Application of LC–MS/MS in the Mycotoxins Studies

The editors are really grateful to all the authors who contributed their high-quality works to this Special Issue. Special thanks go to the expert peer reviewers, whose rigorous evaluations of all of the submitted manuscripts have, no doubt, improved this Special Issue. Last but not least, the editors really appreciate the good organization and constant support of the MDPI editorial team and staff

Cooperative learning at the Analytical Chemistry laboratory

[SPA] El aprendizaje cooperativo es un sistema de organización del trabajo y de motivación, en el que el alumno es responsable de su aprendizaje y el de sus compañeros. A diferencia de los sistemas individualista y competitivo, las metas son grupales. La aplicación sistemática de este sistema da respuesta a tres principios básicos del Espacio Europeo de Educación Superior (EEES): participación y autonomía del estudiante, utilización de metodologías activas y papel del profesorado como agente facilitador del aprendizaje. Brevemente, con esta metodología se divide la clase en pequeños equipos, que reciben unas consignas de actuación a partir de las cuales planifican el trabajo del grupo. Cada miembro del grupo será responsable de áreas específicas que será necesario realizar satisfactoriamente para el éxito del grupo. En esta comunicación se plantea la aplicación del aprendizaje cooperativo para el aprovechamiento efectivo de las clases prácticas de laboratorio de análisis químico, dirigido a alumnos de último curso de la titulación de Ingeniería Química. La práctica en concreto consiste en la verificación de un equipo de HPLC-UV/Vis, para comprobar la conformidad con las especificaciones del fabricante. Además, esta metodología permite trabajar competencias transversales como razonamiento crítico, trabajo en equipo, gestión de proyectos y comunicación entre otras. [ENG] Cooperative learning is a work organizational system to motivate students, who become responsible for their own and their team mates learning. Unlike the individualistic and competitive learning styles, in cooperative learning student achievements occurs only when the other students in the group also achieve the objectives and rewards. Systematic application of this methodology helps reaching three basic principles of the European Higher Education Area (EHEA): student participation and autonomy, active methodologies application and lecturer role as facilitator in the learning process. In short, this methodology involves splitting up the class into small teams, which have to organize their self-workload to complete assignments and reach objectives previously established. Each team member is responsible for specific tasks that must be properly completed for the final success of the group. In this communication we propose the cooperative learning for effective laboratory lessons of analytical chemistry, aimed to last year students of chemical engineering. The specific assignment involves the verification of a whole equipment of liquid chromatography (HPLC) with spectrophotometric detection (UV-Vis) to check compliance with supplier specifications. In addition to specific competences, applying cooperative learning approach, students develop generic competences such as critical thinking, working with other, project management and communication among others

Determination of Aflatoxins by Liquid Chromatography Coupled to High-Resolution Mass Spectrometry

The most common mycotoxins are aflatoxins (AFs), which are produced by strains of various species of molds in the genus Aspergillus (A. flavus, A. parasiticus, A. nomius and A. tamarii) and can grow on many foods, mainly peanuts, maize and cottonseed. AFs are currently considered to be the most hazardous mycotoxins to health, in particular because of their hepatocellular carcinogenic potential. The main aflatoxins are B1 (AFB1), B2 (AFB2), G1 (AFG1) and G2 (AFG2) although many other derivatives have been described. In addition, animals consuming contaminated feeds are able to metabolize them by hydroxylation in a certain position, yield for example aflatoxin M1 (AFM1) and aflatoxin M2 (AFM2) from AFB1 and AFB2, respectively. Nowadays, only the four main AFs and one hydroxylated metabolite (AFM1) are routinely analyzed. High resolution mass spectrometry (HRMS) using Orbitrap or time-of-flight (TOF) mass analysers is a trend for AFs determination, allowing to determine AFs and their derivatives for which there are no commercial standards available, in order to carry out metabolization studies, exposure assessment or monitoring modified AFs in food. The aim of this study is to show the recent trends in analytical methods based on LC-HRMS for determination of AFs

An integrated targeted and untargeted approach for the analysis of ergot alkaloids in cereals using UHPLC - hybrid quadrupole time-of-flight mass spectrometry

An ultra-high performance liquid chromatography hybrid quadrupole time of flight (Q-TOF) mass spectrometry (MS) method is described for the simultaneous quantitative determination of common ergot alkaloids and the screening, detection and identification of unexpected (less studied or novel) members of this class of toxic fungal secondary metabolites. The employed analytical strategy involves an untargeted data acquisition (consisting of full scan TOF MS survey and information dependent acquisition MS/MS scans) and the processing of data using both targeted and untargeted approaches. Method performance characteristics for the quantitative analysis of 6 common ergot alkaloids i.e. ergometrine, ergosine, ergotamine, ergocornine, ergocristine, ergokryptine and their corresponding epimers in rye were comparable to those previously reported for triple-quadrupole (QqQ) MS/MS. The method limits of quantification (LOQ) were in the range from 3 to 19 mu g/kg, and good linearity was observed for the different ergot alkaloids in the range from LOQ to 1000 mu g/kg. Furthermore, the method demonstrated good precision (relative standard deviations at 50 mu g/kg not higher than 14.6 and 16.2% for the intra-day and inter-day precision, respectively), and the trueness values at different concentration levels were all between 89 and 115%. The method was applied for the analysis of a set of 17 rye samples and demonstrated the presence of these ergot alkaloids in the range from <LOQ to 2,811 mu g/kg. Further mining of the same data based on a 'non-targeted peak finding' algorithm and the use of full MS and MS/MS accurate mass data allowed the detection and identification of 19 ergot alkaloids that are commonly not included in most analytical methods using QqQ instruments. Some of these alkaloids are reported for the first time in naturally contaminated samples

Effect of Allium Extract Supplementation on Egg Quality, Productivity, and Intestinal Microbiota of Laying Hens

Simple Summary: The growing interest in phytogenic products for use in feed, especially in the poultry sector, is mainly due to the improvement in the productivity parameters and gut microbiota modulation properties. For this reason, phytogenic products are becoming excellent candidates as alternatives to the use of antibiotics in animal production to mitigate the negative effects derived from their use. The aim of this study is to explore the ability of allium extract (containing garlic and onion), used as an ingredient in laying hen feed, to improve performance. The promising results obtained in the present study suggest that Allium spp. extracts had the potential to be used in feeding laying hens to improve productivity, without affecting egg quality, and to modulate the gut microbiota. Abstract: The use of allium extract containing propyl propane thiosulfonate (PTSO) as hen feed supplement was evaluated to demonstrate its positive effect on egg production and intestinal microbiota modulation. The study was carried out on 90 laying hens whose feed was supplemented with allium extract for 28 days. Nutritional properties of eggs were not affected, whereas an improvement in productivity was observed based on the increase weight of eggs. In addition, a modulator effect on intestinal microbiota was confirmed by the increase in Lactobacillus spp. and Bifidobacterium spp., as well as by the reduction in Enterobacteriaceae populations. Finally, the preservation of egg composition was checked by monitoring the content of PTSO, using a new analytical method consisting of the use of solid phase extraction and ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS). Consequently, based on current results, Allium spp. extract rich in organosulfur compounds such as PTSO added to the diet had a beneficial effect on the microbiota and would seem to be a possible alternative to increase productivity, while not affecting the biochemical composition of egg. However, further studies on the effects of allium extract as feed supplement are necessary.DOMCA S.A.University of Granada FQM-302University of Murcia R-1418/201

Occurrence of Ergot Alkaloids in Barley and Wheat from Algeria

The natural occurrence of six major ergot alkaloids, ergometrine, ergosine, ergotamine, ergocornine, ergokryptine and ergocristine, as well as their corresponding epimers, were investigated in 60 cereal samples (barley and wheat) from Algeria. Ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) and a QuEChERS extraction method were used for sample analysis. The results revealed that 12 out of 60 samples (20%) were contaminated with ergot alkaloids. Wheat was the most contaminated matrix, with an incidence of 26.7% (8 out of 30 samples). The concentration of total ergot alkaloids ranged from 17.8 to 53.9 µg/kg for barley and from 3.66 to 76.0 µg/kg for wheat samples. Ergosine, ergokryptine and ergocristine showed the highest incidences in wheat, while ergometrine was the most common ergot in barley.Spanish Ministry of Science, Innovation and Universities (Project Ref.: RTI2018-097043-B-I00

Occurrence of Mycotoxins in Swine Feeding from Spain

A survey including 228 pig feed samples from Spain has been developed, exploring the occurrence of 19 mycotoxins (aflatoxins B1, B2, G1 and G2, ochratoxin A, fumonisins B1 and B2, citrinin, zearalenone, deoxynivalenol, fusarenon X, sterigmatocystin, T-2 toxin, HT-2 toxin, enniatins A, A1, B and B2, and beauvericin). The samples were analysed by solid-liquid extraction followed by liquid chromatography coupled with fluorescence or mass spectrometry detection. Enniatin B was found in 100% of the samples (up to 1200 ug/kg) and beauvericin in more than 90%. Moreover, 40% of samples were contaminated with more than five mycotoxins. This high occurrence is insurmountable and surpasses all previous studies, probably due to the inclusion of emerging mycotoxins, scarcely explored. The majority of the samples (96.9%) were in accordance with EU regulations, which do not address emerging mycotoxins or co-occurrence. These results show that in order to ensure mycotoxin absence, emerging mycotoxins should always be considered.This research was funded by SPANISH MINISTRY OF ECONOMY AND COMPETITIVENESS and EUROPEAN REGIONAL DEVELOPMENT FUND (MINECO/FEDER, UE), grant number AGL2015-70708-R

Multi-Mycotoxin Occurrence and Exposure Assessment Approach in Foodstuffs from Algeria

A survey on 120 cereal samples (barley, maize, rice and wheat) from Algerian markets has been carried out to evaluate the presence of 15 mycotoxins (ochratoxin A, deoxynivalenol, fumonisin B1 and B2, T-2 and HT-2 toxins, zearalenone, fusarenon X, citrinin, sterigmatocystin, enniatins A, A1, B and B1, and beauvericin). With this purpose, a QuEChERS-based extraction and ultra-high performance liquid chromatography coupled to tandem mass spectrometry (UHPLC-MS/MS) were used. Analytical results showed that 78 cereal samples (65%) were contaminated with at least one toxin, while 50% were contaminated with three to nine mycotoxins. T-2 toxin, citrinin, beauvericin and deoxynivalenol were the most commonly found mycotoxins (frequency of 50%, 41.6%, 40.8% and 33.3%, respectively). Fumonisins (B1 + B2), enniatins B and B1, deoxynivalenol and zearalenone registered high concentrations (289–48878 ug/kg, 1.2–5288 ug/kg, 15–4569 ug/kg, 48–2055 ug/kg and 10.4–579 ug/kg, respectively). Furthermore, concentrations higher than those allowed by the European Union (EU) were observed in 21, 8 and 1 samples for fumonisins, zearalenone and deoxinivalenol, respectively. As a conclusion, the high levels of fumonisins (B1 + B2) in maize and deoxynivalenol, zearalenone and HT-2 + T-2 toxins in wheat, represent a health risk for the average adult consumer in Algeria. These results pointed out the necessity of a consistent control and the definition of maximum allowed levels for mycotoxins in Algerian foodstuffs.This research was funded by SPANISH MINISTRY OF ECONOMY AND COMPETITIVENESS and EUROPEAN REGIONAL DEVELOPMENT FUND (MINECO/FEDER, UE) (Project ref: AGL2015-70708-R, MINECO/FEDER, UE). Choukri Khelifa Mahdjoubi would like to thanks the University of Granada for ERASMUS Mundus Doctoral Fellowship

Aphids transform and detoxify the mycotoxin deoxynivalenol via a type II biotransformation mechanism yet unknown in animals

Biotransformation of mycotoxins in animals comprises phase I and phase II metabolisation reactions. For the trichothecene deoxynivalenol (DON), several phase II biotransformation reactions have been described resulting in DON-glutathiones, DON-glucuronides and DON-sulfates made by glutathione-S-transferases, uridine-diphosphoglucuronyl transferases and sulfotransferases, respectively. These metabolites can be easily excreted and are less toxic than their free compounds. Here, we demonstrate for the first time in the animal kingdom the conversion of DON to DON-3-glucoside (DON-3G) via a model system with plant pathogenic aphids. This phase II biotransformation mechanism has only been reported in plants. As the DON-3G metabolite was less toxic for aphids than DON, this conversion is considered a detoxification reaction. Remarkably, English grain aphids (Sitobion avenae) which cooccur with the DON producer Fusarium graminearum on wheat during the development of fusarium symptoms, tolerate DON much better and convert DON to DON-3G more efficiently than pea aphids (Acyrthosiphon pisum), the latter being known to feed on legumes which are no host for F. graminearum. Using a non-targeted high resolution mass spectrometric approach, we detected DON-diglucosides in aphids probably as a result of sequential glucosylation reactions. Data are discussed in the light of an eventual co-evolutionary adaptation of S. avenae to DON

Thermal desorption-ion mobility spectrometry: A rapid sensor for the detection of cannabinoids and discrimination of Cannabis Sativa L. chemotypes

Existing analytical techniques used for the determination of cannabinoids in Cannabis sativa L. (Cannabis) plants mostly rely on chromatography-based methods. As a rapid alternative for the direct analysis of them, thermal desorption (TD)-ion mobility spectrometry (IMS) was used for obtaining spectral fingerprints of single cannabinoids from Cannabis plant extracts and from plant residues on hands after their manipulation. The ionization source was 63Ni, with automatic switchable polarity. Although in both ionization modes there were signals in the TD-IMS spectra of the plant extracts and residues that could be assigned to concrete cannabinoids and chemotypes, most of them could not be clearly distinguished. Alternatively, the global spectral data of the plant extracts and residues were pre-processed and then, using principal component analysis (PCA)-linear discriminant analysis (LDA), grouped in function of their chemotype in a more feasible way. Using this approach, the possibility of false positive responses was also studied analyzing other non-Cannabis plants and tobacco, which were clustered in a different group to those of Cannabis. Therefore, TD-IMS, as analytical tool, and PCA-LDA, as a strategy for data reduction and pattern recognition, can be applied for on-site chemotaxonomic discrimination of Cannabis varieties and detection of illegal marijuana since the IMS equipment is portable and the analysis time is highly short