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BACKGROUND: Mineral oil aromatic hydrocarbon (MOAH) analysis in foods is a major analytical challenge. Quantification is associated with a high uncertainty. The sources of uncertainty are multiple, but the major one is related to data interpretation and integration, which is partially derived from insufficiently efficient sample preparation. Recently, an updated ISO method for the analysis of mineral oil in fats and oils and a standard operating procedure for infant formula analysis have been published. Both methods reported significantly different (up to 1.25) distributions of the internal standards used for quantification (i.e., tri-tert-butyl benzene (TBB) and 2-methyl naphthalene (2-MN)) over the different solvent phases used in the saponification step. RESULTS: In this work, a microwave-assisted saponification and extraction method was optimized for MOAH analysis to solve the problem related to the MOAH internal standards partition. The paper examines the impact of the solvent mixture used, the concentration of KOH on the partition of TBB and 2-MN, and the effect of the matrix and the washing step to extract the unsaponifiable fraction containing the mineral oils. SIGNIFICANCE: The optimized procedure achieved a TBB/2-MN ratio of 1.05 ± 0.01 tested in five different fats and oils, namely, sunflower, rapeseed, coconut, palm, and extra virgin olive oils. The method can significantly contribute to reducing the uncertainty of the MOAH quantification when saponification is applied.

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The present manuscript reports and discusses critical issues related to the determination of mineral oil hydrocarbon contamination in Citrus essential oils (EOs); an on-line liquid-gas chromatography system equipped with a Y-interface was used (with no additional off-line step for pre-concentration). In total, eighteen samples were analyzed, specifically eleven cold-pressed (CP) and seven distilled EOs. With regard to the CP EOs, various degrees of mineral oil saturated hydrocarbon (MOSH) contamination were detected, ranging between 10.7 and 338.4 mg kg-1 (only one sample was MOSH-free); different MOSH sub-fractions were determined, with the > C25- ≤ C35 sub-fraction always present, with an average concentration of 74.5 mg kg-1. Based on the EO composition, different sample amounts were injected to avoid the overloading of the LC column and consequently the GC one, thus leading to different limits of quantification (LoQ), which were either 2 mg kg-1 (for bergamot EO) or 5 mg kg-1 (for all the other investigated samples). For all samples, the mineral oil aromatic hydrocarbon level was always lower than the LoQ.

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Methods for determining MOSH and MOAH in edible oils showed major problems with interlaboratory comparability of analytical results, especially in the lower concentration range below 10 mg/kg. However, a method with improved sensitivity and reproducibility is urgently needed to obtain a valid data basis for minimization efforts. To cope this problem a new method was created in 2020. The method was established as the standard method DGF C-VI 22 (20) of the German Society for Fat Science e.V. (DGF). For the development of this method different sample epoxidation approaches have been performed, evaluated and improved. Additionally, a saponification, a decision tree for sample preparation, an upstream clean-up column and a system suitability test were introduced. The focus was on reliability and interlaboratory comparability over all edible oil matrices up to a LOQ of 1 mg/kg. The optimized method was validated in terms of trueness and precision in a collaborative trail with 11 laboratories. The achieved recovery rates of 89-105% MOSH and 70-105% MOAH met the JRC requirements. Method and validation results were obtained with HorRat values between 1.3 and 1.8 for MOSH and MOAH.

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COVID-19 currently represents one of the major health challenges worldwide. Albeit its infectious character, with onset affectation mainly at the respiratory track, it is clear that the pathophysiology of COVID-19 has a systemic character, ultimately affecting many organs. This feature enables the possibility of investigating SARS-CoV-2 infection using multi-omic techniques, including metabolomic studies by chromatography coupled to mass spectrometry or by nuclear magnetic resonance (NMR) spectroscopy. Here we review the extensive literature on metabolomics in COVID-19, that unraveled many aspects of the disease including: a characteristic metabotipic signature associated to COVID-19, discrimination of patients according to severity, effect of drugs and vaccination treatments and the characterization of the natural history of the metabolic evolution associated to the disease, from the infection onset to full recovery or long-term and long sequelae of COVID.

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During the 2020-21 olive oil campaign, the contribution of harvesting operations to mineral oil saturated (MOSH) and aromatic hydrocarbon (MOAH) contamination was studied. Oils extracted from hand-picked olives (15 different olive groves) generally had background MOSH (<2.7 mg/kg), and no quantifiable MOAH. In 40% of the cases, an important contamination increase was observed after harvesting operations. Except for one sample (325.8 and 111.0 mg/kg of MOSH and MOAH, respectively), other samples reached 4.3-33.7 mg/kg of MOSH and 1.1-11.3 mg/kg of MOAH. Accidental leaks of lubricants and/or contact with lubricated mechanical parts, were identified as important sources of contamination. Chromatographic traces obtained by on-line high-performance liquid chromatography (HPLC)-gas chromatography (GC)-flame ionization detection (FID) allowed for source identification. A comprehensive two-dimensional gas chromatographic platform (GC × GC) with parallel FID/MS detection was implemented for confirmation and to attempt the characterization of the contaminations. Good harvesting practices are suggested to minimize contamination risks.

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Mineral oil is an ubiquitous food contaminant potentially toxic. It is generally divided into aromatic hydrocarbons (MOAH) and saturated hydrocarbons (MOSH). These compounds are currently under investigation by the European Union to determine their occurrence and their toxicity before legislating on the matter. Although the discussion mainly focuses on food, animal feed can indirectly contribute to human exposure to such a contaminant. In this study, seven commercial feeds were analyzed. The analyses were carried out in two different Universities (Udine-IT and Liège-BE), performing the same sample preparation protocol: microwave-assisted saponification and extraction followed by epoxidation for the MOAH fraction. The final determination was performed by hyphenated liquid-gas chromatography (LC-GC) and LC coupled to comprehensive multidimensional gas chromatography (LC-GC × GC) with parallel detection, namely flame ionization detector (FID) and time-of-flight mass spectrometer (ToFMS). The results obtained by the two laboratories were generally in good agreement. The results obtained by LC-GC × GC-ToFMS/FID platform provided consistent results, with the advantages of more robust data interpretation that can compensate for problems occurring during purification. Moreover, the coupling of enhanced separation obtained by GC × GC and the MS information allowed for a more in-depth characterization of the contamination.

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A rapid and solvent-saving method, based on microwave-assisted saponification (MAS) followed by epoxidation and on-line liquid chromatography (LC) - gas chromatography (GC) - flame ionization detection (FID), was optimized and validated for high-sensitivity MOAH determination in extra virgin olive oils. Quantitative recoveries and good repeatability were obtained even at concentrations of added mineral oils close to the LOQ (0.5 mg/kg for the total hump, 0.2 mg/kg for each single C-fraction). The validated method, also applied for MOSH determination (C-fraction LOQ: 0.5 mg/kg), was used to analyse 18 extra virgin olive oils from the Italian market or oil mills, and 10 additional samples extracted in the laboratory (with an Abencor apparatus) from hand-picked olives. The former resulted contaminated with variable amounts of MOSH and MOAH (on average 19.0 mg/kg and 2.5 mg/kg, respectively), while the latter showed no detectable MOAH, and low and rather constant MOSH (generally below 2.0 mg/kg).

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Agrifood by-products and microalgae represent a low-cost and valuable source of bioactive compounds with neuroprotective properties. However, the neuroprotective effectiveness of therapeutic molecules can be limited by their capacity to cross the blood-brain barrier (BBB) and reach the brain. In this research, various green extracts from Robinia pseudoacacia (ASFE), Cyphomandra betacea (T33), Coffea arabica (PPC1), Olea europaea L., (OL-SS), Citrus sinensis (PLE100) by-products and from the microalgae Dunaliella salina (DS) that have demonstrated in vitro neuroprotective potential were submitted to an in vitro BBB permeability and transport assay based on an immortalized human brain microvascular endothelial cells (HBMEC) model. Toxicity and BBB integrity tests were performed, and the transport of target bioactive molecules across the BBB were evaluated after 2 and 4 h of incubation using gas and liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry (GC/LC-Q-TOF-MS). The HBMEC-BBB transport assay revealed a high permeability of representative neuroprotective compounds, such as mono- and sesquiterpenoids, phytosterols and some phenolic compounds. The obtained results from the proposed in vitro BBB cellular model provide further evidence of the neuroprotective potential of the target natural extracts, which represent a promising source of functional ingredients to be transferred into food supplements, food additives, or nutraceuticals with scientifically supported neuroprotective claims.

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The production of renewable fuels as biodiesel and bio-jet fuel is usually originated by the transformation and processing of oleaginous feedstocks, mainly composed of triacylglycerols. Currently, a significant part of the triacylglycerol production relies on grassy oil crops or other woody oil plants, representing more than 120 million metric tons every year. Considering that the worldwide triacylglycerol demand is expected to rise in the future, alternative routes are necessary to ensure a sustainable biodiesel industry and limit diesel price volatility. In this context, the use of animal fats could be an interesting alternative for biodiesel production as the production of animal byproducts represents nearly 17 million tons per year in the European Union only (2020). Animal fats, however, contain large amounts of no-esterified fatty acids and other oxygen compounds, reducing the yield of biodiesel. Therefore, a specific pretreatment is needed before the trans-esterification process. The setup of such appropriate pretreatments requires detailed upstream characterization of the minor components present in the feedstock. For this purpose, the minor component profile of animal fat was investigated by comprehensive two-dimensional gas chromatography coupled with high-resolution time-of-flight mass spectrometry. This was preceded by an innovative sample fractionation and focalization of these minor components by a preparative liquid chromatographic column method. The overall method permitted to extract different levels of information from the two-dimensional chromatograms, leading to a tentative identification of more than 150 compounds, mainly oxygenated, belonging to different chemical classes.

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Consumers are daily exposed to a range of mineral oil hydrocarbons via food consumption. Major sources of MOH in food are packaging and additives, processing aids, and lubricants. In 2019, an EU guidance was released covering specific directions for sampling and analysis of mineral oil saturated hydrocarbons (MOSH) and mineral oil aromatic hydrocarbons (MOAH) in food and food contact materials within the frame of Recommendation (EU) 2017/84 for the monitoring of mineral oils. The parameters required by the guide are increasingly stringent, and coping with this type of analysis is now very challenging. It is within such a context that the present research is confined, inasmuch that it is focused on the construction of a low-cost, lab-made Y-interface for liquid-gas chromatography coupling used for the determination of MOSH and MOAH in foodstuffs. The response ratios of alkanes comprised between C10 and C50 were measured and were comprised between 0.9 and 1.1, with a maximum coefficient of variation of 4% (n = 5). Intermediate precision was evaluated for the fat/oil category during a period of 48 days obtaining a value of 10%. Qualitative and quantitative analysis of both MOSH and MOAH were performed in a single run and in a fully-automated manner. Seventeen different foods were analyzed in order to cover the categories reported in the EU guide. Saturated hydrocarbon contamination was detected only in a few samples (in the range 1-153 ppm); MOAH contamination was found only in one sample (sunflower oil: 15 ppm).

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The determination of the level of mineral oil contamination in foods is a well-known problem. This class of contaminants is generally divided into mineral oil saturated and aromatic hydrocarbons with different toxicological relevance and analytical challenges. Among the many challenges, data interpretation and integration represent an important source of uncertainty in the results provided by different laboratories leading to a variation evaluated on the order of 20%. The use of multidimensional comprehensive gas chromatography (GC × GC) has been proposed to support the data interpretation but the integration and the reliability of the results using this methodology has never been systematically evaluated. The aim of this work was to assess the integration and quantification performance of a two-dimensional (2D) software. The data were generated using a novel, completely automated platform, namely LC-GC × GC coupled to dual detectors, i.e., time-of-flight mass spectrometer (MS) and flame ionization detector (FID). From a systematic study of the failures of the two-dimensional quantification approach a novel solution was proposed for simplifying and automating the entire process. The novel algorithm was tested on ad hoc created samples (i.e. a paraffin mixture added of n-alkanes) and real-world samples proving the agreement of the results obtained by LC-GC × GC and the traditional mono-dimensional approach. Moreover, the benefits of using an entirely integrated platform were emphasized, particularly regarding the identity confirmation capability of the MS data, which can be easily translated into the 2D FID quantification feature.

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A mild mixed-solvent of n-hexane/isopropanol is proposed for extracting total mineral oil hydrocarbons (MOH) from commercial milk powder products. Unlike acid-hydrolysis, the mixed-solvent extraction was performed at ambient temperature and the low-boiling-point hydrocarbons were retained to the greatest extent. After extraction, total MOH was determined by on-line liquid chromatography-gas chromatography with a flame ionization detector (LC-GC-FID). The validation of the proposed extraction method revealed a recovery efficacy of 83.0-107.5% and a limit of quantification of 0.5 mg/kg. Then, the total MOH in ten commercial milk powders was determined and mineral oil saturated hydrocarbons (MOSH)/polyolefin oligomeric saturated hydrocarbons (POSH) were found to be within the range of 0.61-5.46 mg/kg. The comparison of the total and surface MOSH/POSH indicated that a major part of the contamination was derived from sources before packaging. The present study provides a robust method for the extraction and determination of total MOH in milk powders.

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The main focus of the present research was the on-line coupling of two separation techniques, namely liquid chromatography (LC) and gas chromatography (GC). For such an analytical combination, a dedicated interface is required to remove solvent from the sample, leaving the latter in a sharp band at the head of the GC column. Considering such an objective, a lab-developed LC-GC interface is herein presented, based on the use of a six-port two-position valve and a programmed-temperature-vaporizing (PTV) injector. The PTV injector was derived from a commercial split/splitless injector body, heated using a resistance heating wire, and enabled a satisfactory recovery of low boiling compounds (≤ C13), working in the normal-phase mode. The lab-developed PTV injector allowed the use of a larger-volume liner (compared to the commercial one initially used), it being characterized by dimensions 95 mm length × 5.0 mm O.D. × 3.4 mm I.D. and a volume of 862 µL, thus facilitating the transfer of larger LC fractions. The developed system is fully automatized and controlled without the use of additional software. The interface was evaluated and used for the analysis of mineral oil saturated hydrocarbons in vegetable oils. Detection was carried out by using a flame ionization detector (FID), with quantification performed through external calibration, across the 5-1000 mg kg-1 range. The LC-GC-FID method linearity, limits of detection and quantification, accuracy and precision were measured. The resulting limits of detection and quantification values were 0.4 and 1.3 mg kg-1, respectively. The average accuracy at the 100 mg kg-1 level was 95.5% (ranging between 93.3 and 99.7%). Intra-day repeatability at levels of 5 and 100 mg kg-1 were 2.4% and 3.5%, respectively.

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In this work, we study the effect of different variables affecting elution profile distortion on the enantiomeric resolution eventually achievable when working with on-line coupled liquid chromatography to gas chromatography (LC-GC). Specifically, the proposed configuration combines achiral reversed-phase liquid chromatography (RPLC) and chiral gas chromatography (enantio-GC), with heptakis-(2,3,6-tri-O-methyl)-ß-cyclodextrin as enantioselective stationary phase to analyse target fractions transferred (from LC to GC) via the through oven transfer adsorption desorption (TOTAD) interface. The high degree of orthogonality resulting from the combination of two chromatographic columns having very different separation mechanisms (and also requiring mobile phases in distinct physical states), as well as integration of the sample preparation step in the first dimension of the system, significantly contributed to exploit the performance of the proposed two-dimensional approach. Occasional adverse effects, which may result in severe peak distortions during LC-GC analysis and could be explained by flow instabilities due to viscous fingering, are circumvented by using the outstanding capacity of the TOTAD interface for achieving effective elimination of the eluent arriving from the LC preseparation.

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On-line high-performance liquid chromatography coupled with gas chromatography with flame ionization detector (LC-GC) has become an important technique for the determination of mineral oil aromatic hydrocarbons (MOAH) in foods. However, the analysis is complicated and time-consuming. The present study aimed to develop a fluorescence spectrophotometry approach for the fast screening of MOAH in grains. A reference material, which is a mixture of diesel oils and lubricants, was selected for MOAH calibration. The excitation/emission wavelength of EX230 nm/EM330 nm was determined and the linear range was 0.125-2.5 mg/kg with an R2 of 0.999. The MOAH content determined by the fluorescence method showed good correlation with that determined by LC-GC (r = 0.946). This result suggested that the performance of these two methods was comparable, and thus fluorescence spectroscopy can be used for screening MOAH. A MOAH content of >0.1 mg/kg in grains could be rapidly determined by fluorescence spectroscopy.

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An on-line liquid chromatography-comprehensive two-dimensional gas chromatography (LC-GC×GC) separation process, combined with a dual detection system, namely triple quadrupole mass spectrometry (used as a single quadrupole - QMS) and flame ionization detection (FID), was developed for the analysis of cosmetic lip care products. The LC step was carried out by using a fused-core silica column, with this enabling the separation of mineral oil saturated hydrocarbons (MOSH), as well as polyolefin oligomeric saturated hydrocarbons (POSH), from the mineral oil aromatic hydrocarbon (MOAH) families. Each chemical class was then subjected to GC×GC-QMS/FID analysis, using a medium polarity-low polarity column combination. Notwithstanding the utility of the flame ionization detector for quantification purposes, it is obviously also desirable to obtain information on the type of hydrocarbons present (of mineral or synthetic origin), in order to identify a potential contamination source. Following method optimization, various analytical figures of merit (method linearity, intra- and inter-day repeatability, limits of detection and quantification, and injector discrimination) were measured. Specifically, for MOSH and MOAH, linearities were evaluated in the 32.5-2080 and 50-500 mg L-1 range, respectively; furthermore, intra- and inter-day repeatability values were 6.2 and 8.3%, respectively, while limits of detection and quantification were 3.5 and 11.8 mg L-1, respectively. The proposed method, enables for the first time (to the best of the authors' knowledge), the detailed qualitative and quantitative analysis of saturated and aromatic hydrocarbons, in a single run and in a fully-automated manner. Quantities of MOSH + POSH in the cosmetic products ranged from 8.7 to 44.4% (CV values were in the 1.5-5.4% range) while MOAH contamination was found in four samples, ranging from 386 to 5869 mg kg-1 (CV values were in the 0.5-4.5% range).

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Recently, migration of mineral oil components from food contact materials into various foods has been reported. The analysis of mineral oil in food is complicated since it consists of mineral oil saturated hydrocarbons (MOSH) comprising a complex mixture of linear, branched and cyclic compounds and variable amounts of mineral oil aromatic hydrocarbons (MOAH), mainly alkylated. Both MOSH and MOAH form 'humps' of unresolved peaks in the chromatograms with the same range of volatility. Since these two fractions have a different toxicological relevance, it is important to quantify them separately. Occurrence data on mineral oil are available only for a limited number of food groups and only from few countries. In Belgium, data on the contamination of food by mineral oil are lacking. In this contribution, an in-house validated online combination of liquid chromatography with gas chromatography (LC-GC) with flame ionisation detection (FID) was used for the quantification of MOSH and MOAH. Totally, 217 packed food samples were selected using a well-defined sampling strategy that targeted food categories which are highly consumed and categories suspected to contain mineral oil. For 19 samples, the method was not applicable. For the 198 remaining samples, MOSH was detected in 142 samples with concentrations up to 84.82 mg kg-1. For the MOAH fraction, there are 175 samples with a concentration below the limits of quantification (LOQ), while 23 samples had a higher concentration ranging from 0.6 to 2.24 mg kg-1. Finally, these results were compared with the action thresholds as proposed by the Scientific Committee (SciCom) of the Belgian Food Safety Agency (FAVV-AFSCA). Only one sample exceeded the threshold for MOSH, while the threshold for MOAH was exceeded in 23 samples. For the samples exceeding the action threshold, further investigation is needed to identify the contamination source.

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In this work, a strategy of in-series combination of ultrasound-assisted extraction and online LC-GC/MS was constructed for effective separation and analysis of sucrose esters in tobacco. Sucrose esters were first extracted by ultrasound-assisted extraction with high efficiency and easyhandling. Online LC-GC/MS was then applied for sucrose ester clean-up and analysis. To better evaluate the effectiveness of this strategy, we limited our focus to five groups of sucrose ester isomers. Each group differed in mass from the next by 14 Da. The obtained coefficient of the calibration curve was 0.9986. Limit of detection (LOD) and limit of quantitation (LOQ) were 0.05 and 0.16 µg/ mL, respectively. The recovery was above 90% and the reproducibility was below 4%. This strategy was subsequently applied to the comparison of relative amounts of five groups of sucrose esters extracted from three different parts of aromatic tobacco. The satisfactory performance indicated that this strategy has great prospect for the rapid and high-throughput analysis of sucrose esters in tobacco.

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Mineral oil hydrocarbons (MOH) can be found in detectable levels in a multitude of foodstuffs. Therefore, chemical analysis of food for MOH gains importance. Different proficiency testing (PT) rounds on mineral oil testing have been performed in different matrices: cereals and rice as well as cardboard samples were examined. The laboratories participating in the PT rounds had to follow specific requirements for examination. The sample materials used contained different concentrations of mineral oil saturated hydrocarbons (MOSH) and mineral oil aromatic hydrocarbons (MOAH). The PT results were statistically evaluated according to ISO 13528:2005 and additionally the HorRat(R) value was calculated to gain information on the comparability of the mineral oil testing. It could be shown that for the examined sample materials and under the chosen specifications for testing a comparable determination of the mineral oil content is possible within the required relative standard deviations. A useful analytical determination can be achieved with an acceptable relative standard deviation of <50% from a concentration of defined mineral oil fractions at ≥1 mg/kg in food. In the concentration range for MOH in food of between 1 mg/kg and 2 mg/kg, relative standard deviations of 20-40% were achieved. MOH concentrations of ≥ 2 mg/kg food were determined with good relative standard deviations of around 20%. Moreover, due to the results gained within this work a statement concerning the comparability for MOSH and MOAH contents below concentrations of 1 mg/kg food is possible: under the chosen conditions for examination as part of this work, mineral oil determination below 1 mg/kg food showed high variability. To gain reliable information with regard to consumer protection on the risk of mineral oil contents in this low concentration range further standardisation of the test method is indicated.

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The contamination of food by mineral oil hydrocarbons (MOHs) found in packaging is a long-running concern. A main source of MOHs in foods is the migration of mineral oil from recycled board into the packed food products. Consequently, the majority of food manufacturers have taken protective measures, e.g., by using virgin board instead of recycled fibres and, where feasible, introducing functional barriers to mitigate migration. Despite these protective measures, MOHs may still be observed in low amounts in certain food products, albeit due to different entry points across the food supply chain. In this study, we successfully apply gas chromatography coupled to mass spectrometry (GC-MS) to demonstrate, through marker compounds and the profile of the hydrocarbon response, the possible source of contamination using mainly chocolate and cereals as food matrices. The conventional liquid chromatography-one-dimensional GC coupled to a flame ionisation detector (LC-GC-FID) is a useful screening method, but in cases of positive samples it must be complemented by a confirmatory method such as, for example, GC-MS, allowing a verification of mineral oil contamination. The procedural approach proposed in this study entails profile analysis, marker identification, and interpretation and final quantification.

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