Profiling of volatile substances by direct thermal desorption gas chromatography high-resolution mass spectrometry for flagging a characterising flavour in cigarette tobacco.

This paper describes an analytical method that supports the implementation of articles 9 and 10 of the WHO Framework Convention on Tobacco Control (FCTC) regarding the provisions on the reduction of the palatability and attractiveness of tobacco products regarding flavour ingredients. This study aimed to develop a screening method to identify cigarettes that may have a characterising flavour to support the implementation of the ban of characterising flavours of tobacco products, as laid down in the US and EU law. An analytical method combining direct thermal desorption and GC-QTOF MS was developed for acquiring the profile of volatile and semi-volatile substances in tobacco. A database of flavour additives was created comprising 133 compounds. A group of cigarettes without a declared characterising flavour was used to establish a reference profile of flavouring chemicals commonly present in tobacco products. A reference profile was modelled both by the means of principal component analysis (PCA) and based on the calculation of threshold values specified as 95th percentile of measured compounds' relative responses. Cigarettes and roll-your-own tobacco labelled as flavoured were analysed to evaluate the discrimination power of the method. A constructed model of the reference cigarettes allowed the differentiation of the flavoured tobacco products from the reference group. The method allows drawing conclusions on the chemical profiles of flavour constituents of tobacco products at even sensorial subliminal concentration levels and is suitable for both the initial screening of products on the market for characterising flavours and for confirmatory purposes after sensory analysis.

Identification of Cigarette Brands by Soft Independent Modeling of Class Analogy of Volatile Substances.

INTRODUCTION: This study aimed to develop a method for discriminating cigarette brands based on the profiles of volatile components extracted from the tobacco fraction of the finished cigarettes to authenticate branded cigarettes of unknown origin. METHODS: An analytical method comprising direct thermal desorption coupled with gas chromatography-quadrupole time-of-flight mass spectrometry was developed for acquiring volatile profiles of cigarettes. About 290 samples of commercially available cigarettes were analyzed. Within this batch, 123 samples represented four popular cigarette brands. They were selected for in-depth characterization. Multivariate analysis was used to investigate the interrelations among volatile compounds of cigarettes and to identify characteristic markers for the cigarette discrimination. Supervised pattern recognition techniques were used for designing classification models. RESULTS: Principal component analysis covering all detected volatiles allowed the differentiation of cigarettes based on the brand. A number of 56 volatile components were identified as markers with high discrimination power. These compounds were used for establishing classification models. A method of soft independent modeling of class analogy developed for the four studied cigarette brands proved to be efficient in the classification of unknown cigarettes, with accuracy between 95.9% and 100%. CONCLUSIONS: The data evaluation by soft independent modeling of class analogy was highly accurate in classification of unknown cigarettes with a low rate of false positives and false negatives. The developed models can be used for discrimination of genuine from non-genuine products with high level of probability. IMPLICATIONS: Profiling of volatiles, which is commonly used for authentication of different food commodities, was applied for the characterization of cigarette tobacco for the purpose of authentication a cigarette brand. Volatile components with a high discrimination power were identified by means of multivariate statistical methods and used for establishing of a classification model. The classification model was able to discriminate genuine from non-genuine cigarettes with a high level of prediction accuracy. This model could be a powerful tool for tobacco control to judge the authenticity of cigarettes.

Assessment of critical steps of a GC/MS based indirect analytical method for the determination of fatty acid esters of monochloropropanediols (MCPDEs) and of glycidol (GEs).

Fatty acid esters of 2- and 3-chloropropanediol (MCPDEs) and fatty acid esters of glycidol (GEs) are commonly monitored in edible fats and oils. A recommendation issued by the European Commission emphasizes the need of generating data on the occurrence of these substances in a broad range of different foods. So far, analytical methods for the determination of MCPDEs and GEs are fully validated only for oils, fats and margarine. This manuscript presents the assessment of critical steps in the AOCS Cd 29a-13 method for the simultaneous determination of MCPDEs and GEs in the fat phase obtained from bakery and potato products, smoked and fried fish and meat, and other cereal products. The trueness of the method is affected by the additional formation of 3-MBPD esters from monoacylglycerols (MAGs), which are frequently present in food. The overestimation of GE contents for some samples was confirmed by the comparison of results with results obtained by an independent analytical method (direct analysis of GE by HPLC-MS/MS). An additional sample pre-treatment by SPE was introduced to remove MAGs from fat prior to the GEs conversion, while the overall method sensitivity was not significantly affected. Trueness of the determination of GEs by the modified analytical procedure was confirmed by comparison with a direct analysis of GEs. The potential impact on accuracy of results of the final sample preparation step of the analytical procedure, the derivatization of free forms MCPD and MBPD with PBA, was evaluated as well. Different commercial batches of PBA showed differences in solubility in a non-polar organic solvent. The PBA derivatization in organic solvent did not affect precision and trueness of the method due to the isotopic standard dilution. However, method sensitivity might be significantly compromised.

Optimization of a Differential Ion Mobility Spectrometry-Tandem Mass Spectrometry Method for High-Throughput Analysis of Nicotine and Related Compounds: Application to Electronic Cigarette Refill Liquids.

Fast market penetration of electronic cigarettes is leading to an exponentially growing number of electronic refill liquids with different nicotine contents and an endless list of flavors. Therefore, rapid and simple methods allowing a fast screening of these products are necessary to detect harmful substances which can negatively impact the health of consumers. In this regard, the present work explores the capabilities of differential ion mobility spectrometry coupled to tandem mass spectrometry for high-throughput analysis of nicotine and 11 related compounds in commercial refill liquids for electronic cigarettes. The influence of main factors affecting the ion mobility separation, such as modifier types and concentration, separation voltage, and temperature, was systematically investigated. Despite small molecular weight differences among the studied compounds, a good separation was achieved in the ion mobility cell under the optimized conditions, which involved the use of ethanol as a polar gas-phase chemical modifier. Indeed, differential ion mobility was able to resolve (resolution >4) nicotine from its structural isomer anabasine without the use of any chromatographic separation. The quantitative performance of the proposed method was then evaluated, showing satisfactory precision (RSD ≤ 16%) and recoveries ranging from 85 to 100% for nicotine, and from 84 to 126% for the rest of the target analytes. Several commercial electronic cigarette refill liquids were analyzed to demonstrate the applicability of the method. In some cases, significant differences were found between labeled and measured levels of nicotine. Anatabine, cotinine, myosmine, and nornicotine were also found in some of the analyzed samples.

Derivatization of bisphenol A and its analogues with pyridine-3-sulfonyl chloride: multivariate optimization and fragmentation patterns by liquid chromatography/Orbitrap mass spectrometry.

RATIONALE: Due to the growing restrictions on the use of bisphenol A (BPA), several other bisphenols are gaining importance as substitutes for BPA in a variety of applications. There is, therefore, a real need for selective and sensitive methods based on mass spectrometry which will allow the human exposure to these new bisphenols to be assessed. METHODS: Derivatization of BPA and its substitutes with pyridine-3-sulfonyl chloride is used to enhance the detection capability of bisphenols by electrospray ionization mass spectrometry. A multivariate experimental design, Box-Behnken response surface, was used to evaluate the influence of the main variables potentially affecting the derivatization yield. Fragmentation patterns for all the derivatized bisphenols were acquired by high-resolution/accurate-mass Orbitrap mass spectrometry. RESULTS: Temperature and pH were identified as the most important factors affecting the derivatization yield of bisphenols. Fragmentation of the protonated molecules produced abundant analyte-specific product ions. Most of the derivatized bisphenols showed significant improvements in their signal-to-noise ratios compared with the underivatized forms. The stability of these derivatives was demonstrated through several freeze/thaw cycles, short-term room temperature and long-term cold storage. CONCLUSIONS: Derivatization of BPA and its structural analogues with pyridine-3-sulfonyl chloride is proposed as a specific, sensitive, high-throughput approach to their analysis by liquid chromatography coupled to electrospray ionization mass spectrometry.

Evaluation of the quality of postharvest rapeseed by means of an electronic nose.

BACKGROUND: Rapeseed is a valuable source of edible oil. The presence of even a small amount of mouldy or burnt rapeseed in a particular production batch deteriorates the quality of the edible oil obtained from it. Since the traditional method of using a panel of experts is time-consuming, there is a need for fast and easy methods for rapeseed quality evaluation by intelligent devices to replace human labour. RESULTS: For rapeseed quality evaluation, an electronic nose equipped with an array of eight quartz microbalance sensors and four metal oxide semiconductor sensors was used. Signals generated by the sensors were analysed by principal component analysis and discriminant function analysis. Identification of samples that contained small proportions of mouldy or burnt rapeseed was possible despite the differences between the particular varieties studied. CONCLUSION: Electronic nose technology has shown the possibility of detecting samples of faulty rapeseed at very low contamination levels and distinguishing them with high probability from sound rapeseed.

Influence of battery power setting on carbonyl emissions from electronic cigarettes.

INTRODUCTION: Although e-cigarettes share common features such as power units, heating elements and e-liquids, the variability in design and possibility for customization represent potential risks for consumers. A main health concern is the exposure to carbonyl compounds, which are formed from the main components of e-liquids, propylene glycol and glycerol, through thermal decomposition. Levels of carbonyl emissions in e-cigarette aerosols depend, amongst others, on the power supplied to the coil. Thus, e-cigarettes with adjustable power outputs might lead to high exposures to carbonyls if the users increase the power output excessively. The aim of this work was to elucidate the generation of carbonyls in relation to undue battery power setting. METHODS: Carbonyl emissions were generated by two modular e-cigarettes equipped with two atomizers containing coils of different resistance following the ISO 20768:2018 method. The battery power output was increased from the lower wattage level to above the power range recommended by the producer. Carbonyls were trapped by a 2,4-dinitrophenylhydrazine (DNPH) solution and analysed by LC-MS/MS. RESULTS: The amount of carbonyl emissions increased with increasing power setting. An exponential incline was observed when the applied power level exceeded the recommended power range. Exceeding the recommended power range by just 5 watts resulted in up to twenty times the amount of carbonyls emitted at the recommended upper power level. Generation of acetaldehyde and acrolein next to other carbonyls was prominent at high power outputs. CONCLUSIONS: E-cigarettes with customisable power setting might generate high amounts of carbonyls if the battery power output is set by the consumer to levels above the recommended range. This represents a high risk of exposure to carbonyls and thus should be avoided by integrating safety features in e-cigarette devices to limit the possible power settings to the range specified by the manufacturer.

Validation by collaborative trial of a method for the determination by GC-MS and LC-MS/MS of boar taint marker compounds in pork tissue.

Meat from male pigs may develop an off-flavour, commonly known as boar taint. Castration of male piglets prevents the potential formation of off-flavour. In the suggested method, three marker compounds for boar taint (skatole, androstenone and indole) are quantified in pork fat by isotope dilution gas chromatography mass spectrometry (GC-MS) or by isotope dilution liquid chromatography tandem mass spectrometry (LC-MS/MS). This method was validated by collaborative trial according to ISO 5725-2:1994. The studied concentration ranges included sensorial thresholds. The repeatability relative standard deviation (RSDr) ranges from 3% to 10% and the reproducibility relative standard deviation (RSDR) from 10% to about 30%. The method has proven to be robust and free from matrix interferences. The method performance characteristics are compliant with requirements for official control methods in the area of food contaminants; therefore, the method is regarded as fit for its intended purpose.

Evaluation of gas chromatography columns for the analysis of the 15 + 1 EU-priority polycyclic aromatic hydrocarbons (PAHs).

Three different stationary phases were investigated for the analysis of the 15 + 1 EU-priority polycyclic aromatic hydrocarbons (PAHs) by gas chromatography-mass spectrometry. In addition to the most commonly used 5% phenyl methylpolysiloxane, a mid-polar phase (50% phenyl methylpolysiloxane) and a recently commercialised mid-polar to polar phase (Optima delta-6), were evaluated. Challenging groups of PAHs in terms of separation, such as the pair dibenz[a,h]anthracene-indeno[1,2,3,-cd]pyrene and the two groups benzo[b]fluoranthene-benzo[k]fluoranthene-benzo[j]fluoranthene and cyclopenta[cd]pyrene-benz[a]anthracene-chrysene, were satisfactorily separated by using the mid-polar phase. Moreover, discrimination in terms of peak height for the heaviest PAHs (caused from the strong interaction of these compounds with the stationary phase) was reduced without compromising the resolution of the other target analytes when applying the mid-polar phase in a tailor-made column geometry (20 m x 0.18 mm internal diameter and 0.14 microm film thickness) in combination with optimised chromatographic conditions. A significant enhancement of the analytical sensitivity for dibenzopyrenes is demonstrated with an almost threefold increase of the signal-to-noise (S/N) ratio for dibenzo[a,h]pyrene, the last eluting PAH. The ability of the selected column to separate potentially interfering PAHs from the target analytes in both solvent solutions and food extracts is demonstrated.

Results of a European inter-laboratory comparison study on the determination of EU priority polycyclic aromatic hydrocarbons (PAHs) in edible vegetable oils.

A collaborative study on the analysis for 15 + 1 EU priority PAHs in edible oils was organised to investigate the state-of-the-art of respective analytical methods. Three spiked vegetable oils, one contaminated native sunflower oil, and one standard solution were investigated in this study. The results of 52 laboratories using either high-performance liquid chromatography with fluorescence detection or gas chromatography with mass-selective detectors were evaluated by application of robust statistics. About 95% of the laboratories were able to quantify benzo[a]pyrene together with five other PAHs included in the commonly known list of 16 US-EPA PAHs. About 80% of the participants also quantified seven additional PAHs in most samples, two of which were benzo[b]fluoranthene and benzo[k]fluoranthene, which were also known from the EPA list. Only about 50% of the participants quantified cyclopenta[cd]pyrene, benzo[j]fluoranthene, and benzo[c]fluorene. The robust relative standard deviations of the submitted results without discrimination between the methods applied ranged between 100% for 5-methylchrysene in spiked olive oil and 11% for the same analyte in spiked sunflower oil. The results clearly showed that for these analytes the methods of analysis are not yet well established in European laboratories, and more collaborative trials are needed to promote further development and to improve the performances of the respective methods.

Experimental design-based isotope-dilution SPME-GC/MS method development for the analysis of smoke flavouring products.

For the implementation of Regulation (EC) No 2065/2003 related to smoke flavourings used or intended for use in or on foods a method based on solid-phase micro extraction (SPME) GC/MS was developed for the characterisation of liquid smoke products. A statistically based experimental design (DoE) was used for method optimisation. The best general conditions to quantitatively analyse the liquid smoke compounds were obtained with a polydimethylsiloxane/divinylbenzene (PDMS/DVB) fibre, 60°C extraction temperature, 30 min extraction time, 250°C desorption temperature, 180 s desorption time, 15 s agitation time, and 250 rpm agitation speed. Under the optimised conditions, 119 wood pyrolysis products including furan/pyran derivatives, phenols, guaiacol, syringol, benzenediol, and their derivatives, cyclic ketones, and several other heterocyclic compounds were identified. The proposed method was repeatable (RSD% <5) and the calibration functions were linear for all compounds under study. Nine isotopically labelled internal standards were used for improving quantification of analytes by compensating matrix effects that might affect headspace equilibrium and extractability of compounds. The optimised isotope dilution SPME-GC/MS based analytical method proved to be fit for purpose, allowing the rapid identification and quantification of volatile compounds in liquid smoke flavourings.

Collaborative trial validation study of two methods, one based on high performance liquid chromatography-tandem mass spectrometry and on gas chromatography-mass spectrometry for the determination of acrylamide in bakery and potato products.

A European inter-laboratory study was conducted to validate two analytical procedures for the determination of acrylamide in bakery ware (crispbreads, biscuits) and potato products (chips), within a concentration range from about 20 microg/kg to about 9000 microgg/kg. The methods are based on gas chromatography-mass spectrometry (GC-MS) of the derivatised analyte and on high performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) of native acrylamide. Isotope dilution with isotopically labelled acrylamide was an integral part of both methods. The study was evaluated according to internationally accepted guidelines. The performance of the HPLC-MS/MS method was found to be superior to that of the GC-MS method and to be fit-for-the-purpose.

Investigation of the correlation of the acrylamide content and the antioxidant activity of model cookies.

The purpose of this study was to evaluate the correlation between the acrylamide (AA) content and the antioxidant activity (AOA) of self-prepared cookies. Cookies were baked in the laboratory under defined conditions following four different recipes. The parameters of investigation were the influence of the type and relative content of sugar (glucose and fructose) and the baking time on the AA content as well as AOA of the final products. Parameters depending on the recipe and baking conditions such as the moisture content, the total nitrogen concentration, and the color of the products were evaluated for all samples as well. To prove the transferability of the findings gained with model cookies to samples from industry, the same measurements were performed on seven different types/brands of cookies that were purchased in local markets. A direct correlation was found between the concentration of AA and the AOA. With increasing baking time, the moisture content of the cookies decreased. The latter parameter correlated well with the AA concentration and AOA. The use of fructose enhanced the concentration of AA and the AOA of the final products, when compared with the use of sucrose. However, a simple model for the prediction of acrylamide contents and the AOA of samples from the baking time, color, protein, or moisture content of the samples was not found.

Single-laboratory validation of a saponification method for the determination of four polycyclic aromatic hydrocarbons in edible oils by HPLC-fluorescence detection.

An analytical method is reported for the determination of four polycyclic aromatic hydrocarbons (benzo[a]pyrene (BaP), benz[a]anthracene (BaA), benzo[b]fluoranthene (BbF) and chrysene (CHR)) in edible oils (sesame, maize, sunflower and olive oil) by high-performance liquid chromatography. Sample preparation is based on three steps including saponification, liquid-liquid partitioning and, finally, clean-up by solid phase extraction on 2 g of silica. Guidance on single-laboratory validation of the proposed analysis method was taken from the second edition of the Eurachem guide on method validation. The lower level of the working range of the method was determined by the limits of quantification of the individual analytes, and the upper level was equal to 5.0 µg kg(-1). The limits of detection and quantification of the four PAHs ranged from 0.06 to 0.12 µg kg(-1) and from 0.13 to 0.24 µg kg(-1). Recoveries of more than 84.8% were achieved for all four PAHs at two concentration levels (2.5 and 5.0 µg kg(-1)), and expanded relative measurement uncertainties were below 20%. The performance of the validated method was in all aspects compliant with provisions set in European Union legislation for the performance of analytical methods employed in the official control of food. The applicability of the method to routine samples was evaluated based on a limited number of commercial edible oil samples.

Rapid and sensitive method for the determination of four EU marker polycyclic aromatic hydrocarbons in cereal-based foods using isotope-dilution GC/MS.

A rapid and sensitive method has been developed for the determination of the four European Union marker polycyclic aromatic hydrocarbons (PAHs; benz[a]anthracene, chrysene, benzo[b]fluoranthene and benzo[a]pyrene) in some cereal-based foods. The method is based on pressurised liquid extraction (PLE), solid-phase extraction clean-up (SPE) and isotope-dilution gas chromatography with mass-spectrometric detection (GC/MS). The developed method was calibrated for the content range of 0.05-12.5 µg kg(-1) (expressed on a product basis). Recoveries of PAH were monitored in each sample via the recovery of (13)C-labelled PAHs. Recovery values were in the range between 86% and 91%, with relative standard deviations (RSDs) between 5% and 9%. The achieved limits of detection for all analytes were below 0.05 µg kg(-1). The applicability of the method for the analysis of routine samples was studied by the analysis of a set of commercial bread and breakfast cereal samples. In all analysed samples, benzo[a]pyrene (BAP) was the most prevalent PAH with the content between 0.09 and 0.30 µg kg(-1). On average, samples showed low levels of the sum of the four EU marker PAHs (ΣPAH4) that ranged between 0.11 and 0.22 µg kg(-1) for bread samples and between 0.23 and 0.87 µg kg(-1) for breakfast cereal samples. The developed method was found suitable for the determination of PAHs in cereal-based foods like cornflakes and breads with total relative fat contents below 3.5%.

Analytical method for the trace determination of esterified 3- and 2-monochloropropanediol and glycidyl fatty acid esters in various food matrices.

Fatty acid esters of 3-monochloro-1,2-propanediol (3-MCPDEs), of 2-monochloro-1,3-propanediol (2-MCPDEs) and of 2,3-epoxy-1-propanol or glycidol (GEs), which are considered to be deleterious to human health, may occur in a broad variety of food samples. A proper risk assessment of those substances requires the availability of robust occurrence data; in this respect concerns have been raised regarding the reliability of results obtained with the currently available methods to determine those substances in processed food. This article presents an indirect analytical procedure for the simultaneous determination of 3-MCPDEs, 2-MCPDEs and GEs in a wide variety of food products after extraction by pressurised liquid extraction (PLE) and determination by gas chromatography mass-spectrometry (GC-MS). For the differentiation of MCPDEs and GEs, the latter were first converted to monobromopropanediol esters (MBPDEs) in acid aqueous solution of sodium bromide. MCPDEs and MBPDEs were then hydrolysed under acidic conditions followed by derivatisation of the released free (non-esterified) form in ethyl acetate with phenyl boronic acid (PBA). Quantification of the analytes was carried out using the isotopic labelled analogues of both MCPDEs and GEs. Limits of detection (LODs) and limits of quantitation (LOQs) were in the range of 7-17mgkg(-1) and 13-31mgkg(-1) respectively, while the working range of the method was between LOQ and 1850mgkg(-1) expressed on fat basis. The developed method was successfully applied for the analysis of the target compounds in more than 650 different food samples covering the following commodities: bread and rolls, fine bakery wares, smoked fish products, fried and roasted meat, potato based snacks and fried potato products, cereal-based snacks and margarines.

Evaluation of results of an interlaboratory comparison test on determination of acrylamide in crispbread samples.

The European Commission's Directorate General Joint Research Centre has organized several proficiency tests on the determination of acrylamide (AA) in food. This paper presents the results and outcome of a proficiency test that focused on the determination of AA in crispbread samples. One of the goals was the identification of the influence of different parameters such as analyte extraction or instrument calibration on the analytical results. A set of samples, containing 3 different crispbread samples as well as extracts of one crispbread sample and AA standard solutions, was shipped to each participant. A total of 42 European laboratories reported analytical results that were evaluated by applying internationally accepted protocols and procedures. The study found that, for each sample, the results of 4-8 laboratories were outside the range formed by the target value plus or minus the 2-fold of the target standard deviation; thus, they did not perform satisfactorily. In transferring this knowledge to the data of monitoring databases of AA in food, care must be taken that data are quality controlled, as it is likely that some of them may be biased.

Overview of acrylamide monitoring databases.

Since high acrylamide levels in carbohydrate-rich food were reported in 2002, many research activities were started in order to gain knowledge on occurrence, formation, and prevention of this compound in food products. Among them, monitoring programs were conducted in many countries worldwide by official bodies as well as by the food industry. National and international bodies set up monitoring databases. In 2003, both the European Commission and the World Health Organization posted calls for data and placed their spreadsheets for the submission of data on the Web. The goal of the databases is to collect data for a reliable estimation of the exposure of consumers to acrylamide via the food chain. This paper describes the assessment of the data quality and outlines the composition of the data in the 2 databases, to date.

Results from two interlaboratory comparison tests organized in Germany and at the EU level for analysis of acrylamide in food.

After the publication of high levels of acrylamide (AA) in food, many research activities started all over the world in order to determine the occurrence and the concentration of this substance in various types of food. As no validated methods were available at that time, interlaboratory studies on the determination of AA in food were of the highest priority. Under the boundary conditions of applying well-established evaluation schemes, the results of 2 studies conducted by the Federal Institute for Risk Assessment (BfR) in Germany and by the European Commission's Directorate General Joint Research Center (JRC) exhibited an overall acceptable performance of the participants in these studies. Nevertheless, many laboratories showed problems in determining AA in food with a complex matrix such as cocoa. The results of analysis also showed a broader variation of AA for samples with low AA concentrations and indicated a bias of the results obtained by gas chromatography-mass spectrometry without derivatization. Improvements of the performance of some laboratories appeared to be necessary.