled to gas chromatography with flame ionization detector for determination of EC in breads is proposed. Owing to the high polarity and hydrophilia of EC, adsorption between the analyte and water causes low transport of EC to the headspace and invalidates MHS-SPME for quantification. Efforts were made to reduce the interference of water in the samples, and results showed that desiccating the sample by mixing it with anhydrous NaaSO4 improved the sensitivity of the method without loss of EC. This approach is seldom used in SPME for this purpose. This novel PEG/OH-TSO fiber was used because of its high extraction ability. After optimization of the experimental parameters that affects MHS-SPME performance, the method was validated. Low limit of detection (0.041μg g-1), wide linearity (0.15-1500μg g-1), good repeatability (with a relative standard deviation of 1.6%) and satisfactory recoveries (92.5-103.4%) were obtained. The developed method was successfully applied to several bread samples.3. MHS-SPME combined with gas chromatography-nitrogen phosphorus detector is proposed to determine EC in various alcoholic beverages after matrix modification. To increase the sensitivity of the method, the PEG/OH-TSO fiber was applied for the analysis. Mixing with anhydrous Na2SO4, the sensitivity of the method can be improved. And the employment of this new fiber and the addition of anhydrous Na2SO4 validated the MHS-SPME for quantification. A Taguchi’s L16 (45) orthogonal array design was employed to evaluate potentially significant factors and screen the optimum conditions for MHS-SPME of EC. The optimized extraction protocol was as follows:35℃of the extraction temperature,10 min of the extraction time,20μL of sample volume and 80 mg of Na2SO4 addition. Under the optimized conditions, limit of detection of 34μg/L was obtained. Relative standard deviation of replicate samples (n=6) was 2.19%. The proposed method was linear in the range of 40μg/L-100 000μg/L, and the coefficient of determination was 0.9997. The method was used to determine EC in various alcoholic beverages. The concentrations obtained were compared with those obtained by standard addition method and no statistically significant differences were observed.