Caffeine Decreases Topiramate Levels in Zebrafish Larvae in a Pentylenetetrazol-Induced Seizure Model.

Epilepsy ranks as the second-most prevalent neurological disease, and is characterized by seizures resulting in neurobiological and behavioral impairment. Naturally occurring in coffee beans or tea leaves, the alkaloid caffeine (CAF) is the most prevalent global stimulant. Caffeine has been observed to influence epileptic seizures and the efficacy of antiepileptic medications, with a notable impact on topiramate (TPM). This study aimed to explore the influence of CAF on TPM's anticonvulsant effects in zebrafish larvae within a PTZ-induced seizure model, concurrently determining TPM concentrations through a sophisticated analytical approach based on ultrahigh-performance liquid chromatography and subsequent mass spectrometric detection. Zebrafish larvae four days post-fertilization were incubated for 18 h with varying doses of TPM or combinations of CAF + TPM, and locomotor activity was then assessed. Seizures were induced by introducing a PTZ solution to achieve a final concentration of 20 mM. Utilizing liquid chromatography-mass spectrometry (LC-MS/MS), TPM levels in the larvae were quantified. CAF co-administration (especially in higher doses) with TPM caused a decrease in the average locomotor activity in the larvae compared to TPM alone. Moreover, CAF decreased TPM levels in the larvae at all investigated doses. In conclusion, these findings offer a novel perspective on the interplay between CAF and TPM, shedding light on previously unexplored facets. The potential impact of CAF consumption in assisting with epileptic seizure control, unless proven otherwise, suggests a noteworthy consideration for future research and clinical practices.

LC-MS Metabolomic Profiling of Five Types of Unrefined, Cold-Pressed Seed Oils to Identify Markers to Determine Oil Authenticity and to Test for Oil Adulteration.

The authenticity of food products marketed as health-promoting foods-especially unrefined, cold-pressed seed oils-should be controlled to ensure their quality and safeguard consumers and patients. Metabolomic profiling using liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (LC-QTOF) was employed to identify authenticity markers for five types of unrefined, cold-pressed seed oils: black seed oil (Nigella sativa L.), pumpkin seed oil (Cucurbita pepo L.), evening primrose oil (Oenothera biennis L.), hemp oil (Cannabis sativa L.) and milk thistle oil (Silybum marianum). Of the 36 oil-specific markers detected, 10 were established for black seed oil, 8 for evening primrose seed oil, 7 for hemp seed oil, 4 for milk thistle seed oil and 7 for pumpkin seed oil. In addition, the influence of matrix variability on the oil-specific metabolic markers was examined by studying binary oil mixtures containing varying volume percentages of each tested oil and each of three potential adulterants: sunflower, rapeseed and sesame oil. The presence of oil-specific markers was confirmed in 7 commercial oil mix products. The identified 36 oil-specific metabolic markers proved useful for confirming the authenticity of the five target seed oils. The ability to detect adulterations of these oils with sunflower, rapeseed and sesame oil was demonstrated.

Identification of sunflower, rapeseed, flaxseed and sesame seed oil metabolomic markers as a potential tool for oil authentication and detecting adulterations.

Testing the composition, quality and authenticity of edible oils is crucial to safeguard the consumers' rights and health. The aim of our study was to identify oil-specific markers to enable the differentiation and authentication of sunflower, sesame, flaxseed and rapeseed oils, and to evaluate their antioxidant activity, total phenolic and carotenoid content. A metabolomic approach based on liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry was employed for marker discovery. Spectrophotometric method was used for determination of antioxidant activity, total phenolic and carotenoid content. 76 oil samples from the four different manufacturers were examined. We identified 13 oil-specific markers for sunflower seed oil, 8 for rapeseed oil, 5 for sesame seed oil and 3 for flaxseed oil, their retention times, accurate masses, and characteristic fragment ions are reported. The abundances of the markers for each plant species were found to vary depending on the oil producer and the product batch. Significant differences in antioxidant activity, total phenolic and carotenoid content were also observed both between oils and within oil type. The highest total phenolic content (84.03 ± 4.19 to 103.79 ± 3.67 mg of gallic acid/kg) and antioxidant activity (245.67 ± 7.59 to 297.22 ± 2.32 mg Trolox/kg) were found in sesame seed and flaxseed oils, respectively. Identified metabolic markers can be used as qualitative markers to confirm the authenticity or to detect adulterations of oils. Composition, properties and authenticity testing should be more rigorous for food products marketed as health-promoting.

Unexpected content of kynurenine in mother's milk and infant formulas.

Mother's milk is widely recommended as complete food for the offspring in earliest postnatal time. However, the knowledge about detailed composition and the physiological role of bioactive components of breast milk is incomplete. Therefore, the aim of our study was to determine the content of kynurenine (KYN) in human breast milk during lactation and to explore the effects exerted by intragastric KYN administration from birth to weaning on physical and psychomotor development of adult rats. We found that KYN is consistently present in human milk and its content gradually increased from day 4 to 28 after delivery and that it is present in commercial baby formulas in amounts noticeably exceeding its physiological range. Animal studies showed that KYN supplementation resulted in a marked elevation of absorptive surface of rat intestine and in enhanced expression of both, aryl hydrocarbon receptor and G protein-coupled receptor 35 in the intestinal tissue in rats. Moreover, we discovered that KYN administration from birth to weaning resulted in neurobehavioral changes in adult rats. Therefore, we postulate that further research is required to thoroughly understand the function of KYN in early developmental stages of mammals and to ensure the safety of its presence in baby food products.

Multispecies Identification of Oilseed- and Meat-Specific Proteins and Heat-Stable Peptide Markers in Food Products.

Consumer demand for both plant products and meat products enriched with plant raw materials is constantly increasing. Therefore, new versatile and reliable methods are needed to find and combat fraudulent practices in processed foods. The objective of this study was to identify oilseed species-specific peptide markers and meat-specific markers that were resistant to processing, for multispecies authentication of different meat and vegan food products using the proteomic LC-MS/MS method. To assess the limit of detection (LOD) for hemp proteins, cooked meatballs consisting of three meat species and hemp cake at a final concentration of up to 7.4% were examined. Hemp addition at a low concentration of below 1% was detected. The LOD for edestin subunits and albumin was 0.9% (w/w), whereas for 7S vicilin-like protein it was 4.2% (w/w). Specific heat-stable peptides unique to hemp seeds, flaxseed, nigella, pumpkin, sesame, and sunflower seeds, as well as guinea fowl, rabbit, pork, and chicken meat, were detected in different meat and vegan foods. Most of the oilseed-specific peptides were identified as processing-resistant markers belonging to 11S globulin subunits, namely conlinin, edestin, helianthinin, pumpkin vicilin-like or late embryogenesis proteins, and sesame legumin-like as well as 2S albumins and oleosin isoforms or selected enzymic proteins.

Amino- and polyaminophthalazin-1(2H)-ones: synthesis, coordination properties, and biological activity.

Amino- and polyaminophthalazinones were synthesized by the palladium-catalyzed amination (alkyl- and arylamines, polyamines) of 4-bromophthalazinones in good yields. The coordinating properties of selected aminophthalazinones towards Cu(II) ions were investigated and the participation of the nitrogen atoms in the complexation of the metal ion was shown. A biological screening of the potential cytotoxicity of selected synthesized compounds on HT-29 and PC-3 cell lines, as well as on the L-929 cell line, proved that some amino derivatives of phthalazinone show interesting anticancer activities. The detailed synthesis, spectroscopic data, and biological assays are reported.

Peptide markers for distinguishing guinea fowl meat from that of other species using liquid chromatography-mass spectrometry.

The inability to easily identify the animal species in highly processed meat products makes them highly susceptible to adulterations. Reliable methods for detecting the species origin of meat used in processed food are required to ensure adequate labelling and minimize food fraud and allergenic potential. Liquid chromatography high resolution mass spectrometry was employed to identify new heat-stable guinea-fowl-specific peptide markers that can differentiate guinea fowl meat from other commonly consumed animal species, including closely related poultry species, in highly processed food products. We identified 26 unique guinea-fowl-specific markers. The high stability of guinea-fowl-specific peptides was confirmed by analysing food products with guinea fowl meat content ranging from 4% to 100%. The findings indicate that sensitive and reliable LC-MS/MS methods can be developed for the targeted detection and quantification of guinea fowl meat in highly processed meat products.

Amiodarone Enhances Anticonvulsive Effect of Oxcarbazepine and Pregabalin in the Mouse Maximal Electroshock Model.

Accumulating experimental studies show that antiarrhythmic and antiepileptic drugs share some molecular mechanisms of action and can interact with each other. In this study, the influence of amiodarone (a class III antiarrhythmic drug) on the antiseizure action of four second-generation antiepileptic drugs was evaluated in the maximal electroshock model in mice. Amiodarone, although ineffective in the electroconvulsive threshold test, significantly potentiated the antielectroshock activity of oxcarbazepine and pregabalin. Amiodarone, given alone or in combination with oxcarbazepine, lamotrigine, or topiramate, significantly disturbed long-term memory in the passive-avoidance task in mice. Brain concentrations of antiepileptic drugs were not affected by amiodarone. However, the brain concentration of amiodarone was significantly elevated by oxcarbazepine, topiramate, and pregabalin. Additionally, oxcarbazepine and pregabalin elevated the brain concentration of desethylamiodarone, the main metabolite of amiodarone. In conclusion, potentially beneficial action of amiodarone in epilepsy patients seems to be limited by neurotoxic effects of amiodarone. Although results of this study should still be confirmed in chronic protocols of treatment, special precautions are recommended in clinical conditions. Coadministration of amiodarone, even at low therapeutic doses, with antiepileptic drugs should be carefully monitored to exclude undesired effects related to accumulation of the antiarrhythmic drug and its main metabolite, desethylamiodarone.

LIQUID CHROMATOGRAPHY-MASS SPECTROMETRY BOTTOM-UP PROTEOMIC METHODS IN ANIMAL SPECIES ANALYSIS OF PROCESSED MEAT FOR FOOD AUTHENTICATION AND THE DETECTION OF ADULTERATIONS.

This review offers an overview of the current status and the most recent advances in liquid chromatography-mass spectrometry (LC-MS) techniques with both high-resolution and low-resolution tandem mass analyzers applied to the identification and detection of heat-stable species-specific peptide markers of meat in highly processed food products. We present sets of myofibrillar and sarcoplasmic proteins, which turned out to be the source of 105 heat-stable peptides, detectable in processed meat using LC-MS/MS. A list of heat-stable species-specific peptides was compiled for eleven types of white and red meat including chicken, duck, goose, turkey, pork, beef, lamb, rabbit, buffalo, deer, and horse meat, which can be used as markers for meat authentication. Among the 105 peptides, 57 were verified by multiple reaction monitoring, enabling identification of each species with high specificity and selectivity. The most described and monitored species by LC-MS/MS so far are chicken and pork with 26 confirmed heat-stable peptide markers for each meat. In thermally processed samples, myosin, myoglobin, hemoglobin, l-lactase dehydrogenase A and ß-enolase are the main protein sources of heat-stable markers. © 2019 John Wiley & Sons Ltd. Mass Spec Rev.

Proteomic analysis of oilseed cake: a comparative study of species-specific proteins and peptides extracted from ten seed species.

BACKGROUND: In recent years there has been a visible trend among consumers to move away from consuming meat in favor of plant products. Meat producers have therefore been trying to meet the expectations of consumers by introducing new products to the food market with a greater proportion of plant ingredients. Meat products are enriched not only by the addition of vegetable oils but also by ground or whole oilseeds or their preparation. In this study, we present in-solution tryptic digestion and an ultra-performance liquid chromatography coupled to quadrupole time-of-flight mass spectrometry (UHPLC-Q-TOF-MS/MS)-based proteomics approach to investigate specific proteins and peptides of ten oilseed cakes, by-products of cold pressing oil from coconut, evening primrose, hemp, flax, milk thistle, nigella, pumpkin, rapeseed, sesame, and sunflower seeds, for authentication purposes. RESULTS: We identified a total of 229 unique oilseed proteins. The number of specific proteins varied depending on the sample, from 4 to 48 in evening primrose and sesame. Moreover, we identified approximately 440 oilseed unique peptides in the cakes of all the analyzed oilseeds; the largest amounts were found in sesame (107 peptides), sunflower (100), pumpkin, hemp (42), rapeseed (36), and flax cake (35 peptides). CONCLUSIONS: We provide novel information on unique / species-specific peptide markers that will extend the scope of testing the authenticity of a wide range of foods. The results of this peptide discovery experiment may further contribute to the development of targeted methods for the detection and quantification of oilseed proteins in processed foods, and thus to the improvement of food quality. © 2020 Society of Chemical Industry.

Identification of species-specific peptide markers in cold-pressed oils.

In recent years, cold-pressed vegetable oils have become very popular on the global market. Therefore, new versatile methods with high sensitivity and specificity are needed to find and combat fraudulent practices. The objective of this study was to identify oilseed species-specific peptide markers, using proteomic techniques, for authentication of 10 cold-pressed oils. In total, over 380 proteins and 1050 peptides were detected in the samples. Among those peptides, 92 were found to be species-specific and unique to coconut, evening primrose, flax, hemp, milk thistle, nigella, pumpkin, rapeseed, sesame, and sunflower oilseed species. Most of the specific peptides were released from major seed storage proteins (11 globulins, 2S albumins), and oleosins. Additionally, the presence of allergenic proteins in the cold-pressed oils, including pumpkin Cuc ma 5, sunflower Hel a 3, and six sesame allergens (Ses i 1, Ses i 2, Ses i 3, Ses i 4, Ses i 6, and Ses i 7) was confirmed in this study. This study provides novel information on specific peptides that will help to monitor and verify the declared composition of cold-pressed oil as well as the presence of food allergens. This study can be useful in the era of widely used unlawful practices.

LC-QTOF/MS determination of tryptophan and kynurenine in infant formulas.

A rapid, reliable and sensitive liquid chromatography quadrupole time-of-flight mass spectrometry method for the determination of tryptophan and its metabolite kynurenine in milk formulas for neonates and infants was developed and validated. Two extraction techniques based on EMR Lipid QuEChERS and liquid-liquid extraction with diethyl ether to extract lipids and methanol to precipitate the protein were tested and compared. Four different infant formula products were randomly selected and evaluated for the effect of co-extracted matrix components on the quantitative analysis results. The influence of matrix components on analytical signals was normalized by the use of stable isotope-labeled standards and matrix-matched calibration. The developed method was found to be sensitive and effective for both analytes in all the examined infant formulas with satisfactory linearity (R2 ≥ 0.9995), recovery in the range of 75.7% ± 4.5 - 99.0% ± 1.1, and intra- and inter-day precision with the coefficient of variation below 6.3% and 17.9%, respectively. The limits of detection (LOD) and quantification (LOQ) for both compounds differed significantly between the examined formulas. The LOD and LOQ values were found to be in the range of 2.18-9.85 µg/g and 6.61-29.84 µg/g for the determination of tryptophan and in the range of 0.21-2.71 µg/g and 0.63-8.23 µg/g for the determination of kynurenine, respectively. The method was proved to be suitable for the determination of tryptophan and kynurenine in infant formulas, and it can be used to study the link between tryptophan metabolism via kynurenine pathway and metabolic disorders in infants.

LC-QTOF-MS identification of rabbit-specific peptides for authenticating the species composition of meat products.

Rabbit is a healthy meat, with low allergenicity and excellent nutritional properties. The global popularity of rabbit meat makes it a target for food fraud. We present a LC-QTOF-MS/MS approach for detecting and identifying rabbit-specific peptide-markers from thermally processed meat products to differentiate rabbit from other commonly-consumed animal species. We identified 49 heat-stable specific peptides. We selected the most stable markers for testing complex meat matrices by analysing pâtés-type products with a rabbit meat content ranging from 5% to 85%. Of the 49 heat-stable peptides detected in pure cooked rabbit meat, three were consistently detected in all investigated pâté samples i.e., SSVFVADPK, AFFGHYLYEVAR and PHSHPALTPEQK. Monitoring meat species other than rabbit in the examined pâtés using pork-, lamb- and chicken-specific peptides identified the presence of undeclared chicken in two samples. The results confirm that LC-QTOF-MS/MS is a suitable tool for multi-species detection in processed meat products, particularly for authentication purposes.