Ultrasound-Assisted Extraction of Total Phenolic Compounds and Antioxidant Activity Evaluation from Oregano (Origanum vulgare ssp. hirtum) Using Response Surface Methodology and Identification of Specific Phenolic Compounds with HPLC-PDA and Q-TOF-MS/MS.

Oregano is native to the Mediterranean region and it has been reported to contain several phenolic compounds particularly flavonoids that have been related with multiple bioactivities towards certain diseases. Oregano is cultivated in the island of Lemnos where the climate promotes its growth and thus it could be further used in promoting local economy. The aim of the present study was to establish a methodology for the extraction of total phenolic content along with the antioxidant capacity of oregano by using response surface methodology. A Box-Behnken design was applied to optimize the extraction conditions with regard to the extraction time, temperature, and solvent mixture with the use of ultrasound-assisted extraction. For the optimized extracts, identification of the most abundant flavonoids (luteolin, kaempferol, and apigenin) was performed with an analytical HPLC-PDA and UPLC-Q-TOF MS methodology. The predicted optimal conditions of the statistical model were identified, and the predicted values confirmed. The linear factors evaluated, temperature, time, and ethanol concentration, all showed significant effect (p < 0.05), and the regression coefficient (R2) presented a good correlation between predicted and experimental data. Actual values under optimum conditions were 362.1 ± 1.8 and 108.6 ± 0.9 mg/g dry oregano with regard to total phenolic content and antioxidant activity based on 2,2-Diphenyl-1-picrylhydrazyl (DPPH) assay, respectively. Additionally, further antioxidant activities by 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid (ABTS) (115.2 ± 1.2 mg/g dry oregano), Ferric Reducing Antioxidant Power (FRAP) (13.7 ± 0.8 mg/g dry oregano), and Cupric Reducing Antioxidant Capacity (CUPRAC) (1.2 ± 0.2 mg/g dry oregano) assays were performed for the optimized extract. The extract acquired under the optimum conditions contain an adequate quantity of phenolic compounds that could be used in the production of functional foods by food enrichment procedure.

Thorough Investigation of the Phenolic Profile of Reputable Greek Honey Varieties: Varietal Discrimination and Floral Markers Identification Using Liquid Chromatography-High-Resolution Mass Spectrometry.

Honey is a highly consumed commodity due to its potential health benefits upon certain consumption, resulting in a high market price. This fact indicates the need to protect honey from fraudulent acts by delivering comprehensive analytical methodologies. In this study, targeted, suspect and non-targeted metabolomic workflows were applied to identify botanical origin markers of Greek honey. Blossom honey samples (n = 62) and the unifloral fir (n = 10), oak (n = 24), pine (n = 39) and thyme (n = 34) honeys were analyzed using an ultra-high-performance liquid chromatography hybrid quadrupole time-of-flight mass spectrometry (UHPLC-q-TOF-MS) system. Several potential authenticity markers were revealed from the application of different metabolomic workflows. In detail, based on quantitative targeted analysis, three blossom honey markers were found, namely, galangin, pinocembrin and chrysin, while gallic acid concentration was found to be significantly higher in oak honey. Using suspect screening workflow, 12 additional bioactive compounds were identified and semi-quantified, achieving comprehensive metabolomic honey characterization. Lastly, by combining non-targeted screening with advanced chemometrics, it was possible to discriminate thyme from blossom honey and develop binary discriminatory models with high predictive power. In conclusion, a holistic approach to assessing the botanical origin of Greek honey is presented, highlighting the complementarity of the three applied metabolomic approaches.

Development of a Wine Metabolomics Approach for the Authenticity Assessment of Selected Greek Red Wines.

Wine metabolomics constitutes a powerful discipline towards wine authenticity assessment through the simultaneous exploration of multiple classes of compounds in the wine matrix. Over the last decades, wines from autochthonous Greek grape varieties have become increasingly popular among wine connoisseurs, attracting great interest for their authentication and chemical characterization. In this work, 46 red wine samples from Agiorgitiko and Xinomavro grape varieties were collected from wineries in two important winemaking regions of Greece during two consecutive vintages and analyzed using ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UHPLC-QToF-MS). A targeted metabolomics methodology was developed, including the determination and quantification of 28 phenolic compounds from different classes (hydroxycinnamic acids, hydroxybenzoic acids, stilbenes and flavonoids). Moreover, 86 compounds were detected and tentatively identified via a robust suspect screening workflow using an in-house database of 420 wine related compounds. Supervised chemometric techniques were employed to build an accurate and robust model to discriminate between two varieties.

Honey Phenolic Compound Profiling and Authenticity Assessment Using HRMS Targeted and Untargeted Metabolomics.

Honey consumption is attributed to potentially advantageous effects on human health due to its antioxidant capacity as well as anti-inflammatory and antimicrobial activity, which are mainly related to phenolic compound content. Phenolic compounds are secondary metabolites of plants, and their content in honey is primarily affected by the botanical and geographical origin. In this study, a high-resolution mass spectrometry (HRMS) method was applied to determine the phenolic profile of various honey matrices and investigate authenticity markers. A fruitful sample set was collected, including honey from 10 different botanical sources (n = 51) originating from Greece and Poland. Generic liquid-liquid extraction using ethyl acetate as the extractant was used to apply targeted and non-targeted workflows simultaneously. The method was fully validated according to the Eurachem guidelines, and it demonstrated high accuracy, precision, and sensitivity resulting in the detection of 11 target analytes in the samples. Suspect screening identified 16 bioactive compounds in at least one sample, with abscisic acid isomers being the most abundant in arbutus honey. Importantly, 10 markers related to honey geographical origin were revealed through non-targeted screening and the application of advanced chemometric tools. In conclusion, authenticity markers and discrimination patterns were emerged using targeted and non-targeted workflows, indicating the impact of this study on food authenticity and metabolomic fields.

Authentication of Greek PDO Kalamata Table Olives: A Novel Non-Target High Resolution Mass Spectrometric Approach.

Food science continually requires the development of novel analytical methods to prevent fraudulent actions and guarantee food authenticity. Greek table olives, one of the most emblematic and valuable Greek national products, are often subjected to economically motivated fraud. In this work, a novel ultra-high-performance liquid chromatography-quadrupole time of flight tandem mass spectrometry (UHPLC-QTOF-MS) analytical method was developed to detect the mislabeling of Greek PDO Kalamata table olives, and thereby establish their authenticity. A non-targeted screening workflow was applied, coupled to advanced chemometric techniques such as Principal Component Analysis (PCA) and Partial Least Square Discriminant Analysis (PLS-DA) in order to fingerprint and accurately discriminate PDO Greek Kalamata olives from Kalamata (or Kalamon) type olives from Egypt and Chile. The method performance was evaluated using a target set of phenolic compounds and several validation parameters were calculated. Overall, 65 table olive samples from Greece, Egypt, and Chile were analyzed and processed for the model development and its accuracy was validated. The robustness of the chemometric model was tested using 11 Greek Kalamon olive samples that were produced during the following crop year, 2018, and they were successfully classified as Greek Kalamon olives from Kalamata. Twenty-six characteristic authenticity markers were indicated to be responsible for the discrimination of Kalamon olives of different geographical origins.

Quality and Authenticity Control of Fruit Juices-A Review.

Food fraud, being the act of intentional adulteration of food for financial advantage, has vexed the consumers and the food industry throughout history. According to the European Committee on the Environment, Public Health and Food Safety, fruit juices are included in the top 10 food products that are most at risk of food fraud. Therefore, reliable, efficient, sensitive and cost-effective analytical methodologies need to be developed continuously to guarantee fruit juice quality and safety. This review covers the latest advances in the past ten years concerning the targeted and non-targeted methodologies that have been developed to assure fruit juice authenticity and to preclude adulteration. Emphasis is placed on the use of hyphenated techniques and on the constantly-growing role of MS-based metabolomics in fruit juice quality control area.

Reflection of Socioeconomic Changes in Wastewater: Licit and Illicit Drug Use Patterns.

The economic crisis plaguing Greece was expected to impact consumption of pharmaceuticals and illicit drugs - a priori to an unknown extent. We quantified the change of use for various classes of licit and illicit drugs by monitoring Athens' wastewater from 2010 to 2014. A high increase in the use of psychoactive drugs was detected between 2010 and 2014, especially for antipsychotics (35-fold), benzodiazepines (19-fold), and antidepressants (11-fold). This directly reflects the perceived increase of incidences associated with mental illnesses in the population, as a consequence of severe socioeconomic changes. Other therapeutic classes, like antiepileptics, hypertensives, and gastric and ulcer drugs also showed an increase in use (from 2-fold increase for antiepileptics to 13-fold for hypertensives). In contrast, the overall use of antibiotics and NSAIDs decreased. For mefenamic acid, an almost 28-fold decrease was observed. This finding is likely related to the reduction in drug expenditure applied in public health. A 2-fold increase of methamphetamine use was detected, associated with a cheap street drug called ″sisa″ (related to marginal conducts), which is a health concern. MDMA (5-fold) and methadone (7-fold) use showed also an increase, while cocaine and cannabis estimates did not show a clear trend.

Multianalyte method for the determination of pharmaceuticals in wastewater samples using solid-phase extraction and liquid chromatography-tandem mass spectrometry.

A fast and sensitive multianalyte/multiclass high-performance reversed-phase liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous analysis of 89 pharmaceuticals in influent and effluent wastewater samples. The method developed consists of solid-phase extraction (SPE) using a hydrophilic-lipophilic-balanced polymer followed by LC-MS/MS with electrospray ionization in both positive mode and negative mode. The selected pharmaceuticals belong to different classes--analgesic/anti-inflammatory drugs, antibiotics, antiepileptics, ß-adrenoceptor-blocking drugs, lipid-regulating agents, statins, and many others. The influence of the mobile phase composition on the sensitivity of the method, and the optimum conditions for SPE in terms of analyte recovery were extensively studied. Chromatographic separation was performed on an Atlantis T3 (100 mm × 2.1 mm, 3-µm) column with a gradient elution using methanol-0.01% v/v formic acid as the mobile phase in positive ionization mode determination and methanol-acetonitrile-1 mM ammonium formate as the mobile phase in negative ionization mode determination. Recoveries for most of the compounds ranged from 50 to 120%. Precision, expressed as relative standard deviations, was always below 15%, and the method detection limits ranged from 1.06 ng/L (4-hydroxyomeprazole) to 211 ng/L (metformin). Finally, the method developed was applied to the determination of target analytes in wastewater samples obtained from the Psyttalia wastewater treatment plant, Athens, Greece. Although SPE of pharmaceuticals from wastewater samples and their determination by LC-MS/MS is a well-established technique, the uniqueness of this study lies in the simultaneous determination of a remarkable number of compounds belonging to more than 20 drug classes. Moreover, the LC-MS/MS method has been thoroughly optimized so that maximum sensitivity is achieved for most of the compounds, making the proposed method a valuable tool for pharmaceutical analysis in influent and effluent wastewater at the sub-nanogram per liter level.

Simultaneous determination of 148 pharmaceuticals and illicit drugs in sewage sludge based on ultrasound-assisted extraction and liquid chromatography-tandem mass spectrometry.

This paper describes the development and validation of a new method for the simultaneous determination of 148 substances in sewage sludge. The selected compounds belong to different classes of pharmaceuticals, including antibiotics, analgesic and/or anti-inflammatory drugs, antiepileptics, benzodiazepines, antipsychotics, and antidepressants, among others, and illicit drugs, including opiates, opioids, cocaine, amphetamines, cannabinoids, and their metabolites. As far as the authors are aware, this is the first method in the peer-reviewed literature covering such a large number of target drugs for determination in a complex matrix like sewage sludge. The method presented herein combines ultrasound-assisted extraction (USE) and liquid chromatography coupled to tandem mass spectrometry. Good analytical performance was achieved, with limit-of-detection values below 10 ng g(-1) d.w. for 91% of the analytes and absolute recovery in the range 50-110% for more than 77% of the studied compounds. A combination of methanol and acidified water, also containing EDTA, proved to be the optimum solvent mixture to perform the extractions. An extra solid-phase-extraction clean-up step was not required, substantially reducing sample-preparation time and solvent consumption. Finally, the developed method was applied to the analysis of different sewage-sludge samples from five wastewater treatment plants of Santorini Island (Greece). Out of the 148 target compounds, 36 were detected. Several compounds, including acetylsalicylic acid, citalopram, and ciprofloxacin among others, had maximum concentrations above 100 ng g(-1) d.w.

A comprehensive HRMS methodology using LC-(ESI)-/GC-(APCI)-QTOF MS complementary platforms for wide-scope target screening of >750 pesticides in olive oil.

This study presents the development and validation of a comprehensive high-resolution mass spectrometry (HRMS) methodology for the detection of 771 pesticides in olive oil, using liquid chromatography with electrospray ionization, operating in positive and negative mode, and gas chromatography with atmospheric-pressure chemical ionization in positive mode, both coupled to quadrupole-time-of-flight mass spectrometry (LC-(ESI)-/GC-(APCI)-QTOF MS). Special reference is made to the post-acquisition evaluation step, in which all LC/GC-HRMS analytical evidence (i.e. mass accuracy, retention time, isotopic pattern, MS/MS fragmentation) is taken into account in order to successfully identify the compounds. The sample preparation of the method involves a QuEChERS-based protocol, common for both techniques, differentiated only on the reconstitution step, making the method highly applicable in routine analysis. A smart evaluation of method's performance was carried out, with 65 representative analytes comprising the validation set. The method was validated in terms of linearity, accuracy, matrix effect and precision, while the limits of detection and quantification of the method were estimated. Finally, twenty Greek olive oil samples were analysed in both analytical platforms and the findings included the pesticides lambda-cyhalothrin, chlorpyrifos, phosphamidon, pirimiphos-methyl and esprocarb at low ng g-1 level.

Development and validation of a high-throughput headspace solid-phase microextraction gas chromatography-mass spectrometry methodology for target and suspect determination of honey volatiles.

The determination of volatile compounds is essential for the chemical characterisation of honey's aroma and its correlation to its sensory profile and botanical origin. The present study describes the development, optimization and validation of a new, simple and reliable method for the determination of volatile compounds in honey using headspace solid-phase microextraction combined with gas chromatography/mass spectrometry (HS-SPME-GC-MS). The optimization of the SPME conditions showed that the ratio of honey: water (2:1) and the incubation temperature (60 °C) are the most critical parameters. Gas chromatography was performed with medium polar Varian CP-Select 624 column and the experimental Retention Index for a number of compounds was determined as an additional identification feature for suspect analysis. The simultaneous use of four internal standards chlorobenzene, benzophenone, 2-pentanol and 4-methyl-2-pentanone and matrix matched calibration enhanced method accuracy achieving recoveries 73-114 % and repeatability ranging between 3.9 and 19 % relative standard deviations. Furthermore, the superiority of the HS-SPME to static head space technique was verified exhibiting four-to nine-fold higher sensitivity. Target and suspect screening were applied to 30 Greek honey samples and 53 volatile compounds belonging to different chemical classes, such as alkanes, aldehydes, ketones, alcohols, and esters were identified with quantified concentrations ranging between 3.1 µg kg-1 (Limonene) up to 20 mg kg-1 (Benzeneacetaldehyde). Among the new findings is the detection of Myrtenol in Greek pine honey and 2,3-butanediol in Greek oak honey. The developed analytical protocol can be a valuable tool in order to chemically characterize honey based on the volatile content.

Comparative Assessment of Antibiotic Residues Using Liquid Chromatography Coupled with Tandem Mass Spectrometry (LC-MS/MS) and a Rapid Screening Test in Raw Milk Collected from the North-Central Algerian Dairies.

Antibiotic residues in milk are a major health threat for the consumer and a hazard to the dairy industry, causing significant economic losses. This study aims to assess the presence of antibiotic residues in raw milk comparatively by a rapid screening test (BetaStar® Combo) and Liquid Chromatography coupled with Tandem Mass Spectrometry (LC-MS/MS). A total of 445 samples were collected from 3 dairy companies of north-central Algeria (Algiers, Blida, Boumerdes), and they were rapidly screened for ß-lactams and tetracyclines; 52 samples, comprising 34 positive tanker-truck milk and 18 negative bulk-tank milk were tested by LC-MS/MS, which revealed 90.4% were contaminated (n = 47) and 55.3% exceeded the Maximum Residue Limit (MRL). The ß-lactams as parent compounds and their metabolites were the most frequently detected with maximum value for cloxacillin (1231 µg/kg) and penicillin G (2062 µg/kg). Under field condition, the false-positive results, particularly for tetracyclines, seems to be related to milk samples displaying extreme acidity values (≥19°D) or fat-level fluctuations (2.7 g/100 mL and 5.6-6.2 g/100 mL). Despite a relatively low prevalence (7.64%) of residues using the rapid test, the detection by LC-MS/MS of flumequine (52 µg/kg), cefaclor (maximum 220 µg/kg) and metabolites of ß-lactams at high levels should lead to reflections on the control of their human and environmental toxicological effects.

FoodOmicsGR_RI. A Consortium for Comprehensive Molecular Characterisation of Food Products.

The national infrastructure FoodOmicsGR_RI coordinates research efforts from eight Greek Universities and Research Centers in a network aiming to support research and development (R&D) in the agri-food sector. The goals of FoodOmicsGR_RI are the comprehensive in-depth characterization of foods using cutting-edge omics technologies and the support of dietary/nutrition studies. The network combines strong omics expertise with expert field/application scientists (food/nutrition sciences, plant protection/plant growth, animal husbandry, apiculture and 10 other fields). Human resources involve more than 60 staff scientists and more than 30 recruits. State-of-the-art technologies and instrumentation is available for the comprehensive mapping of the food composition and available genetic resources, the assessment of the distinct value of foods, and the effect of nutritional intervention on the metabolic profile of biological samples of consumers and animal models. The consortium has the know-how and expertise that covers the breadth of the Greek agri-food sector. Metabolomics teams have developed and implemented a variety of methods for profiling and quantitative analysis. The implementation plan includes the following research axes: development of a detailed database of Greek food constituents; exploitation of "omics" technologies to assess domestic agricultural biodiversity aiding authenticity-traceability control/certification of geographical/genetic origin; highlighting unique characteristics of Greek products with an emphasis on quality, sustainability and food safety; assessment of diet's effect on health and well-being; creating added value from agri-food waste. FoodOmicsGR_RI develops new tools to evaluate the nutritional value of Greek foods, study the role of traditional foods and Greek functional foods in the prevention of chronic diseases and support health claims of Greek traditional products. FoodOmicsGR_RI provides access to state-of-the-art facilities, unique, well-characterised sample sets, obtained from precision/experimental farming/breeding (milk, honey, meat, olive oil and so forth) along with more than 20 complementary scientific disciplines. FoodOmicsGR_RI is open for collaboration with national and international stakeholders.

Determination of isoascorbic acid in fish tissue by hydrophilic interaction liquid chromatography-ultraviolet detection.

A new hydrophilic interaction liquid chromatographic (HILIC) method for the simultaneous determination of isoascorbic (IAA) and ascorbic acid (AA) was developed. The separation of IAA and AA was studied in various HILIC stationary phases and the influence of the composition of the mobile phase, the ionic strength and the column temperature to the chromatographic characteristics is presented. The final method used an aminopropyl column under isocratic elution with acetonitrile-100 mM ammonium acetate solution (90:10, v/v) at a flow rate of 0.4 mL/min and a detection wavelength of 240 nm. This method was validated and the calibration curves were found to be linear in the range of 1.0-65 microg/mL for both IAA and AA. The method limit of detection for IAA determination in fish tissue was 2.3 microg/g. Inter-day precision (as %RSD(R)) was ranged between 0.56% and 8.3% at three concentration levels, whereas the respected recoveries ranged between 82% and 98%. This method was applied to the determination of IAA (as additive E315) in frozen redfish samples. The hyphenation of the HILIC separation with a tandem mass spectrometer was also studied and the problems encountered with negative electrospray ionization under HILIC separation conditions are discussed.

Application of High Resolution Mass Spectrometric methods coupled with chemometric techniques in olive oil authenticity studies - A review.

Extra Virgin Olive Oil (EVOO), the emblematic food of the Mediterranean diet, is recognized for its nutritional value and beneficial health effects. The main authenticity issues associated with EVOO's quality involve the organoleptic properties (EVOO or defective), mislabeling of production type (organic or conventional), variety and geographical origin, and adulteration. Currently, there is an emerging need to characterize EVOOs and evaluate their genuineness. This can be achieved through the development of analytical methodologies applying advanced "omics" technologies and the investigation of EVOOs chemical fingerprints. The objective of this review is to demonstrate the analytical performance of High Resolution Mass Spectrometry (HRMS) in the field of food authenticity assessment, allowing the determination of a wide range of food constituents with exceptional identification capabilities. HRMS-based workflows used for the investigation of critical olive oil authenticity issues are presented and discussed, combined with advanced data processing, comprehensive data mining and chemometric tools. The use of unsupervised classification tools, such as Principal Component Analysis (PCA) and Hierarchical Clustering Analysis (HCA), as well as supervised classification techniques, including Linear Discriminant Analysis (LDA), Support Vector Machine (SVM), Partial Least Square Discriminant Analysis (PLS-DA), Orthogonal Projection to Latent Structure-Discriminant Analysis (OPLS-DA), Counter Propagation Artificial Neural Networks (CP-ANNs), Self-Organizing Maps (SOMs) and Random Forest (RF) is summarized. The combination of HRMS methodologies with chemometrics improves the quality and reliability of the conclusions from experimental data (profile or fingerprints), provides valuable information suggesting potential authenticity markers and is widely applied in food authenticity studies.

Liquid Chromatographic Determination of Biogenic Amines in Fish Based on Pyrene Sulfonyl Chloride Pre-Column Derivatization.

Monitoring of biogenic amines in food is important for quality control, in terms of freshness evaluation and even more for food safety. A novel and cost-effective method was developed and validated for the determination of the main biogenic amines: histamine, putrescine, cadaverine, spermidine and spermine in fish tissues. The method includes extraction of amines with perchloric acid, pre-column derivatization with Pyrene Sulfonyl Chloride (PSCl), extraction of derivatives with toluene, back-dissolution in ACN after evaporation and determination by reversed phase high performance liquid chromatography with UV and intramolecular excimer fluorescence detection. The structure of the pyrene-derivatives was confirmed by liquid chromatography-mass spectrometry with electrospray ionization. The standard addition technique was applied for the quantitation due to significant matrix effect, while the use of 1,7-diaminoheptane as internal standard offered an additional confirmation tool for the identification of the analytes. Method repeatability expressed as %RSD ranged between 7.4-14% for the different amines and recovery ranged from 67% for histamine up to 114% for spermine. The limits of detection ranged between 0.1-1.4 mg kg-1 and the limits of quantification between 0.3-4.2 mg kg-1. The method was applied to canned fish samples and the concentrations of the individual biogenic amines were below the detection limit up to 40.1 mg kg-1, while their sum was within the range 4.1-49.6 mg kg-1.

Multi-residue methodology for the determination of 16 coccidiostats in animal tissues and eggs by hydrophilic interaction liquid chromatography - Tandem mass spectrometry.

A simple, sensitive and efficient confirmatory method was developed and validated for the determination of 16 coccidiostats in animal tissues and eggs using hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS). The sample preparation consisted of a solid-liquid extraction with ACN and dispersive SPE cleanup with MgSO4 and C18. Analysis was realized in an Acquity BEH HILIC silica column, in SRM mode. Both positive and negative ionization was performed, using polarity switching. Isocratic elution was used with a mobile phase of ACN: aqueous ammonium formate 1 mM with 0.1% formic acid (80:20, v/v). Method validation was performed in eggs, poultry, bovine, ovine, porcine and rabbit tissue and exceptionally low LODs were achieved, varying from 0.004 µg kg-1 (decoquinate in porcine tissue) to 0.560 µg kg-1 (halofuginone in eggs). The developed methodology was applied in 82 muscle and egg samples through the Greek National Residue Control Plan for coccidiostats.

Ionization study and simultaneous determination of avermectins and milbemycines in fish tissue by LC-ESI-MS/MS.

In this study, the simultaneous determination of avermectins (emamectin, eprinomectin, abamectin, doramectin and ivermectin) and milbemycines (moxidectin) in fish tissue with LC-ESI-MS/MS, was studied. Optimum chromatographic separation of target analytes was achieved using a Waters Acquity UPLC BEH C18 (100 mm × 2.1 mm, 1.7 mm) analytical column, operated at 40 °C and the composition of the mobile phase used was (A): ACN-MeOH (0.1% HCOOH) (1:1) and (B): 1 mM HCOONH4 (0.1% HCOOH). Various mobile phases were tested and the effect of the mobile phase composition on the analytes ionization was thoroughly examined in an extensive ionization study, aiming to increase the analytes' sensitivity. Deuterated ivermectin (IVR-d2) was used as an internal standard (IS). Avermectin's and milbemycine's extraction from the fish matrix was conducted with acidified ACN (0.1% HCOOH), followed by QuEChERS methodology. The method developed herein was validated according to the European Legislation requirements (Commission Decision 657/2002/EC) and recoveries ranged from 86 to 106% for all target analytes, with relative standard deviations < 20%. LODs ranged from 0.07 µg/kg (emamectin) to 1.3 µg/kg (doramectin), indicating the excellent sensitivity of the method. The developed methodology was successfully applied to fish samples obtained from aquaculture.

Targeted and Untargeted Metabolomics as an Enhanced Tool for the Detection of Pomegranate Juice Adulteration.

Pomegranate juice is one of the most popular fruit juices, is well-known as a "superfood", and plays an important role in healthy diets. Due to its constantly growing demand and high value, pomegranate juice is often targeted for adulteration, especially with cheaper substitutes such as apple and red grape juice. In the present study, the potential of applying a metabolomics approach to trace pomegranate juice adulteration was investigated. A novel methodology based on high-resolution mass spectrometric analysis was developed using targeted and untargeted screening strategies to discover potential biomarkers for the reliable detection of pomegranate juice adulteration from apple and red grape juice. Robust classification and prediction models were built with the use of unsupervised and supervised techniques (principal component analysis (PCA) and partial least squares discriminant analysis (PLS-DA)), which were able to distinguish pomegranate juice adulteration to a level down to 1%. Characteristic m/z markers were detected, indicating pomegranate juice adulteration, and several marker compounds were identified. The results obtained from this study clearly demonstrate that Mass Spectrometry (MS)-based metabolomics have the potential to be used as a reliable screening tool for the rapid determination of food adulteration.

Analysis of 76 veterinary pharmaceuticals from 13 classes including aminoglycosides in bovine muscle by hydrophilic interaction liquid chromatography-tandem mass spectrometry.

A multiresidue/multiclass method for the simultaneous determination of 76 veterinary drugs and pharmaceuticals in bovine muscle tissue has been developed and validated according to the requirements of European Commission Decision 2002/657/EC. The analytes belong in 13 different classes, including aminoglycoside antibiotics, whose different physicochemical properties (extremely polar character) render their simultaneous determination with other veterinary drugs quite problematic. The method combines a two-step extraction procedure (extraction with acetonitrile followed by an acidic aqueous buffer extraction) with hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) determination, allowing confirmation and quantification in a single chromatographic run. Further cleanup with solid phase extraction was performed using polymeric SPE cartridges. A thorough ionization study of aminoglycosides was performed in order to increase their sensitivity and significant differences in the abundance of the precursor ions of the analytes were revealed, depending on the composition of the mobile phase tested. Further gradient elution optimization and injection solvent optimization were performed for all target analytes.The method was validated according to the European Commission Decision 2002/657. Quantitative analysis was performed by means of standard addition calibration. Recoveries varied from 37.4% (bromhexine) to 106% (kanamycin) in the lowest validation level and 82% of the compounds showed recovery >70%. Detection capability (CCß) varied from 2.4 (salinomycin) to 1302 (apramycin) µgkg(-1).