Principles and applications of porous graphitic carbon stationary phase in liquid chromatography: An update.

The introduction of carbon black particles as packaging material for liquid chromatography columns dates back to the late 70's, in an attempt to overcome common drawbacks associated with silica-based packings. The latter consisted of the difficulty in eliminating or shielding the polar residual silanol groups, responsible for secondary interactions with non-polar ligands, but also the fragility and instability of the bonded ligands. Since then, numerous advances have been made in the synthesis of carbon-based stationary phases, achieving excellent objectives in terms of chromatographic performance and versatility, mainly related to the possibility of working under a wide range of pH (1-14) and temperature (higher than 200 °C). The purpose of this review is to summarize the most significant advances in the synthesis and application of the porous graphitic carbon phase (PGC), in the last decade. Literature reports based on the use of PGC columns are focused on the analysis of a wide range of chemicals, spanning from polar compounds to apolar polymers. More in detail, polar analytes have included both small molecules and larger biomolecules (such as oligo- and polysaccharides, peptides, and glycopeptides), with special emphasis on additional selectivity for isomer separation. On the other hand, applications devoted to the analysis of non-polar analytes could benefit from the use of high temperatures, allowing for the achievement of satisfactory separations within reduced analysis time.

Determination of Oxygen Heterocyclic Compounds in Foods Using Supercritical Fluid Chromatography-Tandem Mass Spectrometry.

The aim of this research was to determine oxygen heterocyclic compounds in twenty-six Citrus- and cinnamon-flavoured foods using supercritical fluid chromatography in combination with triple-quadrupole mass spectrometry (SFC-QqQ-MS). According to the authors' knowledge, this is the first report on the determination of these molecules in foods by means of the SFC-QqQ-MS technique. The analytical technique normally used for their determination in foods is liquid chromatography coupled with a photodiode array detector. However, supercritical fluid chromatography is proving to be a valid alternative approach to investigating coumarins, furocoumarins and polymethoxyflavones. According to the results presented herein, each sample analysed showed the presence of molecules of interest. Coumarin was found in all the cinnamon-flavoured samples analysed in a low concentration. The presence of oxygen heterocyclic compounds in all the Citrus-flavoured samples, according to the label, comfirmed that the foods selected for this research article were prepared with Citrus fruits. Among the samples analysed, mandarin juice was the richest in bioactive compounds, representing a good source of polymethoxyflavones in a diet.

Mechanisms underlying the anti-aging activity of bergamot (Citrus bergamia) extract in human red blood cells.

Introduction: Aging is a process characterised by a decline in physiological functions. Reactive species play a crucial role in the aging rate. Due to the close relationship between aging and oxidative stress, functional foods rich in phytochemicals are excellent candidates to neutralise age-related changes. Aim: This investigation aims to verify the potential protective role of bergamot (Citrus bergamia, Femminello cultivar) peel and juice extract in a model of aging represented by human red blood cells (RBCs) exposed to D-Galactose (DGal). Methods: Bergamot peel and juice extracts were subjected to RP-HPLC/PDA/MS for determination of their composition in bioactive compounds. Markers of oxidative stress, including ROS production, thiobarbituric acid reactive substances (TBARS) levels -a marker of lipid peroxidation, oxidation of total protein sulfhydryl groups, as well as the expression and anion exchange capability of band 3 and glycated haemoglobin (A1c) production have been investigated in RBCs treated with D-Gal for 24 h, with or without pre-incubation for 15 min with 5 µg/mL peel or juice extract. In addition, the activity of the endogenous antioxidant system, including catalase (CAT) and superoxide dismutase (SOD), as well as the diversion of the RBC metabolism from glycolysis towards the pentose phosphate pathway shunt, as denoted by activation of glucose-6-phosphate dehydrogenase (G6PDH), have been explored. Results: Data shown here suggest that bergamot peel and juice extract i) prevented the D-Gal-induced ROS production, and consequently, oxidative stress injury to biological macromolecules including membrane lipids and proteins; ii) significantly restored D-Gal-induced alterations in the distribution and ion transport kinetics of band 3; iii) blunted A1c production; iv) effectively impeded the over-activation of the endogenous antioxidant enzymes CAT and SOD; and v) significantly prevented the activation of G6PDH. Discussion: These results further contribute to shed light on aging mechanisms in human RBCs and identify bergamot as a functional food rich in natural antioxidants useful for prevention and treatment of oxidative stress-related changes, which may lead to pathological states during aging.

Direct analysis in real time of high-quality extra virgin olive oils for the rapid and automatic identification of origin trademark.

BACKGROUND: Following our previous research on the differentiation of Italian extra virgin olive oils (EVOOs) by rapid evaporative ionization mass spectrometry coupled to a tandem high resolution mass analyser, the present study deals with the evaluation of another direct mass spectrometry (direct-MS) approach for the rapid and automatic discrimination of EVOOs. In particular, direct analysis in real time (DART-MS) was explored as an ambient MS (AMS) source for the building of a top-quality Italian EVOOs database and fast identification of unknown samples. A single quadrupole detector (QDa) was coupled with DART, taking advantage of a cost-saving, user-friendly and less sophisticated instrumental setup. Particularly, quickstrip cards, located on a moving rail holder, were employed, allowing for the direct analysis of 12 EVOO spots in a total analysis time of 6 min. The aim was to develop a reliable statistical model by applying principal component and linear discriminant analyses to clusterize and classify EVOOs according to geographical origin and cultivar, as main factors determining their nutritional and sensory profiles. RESULTS: Satisfactory results were achieved in terms of identification reliability of unknown EVOOs, as well as false positive risk, thus confirming that the use of AMS combined with chemometrics is a powerful tool against fraudulent activities, without the need for mass accuracy data, which would increase the analysis cost. CONCLUSION: A DART ionization source with a compact and reliable QDa MS analyser allowed for rapid fingerprinting analysis. Furthermore, MS spectra provided quali-quantitative information successfully related to EVOO differentiation. © 2023 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Hidden threat lurking in extensive hand hygiene during the Covid-19 pandemic: investigation of sensitizing molecules in gel products by hyphenated chromatography techniques.

During the Covid-19 pandemic, health agencies worldwide have recommended frequent handwashing and sanitizing. A variety of hand gel products were made available on the market, often with fragrances added to curtail the strong smell of alcohol. Commonly used Citrus fragrances contain volatile aroma constituents and non-volatile oxygen heterocyclic compounds (OHCs), consisting mostly of polymethoxyflavones, coumarins, and furocoumarins. The latter have long been investigated for their phototoxic properties, and their safety as cosmetic product ingredients has been debated recurrently. To this concern, twelve commercial Citrus-scented products were investigated in this study. An extraction method was optimized for thirty-seven OHC compounds, obtaining absolute mean recovery values in the 73.5-116% range with only few milliliters of solvent consumption. Analysis by ultra-high-pressure liquid chromatography with tandem mass spectrometry detection evidenced that three samples did not conform to the labeling requirements for fragrance allergens (coumarin) laid down by the European Union Regulation on Cosmetic Products. The total furocoumarin (FC) content of the samples investigated was in the 0.003-3.7ppm range, with some noteworthy exceptions. Specifically, in two samples, the total FCs were quantified as 89 and 219 ppm, thus exceeding the safe limits recommended up to a factor of 15. Finally, the consistency of the volatile fingerprint attained by gas chromatography allowed drawing conclusions on the authenticity of the Citrus fragrances labeled, and several products did not conform to the information reported on the label concerning the presence of essential oils. Besides the issue of product authenticity, analytical tools and regulatory actions for widespread testing of hand hygiene products are urgent, to protect consumers' health and safety.

Development and validation of a multi-class analysis of pesticides in corn products by comprehensive two-dimensional liquid chromatography-tandem mass spectrometry.

Due to the growing trend of organic food, there is still concern over the use of chemicals and pesticides in agriculture. In recent years, several procedures have been validated for the control of pesticides in food. In the present research, a comprehensive two-dimensional liquid chromatography coupled with tandem mass spectrometry is proposed for the first time for a multi-class analysis of 112 pesticides in corn-based products. Notably, a "reduced" QuEChERS-based method as extraction and clean-up procedure prior to the analysis, was successfully employed. Limits of quantification values were lower than the ones fixed by the European legislation; intra-day and inter-day precision were lower than 12.9% and 15.1%, respectively (at the 500 µg/kg concentration levels). Over 70% of the analytes provided recoveries between 70% and 120% range (at 50, 500 and 1000 µg/kg concentration levels) with standard deviation values below 20%. In addition, matrix effect values were in the range between 13% to 161%. The method was applied to the analysis of real samples, and three pesticides were detected at trace levels in both samples. The findings of this work pave the way for the treatment of complex matrices such as corn products.

Supercritical Fluid Extraction of Oils from Cactus Opuntia ficus-indica L. and Opuntia dillenii Seeds.

This study aimed to assess the capability of supercritical fluid extraction (SFE) as an alternative and green technique compared to Soxhlet extraction for the production of oils from Opuntia ficus-indica (OFI) seeds originating from Yemen and Italy and Opuntia dillenii (OD) seeds from Yemen. The following parameters were used for SFE extraction: a pressure of 300 bar, a CO2 flow rate of 1 L/h, and temperatures of 40 and 60 °C. The chemical composition, including the fatty acids and tocopherols (vitamin E) of the oils, was determined using chromatographic methods. The highest yield was achieved with Soxhlet extraction. The oils obtained with the different extraction procedures were all characterized by a high level of unsaturated fatty acids. Linoleic acid (≤62% in all samples) was the most abundant one, followed by oleic and vaccenic acid. Thirty triacylglycerols (TAGs) were identified in both OFI and OD seed oils, with trilinolein being the most abundant (29-35%). Vanillin, 4-hydroxybenzaldehyde, vanillic acid, and hydroxytyrosol were phenols detected in both OFI and OD oils. The highest γ-tocopherol content (177 ± 0.23 mg/100 g) was obtained through the SFE of OFI seeds from Yemen. Overall, the results highlighted the potential of SFE as green technology to obtain oils suitable for functional food and nutraceutical products.

A Bacillus-based biofungicide agent prevents ochratoxins occurrence in grapes and impacts the volatile profile throughout the Chardonnay winemaking stages.

Bacillus-based biocontrol agents have emerged as a strategy to eliminate or reduce the use of synthetic fungicides that are detrimental to health and the environment. In vineyards, a special concern arises from the control of Aspergillus carbonarius, a fungus known for its potential to produce ochratoxins. Ochratoxin A (OTA) is the most toxic form among ochratoxins and its maximum limit in wine has been established in Europe and Brazil as 2 µg/kg. Wine quality, especially the volatile profile, may be influenced by the antifungal strategies, since fungicide residues are transferred from grapes to must during winemaking. The objective of this study was to evaluate, for the first time, the impact of a biocontrol strategy containing Bacillus velezensis P1 on the volatile profile and occurrence of ochratoxins when grapes infected with A. carbonarius were used in winemaking. The evaluation of ochratoxins was carried out by liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (LC-QToF-MS), and volatile compounds were analyzed using comprehensive two-dimensional gas chromatography coupled to quadrupole mass spectrometry (GC × GC/qMS). Six ochratoxins were identified in must prepared with Chardonnay grapes inoculated with A. carbonarius (ochratoxin α, ochratoxin ß, ochratoxin α methyl-ester, ochratoxin α amide, N-formyl-ochratoxin α amide and OTA). Although winemaking causes a decrease in the levels of all forms of ochratoxins, the co-occurrence of these mycotoxins was verified in wine made with grapes containing A. carbonarius. B. velezensis P1 prevented the occurrence of ochratoxins in must, ensuring the safety of wines. Regarding the volatile profile, a predominant presence of terpenic compounds was verified in samples treated with B. velezensis when compared with those not treated with the biocontrol strategy, whereas the presence of A. carbonarius resulted in a higher concentration of volatile compounds with an odor described as fatty/waxy, possibly compromising wine quality. Therefore, B. velezensis P1 is a new biofungicide possibility to produce ochratoxin-free grapes and high-quality wines.

Determination of the polyphenolic content of berry juices using focusing-modulated comprehensive two-dimensional liquid chromatography coupled to mass spectrometry detection.

In this work, a comprehensive two-dimensional liquid chromatography system, comprised of a ZIC-HILIC and C18 columns in the first and second dimensions, respectively, was tuned and employed for attaining high resolution profiles of the polyphenolic pattern in seven commercial berry juices. The developed HILIC × RP-LC method was validated in terms of linearity range, correlation coefficients, limit of detection, limit of quantification, precision (intra- and inter-day), and recovery. A total of 104 polyphenolic compounds belonging to different chemical classes (hydroxybenzoic and cinnamic acid derivatives, flavone glycosides, flavonols, flavonol glycosides, dihydroflavonols, and anthocyanin glycosides) have been characterized and quantified in the juices investigated. Despite the constituents being similar, a notable quantitative variation among the analyzed berry species was observed. Elderberry contained the highest amount of polyphenols (918 ± 1.10 mg 100 mL-1), followed by chokeberry (516 ± 0.08 mg 100 mL-1). On the other hand, raspberry contained the lowest amount (104 ± 1.21 mg 100 mL-1). Further, total phenolic, flavonoid, and anthocyanin contents were determined spectrophotometrically, yielding consistent results. The free-radical scavenging activity (DPPH test) and reducing power of the juices, expressed as IC50 (µL mL-1) and mg ASE mL-1, varied from 2.79 ± 0.03 (honeyberry) to 31.66 ± 0.02 (blueberry) and from 1.71 ± 0.01 (blueberry) to 8.89 ± 0.12 (chokeberry), respectively. Such a ZIC-HILIC × C18 platform based on focusing modulation, never employed so far for berry juices, showed a remarkable separation capability with high values of corrected peak capacity (up to 1372) and orthogonality (Ao up to 0.80), thus providing a great applicability to be advantageously employed for other complex food samples.

Determination of the polyphenolic content of Ammodaucus leucotrichus Cosson and Durieu by liquid chromatography coupled with mass spectrometry and evaluation of the antioxidant and antiglycation properties.

Ammodaucus leucotrichus Cosson and Durieu, known as Sahara cumin, is a plant belonging to the Apiaceae family with a very strong smell of anise growing in the maritime sands in the countries of North Africa. The present work aims to study the polyphenolic profile of its seeds hydroalcoholic extract along with the determination of the antioxidant and antiglycation properties. The phytochemical screening revealed the presence of 16 compounds, out of which 15 have been detected in this extract for the first time. Luteolin-glucoside turned out to be the most abundant one (281.32 ± 0.34 mg/kg), followed by apigenin-hexoside (235.06 ± 0.29 mg/kg) and luteolin (202.41 ± 0.40 mg/kg). In terms of antioxidant activity, a half-maximal inhibitory concentration value as high as 0.39 ± 0.003 mg AAE/ml (w/v) was attained. Further, the antiglycation activity was determined to yield interesting results: at a concentration of 1.5 mg/ml, the extract showed an antiglycation activity (%I) of 61.86 compared to metformin as a positive control (%I = 84.01); on the other hand, increasing the concentration to 10 mg/ml, the inhibition activity switched to advanced glycation end products formation activation (%I = 41.71).

Elucidation of the Lipid Composition of Hemp (Cannabis sativa L.) Products by Means of Gas Chromatography and Ultra-High Performance Liquid Chromatography Coupled to Mass Spectrometry Detection.

The growing demand in natural matrices that represent a source of dietary and nutraceutical molecules has led to an increasing interest in Cannabis sativa, considered to be a multipurpose, sustainable crop. Particularly, the considerable content in essential fatty acids (FAs) makes its derived-products useful food ingredients in the formulation of dietary supplements. In this research, the FA and triacylglycerol (TAG) composition of hempseed oils and flours were investigated using gas chromatography coupled to mass spectrometry and flame ionization detection as well as liquid chromatography coupled to mass spectrometry (LC-MS), respectively. Furthermore, a recently introduced linear retention index (LRI) approach in LC was successfully employed as a useful tool for the reliable identification of TAG species. A total of 30 FAs and 62 glycerolipids were positively identified in the investigated samples. Relative quantitative analyses confirmed linoleic acid as the most abundant component (50-55%). A favorable omega6/omega3 ratio was also measured in hemp-derived products, with the α-linolenic acid around 12-14%. Whereas, γ-linolenic acid was found to be higher than 1.70%. These results confirm the great value of Cannabis sativa as a source of valuable lipids, and the further improvement of the LRI system paves the way for the automatization of the identification process in LC.

Supercritical fluid chromatography-tandem mass spectrometry of oxygen heterocyclic compounds in Citrus essential oils.

Oxygen heterocyclic compounds are secondary metabolites mainly present in the non-volatile fraction of cold-pressed Citrus essential oils. Under this denomination are included coumarins, furocoumarins, and polymethoxyflavones. These compounds possess numerous beneficial properties for human health, but the ingestion of large amounts of coumarins is often related to toxic effects, whereas the phototoxicity caused by furocoumarins and UVA exposure has been well known for a long time. This research has been aimed at the validation of an analytical approach, based on supercritical fluid chromatography coupled to tandem mass spectrometry, for the analysis of OHCs in Citrus essential oils. Among eight columns tested, packed with different stationary phases, the pentafluorophenyl allowed the best baseline separation in 8 min and by using less than 10% of methanol. Calibration curves of twenty-eight standards (coumarins, furocoumarins, polymethoxyflavones) were constructed on spiked lemon distilled essential oil and the method was validated according to the EURACHEM guidelines, by calculating linearity, limit of detection (LoD), limit of quantification (LoQ), accuracy, intra-day, and inter-day precision. Specifically, recoveries were in the 80.0-118.6% range, regression coefficients were between 0.9904 and 0.9998, the LoDs were in the 0.0004-0.0470 mg kg-1 range, the LoQs were in the 0.0014-0.1536 mg kg-1 range, and coefficients of variation were between 0.3 and 2.6% (intra-day) and 1.1 and 7.4% (inter-day). The quantitative profiles of thirteen cold-pressed Citrus essential oils were determined.

Elucidation of Analytical-Compositional Fingerprinting of Three Different Species of Chili Pepper by Using Headspace Solid-Phase Microextraction Coupled with Gas Chromatography-Mass Spectrometry Analysis, and Sensory Profile Evaluation.

The aim of the present study was to determine the volatile compounds of three different species of chili peppers, using solid-phase microextraction (SPME) methods in combination with gas chromatography-mass spectrometry (GC-MS). The detection of marker aroma compounds could be used as a parameter to differentiate between species of chili peppers for their detection and traceability in chili pepper food. The sensorial contribution was also investigated to identify the predominant notes in each species and to evaluate how they can influence the overall aroma. Three different pepper species belonging to the Capsicum genus were analyzed: Chinense, Annuum, and Baccatum. A total of 269 volatile compounds were identified in these species of chili peppers. The Capsicum annum species were characterized by a high number of acids and ketones, while the Capsicum chinense and Capsicum baccatum were characterized by esters and aldehydes, respectively. The volatile profile of extra virgin olive oils (EVOOs) flavored with chili peppers was also investigated, and principal component analysis (PCA) and hierarchical cluster analysis (HCA) of the volatile profiles were demonstrated to be a powerful analytical strategy for building a model that highlights the potential of a volatile characterization approach for use in evaluating food traceability and authenticity.

Phytochemical Characterization of Rhus coriaria L. Extracts by Headspace Solid-Phase Micro Extraction Gas Chromatography, Comprehensive Two-Dimensional Liquid Chromatography, and Antioxidant Activity Evaluation.

Rhus coriaria L. (Anacardiaceae), commonly known as sumac, has been used since ancient times for many different applications, and nowadays is used mostly as a spice obtained from its in the Mediterranean and the Middle ground fruits and employed for flavoring and garnishing food, predominantly Eastern regions. Traditionally, sumac has been also used in popular medicine for the treatment of many ailments including hemorrhoids, wound healing, diarrhea, ulcers, and eye inflammation. Sumac drupes are indeed rich in various classes of phytochemicals including organic acids, flavonoids, tannins, and others, which are responsible of their powerful antioxidant capacity, from which treatment of many common diseases such as cardiovascular disease, diabetes, and cancer could benefit. In this work we evaluated the influence of fruit ripeness, conservation, and processing. To this aim, a phytochemical characterization of six different samples of Rhus coriaria L. was carried out. Specifically, headspace solid-phase micro extraction gas chromatography coupled to mass spectrometry and comprehensive two-dimensional liquid chromatography coupled to photodiode array and mass spectrometry detection, were employed. A total of 263 volatile compounds, including terpene hydrocarbons, acids, and aldehydes, as well as 83 polyphenolic compounds, mainly gallic acid derivatives, were positively identified. All samples showed a significant antioxidant activity by means of oxygen radical absorbance capacity, in line with their polyphenolic content and composition. Such findings set a solid ground to support the utilization of this plant as an attractive target for novel nutraceutical approaches and for drug discovery.

Non-psychoactive cannabinoids identification by linear retention index approach applied to a hand-portable capillary liquid chromatography platform.

The aim of the present research was the application of the linear retention index (LRI) system for the identification of non-psychoactive cannabinoids using a portable LC instrument. The miniaturization, viz. the use of very low quantities of mobile phase, enabled the development of a compact mobile system to be used for in situ analysis, also according to a green and cost-saving approach. In particular, new capillary LC (cap-LC) methods coupled with UV detection were developed for the analysis of extracts of Cannabis sativa L. Two setups were explored to achieve the efficient separation of twenty-four cannabinoids: a single column setup which exploited a sub-2 µm packing to increase the chromatographic resolution, and a dual-column setup based on the serial connection of two different stationary phases, each coupled to an UV detector. The latter allowed the determination of two LRI values for each analyte, thus increasing the identification power. Moreover, since two different wavelengths were used on the LED-based UV detectors, the ratio of the absorbances measured on each chromatographic trace represented a third identification criterion, thus fulfilling the recommendations of the Scientific Working Group for The Analysis of Seized Drugs (SWDRUG) about the categories of analytical techniques to be used and the minimum number of parameters required for the unambiguous identification of drugs. The obtained results could be used for the development of a novel analytical method for fast and automatic in situ forensic investigations and hemp breeding programs, also minimizing the consumption of both sample and solvent.

Comprehensive Chemical Characterization of Chia (Salvia hispanica L.) Seed Oil with a Focus on Minor Lipid Components.

A comprehensive chemical characterization of different lipid components, namely fatty acid composition after derivatization in fatty acid methyl esters (FAMEs), triacylglycerols (TAGs), phospholipids (PLs), free fatty acids (FFAs), sterols, carotenoids, tocopherols, and polyphenols in Chia seed oil, obtained by Soxhlet extraction, was reported. Reversed phase liquid chromatography (RP-LC) coupled to UV and mass spectrometry (MS) detectors was employed for carotenoids, polyphenols, and TAGs determination; normal phase-LC in combination with fluorescence detector (FLD) was used for tocopherols analysis; PL and FFA fractions were investigated after a rapid solid phase extraction followed by RP-LC-MS and NanoLC coupled to electron ionization (EI) MS, respectively. Furthermore, gas chromatography (GC)-flame ionization (FID) and MS detectors were used for FAMEs and sterols analysis. Results demonstrated a significant content of bioactive compounds, such as the antioxidant tocopherols (22.88 µg mL-1), and a very high content of essential fatty acids (81.39%), namely α-linolenic (62.16%) and linoleic (19.23%) acids. In addition, for the best of authors knowledge, FFA profile, as well as some carotenoid classes has been elucidated for the first time. The importance of free fatty acids in vegetable matrices is related to the fact that they can be readily involved in metabolic processes or biosynthetic pathways of the plant itself. For a fast and reliable determination of this chemical class, a very innovative and sensitive NanoLC-EI-MS analytical determination was applied.

On-line coupling of supercritical fluid extraction with enantioselective supercritical fluid chromatography-triple quadrupole mass spectrometry for the determination of chiral pesticides in hemp seeds: A proof-of-principle study.

The present research can be considered as a proof-of-principle study focused on the determination of chiral pesticides using a supercritical fluid extraction instrument coupled on-line with an enantioselective supercritical fluid chromatography-triple quadrupole mass spectrometry. To the best of Authors' knowledge, this is the first description of an on-line approach for the extraction and determination of chiral pesticides. Metalaxyl, benalaxyl and dimethenamid were investigated in nine hemp seed samples belonging to four varieties of Cannabis sativa; only in one case a pesticide was found at levels above the method limit of quantification (LoQ), though within the EU maximum residue level value. The figures-of-merit determined were linearity, precision, limit of detection (LoD), and LoQ. Regression coefficients were between 0.9856 and 0.9973, the LoDs were in the 0.04-0.41 µg kg-1 range, the LoQs were in the 0.12-1.38 µg kg-1 range, while coefficients of variation were between 1 and 3% (10 µg kg-1 level).

Dietary Intake of Coumarins and Furocoumarins through Citrus Beverages: A Detailed Estimation by a HPLC-MS/MS Method Combined with the Linear Retention Index System.

Official regulations concerning the maximum number of substances in food are introduced as a consequence of possible adverse effects, after oral administration. In this regard, analytical methods are necessary in order to determine specific targets. Among oxygen heterocyclic compounds (OHCs, that are furocoumarins, coumarins and polymethoxyflavones), only coumarin is subject to restriction by the Regulation (EC) No 1334/2008 of the European Parliament. Furocoumarins are known for their phototoxicity and other side effects due to their dietary intake; however, an official limit about the maximum content of these compounds in food is still missing. The lack of information about the real amount of these compounds in food is responsible for the conflicting opinions about the introduction of an official limit. The HPLC-MS/MS method here proposed, in combination with the linear retention index system, represents an innovative analytical strategy for the characterization of OHCs in citrus beverages. Several types of drinks were analysed in order to quantify 35 OHCs in total. This method is suitable for the quality control of OHCs in food and the obtained results may be considered as informative data useful for the regulatory authorities in the emission of new opinions and for a potential new regulation in this field.